Transgenic Plants in Agriculture

Proceeding of the UGC Sponsored National Seminar on ''THE ROLE OF BIOLOGY IN BRINGING SECOND GREEN REVOLUTION"

Organised by Department of Botany, M. S. College, Saharanpur, (CCS University, Meerut, UP.) on 11 & 12 October, 2015

Editor Dr. Vijai Malik M.Sc., M.Phil., Ph.D., F.A.P.S., F.A.P.T., F.B.S. Assistant Professor, Department of Botany M. S. College, Saharanpur (UP) India

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ISBN: 978-93-84659-19-6

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TABLE OF CONTENT S. No. 1

Title Green Revolution for Qualitative Traits in Plants: Biofortification of Nutrients

Name of the Authors

Page No.

Rajni Shukla, Rashmi Upadhyay and Yogesh Kumar

1-8

Sharma 2 3

Shoving Towards a New Green Revolution Biopesticide: An Ecofriendly Approach For Second

Vijai Malik Rashmi Nigam

9-13 14-18

Vishal Kaushik

19-25

Green Revolution in Agriculture 4

Rhododendron Diversity and their Conservation in Sikkim Himalayas

5

Changes in Micromorphology of Plant Sida veronicaefolia in Response to Air Pollution Stress in Meerut City

Shiv Kumari and Ila Prakash

26-32

6

In vitro study of an endangered “Miracle plant”: Mandookparni (Centella asiatica (L.) Urb.) Role of Nanotechnology in Pollution Control:

Shalini Sharma and Y. Vimala

33-40

Shalini Singh

41-45

Shail Pande

46-50

7

Review 8

Identification of Geminivirus in Cowpea (Vigna unguiculata) Through Amplification of Selective DNA Fragment Using Degenerate Primers

9

Science and Technology in Rural India

Richa Atreya

51-55

10

Effects of Distellery Effluent Irrigation on Mustard (Brassica

Renu Choudhary, Naresh

56-66

campestris)

Kumar and Harendra Malik

11

Role of Transgenic Plants in Agriculture

Renu Rani

67-71

12

Depression in Serum Zinc Concentration and Elevation in

Punam Yadav

72-79

Namita Yadav & Y.K. Sharma

80-83

Serum Potassium Concentration in Chronic Renal Failure Patients 13

Enhancement of Fe and Zn in Cowpea grain by applying soil and foliar application

14

Adoption of Improved Technologies in Black Gram Crop

Lokendra Kumar Singh

84-88

15

Study on SO2 Induced Effects and their Amelioration in Arhar (Cajanus cajan)

Harendra Malik, Naresh Kumar

89-98

and Renu Choudhary

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Synthesis, Characterization and Biological Activities of Some

Dinkar Malik

99-105

Nutritional sink formation in galls of Alstonia scholaris Linn. (Apocynaceae) by the insect Pauropsylla tuberculata (Homoptera: Psyllidae). Perspective of Stem Cell Research in India

Deepak Kumar, Vijai Malik and S.C. Dhiman

106-114

Bindu Sharma

115-120

19

Study of Medicinal Angiosperms of Rampur District (UP), India With Special Reference to Their Sustainable Use

Beena Kumari

121-129

20

Water Quality Control of Industrial Effluents Using

Anuja Agarwal & Vaishali

130-137

Abhishek Gupta & Bindu

138-143

Metal Complexes with 2- Amino -4- (p- Ethoxy Phenyl) Thiazoline Ligand 17

18

Crosslinked Chitosan Hydrogel Beads. 21

Clinical Application and Potential Role of Stem Cells

Sharma 22

‘‘Wild Life and Biodiversity in the Plays of Shakespeare’’

Rashmi Rana

144-148

23

Conservation of Withania somnifera and Commiphora wightii

Anshu Dhaka & Vinit Kumar

149-152

Shyam Singh

153-162

Meenakshi

163-164

Om Dutta

165-171

27

Morphological Studies of Certain Plant Parasitic Nematodes Associated with Vegetable Crops of District Bulnadshahr Genetics of Hg++ -tolerance in Barley

Ritu Aggarwal

172-179

28

Stem Cell Therapy

Pragati

180-182

medicinal plants 24

The zero energy sewage treatment plant (ZESTP). A conventional or alternative method of sewage treatment to enhance the livelihood of marginal formers in peri urban region of Gorakhpur (U.P.)

25

Health Issues in Old Women: Reasons and Suggestion for Improvement

26

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Themes to be Covered 

Role of Biotechnology and Nanotechnology to revolutionize Agriculture and Food Industry.



Means of enhanced productivity by employing high yielding varieties of crop plants.



Pest and disease control.



Soil chemistry.



Conventional manures and recent thoughts.



Water management-water scarcity and water security.



Mechanization.



Due stress on forestry and ecological balance.



Conservation of plants and animals.



Agricultural management.



Human health.



Biodiesel and alternate energy sources.



Generating gainful self employment.



Government policies and recent initiatives.



Climate change and food security.



Food and nutrition security.



Stem cell therapy.



Science education in rural areas

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Green Revolution for Qualitative Traits in Plants: Biofortification of Nutrients Rajni Shukla, Rashmi Upadhyay and Yogesh Kumar Sharma Department of Botany, University of Lucknow, Lucknow (226 007) The Agricultural Revolution was a period of technological improvement and increased crop productivity that occurred during the 18th and early 19th centuries in Europe. The need of food is based on nutrient requirement. More than 12% of soils are deficient in available iron. Zinc deficiency also occurs in crops and human on a world scale and is now regarded as next to iron. Iron deficiency is growing a health concern in the developing world, and responsible for diverse of health complications including anemia and impairments in immune system. Zinc is an essential micronutrient for plant and human, and over 40% of children in the world are suffering from Zn malnutrition. Iron and Zinc deficiency in human is mainly caused from low Fe or Zn concentration and their bioavailability in foods or edible parts of crop plants. Cereal grains are the most important dietary source of micronutrients in many developing countries. Micronutrient concentrations and bioavailability in cereal grain is generally low. Biofortification appears to be a most sustainable and cost-effective approach to solve human micronutrient malnutrition. The genetic capacity of the newly developed (biofortified) cultivars to absorb sufficient amount of Fe and Zn from soil and to accumulate it in the grain may not be expressed to the full extent. It is, therefore, essential to have a short term approach to improve Fe and Zn concentration in cereal grains. Application of FeSO 4 as foliar spray, Zn fertilizers or Zn-enriched NPK fertilizers (e.g., agronomic biofortification) offer a rapid solution to the problem, and represents useful complementary approach to on-going breeding programs. There is increasing evidence showing that foliar or combined soil+foliar application of Fe and Zn fertilizers under field conditions are highly effective and very practical way to maximize uptake and accumulation of Fe and Zn in whole cereal grains, raising concentration up to 20mg kg-1 Fe and 60 mg kg−1 Zn. Increasing the micronutrient concentration of cereal grains has been identified as a way of addressing human micronutrient deficiencies. Revolution The Agricultural Revolution was a period of technological improvement and increased crop productivity that occurred during the 18th and early 19th centuries in Europe. In this lesson, learn the timeline, causes, effects and major inventions that spurred this shift in production. There have been so many revolutions in the system time to time as per the need of human kind* Black Revolution - Petroleum Production : 1970 * Blue Revolution - Fish Production : 1960 Father of Blue revolution Prof. Hiralal Chaudhuri. * Brown Revolution - Leather/non-conventional/Cocoa production * Golden Fiber Revolution - Jute Production * Golden Revolution - Fruits/Overall Horticulture development/Honey Production

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in * Green Revolution - Food grains : Father of Green Revolution M.S. Swaminathan. * Grey Revolution - Fertilizer : 1964 * Pink Revolution - Onion production/Pharmaceutical/Prawn production * Red Revolution - Meat & Tomato Production * Round Revolution – Potato * Silver Fiber Revolution – Cotton Production * Silver Revolution - Egg/Poultry Production * White Revolution (In India: Operation Flood) - Milk/Dairy production : 1970 Father of White revolution Dr. Verghese Kurien * Yellow Revolution - Oil Seeds production : Father of Yellow Revolution Sam Pit Roda Orange Revolution: After 20 years of persistent research, city-based National Research Centre for Citrus (NRCC) has sown the seeds of a new orange revolution in near future. The Centre has succeeded in increasing the productivity and tolerance to diseases of Nagpur orange or mandarin and acid lime manifold by simply replacing the conventionally used root stock of Rangpur lime and rough lemon with an exotic root stock from USA called as Alemow (Citrus macrophylla) during budding/grafting. The process of bringing this change and actual scientific trials took over 18-20 years. "Alemow is being used worldwide as a root stock for improving productivity of fruits, especially citrus family. Our Centre began work in this direction as an in-house research way back in 1992. The root stock has much superior horticultural characteristics like smaller canopy, higher productivity and better disease tolerance compared to the domestic root stocks," said the NRCC director V J Shivankar. The productivity of orange has increased from just 9-10 tonnes/ha to 21 tonnes/ha in orange and from five tonnes/ha to 13 tonnes/ha in acid lime. Both orange and acid lime developed using Alemow root stock also have increased tolerance towards main citrus diseases, especially soil borne diseases like phytophthora fungus, the biggest threat to orange. Right now, the success of technology is also being tested at national level through an All India Coordinated Research Project (AIRCP) on orange by the Indian Council of Agricultural Research (ICAR) in all the citrus growing belts of the country. Principal scientists in horticulture and the project leader of the research R K Sonkar, who was recently honoured with Vidarbha Bhushan award for leading the work, told TOI that the centre was also planning to try the technology on other orange varieties like the Kinno, Khasi and Kurg mandarins if these national level trials yielded good results. "The technology is not new. World over, Alemow is being used as root stock for many citrus crops. It is being tried in India for the first time. We started experimenting with 12 different root stocks but Alemow turned out to be the best," said Sonkar. To develop planting material for orange, nurseries use Rangpur lime and rough lemon as root stocks on which the buds of Nagpur orange are put. Both these root stocks have much less yield and are susceptible to diseases. A good root stock provides the grower a useful tool to manipulate the vigour and performance of the orchard. Though even Alemow is not completely resistant to phytophthora, the fungus attack is delayed by many

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in years and is comparatively milder. Also, since the canopy size is reduced with Alemow use, sunlight penetrates the lowest part of the tree and increases fruit bearing ability. This makes the new trees more suitable for 'high density plantation' the technique being promoted for all horticultural crops now. Instead of conventional 6mX6m distance these can be planted at a distance of 6mX4m. Sonkar explained that acid lime was generally grown through seedling but in this technology, budding is used. The production through seedling is just 5 tonnes/ha whereas budding on Alemow root stock increased this yield to 13 tonnes/ha due to vigorous growth that occurs. Some farmers who regularly visit NRCC on their own decided to take up cultivation using Alemow root stock. Bandu Wasankar from Paratwada has tried it successfully. Mohan Tambi has gone step ahead and imported the root stock from USA for his nursery and is developing planting material from them. Some farmers in Solapur too are trying it. Centre in the long run will try to work on drought tolerance capacity too. Red Revolution: The hill state of Uttarakhand is silently scripting a 'red revolution' of sorts on the Himalayan reaches. Growing a lesser known red cereal crop, amaranth, is opening up new vistas for the farmers settled in the upper areas where staple plantations of wheat and paddy are not possible because of extreme climate conditions and mountainous topography. Amaranth is a traditional fibre-rich plant used in baby food products, breads, etc. for its high protein content and other nutritional values. In the past two years, international demand for amaranth has risen to the extent that the farmers in the state are unable to meet the supply orders. "Demand for amaranth is largely from the overseas markets of South Africa and the Netherlands. The cereal is grown only in this state. Even as the area under amaranth cultivation has increased over the years, the demand and supply is a mismatch, perhaps to the advantage of the farmers," said Binita Shah, senior programme manager, Uttarakhand Organic Commodity Board (UOCB). Shah said the state, which acts as a facilitator, has an estimated standing demand of over 1,000 tonnes from overseas buyers, against an estimated production of close to 600 tonnes expected this year. "A few years ago, our visit to higher areas in the state brought to the fore a sordid tale. Farmers used to barter amaranth for 2 kg of wheat, salt and a little more. Today, their income has increased manifold," she said. To augment the demand, both in terms of export orders and domestic supplies, the government is laying more emphasis on plantation of organic amaranth to empower farmers, said Chief Minister Ramesh Pokhriyal "Nishank". From insignificant gains till some years ago, farmers this year got a handsome Rs 3,500 per quintal of amaranth, up nearly 30 per cent since last year. Close to 5,000 hectares of area in upper areas of Garhwal, Uttarakashi, Chamoli, Rudraprag is under amaranth production and just about 500 hectares under certified organic produce in Uttarakhand. An increase in food production, especially in underdeveloped and developing nations, through the introduction of high-yield crop varieties and application of modern agricultural techniques. The introduction of high-yielding varieties of seeds and the increased use of chemical fertilizers and irrigation are known collectively as the Green Revolution, which provided the increase in production needed to make India self-sufficient in food grains, thus improving agriculture in India. High-yielding wheat was first introduced to India in 1968 by American agronomist Norman Borlaug. Borlaug has been hailed as the Father of the Green Revolution but M.S. Swaminathan is known as the "Father of the Green Revolution in India". The methods adopted included the use of high yielding varieties

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in (HYV) of seeds. The production of wheat has produced the best results in fueling self-sufficiency of India. Along with high yielding seeds and irrigation facilities, the enthusiasm of farmers mobilized the idea of agricultural revolution and is also credited to M. S. Swaminathan and his team had contributed towards the success of green revolution. Due to the rise in use of chemical pesticides and fertilizers there were many negative effects on the soil and the land such as land degradation. Blue Revolution Blue Revolution means the adoption of a package programme to increase the production of fish and marine products. The Blue Revolution in India was started in 1970 during the Five-Year Plan when the Central Government sponsored the Fish Farmers Development Agency (FFDA). Subsequently, the Brakish Water Fish Farms Development Agency were set up to develop aquaculture. The Blue Revolution has brought improvement in aquaculture by adopting new techniques of fish breeding, fish rearing, fish marketing, and fish export. Under the Blue Revolution programme, there had been a tremendous increase in the production of shrimp. Andhra Pradesh and Tamil Nadu have developed shrimp in a big way. The Nellore District of Andhra Pradesh is known as the 'Shrimp Capital of India'. Pink Revolution Pink Revolution is a term used to denote the technological revolutions in the meat and poultry processing sector. India has already seen the ‘green’ and ‘white’ revolutions in its food industry – related to agriculture and milk respectively, now thrust is upon meat and poultry sector. India being a country of huge cattle and poultry population, has high potential for growth if this sector is modernized. Green Revolution: The Green Revolution refers to a series of research and development and technology transfer initiatives, occurring between the 1930s and the late 1960s (with prequels in the work of the agrarian genetist Nazareno Strampelli in the 1920s and 1930s), that increased agricultural production worldwide, particularly in the developing world, beginning most markedly in the late 1960s. The initiatives, led by Norman Borlaug, the "Father of the Green Revolution," who won the Nobel Peace Prize in 1970, credited with saving over a billion people from starvation, involved the development of high-yielding varieties of cereal grains, expansion of irrigation infrastructure, modernization of management techniques, distribution of hybridized seeds, synthetic fertilizers, and pesticides to farmers. The term "Green Revolution" was first used in 1968 by former United States Agency for International Development(USAID) directorWilliam Gaud, who noted the spread of the new technologies: "These and other developments in the field of agriculture contain the makings of a new revolution. It is not a violent Red Revolution like that of the Soviets, nor is it a White Revolution like that of the Shah of Iran. I call it the Green Revolution." Effect of Green Revolution on Crop Production: Cereal production more than doubled in developing nations between the years 1961–1985. Yields of rice, maize, and wheat increased steadily during that period. The production increases can be attributed roughly equally

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in to irrigation, fertilizer, and seed development, at least in the case of Asian rice. While agricultural output increased as a result of the Green Revolution, the energy input to produce a crop has increased faster, so that the ratio of crops produced to energy input has decreased over time. Green Revolution techniques also heavily rely on chemical fertilizers, pesticides and herbicides and rely on machines, which as of 2014 rely on or are derived from crude oil, making agriculture increasingly reliant on crude oil extraction. Proponents of the Peak Oil theory fear that a future decline in oil and gas production would lead to a decline in food production or even a Malthusian catastrophe . Technologies used in Green Revolution: Technology is the application of organized and scientific knowledge to solve practical problems. In agriculture, the success of a technology can be measured only when it gets transferred for increased crop production. The following technologies have helped in increasing crop production in the World: 1.

Dwarfing gene

2.

Hybrid Technology

3.

Biotechnology particularly cry genes in cotton

4.

Seed Technologies: Seed Vigor, seed coating

5. Agronomic Technologies: Row-Plant Spacing minimum tillage etc Introduction: The diets of over two-thirds of the world’s population lack one or more essential mineral elements. This can be remedied through dietary diversification,mineral supplementation, food fortification, or increasing the concentrations or bioavailability of mineral elements in produce (biofortification). Humans require at least 22 mineral elements for their wellbeing(Welch & Graham, 2004; White & Broadley, 2005a; Graham et al., 2007). These can be supplied by an appropriate diet. However, it is estimated that over 60% of the world’s 6 billion people are iron (Fe) deficient, over 30% are zinc (Zn) deficient, 30% are iodine (I) deficient and c. 15% are selenium (Se) deficient (see Supporting Information References S1). In addition, calcium (Ca), magnesium (Mg) and copper (Cu) deficiencies are common in many developed and developing countries (Frossard et al., 2000; Welch & Graham, 2002,2005; Rude & Gruber, 2004; Grusak & Cakmak, 2005;Thacher et al., 2006). Currently, mineral malnutrition is considered to be among the most serious global challenges to humankind and is avoidable (Copenhagen Consensus 2004;http://www.copenhagenconsensus.com). Mineral malnutrition can be addressed through dietary diversification, mineral supplementation, food fortification and/or increasing mineral concentrations in edible crops (biofortification). Because the seeds of many cereals are often consumed after milling or polishing, it is pertinent to consider whether genetic variation in the distribution of mineral elements within the seed can be utilized in biofortification strategies. Mineral elements are nonhomogenously distributed within the seed and the concentrations of many mineral elements are highest in the husk and/or aleurone layers (see References S12). Milling or polishing cereal seeds can, therefore, remove large quantities of mineral elements from the diet and the extent of these losses is genotype dependent (Gregorio et al., 2000; Vasconcelos et al., 2003; Ma et al., 2004; Lyons et al., 2005; Prom-uthai et al., 2007).

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Improving the crop yield and quality through Biofortification: Regarding the nutritional traits, one of the most promising application of transgenic technology has been the development of vitamin A enriched varieties, popularly known as Golden Rice due to the slightly yellow colour conferred to the endosperm. First successful product of GM based biofortification with pro-vitamin A trait. Researchers have found that in Japonica Golden Rice, an intake of only 120 g is required for meeting the Vit A’s RDA. (Paine et al, 2005;Tang et al, 2009) Bio fortification of rice with iron and zinc are also being carried out by several public and private sector organizations. Increasing the concentration of micronutrient (especially Zn) in food crop plants is a growing global challenge, with potentially great implications for both crop production and human health. It is believed thatZn deficiency is the most widespread micronutrient deficiency in crop plants and human beings (Alloway, 2004; Hotz and Brown, 2004). Zn deficiency in humans causes a wide range of health complications, including impairments in the immune system, learning ability and physical growth, and increases in mortality and infections (Hotz and Brown, 2004; Cunnigham-Rundles et al., 2005). Zinc deficiency also induces DNA damage and increases the risk of cancer occurrence (Ho, 2004). Zinc and Fe deficiencies are a growing public health and socioeconomic issue, particularly in the developing world (Welch and Graham 2004). Recent reports indicate that nearly 500,000 children under 5 years of age die annually because of Zn and Fe deficiencies (Black et al.2008). Zinc and Fe deficiencies together with vitamin A deficiency have been identified as the top priority global issue to be addressed to achieve a rapid and significant return for humanity and global stability (www.copenhagenconsensus.com). Low dietary intake of Fe and Zn appears to be the major reason for the widespread prevalence of Fe and Zn deficiencies in human populations. In countries with a high incidence of micronutrient deficiencies, cereal-based foods represent the largest proportion of the daily diet (Cakmak 2008). Cereal crops are inherently very low in grain Zn and Fe concentrations, and growing them on potentially Zn- and Fe-deficient soils further reduces Fe and Zn concentrations in grain (Cakmak et al. 2010). Thus, biofortification of cereal crops with Zn and Fe is a high-priority global issue. HarvestPlus (www.harvestplus.org) is the major international consortium to develop new plant genotypes with high concentrations of micronutrients by applying classical and modern breeding tools (i.e. genetic biofortification). Although plant breeding is the most sustainable solution to the problem, developing new micronutrient-rich plant genotypes is a protracted process and its effectiveness can be limited by the low amount of readily available pools of micronutrients in soil solution (Cakmak 2008). Application of Zn- and Fe-containing fertilizers (i.e. agronomic biofortification) is a short-term solution and represents a complementary approach to breeding. Cereals (e.g.wheat, rice and maize) with inherently low Zn concentrations in the grain are the most important source of calories in the developing world. In the present study, we used a range of wheat genotypes with different concentrations of Zn and other micronutrients to determine whether the DTZ method could be useful as a rapid screening method for seed Zn concentration.

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GW-366T1SEED (Control)

DL-803(T1)SEED (Control)

GW-366T2SEED

GW-366T3SEED

High Efficient Variety

DL-803(T2)SEED

DL-803(T3)SEED

Low Efficient Variety Biofortification of Zinc in two different variety of Wheat through soil application (T2)and (T3)Foliar application

Here we have taken two different variety of wheat where one is High efficient variety name GW-366 mean that variety which perform better in sense of yield in case of adverse condition and the second variety is Low efficient variety name DL-803 means that variety is not performing better in unfavourable condition. Now the biofortification of zinc is provided in two ways the first one is through soil application (T1) and second is soil + Foliar application (T3). Here through analytical study biofortification of

zinc is occurs more in foliar

application.variety GW-366 is much better highly fortified in comparision to the variety DL-803. The localization and accumulation of zinc is found in foliar application of mature seed of GW-366 variety. References: 1.

Black RE, Lindsay HA. Bhutta ZA, Caulfield LE, De Onnis M, Ezzati M, Mathers C, Rivera J (2008). Maternal and child undernutrition: global and regional exposures and health consequences. Lancet 371,243-260.

2.

Cakmak I (2008) Enrichment of cereal grains withzinc: Agronomic or genetic biofortification? Plant and Soil 302,1-17.

3.

Cakmak I, Pfeiffer WH, McClafferty B (2010) Biofortification of durum wheat with zinc and iron. Cereal Chemistry 87,10-20.

4.

Welch RM, Graham RD (2004) Breeding for micronutrients in staple food crops from a human nutrition perspective. Journal of Experimental Botany 55,353–364.

5.

White PJ, Broadley MR. 2005. Biofortifying crops with essential mineral elements. Trends in Plant Science 10: 586–593.

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 6.

Hotz C, Brown KH (2004) Assessment of the risk of zinc deficiency in populations and options for its control. Food Nutr Bull 25: 94–204.

7.

Alloway BJ (2004) Zinc in Soils and Crop Nutrition.International Zinc Association Communications. IZAPublications, Brussel.

8.

Graham RD, Welch RM, Saunders DA, Ortiz-Monasterio I, Bouis HE Bonierbale M, de Haan S, Burgos G, Thiele G, Liria R et al. 2007. Nutritious subsistence food systems. Advances in Agronomy 92: 1–74.

9.

Frossard E, Bucher M, Mächler F, Mozafar A, Hurrell R. (2000). Potential for increasing the content and bioavailability of Fe, Zn and Ca in plants for human nutrition. Journal of the Science of Food and Agriculture 80:861–879.

10. Welch RM, Graham RD. 2002. Breeding crops for enhanced micronutrient content. Plant and Soil 245: 205–214. 11. Welch RM, Graham RD. 2005. Agriculture: the real nexus for enhancing bioavailable micronutrients in food crops. Journal of Trace Elements in Medicine and Biology 18: 299–307. 12. Rude RK, Gruber HE. 2004. Magnesium deficiency and osteoporosis: animal and human observations. Journal of Nutritional Biochemistry 15: 710–716. 13. Grusak MA, Cakmak I. 2005. Methods to improve the crop-delivery of minerals to humans and livestock. In: Broadley MR, White PJ, eds. Plant nutritional genomics. Oxford, UK: Blackwell, 265–286. 14. Thacher TD, Fischer PR, Strand MA, Pettifor JM. (2006). Nutritional rickets around the world: causes and future directions. Annals of Tropical Paediatrics 26: 1–16.

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Shoving Towards a New Green Revolution Vijai Malik Department of Botany, M. S. College, Saharanpur Abstract India is an agricultural country due to its strategic geographical, climatic and topographical location on the world map. Even with industrial revolution, the importance of agriculture has not reduced. Agricultural selfsustenance has brought India pride during the first green revolution, as the country had sustained several famines from British ruled period till just after independence. The seat of first green revolution was north- west India mainly, with wheat being the focal staple crop. Second Green Revolution in fact should be focused to practice sustainable agriculture at regional level and crop wise. It must aim at raising the productivity of other important food crops such as Sorghum, Millets and Cassava- food produced and consumed mainly by the world’s poor. The present article includes aim, calls, and some of the approaches that may push India towards Second Green Revolution. Key Words: Second Green Revolution, Food Security, Nanotechnology Introduction: Research in applied Botany is playing and will play important role in self sufficiency towards food production. Advance research in Genetics, Cytogenetics, Plant Breeding, Molecular biology, Biotechnology etc. are helping a lot of in this direction. Not only Yoga and use of medicinal plants are helping in building good character and health of human beings but also recent advances in Molecular Biology, Molecular Cytogenetics and Pant Biotechnology are further enhancing food production and human health. We have seen a revolution in the past concerning with food production, i.e. Green Revolution. Due to shrinkage of agricultural land by industrialization, urbanization and many fold increase in population, this revolution has no means. We need big revolution at this juncture. The term Green revolution is applied to the period 1967-1978. In 1968, William S. Gaud coined the term “Green Revolution” to describe phenomenal growth in agriculture. The first green revolution relied heavily on the use of large amounts of fertilizers, pesticides and other agricultural inputs. This coupled with continued expansion of farming areas led to self-sufficiency in food production. A quantum jump in the productivity and production of wheat and then rice transformed the image of India as a begging bowl to a bread basket. Second Green Revolution in fact refers to practicing sustainable agriculture. That is, protecting natural resources from becoming increasingly degraded and polluted and using production technologies that conserve and enhance the natural resource base of crops, forests, inlands and marine fisheries [1]. Aim of Second Green Revolution: Declining productivity due to unstable agriculture practices over the years and galloping rate of population rate have both put a severe strain on the food supply situation in the country. To meet the food requirements of the

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in growing population a second green revolution has become imperative. Second Green Revolution must aim at raising the productivity of other important food crops such as Sorghum, Millets and Cassava- food produced and consumed mainly by the world’s poor. Calls for a Second Green Revolution The Second Green Revolution is a change in agricultural production widely thought necessary to feed and sustain the growing population on Earth. These calls have precipitated in part, as a response to rising food commodity prices, and fears of peak oil among other factors. Calling for a second Green Revolution, Prime Minister Sh. Narendra Modi has asked farmers to adopt scientific methods to enhance food grain production and reduce imports by using one-fifth of their farming land to cultivate lentils. "Unless we prepare a balanced and a comprehensive integrated plan, we will not be able to change the lives of farmers," PM said, adding that Indian farmers are still lagging behind in terms of availability of good quality seeds, adequate water, power, right price and market for their produce. Invoking former Prime Minister Lal Bahadur Shastri's slogan "Jai Jawan, Jai Kisan," Modi asked farmers to try and grow pulses on part of their land. "The production of pulses in the country is very low. I (PM) urge farmers that if they have five acres of farming land, use four acres for other crops but cultivate pulses on at least one acre. This basic source of protein should be available to the poor at affordable prices," he said. The Centre is willing to pay more than the announced minimum support price to farmers for pulse production, he said. "We have seen the first Green Revolution, but it happened several years ago. Now it is the demand of time that there should be a second Green Revolution without any delay. And where is it possible? It is possible in eastern UP, Bihar, West Bengal, Jharkhand, Assam, and Odisha", PM said. "The government is making every effort to make the farmers aware of modern scientific advancements," he added. The PM said only one model of farm technology cannot be adopted uniformly in the country as there are variations in soil and climatic conditions to consider. Pitching for 'per drop, more crop', he stressed the need for research in the field of agriculture to determine the health of soil and its needs in terms of seeds, water quantity, amount of fertilization, etc. "We want private individuals to own laboratories, similar to pathological laboratories, to test soil and issue a health card so that a farmer is aware of the deficiency, fertilizer requirements and type of crop suitable for his plot of land," he said, adding this will also lead to job creation. He also added that students enrolling in IARI from different parts of the country would come for higher education and research and would then return to their respective states after developing a model suitable for that geographical region [2]. Role of Nanotechnology to Revolutionize Agriculture

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in The technology applied in agriculture is referring as green technology. The green technology subjects are energy, green buildings, environmentally preferred purchasing, green chemistry and green nanotechnology. Nanotechnology in agriculture has gained momentum in the last decade with an abundance of public funding, but the pace of development is modest, even though many disciplines come under the umbrella of agriculture. This could be attributed to: a unique nature of farm production, which functions as an open system whereby energy and matter are exchanged freely; the scale of demand of input materials always being gigantic in contrast with industrial nanoproducts; an absence of control over the input nanomaterials in contrast with industrial nanoproducts (eg, the cell phone) and because their fate has to be conceived on the ecosphere-biosphere-hydrosphere-atmosphere continuum; the time lag of emerging technologies reaching the farmers’ field, especially given that many emerging economies are unwilling to spend on innovation; and the lack of foresight resulting from agricultural education not having attracted a sufficient number of brilliant minds the world over, while personnel from kindred disciplines might lack an understanding of agricultural production systems. If these issues are taken care of, nanotechnologic intervention in farming has bright prospects for improving the efficiency of nutrient use through nanoformulations of fertilizers, breaking yield barriers through bionanotechnology, surveillance and control of pests and diseases, understanding mechanisms of host-parasite interactions at the molecular level, development of new-generation pesticides and their carriers, preservation and packaging of food and food additives, strengthening of natural fibers, removal of contaminants from soil and water, improving the shelf-life of vegetables and flowers, clay-based nanoresources for precision water management, reclamation of salt-affected soils, and stabilization of erosion-prone surfaces, to name a few. Time for a Second Green Revolution After Green Revolution India had become self-sufficient in basic food grains (wheat and rice), but recently we are facing recurrent spells of shortages in essential items like lentils, edible oil, sugar and onions. This has resulted in their knee-jerk imports at exorbitant prices. According to experts if we do not focus immediately on increasing food production by rationalizing priorities in the agriculture sector, we may be in for a rude shock of unmanageable food scarcity. In the present scenario cultivable land and water availability are shrinking. We have almost dried our rivers and are emptying groundwater reservoirs by reckless extraction. Extensive use of chemical fertilizers and pesticides has polluted surface water and but groundwater resources. Researchers should focus on alternative organic fertilizers and pesticides which will need less water. Irrigation should aim at giving only the required amount of water in the root zone of plants for which drip and/or sprinkler system must be applied in the entire cultivable area—through subsidy, wherever necessary. Provide water to every field. This has to be done by making field channels, wherever terrain permits. At all other places sprinklers with minimal extraction of groundwater should be used to meet the target of total irrigation coverage to cultivated land. We will have to collect and store each drop of water, check its misuse and wastage, and use it frugally yet efficiently—especially in agriculture, because that is the biggest user sector of water. This should be the target of the second Green Revolution.

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in For Second Green Revolution we will have to develop techniques which would deliver more crops for each drop of irrigation as well as more yield per field of cultivated land. Although land available for agriculture is declining there is still scope to increase productivity from it, mainly because we are far behind most countries in per hectare yield. With new technology, improved seeds, rationalized crop rotation and balancing soil chemistry etc., we can obtain much more crop from the available land. Genetically improved seeds, which are being scoffed at—due to lack of proper education and information—can enhance productivity greatly. Time has come to launch a second Green Revolution to provide long-term food security to the nation, and it is the direction in which agriculture scientists and institutions need to focus. Agriculture scientists should start their research by accepting that whatever water is available for irrigating crops is not only limited but also mostly polluted. Their target should be to obtain maximum production from frugal use of this water in irrigation. Of course, reuse and recycling of unclean water would be attempted after treating it to acceptable parameters for agricultural application [3]. The Second Green Revolution through Biotechnology: The world’s food grain production was tripled and saved millions of life from famine. This could happen because of the Green Revolution during 1960s. Despite of that, feeding an estimated worldwide human population of over 9 billion by 2050 and alleviating the malnutrition in the poor nations are still the massive global challenges. Conventional breeding strategies alone may not be able to increase the productivity without disturbing the environment. The biotechnological interventions including tissue culture, genetic engineering and molecular breeding have proved as a game changer. In India, Bt-cotton the only transgenic crop is the live demonstration of the importance of genetic engineering that has revolutionized cotton production by escalated yields, reduced insecticide applications, and improved socioeconomic livelihood of farmers. But misinformation, lack of public awareness and current national policies are the major bottleneck in the adoption of this technology. A second revolution has been made possible by the sequencing of the rice genome in 2005 (the first cereal crop to be sequenced). This enabled breeders to discover the genes for flood resistance in one obscure variety from eastern India and transfer them to varieties all round the world. Breeders will soon do the same for genes that provide other valuable traits. Conclusion: 70% of Indian population lives in rural areas and 60% of its workforce is agriculture. As we all know current state of agriculture in India is result of green revolution which is in place since late 1960’s, which was heavily backed by government. It has delivered India food security and sufficiency which was critical at that time. This progress and security had its own costs in terms of environment and economic viability. Green revolution rampantly used fertilizers and other chemicals, which made food and water, toxic to some extent. India’s 80% of fresh water is

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in consumed by agriculture, more particularly by rice farming. Consequently, new agriculture policy of India aims at sustainable agriculture, which is popularly called ‘Second Green Revolution’ or ‘Evergreen Revolution’. Declining Indian economy and growing food insecurity policy and environmental experts has called for a Second Green Revolution. The only way to attain food and nutritional security is by improving marginal and dry land agriculture in a sustainable manner by implementing Second Green Revolution. References: Kumar S. Towards a Second Green Revolution. Science Reporter (June) 9-15 (2008). Deogharia J. PM calls for Second Green Revolution. The Times of India, June 29, (2015) Narain Y. and Kumar S. K. Time for a Second Green Revolution. Indian Express, June 26, (2015).

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Biopesticide: An Ecofriendly Approach For Second Green Revolution in Agriculture Rashmi Nigam Assistant Professor, Department of Plant Pathology, Janta Vedic College, Baraut, Baghpat(U.P.) ABSTRACT Agriculture plays a vital role in a developing country like India. Apart from fulfilling the food requirement of the growing Indian population, it also plays a role in improving economy of the country. Emphasis of present day agriculture is to produce more with lesser land, water and man power. The Green Revolution technology adoption during 1960 to 2000 has increased wide varieties of agricultural crop yield per hectare which increased 12-13% food supply in developing countries. Inputs like fertilizers, pesticides helped a lot in this regard. But in spite of this fact, food insecurity and poverty still prevails prominently in our country. Use of chemical pesticides and fertilizers have caused negative impact on environment by affecting soil fertility, water hardness, development of insect resistance, genetic variation in plants, increase in toxic residue through food chain and animal feed thus increasing health problems and many more which makes bio pesticides to come into picture. The present study deals with different type of biopesticide used in agriculture. Bio-pesticides are ecofriendly pesticides which are obtained from naturally occurring substances (biochemicals), microbes and plants. They are also biochemical pesticides that are naturally occurring substances that control pests by nontoxic mechanisms. Biopesticides are living organisms (natural enemies) or their products (phytochemicals, microbial products) or by products (semiochemicals) which can be used for the management of pests that are injurious to plants. The most commonly used biopesticides are living organisms, which are pathogenic for the pest of interest. These include biofungicides (Trichoderma), bioherbicides (Phytopthora) and bioinsecticides (Bacillus thuringiensis). There are few plant products also which can now be used as a major biopesticide source. The stress on organic farming and on residue free commodities would certainly increased adoption of biopesticides by the farmers. Keywords: Biopesticide, Ecofriendly, Biofungicide, Bacillus thuringensis. Introduction Agriculture plays a vital role in a developing country like India as it plays a important role in improving economy of the country. Agriculture is adversely affected by destructive activities of numerous pests like bacteria, fungi, weeds and insects, leading reduced yields. Since 1960s, the most common method for pest control has been based on the intensive use of synthetic organic pesticides. The Green Revolution technology adoption during 1960 to 2000 has increased wide varieties of agricultural crop yield per hectare which increased 12-13% food supply in developing countries. Inputs like fertilizers, pesticides helped a lot in this regard. The indiscriminate use of chemical pesticides in modern agriculture resulted in the development of several problems such as pesticide resistance insects, resurgence of target and non target pests, destruction of beneficial organisms like honey bees, pollinators, parasitoids and predators and pesticide residue in food, feed and fodder (Al-Zaidi et al., 2011). Hence, the today’s need is to produce maximum from the decreasing availability of natural resources without adversely affecting the environment. Therefore, alternative, environmentally safe methods are needed for pest management. As a result,

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in bio-pesticides using eco-friendly, plant or microbial derived ingredients showing broad insecticidal effects with minimal damage to the environment have been increasingly developed. Biopesticides are generally less toxic than chemical pesticides, often target specific, cause minimal harm to birds, insects and mammals. In addition, even if used in an open field, they decompose quickly, thereby minimizing the risk of environmental pollution or residual toxicity.

Bio-pesticides are ecofriendly pesticides which are obtained from naturally occurring substances

(biochemicals), microbes and plants (Dutta, 2015). Biopesticides are certain types of pesticides derived from such natural materials as animals, plants, bacteria, and certain minerals. The most commonly used biopesticides are livingorganisms, which are pathogenic for the pest of interest. These include biofungicides (Trichoderma), bioherbicides (Phytopthora) and bioinsecticides (Bacillus thuringiensis).There are few plant products also which can now be used as a major biopesticide source (Salma et al., 2011). Plant-incorporated protectants include substances that are produced naturally on genetic modification of plants. Such examples are incorporation of BT gene, protease inhibitor, lectines, chitinase etc into the plant genome so that the transgenic plant synthesizes its own substance that destroys the targeted pest. In India, some of the biopesticides like Bt, NPV, neem based pesticides, etc. have already been registered and are being practiced. (Gupta, 2010). In the present study, an attempt has been made to provide a comprehensive study or review on biopesticide, its ecofriendly approach on the sustainable agriculture or for an evergreen revolution in agriculture. The study deals with different type of biopesticide, used in agriculture, its application and advantages. The study is based on secondary data which has been collected from the different sources. Types of Biopesticide Biopesticides are an important ingredient of Integrated Pest Management (IPM) packages due to their capability in maintaining the natural diversity without the use of any artificial or synthetic residues. The origin of Biopesticides can be microbial (bacteria, fungi or virus), herbal (plant extracts) or genetically modified plants (GM). Beauveria spp., Trichoderma spp., and Bacillus spp., are some of the microbial biopesticides. Biopesticides may be broadly categorized into three major groups: 1) Biochemical biopesticides 2) Microbial pesticides and 3) Plant-Incorporated Protectants (PIPs). 1. Biochemical pesticides: Biochemical pesticides are naturally occurring substances that control pests by non-toxic mechanisms. Conventional pesticides are generally synthetic materials that directly kill or inactivate the pest. Biochemical pesticides are herbal-based substances that are naturally produced by a plant or an organism. They are non-toxic and biodegradable. They help the plant in counter-attacking its pests or producing chemicals that would prevent pest attack on the plant. Biochemical pesticides include substances that interfere with growth or mating, such as plant growth regulators, or substances that repel or attract pests, such as pheromones. Because it is sometimes difficult to determine whether a substance meets the criteria for classification as a biochemical pesticide, EPA has established a special committee to make such decisions. Plant biochemicals are collectively called botanicals and the most important botanical is pyrethrum, followed by neem, rotenone and essential oils, typical used as insecticides (e.g. pyrethrum, rotenone, rape seed oil, quassia extract, neem oil, nicotine), repellents (e.g. citronella), fungicides (e.g. laminarine, fennel oil, lecithine), herbicides (e.g. pine oil), sprouting inhibitors (e.g. caravay seed oil) (Isman, 2006).

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 2. Microbial pesticides: These pesticides originate from micro-organisms such as bacteria, fungi or other protozoan groups. These are mostly target-specific organisms that are aimed at killing one or a group of pests. Microbial pesticides consist of a microorganism (e.g., a bacterium, fungus, virus or protozoan) as the active ingredient. Desai, (1997) concluded that microbial pesticides can control many different kinds of pests, although each separate active ingredient is relatively specific for its target pest. Forexample, there are fungi that control certain weeds, and other fungi that kill specific insects. Microbial pesticides can control many different kinds of pests these include biofungicides (Trichoderma), bioherbicides (Phytopthora) and bioinsecticides (Bacillus thuringiensis and Baculovirus). The most widely used microbial pesticides are subspecies and strains of Bacillus thuringiensis, or Bt., can control certain insects in cabbage, potatoes, and other crops Each strain of this bacterium produces a different mix of proteins, and specifically kills one or a few related species of insect larvae. Some Bt's control moth larvae found on plants, other Bt's are specific for larvae of flies and mosquitoes. The target insect species are determined by whether the particular Bt produces a protein that can bind to a larval gut receptor, thereby causing the insect larvae to starve. (Kalra, and Khanuja, 2007). The widely used microbial pesticides are Trichoderma viride, Bacillus thuringenesis, Bacillus sphaericus, Pseudomonas fluorescence (Bacteria), Beauveria bassiana (Fungi), Baculo virus and Nucleopolyhedrosis Virus. 3. Plant-Incorporated-Protectants (PIPs): These are genetically modified materials produced by modifying a protein and introduced into the plant so that it produces its own pesticide. Plant-Incorporated-Protectants are pesticidal substances that plants produce from genetic material that has been added to the plant. For example, the gene for Bt pesticidal protein, was introduced into the genetic material of cotton plant and plant manufactures the substance that destroys the pest. The protein and its genetic material, but not the plant itself, are regulated by EPA. (Thakore, 2006). Transgenic plant produces biodegradable protein with no harmful effect on animals and human beings, and thus minimizes the use of hazardous pesticides.

Applications of Biopesticide Biopesticides are usually applied in a manner similar to chemical pesticides, but achieve pest management in an environmentally friendly way. With all pest management products, but especially microbial agents, effective control requires appropriate formulation (Burges,1998) and application (Matthews et al., 2014, Lacey and Kaya, 2007). Biopesticides for use against crop diseases have already established themselves on a variety of crops. For example, biopesticides already play an important role in controlling downy mildew diseases. A major growth area for biopesticides is in the area of seed treatments and soil amendments. Fungicidal and biofungicidal seed treatments are used to control soil borne fungal pathogens that cause seed rots, damping-off, root rot and seedling blights. They can also be used to control internal seed–borne fungal pathogens as well as fungal pathogens that are on the surface of the seed. Many biofungicidal products also show capacities to stimulate plant host defence and other physiological processes that can make treated crops more resistant to a variety of biotic and abiotic stresses. Biopesticide Technology in India The Bio pesticide segment occupies a small portion of the large pesticide market in India. In 2005, it accounted for just 2.89%, which was expected to increase by 2.3%. Globally, there are 175 registered biopesticide active-

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in ingredients and 700 products available in the market (Hajeck and Leger, 1994). In India, so far only 12 types of biopesticides have been registered under the Insecticide Act, 1968 (Thakore, 2006). Neem based pesticides, Bacillus thuringensis, NPV and Trichoderma are the major biopesticides produced and used in India.

Last decade, has

witnessed a rapid growth in this segment, especially on standardization of production techniques of Trichoderma, Gliocladium, Paecilomyces, Pseudomonas, Trichogramma, NPV and Bacillus to use them against many insect pests and diseases. Today, these biological control agents have been successfully employed in India. Trichogramma, which is a stingless wasp that feeds on the eggs of sugarcane borers, has been used against borers in the states of Tamil Nadu, Rajasthan, UP, Bihar and Haryana. Similarly Trichogramma, Bracon, Chelonus and Chrysopaspp are being used for the control of cotton bollworms. Trichogramma has also been used against rice stem borer and leaf folder. The sugarcane scale insect has been controlled with the help of predatory Coccinellid beetles in UP, West Bengal, Gujarat and Karnataka. Most of the biopesticides find use in public health, except a few that are used in agriculture as transgenic plants and beneficial organisms called bio-agents: are used for pest management in India (Kalra, and Khanuja, 2007). Important biopesticide registered in India are Bacillus thuringiensis var. israelensis, Bacillus thuringiensis var. kurstaki, Bacillus thuringiensis var. galleriae, Bacillus sphaericus, Trichoderma viride, Trichoderma harzianum and Pseudomonas fluoresens. Shia and Feng, (2004) reported some successful utilization of biopesticides and bio-control agents in Indian agriculture like control of rots and wilts disease in various crops by Trichoderma based products, control of mango hoppers and mealy bugs and coffee pod borer by Beauveria, control of Helicoverpa on cotton, pigeon-pea, and tomato by Bacillus thuringiensis, control of white fly on cotton by neem products, control of diamondback moths by Bacillus thuringiensis, control of sugarcane borers by Trichogramma and control of Helicoverpa on gram by N.P.V. Advantages of Biopesticide 

Biopesticides are usually inherently less toxic than conventional pesticides. No harmful residues detected.

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Biopesticides generally affect only the target pest and closely related organisms, in contrast to broad spectrum, conventional pesticides that may affect organisms as different as birds, insects and mammals.

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Biopesticides often are effective in very small quantities and often decompose quickly, resulting in lower exposures and largely avoiding the pollution problems caused by conventional pesticides. It can be cheaper than chemical pesticides when locally produced.

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When used as a component of Integrated Pest Management (IPM) programs, biopesticides can greatly reduce the use of conventional pesticides, while crop yields remain high. Biopesticides can be more effective than chemical pesticides in the long-term

Conclusion India has a vast potential for biopesticides. To develop eco-friendly pest control technologies are the need of day and challenging tasks for developing countries to improve agricultural productivity in a sustainable manner. Biopesticides are considered as one of the eco-safe alternatives due to their biodegradation in nature, multiple mode of action on target pests and may not leave toxic residues,are used globally for controlling insect pests and diseases. Thus bio-pesticides would achieve the target of evergreen revolution or second green revolution if these bio-

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in molecules can be characterized for their potency using available modern biotechnological tools, to develop low cost technology for compound isolation, formulation and commercial scale establishment of plant resource and microbial pesticides. The stress on organic farming and on residue free commodities would certainly warrant increased adoption of biopesticides by the farmers. References 1.

Al-Zaidi, A. A., Elhag, E. A., Al-Otaibi, S. H. and Baig, M. B. (2011). Negative effects of pesticides on the environment and the farmers awareness in Saudi Arabia: a case study. J. Anim. Plant Sci. 21(3): 605611.

2.

Burges, H.D. (1998). Formulation of Microbial Biopesticides, beneficial microorganisms, nematodes and seed treatments Publ. Kluwer Academic, Dordrecht, 412 pp.

3.

Desai, S. T. (1997). Chemical industry in the post independence era: a finance analysis point of view. Chemical Business, 11(1): 25 - 28.

4.

Hajeck, A. E and Leger, St. (1994) Interactions between fungal pathogens and insect hosts, Annual Review of Entomology, 39: 293 - 322.

5.

Isman, M. B., (2006) Botanical insecticides, deterrents, and repellents in modern agriculture and an increasingly regulated world, Annu. Rev. Entomol. 51 45–66.

6.

Kalra, A. and Khanuja, S. P. S. (2007). Research and Development priorities for biopesticide andbiofertiliser products for sustainable agriculture in India. In. Business Potential for Agricultural Biotechnology (Teng, P. S. ed.), Asian Productivity Organisation, 2007; 96-102.

7.

Lacey, L. and Kaya, H. (2007). Field Manual of Techniques in Invertebrate Pathology2nd edition. Kluwer Academic, Dordrecht, NL

8.

Matthews, G. A., Bateman, R. P., Miller, P. C. H. (2014). Pesticide Application Methods (4th Edition), Chapter 16. Wiley, UK.

9.

Salma Mazid,Ratul Rajkhowa, and Jogen Kalita ( 2011). Article “A Review on use of Biopesticides in Insect Management”, International Journal of Science and Advanced Technology: 1 (7).

10. Shia, W.B. and. Feng, M.G. (2004) Lethal effect of Beauveria bassiana, Metarhizium anisopliae, and Paecilomyces fumosoroseus on the eggs of Tetranychus cinnabarinus (Acari:Tetranychidae) with a description of a mite egg bioassay system, Biological Control; 30: 165– 173. 11. Dutta, S. (2015) Biopesticide: An ecofriendly approach for pest control.World Journal of Pharmacy and Pharmaceutical Sciences. 4,( 06),, 250-265 12. Gupta, S. (2010). Biopesticides: An eco-friendly approach for pest control, Journal of Biopesticides,; 3(1 Special Issue): 186 - 188. 13. Thakore, Y. (2006). The biopesticide market for global agricultural use. Industrial Biotechnology; 194208.

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Rhododendron Diversity and their Conservation in Sikkim Himalayas Vishal Kaushik, Department of Botany, N. R. E. C. College, Khurja. [email protected] Abstract The genus belongs to family Ericacae of order Ericales. In India the members of the family are confined to higher altitudes and grow from 1,500 to 6,000 meters. But most of the species are found at altitudes between 2,500 to 4,500 meters. The Rhododendron are found along with Pines and Junipers Elnus, Popular, Betula and Quercus in Eastern Himalayas. The plants sprout in February, March and profuse flowering continues till July. Flowers with beautiful shapes and shades add to the beauty of the high altitudes. Forty three species of Rhododendron are found in India out of which thirty six species are confined to Sikkim alone. The Rhododendron is a keystone species in the higher altitudes of Himalayas. It is a sacred plant and has a lot of social importance attached to it. It also has aesthetic, ethano - medicinal, medicinal, and commercial value. The plant is also used as fuel wood. Due to anthropogenic activities the natural populations and diversity of Rhododendron species in the entire Himalayas are gradually diminishing. The major threats to Rhododendron population and diversity are deforestation, and unsustainable extraction for firewood and incense by local people. Many species of Rhododendron which are classified as rare or endangered may soon become extinct and will we lost forever if proper conservation measures are not taken. The present study deals with the study of diversity of the plant, its conservation status and strategies being used to tackle the situation. Key Words: Rhododendron, Diversity, Conservation, Sikkim, Himalayas. Introduction: The genus Rhododendron, belongs to the family Ericaceae and was founded by Linnaeus [1]. The word Rhododendron itself is derived by two Greek words rhodon (rose) and dendron (tree) which mean, rose tree. This genus is represented by 850 species in the world [2], which are mostly distributed at higher elevations in the Sino-Himalayan region with maximum concentration in Western China [3]. In India, the genus is mostly confined to the Himalayan region, especially in the Eastern Himalayas. A revision of the genus was carried out by Cullen [4], Chamberlain [5], Philipson and Philipson [6],Chamberlain and Rae [7], Kron [8] and, Judd and Kron [9]. Some inventories of the genus were also made by Pradhan [3,10], Ghosh and Samaddar [11], Bhattacharyya and Sanjappa [12]. Sastry and Hajra [13] and Mao et al. [14] contributed towards the study of rare and endemic Rhododendrons of India. Extensive study of Rhododendrons of the Sikkim-Himalayan region were conducted by Pradhan and Lachungpa [15] and Singh et al. [16]. The Indian Himalayas are one of the most recent and fragile mountain regions of the world. They house a large number of biological species, and harbour one of the biological hot spots of the world. The region is quite rich in a variety of flora, fauna, human settlements, tribes and culture. Of the estimated 8,000 species of vascular plants in the Himalayan region, around 3,160 are endemic and 450 species are endangered [17, 18].

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in This diversity is constantly and increasingly under pressure from anthropogenic activities. The indiscriminate exploitation of the flora, fauna and other natural resources present in the region, destruction of natural habitats, spread of non biodegradable organic chemicals, introduction of alien species, have taken a toll on the number of a number of plant and animal species, which have disappeared completely while others await the same fate [19]. Due to this disappearance of the natural flora and fauna, an alarm has been raised by the enlightened gentry of the world towards the conservation of the various species present in the world. In this regard, efforts are being made to conserve the various species and genera endemic to the respective areas. One such plant species endemic to the Himalayas is the Rhododendron. It is found at high altitudes in Himalayan regions of both the Western and Eastern Himalayas at very high altitudes. Again in the North - Eastern states a very large number of species of the plant ( Rhododendron ) are present. Due to human interference the natural populations of Rhododendrons in the entire Himalaya are gradually diminishing. The major threats to Rhododendrons are deforestation and unsustainable extraction for firewood and incense by local people. A set of Rhododendrons which are classified as rare/endangered may be wiped out from the biota in the near future if proper conservation measures are not made. [20] In the present paper we deal with the diversity and conservation status of Rhododendron in the state of Sikkim. Material And Methods: The present work is based on extensive literature surveys made in the state of Sikkim., Published scientific papers, monographs, red-list documents, IUCN list, etc. were consulted, for the threat categories. All the taxa observed and studied, have been meticulously listed in table form with their vital information and their threat categories have also been listed. Results and Discussion: A total of 38 species, 3 subspecies and 2 varieties of Rhododendrons were recorded in the state of Sikkim in the study, the details are provided in the table. The distribution of species in relation to altitude is as follows: The maximum numbers of Rhododenrons are present at an altitude of 3001-3500 mts. Minimum number of the said plant are found at the lower altitudes of 500-1000 mts. and above 5000 mts. The genera was absent at an altitude of less than 500 mts. Only one species viz., namely R. arboreum Sm. is only found at an altitude of less than 1000 mts. (800 m onwards). R.nivale Hook. f. was the single species found above the altitude of 5000 mts. The three taxa found endemic to Sikkim were R. candelabrum Hook. f. ,

R. decipiens Lacait. and R.

sikkimense U. C. Pradhan & S. T. Lachungpa. The following four taxa were found to be endangered, these are R. leptocarpum Nutt. , R. maddenii Hook. f., R. niveum Hook. f. and R. pumilum Hook. f.. of the observed taxa Nine were found to be rare which are, R. baileyi Balf. f., R. campylocarpum Hook. f., R. edgeworthii Hook. f., R. keysii Nutt., R. maddenii Hook. f., R. papillatum Balf. f. & Copper, R. pendulum Hook. f., R. wightii Hook. f. and R. xanthostephanum Merrill out of which R. maddenii Hook. f. is both endangered and rare. The species of Rhododendrons exhibits significant diversity in habit and broad range of distribution from the altitude of 800-6000 mts. A total of 38 species, 3 subspecies and 2 varieties of Rhododendrons were recorded in the state of Sikkim in the study. Out of these species, 3 species were endemic to Sikkim; four

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in species were found to be endangered, while nine species were found to be rare and 23 were not yet evaluated till the time of study[14,16,20]. On comparison with neighboring states the maximum concentration of species is observed in Arunachal Pradesh , where 75of the total 87 species of Rhododendron, found in the eastern Himalayas are observed, after that the states in the order of decreasing diversity of Rhododendron species are Sikkim, Manipur, Mizoram, Nagaland and Meghalaya. Compared with other Rhododendron rich regions China has a total of 571 species of Rhododendrons, out of which 409 species are endemic [21]. Countries like Pakistan, Bhutan, Nepal, etc. are have very little species diversity of Rhododendrons. Considering the altitude the maximum number of Rhododendron species are present in the altitudes of 3001-3500 mts. These altitudes are considered as optimum suitable heights for the growth the Rhododendrons and for their conservation and multiplication. In the present times, the diversity as well as the number of species of Rhododendrons is adversely affected due to threats posed by natural calamaties and anthropogenic activities. The rise in population with demand on land for farming, increased animal husbandry practices, construction of roadways, hydel-power stations and allied works, army personnel garrisoned at alpine locations and lately the tourist influx have collectively resulted in the building up of considerable pressure on the availability of Rhododendron species [20,21]. The major threats to Rhododendrons are deforestation and unsustainable extraction for firewood and incense by local people. Due to the presence of polyphenols and flavonoids, Rhododendrons make excellent firewood that burns even under wet conditions. Rhododendron firewood is also being used in the high-altitude trekking corridor for the purpose of tourism. Some of the species have already become scarce, for example, R. leptocarpum is endangered and reported to have only 16 surviving individuals at present in the Sikkim [16, 20]. The conservation of Rhododendron species can be done by, the in-situ and ex-situ modes of conservation. Insitu conservation can be brought about by establishing Rhododendron sanctuaries, Parks, etc. which is being looked into by the Sikkim government and the Government of India. Two Rhododendron sanctuaries have already been established at Barsey in West Sikkim and at Shingba in North Sikkim[22]. Other than that various governmental agencies and NGO’s also take up programs from time to time to create awareness for the environment. Some efforts by Sikkim forest department and Sikkim Rhododendron Society have been made by fencing the Rhododendron rich sites and declaring them as Rhododendron Sanctuary between Lachung and Yumthang in the State. The ex-situ conservation can be made by cultivating Rhododendron species in the gardens and parks under suitable climatic conditions or by using tissue culture techniques. The plants can be introduced in Botanical Gardens and Parks also. The species of Rhododendron arboreum is being propagated through cuttings [23,24]. Tissue culture studies of Indian Rhododendrons are also being used to propagate the species of which Rhododendron maddeni is a successful example [25]. In many countries, these techniques are already in use for commercial cultivation of Rhododendrons [26-32]. The success of the conservation programs depends as much, on Govermental agencies, NGO’s, as much as on the awareness of local people. But an onus also lies with common tourist also. It is imperative to educate the local inhabitants and the tourism industry operators and

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in tourists about the wealth of Rhododendrons and importance towards the conservation of biodiversity in the region. References [1] Linnaeus, C. “Species Plantarum,” Vol. 1, London, pp. 392. (1753) [2] Mabberley, D. J. “Mabberley’s Plant-Book. A Portable Dictionary of Plants,

Their Classifications and

nd

Uses,” 2 Edition, Cambridge University Press, Cambridge, (2008). [3] Pradhan, U. C. “A Preliminary enumeration of Rhododendrons of the Indian region—Part 1,” Himalayan Plant Journal, Vol. 3, No. 8, , pp. 110-123, (1985). [4] Cullen, J. “A Revision of Rhododendron. I. Subgenus Rhododendron Sections Rhododendron and Pogonanthum,” Notes from the Royal Botanic Garden Edinburgh Vol. 39, pp. 1-207, (1980). [5] Chamberlain, D. F. “A Revision of Rhododendron II. Subgenus Hymenanthes,” Notes from the Royal Botanic Garden Edinburgh, Vol. 39, pp. 209-486, (1982). [6] Philipson, W. R. and Philipson, M. N. “A Revision of Rhododendron III. Subgenera Azaleastrum, Mumeazalea, Candidastrum and Therorhodion,” Notes from the Royal Botanic Garden Edinburgh, Vol. 44, pp. 1-23, (1986). [7] Chamberlain, D.F. and Rae, S. J. “A revision of RhododendronIV. Subgenus Tsutsusi,” Edinburgh Journal of Botany, Vol. 47, pp. 89-200,(1990). [8] Kron, K. A. “A Revision of Rhododendron Section Pentanthera,”Edinburgh Journal of Botany, Vol. 50, pp. 249-364, (1993). [9] Judd, W. S. and Kron, K. A. “A Revision of RhododendronVI. Subgenus Pentanthera (Sections Sciadorhodion, Rhodora, and Viscidula),” Edinburgh Journal of Botany,Vol. 52, pp. 1-54, (1995). [10] Pradhan, U.C. “A Preliminary Enumeration of Rhododendrons of the Indian region—Part 2,” Himalayan PlantJournal, Vol. 4, No. 11-12, pp. 73-76, (1986). [11] Ghosh, R.B. and Samaddar, U.P. “The Rhododendrons of the North-East India,” Journal of Economic and Taxonomic Botany, Vol. 13, No. 1, pp. 205-220, (1989). [12] Bhattacharyya, D. and Sanjappa, M. “Rhododendrons Habitats in India,” Journal of American Rhododendron Society Vol. 62, No. 1, pp. 14-18, (2008). [13] Sastry, A. R. K. and Hajra, P.K. “Rare and endemic species of Rhododendrons in India—A Preliminary Study,” In: S. K. Jain and R. R. Rao, Eds., An Assessment of Threatened Plants of India, BSI, Calcutta, pp. 222-231, (1983). [14] Mao, A. A. Singh, K. P. and Hajra, P. K. “Rhododendrons,” In: N. P. Singh and D. K. Singh, Eds., Floristic Diversity and Conservation Strategies in India, BSI, Calcutta, pp. 2167-2202, (2002). [15] Pradhan, U. C. and Lachungpa, S. T. “Sikkim Himalayan Rhododendrons,” Primulaceae Books, Kalimpong, (1990). [16] Singh, K. K., Kumar, S., Rai, L. K. and Krishna, A. P. “Rhododendron Conservation in Sikkim Himalaya,” Current Science, Vol. 85, pp. 602-606,(2003)

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in [17] Singh, D. K. and Hajra, P. K. “Floristic Diversity,” In: G. S. Gujral and V. Sharma, Eds., Changing Perspectives of Biodiversity Status in the Himalaya, British Council, New Delhi, (1996). [18] Samant, S. S., Dhar, U. and Palni, L. M. S. “Medicinal Plants of Indian Himalaya: Diversity Distribution Potential Values,” Gyanodaya Prakashan, Nainitial, (1998). [19] Singh, N. P., Sharma, J. R., Singh, K. P. and V. Mudgal, “Species Diversity in Angiosperms,” In: N. P. Singh and D. K. Singh, Eds., Floristic Diversity and Conservation Strategies in India, Botanical Survey of India, Kolkata, Vol. 5, pp. 1631-1674.( 2002) [20] Chandra Sekar, K. and Srivastava, S. K. “Rhododendrons in Indian Himalayan Region: Diversity and Conservation” American Journal of Plant Sciences, , 1, 131-137, (2010). [21]Paul, A., Khan, M.L., Arunachalam, A. and Arunachalam, K. Biodiversity and conservation of rhododendrons in Arunachal Pradesh in the IndoBurma biodiversity hotspot. Current Science 89(4): 623634, (2005). [22] http://nbaindia.in/uploaded/state-wise/sikkim/1.list_Protectedareas_SIKKIM.pdf [23] Thakur, P., Sharma, Y. D., Kashyap B. and Thakur, A. “Vegetative propagation of native ornamentals of Himachal Pradesh in India,” Abstract of XXVII International Horticultural Congress—IHC2006, Himachal Pradesh, (2006). [24] Singh, K. K., Kumar ,S. and Shanti, R. “Raising Planting Materials of Sikkim Himalayan Rhododendron through Vegetative Propagation Using Air-Wet Technique,” Journal of American Rhododendron Society, Vol. 62, pp. 136-138, (2008). [25] Singh, K. K. and Gurung, B. “In vitro Propagation of R.maddeni Hook. f. an Endangered Rhododendron Species of Sikkim Himalaya,” Notulae Botanicae Horti Agrobotanici Cluj-Napoca, Vol. 37, No. 1, pp. 7983, (2009). [26] Lloyd, G. and McCown, B. “Community-Feasible Micropropagation of Mountain Laurel, Kalmia latifolia, by Use of Shoot Tip Culture,” Proceedings of International Plant Propagation Society, Vol. 30, pp. 421427, (1981). [27] W. C. Anderson, “A revised Medium for Shoot Proliferation of Rhododendron,” Journal of American Society & Horticultural Science, Vol. 109, pp. 343-347, (1984). [28] Douglas, G. C. “Propagation of Eight Cultivars of Rhododendron in vitro Using Agar-Solidified and Liquid Media and Direct Rooting of Shoots in vivo,” Scientia Horticulturae,Vol. 24, pp. 337-347, (1984). [29] McCown, B.H. and Lloyd, G.B. “A Survey of the Response of Rhododendrons to in vitro Cultures,” Plant cell tissue and organ culture, Vol. 2, pp. 77-85, (1985). [30] Briggs, B. A., McCulloch, S. M. and Caton, L. A. “In vitropropagation of Rhododendron,” Acta Horticulturae, Vol.364, pp. 21-26, (1994). [31] Evers, P. W., Donkers, J., Prat, A. and Vermeer, E. “Micropropagation of forest trees through tissue culture,” Centre for Agricultural Publishing and Documentation, Wageningen, (1988).

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in [32] R. Almeida, S. Gonçalves and A. Romano, “In vitro Micropropagation of Endangered Rhododendron ponticum L.subsp. baeticum (Boissier & Reuter) Handel-Mazzetti,” Biodiversity and Conservation Vol. 14, No. 5, pp. 1059-1069, (2005). Table 1: Rhododendrons of Sikkim S.

NAME OF THE TAXA

No.

ALTITUDE

STAT

(MTS)

US

1

R. anthopogon D. Don

3350-5000

N.E.

2

R. anthopogon D. Don subsp. hypenanthum (Balf. f.) J. Cullen

3350-5000

N.E.

3

R. arboreum Sm.

800-3000

N.E.

4

R. arboreum Sm. subsp. Cinnamomeum (Wall. ex G. Don) Tagg c.

2500

N.E.

5

R. arboreum Sm. var. roseum Lindl.

2500-3600

N.E.

6

R. baileyi Balf. f.

3000-4000

R.A.1

7

R. barbatum G. Don

2500-3700

N.E.

8

R. camelliaeflorum Hook. f.

2700-4000

N.E.

9

R. campanulatum D. Don

2500-4300

N.E.

10

R. campanulatum D. Don subsp. aeruginosum (Hook. f.) D. F. Chamb.

4500-5000

N.E.

11

R. campanulatum D. Don var. wallichii Hook. f.

4000

N.E.

12

R. campylocarpum Hook. f.

3300-4300

R.A.1

13

R. candelabrum Hook. f.

3600-4300

E.N.1

14

R. ciliatum Hook. f.

2700-3400

N.E.

15

R. dalhousiae Hook. f.

1800-2300

N.E.

16

R. decipiens Lacait.

2500-3000

E.N.1

17

R. edgeworthii Hook. f.

2100-3300

R.A.1

18

R. falconeri Hook. f.

2100-4000

N.E.

19

R. glaucophyllum Rehder

3080-3700

N.E.

20

R. grande Wight

2160-3385

N.E.

21

R. griffithianum Wight

2160-2770

N.E.

22

R. hodgsonii Hook. f.

3080-3690

N.E.

23

R. keysii Nutt.

2440-3650

R.A.1

24

R. lanatum Hook. f.

3080-4000

N.E.

25

R. lepidotum Wall. ex D. Don

2160-4620

N.E.

26

R. leptocarpum Nutt.

2300-4310

E.D.2

27

R. lindleyi T. Moore

1850-3080

N.E.

28

R. maddenii Hook. f.

2400-3650

RA1, ED2

29

R. nivale Hook. f.

4000-6000

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NE

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 30

R. niveum Hook. f.

3080-3700

ED2

31

R. papillatum Balf. f. & Copper

1800-3300

RA1

32

R. pendulum Hook. f.

2270-3650

RA1

33

R. pumilum Hook. f.

3500-4500

ED2

34

R. setosum D. Don

2160-4950

NE

35

R. sikkimense U. C. Pradhan & S. T. Lachungpa

3700

EN1

36

R. smithii Nutt.

2160-3700

NE

37

R. thomsonii Hook. f.

3390-4000

NE

38

R. triflorum Hook. f.

2160-2930

NE

39

R. vaccinioides Hook. f.

1850-3700

NE

40

R. virgatum Hook. f.

2160-2770

NE

41

R. wallichii Hook. f.

4000-4500

NE

42

R. wightii Hook. f.

3050-4310

RA2

43

R. xanthostephanum Merrill

1500-3000

RA1

1 - Mao et al., 2002; 2 – Singh et al., 2003; ED - Endangered; EN – Endemic; NE – Not Evaluated; RA – Rare;

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Changes in Micromorphology of Plant Sida veronicaefolia in Response to Air Pollution Stress in Meerut City Shiv Kumari* and Ila Prakash Department of Botany D.N. College Meerut, (U.P.) India [email protected] ABSTRACT The race for rapid development has resulted in unscrupulous exploitation of natural resources. This has disturbed the delicate ecological balance between living and nonliving components of the biosphere. Of the various changes disturbing the ecological balance most originate from industries. Emissions from various industrial units, thermal power plants, etc. have immensely contributed to environmental pollution. Entire global biota (flora and fauna) has become vulnerable to this ever increasing menace. Present study deals with the observation made on the plants on roadside in comparison to plants in controlled area. On roadside plants having heavy auto-exhaust pollution load. Present studies were made on Sida veronicaefolia taken from Garh road, Railway road, Delhi road and University road. These changes have been worked out on the basis of percentage of reduction in various parameters in tested plants. Due to high concentration of automobile pollution, the number of stomata, stomatal index and stomatal density were reduced and Number of epidermal cells and epidermal density were increased. Variation in stomatal index and stomatal density on the adaxial and abaxial surface has been observed in these plants. Key Words: Stomatal Density, Stomatal index, epidermal density, Vehicular exhaust pollution. INTRODUCTION: Our environment is a complex mixture of a number of constituents like air, water, soil, plants and animals, all of which maintain a dynamic inter-relationship and interdependence. The earth is the only planet known in the entire universe capable of supporting life which is due to its unique environment. Any undesirable change in the environment, which may be due to addition of unwanted substances results in atmospheric pollution and disturbs the normal functioning of the ecosystem. Zielinska et al., 2004 reported that the composition of emissions from automobiles highly depend on the fuel type, the state of vehicular maintenance and ambient conditions. Sarkar et al. 1986 observed the high effects of automobile exhaust pollution on Clerodendron incerme, Solanum torum and Calotropis procera along a road carring dense traffic. And found visible injury, necrosis, chlorosis and reduction in leaf area due to air pollution. Decrease in stomatal frequency and occurence of aborted stomata were reported in the leaves of some woody perennials as a result of air pollution (Chattopadhyay, 1996 ). Similarly, decrease in stomatal index, density and coverage area was observed in Dahlia and Tagetus (Dhaka, 1999; Prakash et al., 2008; Helianthus annus (Goswami, 2002; Zea mays (Jeyakumar et al., 2003) and Mammillaria fragilis (Joshi et al.,2004); in sunflower and napier grass (Marie and Romeo, 2008); and Miyazawa et al. 2006.

MATERIAL AND METHOD:

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Leaves collected from different sites and studied. For the study of stomata parameters, the replica technique (conservative facsimile technique) was adoptd (Prakash and Kumar, 1995). In this method, an adhesive such as quick-fix was applied on the surface of the leaf. It was allowed to dry for a few seconds and then a cellotape was placed on the leaf surface. The tape was then pulled off and the impression of leaf epidermis than was obtained under the microscope. Stomatal index was determined as follows.

Stomatal density 

Epidermal density 

Stomatal index 

Number of stomata Field area mm 2





Number of epidermal cells 100 Field area mm2

S 100 E S





(Salisbury, 1927)

Where, S = no. of stomata in microscope field area-1, E = no. of epidermal cells microscopic field area-1

RESULTS & DISCUSSION: The reduction in the number of stomata on adaxial and abaxial surface was Sida veronicaefolia 32.0% and 15.75 at Delhi road (Table: 1, 2). The number of epidermal cells however recorded at increase. The number of epidermal cells was higher at high polluted sites. In Sida veronicaefolia plant the stomatal index at Delhi road was 15.3783 on adaxial surface and 20.6632 on abaxial surface (Table 1, 3). Stomatal density also recorded a decrease at different polluted sites. In In Sida veronicaefolia the stomatal density was 120.3966/ 180.594 on adaxial/ abaxial surface at Delhi road and 173.5127/ 254.957 on adaxial/ abaxial surface at University road (Table 1, 3). Number of epidermal cells got increased in number due to reduction in number stomata was observed with an increase in concentration of pollutants at polluted site. As a result of reduction in the number of stomata, the epidermal density was increased. In Sida veronicaefolia the epidermal density was recorded as 662.1813 mm-2 / 686.968 mm-2 on adaxial / abaxial surfaces at Delhi road. In Sida veronicaefolia, the stomatal coverage area is 2521.2464/ 3271.9546 on adaxial/ abaxial surface at Delhi road and 5545.3257/ 7963.8810 on adaxial/ abaxial surface at University road (Tables 2, 4). In Sida veronicaefolia the number of trichomes recorded an increase of 3.25/1.5 on adaxial/ abaxial surface at Delhi road and 1.0/0.75 on adaxial/ abaxial at University road. T he results were statistically analyzed and interpreted by three way ANOVA. All the data were subjected to statistical analysis to find out Critical Difference at (CD) 5% and 1% level (Fisher, 1951) is superscripted with single star (*) and double star (**)respectively.

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Effect of pollution on various other parameters of plants growth growing under polluted condition might be attributed to the disturbances caused in habitat due to pollutants emitted by diesel engines of locomotives, deposition of smoke, carbon, dust and gaseous pollutant, etc. present study number of stomata decreases with increase in pollution on roadside. As the number of stomata decreases, consequently the number of epidermal cells increases. Decrease in stomata could be regarded as an adaptive feature developed by plants in order to cope up with the effect of the gaseous pollutant which enters the leaf, injuries the tissue and causes death (Chattopadhyay, 1996); Marie et. al., 2008; Prakash et al., 2008. Gaseous pollutant enters the leaves through stomata following the same diffusion pathway as CO2. Stomatal opening is checked by the entry of gases in leaf. The pollutants after entering the leaf dissolve in the apoplastic water to produce mainly sulphite and bisulphite ions as (SO 3-2, HSO3-) which are toxic at high concentrations but at low concentrations are effectively detoxified by plants to sulphate ions which then work as sulphur source for the plant. Kulshreshtha et. al., 2005); Salsbury 2006; Sunstar.com 2009. Urban areas are characterized by higher concentration of SO2, hence the plants in these areas cannot be detoxified rapidly and adapt themselves by following some line of defense against SO 2 stress. It may include stomotal closure and reduction in number of stomata was observed at all sites, so stomatal index and stomatal Density was also reduced accordingly. The number of trichomes was high at upper surface and low at lower surface. Higher number of trichomes was found on the upper surface of the leaf in order to keep the pollutants away from the leaf surface.The gradual increase in trichome number may be due to the tendency of plants to increase the resistance to air pollutants as the trichomes may increase boundary layer resistance and hence reducing the gaseous diffusion into leaves. The effect was more severe on adaxial surface than on abaxial surface. This might be due to direct exposure of the adaxial surface to the pollutant. As the number of stomata decreases, consequently number of epidermal cells increases. This resulted in increase in epidermal density. CONCLUSION: Present paper shows an overview that high concentration of pollutants was observed at Delhi road and low at University and moderate at Garh road and Railway road in comparison to control site. The adverse effects of air pollution on plants were greater in the area receiving higher pollution load and vice-versa. Due to higher level of dust on upper surface of plant, stomatal process and food synthesis in leaves were highly affected. According to the study it can be concluded that Delhi road was most polluted site followed by Garh road, Railway road, and University road in comparison to control. It can be concluded that the high level of pollution in busy roadsites hampers the plant growth and development to an extent that the plant is disturbed. ACKNOWLEDGEMENT: The author is thankful to centre for the study of D.N.College laboratory, Meerut, U.P. India to providing me all the facilities for experimental work. author is also thankful to Dr. (Smt) Ila Prakash for her guidance and supervision for this work. REFERENCES:

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 1) Chattopadhyay, S.P. Leaf surface effects of air pollution on certain tree species in Calcutta, Adv Plant Sci., 9(1), 1-4 (1996). 2) Dhaka, R. Impact of air pollution on some plants. A ph.D. Thesis, C.C.S. Uni. Meerut India, (1999). 3) Fisher, R.A. The Design of Experiments. Six Eds. Oliver and Boyd, London, (1951). 4) Goswami, R. Toxicity of air pollution to plants. A Ph.D Thesis. C.C.S.Univ., Meerut India, (2002). 5) Jeyakumar, M., Jayabalan, N. and Arockiasamy, D.I. Effect of sulphur dioxide on maize (Zea mays L.var. Co-1) seedling at lethal dose 50. Physiol. Mol Biol Plants 9 (1), 147-151 (2003). 6) Joshi, R. Choudhary, M., Tyagi, K. and G. Prakash Experimental analysis on stomatal behaviour in cacti exposed to sulphur dioxide pollution. Prog Agric 4(1), 22-24 (2004). 7) Kulshreshtha, K., K. Srivastava and K. J. Ahmed Effect of automobile exhaust pollution on leaf surface structures of Calotropis procera L. and Nerium indicum L. Feddes repertorium, Environ. Sci. Hlth. Part A, Environ. Sci and Egg. (105), 185- 189 (2005). 8) Marie C.G. Duldulao and Romeo, A. Gomez Effect of vehicular emission on morphological characteristics of young and mature leaves of sunflower and Napier grass. Benguet State University, Research Journal, Volume (xvi), 142-151 (2008). 9) Miyazawa, S., Livingston, N. J. and Turpin, D. H. Stomatal development in new leaves is related to the stomatal conductance of mature leaves in Populas trichocarpa. J. of Experimental Botany, 57 (2), 373-380 (2006). 10) Prakash., Joshi, C. and Chauhan, A. Performance of locally grown rice plants (Oryza sativa) exposed to air pollutants in a rapidly growing industrial area of Haridwar. Life Science Journal. 5(3), 57-61 (2008). 11) Prakash, G. and Kumar, V. Stomatal study on defense organs by conservative facsimile method. J. Ind. Bot. Soc. (74), 263-268 (1995). 12) Prakash, G., Poonia, S., Sharma, S., Sangita and Dhaka, R. Stomatal response to sulphur dioxide exposure in Dahlia variabilis L. and Tagetes patula L. Role of Biological Sciences in New Millennium 39-44 (2001). 13) Salisbury, E.J. On the causes and ecological significance of stomatal frequency with special reference to woodland flora. Phill. Trans, R. Soc. B. Vol. (216), PP. 1-65 (1927, 2006 a). 14) Sarkar, R.K., Banerjee, A. and Mukherji, S. Acute ration of peroxidase and catalase activities in leaves of wild dicotyledonous plants as an indication of automobile exhaust pollution. Environ. Pollut (42), 289-295 (1986). 15) Sunstar .com. ph/ Static/ bag/ news, 2009 /02/10. 16) Zielinska, B., Sagebiel, J., Donald, MC., Whitney, J. D.K., Lawson, D.R. J. Air Manag. Assoc.54 (9), 113850 (2004). TABLES: Table. 1 Stomatal response in terms of no. of epidermal cells, no. of stomata, epidermal density (mm-2),

stomatal

-2

density (mm ) and stomatal index on adaxial surface in Sida veronicaefolia plant.

'THE ROLE OF BIOLOGY IN BRINGING SECOND GREEN REVOLUTION

29

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Different Study Sites

Statistical Value

Attribute

Railway

University

Road

Road

40.25

39.25

38.5

+ 0.9574

+ 1.5

+ 2.5

+ 1.732

12.5

8.5

9.25

10.75

12.25

+ 0.5773

+ 0.5773

+ 0.5

+ 0.5

+ 0.5

382.4362

662.1813**

570.1133**

555.9490**

545.3257**

density (mm-2)

+ 16.355

+ 13.5612

+ 21.246

+ 35.4107

+ 24.533

Stomatal density

177.0538

120.3966*

131.01983*

152.2662*

173.5127

(mm-2 )

+ 8.1777

+ 8.177

+ 7.0821

+ 7.0821

+ 7.0821

31.6548

15.3783**

18.6826**

21.5339**

24.1515*

No. of epidermal cells

Control

Delhi Road

Garh Road

27.0

46.75

+ 1.1547

CD 5%

CD 1%

47.756

113.830

23.878

56.916

3.976

9.478

No. of stomata

Epidermal

Stomatal index + 1.36190

+ 0.8467

+ 0.5084

+ 1.3383

+ 0.9969

Table 2. Stomatal response in terms of length and breadth of stomata and stomatal coverage area (mm -2) on adaxial surface in Sida veronicaefolia plant. Different Study Sites Attribute

Length of stomata(

Breadth of

Statistical value Railway

University

Road

Road

180.0

195.0

202.5

+ 15.0

+ 24.494

+ 17.320

+ 15.0

150.0

120.0

127.5

135.0

142.5

+ 24.494

+ 0.0

+ 15.0

+ 17.320

+ 15.0

Control

Delhi Road

Garh Road

210.0

157.5

+ 24.494

'THE ROLE OF BIOLOGY IN BRINGING SECOND GREEN REVOLUTION

CD 5%

CD 1%

30

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Stomatal coverage area (mm-2 )

6157.932

2521.2464*

3349.8583*

4472.3796

5545.3257 2755.393

+ 943.628

+ 187.9107

+ 665.7223

+ 849.730

6567.650

+ 561.0567

Table. 3 Stomatal response in terms of no. of epidermal cells, no. of stomata, epidermal density (mm-2), stomatal density (mm-2) and stomatal index on abaxial surface in Sida veronicaefolia plant. Different Study Sites Attribute

No. of epidermal cells

Statistical Value Railway

University

Road

Road

46.0

44.0

40.75

+ 0.5773

+ 0.816

+ 0.816

+ 1.5

20.75

12.75

13.5

15.75

18.0

+ 0.9575

+ 2.2173

+ 2.516

+ 0.9574

+ 1.4142

570.113

686.968**

651.558*

623.229*

577.195

+ 17.823

+ 8.1777

+ 11.565

+ 11.565

+ 21.246

293.909

180.594**

191.218**

223.087*

254.957

+ 13.561

+ 31.407

+ 35.646

+ 13.561

+ 20.031

34.008

20.6632**

22.6100**

26.346*

30.631

+ 0.508

+ 3.005

+ 3.642

+ 1.067

+ 2.177

Control

Delhi Road

Garh Road

40.25

48.5

+ 1.258

CD 5%

CD 1%

52.043

124.048

39.598

94.384

1.483

3.535

No. of stomata

Epidermal density (mm-2 )

Stomatal density (mm-2 )

Stomatal index

'THE ROLE OF BIOLOGY IN BRINGING SECOND GREEN REVOLUTION

31

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in

-2

Table. 4 Stomatal

) on abaxial surface

in Sida veronicaefolia plant. Different Study Sites Attribute

Length of

Breadth of

Stomatal coverage area (mm-2 )

Statistical value Railway

University

Road

Road

142.5

165.0

187.5

+ 15.0

+ 15.0

+ 17.320

+ 15.0

16.5

127.5

127.5

135.0

150.0

+ 17.320

+ 15.0

+ 28.722

+ 17.320

+ 24.494

10949.008

3271.9546*

3788.951*

5541.784*

7963.8810

+ 1759.7914

+ 814.991

+ 906.035

+ 1168.555

+ 1550.862

Control

Delhi Road

Garh Road

202.5

127.5

+ 15.0

32

CD 5%

CD 1%

5138.590

12248.148

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In vitro study of an endangered “Miracle plant”: Mandookparni (Centella asiatica (L.) Urb.) Shalini Sharma* and Y. Vimala Department of Botany, C. C. S. University, Meerut [*[email protected]] Abstract Plants have been used as treatments for thousands of years, based on experience and folk remedies and continue to draw wide attention for their role in the treatment of chronic diseases. Mandookparni (Centella asiatica (L.) Urb.) or Indian pennywort medicinal herb belongs to family Apiaceae. It is valued in Indian systems of medicines for improving memory, for treatment of nervine disorders, skin diseases and wound healing. The bioactive compounds responsible for its medicinal properties are triterpene saponins as Asiaticoside, Asiatic acid, Madecassocide etc. Rapid urbanization, degradation of plant habitat, ruthless collection of herb and other anthropological activities have markedly depleted the wild stocks of plant. It has been listed as Threatened plant species by IUCN and an endangered species. Present study reports the standardized protocol for callus induction from Centella plant and comparative account of yield of asiaticoside from this callus by treating with various growth regulators, salt and colchicine treatment. Present study reports significant amount of yield of asiaticoside from callus with that of 60 days old leaves of plants, so that overexploitation of this herb can be avoided. Key words: Mandookparni, Callus induction, Salt treatment, Colchicine treatment, Asiaticoside. Introduction: Plants have been enumerated as an efficient basis of medicine since immemorial past. Drugs based on the plants are of prime importance for several remedies in traditional and conventional medicine throughout the world and serve as a substitute for drug supply in modern medicine. Now a days world markets are turning towards plants as the source of ingredients in manufacturing health care products. Secondary metabolites obtained from the plants are found to be an important source of various phytochemicals that could be used directly or as an intermediate for the production of pharmaceuticals. Approximately 80% of the population still relies on the traditional medicine derived from the plants for health care needs 3-5. Thus the demand for herbal medicines is continuously increasing day by day due to lesser side effects in comparison to the synthetic drugs. Unfortunately, in the present day, precipitous economic development and suburbanization resulted in overexploitation and loss of valuable natural resources, including the medicinally important plants. As a result, many of the plant species are endangered or threatened with extinction leading to severe depletion of biodiversity. C. asiatica L. (Fig. 1) is one of the threatened medicinal herbs, generally endemic to Western Ghats of South India. The medicinal properties of this plant are due to secondary metabolites triterpenoidsaponins, that can be defensive substances such as phytoanticipins, antifeedant, attractants, phytoalexins and pheromones. The triterpene of C. asiatica composed of several compounds which include asiatic acid, madecassic acid, asiaticoside, madecassoside, brahmic acid, thankunoside, isothankunoside, centelloside, sceffoleoside, madsiatic acid, centic acid and centillic acid and alkaloid hydrocotylin. In addition to these bioactive components it also contains high phenolic content which is contributed by the flavonoids such as quercetin, kaempherol, catechin,

33

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in rutin, apigenin and naringin and sugars, tannins and Vellarine. Among the triterpenes, the most important biologically active compounds are the Asiatic acid, madecassic acid, asiaticoside and madecassoside.The development of plant tissue culture technology holds great promise for conservation and enhancement of valuable medicinal plants Material and Methods: (A) Materials: Plants of Centella were procured from Forest Research Institute, Dehradun. The procured plantlets of Centella were allowed to multiply in damp and shaded area of garden. Plantlets of equal size and age were selected for transfer to experimental pots for in vivo propagation. For the present study leaves and petiole of Centella asiatica were used as experimental materials. C. asiatica L. is a small stonoliferous perennial creeping aromatic herb belonging to the family Apiaceae (Umbelliferae). The leaves are 2-5 cm wide, hastate or cordate or palmately lobed or reniform, arranged in an alternate fashion in form of clusters at stem nodes having long stalk and sheathing leaf bases. The petiole is long and stipules are small in size.

Fig.1 showing C. asiatica L. (Urb.) plant

C. asiatica L. has been used for several hundred years in folk medicines. In Indian Ayurveda literature, C. asiaticais considered as one of the recognised drugs used for “Rasayana’’ purpose. In Chinese medicine, C. asiaticais used for treatment of vomiting, epistaxis, urinary calculi, scabies and jaundice. In homeopathic medicine, it is used for treating ascariasis, elephantiasis and in granular cervicites. Clinical tests have formulated several benefits of C. asiatica extracts in terms of wound healing, burns and in skin diseases in gastrointestinal disorders and in treatment of leprosy, lupus, scleroderma, eczema, veins diseases. It is also used for treatment of psoriasis. It gives protection against diseases by enhancing immunity of the body. The extract of the whole plant is reported to have anticancerous activity and the methanolic extract of aerial parts of C. asiatica inhibit the growth of human uterine carcinoma, human gastric carcinoma, and murine melanoma cells in vitro . C. asiatica is also used as nervine tonic, along with antibacterial, antifeedant and antileptic property. It is also efficient in promoting fast growth of skin and keratinization. It also possesses anti-inflammatory and memory enhancing property. It also finds application in controlling anxiety and thereby imparting mental calmness. Wound healing, memory enhancement, neuroprotective, immunomodulatory, hepatoprotective, cardiovascular, antidepressant and anticancer.

METHODS: STANDARDIZATION OF CULTURE ESTABLISHMENT MEDIUM:

34

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Chemicals: All the chemicals used for preparation of basal medium as well as for biochemical analyses, were procured from Qualigens Pvt. Ltd., Mumbai, India and vitamins and plant growth regulators from Merck India limited and Sigma Chemicals Preparation of culture media: Standard medium [6] was used as basal medium throughout the present tissue culture studies. Young leaves and petioles of Centella were surface sterilized with 0.1% HgCl2 and 70% alcohol before inoculation.

Explant: In Centella young leaves and their petioles were used as explant. The explants were inoculated for establishing callus culture. The callus induced from different explants was maintained on same medium for minimum of 2-3 passages and was used for subsequent experiments. The undifferentiated stock calli were routinely subcultured at an interval of every 4 weeks for subsequent experimentation.

Incubation of cultures: The cultures were incubated in culture room and provided with light of 1400 lux intensity with a photo period of 16/8 h light/ dark cycles at 25 2oC temperature maintained by automated photoperiod controlled device and air conditioner or room heater.

Standardization of basal medium: For culture initiation MS medium supplemented with growth regulators was studied for culture establishment. BAP, Kn individually or in combination with NAA and IBA along with 30 g/l sucrose and 8.0 g/l agar – agar were used.

Sub culture procedure : The calli were regularly transferred every 3-4

weeks in their exponential phase of

growth on the fresh media. In subsequent passages, the non-morphogenic callus was transferred on auxin or cytokinin alone or in combination of both, with three salt treatments (50, 75 & 100 mM NaCl) and without salt added media (as control). Actively dividing cells of calli were treated with Colchicine (Ctrl and 0.4%) in liquid medium for 24 hours. Then these were analyzed with their respective controls for growth and type of callus after fourth week as treated calli showed various growths under different hormonal treatments, salts as well as colchicine treatment. PHYSICOCHEMICAL ANALYSIS:  Weight analysis (Fresh weight, Dry weight) and Moisture content- Moisture percent was calculated according to the following formula: -

 Growth Index: Growth index of callus was calculated by the following formula:-

 Determination of Total Proteins [2], Reducing, Non-reducing and Total sugar [8], Proline content [1] and Total Phenolic content [9] was carried out by standard methods mentioned in parentheses.

35

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in  Extraction Procedure for Asiaticoside and its estimation: Each of the dried 1.0 g samples was dried and soxhlet extracted in Methanol for 24 hours. Each of the Methanol extracts of various test samples was separately dried in vacuo [4]and taken up in methanol for further analysis of Asiaticoside estimation. Further quantitative estimation was done by HPLC. (Kind courtesy of Natural Remedies Private Ltd., Bangalore). Every estimation was done in three extractions done in triplicates and subjected to statistical analysis in terms of mean and standard deviation for testing significance of data. RESULTS AND DISCUSSION: As explants, leaves were selected for callus initiation because of the superiority of callus from the same, over callus obtained from petioles. Callus growth index was best in medium supplemented with combinations of auxins and cytokinins, instead of when they were used alone. BAP (2.0mg/l)+ NAA (0.1mg/l) supplementation to MS medium using 7 days old leaves as explant produced superior callus in C.asiatica, that was used for further studies. Callus Growth index under 50 to 100 mM salt and 0.5 to 2.0 mg/l concentrations of various Auxin (Fig.1-a,b,c), Cytokinins (Fig.2-a,b) and their combinations (Auxin+ Cytokinin) supplemented media (Fig.3)

Growth Index

1.5

Fig-1(a) Effect of NAA and Salt on four weeks old callus of C. asiatica

1 0.5 0 DW

50mM

0.5 NAA

Growth Index

1

75mM 1.0 NAA

100mM 2.0 NAA

Fig-1 (b) Effect of 2,4-D and Salt on four weeks old callus of C. asiatica

0.5

0

DW 0.5 2,4-D

50mM

75mM

1.0 2,4-D

100mM 2.0 2,4-D

36

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Fig-1 (c) Effect of IAA and Salt on four weeks old callus of C. asiatica Growth Index

1.5 1 0.5 0 DW

50mM

0.5 IAA

100mM

1.0 IAA

2.0 IAA

Fig-2 (a) Effect of BAP and Salt on four weeks old callus of C. asiatica

1.5 Growth Index

75mM

1 0.5 0 DW

50mM

0.5 BAP

1.0 BAP

100mM 2.0 BAP

Fig-2 (b) Effect of Kinetin and Salt on four weeks old callus of C. asiatica

1.5 Growth Index

75mM

1

0.5 0

DW

50mM

75mM

0.5 Kn

Growth Index

2

100mM

1.0 Kn

2.0 Kn

Fig-3 Effect of Kinetin and Salt on four weeks old callus of C. asiatica

1

0 DW 0.5 2,4D+ 0.5Kn

50mM 1.0 2,4D+ 0.5Kn

75mM 2.0 2,4D+ 0.5Kn

100mM 2 BAP+ 0.1NAA

High protein content in callus of C.asiatica indicates its good nutritional value that is much higher than 60 days old leaves grown in vivo. Callus is able to maintain good growth index even in salt treatment by accumulating Osmoprotectants as proline as well as phenolics in it as shown in Table- 1. With Salt stress Proline content significantly increased in callus in comparison to control (DW). Phenolic content also significantly increased with salinity stress as well as in colchicines treated callus. This indicates that callus has significant antioxidant activity which contributes towards its medicinal and nutritional value for human beings, yet not as good as found in in vivo grown 60 days old leaves. Dietary antioxidant supplementation is a promising mean to strengthen the antioxidant defense and repair systems [5]. However, antioxidants from natural sources are of

37

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in great

value

as

most

commonly

used

synthetic

antioxidants

(e.g.

butylatedhydoxyanisole,

butylatedhydoxytoluene and propylgallate) have health hazardous side effects like liver damage and carcinogenesis. Several reasons may explain the differences in concentration of bioactive compounds; genetic diversity, environmental conditions, and nutrients present in the soil/culture medium may alter the metabolic activity of plants [7]. Alternatively, climatic conditions and bioavailability of nutrients, may alter the growth rate of explants, and thereby plant metabolic activity. Table:1 S. No.

Media for

Proline (mg

Phenolics ( mg trans

Protein (mg

Proline/ g f wt)

cinnamic acid eq g/ f

casein eq /g fwt)

growth 1. 2BAP+0.1NAA

wt) 0.017

1.17

20.98

0.036

1.466

21.46

0.026

1.254

9.19

0.025

0.868

12.96

0.007

1.31

10.49

0.004

4.64

16.36

(DW) 2. 2BAP+0.1NAA (50Mm) 3. 2BAP+0.1NAA (75mM) 4. 2BAP+0.1NAA (100mM) 5. 0.4% Colchicines treated callus 6. Soil grown 60 days old leaves

With increasing salt stress callus accumulated osmoprotectants (Table -2) that is apparent in the form of Total sugar content accumulated in callus. Non reducing Sugars also accumulated in callus in 100mM NaCl treatment in comparison to control that signifies increased level of triterpenoid saponins that are responsible for medicinal properties of C. asiatica plant. This fact is confirmed by the Asiaticoside content measured by HPLC, where 100mM NaCl treated callus accumulated, yet remained lower in comparison to in vivo grown 60 days old leaves. In comparison to salt treated calli, Colchicine treatment of callus for Asiaticoside content proved to be beneficial as it showed accumulation of sugars as well as asiaticoside content equitable with soil grown 60 days old leaves (in vivo). This may be due to the stress experienced by the callus and tolerance response exhibited. Climatic conditions, bioavailability of nutrients, may alter the growth rate of explants, and thereby plant metabolic activity [3].

Table-2

S.No.

TS Media used

RS

NRS

(mg glucose eq g/ dw ± SD)

38

Asiaticoside %

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 1

2BAP+0.1NAA (DW) treated callus

0.19

0.155

0.035

0.0045

2

2BAP+0.1NAA (50Mm) treated callus

0.144

0.122

0.022

-

3

2BAP+0.1NAA (75mM) treated callus

0.154

0.131

0.023

-

2BAP+0.1NAA (100mM) treated callus

0.237

0.186

0.051

0.07

4

0.4% Colchicines treated callus

2.05

1.114

0.91

0.107

5

Soil grown 60 days old leaves (in vivo)

0.837

0.031

0.806

0.118

CONCLUSION: Thus, it can be stated that research on plants have been enthralled throughout the world to emblematize the tremendous potential of medicinal plants in recent years. Due to its wide prospects and potential, its demand has led to a quantum increase which plays a vital role in alleviating human sufferings due to lesser side effects, easy availability at affordable cost and being non-narcotic. Sometimes, it is the only source of health care available to the poor. C. asiatica L. (Urb.) is one such medicinally important herb with well known biological activities in terms of immunomodulatory, memory enhancer, antidepressant, etc., proved by clinical studies. Thus, present in vitro studies of C. asiatica provide a promising strategy to conserve the green cover of Mandookparni, without overexploiting it and providing callus with medicinal, antioxidant activity, nutritional value as good as given by soil grown 60 days old leaves. However, further research on C. asiatica must be explicit in terms of exploring its immense potential as nutraceutical. In addition to this, studies should be premeditated regarding the investigation of underutilized green leafy vegetables, substantial to permeate nutritional ailments. ACKNOWLEDGEMENT: Authors are thankful to Late Professor C.M. Govil. and Prof A. K. Srivastava for their valuable advice. One of the authors (Shalini Sharma) extends her gratitude to C.S.I.R. for the award of Senior Research Fellowship.

REFERENCES 1.

Bates, C. A, Waldern, R. P and Taleve, I. D. Rapid determination of free Proline or water stress studies. Plant Soil.39: 205-207(1973).

2.

Bradford, M. M A rapid and sensitive method for quantitation of microgram quantities of protein utilizing the principles of dye binding. Annal.Biochem.72 : 248-254(1976).

3.

F. Bourgaud, A. Gravot, S. Milesi, E. Gontier, Production of plant secondary metabolites: a historical perspective :Plant Sci. 161: 825-1043(2001).

4.

F.

Gafner,

J.D.

Msonthi,

K.

Hostettmann,

Helv.

Chim.

Molluscicidalsaponins

from

Talinumtennuisimum: Acta 68: 555–558(1985). 5.

Mijanur Rahman, Shahdat Hossain, AsiqurRahaman, Nusrat Fatima, TaslimaNahar, Borhan Uddin and Mafroz Ahmed Basunia Antioxidant Activity of Centella asiatica(Lin.) Urban: Impact of Extraction Solvent PolarityJournal of Pharmacognosy and Phytochemistry, 1(6): 27(2013).

6.

Murashige, T. and Skoog, F. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol. Plantarum15: 473-497(1962).

39

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 7.

MuthusamyGovarthanan,

RathikaRajinikanth,

Seralathan

Kamala-Kannan

and

ThangasamySelvankumarA comparative study on bioactive constituents between wild and in vitro propagated Centellaasiatica:Journal of Genetic Engineering and Biotechnology 13: 25–29(2015). 8.

Nelson, N. A photometric adaptation of Somogyi method for the determination of glucose. J. Biochem.153: 375-380(1944).

9.

Sadasivam, S. and Manickam, A. Phenolics in: Biochemical method for Agricultural Sciences. Wiley Eastern Limited. New Delhi (India). 187-189(1992).

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Role of Nanotechnology in Pollution Control: Review Shalini Singh Department of Zoology, M.K.R. Govt. Degree College, Ghaziabad [email protected] Human activity and industrialization leads to the environment filled with different kinds of pollutants. Air is filled with cabonmonooxide (CO), Cholofluorocarbons (CFC), heavy metals (lead, arsenic, chromium, cadmium, mercury, zinc), hydrocarbons, nitrogen oxide, sulphur oxide and particulates. Water pollution is caused by numerous factors including sewage, oils spills, leaking of fertilizers, herbicides and pesticides from land etc. Contaminants are most often measured in parts per million (ppm) or parts per billion (ppb) and their toxicity defined by a ‘toxic level’. The toxic level for arsenic, for instance is 10 ppm in soil whereas for mercury is 0.002 ppm in water. Therefore, very low concentrations of a specific contaminant can be toxic. There is need for technologies are capable of monitoring recognizing and ideally treating such small amount of contaminants in air, water and soil. Environmental nanotechnology is considered to play a key role in the shaping of current environmental engineering and science. The nanotechnological applications and products can lead to a cleaner and healthier environment [1]. Maintaining and re-improving the quality of water, air and soil, so that the Earth will be able to support human and other life sustainably, are one of the great challenges of our time. Nanotechnology can play a vital role in providing clean air, water and soil in an efficient and cheap way [2]. Nanoscience allows designing and manipulating materials at the atomic and molecular level. Nanomaterials can be fabricated with specific properties that can recognize a particular pollutant within a mixture. The small size of nonmaterials together with their high surface to volume ratio can lead to very sensitive detection. These properties will allow developing highly miniaturize, accurate and sensitive pollutionmonitoring devices (‘nano-sensors’). Nanomaterials can also be engineered to actively interact with a pollutant and decompose it in less toxic species. Thus, in the future nanotechnology could be used not only for detecting contaminated sites but also treating them.

POLLUTION DETECTION AND SENSING Fortification of the human health and protection of the environment requires the rapid, sensitive detection of pollutants and pathogens with molecular precision. Sensors are needed for in situ detection, as miniaturized portable devices, and as remote sensors, for the real-time monitoring of large areas in the field. A sensor is a device built to detect a specific biological or chemical compound, usually producing a digital electronic signal upon detection. Sensors are now used for the identification of toxic chemical compounds at ultra low levels (ppm and ppb) in industrial products, chemical substances, water, air and soil samples, or in biological systems. Nanotechnology can improve current sensing technology in various ways. First, by using nanomaterials with specific chemical and biological properties, the sensor selectivity can be improved, thus allowing isolating a specific chemical or biological compound with little interference. Hence, the accuracy of the sensors is improved. As with other nano-engineered products discussed in this document, the high surfaceto-volume ratio of nanomaterials increases the surface area available for detection, which in turn has a positive effect on the limit of detection of the sensor, therefore improving the sensitivity of the device. Scaling down

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in using nanomaterials allows packing more detection sites in the same device, thus allowing the detection of multiple analytes. This scaling-down capability, together with the high specificity of the detection sites obtainable using nanotechnology, will allow the fabrication of super-small ‘multiplex’ sensors, this way lowering the cost of the analysis and reduce the number of devices needed to perform the analysis with an economic benefit. Advancements in the field of nanoelectronics will also allow the fabrication of nanosensors capable of continuous, real time monitoring [3]. Various nanostructured materials have been explored for their use in sensors for the detection of different compounds [4]. An example is silver nanoparticle array membranes that can be used as flow-through Raman scattering sensors for water quality monitoring [5]. The particular properties of carbon nanotubes (CNTs) make them very attractive for the fabrication of nanoscale chemical sensors and especially for electrochemical sensors [6-9]. A majority of sensors described so far use CNTs as a building block. Upon exposure to gases such as NO2, NH3 or O3, the electrical resistance of CNTs changes dramatically, induced by charge transfer with the gas molecules or due to physical adsorption [10, 11]. The possibility of a bottom-up approach makes the fabrication compatible with silicon microfabrication processes. The sensor is made of an array of electrode pairs fabricated on a silicon chip and separated by few nanometres. When the electrodes are exposed to a solution of water containing metal ions, these deposit inside the nano-gap in between the electrodes. Once the deposited metal bridges the gap a ‘jump’ in conductance between the electrodes is registered. The size of the gap, being only few nanometres, allows the detection of a very low concentration of metal ions. This type of sensor is called ‘nanocontact sensor’. [12]. The connection of CNTs with enzymes establishes a fast electron transfer from the active site of the enzyme through the CNT to an electrode, in many cases enhancing the electrochemical activity of the biomolecules [8]. In order to take advantage of the properties of CNTs, they need to be properly functionalized and immobilized. CNT sensors have been developed for glucose, ethanol, sulfide and sequence-specific DNA analysis [8]. Trace analysis of organic compounds, e.g. for the drug fluphenazine, has also been reported [13]. Nanoimmunomagnetic labeling using magnetic nanoparticles coated with antibodies specific to a target bacterium have been shown to be useful for the rapid detection of bacteria in complex matrices [14]. Materials that are more environment-friendly fabricated using nanotechnology include biodegradable elf-cleaning glasses, such as Activ™ Glass [15], the glass is composed of a special coating made of nanocrystals of TiO2 which, once exposed to daylight, reacts in two ways. First, it breaks down any organic dirt deposits on the glass and second, when exposed to water, it allows rain to 'sheet' down the glass easily and washes the loosened dirt away.

NANOCATALSIS A catalyst is a substance that increases a chemical reaction rate without being consumed or chemically altered. One of the most important properties of a catalyst is its ‘active surface’ where the reaction takes place. The ‘active surface’ increases when the size of the catalysts is decreased . The higher is the catalysts active surface, the greater is the reaction efficiency. Also, research has shown that the spatial organization of the active sites in a catalyst is important as well [16]. Both properties (nanoparticle size and molecular structure/distribution) can be controlled using nanotechnology. In the environmental field, nanocatalysis is being investigated for desulphurizing fuels, with the aim of developing ‘clean’ fuels containing very low sulphur products (produced in the fuel during its refining process and responsible for generating sulphuric acid upon fuel

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in combustion). Another area where nanotechnology is making a contribution is the development of fertilizers and wood treatment products that are more stable and leach less into the environment. For instance, researchers at the Michigan State University have incorporated biocides for wood treatment inside polymeric nanoparticles. Their small size allows them to efficiently travel inside the very fine, sieve-like structure of wood. At the same time, the biocide, being safely trapped inside a ‘nanoshell’, is protected from leach and random degradative processes [17].

GREEN MANUFACTURING Manufacturing processes are always accompanied by the production of diverse waste products, many of which pose a threat to the environment and thus need to be removed and treated. Green manufacturing includes the development of new chemical and industrial procedures (for instance water-based rather the solvent-based processes); reduction in the use of unsafe compounds (such as metals); development of ‘green’ chemicals that are more environment-compatible; and efficient use of energy. In terms of its application to the reduction of manufacturing waste, nanotechnology can contribute in two ways: by directing the manufacturing to be more controlled and efficient, and by using nanomaterials (such as catalysts) that can raise the manufacturing efficiency while reducing or eliminating the use of toxic materials. Overall, nanotechnology has the potential of making industrial processes more efficient in terms of energy usage and material usage, while minimizing the production of toxic wastes. The application of ‘green nanotechnology [18] to manufacturing includes bottom-up, atomic-level synthesis for developing improved catalysts; inserting information into molecules to build new materials (such as DNA) through highly specific synthetic routes; scaling down material usage during chemical reaction by using nanoscale reactors; and improving manufacturing to require less energy and less toxic materials. An example of ‘green nanotechnology’ is the development of aqueous-based microemulsions to be used in alternative to volatile organic compounds (VOCs) in the cleaning industry. These toxic and potentially carcinogenic compounds, such as chloroform, hexane, percholoroethylene, are conventionally used in the cleaning industry (like the textile industry) as well as in the oil extraction industry. Microemulsions contain nano-sized aggregates that can be used as ‘receptors’ for extracting specific molecules at a nanoscale level. Other example is microemulsions having water-attractive and water-repellent “linkers” inserted between the head and tail parts of a surfactant molecule [19]. The result is a surfactant that has a very low interfacial tension with a wide range of oils. When tested for cleaning textiles from motor oil residues, as well as for extracting edible oil from oilseeds, the microemulsions were found to be very competitive with conventionally used VOCs, both in terms of extraction yield and simplicity of the process. Nanowires of semiconductors such as silicon has established knowledge for the chemical modification of their surface. Boron-doped silicon nanowires (SiNWs) have been used for the sensitive real-time electrical detection of proteins, antibodies the metabolic indicator calcium[20, 21] and glucose in water [22]. The small size and the capability of these semiconductor nanowires to detect in real-time a wide range of analytes could be used for developing sensors for detecting pathogens, chemical and biological agents in water, air and food. Nanotechnology’s potential and promise have steadily been growing throughout the years. The world is quickly accepting and adapting to this new addition to the scientific toolbox. Although there are many obstacles

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in to overcome in implementing this technology for pollution control, science is constantly refining, developing, and making breakthroughs. REFERENCES 1. Masciangioli T. and Zhang W. X. Environmental technologies at the nanoscale, Environ. Sci. Technol. 37(5), 102A-108A (2003) 2. Hillie T., Munasinghe M., Hlope M. and Deraniyagala Y. Nanotechnology, Water and Development, Meridian Institute (2006) 3. Report from Applications of Nanotechnology: Environment-Luisa Filipponi & Duncan Sutherland, Nanocap (2007) 4. Vaseashta A., Vaclavikova M., Vaseashta S., Gallios G., Roy P. and Pummakarnchana O. Nanostructures in environmental pollution detection, monitoring, and remediation, Sci. Technol. Adv. Mater. 8 (1), 47-59 (2007) 5. Taurozzi, J. S. and Tarabara, V. V. Silver nanoparticle arrays on track etch membrane support as flowthrough optical sensors for water quality control, Environ. Eng. Sci. 24 (1), 122-137 (2007) 6. Wang J. Carbon-Nanotube Based Electrochemical Biosensors: A Review, Electroanalysis 17 (1), 7-14 (2005) 7. Trojanowicz M. Analytical applications of carbon nanotubes: A review, Trends Anal. Chem. 25 (5), 480-489 (2006) 8. Valcarcel M. Simonet B.M., Cardenas S. and Suarez B. Present and future applications of carbon nanotubes to analytical science, Anal. Bioanal. Chem. 382 (8), 1783-1790 (2005) 9. Merkoci A. Carbon Nanotubes in Analytical Sciences, Microchim. Acta 152 (3), 157-174(2006) 10. Dai L., Soundarrajan P. and Kim T. Sensors and sensor arrays based on conjugated polymers and carbon nanotubes, Pure Appl. Chem. 74 (9), 1753-1772 (2002) 11. Sano N. and Ohtsuki F. Carbon nanohorn sensor to detect ozone in water, J. Electrostat. 65 (4), 263-268 (2007) 12. Li J., Koehne J.E., Cassell A.M., Chen H., Ng H.T., Ye Q., Fan W, Han J. and Meyyappan M. Inlaid Multi-Walled Carbon Nanotube Nanoelectrode Arrays for Electroanalysis, Electroanalysis, 17(1), 15-27 (2005) 13. Zeng B.Z. and Huang F. Electrochemical behavior and determination of fluphenazine at multi-walled carbon nanotubes/(3-mercaptopropyl) trimethoxysilane bilayer modified gold electrodes,

Talanta 64 (2), 380-386

(2004) 14. Chang S. C. and Adriaens P. Nano-immunodetection and quantification of mycobacteria in metalworking fluids, Environ. Eng. Sci. 24 (1), 58-72 (2007) 15. Activ™ Glass, Pilkington, www.pilkington.com 16. Gemming S. and Seifert G. Catalysts on the edge, Nature 2, 21-22 (2007) 17. Liu Y., Yan L., Heiden P. and Laks P. Use of nanoparticles for controlled release of biocides in solid wood’, J. Appl. Poly. Sci. 79 (3), 458-465 (2001)

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 18.For a review on ‘Green nanotechnology’ covering definition, concepts and applications see K. Schmidt, Green Nanotechnology (PEN8), Woodrow Wilson Center International Center for Scholars, free to download from www.nanotechproject.org/reports . 19. Acosta E.J. , Nguyen T. , Witthayapanyanon A. , Harwell J. H., and Sabatini D.A. Linker-based biocompatible microemulsions, Environ. Sci.Tecnol. 39 (5), 1275-1282 (2005) 20. Cui Y., Park H. and Lieber C.M. Nanowire nanosensors for highly sensitive and selective detection of biological and chemical species, Science 293 (5533), 1289-1292 (2001) 21.Patolsky F. and LieberC.M. Nanowire nanosensors, Materials Today 8 (5) 20-28(2005) 22 Shao M., Shan Y, Wong N. and Lee S. Silicon nanowire sensors for bioanalytical applications: glucose and hydrogen peroxide detection, Adv. Func. Mater. 15 (9), 1478-1482 (2005)

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Identification of Geminivirus in Cowpea (Vigna unguiculata) Through Amplification of Selective DNA Fragment Using Degenerate Primers Shail Pande Mahatma Gandhi P G College, Gorakhpur ABSTRACT Mostly Gemini viruses have been characterized by symptomatology and host range tests,reliance on symptoms only; may impede diagnosis, as symptoms are affected by several variables such as virus strain, the growth stage of the plant at the time of infection, the plant cultivar and environment conditions.The particles of gemini viruses occur in plants in only low to moderate concentrations, which can be inadequate for detection by the conventional serological tests but are adequate for some of the newer, more sensitive methods like polymerase chain reaction (PCR).DNA probes can be valuable especially those derived from the DNA-2 of bipartite genomes they are used to detect virus in homologous sequences.PCR is especially useful for detecting gemini viruses, because of its sensitivity to the viral template in low titers. In addition, taxonomically informati ve domains may be amplified by PCR. Annealing temperature, type and concentration of PCR additives and MgCl 2 concentration were first adjusted to improve sensitivity and reproducibility. DNA concentration required for amplification was assessed by detection of the initial and end points of dilution and the intensity of the amplified bands. KEYWORDS- Geminivirus, degenerate primer, PCR, DNA dilutions. INTRODUCTION Historically, Gemini viruses have been characterized by symptomatology and host range tests. A reliance on symptom development may impede diagnosis, as symptoms are affected by several variables such as virus strain, the growth stage of the plant at the time of infection, the plant cultivar and environment conditions [1]. In general, the particles of gemini viruses occur in plants in only low to moderate concentrations, which can be inadequate for detection by the conventional serological tests but are adequate for some of the newer, more sensitive methods for e.g. Immunosorbent electron microscopy is possible, although it is not straight forward because the virus particles are disrupted or damaged in some buffers and they show relatively poor contrast in most electron dense strains. DNA probes can be valuable especially those derived from the DNA-2 of bipartite genomes they are used to detect virus in homologous sequences [2]. Among several diagnostic tools available, the polymerase chain reaction (PCR) using degenerate primers has been the most useful for the detection of Gemini viruses. PCR is especially useful for detecting Gemini viruses, because of its sensitivity to the viral template in low titers. In addition, taxonomically informative domains may be amplified by PCR and used in identification and phylogenetic studies [3-4]. MATERIALS & METHODS Experimental Strategies for PCR The isolation of DNA from plant is often a limiting factor; this is because of the special nature of plant tissue and cells. A tough cell-wall, abundant secondary metabolites, presence of several classes of chemical compounds with varying properties are the feature unique to plant cells and these have led to the development of several plant specific DNA isolation procedures. Once the DNA is isolated it can be subjected to PCR analysis.

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in The PCR in general is a technique requiring several rigorous conditions. Thus the experimental strategy for carrying out studies in case of plants using PCR-based approach is mainly influenced by the PCR-conditions. As a technique, though PCR is simple, it is also influenced by a number of factors, these include. (i)

Commercial and biological source and quality of the thermostable polymerase.

(ii)

Length and base composition of the primers.

(iii)

Concentration of the template and primer.

(iv)

Concentration of Mg++ions.

(v)

Presence or absence of K+ ions.

(vi)

Stringency of Primer annealing conditions.

(vii)

Thermocycler parameters such as ramping and its duration.

(viii)

Cycle parameters such as number and duration of cycles.

(ix)

Cleanliness of work, especially the presence or absence of other contaminating DNAs. Several workers have described the role of above parameters and have also detailed the optimization of

the experimental protocols [5-6]. After the PCR has been carried out, the amplification product can be analysed by gel electrophoresis on 1.4% agarose gel stained with ethidium bromide and visualized by UV lamp. The patterns of bands revealed are used for comparison amongst healthy and diseased plants. This technology is proving to be invaluable in pathogen analysis, especially in case of viruses. This technology is being used for detection of pathogens present in low concentration. Reaction setup for PCR amplification 

DNA isolated was checked on 0.8% agarose get by electrophoresis for quality and yield.



Isolated DNA was given RNAase treatment to remove RNA by treating it with RNAase (100mg/ml stock - Solution of which 2l was used in 5l of DNA) at 370C for 1 hour.



Yield was again checked on 0.8% agarose by electrophoresis.



Optimization of the PCR amplification was done in order to maximize sensitively and reproducibility of detection of virus templates in DNA from symptomatic leaves using degenerate primers. Annealing temperature, type and concentration of PCR additives and concentration of PCR additives and MgCl2 concentration were first adjusted to improve sensitivity and reproducibility sensitivity was assessed by detection of the initial and end points of dilution and the intensity of the amplified bands.

Table-1: Detection of initial and end points of dilution and respective amplified band intensity

Replicates

++++

Dilutions 30ng

20ng

10ng

5ng

1

+++

++

+

-

2

++++

++

+

-

3

+++

+++

+

-

4

+++

++

+

-

-

Very intense amplified band

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in +++

-

Intense amplified band

++

-

Visible amplified band

+

-

Barely visible band

When DNA was used at concentration above 30ng/l, shearing in bands were seen. From this table it is evident that 30ng/l of DNA is most suitable for the PCR so we used 30ng/l concentration of DNA for PCR. DNA was diluted to get 30ng/l of concentration and following PCR reaction was set. dNTPs

- 1.0l

Taq DNA Polymerase

- 0.5l

10X Buffer

- 2.5l

Primer A (Forward)

- 1.0l (~ 25ng/l)

Primer B (Reverse)

- 1.0l (~ 25ng/l)

DNA from diseased and healthy leaf

- 1.0 l

The rest of the volume (18l) was made up by water up to 25l. This reaction mixture was put in DNA engine (PCR machine) for the following cycles. 940C

-

5 Minute

0

94 C

-

1 Minute

0

52 C

-

1 Minute

720C

-

1 Minute

0

72 C

-

5 Minute

40C

-

for ever

One Cycle

35 times

When reaction cycle is completed then amplified product was checked in 1.2% agarose gel by taking 10l of PCR product with 2l of dye, bromophenol blue. 

This PCR product is run along the marker -DNA ECQRI and Hind-III double digest to determine the size or base pairs of bands.



For control, DNA from the healthy leaf is also given the same reaction mixture for PCR amplification and run on the same gel.



The PCR amplification product were resolved in 1.2% agarose gel stained with ethidium bromide and screened on UV transilluminator Gel documentation system for analysis was used to take photograph of gel for analysis.

Degenerate primer pairs [7] is used for DNA amplification. The two degenerate oligonucleotide primers had the following sequences. Primer

PA - 5' TAA TAT TAC CGG AGG AGG CCC CC3' PB - 5' TGG ACC TAA CAA GGG CCT TCA CA 3'

These degenerate primers permit detection of sub-group-III Gemini viruses by annealing to sequences that flank the conserved core region of the coat protein gene, yielding a diagnostic about 565bp virus fragment [8]. This means primer will bind to the DNA of virus only and amplification of that particular DNA will be done by PCR.

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Plate-I of gel shows marker in first and last lane. Second and third lane represents control plant. Fourth, fifth and sixth lane contain amplification product of diseased leaf. As it is evident from the photograph of gel that the lane containing amplification product of diseased plant are showing a specific band of about 500 bp, no such band is seen in control lane making it clear that the virus under study belongs to family Gemini viridae and subgroup-III genus begomovirus species cowpea golden mosaic virus.

Figure: 1 First and last lane- Molecular weight markers, second and third lane- control plants, fourth fifth and sixth row- diseased plant with 500bp band. DISCUSSION : For identification of gemini viruses, serology has never been a preferred tool for the simple reason that the particles are difficult to purify, making it hard to produce good antisera. On the other hand begomo virus is the close serological relationship due to conservation in coat protein gene [9], as a result polyclonal antibody raised against one begomovirus could detect all the begomoviruses in double antibody sandwich ELISH or ISEM. Because of the difficulty in purifying begomoviruses virion particles, nucleic acid based approaches like PCR are being widely preferred for the diagnosis. Advent of PCR technique revolutionized research on begomoviruses. Full length genome or specific regions in the viral genome are amplified with degenerate primers or specific primers. Diagnostic PCR should fulfill several qualitative characteristics, of which the most important are specificity, sensitivity, efficiency and reproducibility. Designed degenerate primers flank the 5' terminal region of the coat protein gene to specifically detect begomoviruses and when tested with degenerate primers it showed the specific band in the diseased plant DNA corresponding to the coat protein gene of begomovirus confirming the presence of begomovirus in diseased plants. REFERENCE 1.

1.Goodman, R.M. Geminivirus. In Hand book of Plant virus Infections and comparative Diagnosis, ed. E.Kurstak Amsterdem : Elsevier / North-Holland Biomed. 879-910,(1981).

2.

Harrison, B.D. Advances in Geminivirus research, Ann. Rev. Phytopathol., 23, 55-82. (1985).

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 3.

Fauquet, G.M., Stanley, J. Geminivirus classification and nomenclature-progress and problems, Ann Appl. Biol., 142, 165-189 (2003).

4.

Malathi, V.G., Usharani, J.S., Sivalingam, P.N., Rouhibaksh, A., Padma Latha, K.V. and Periasamy, M. Diversity and complexity of Begomoviruses, Annu. Rev. Plant Pathal., 3, 225-270 (2004).

5.

Samec, P. DNA Polymorphism and RAPD technology, Priloha Casopisu Genetika a Slechteni 29 , 291230. (1993).

6.

Williams, J.G.K., Manafey M.K., Rafalkl J.A. and Tingey S.V. Genetic analysis using random amplified polymorphic DNA markers, Methods in Enzymology, 218, 704-740 (1993).

7.

Deng D., McGrath P.F., Robinson D.J., Harrison B.D. Detction and differentiation of whitefly transmitted geminiviruses in plant and vector insect by the polymerase chain reaction with degenerate primers, Ann. Appl. Biol., 125 , 327 - 336. (1994).

8.

Wyatt S D & Brown J K. Detection of subgroupIII Geminivirus isolates in leaf extracts by degenerate primers and polymerase chain reaction, Phytopathology ,86(12) ,1288-1293(1996).

9.

Harrison, B.D. and Robinson, D.J. Natural genomic and antigenic variation in whitefly-transmitted geminiviruses, Ann. Rev. Phytopathal., 37, 369-398 (1999).

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Science and Technology in Rural India Richa Atreya Department. of Botany, M.S. College, Saharanpur (India) e-mail address - [email protected] ABSTRACT The importance for science and technology for rural India was appreciated by eminent leaders and scientists in 1930s giving rise to the centre for science for villages and advanced institutions for education. However post independence steps were biased towards urban areas and science & technology turned their attention for rural areas about 40 years later in 1970s. The most well known step was from Indian Institute of Sciences by its program for the application of science and technology to rural areas also called by its acronym “ASTRA”. Present scenario poses promising future of S&T in rural areas with many governmental and nongovernmental organizations participating in process of its development. Ministry of human resource development has started Rashtriya Avishkar Abhiyan that works in conduit with Sarva Shiksha abhiyan, Madhyamik Shiksha Abhiyan and also promotes study of S&T in higher education among rural children. Likelihood for science education can be fostered among rural Indians by making it affordable, technologically assisted, developing rural infrastructure, and creating awareness through rigorous campaigns and fairs. Our present National Education Policy shall be revised for promotion of science and innovations in India. KEYWORDS - S&T, infrastructure, ASTRA, biomethanation, biogasification, STEM. INTRODUCTION Many pioneer leaders like Bal Gangadhar Tilak, Raja Ram Mohan Roy, Swami Dyanand Saraswati along with eminent Indian scientists like J.C. Bose, P.C. Roy and C.V. Raman realized the need of acquaintances with science and technology (S&T) for common Indians including rural people. They envisioned the importance of science aptitude among people for nation building. But post independence era had an upthrust for industrial development and ideas of mainstream scientific and technological development were marginalized. Research and development got a mention in Indian budgets, though R&D budget in1957-58 was mere 18.81 crores. Indian economy is an agro economy with rural areas as its main centers and therefore it is indispensible to promote science education in rural areas and harnessing S&T for them. Problems of lack of easy access, lack of interest, common curricula, gender differentiation, and most important lack of infrastructure in rural areas is holding back promotion of S&T. New & better government policies shall be formulated for rural areas. Yet public private partnership is paving way for better rural India and now we see rural areas that are tech-savvy and have more acceptances for science and innovations. POST INDEPENDENCE S &T IN RURAL AREAS The post independence establishment was preeminently dominated by scientists and engineers who returned from Europe and North America and they were prolifically influenced by their studies and sojourns abroad. They were more open to industrialization and technology and at that time demand of Indian industry and government were the determining force for development. There can be meticulous mention of post

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in independence growth of S&T in India as the R&D budget increased; a number of science and technology institutions like DRDO, IARI, ISRO etc. established; an enormous increase in technically trained personnel; international collaboration for S&T was also achieved. Rural areas faced biased attitude- By and large all scientific development shifted to urban needs and rural areas were unnoticed. Conclusions for such polar development of S&T can be drawn from anti rural bias in R&D expenditure  Establishment of institutions  Distribution of technical personnel  Development of infrastructure  Government policies and future plans Even five years plans of India addressed primary education, health facilities, sanitation, agriculture and irrigation in these areas and to add setbacks to these initiatives many of five year plans collapsed due to natural and political upheaval in the country. So there was no major breakthrough in creating scientific awareness and harnessing S&T for rural people. Later in 1970’s when Indian institute of sciences presented a program called ASTRA for use of S&T for rural people, there arose hopes of creating scientific aptitude among rural Indians [1]. ASTRA – an institutional experiment was against above mentioned backdrops that there arose a need to reorient Indian science and technology towards need for rural India. In 1970 Indian Institute of Sciences Bangalore made a presentation and translated it into a program called “Application of Science and Technology to Rural Areas”, also called by its acronym “ASTRA”. The ASTRA institutional experiment was based on a model of technology & rural society interactions. The rural studies of energy, building, water, health etc. carried out by ASTRA provided an important step for technology development. ASTRA is now known as Center for Sustainable Energy (CST). Energy efficient wood burning devices, biomethanation, biogasification, green building, bioenergy and climate change studies are few examples of fruitful interventions of ASTRA [1]. CAPART (Council for Advancement of People’s Action and Rural Technology), an autonomous body registered in 1986 under aegis of Ministry of Rural Development is envisioned to play dynamic role to strengthen the voluntary movement in the country and facilitate the promotion of innovative rural technology[2]. Several other steps to promote science education among rural people were adopted but yet rural infrastructure was poor and incapable to support S&T for rural Indians. PRESENT SCENARIO FOR S&T IN RURAL AREAS STEM refers to the academic disciplines of science, technology, engineering and mathematics. The term is typically used when addressing education policy and curriculum choices in schools to improve competitiveness in science and technology development. STEM crisis has implications for work force development, national security concerns, immigration policy and S&T development in a country. Some educationist, technocrats, academicians, scientists and economists believe there exist a STEM crisis in India while others do not agree to it. But if we see analytical and statistical details we find STEM crisis does occur in rural India. Annual reports and surveys may be seen as an effort towards strengthening the S&T statistical network within our country. A survey by NCAER (National Council of Applied Economic Research) reveals engineering was the favorite

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in subject chosen by maximum number of students (22%) as the one in which they will like to complete higher education, medicine was next (18%) and pure sciences was marginally low [3]. FIG. 1 An another NCAER survey found that a fourth of those in rural areas said they would like to complete their higher education in arts as compared to 15% in urban areas. Since curiosity is basic nature of human, rural attention and interest towards S&T can be easily nurtured. It is believed that development of rural infrastructure and more awareness for S&T will help in socio-economic as well as technological development of rural areas. Moreover the agro economy of India works with agriculture and the rural people, who constitute more than 70% of our total population, still plays a front role. Today farmers use S&T for agriculture; some of these techniques/technologies included the use of manure/fertilizers, the use of water harvesting or green manuring [3]. FIG. 2. Facilities like 24 hours satellite channel and phone helplines along with internet services are working for farmers and these prove highly beneficial to them. With such instances of S&T use by rural people the spread of mainstream science education may also be imagined. Other silent reforms to enhance associations between rural society and S&T are underway. Recently Planning Commission of India was discarded and a new institution called NITI Aayog was established. NITI Aayog shall provide a critical directional and strategic input into developmental processes. It will work by providing better inter ministry and center-state coordination. It will develop mechanisms to formulate credible plans to the village levels and aggregate these progressively at higher levels. These plans include the rural areas. In 2015 Ministry of Human Resource Development set up Rashtriya Avishkar abhiyan (RAA) to nurture a spirit of enquiry and creativity, love for science and mathematics and effective use of technology among children. RAA will span across MHRD’s schematic interventions of Sarva Shiksha Abhiyan, Rashtryia Madhyamik Shiksha Abhiyan in Department of School Education and Literacy and programs of Department of Higher Education to encourage science, mathematics and technology. Institutes like ICAR, CSIR, DBT, CPRI etc. have established their local institutes in many towns of India. Many private medical and engineering institutes are established in rural areas due to low cost of settlement in these areas. NGOs like Pratham are working on direct instruction model for creating aptitude for science and math education among rural children. Aser-A nongovernmental survey reveals improvement in science and math learning through education camps [4]. FIG. 3. Agastya International Foundation works for developing science aptitude among rural children. They foster mobile labs, conduct science fairs, establish science centers in rural areas and run programs targeting drop out students and communities. Azim premji Foundation helping 2 million students across 16000 students from 14 states, Vidya gyan foundation and Samudaya are few other to mention. CONCLUSIONS Today the socio-economic conditions of India present an environment where rural India may S&T developed. Efforts to create their access to resources and a better infrastructure with proper facilities of electricity and water etc. shall be structured . Since technology prompts interest among rural student, E-learning may be a correct way for it. Results presented in surveys and reports will essentially be an important input for entire scientific community and policy makers to set achievable goals and work out action plans towards S&T development in rural areas. Since 1986 social, economic and scientific atmosphere of India has changed drastically, so our

53

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in education policy needs to be reviewed. A National Education Policy shall be formulated that work on soul motto of science and innovations and consider every child as an integral part of our nation building. This may fulfill Indian aspirations of being a developed economy someday. Promotion and development of S&T in India with special emphasis on rural areas may help on the path of national development. REFERENCES 1. Reddy AKN. Choices of Alternative Technologies. Economic and political weekly. 3(25), 1109-1114. (1973). 2. CAPART (Council for advancement of people’s action and rural technology). Avail from: http://www.capart.nic.in. 2010. [cited 1 oct 2015]. 3. Rajesh Shukla. Indian Science Report - Science education, HR & Public attitude towards S&T. NCAER, New Delhi. ISBN - 81-88830-07-0. (2005). 4. Pratham Education Foundation. Annual Status of Education Report (Rural) 2010. Pratham resource centre, New Delhi. (2011). FIGURES Figures mentioned above are represented here in ascending order.

6-8th 9th 10th 11-12th

Least informed Moderately Most informed

agriculture household communication health

SCIENCE ENGINN MEDICINE ARTS COMMERCE OTHERS

FIG. 1- Preferred subject for higher education by level of students

FIG. 2- Distribution of public by awareness of technology related to

Source: NCAER’s National Survey-2004

selected sectors. Source: NCAER’s National Survey- 2004

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in FIG. 3- Special learning camps in U.P. - Math progress-The percentage of children that can recognize 2-3 digit numbers has increased by 69% in 10 day model. Source: A Non Governmental Survey by Pratham Education Foundation.

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Effects of Distellery Effluent Irrigation on Mustard (Brassica campestris) RENU CHOUDHARY, NARESH KUMAR* AND HARENDRA MALIK * Department of Biology, G.G.I.C. Gagalheri, Saharanpur(U.P.), India. *Department of Botany, C.C.R. (P.G.) College, Muzaffarnagar (U.P.), India. Email- [email protected] Abstract A large network of distilleries has been established in India which have been recognized as one of the most polluting agro-based industries, generating huge quantities of distillery effluent. The present study was conducted to observe the impact of different concentrations i.e. 0.5,1.0,2.0,5,10,25,35,45 and 55% of different distillery effluents i.e. raw spent wash (RSW), biomethanated spent wash (BSW) and lagoon sludge (LS) collected from Shamli distillery and chemical works, Shamli, District-Shamli) on Mustard (Brassica campestris) cv. PAC-401. Distillery effluents (RSW, BSW and LS) considerably affected all the growth parameters in treated plants. Growth parameters improves up to 5% concentrations of RSW and up to 10% concentrations of BSW and LS, while above these concentrations growth is significantly reduced. The improvement or reduction in different biochemical content was observed according to the concentrations of effluents. The chlorophyll a and b, carotenoids and oil contents of plants were increased initially upto 5% concentrations of RSW and upto 10% concentrations of BSW and LS effluents. Higher concentrations (>5% of RSW and >10% of BSW and LS) of distillery effluents were detrimental to all the treated crop plants. Keywords : Distillery effluent, Raw spent wash, Biomethanated spent wash, Lagoon sludge, Germination, Growth, Mustard. Introduction During the last two decades, India has emerged as the largest sugar producer in the world. The sugar factories after extraction of crystalline sugar from cane juice, sell the molasses to distilleries, where it is converted into alcohol by yeast fermentation. The raw spent wash coming out as a result of fermentation process from the distilleries is almost 100 times more concentrated as compared to sewage with regards to its pollutional characteristics. Uttar Pradesh is the highest sugar producing state in India and therefore, here about 126 sugar factories and 61 distilleries are established in public as well as private sector mainly in western Uttar Pradesh. Most of these distilleries are highly polluting units. The effluents of almost all of these units are discharged either directly into water bodies or nearby rivers through drains. Some distilleries discharge their effluents into the adjoining crop areas and farmers use this effluent directly for irrigation. Therefore, in the present study, an attempt has been made to assess the effects of different concentrations (0.5,1.0,2.0,5,10,25,35,45 and 55%) of distillery effluents i.e. raw spent wash (RSW), biomethanated spent wash (BSW) and lagoon sludge (LS) irrigation on three important oil yielding crop plants grown in this locality viz. Mustard (Brassica campestris) cv. PAC-401. Materials and Methods

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in The main object of the present investigation is to study the effects of different distillery effluents (RSW, BSW and LS) irrigation at different concentrations on various aspects of Mustard (Brassica campestris) cv. PAC-401. All three pure effluents of distillery were diluted with tap water to get 0.5%, 1.0%, 2.0%, 5%, 10%, 25%, 35%, 45%, and 55% concentrations. The tap water was used as control. The soil of research plots or pots was irrigated with different concentrations of effluent (RSW, BSW and LS) on alternate days or according to the requirement of the crop. For seed germination studies, healthy seeds were selected for uniformity (criteria being the size and colour of seeds) and were surface sterilized with 0.1% mercuric chloride for two minutes and thoroughly washed with distilled water. Plants grown in research plots were taken out and rinsed their roots repeatedly with unionized water to eliminate undesirable nutrients from the root surface. Excess of moisture, thus created, was wiped out with the help of clean blotting paper and absorbent towels. Ordinary scale was used to measure the length of root and shoot in centimeters. For phytomass determination (g dw/plant), different plant parts were separated and oven dried at 800C for 24 hours or until a constant weight was achieved. Dry weight of different plant parts were added to get the phytomass of each plant. Chlorophyll, carotenoid and oil content calculated using several deeds [1, 3 & 4]. Result and Discussion A slight increase in germination percentage was recorded upto 5% concentration of RSW and 10% concentration of BSW and LS. 100% seed germination occurred at above concentrations of RSW, BSW and LS in Brassica campestris cv. PAC-401. Reduction in seed germination percentage was 42.42, 37.37 and 34.34 percent under 55% concentration of RSW, BSW and LS respectively. The speed of germination index increased upto 10.35 percent in 5% RSW concentration and 15.26 and 12.80 percent in 10% BSW and LS concentration treatments respectively, The reduction percentage was 20.00, 16.84 and 15.61 percent under 55% of RSW, BSW and LS concentration [7,8]. Table 1 : Effect of different concentrations of distillery effluents on seed germination percentage and speed of germination index of 8 days old seedlings of Brassica campestris cv. PAC-401. Treatments Parameters

RSW concentration (%) Contro

0.5

1.0

2.0

5.0

10

25

35

45

55

99

99

100

100

100

95

84

74

66

57

570

576

591

608

629

541

520

503

478

456

86

79

70

62

l Seed germination percentage Speed germination index

BSW concentration (%) Seed germination

99

100

100

100

100

57

100

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in percentage Speed germination

570

581

597

615

636

657

542

520

503

474

index LS concentration (%) Seed germination

99

99

99

100

100

100

91

83

72

65

570

573

588

605

626

643

556

529

511

481

percentage Speed germination index

Growth parameters improves up to 5% concentrations of RSW and up to 10% concentrations of BSW and LS, while above these concentrations growth is significantly reduced. Results of exposure to different concentrations of distillery effluents on various growth parameters of both cultivars are presented in tables 2, 3 and 4. The root length was increased upto 22.00% at 5% of RSW and 25.40 and 23.78% at 10% concentrations of BSW and LS treatments. The reduction percentage were 32.03, 26.05 and 24.43 percent in RSW, BSW and LS respectively at 55% concentration in 25 days old plants. In 50 days old plants the root length increased upto 16.52% at 5% of RSW and 19.80 and 17.90 percent at 10% of BSW and LS treatments respectively, while reduction percentage were 28.58, 24.12 and 22.07 percent at 55% concentrations of RSW, BSW and LS treatments. In 75 days old plants similar pattern was observed. Shoot length also show the similar pattern of increase and decrease in 25, 50 and 75 days old plants. The biomass production (phytomass accumulation) of both cultivars were found to be showing similar changes as found in root and shoot length in 25, 50 and 75 days old plants. Decrease in total dry weight or phytomass accumulation eventually lead to decrease in net primary productivity (NPP) of treated plants [5,6]. Table 2 : Effect of different concentrations of distillery effluents on different growth parameters of 25 days old plants of Brassica campestris cv. PAC-401. Treatments Parameters

RSW concentration (%) Control

Root length (cm)

Shoot length (cm) Biomass production(g)

0.5

1.0

2.0

5.0

10

25

35

45

55

6.18±0. 6.36±0. 6.78±0. 7.11±0. 7.54±0. 5.79±0. 5.34±0. 5.01±0. 4.58±0. 4.20±0. 84

81†

77†

72*

79**

82†

74*

67*

65**

69**

9.88±0. 10.27± 10.76± 11.29± 11.85± 9.19±0. 8.18±0. 7.76±0. 7.38±0. 7.01±0. 62

0.83†

0.88†

0.89*

0.82**

60†

56*

59**

73**

76**

2.66±0. 2.77±0. 2.90±0. 3.09±0. 3.26±0. 2.51±0. 2.36±0. 2.23±0. 2.12±0. 1.97±0. 46

NPP (g/plant/day) 0.106

52†

67†

59*

61**

45†

42*

38*

36**

31**

0.110

0.116

0.123

0.130

0.100

0.094

0.089

0.084

0.078

58

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in BSW concentration (%) Root length (cm) 6.18±0. 6.42±0. 6.88±0. 7.22±0. 7.64±0. 7.75±0. 5.67±0. 5.24±0. 4.82±0. 4.57±0. 84

76†

85†

73*

88**

81**

75†

71*

67**

65**

Shoot length (cm) 9.88±0. 10.50± 10.98± 11.54± 12.04± 12.17± 8.62±0. 8.18±0. 7.76±0. 7.34±0. 62 Biomass production(g)

0.68†

0.60†

0.67*

0.70** 0.63**

59†

55*

63**

54**

2.66±0. 2.79±0. 3.03±0. 3.19±0. 3.35±0. 3.44±0. 2.43±0. 2.30±0. 2.18±0. 2.05±0. 46

NPP (g/plant/day) 0.106

48†

44*

49**

52**

43**

39†

41*

37**

33**

0.111

0.121

0.127

0.134

0.137

0.097

0.092

0.87

0.082

LS concentration (%) Root length (cm) 6.18±0. 6.30±0. 6.68±0. 7.01±0. 7.43±0. 7.65±0. 5.80±0. 5.37±0. 4.94±0. 4.67±0. 84

81†

87†

76*

89**

82**

78†

83*

72**

76**

Shoot length (cm) 9.88±0. 10.23± 10.72± 11.22± 11.69± 12.12± 8.89±0. 8.30±0. 2.39±0. 2.27±0. 2.10±0. 62 Biomass production(g)

0.65†

0.60†

0.58*

0.64** 0.69**

71†

2.66±0. 2.73±0. 2.85±0. 3.01±0. 3.19±0. 3.30±0. 2.51±0. 46

NPP (g/plant/day) 0.106

43

†

0.109

†

†

49

52*

56**

58**

44

0.114

0.120

0.127

0.132

0.100

Values are in mean ± standard deviation. Significance of difference from control; P* < 0.05; P** < 0.01 and † non-significant

59

66*

41*

37*

35**

0.095

0.090

0.084

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Table 3 : Effect of different concentrations of distillery effluents on different growth parameters of 50 days old plants of Brassica campestris cv. PAC-401 Treatments Parameters

RSW concentration (%) Control

Root length (cm)

Shoot length (cm) Biomass

0.5

1.0

2.0

5.0

10

25

35

45

55

13.68±1 14.05±1 14.61±1 15.29±1 15.94±1 13.26±1 12.28±1 11.39±1 10.56±1 9.77±1. .44

.21†

.28†

.42*

.48**

.36†

.31*

.22**

.26**

09**

42.30±1 43.17±1 44.88±1 46.74±1 48.11±1 41.53±1 37.65±1 35.83±1 34.19±1 31.52±1 .78

.65†

.82†

.89*

.91*

.56†

.50*

.43**

.33**

.15**

8.56±0. 8.71±0. 9.08±0. 9.46±0. 9.92±0. 8.37±0. 7.98±0. 7.51±0. 6.90±0. 6.32±0.

production(g)

66

61†

65†

67*

71**

58†

53*

62**

59**

52**

NPP (g/plant/day)

0.171

0.174

0.181

0.189

0.198

0.167

0.159

0.150

0.138

0.126

BSW concentration (%) Root length (cm)

13.68±1 14.16±1 14.83±1 15.58±1 16.20±1 16.39±1 13.11±1 12.43±1 11.65±1 10.38±1 .44

.34†

.38†

.30*

.26**

.15**

.11†

.24*

.32**

.19**

Shoot length (cm) 42.30±1 43.46±1 45.50±1 47.6±1. 48.91±1 49.38±1 41.69±1 38.52±1 36.15±1 33.81±1 .78 Biomass

.81†

.86†

92*

.94**

.26**

.32†

.39*

.40**

.33**

8.56±0. 8.78±0. 9.22±0. 9.68±0. 10.13±0 10.35±0 8.24±0. 7.83±0. 7.39±0. 6.75±0.

production(g)

66

56†

63†

65*

.69**

.61**

58†

54*

51**

55**

NPP (g/plant/day)

0.171

0.175

0.184

0.193

0.202

0.207

0.164

0.156

0.147

0.135

LS concentration (%) Root length (cm)

13.68±1 13.96±1 14.46±1 15.07±1 15.73±1 16.13±1 13.19±1 12.54±1 11.81±1 10.66±1 .44

.31†

.42†

.39*

.46**

.36**

.39†

.28*

.24**

.29**

Shoot length (cm) 42.30±1 43.09±1 44.72±1 46.51±1 47.74±1 48.61±1 41.83±1 39.26±1 36.69±1 34.53±1 .78 Biomass

.74†

.81†

.85*

.69*

.86**

.52†

.46*

.62**

.65**

8.56±0. 8.67±0. 896±0.7 9.31±0. 9.78±0. 10.19±0 8.31±0. 7.97±0. 7.54±0. 6.98±0.

production(g)

66

70†

4†

78*

72**

.81**

68†

62*

58**

56**

NPP (g/plant/day)

0.171

0.173

0.179

0.186

0.195

0.203

0.166

0.159

0.150

0.139

Values are in mean ± standard deviation. Significance of difference from control; P* < 0.05; P** < 0.01 and † non-significant

60

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Table 4 : Effect of different concentrations of distillery effluents on different growth parameters of 75 days old plants of Brassica campestris cv. PAC-401. Treatments Parameters

RSW concentration (%) Control

Root length (cm)

Shoot length (cm) Biomass

0.5

1.0

2.0

5.0

10

25

35

45

55

16.08±1 16.29±1 16.85±1 17.43±1 18.26±1 15.52±1 13.24±1 12.47±1 11.61±1 10.52±1. .36

.39†

.42†

.44*

.45*

.33†

.28*

.31**

.24**

27**

123.42± 125.80± 128.63± 132.02± 136.48± 118.65± 104.73± 98.81±3 93.69±3 88.11±3. 3.48

3.56†

3.81†

3.73*

3.79*

3.56†

3.42*

.36**

.29**

48**

44.63±1 45.38±1 46.78±1 48.62±1 51.27±1 42.38±1 38.14±1 36.79±1 35.41±1 33.64±1.

production(g)

.16

.24†

.34†

.48*

.53**

.21†

.26*

.18**

.11**

16**

NPP (g/plant/day)

0.595

0.605

0.623

0.648

0.683

0.565

0.508

0.490

0.472

0.448

BSW concentration (%) Root length (cm)

16.08±1 16.38±1 17.03±1 17.69±1 18.52±1 18.96±1 13.89±1 12.96±1 12.07±1 11.32±1. .36

.41†

.54†

.39*

.48**

.51**

.33*

.27**

.29**

26**

Shoot length (cm) 123.42± 126.49± 129.52± 133.22± 137.29± 140.63±3 111.64± 102.88±3 96.75±3 90.82±3. 3.48 Biomass

3.51†

3.59†

3.77*

3.68*

.76**

3.29*

.15**

.31**

28**

44.63±1 45.64±1 47.26±1 49.36±1 51.94±1 52.88±1 40.19±1 37.53±1 36.04±1 34.77±1.

production(g)

.16

.19†

.27†

.35*

.39**

.42**

.18*

.14**

.08**

12**

NPP (g/plant/day)

0.595

0.608

0.630

0.658

0.692

0.705

0.535

0.500

0.480

0.463

LS concentration (%) Root length (cm)

16.08±1 16.20±1 16.67±1 17.15±1 17.89±1 18.45±1 14.20±1 13.28±1 12.46±1 11.83±1. .36

.34†

.48†

.54†

.31*

.35**

.46*

.39**

.33**

28**

Shoot length (cm) 123.42± 125.61± 128.19± 131.17± 135.81± 137.52±3 112.36± 103.43±3 97.38±3 91.88±3. 3.48 Biomass

3.56†

3.68†

3.85*

3.76*

.82**

3.37*

.61**

.48**

39**

44.63±1 45.27±1 46.56±1 48.09±1 50.68±1 52.14±1 40.78±1 38.21±1 36.69±1 35.49±1.

production(g)

.16

.27†

.31†

.37*

.44*

.50**

.29*

.14**

.22**

21**

NPP (g/plant/day)

0.595

0.603

0.620

0.641

0.675

0.695

0.543

0.509

0.489

0.473

Values are in mean ± standard deviation. Significance of difference from control; P* < 0.05; P** < 0.01 and † non-significant Considerable increase and decrease in chlorophyll a and chlorophyll b content of leaves were observed in different concentrations of RSW, BSW and LS treatments. For example, chlorophyll a and b content increased upto 21.48 and 22.47% at 5% concentration of RSW respectively, while at 10% concentration of BSW and LS these values were 25.65, 27.75 and 21.61, 23.85% respectively in 25 days old plants. Carotenoids, the accessory pigments increased by 19.69, 24.57 and 21.21% at 5% concentration of RSW and 10% concentration of BSW and LS respectively against control of 25 days old plants. Whereas, reduction of carotenoid content

61

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in occurred as 30.97, 24.91 and 23.40% at 55% concentration of RSW, BSW and LS respectively. Similar trends of results also obtained from 50 days and 75 days old plants. Oil content of seeds were increased upto 5% concentration of RSW, i.e. by 6.38% and upto 10% concentration of BSW and LS, i.e. by 8.27 and 7.85% respectively. Above these concentrations of RSW, BSW and LS, oil content decreased regularly and decrease was maximum at 55% concentration of RSW, BSW and LS, i.e. by 38.71, 20.67 and 20.38% respectively [ 2,9].

Chlorophyll-a content (mg/g f.wt.)

25 Days 1 0.8 0.6

RSW

0.4

BSW

0.2

LS

0

Chlorophyll-a contentcontent Chlorophyll-a (mg/g (mg/g f.wt.) f.wt.)

Treatments (%)

1

25 Days 50 Days

1.2 0.8 0.61

RSW

0.8 0.4

BSW

0.2 0.6

LS

RSW

0 0.4

BSW

0.2

LS

0

Treatments (%)

Treatments (%)

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75 Days Chlorophyll-a content (mg/g f.wt.)

1 0.8 0.6 RSW

0.4

BSW 0.2

LS

0

Treatments (%) Figure 1 : Effect of different concentrations of distillery effluents on chlorophyll-a content (mg/g f.wt.) of 25, 50 and 75 days old plants of Brassica campestris cv. PAC-401.

25 Days Chlorophyll-b content (mg/g f.wt.)

0.6 0.5 0.4 0.3

RSW

0.2

BSW

0.1

LS

0

Treatments (%)

50 Days Chlorophyll-b content (mg/g f.wt.)

0.8 0.7 0.6 0.5 RSW

0.4 0.3

BSW

0.2

LS

0.1 0

Treatments (%)

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75 Days Chlorophyll-b content (mg/g f.wt.)

0.7 0.6 0.5 0.4 0.3

RSW

0.2

BSW

0.1

LS

0

Treatments (%)

Figure 2 : Effect of different concentrations of distillery effluents on chlorophyll-b content (mg/g f.wt.) of 25, 50 and 75 days old plants of Brassica campestris cv. PAC-401.

25 Days Carotenoid content (mg/g f.wt.)

2.5 2 1.5 1

LS

0.5

BSW

0

RSW

Treatments (%)

Carotenoid content (mg/g f.wt.)

50 Days 2.5 2 1.5 1

LS

0.5

BSW

0

RSW

Treatments (%)

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75 Days Carotenoid content (mg/g f.wt.)

2 1.5

1 LS

0.5

BSW

0

RSW

Treatments (%)

Figure 3 : Effect of different concentrations of distillery effluents on carotenoid content (mg/g f.wt.) of 25, 50 and 75 days old plants of Brassica campestris cv. PAC-401.

cv. PAC-401 50 45

Oil content (%)

40 35 30 25

RSW

20

BSW

15

LS

10

5 0

Treatments (%) Figure 4 : Effect of different concentrations of distillery effluents on oil percentage of Brassica campestris cv. PAC-401. References 1.

Arnon, D.I. Copper enzymes in isolated chloroplasts, polyphenol oxidase in Beta vulgaris. Plant Physiol. 24 : 1-15 (1949).

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Chandra, R., Kumar, K. and Singh, J. Impact of anaerobically treated and untreated (raw) distillery effluent irrigation on soil microflora, growth, total chlorophyll and protein contents of Phaseolus aureus L. J. Environ. Biol. 25 (4) : 381-385 (2004).

3.

Maclachlan, S. and Zalik, S. Plastid structure, chlorophyll concentration and free amino acid composition of chlorophyll mutant of barley. Can. J. Bot. 43 : 1053-1062 (1963).

4.

Pain, S. K. and Nayek, B. Studies on the physiology of growth and development of Sesamum indicum cv. 13-9 with special reference to yield of seed and oil : The effect of indolyl-3-propeonic acid used as foliar spray. J. Indian Bot. Soc. 60 : 202-207 (1981).

5.

Pandey, S.N., Nautiyal, B.D. and Sharma. C.P. Pollution level in distillery effluent and its phytotoxic effect on seed germination and early growth of maize and rice. J. Environ. Biol. 29 (2) : 267-270 (2008).

6.

Rath ,P., Pradhan, G. and Mishra ,M.K. Effect of Sugar factory distillery spent wash (DSW) on the growth pattern of sugarcane (Saccharum officinarum) crop. J. Phytology. 2 (5) : 33-39 (2010).

7.

Sharma, V., Sharma, R. and Sharma, K.D. Distillery effluent effect on seed germination, early seedling growth and pigment content of Sugarbeet (Beta vulgaris Linn. Var. Mezzanau-Poly). J. Environ. Biol. 23 (1) : 77-80 (2002).

8.

Sunitha, N. S. and Seenappa, C. Indigenous State-of-The-Art Technology Development for Distillery Effluents: An organic biochemical reagent for productive soils by means of Aerobic Sponge Method Vermitechnology (ASMV). J. Chem. Bio. Phy. Sci. 3 (1) : 559-566 (2013).

9.

Zengin, F. K. and Kirbag, S. Effects of copper on chlorophyll, proline, protein and abscisic acid level of sunflower (Helianthus annuus L.) seedlings. J. Environ. Biol. 28 (3) : 561-566 (2007).

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Role of Transgenic Plants in Agriculture Renu Rani Department of Botany, Govt. Degree College, Behat, Saharanpur Email : [email protected] ABSTRACT Environmental Stresses, population explosion and food shortage have caused serious problems to mankind on the globe. The world population is increasing alarmingly and is projected to reach 9 billion by 2050. To fulfill the food demand on every individual from limited natural resources is difficult. This factor has resulted in food deficiency thereby causing malnutrition, which is a serious health problem these days. Food production will need to increase at the same rate or more in order to satisfy the needs of such an enormous number of people in some older centuries. So there is a need to use the genetic techniques to improve crops over the recent decads. Transgenic breeding uses molecular cloning techniques to identify cloned or synthesized genes of interest and directly transforms the recipient genome. This process manipulates plant genomes through insertion of gene(s) from another species. Transgenic plant have been found to have many advantages like, development of high yielding varieties of crop plants and disease resistant, and are plants with improved tolerance to biotic and abiotic stress. Key words: Abiotic stress, Genetically modified Organism, Genetic Engineering and Transgenic plants,. INTRODUCTION Increasing world population and food demands require world agricultural production be increased by 50% by 2030 (The Royal Society, 2009). In the meantime, climate change and shrinking environmental resources are limiting agricultural production over the world [1]. These challenges bring an urgent need to enhance crop productivity. To breed crops with increased yield and resistance to environment stresses, a pivotal consideration is how to effectively utilize genetic diversity. Genetic crossing, selection of natural or artificial mutations, and transgenics, are the main techniques for plant breeding. Traditional plant breeding uses crossing, mutagenesis and somatic hybridization for genome modification to improve crop traits. It introduces new beneficial alleles from crossable species. Due to crossing barriers and linkage drag, however, traditional plant breeding is timeconsuming and requires several generations of breeding and selection. Transgenic breeding uses molecular cloning techniques to identify cloned or synthesized genes of interest and directly transforms the recipient genome. This process manipulates plant genomes through insertion of gene(s) from another species. An organism that is generated this way is considered to be a genetically modified organisms (GMOs). Most genetically modified plants are generated either by particle gun method or by Agrobacterium tumefaciens mediated transformation method. Since 1990s, the major emphasis of agriculture biotechnology can be found on traits for improvement in crops related to insect and herbicide resistance, nutritional quality, virus resistance, shelf life and bio-fuel production [2].Transgenic plant

have been found to have many advantages like,

development of high yielding varieties of crop plants and disease resistant, and are plants with improved tolerance to biotic and abiotic stress [3-6]. Recently, biotechnology has revolutionized crop improvement by producing GM crops with enhanced availability and utilization of important traits [7]. In agriculture, yield is a major output and improvement in yield of plants is a major thrust area by counteracting biotic and abiotic environmental cues. Thus, crop cultivars with enhanced yield and stability are required.

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in ADVANTAGE OF TRANSGENIC PLANTS Environmental factors are essential components which affect crop yield to a great extent. The introduction of resistance to abiotic and biotic stress into crop plants has become a topic of major economic interest for agriculture. Infectious diseases are the most dangerous problems in the present world and each year one third of all deaths are caused by the infectious agents. Genetically engineered plants offer significant benefits by improving yield, transportation costs, enhancing the nutritional content, biotic and abiotic stress and biofuel production. in this review we analyze the role of transgenic plants in combating environmental stress. INSECT RESISTANCE Plants are equipped with the natural plant defense system against insects, fungi, bacteria which is provided by the proteinase inhibitors [8-10]. PIs are ubiquitous in plants especially in Leguminoseae, Gramineae, Compositeae. They are natural defense related proteins often present in seeds and induced in certain plant tissue by herbivory and wounding. The activity of PIs is due to their capacity to form stable complexes with target proteases, blocking, altering or preventing access to enzyme active sites. Bacillus thuringiensis (Bt) insect resistant crops are one of the most astounding achievements in plant transgenic technology. Bt is a potent insecticide which comprises crystal protein endotoxin produced by some strains of soil bacterium B. thuringiensis (a soil bacterium). The Bt crystal (cry) insecticidal protein (δ-endotoxin) genes are toxic to lepidopterans [11], dipterans [12], coleopterans [13]. Bt cry protein is non-toxic to humans and animals, but toxic to insects [14]. When cry protein are ingested by insects, they are dissolved in the alkaline juice present in midgut lumen. The gut proteases process them hydrolytically to release the core toxic fragment. The toxic fragment of cry protein have three domains, domain I function in pore and ion channel formations, domain II is involved in receptor recognition, while domain III bind to receptor. The toxic fragments are believed to bind to specific high affinity receptors present in the brush border of mid gut epithelial cell. As a result, the brush border membrane develops pores, permitting influx into the epithelial cell of ion and water, which cause their swelling and eventual lysis. HERBICIDE RESISTANCE The early herbicides were found to be very destructive for most plants and they created undesirable environmental impacts. New chemicals such as glyphosate have been widely recommended for use because glyphosate is environmental-friendly as soil microorganisms are able to degrade it rapidly. Plants expressing transformed herbicide tolerance accounted for 71% of all transgenic crops grown worldwide in 1998 and 1999 [15]. Herbicide tolerant soybean, corn, cotton and canola represent the major transgenic products [16]. Herbicide resistant Amaranthus palmeri developed by expressing glyphosate-insensitive herbicide target site gene, 5 enol pyruvylshikmate-3 phosphate synthase (EPSP) that is involved in the shikimate cycle where it catalyzes the reversible addition of enolpyruvyl moiety of phosphor enolpyruvate to shikimate 3 phosphate[17]. Generally two approaches have been used to create herbicide tolerant crops either modify the degree of sensitivity of the target enzyme so that the plant sensitivity to the herbicide is inhibited, or to engineer the herbicide-detoxifying pathway into the plant [18]. First approach includes transgenic plants tolerant to the herbicide acifluorfen, which inhibits chlorophyll biosynthesis, have been produced through over-expression of the target enzyme involved in chlorophyll biosynthesis [19]. In comparison, resistance to glufosinate and bromoxynil is based on the second approach. By introducing genes that enhance metabolism of these herbicides the active compound is converted to products that are non-toxic to the crop [20].

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in VIRUS RESISTANCE TMV resistant tobacco and tomato plant are produced by introducing viral coat proteins. Other viral resistant transgenic plants are Potato virus resistant potato, RSV resistant rice, YMV resistant black gram, YMV resistant green gram etc. ABIOTIC STRESS TOLERANCE Abiotic stresses such as salt, drought, flooding, extreme temperature and oxidative stresses often diminish plant growth and final yield. Agricultural productivity could be increased dramatically if crops were redesigned to better cope with environmental stresses. Over-production of a superoxide dismutase (SOD) gene resulted in increased chilling tolerance in plants. This could be due to the reason that different stress environment (high light intensity, pathogens and cold) produce reactive oxygen species (ROS) which can damage to plants. Antioxidant enzymes such as superoxide dismutase, catalase and peroxidase have the capacity to neutralize the effect of ROS [21, 3, 4]. While, observing the constitutive expression of Osmyb4 rice gene in A. thaliana under salinity, drought, temperature (low and high), and oxidative stress, shows that this gene helps in stress tolerance by regulating vital metabolites as well as ROS scavengers [22]. The over-expression of strawberry GalUR gene in transgenic potato resulted in enhanced tolerance to methyl viologen (MV), mannitol and salinity by increasing chlorophyll pigments and 1.6–2-fold high accumulation of AsA in transgenic plants as compared to that in wild type (non-transformed) plants[23]. The levels of AsA in the transgenic potato were significantly associated with enhanced GalUR activity. CONCLUSION AND FUTURE PROSPECTIVE The advent of genetic engineering (GE) and other tools has enabled plant biologists to fight against the prevailing adversaries. GM plants have been generated for their enhanced tolerance to herbicides and pests. Bt cotton is one of the best example of insect resistance transgenic plant. In India, transgenic Bt cotton was approved for commercial cultivation in 2002 and area under Bt cotton increased at the rate of almost 100% every year. In 2007, 131 different Bt cotton hybrids were in cultivation. The productivity of cotton during this period (2002-2007) increased from 300kg/ha to around 500kg/ha and as a result, India has now become a net exporter of cotton from being a net importer till 2003-04. In future, the transgenic crops will be used not only for improved agronomic traits, but also for traits involving food processing, pharmaceuticals (including edible vaccines) and specialty chemicals. Transgenic rubber tree has also been produced and will be used for a variety of purposes. Thus the future of transgenic crop is bright. Undoubtedly, there is a consistent increase in the use of genetically modified organisms for food or other essential commodities. The promoters of GM foods claim that they are environment-friendly, have no risk to human health, profitable for farmers as well as well regulated, many people are still of the firm view that GM foods can be injurious to human and animal health, because they have not been properly tested. Also it is not certain what types of long-term effects GM foods can cause. Critics argue that transferring new genes into a food can alter the chemical composition of that food, which may trigger the human body to respond differently to that food, thereby developing allergies or causing long-term toxicity. Thus, every country needs to frame well defined rules and regulations for the utilization of GM organisms, although many developed and some developing countries have already formulated specific regulations. REFERENCES 1.

Lobell D., Burke M., Tebaldi C., Mastrandrea M., Falcon W. and Naylor R. Prioritizing climate change adaptation needs for food security in 2030, Sci. 319, 607–610, (2008)

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Ahmad P., Ashraf M., Younis M., Hu X., Kumar A., Akram, N.A. and AlQurainy F. Role of transgenic in agriculture and biopharming, Biotechnol. Advances, 30, 524-540 (2012)

3.

Ahmad P., Sarwat M. and Sharma S. Reactive oxygen species, antioxidants and signaling in plants, J. Plant Biol., 51(3), 167-173 (2008)

4.

Ahmad P., Jaleel C.A., Salem M.A., Nabi G. and Sharma S. Roles of Enzymatic and non-enzymatic antioxidants in plants during abiotic stress, Crit. Rev. Biotechnol. 30(3), 161–175 (2010a).

5.

Ahmad P., Umar S. and Sharma S. Mechanism of free radical scavenging and role of phytohormones during abiotic stress in plants, In: M Ashraf, M Ozturk, M.S.A. Ahmad (ed.) Plant adaptation and phytoremediation. Springer Dordrecht Heidelberg, London, New York (2010b).

6.

Ahmad P. and Umar, S. Oxidative stress: Role of antioxidants in plants, Studium Press Pvt. Ltd. New Delhi, India. (2011)

7.

Icoz I. and Stotzky G. Fate and effects of insect-resistant Bt crops in soil ecosystems, Soil Biol. Biochem., 40, 559–586 (2008)

8.

Jongsma M.A. and Bolter C. The adaptation of insects to plant protease inhibitors, J. Insect Physiol, 43, 885–895 (1997)

9.

Larry L.M. and Richard E.S. Lectins and protease inhibitors as plant defenses against insects, J. Agric. Food Chem., 50, 6605–6611 (2002)

10. Kim J.Y., Park S.C., Hwang I., Cheong H., Nah J.W., Hahm K.S. and Park Y. Protease inhibitors from plants with antimicrobial activity, Int. J. Mol. Sci , 10, 2860–2872 (2009) 11. Cohen B.M., Gould F., Bentur J.C. Bt rice: practical steps to sustainable use, Int Rice Res, 25, 4-10 (2000). 12. Andrews R.W., Fausr R., Wabiko M.H. and Roymond K.C. Bulla LA. Biotechnology of Bt: a critical Review, Bio/Technol, 6, 163–232, (1987) 13. Herrnstadt C., George G.S., Edward W.R. and David, L. A new strain of Bacillus thuringiensis with activity against coleopteran insects, Nat Biotechnol., 4, 305–308 (1986) 14. BANR (Board on Agriculture and Natural Resources). Genetically modified pest protected plant: science and regulation, pp. 292, (2000) 15. James, C. Global Review of commercialized Transgenic Crops in 1999, Int Service Acquisition Agric Biotechnol Appl, 12, 1-7(1999) 16. Gaines T.A., Zhang W., Wang D., Bukun B., Chisholm S.T., Shaner D.L. and Nissen S.J. Gene amplification confers glyphosate resistance in Amaranthus palmeri, Proc. Nat. Acad. Sci. USA, 107, 1029-1034 (2010) 17. Simoens C. and Van Montagu M. Genetic engineering in plants, Hum. Reprod. Update, 1, 523-542 (1995) 18. Lermontova I. and Grimm B. Overexpression of plastidic protoporphyrinogen IX oxidase leads to resistance to the diphenyl-ether herbicide acifluorfen, Plant Physiol., 122, 75–84 (2000) 19. Haumann B.E. Bioengineered oilseed acreage escalating, Inform., 8, 804–811 (1997) 20. Hiei Y., Ohta S., Komari T. and Kumashiro T. Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA, Plant J., 6, 271– 282 (1994)

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 21. Vannini C., Locatelli F., Bracale M., Magnani E., Marsoni M. and Osnato M. Overexpression of rice Osmb4 gene increases chilling and freezing tolerance of Arabidopsis thaliana plants, Plant J., 37, 115127, (2004) 22. Hemavathi, Upadhyaya C.P., Young K.E., Akula N., Kim H.S., Heung J.J., Oh O.M., Aswath C.R., Se Chul Chun S.C., Kim D.H. and Park S.W. Over-expression of strawberry D-galacturonic acid reductase in potato leads to accumulation of vitamin C with enhanced abiotic stress tolerance, Plant Sci., 177, 659-667 (2009).

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Depression in Serum Zinc Concentration and Elevation in Serum Potassium Concentration in Chronic Renal Failure Patients Punam Yadav Department of Chemistry, M. S. College, Saharanpur U.P. Corresponding address: Dr. Punam Yadav, New Madhonagar Saharanpur-247001 (U.P.) Email- [email protected] ABSTRACT Abnormal serum zinc and serum potassium levels have been associated with increased mortality in numerous observational studies. Hypozincaemia is defined as a decrease in the serum zinc concentration to a level below 48-96 µg/ 100 ml. Hyperkalaemia occurs when serum potassium concentration is increased in chronic renal failure patients. Hyperkalaemia is dangerous because cardiac arrest can occur when plasma potassium exceeds 7 mmol/L. Both Hypozincaemia and Hyperkalaemia are common conditions, especially in hospitalized patients and in patients with various comorbid conditions such as chronic renal failure disease. The present paper includes the study of serum potassium levels of 200 patients (according to age group and sex) with chronic renal failure (CRF) before and after the process of treatment and it has been compared with 50 normal healthy individuals comprising the control group. Key Words: Serum zinc, Serum Potassium, Hypozincaemia, Hyperkalaemia, Chronic Renal Failure INTRODUCTION Zinc is an important cation in the body. In recent times, zinc has been recognized as a constituent of prime importance in a variety of metallo-enzyme systems and biochemical pathways essential for protein synthesis and metabolism of carbohydrates, fats and proteins. Zinc is therefore, crucial for growth and development. Potassium is the major components of the cations of the extracellular fluid and exists in the body in association with the anion is chloride, bicarbonate, phosphate and lactate. The important functions of potassium are to regulate acid-base equilibrium and maintenance of the osmotic pressure of the body fluid thus protecting the body against excessive fluid loss. It also functions in the preservation of normal irritability of muscles and the permeability of the cells. Conclusive evidence of the essentiality of zinc to the normal growth and development of animals was not reported until 1934 [1]. The first concrete demonstration of a specific biologic function was published in 1939 [2]. More recently DNA department, RNA polymerase has been shown to be zinc dependent enzymes [3].The activity of ribonuclease has been demonstrated to increase in zinc deficient tissue, suggesting that RNA catabolism is regulated by zinc. The chronic renal failure (CRF) is one of the most severe diseases worldwide[4]. The renal failure occurs when the kidneys cannot properly remove wastes that causes buildup of waste and fluid in the body [5].It was first reported that uremics had a factor in plasma that could reduce Na +-K+ ATPase activity of normal erythrocyte using a cross incubation method [6]. The potassium balance is usually maintained in the early stage of chronic renal failure through the increased potassium excretion per functioning nephron and the colon by aldosteron induced increase in Na-K ATPase activity as long as urine output remains adequate [7]. The evidence is also available to suggest a contribution of Potassium recycling to the overall handling of potassium along the loop of henle [8]. The thick ascending limb of the loop of henle is an important site of sodium, potassium, bicarbonate and ammonium transport[9,10]. In patients of chronic renal insufficiency, fractional potassium excretion is greatly increased [11]. The deficiency in the pump’s energy substrate the ATP

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in itself may perhaps be ruled out since the pump is saturated with ATP under physiological conditions [12]. The serum potassium values in the renal failure patients were not significantly different from normal values (4.5 ± 0.2 meq/l vs 4.2 ± 0.1 meq/l) [13].The relationship between changes in potassium balance and electrolyte and fluid transport, particularly with respect to potassium along the loop of henle remains to be fully elucidated. He also observed that no significant changes in plasma sodium concentration were observed. Moderately elevated plasma potassium levels were observed in another study of chronic hyperkalemia [14]. Chronic kidney disease is known to affect by the disturbance in the concentration of serum urea, serum creatinine, serum electrolytes and serum uric acid [15-17]. EXPERIMENTAL Materials and methods: The present study was carried out on 200 adult patients of chronic renal failure attended in the S.V.B.P. hospital attached to L.L.R.M. Medical College, Meerut and also 50 normal healthy individuals with age, sex matched who had no history of renal failure to serve as controls. All the known cases of chronic renal failure were included in this study on the basis of clinical and biochemical criteria. After confirmation of diagnosis on the above parameters, blood samples were drawn from these patients for the estimation of serum zinc and serum potassium levels. Observations: TABLE I SHOWING DISTRIBUTION OF C.R.F. CASES ACCORDING TO AGE GROUP AND SEX

Age Groups

Number of cases

Total

(Years)

Males

Females

10-30

5

2

7(3.5%)

31-50

50

25

75(37.5%)

51-70

63

40

103(51.5%)

71-above

10

5

15(7.5%)

Total

128(64.0%)

72(36.0%)

200(100.0%)

Out of 200 individuals, 128 (64%) controls were male’s individuals and rest 72 (36%) were females. All the 200 individuals were between the age group of 10 above 70 years. The maximum number of cases, 103 (51.5%), were observed in the age group of 51-70 years followed by 75 (37.5%) cases in the age group of 31-50 years, 15(7.5%) cases in the age group of above 70 years and 7 (3.5%) cases in the age group of 10-30 years. It is observed that the incidence of chronic renal failure reaches its maximum strength during middle age and later part of life. TABLE II SHOWING DISTRIBUTION OF CONTROL CASES ACCORDING TO AGE GROUP AND SEX

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Number of cases

Total

(Years)

Males

Females

10-30

4

1

5(10.0%)

31-50

12

6

18(36.0%)

51-70

18

9

27(54.0%)

Total

34

16

50(100.0%)

Out of 50 control cases, 34 (68.0%) cases were males and 16 (32.0%) were females. 54.0% were found in the age group 51-70 years, 36.0% were 31-50 years age group and 10.0% were 10-30 years age group.

TABLE III DISTRIBUTION OF C.R.F. CASES ACCORDING TO DURATION OF ILLNESS

Duration of illness

No. of cases

Percentage %

3 months-6 months

42

21.0%

6 months-1 year

114

57.0%

More than 1 year

44

22.0%

Total

200

100.0%

The majority of chronic renal failure cases were among more than 6 months- 1 year duration (114 cases, 57.0%) and then more than I year children (44 cases, 22.0%). TABLE IV SERUM ZINC AND SERUM POTASSIUM LEVEL IN NORMAL HEALTHY CONTROLS

ZINC

POTASSIUM

Age in years

Age in years

MALE

10-30

31-50

51-70

Total

10-30

31-50

51-70

Total

No.

4

12

18

34

4

12

18

34

Range

76-115

80-114

83-115

76-115

4.5-6.0

2.5-5.9

4.2-6.0

2.5-6.0

Mean±S.D.

103.75±

98.38±

97.61±

98.59±

5.25±

4.22±

5.14±

4.82±

16.08

28.86

9.26

10.41

0.56

0.97

0.52

0.86

FEMALE

10-30

31-50

51-70

Total

10-30

31-50

51-70

Total

No.

1

6

9

16

1

6

9

16

Range

73-80

85-110

80-109

73-110

3.5-4.5

3.0-4.0

3.8-4.9

3.0-5.0

Mean±S.D.

73.00±

97.58±

97.33±

95.88±

4.10±

3.86±

4.40±

4.18±

9.32

8.46

9.44

10.84

0.34

0.98

0.37

0.58

10-30

31-50

51-70

Total

10-30

31-50

51-70

Total

TOTAL

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5

18

27

50

5

18

27

50

Range

73-115

80-114

80-115

73-115

3.5-6.0

2.5-5.9

3.8-6.0

2.5-6.0

Mean±S.D.

97.60±

98.11±

97.52±

97.72±

5.02±

4.10±

4.89±

4.61±

18.92

9.49

9.29

10.79

0.67

0.92

0.59

0.84

The level of serum zinc in healthy subjects was 73-115 µg/100ml (mean 97.72±10.79 µg/100ml). In males, the range was 76-115 µg/100ml (mean 98.59±10.41 µg/100ml) and in females, the range was 73-110 µg/100ml (mean 95.88±10.84 µg/100m)l. The highest serum zinc level was observed in the age group of 31-50 years, ranged as 80-114 µg/100ml (mean 98.11±9.49 µg/100ml) while highest serum potassium level was observed in the age group of 10-30 years, ranged as 3.5-6.0 mmol/L (mean 5.02±0.67mmol/L). The lowest serum zinc level was observed in the age group of 51-70 years, ranged as 80-115 µg/100ml (mean 97.52±9.29 µg/100ml) while lowest serum potassium level was observed in the age group of 31-50 years, ranged as 2.5-5.9 mmol/L (mean 4.10±0.92mmol/L). No significant difference was seen among the serum zinc and potassium levels of different age groups and sexes. Our observations are very close to the observations of many workers (Kavukcu et. al. 1993, normal serum potassium level is 4.1±0.2 mmol/L), (Unwin et. al. 1994, normal serum potassium level is 4.09±0.06 mmol/L), (Price 1978, normal serum potassium level is 3.4-5.4 mmol/L) and (Harper et. al. 1979, normal serum potassium level is 2.5-5.0 mmol/L)[14,18-20]. TABLE V SERUM ZINC AND SERUM POTASSIUM LEVELS BEFORE AND AFTER TREATMENT IN TOTAL CASES OF CHRONIC RENAL FAILURE

Serum Zinc and Potassium ZINC Interval

POTASSIUM

No. of

Range (µg/100

Cases

ml)

Control

50

73-115

97.72±10.79

2.5-6.0

4.61±0.84

Before treatment

200

48-96

81.20±12.84***

3.2-8.0

6.46±1.32***

15 days after treatment

186

50-100

85.60±13.77***

3.0-7.5

5.92±1.15***

30 days after treatment

169

60-108

91.56±10.95***

2.8-6.9

5.27±0.99**

60 days after treatment

145

66-110

95.36±8.74

2.6-6.4

4.85±0.95

90 days after treatment

122

70-114

97.04±10.34

2.4-5.9

4.54±0.89

P- Significance, control vs treatment *p < 0.05, **p < 0.01, ***p < 0.001.

75

Mean ±S.D.

Range (mmol/L)

Mean ± S.D.

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in The range of serum zinc level was significantly low before and after thirty days of treatment while range of serum potassium level was significantly high before and after fifteen days of treatment in the patients of chronic renal failure as compared to controls. The range of serum zinc before, after fifteen and thirty days of treatment were 48-96 µg/100 ml (mean 81.20±12.84 µg/100 ml), 50-100 µg/100 ml (mean 85.60±13.77 µg/100 ml) and 60-108 µg/100 ml (mean 91.56±10.95 µg/100 ml) respectively. Serum zinc level after sixty and ninety days of the treatment ranged as

66-110 µg/100 ml (mean 95.36±8.74 µg/100 ml) and 70-114 µg/100 ml (mean

97.04±10.34 µg/100 ml) respectively. The range of serum potassium level before treatment was 3.2-8.0 mmol/L (mean 6.46±1.32 mmol/L). After fifteen, thirty, sixty and ninety days of treatment serum potassium ranged as 3.0-7.5 mmol/L (mean 5.92±1.15 mmol/L), 2.8-6.9 mmol/L (mean 5.27±0.99 mmol/L), 2.6-6.4 mmol/L (mean 4.85±0.95 mmol/L) and 2.4-5.9 mmol/L (mean 4.54±0.89 mmol/L) respectively. After sixty and ninety days no significant difference was observed in serum zinc and serum potassium as compared to controls. DISCUSSION Hypozincaemia in uremia is rather a result of a shift of zinc from plasma into tissue due to zinc while Hyperkalemia is generally occur in the patients of severe chronic renal failure due to potassium imbalance. The mean whole blood zinc concentration of male and female hemodialysis patients was significantly below control values while mean whole blood potassium concentration of male and female patients was significantly high than control values during chronic renal failure. The treatment of both these conditions is tricky, as over-treatment can lead to potentially dangerous complications and under-treatment is associated with significant mortality and morbidity. It is therefore essential to monitor the serum sodium and serum potassium concentration every 2 – 4 hours to prevent treatment related complications. The present study is conducted on a total of 250 individuals, out of which 50 are normal healthy individuals comprising the control group and rest 200 is of chronic renal failure. Results of biochemical parameter like serum uric acid from this study are discussed belowOut of 200 individuals, 128 (64%) controls were male’s individuals and rest 72 (36%) were females. All the 200 individuals were between the age group of 10 above 70 years. The maximum number of cases, 103 (51.5%), were observed in the age group of 51-70 years followed by 75 (37.5%) cases in the age group of 31-50 years, 15(7.5%) cases in the age group of above 70 years and 7 (3.5%) cases in the age group of 10-30 years (Table I). Out of 50 healthy controls, 34 (68%) controls were male’s individuals and rest 16 (32%) were females (Table II). It is observed that the incidence of chronic renal failure reaches its maximum strength during middle age and later part of life. Biochemical Studies The levels of serum zinc and serum potassium were studied in controls and in all cases of chronic renal failure. In normal healthy subjects serum zinc ranged from 73-115 µg/100ml (mean 97.72±10.79 µg/100ml).In males, it ranged from 76-115 µg/100ml (mean 98.59±10.41 µg/100ml) and in females, 73-110 µg/100ml

(mean

95.88±10.84 µg/100ml) (Table IV). The range of serum potassium in healthy subjects was 2.5-6.0 mmol/L (mean 4.61±0.84mmol/L). In males, it was 2.5-6.0 mmol/L (mean 4.82±0.86mmol/L).and in females, it was 3.05.0 mmol/L (mean 4.18±0.58mmol/L) (Table IV). No significant difference was observed in the serum zinc level of different age groups and sex. In cases of chronic renal failure serum zinc was found to be depressed in 72% cases while serum potassium was found to be elevated in 84% cases. Before treatment serum zinc level

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in was 81.20±12.84 µg/100 ml which was significantly low (p < 0.001) as compared to that of controls (97.72±10.79 µg/100 ml) and serum potassium level was 6.46±1.32 mmol/L which was significantly high (p < 0.001) as compared to that of controls (4.61±0.84 mmol/L). Generally Hypozincaemia occurs due to lower serum zinc level while hyperkalaemia occurs due to high serum potassium level in the patients of chronic renal failure. In Hypozincaemia the decreased intake of zinc due to low protein diet is regarded as a relevant factor in the development of zinc deficiency in chronic renal failure. The plasma zinc concentration is dependent on the balance between anabolic and catabolic processes.

SERUM ZINC LEVELS IN TOTAL CASES OF CRF 120

SERUM ZINC LEVELS

100 80 60

CRF CASES

40 20

0 Control

Before Treatment

After 15 Days

After 30 Days

After 60 Days

After 90 Days

TIME INTERVAL (DAYS)

Hyperkalaemia is caused by various metal disorders, shift of potassium from tissues etc. Hyperkalaemia may also develop rapidly if the potassium load is increased or excretory capacity is limited. Hyperkalaemia is dangerous because cardiac arrest can occur when plasma potassium exceeds 7 mmol/L. So, the corrections of sodium and potassium electrolytes are very important for the improvement of the condition of patients.

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SERUM POTASSIUM LEVELS IN TOTAL CASES OF CRF SERUM POTASSIUM LEVELS

7 6 5 4 CRF CASES

3 2 1 0 Control

Before Treatment

After 15 Days

After 30 Days

After 60 Days

After 90 Days

TIME INTERVAL (DAYS)

SUMMARY AND CONCLUSIONS The decreased levels of serum zinc caused hypozincaemia and increased levels of serum potassium levels caused Hyperkalaemia in chronic renal failure patients. In the study group, the levels of serum zinc were found decreased ranging between 48-96 µg/ 100 ml and mean 81.20±12.84 µg/ 100 ml however the levels of serum potassium were found highly increased, ranging between 3.2-8.0 mmol/L and mean 6.46±1.32 mmol/L. Significant difference (p < 0.01) was observed among the chronic renal failure patients and controls. The serum zinc and serum potassium levels are closely related to the severity of the disease. The following conclusions are derived from this study: 1- There was insignificant difference in the levels of all the above mentioned parameters as regards to age or sex of the healthy controls included in this study. 2- Maximum probabilities of chronic renal failure were found in the age group of 51-70 years (51.5%). 3- Minimum probabilities of chronic renal failure were found in the age group of 10-30 years (3.5%). 4- The levels of serum zinc and serum potassium were found to be significantly depressed and elevated respectively in cases of chronic renal failure as compared to that of controls. 5- The levels of serum zinc and serum potassium remained low and high respectively after thirty days of treatment and then returned to normal. 6-

The fall in the levels of serum zinc and serum potassium is related to the extent of the disease.

7- The levels shifted to normal range as the condition of patients improved clinically.

REFERENCES 1.

Tood W.R., Elvehjem C.A. and Hart E.B. Amer. J. Physiol. (107) 146 (1934)

2.

Keilen D. and Mann T. Nature. 144, 442.(1939)

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 3.

Kichgesser H., Roth H.P. and Weigand E. Biochemical changes in zinc deficiency in trace elements in human health and disease. Vol. I. Prasad A.S. ed. Acad. Press, New York. P-189 (1976)

4.

Stenvinkel P. and Alvestrand A. “Inflammation in end-stage renal disease: Sources, Consequences, and Therapy”. Semin Dial, 15(5) 329-337 (2002)

5.

Lise M., Cassio L. and Richar M.G. "Kidney Failure". The Journal of the American Medical Association, 301(6) 686 (2009)

6.

Cole C.H., Balfe J.W. and Welt L.G. Induction of a Quabain Sensitive ATPase defect by uremic plasma. Trans Assoc. Am. Phys. (81) 213-220 (1968)

7.

Bastl C., Hayslett J.P. and Binder H.J. Increased large intestinal secretion of potassium in renal insufficiency. Kidney Int., (9) 12 (1977)

8.

Jamison R.L., Work J. and Schafer J.A. New pathways for potassium transport in the kidney. Am. J. physiol. (242) 297-312 (1982)

9.

Good D.W., Kneppar M.A. and Burg M.B. Ammonia and bicarbonate transport by thick ascending limb of rat kidney. Am. J. Physiol. ( 247), 35-44 (1984)

10. Greger R., Schlatter and Lang F. Evidence for electroneutral sodium chloride co-transport in the cortical thick ascending limb of Henle’s lop of rabbit kidney. Pfluger Arch., (396) 308-314 (1983) 11. Hene R.J., Koomans H.A., Boer P., Roos J.C. and Mees E.J.D. Relation between plasma Aldosterone concentration and Renal handling of Sodium and Potassium in particular in patients with CRF. Nephron. (37) 94-99 (1984) 12. Mujais S.K., Sabatini S. and Kurtzman N.A. Pathophysiology of the uremic syndrome in the kidney, edited by Brenner BM, Rector FC, (3rd ed.) phiadelphia, Saunders WB. P- 1950 (1986) 13. Ray S., Piraino B., Chong T.K., Shanawy M.E. and Puschett J.B. Acid excretion and serum electrolyte in particular in patients with Advanced CRF. Miner elect. Metab. (16)355-361 (1990) 14. Unwin R., Capasso G. and Giebisch G. Potassium and sodium transport along the loop of Henie, effect of altered dietry potassium intake. Kid Int. (46)1092-1099 (1994) 15. Yadav Punam, Malik Dinkar, Kumar Sandeep, Malik Vijai. A role of serum uric acid in Chronic Renal Failure Patients and its effects. Int. J. Sci. Res. & Edu. 2(3), 434-442 (2014) 16. Yadav Punam, Malik Dinkar, Kumar Sandeep, Malik Vijai. Effect of elevated creatinine level in blood serum of Chronic Renal Failure Patients. Biological Forum, 6(1) 48-52 (2014) 17. Yadav Punam, Malik Dinkar, Kumar Sandeep, Malik Vijai. Study of serum urea in the patients of chronic renal failure. Medical Science, (4) 76-80 (2014) 18. Kavukcu S., Saatci U. and Ozean S. Effects of recombinant human erythropoietin on sodium balance in non dialysed children with CRF. Int. Urol. Nephrol. 144,442 (1993) 19. Price’s. Text book of the practice of Med. Scott, R.B.E.L.B.S. 12 th ed. Oxford, 1948-1950, 388 (1978) 20. Harper HA, Victor WR and Peter A Mayes. Review of physiological chemistry, 17 th edition. Maruzen Asian ed. Lange Medical Publications. ( 215) 579 (1979)

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Enhancement of Fe and Zn in Cowpea grain by applying soil and foliar application Namita Yadav* & Y.K. Sharma Plant Physiology lab, Botany Department, University of Lucknow, Lucknow ABSTRACT This pot experiment was carried out in green house condition to investigate Fe, Zn and Fe+Zn soil and foliar application effects on their concentration and protein in cowpea (vigna unguiculata) grain. The experiment was designed with three replicate. In soil application concentration of Fe and Zn (10ppm) were applied with N: P: K and farm yard manure in presowing soil for proper status of soil. In foliar application 0.5% of Fe and Zn were sprayed at four different stages of plant growth. The results showed that seed protein and tissue concentration of elements were more significant in individual and combined application of Fe and Zn than control. The study of results explains that soil and foliar application with micronutrients may have an important role in increasing cowpea yield. Keyword: cowpea, soil application, foliar application, tissue concentration and protein *Corresponding Author [email protected] Introduction: Cowpea is an important crop of legume family. Legumes are rich source of protein and energy for human food. Cowpea seeds are a nutrition component in the human diet as well as a nutritious livestock feed. The protein in cowpea seeds is rich in lysine and tryptophan amino acids compared to cereal grains [1]. Due to nutritional value and economic importance, it is necessary to focus on improving crop yield and production by applying fertilization strategies. Using soil and foliar application of micronutrients is one of most important strategies involved in improving plant growth, yield and quality of cowpea crop. Iron and zinc are essential micronutrient, need in small amount for plants. They can maintain crop physiology balance and play vital role in plant metabolism such as photosynthesis respiration, nitrogen fixation, chlorophyll development and function and reproductive physiology [2]. El-Fouly [3] reported that the availability of micronutrients such as Fe, Mn and Zn is much affected by pH and CaCO3 content as well as soil texture. The availability and uptake of micronutrients by plants decrease with increasing soil pH [4]. The mobility of microelements in soil and their translocation in plants as well as interaction among themselves play an important role in plants nutrition [2]. The deficiency of microelements in higher plants impaired the important metabolic functions and resulted in poor growth and yield of crops [5]. They also play vital role in enzyme metabolism being the cofactor and constituent of some enzymes and protein [2]. To sustain high crop yield the application of nutrients are required. In soil application nutrients are applied to plants with soil mixing while in foliar application nutrient are applied to plants in soluble form by spray. The aim of this study is to investigate the effect of micronutrient foliar and soil application on cowpea yield and quality of grain in soil.

Materials and methods:

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Cowpea (Vigna unguiculata) var. rituraj plants were grown in pots containing 10 Kg soil in a glass house at ambient temperature (15.C–30.C). The crop was raised in soil prepared with recommended dose of nitrogen (N), phosphorous (P) and potassium (K) 120: 60: 60 and farmyard manure. Experiment included a soil application for proper status of soil and four foliar treatments of iron and zinc at different stages of plant growth as follows: Soil application 1) NPK + farmyard manure ( control) 2) FeSO4 (10ppm) with NPK + farmyard manure 3) ZnSO4 (10ppm) with NPK + Farmyard manure 4) ZnSO4 (10ppm) + FeSO4 (10ppm) with NPK + farmyard manure Foliar application ZnSO4 (0.5%) and FeSO4 (0.5%) applied on 35D, 45D, 55D and 65D. The experiment was designed with three replicate. Initially two plants were maintained in each pot. Apart from the visual effect of Fe and Zn, the concentrations of iron, zinc and protein were observed at 35D, 45D, 55D and 65D in cowpea seeds. Tissue concentration was determined in oven dried seed material after wet digestion [6] and estimated in AAS and seed protein was estimated by Lowry et al [7] method. Results and Discussions: Vegetative growth of crop was promoted with the foliar application of iron and zinc. Plant height, pod length, number of pods per plant, number of seed per pod, weight of 100 seeds were more significantly increase as compare to control.

Cowpea Results showed that individual and combined treatment of iron and zinc significantly increased nutrient concentration of cowpea seeds as compared to control treatment (Figure 1, 2). The results indicate that highest concentration of iron was accumulated at vegetative phase (35D) in individual treatment but in combined treatment iron concentration suppressed by zinc application. Zinc concentration was more accumulated at reproductive phase (65D) in individual application and in combined application zinc concentration was more as compared to control treatment. Seed protein result showed significant differences among treatments as well as difference for interaction of iron and zinc on the protein percentage of seed. The highest protein percentage was obtained when cowpea treated with iron and zinc individual treatment and in combined treatment of iron and zinc the protein concentration was more as compared to control treatment (Figure 3). Zeidan [8] observed that Zn application significantly increased the grain Zn concentration, while simultaneously reduced the grain P concentration.

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Graph Values are mean of 3 replications-

ppm Fe in dry matter

120 100 80 60

Fe(contol)

40

Fe

20

Fe(Zn+Fe)

0

35D

45D

55D

65D

Days of foliar application

Figure 1: Effect of foliar concentration of iron in Cowpea seed at different days

ppm Zn in dry matter

40 Zn(control)

35 30 25 20

Zn

15 10 5 0

35D

45D

55D

65D

Zn(Zn+Fe)

Days of foliar application

Figure 2: Effect of foliar concentration of zinc in cowpea seed at different days

% protein in dry matter

20 19 18

control

17

Zn

16

Fe

15

Zn+Fe

14 35D

45D 55D 65D Days of foliar application

Figure 3: Effect of foliar concentration of Fe and Zn on protein percentage in cowpea seed Conclusion: Zinc and iron deficiency in human are important malnutrition problem worldwide. Human Zinc deficiency is major cause of disease and deaths in developing countries. Zinc deficiency is associated with poor growth and

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in development and impaired immune system. By applying soil and foliar application can improve the quality and value of grain. This is good implication for human health because these elements are required in human. Acknowledgements: Authors are very grateful to Dr. Joba Chatterjee, Department of Botany, University of Lucknow, Lucknow for her valuable help and guidance. Corresponding author is grateful to CSIR, New Delhi for financial help (ref no 21/12/2014(ii) EU-V). References: 1. Esia G. S. A and Ali T. B. Impact Spraying of Some Microelement on Growth, Yield, Nitrogenase Activity and Anatomical Features of Cowpea Plants, World Journal of Agricultural Sciences, 10(2), 57-67 (2014). 2..Marschner, H., Mineral Nutrition of Higher Plants. 2nd ed. Academic Press, Harcourt Brace Company, Publisher Lodon, pp. 313- 396 (1995). 3. Fouly, M.M., Micronutrients in arid and semiarid areas: Level in soils and plants and the need for fertilizers with reference to Egypt. Proc. 17th Colloquium of the International Potash Institute Bern, Switzerland, pp: 163173 (1983). 4.. Mortvedt, J .J., F.R. Cox, L.M. Shuman and R.M. Welch, Micronutrients in Agriculture,Published by Soil Soc. Amer. Inc. Madison, Wisconsin, USA, pp: 760 (1991). 5. Srivastava, P.C. and Gupta, U. C. Trace Elements in Crop Production. Science Pub. Inc. Lebanon, NH03766 USA, pp: 366 (1996). 6.. Piper, C. S. Soil and Plant analysis monograph from waite Agric. Res.Inst.TheUniv. Adelaide .Adelaide (1942). 7. Lowry, O. H., Rosenbrough, N. J., farr,A.L. and Randell,R.J. Protein measurement with Folin-Phenol. J. Biol. Chem. 193, 265-275 (1951). 8. Zeidan, M.S.,. Response of wheat plants (Triticum aestivum L) to different methods of Zinc fertilization in reclaimed soils of Egypt. Plant Nutrition - Food Security and Sustainability ofAgro-ecosystems (Eds W.J. Horst, et al.), Kluwer,Dordrecht, The Netherlands, pp: 1048-1049 (2001).

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Adoption of Improved Technologies in Black Gram Crop LOKENDRA KUMAR SINGH Department of Agricultural Extension Janta Vedic Collage, Baraut, Baghpat. ABSTRACT India is largest producer of pulses in world with 25 per cent share in global production. Chickpea, Pigeon Pea.MungBean, UradBean, lentil and Field Pea are important pulses crop contributing 39.00 per cent,21.00 per cent, 11.00 per cent, 7.00 per cent and 5.00 per cent to the total production of pulses in the country. The total production was estimated 14.56 million tones and an area of 23.63 million hectares with average producti vity 625kg/ha. (www agro pedia. Nic in.2015) Over 75% of the total of Black gram is generally known as dal milling or, dehulling. Milling means removal of the outer husk and splitting the grain into two equal halves. Dalmilling is one of the major food processing industries in the country, next only to rice milling. Black gram used as daal and as ingredient in snacks like idly, dosa, vada and papad etc. Adoption of Improved practice is the more essential part of increasing the production, because the population is increase day to day and holding size is decrease, so dal price is highly explosion The first week of October 2015 noted of cost of Black Gram dal 150- 180 Rupees per kg in the retails just before one year the cost October 2014 was 75-85 Rupees per kg(sources individual market survey 2015 ). We need the more production of pulses, we solve this problems by the applied more technologies in their field . Key Word : Black Gram. Improved Practice, Agriculture. Introduction No doubt India lives in the villages and about 50 per cent of the 6.41 lac villages of the country are situated in different terrain characterized by poor socio-economic condition. Even a casual glimpse at the sub continent of India is sufficient to convin ce that ours is a land of villages. Good majorities of her people i.e. nearly 68.84 per cent lives in villages and are occupied in the agriculture. According to the latest census figures, there are only 7936 towns in India; whereas the numbers of villages are 6.41 Lac. India is largest producer of pulses in world with 25 per cent share in global production. Chickpea, PigeonPea.MungBean,UradBean, lentil

and FieldPea are important pulses crop

contributing 39.00 per cent,21.00 per cent, 11.00 per cent, 7.00 pe r cent and 5.00 per cent to the total production of pulses in the country.The total production was estimated 14.56 million tones and an area of 23.63 million hectares with average productivity 625kg/ha. (www agro pedia. Nic in.2015) Over 75% of the total of Black gram is generally known as dal milling or, dehulling. Milling means removal of the outer husk and splitting the grain into two equal halves. Dalmilling is one of the major food

84

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in processing industries in the country, next only to rice milling. Black gram used as daal and as ingredient in snacks like idly, dosa, vada and papad etc. Rhyzobium bacteria are present on the root nodules of Black gram. The Black-Gram crop fixes atmospheric nitrogen in symbiotic association with Rhyzobium bacteria and maintains the soil fertility Black gram is a member of the Asiatic Vigna crop group. It is an annual pulse grown mostly as a fallow crop in rotation with rice. Similar to the other pulses, black gram, being a legume, enriches soil nitrogen content and has relatively short (90-120 days) duration of maturity. Materials and Methods’: Research methodology is of paramount importance in any scientific study as the validity and reliability of the facts depend upon the system of investigation. It provides the details of the various aspects concerning the research methodology. The scientific steps required to carry out the research are being discussed below in depth. A. Selection of the state: In 1947, when India gained independence, the state of United Provinces was renamed as Uttar Pradesh. Uttar Pradesh is the biggest of 31 States in India. Uttar Pradesh is now divided into seventy five districts under eighteen divisions . B. Selection of the district : There are 75 districts in the state of U.P. The Research was conducted in the District of Baghpat (U.P.). The District of Baghpat was purposively selected. C. Selection of the block : Baghpat District has the privilege of having six blocks namely Baghpat, Baraut, Binauli, Chaprauli, Khekra and Pilana. Out of six blocks, Baraut blocks were selected purposively keeping in view of the nature of study. D. Selection of the village : There were 54 villages in the C.D. block Baraut, but the the five villages were selected by randomly the selected villages are 1 Malakpur,Sinoli,Hilwari, Baoli and Lohari. E. Selection of the respondents : From each Villages 30 farmers were finally selected for the study. Who had adopted improved farm practices of Black-Gram.The total 150 respondents to be interviewed under the study. RESULTS AND DISCUSSION The finding of the research of investigation is presented in the following table. Adoption of Improved Agricultural Practice by the Black Gram growers. S.No.

Practice

Numbers

85

Per

cent

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in (%) 1

Knowledge about Improved variety.

2

Sowing of Improved Variety Type-9, Pusa-1&Pant-19.

3

122

81.33

102

68.00

68

45.33

82

54.64

Field Preparation (I)

By Scientific methods.- Summer Plowing, Leveling, 2Plowing of harrows &2-4 Plowing of Cultivators.

(II) By traditions methods.

4

5

Sowing time (I)

Early Time-(February-march)

86

57.33

(II)

Timely-(May-June)

64

42.66

35

23.33

22

14.66

55

36.66

(I) Correct- 10-12 kg/ha.

65

43.33

(II) In Correct 12-16kg/ha.

85

56.66

Soil treatment (I)

Weed control-At the time of field preparation and before the germination of seed. Spray 1.00kg Basalineadd 1000liter water.

(II)

Nutrient test- NO2,P2O5 ,Ca,Mg,etc

(III)

Insect treatment 5% Aldrin dust @-25 kg to add in the soil at the time of field preparation,to control the eggs and pupa of insects.

6

7

8

9

10

11

Seed rate

Seed treatment Practice (I)

Sowing treated seed(packets seed)

110

73.33

(II)

Sowing Un treated seed

40

26.66

Methods of sowing (I)

Sowing in the line (furrow method)

58

38.66

(II)

Traditions method By broad casting

92

61.33

(I)

Spring season 2-3 Irrigation apply

85

58.66

(II)

Rainy season 1-2 Irrigation apply

68

45.33

(I)

20-30 Kg Nitrogen

90

60.00

(II)

40-50 Kg Phosphorus

55

36.66

(III)

30-40 Kg potash

34

22.66

135

90.00

Irrigation

Fertilizers

Crop Protection (A)Weed Control (I)

Physical methods (by hands)

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in (II)

Chemical methods(by herbicides)

15

10.00

57

38.00

(B) Insect Control (I) To control Bihar hairy Cutter Pillar and Aphid

to spray

Indosulphan 35 [email protected]% and 800 liter water two spray to interval of 8-10days. 12

13

Harvesting of Crop (I)

Maturity of pod -75-85%

64

42.66

(II)

Maturity of pod 85-95%

86

57.33

Threshing of Crop (I)

By threshing machine

65

43.33

(II)

By Bullocks and men’s

85

56.66

Above table reveal that 81.00 per cent respondents have well awareness of about improved varieties of black, but only 68.00 per cent farmers finally apply of improved varieties on their field. In case of field preparation 54.64 per cent farmers are use of tradition methods and 45.33 per cent respondents have apply scientific methods in their field preparation of Black Gram. While the 36.66 per cent, 23.33 and 14.66 per cent respondents have apply soil treatment practice under the Insect treatment, weed control, and Nutrient test in their field before the sowing. In case of seed rate and seed treatment majority i e 56.66 per cent , 73.33 per cent respondents have adopted correct seed rate and sowing treated seed available in the market and other seed stores. Only 38.66 per centrespondents have applied scientific methods of sowingof black Gram crop, majority of respondents do not apply the scientific recommendation of sowing methods. In case of irrigation practice 58.66 per cent and 45.33 per cent respondents provide irrigation facilities their crop 2-3 irrigation in spring season and 1-2 irrigation in rainy season in lack of rain. While 60.00 per cent,36.66 per cent 22.66 per cent respondents have applied 20-30kg Nitrogen,40-50 kg

Phosphorus and 30-40 Kg Potash in their field. The

very low amount of respondents is 22.66 per cent to have applied Potashic fertilizer in their crops of Black Gram. 90.00 per cent respondents have applied physical methods ( by hands) to control weed their won standing crops and 38.00 per cent respondents have applied chemical methods recommended by the scientist to control hairy cutter pillar or aphid .While in case of harvesting and threshing of crop the majorities of i.e.57.33 per cent respondents harvest our crops when the 85-95 per cent pod mature, and 42.66 per cent respondents have harvest won crop when the pod mature is 75-85 per cent, majorities of respondents 56.66 per cent have thresh their crop by the men’s power and Bullock and 43.33 per cent respondents are applied threshing machine in duration threshing of our crops in case of Black Gram. CONCLUSION Overall majorities i.e. 81.00 per cent, respondents have knowledge about Improved varieties,68.00 per cent respondents have applied Improved varieties wne their field,57.00 per cent and 57.00 per cent,respondents adopted Early sowing and incorrect seed rate of Black –Gram, 73.00per cent and 61.00 per cen,t respondents

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in have sowing the treated seed, and were used traditional methods at the time of sowing, 60.00 per cent, and 90.00 per cent, grower apply the recommended dose of Nitrogen fertilizer and farmers control of weeds in our crop by hands helps by the Khurpy,(weeding tools) 57.00 per cent respondents harvesting of their crop when the pod mature 85-95 per cent and threshing of their crop by men hands ,Bullock and tractors. REFERENCES 1. Jeswani, L.M. and Baldev, B., (1988). Advances in Pulse Production Technology, Indian Council of Agricultural Research Publication 2. Pandey ,P.H.(1988). Principles and Practices of Post Harvest Technology, 3. Acharya, S.S.andAgarwal,N.L.(1999). Agricultural Marketing in India, 4. Chakraverty, A.(1988). Post Harvest Technology of Cereals, Pulses and Oil seeds, 5. Annual Report, 2003-2004, National Cooperative Development Corporation, New Delhi. 6. Annual Report, 2004-2005,Central Warehousing Corporation, New Delhi. 7. Agmark Grading Statistics, 2003-2004 and 2004-2005, Directorate of Marketing and, FaridabadInspection. 8. Agarwal, P.K., Agricultural Marketing, ( 2002).. Establishing Regional and Global Marketing Network for Small holders’ Agricultural Produce / Products with reference to Sanitary and Phytosanitary (SPS) Requirement, PP.15-23

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Study on SO2 Induced Effects and their Amelioration in Arhar (Cajanus cajan) HARENDRA MALIK, NARESH KUMAR AND RENU CHOUDHARY* Department of Botany, C.C.R. (P.G.) College, Muzaffarnagar (U.P.), India. * Department of Biology, G.G.I.C. Gagalheri, Saharanpur(U.P.), India. Email- [email protected] Abstract Sulphur dioxide is a well known wide spread air pollutant and most harmful to plants. The main aim of present investigation is to study the extent and magnitude of damage induced by sulphur dioxide and amelioration of this damage with the help of some chemical agents like calcium hydroxide and sodium benzoate on Cajanus cajan cv. Manak. The present study was conducted to observe the impact of exposure of four different concentrations of sulphur dioxide i.e. 653, 1306, 2612 and 3918 µg m-3 alone and after calcium hydroxide treatment and sodium benzoate treatment on Arhar (Cajanus cajan) cv. Manak. Exposure to SO

2

inhibited the seed germination, foliar injuries, root, shoot and whole plant growth, chlorophyll a, chlorophyll b and carotenoids contents of leaves. However, the accumulation of anthocyanin and total phenolics contents of leaves were increased significantly in higher SO concentrations. After treatment of two ameliorating agents i.e. 2

calcium hydroxide and sodium benzoate, better growth and development was observed in SO treated plants. 2

Keywords : Pollutant, Sulphur dioxide, Amelioration, Germination, Growth, Calcium hydroxide, Sodium benzoate, Arhar. Introduction Major air pollutants are oxides of sulphur (SOx), oxides of nitrogen (NOx), carbon monoxide (CO), hydrogen sulphide (H2S), hydrocarbons, ozone (O3 ), fluorides, lead, mercury and particulates etc. Excessive quantity of these all air pollutants can impair normal healthy life of all organisms. Amongst these air pollutants oxides of sulphur including sulphur monoxide (SO), sulphur dioxide (SO2), sulphur trioxide (SO3), sulphur tetra oxide (SO4), sulphur sesquioxide (S 2O3) and sulphur heptaoxide (S 2O7). SO2 is one of most toxic air pollutant and major sources of SO2 emission are automobile exhaust, burning of fossil fuels in thermal power plants, smelting industries, other process such as manufacture of sulphuric acid and fertilizers and refining of crude petroleum. The hazardous effects of SO2 create a big threat to the survival and sustenance of the living systems. As the total control of SO2 air pollution is technically and economically not feasible, various crop management practices, such as nutrient supplementation and spraying of chemical protectants have been used to reduce air pollution injury in plats. Materials and Methods Germination studies were carried out on selected plants. Seeds were selected uniformly (criteria being the size and colour of seeds). For long-term treatments the exposure to SO2 was initiated when the seedlings were 11 days and stop at 60 days old plants. The seedlings of each cultivars of the plants were grouped into thirteen research plots so that four of them treated with. 653, 1306, 2612 and 3918 µg m-3 concentrations of

89

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in SO2, remaining four treated with these four SO2 concentration and then sprayed with calcium hydroxide (denoted by TC ) and last four sprayed with sodium benzoate after SO2 treatments (TS), while the last one was set aside as control. The plants were given the treatments of SO2 for 2 h. on alternate days. All the normal agronomic practices, other than fertilizer and pesticide application, were followed. Chlorophyll, carotenoid, anthocyanin and phenolics content calculated using several deeds [1, 6, 7 & 9]. Result and Discussion Exposure to SO2 inhibited the seed germination of the Cajanus cajan cv. Manak. Reduction in seed germination percentage was as 0.96, 5.73, 11.20 and 16.02 percent under 653, 1306, 2612 and 3918 ug m-3 concentration of SO2, respectively. Exposure of seeds to varying concentrations of SO2 with treatment of calcium hydroxide (Tc) and sodium benzoate (Ts) exhibited lesser reduction as compared to SO2 alone. These results are in agreement with the findings of [3,11] in different plants. Table 1. Seed germination percentage, mean germination percentage and seedling survival percentage of Cajanus cajan cv. Manak exposed to different concentrations of SO 2 alone and treated with calcium hydroxide and sodium benzoate (Tc and Ts). Parameters

SO2 treatment (µg m-3) Tc/Ts

Control

653

1306

2612

3918

-

94.24

93.33

88.84

83.68

79.14

germination

Tc

-

93.85

91.23

88.42

86.04

percentage

Ts

-

93.92

91.86

89.28

86.57

-

20.86

20.24

19.63

18.83

17.88

germination

Tc

-

20.46

20.13

19.56

19.01

percentage

Ts

-

20.49

20.18

19.63

19.07

Seedling

-

87.82

86.78

85.58

84.24

82.12

survival

Tc

-

87.08

86.73

86.32

85.89

percentage

Ts

-

87.15

86.81

86.54

86.11

Seed

Mean

Plants exposed to varying concentrations of SO2 and with calcium hydroxide and sodium benzoate showed detectable foliar injuries. Such type of observations were also made by [2,13]. In 30 days old plants, chlorotic patches at the margin of the lamina and at the tips of the lower leaves were exhibited. But when the SO2 treatment duration was prolonged these patches become dark brown bifacial necrotic lesions. The injury was mostly confined in mature leaves and new leaves develop normally or less affected. The extent of injury increased with increasing concentration of SO2.

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Table 5

:

Growth responses of 30 days old plants of Cajanus cajan cv. Manak exposed to different concentrations of SO2 alone and treated with calcium hydroxide and sodium benzoate (Tc and Ts).

Parameters

Plant height (cm)

Length of root (cm)

Length of shoot (cm)

Fresh weight of root (g)

Fresh weight of shoot (g)

Dry weight of root (g)

Dry weight of shoot (g)

No. of leaves per plant

No. of primary branches per plant

No. of nodules per plant Dry matter production (phytomass accumulation) Net primary productivity (g/plant/ day)

Cajanus cajan cv. Manak SO2 treatment (g m-3) 1306 2612

Tc Ts

Control

-

89.66+4.72

85.88+4.19*

79.32+2.78**

70.19+3.34**

64.31+3.73**

Tc

-

87.37+4.42

82.00+3.76*

77.89+4.17**

75.86+3.42**

Ts

-

87.87+5.11

82.60+4.63*

78.64+3.25**

76.56+3.35**

-

13.28+1.24

12.62+1.47

11.76+1.19*

10.48+1.51**

9.42+1.40**

Tc

-

12.78+1.31

12.08+1.28*

11.58+1.22*

11.09+1.33**

Ts

-

12.84+1.54

12.17+1.37*

11.76+1.13*

11.18+1.52**

-

76.38+3.48

73.26+2.72*

67.56+1.59**

59.71+1.83**

54.89+2.33**

Tc

-

74.59+3.11

69.92+2.48*

66.31+2.95**

64.77+2.09**

Ts

-

75.03+3.57

70.43+3.26*

66.88+2.12**

65.38+1.83**

-

3.94+0.62

3.79+0.71

3.52+0.56*

3.08+0.80**

2.68+0.59**

Tc

-

3.83+0.58

3.63+0.42

3.49+0.74*

3.28+0.34**

Ts

-

3.85+0.62

3.67+0.33

3.53+0.67*

3.33+0.28**

-

24.57+1.22

23.44+1.51

21.84+1.27*

19.26+1.38**

17.13+1.12**

Tc

-

23.76+1.36

22.68+1.45*

21.18+1.60*

20.11+1.58**

Ts

-

23.84+1.43

22.81+1.66*

21.37+1.83*

20.35+1.72**

-

0.436+0.072

0.428+0.063

0.396+0.048*

0.347+0.039**

0.303+0.045**

Tc

-

0.431+0.041

0.403+0.052*

0.371+0.044*

0.354+0.038*

Ts

-

0.432+0.069

0.408+0.029*

0.379+0.061*

0.359+0.074*

-

3.88+0.58

3.75+0.36

3.48+0.61*

3.07+0.49**

2.73+0.35**

Tc

-

3.80+0.52

3.68+0.31

3.42+0.52*

3.11+0.48**

Ts

-

3.81+0.47

3.71+0.41

3.48+0.73*

3.19+0.41**

-

56.08+1.29

54.22+1.15

51.92+1.24**

46.73+1.36**

42.41+1.46**

Tc

-

55.47+0.95

53.61+1.31*

50.64+1.27**

49.09+1.31**

Ts

-

55.80+0.89

53.93+1.21*

50.97+0.98**

49.78+1.24**

-

7.10+1.48

6.96+1.53

6.77+1.34

6.40+1.46**

6.19+1.72**

Tc

-

7.01+1.31

6.82+1.18

6.68+1.12*

6.58+1.28*

Ts

-

7.02+1.26

6.84+1.22

6.71+1.08*

6.63+1.19*

-

24.52+1.89

23.92+2.16

21.83+2.25*

20.63+1.68**

19.80+1.51**

Tc

-

24.08+1.69

22.65+1.58*

22.09+1.46**

21.67+1.36*

Ts

-

24.13+1.82

22.81+1.76*

22.15+1.70**

21.82+1.22*

-

4.31+1.30

4.17+0.99

3.87+1.09*

3.41+0.88**

3.03+0.80**

Tc

-

4.23+1.21

4.08+0.83

3.79+0.96*

3.46+0.86**

Ts

-

4.23+0.88

4.11+0.70

3.85+1.34*

3.54+1.15**

-

0.143

0.139

0.129

0.113

0.101

Tc

-

0.141

0.136

0.126

0.115

Ts

-

0.141

0.137

0.128

0.118

653

3918

Values are in mean + standard deviation Significance of difference from control; P* < 0.05; P** < 0.01 and  non-significant

It was observed that root, shoot and whole plant growth reduced in SO2 exposures. At SO2 concentration of 653, 1306, 2612, and 3918 µg m-3, the reduction percentage in plant height were 4.21, 11.53, 21.71, and 28.27 in 30 days old plants. In Tc plants, the reduction were 2.55, 8.54, 13.12 and 15.39 percent similarly in Ts plants, these reductions were 1.99, 7.87, 12.29 and 14.61 percent, respectively. Similar pattern of reduction was observed in plant height of 60 days old plants. The reduction values for root length were 4.96, 11.44, 21.08 and 29.06 percent, respectively in 30 days old plants. In comparison to control, 60 days old plants exhibited maximum reduction in root length at 3918 µg m-3 concentration of SO2.

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:

Growth responses of 60 days old plants of Cajanus cajan cv. Manak exposed to different concentrations of SO2 alone and treated with calcium hydroxide and sodium benzoate (Tc and Ts).

Parameters

Plant height (cm)

Length of root (cm)

Length of shoot (cm)

Fresh weight of root (g)

Fresh weight of shoot (g)

Dry weight of root (g)

Dry weight of shoot (g)

No. of leaves per plant

No. of primary branches per plant

No. of nodules per plant Dry matter production (phytomass accumulation) Net primary productivity (g/plant/ day)

Cajanus cajan cv. Manak SO2 treatment (g m-3) 1306 2612

Tc Ts

Control

-

175.21+6.01

168.90+6.93*

153.99+6.50**

138.40+6.18**

123.32+4.98**

Tc

-

171.81+7.25

160.27+5.92*

150.30+5.68**

147.34+5.26**

Ts

-

172.25+6.45

160.74+5.57*

151.22+5.83**

149.67+5.56**

-

22.86+1.33

21.46+1.81*

19.68+1.52**

17.76+1.55**

15.95+1.37**

Tc

-

22.13+1.78

20.54+1.47*

19.72+1.14**

19.00+1.49**

Ts

-

22.19+1.69

20.62+1.63*

19.81+1.68**

19.15+1.65**

-

152.35+4.68

147.44+5.12*

134.31+4.98**

120.64+4.63**

107.37+3.61**

Tc

-

149.68+5.47

139.73+4.45*

130.58+4.54**

128.38+3.77**

Ts

-

150.06+4.76

140.12+3.94*

131.41+4.15**

130.52+3.91**

-

4.13+0.88

3.96+0.73

3.68+0.56*

3.10+0.60**

2.81+0.83**

Tc

-

4.04+0.58

3.81+0.76

3.56+0.47*

3.43+0.74**

Ts

-

4.06+0.81

3.83+0.67

3.59+0.51*

3.44+0.69**

-

65.42+2.36

62.34+2.41*

56.81+2.48**

50.73+2.58**

46.09+2.33**

Tc

-

63.85+2.53

59.44+2.54*

56.65+2.64**

54.63+2.29**

Ts

-

63.91+2.23

59.68+2.61*

56.82+2.50**

54.76+2.46**

-

0.762+0.037

0.749+0.056

0.676+0.073**

0.589+0.064**

0.519+0.037**

Tc

-

0.753+0.048

0.698+0.075*

0.668+0.052**

0.623+0.033**

Ts

-

0.754+0.063

0.703+0.067*

0.673+0.049**

0.626+0.041**

-

12.65+0.73

11.89+0.69*

10.74+0.86**

9.31+0.65**

8.66+0.47**

Tc

-

12.09+0.57

11.29+0.61*

10.54+0.58**

9.80+0.76**

Ts

-

12.14+0.82

11.35+0.90*

10.66+0.72**

9.89+0.63**

-

193.11+5.61

186.34+5.19*

161.58+4.87**

129.36+3.54**

103.65+4.66**

Tc

-

188.45+4.33

171.92+5.26*

149.88+3.71**

133.51+3.51**

Ts

-

189.72+3.93

172.69+4.36*

151.43+5.12**

139.72+3.28**

-

12.41+2.27

12.08+2.16

11.51+1.96*

10.89+2.11**

10.35+2.46**

Tc

-

12.21+1.53

11.91+1.32

11.55+1.14*

11.29+1.81*

Ts

-

12.23+1.37

11.93+1.28

11.58+1.21*

11.38+1.64*

-

34.52+2.11

32.21+1.79*

29.49+2.31**

26.88+1.77**

25.02+1.56**

Tc

-

33.09+1.93

31.28+1.81*

29.53+1.69**

28.35+1.65**

Ts

-

33.19+2.25

31.44+2.07*

29.68+1.62**

28.79+1.72**

-

13.41+1.10

12.63+1.25

11.41+1.56*

9.89+1.20**

9.17+0.84**

Tc

12.84+1.05

11.98+1.36*

11.20+1.10*

10.42+1.09**

Ts

12.89+1.45

653

3918

12.05+1.57*

11.33+1.21*

10.51+1.04**

-

0.223

0.210

0.190

0.164

0.152

Tc

-

0.214

0.199

0.186

0.173

Ts

-

0.214

0.200

0.188

0.175

Values are in mean + standard deviation Significance of difference from control; P* < 0.05; P** < 0.01 and  non-significant

The reductions in shoot length in 30 days old plants were 4.08, 11.54, 21.82 and 28.13 percent, respectively. In Tc plants reduction was observed upto 15.20 percent at 3918 µg m-3 SO2 concentration. Corresponding values in Ts plants were 14.40 percent. Reductions in fresh weight of shoot were 30.28, 18.15 and 17.17 percent against 3918 µg m-3 concentration of SO2 alone, Tc and Ts plants, respectively in 30 days old plants. 60 days old plants also showed similar pattern of reduction in fresh weight of shoot. Reductions in dry weight fractions of both root and shoot were recorded in all treated plants. Similar findings were also made by [12,10,4,11] in Vigna radiata.

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Chlorophyll a, chlorophyll b and total chlorophyll contents of leaves reduced considerably in all the treatments. The reduction percentage in 30 days old plants in total chlorophyll content were 16.74, 9.88 and 9.10 in 3918 µg m-3 of SO2 treated, Tc and Ts plants, respectively. Almost similar trend of reduction was observed in 60 days old plants. Carotenoids, the accessory pigments also exhibited reduction in their content in all treatments. Such studies relate to the findings of [8,5]. SO2 induced the accumulation of anthocyanin. Both, the age of plant and concentration of fumigant (SO2) were found to have a direct relation with the amount of anthocyanin accumulated in the cultivars. The increase in anthocyanin content was found to be maximum at 3918 µg m-3 of SO2 treated plants. In comparison to control, total phenolic contents of leaves were increased significantly in higher SO2 concentrations. In 60 days old plant, percent increase in total phenolic contents, were 64.33, 29.72 and 31.46 percent in 3918 µg m-3 of SO2 treated, Tc and Ts plants, respectively. cv. T-21

Chlorophyll content (mg/g f.wt.)

1.2 1.0 0.8 0.6 0.4

Control 2612  g m -3 Tc Ts

Control 3918  g m -3 Tc Ts

Control 2612 g m -3 Tc Ts

Control 3918 g m -3 Tc Ts

Control 1306  g m -3 Tc Ts

Control 653  g m -3 Tc Ts

0.2

cv. Manak

Chlorophyll content (mg/g f.wt.)

1.2 1.0 0.8 0.6 0.4

Control 1306 g m -3 Tc Ts

Control 653 g m -3 Tc Ts

0.2

Chl. a Chl. b

Figure 2 :

Effect of different concentrations of SO 2alone, after calcium hydroxide treatment (Tc) and sodium benzoate treatment (Ts) on Chl. a, Chl. b and total chlorophyll content of 30 days old plants of Cajanus cajan cv. T-21 and Manak.

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1.4

Chlorophyll content (mg/g f.wt.)

1.2 1.0 0.8 0.6 0.4

Control 2612 g m -3 Tc Ts

Control 3918 g m -3 Tc Ts

Control 2612 g m -3 Tc Ts

Control 3918 g m -3 Tc Ts

Control 1306 g m -3 Tc Ts

Control 653 g m -3 Tc Ts

0.2

cv. Manak 1.4

Chlorophyll content (mg/g f.wt.)

1.2 1.0 0.8 0.6 0.4

Chl. a

Control 1306 g m -3 Tc Ts

Control 653 g m -3 Tc Ts

0.2

Chl. b

Figure 3 :

Effect of different concentrations of SO 2alone, after calcium hydroxide treatment (Tc) and sodium benzoate treatment (Ts) on Chl. a, Chl. b and total chlorophyll content of 60 days old plants of Cajanus cajan cv. T-21 and Manak.

Figure 4 :Effect of different concentrations of SO 2 alone, after calcium hydroxide treatment (Tc) and sodium benzoate treatment (Ts) on carotenoid content (mg/g f.wt.) of 30 and 60 days old plants of Cajanus cajan cv. Manak.

30 Days Carotenoid content (mg/g f.wt.) 0.5 0.45

Series1

0.4 0.35

Series2

0.3

0.25 0.2

Series3

0.15 0.1

Series4

0.05 0 1

2

3

60 Days Carotenoid content (mg/g f.wt.)

94

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0.7 Series1

0.6 0.5

Series2

0.4 0.3

Series3

0.2 0.1

Series4

0

1

2

3

4

Figure 5 : Effect of different concentrations of SO2 alone, after calcium hydroxide treatment (Tc) and sodium benzoate treatment (Ts) on anthocyanin content (mg/g f.wt.) of 30 and 60 days old plants of Cajanus cajan cv. Manak. 30 Days Anthocyanin content (mg/g f.wt.) 0.4 0.35 Series1

0.3 0.25

Series2

0.2 0.15

Series3

0.1 Series4

0.05 0 1

2

3

95

4

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0.7 0.6

Series1

0.5 Series2

0.4 0.3

Series3

0.2 0.1

Series4

0 1

2

3

4

Figure 6 : Effect of different concentrations of SO2 alone, after calcium hydroxide treatment (Tc) and sodium benzoate treatment (Ts) on phenolics content (mg/g f.wt.) of 30 and 60 days old plants of Cajanus cajan cv. Manak.

30 Days Phenolics content (mg/g f.wt.) 3.5 Series1

3 2.5

Series2

2 1.5

Series3

1 Series4

0.5 0

1

2

3

4

60 Days Phenolics content (mg/g f.wt.)

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4 3.5

Series1

3 Series2

2.5 2

Series3

1.5 1

Series4

0.5 0 1

2

3

4

Conclusion The present study was conducted to observe the impact of exposure of four different concentrations of sulphur dioxide i.e. 653, 1306, 2612 and 3918 µg m-3 alone and after calcium hydroxide treatment and sodium benzoate treatment on Arhar (Cajanus cajan) cv. Manak. The effect of SO2 alone and with treatments of chemicals (calcium hydroxide and sodium benzoate) were studied in respect of seed germination, visible foliar injury, different growth parameters and some biochemical changes like chlorophyll content, carotenoid content, anthocyanin content and phenolics content. Thus, all the treated plants under study were sensitive to SO2 pollution, though their responses vary with SO2 concentration, age of plant and stage of development. SO2 causes reduction in all growth and biochemical components (except anthocyanin and phenolics content). After treatment of two ameliorating agents i.e. calcium hydroxide and sodium benzoate, better growth was observed in all Tc and Ts plants under study. References 1.

Arnon, D.I. Copper enzymes in isolated chloroplasts, polyphenol oxidase in Beta vulgaris. Plant Physiol. 24 : 1-15 (1949).

2.

Ayer, S.K. and Bedi, S.J. Effect of artificial fumigation of sulphur dioxide on growth and yield of Zea mays L. var. American Sweet Corn. Pollut. Res. 9 : 33 -37 (1990).

3.

Goswami, R. Toxicity of air pollution to plants. A. Ph.D. Thesis, C.C.S. University, Meerut, India (2002).

4.

Jeyakumar, M., Jayabalan, N. and Arockiasamy, D.I. Effect of sulphur dioxide on maize (Zea mays L.) var. (CO-1) seedlings at lethal dose 50. Physiol. Mol. Biol. Plants. 9 (1) : 147-151(2003).

5.

Joshi, P.C. and Swami, A. Air pollution induced changes in the photosynthetic pigments of selected plant species. J. Environ. Biol. 30(2) : 295-298 (2009).

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Maclachlan, S. and Zalik, S. Plastid structure, chlorophyll concentration and free amino acid composition of chlorophyll mutant of barley. Can. J. Bot. 43 : 1053-1062 (1963).

7.

Mancinelli, A.L., Yang, C.P.H., Lindquist, T., Anderson, O.R. and Robion, I. Photocontrol of Anthocyanin III. The action of streptomycin on synthesis of chlorophyll and anthocyanin. Plant Physiol. 155 : 251-257 (1975).

8.

Ranieri, A., Pieruccetti, F., Panicucci, A., Castagna, A., Lorenzini, G. and Soldatini, G.F. SO 2-induced decrease in photosynthetic activity in two barley cultivars. Evidence against specific damage of the protein-pigment complex level. Plant Physiol. Biochem. 37 : 919-929 (1999).

9.

Sadasivam, S. and Manickam, A. Phenolics : In biochemical methods for agricultural sciences. Wiley Eastern Limited, New Delhi, India. pp. 187-189 (1992).

10. Saxena, D.K., Saxena, A., Gupta, P. and Kamakshi. Effect of short-term fumigation of SO2 on seedling growth of Cicer arietinum L. J. Indian Bot. Soc. 80 : 63–66 (2001). 11. Tyagi, A. Sulphur dioxide damage to plants. A Ph.D. Thesis, C.C.S. University, Meerut, India (2006). 12. Verma, M. and Agrawal, M. Response of wheat plants to sulphur dioxide and herbicide interaction at different fertility regiones. J. Indian Bot. Soc. 80 : 67-72 (2001). 13. Wang, C., Da Xing, D., Zeng, L., Ding, D. and Chen. Effect of artificial acid rain and SO 2 on characteristics of delayed light emission. Luminescence. 20 : 51-56 (2005).

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Synthesis, Characterization and Biological Activities of Some Metal Complexes with 2Amino -4- (p- Ethoxy Phenyl) Thiazoline Ligand Dinkar Malik Department of Chemistry, M. S. College, Saharanpur U.P. Corresponding address: Dr. Dinkar Malik, 1800/1, Mission Compound, Saharanpur-247001 (U.P.) Email- [email protected] ABSTRACT The ligand complexes of Ni(II), Co(II) and Cu(II) with 2 - amino -4- (p- ethoxy phenyl) thiazoline have been synthesized and characterized with the help of their elemental analysis, IR, electronic and magnetic susceptibility studies. From the analytical and spectral data the stoichiometry of these complexes have been found to be of the type ML2X2 (where M = Cu (II), Co (II) and Ni (II)}. It is found that Ni(II), Cu(II) and Co(II) complexes exhibit octahedral and square planar geometry. The fungicidal activities of ligands and metal complexes were screened by growth method against various fungi i.e. Drechslere setramera, Fusarium oxyporum, Macrophomera phaseoli at different concentrations. It is found that the activity decreases with decrease of concentration and the metal complexes are less toxic than the parent ligand. Key Words: Thiazoline Complexes, Fungicidal Activity, Heterocyclic Compounds, Toxicity,

Fungicidal

activity, INTRODUCTION Complexes of transition metals ions containing ligands with N, S and N, S, O donors are known to exhibit interesting stereo chemical, electrochemical and electronic properties. Semicarbazones and thio semicarbazones are amongst the most widely studied nitrogen and oxygen/sulphur donor ligands. Besides, thio semicarbazones, in the last two decades, have emerged as an important class of sulphur ligands particularly for transition metal ions . The real impetus towards developing their co-ordination chemistry i.e. their physiochemical properties and significant biological activities. Thiazolines and their derivatives have created an interest due to their wide range of activity. Thiazole and their derivatives possess anti-malarial, anti-theminitic, anti-fungal, anti-bacterial, anti-sparodic and anti-tubercular activities. Such compounds can also be used as local anaesthetic, anti-radiation drugs,anti-viral and anti-protozoan agents and also in the rubber industry as vulcanization accelerators. Transition metal complex formed by organic ligands are essential in plant nutrition, they have been studied which induced several amines containing sulphur and mercaptoacetates. The survey of literature revealed that metal complex play an important role in biological activity of drugs. The mechanisms of the activity of such drugs have been explained on the basis of complex formation. Attempts have been made to study their structure with the help of elemental analysis, megnatic mesurments, spectral studies and conductance measurements. The synthesis, spectral characterization and biological activity of Schiff’s base derived metal complexes were studied by many workers[1-3]. Similar experiments on fungicidal and antimicrobial activites of Cu (II), Co (II) and Ni (II) Complexes with O, N, and S donor, their EPR and electronic spectral studies were also conducted by many workers [4-8] Schiff’s base derived complexes of derivatives of DHA, their spectra and synthesis under microwave irradiation were also studied by many workers [9.10]. The present paper deals with the preparation

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Complexes

%Calc./ Obs.

C23H20ON2S [Cu(C23H20ON2S)2Cl2]

[Ni(C23H20ON2S)2Cl2]

[Co(C23H20ON2S)2Cl2]

[Cu(C23H20ON2S)2(CH3COO)2] -

[Ni(C23H20ON2S)2(CH3COO )2] [Co(C23H20ON2S)2(CH3COO-)2]

C

H

S

N

O

M

74.19

5.37

8.60

7.52

4.30

-----

74.17

5.31

8.56

7.51

4.29

62.83

4.55

7.28

6.37

3.64

7.22

62.73

4.52

7.26

6.33

3.61

7.21

63.17

4.56

7.35

6.42

3.65

6.77

63.14

4.52

7.29

6.39

3.61

6.73

63.15

4.57

7.32

6.40

3.66

6.75

63.13

4.52

7.26

6.39

3.64

6.71

64.82

4.97

6.91

6.05

10.37

6.87

64.76

4.94

6.90

6.02

10.34

6.78

65.16

4.98

6.97

6.11

10.45

6.43

65.14

4.94

6.95

6.05

10.39

6.41

65.14

4.99

6.94

6.09

10.42

6.40

65.12

4.92

6.91

6.06

10.36

6.38

The ligand 2-amino-4-(p-ethoxy phenyl) thiazoline was prepared using the procedure reported in the literature[14].

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TABLE 2 CHARACTERISTIC IR BANDS OF LIGANDS AND COMPLEXES IR Bands (cm-1)

Complexes

C23H20ON2S

νN-H

νC-S

νC-H

νC=C

νC=N

νM-S

3468-

834-682

3090-

1620-

1639-

--

3082

1590

1620

3108-

1623-

1633-

3098

1592

1618

3106-

1621-

1630-

3099

1597

1619

3105-

1618-

1627-

3097

1599

1611

3107-

1602-

1628-

3091

1582

1609

3103-

1618-

1632-

3094

1599

1607

3104-

1604-

1625-

3096

1597

1613

3286 [Cu(C23H20ON2S)2Cl2]

3381-

855-690

3262 [Ni(C23H20ON2S)2Cl2]

3391-

859-677

3264 [Co(C23H20ON2S)2Cl2]

3399-

860-683

3283 [Cu(C23H20ON2S)2(CH3COO)2]

3406-

866-690

3287 [Ni(C23H20ON2S)2(CH3COO-)2]

3396-

853-684

3267 -

[Co(C23H20ON2S)2(CH3COO )2]

3391-

867-685

3276

315-309

326-318

335-322

319-315

330-321

338-325

A shift in the νC-S and νN-H band frequencies is observed in all the complexes. This shows that the lone pair of electron presents on the sulphur atom of thiazoline ring and nitrogen atom of free amino group is taking part in co-ordination (Table 2).

TABLE 3 (a) ELECTRONIC REFLECTANCE SPECTRAL DATA AND THEIR ASSIGNMENTS OF NI(II) COMPLEX ν1

ν2

ν3

Dq

B

ν2/ ν1

ν3(Calc.)

[Ni(C23H20ON2S)2Cl2]

8523

14518

24448

1285.6

600

1.70

28966

[Ni(C23H20ON2S)2(CH3COO)2]

8515

14511

24417

1283.5

662.4

1.70

28997

Complexes

ν1 = 3A2g (F) → 3T2g (F), ν2 = 3A2g (F) → 3T1g (F) and ν3 = 3A2g (F) → 3T1g (P). (b) ELECTRONIC REFLECTANCE SPECTRAL DATA AND THEIR ASSIGNMENTS OF CO(II) COMPLEX Complexes

ν1

ν2

ν3

101

Dq

B

ν2/ ν1

ν2(Calc.)

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in [Co(C23H20ON2S)2Cl2]

8810

14245

20080

1198.1

667

1.61

27226

[Co(C23H20ON2S)2(CH3COO)2]

8798

14242

20081

1195.4

666

1.61

27199

ν1 = 4T1g (F) → 4T2g(F), ν2 = 4T1g (F) → 4 A2g (F) and ν3 = 4T1g (F) → 4T1g (P). (c) ELECTRONIC REFLECTANCE SPECTRAL DATA AND THEIR ASSIGNMENTS OF CU(II) COMPLEX ν1

ν2

ν3

Dq

B

ν2/ ν1

ν2(Calc.)

[Cu(C23H20ON2S)2Cl2]

15320

19118

--

--

--

--

--

[Cu(C23H20ON2S)2(CH3COO)2]

15325

19124

--

--

--

--

--

Complexes

ν1 =2B1g → 2A1g and ν2 = 2B1g → 2Eg CZ-record UV-Viz. spectrometer provided with an automatic recorder was used to record the electronic spectra of the complexes in ethanol at room temperature (Table 3). PREPARATION OF METAL COMPLEXES In general all these complexes were synthesized by refluxing the respective metal salts with ligand 2-amino-4(p-ethoxy phenyl) thiazoline in 1:2 molar ratio in ethanolic medium on water bath for one hour. The solution was concentrated to half of its volume then it was kept for some time. The crystals of complexes separated out which were filtered, washed with alcohol and dried in vacuum. Similarily some complexes of thiazoline were also synthesized by many workers [15-21]. RESULTS AND DISCUSSION Adducts of all the complexes were prepared by refluxing the respective metal salts with ligands in 1:2 molar ratio in ethanolic medium. The crystals of complexes separated out which were filtered, washed with alcohol and dried in vacuum. IR Studies: The ѵ (C=N) band frequencies in the free ligand are completely unaffected on complexation. The unchanged position of the band indicates that the ring nitrogen does not take any part in the coordination. The band observed at 834 cm-1 in the free ligand assigned to asymmetric ѵ(C-S) is shifted to lower frequency after complexation. But the symmetric ѵ (C-S) frequency completely disappears or intensity of this band is reduced after complexation. These facts confirm that the ring sulphur is taking part in complex formation. The ѵ(N-H) asymmetric and symmetric stretching frequencies appearing in the region 3468 and 3286 cm-1 respectively, also decreases in the complex. This shows that the lone pair of electron available on nitrogen atom took part in coordination. From the above observation it is clear that the nitrogen of the –NH2 group and ring sulphur take part in coordination. Electronic Reflectance Spectral Studies:

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in In the electronic spectra of Ni (II) complexes three bands at 8515-8523, 14511-14518 and 24417-24448 cm-1 were observed which may be assigned for 3 A2g (F) → 3T2g (F) (ν1), 3 A2g (F) → 3 T1g (F) (ν2) and 3A2g (F) → 3T1 g (P) (ν3) which are characteristic of octahedral Ni(II) ion. The magnetic moment values are found in the range 2.90-3.20 B.M. This is in support of high spin octahedral complex. The value is however is raised only to a small extent suggesting that the splitting is weak and that the environment is quite close to an octahedral one [22]. The observed value of magnetic moment is found in the range 2.97-3.55 B.M. which is expected for octahedral Co(II) complex. Three bands were observed at 8798-8810, 14242-14245 and 20080-20081 cm-1 which may be assigned to 4T1g (F) → 4 T2g (F) (ν1), 4T1g (F) → 4 A2g (F) (ν2) and 4 T1g (F) → 4 T1g (P) (ν3) respectively for octahedral complexes. Two bands were observed in the electronic spectra of Cu (II) complexes in the region 15320-15325 and 1911819124 cm-1 which may be assigned to 2B1g → 2A1g and 2B1g → 2Eg respectively in a planar field. The magnetic moment value foe the Cu (II) complexes lie in the range 1.54-1.58 B.M. which support square planar geometry. The fungicidal activities of the ligand as well as of metal complexes were screened against different fungi at different concentrations 100, 50 and 20 ppm in Czapek’s dox agar medium. It has been observed that the fugitoxicity of the metal complexes are lesser than the free ligand. This might be due to the fact that the group which is responsible for toxicity is not free in complexes due to co-ordination however it is free in ligand. The ligand as well as the metal complexes is most toxic at higher concentration i.e. the fungicidal activity decreases with the decrease of concentration. REFERENCES 1.

Khamamkar Ashwini and Pallapothula Rao Venkateshwar. Synthesis, Spectral Characterization and Biological activity of Schiff’s base derived metal complexes. J. Ind. Council Chem., 29(1&2) 71-76 (2012)

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Mane P.S., Shirodkar S.G., Arbad B.R. and Chondhekar T.K. Ind.. J. Chem, Sec A; Inorganic, Bio-inorganic, Physical & Analytical Chemistry, 40A(6) 648(2001)

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Malik Dinkar, Yadav Punam, Kumar Sandeep and Malik Vijai Studies on Structural and biological aspects of transition metal complexes of the ligand 2-amino-4-(p-hydroxy phenyl) thiazole. Discovery Pharmacy, 5(15) 15-17 (2013)

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Chandra S. and Sangeetika J., EPR and electronic spectral studies on copper(II) complexes of some N-O donor ligands, J. Indian Chem. Soc., (81) 203(2004)

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Shriodkar S.G., Mane P.S. and Chondhekar T.K. Synthesis and fungitoxic studies of Mn(II), Co(II), Ni(II) and Cu(II) with some heterocyclic Schiff base ligands, Indian J. Chem, (40A) 11141117(2001)

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Belaid S., Landreau A., Benali-Baitich O., Khan M.A. and Bouet G., Synthesis, characterisation and antifungal activity of a series of cobalt(II) and nickel(II) complexes with ligands derived from reduced N, N'-ophenylenebis (salicylideneimine), Trans. Met. Chem., (33) 511(2008)

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Mapari A.K. and Mangaonkar K.V. Synthesis, Characterization and Antimicrobial Activity of Mixed Ligand Complexes of N-(2-ethoxy-1-naphthylidene)-2,6-diisopropylaniline and N-(2ethoxybenzylidene)-2,3-dimethylaniline with Co(II), Ni(II), Cu(II) and Zn(II) ions. International Journal of ChemTech Research., 3(2) 636-641(2011)

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Ravanasiddappa M., Sureshg T., Syed K., Radhavendray S. C., Basavaraja C. and Angadi S. D., Transition

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di-iminoazine:

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Manch W. and Conard Fernelius W., The Structure and Spectra of Nickel(II) and Copper(II) Complexes, Journal of Chemical Education., 38 (4) 192-201(1961)

10. Naik B. and Desai K., Novel approach for the rapid and efficient synthesis of heterocyclic Schiff bases and azetidinones under microwave irradiation, Indian journal of chemistry., (45B) 267271(2006) 11. Bharti S.K., Nath G., Tilak R. and Singh SK. Synthesis, anti-bacterial and anti-fungal activities of some novel Schiff bases containing 2,4-disubstituted thiazole ring. Eur J Med Chem., (45) 651-660 (2010) 12. Vogal A.I. Quantitative Organic Analysis., (1958) 13. Vogal A.I. A Text Book of Quantitative Inorganic Analysis. 3rd ed. (English Language Book Society and Longman)., (1961) 14. Dodson R.M. and King L.C. The reaction of ketones with halogens and thiourea J. Am. Chem. Soc., (67) 2242 (1945) 15. Adibpour N., Khalaj A. and Rajabalian S. Synthesis and antibacterial activity of isothiazolyloxazolidinones and analogous 3(2H)-isothiazolones. Eur J Med Chem., (45) 19-24 (2010 ) 16. Aridoss G., Amirthaganesan S., Kim M.S., Kim J.T. and Jeong Y.T. Synthesis, spectral and biological evaluation of some new thiazolidinones and thiazoles based on t-3-alkyl-r-2, c-6diarylpiperidin- 4-ones. Eur J Med Chem., (44) 4199-4210 (2009) 17. Arshad A., Osman H., Bagiey M.C., Lan C.K., Mohamad S., Safirah A. and Zahariluddin M. Synthesis and antimicrobial properties of some new thiazolyl coumarin derivatives. Eur J Med Chem.,1-7 (2011) 18. Dawane B.S., Konda S.G., Mandawad G.G. and Shaikh BM. Poly(ethylene glycol) (PEG-400) as an alternative reaction solvent for the synthesis of some new 1-(4- (4-chlorophenyl)-2-thiazolyl)-3aryl-5-(2-butyl-4-chloro-1H-imidazol-5yl)-2-pyrazolines

and

their

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vitro

antimicrobial

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 21. Reddy V., Patil N. and Angadi S. D., Synthesis, Characterization and Antimicrobial Activity of Cu(II), Co(II) and Ni(II) Complexes with O, N, and S Donor Ligands, E-J. Chem., 5(3) 577-583 (2008) 22. Earnshaw A, Introductio to Magnetochemistry. Academic Press. New York., (1968)

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Nutritional sink formation in galls of Alstonia scholaris Linn. (Apocynaceae) by the insect Pauropsylla tuberculata (Homoptera: Psyllidae). Deepak Kumar, Vijai Malik and S.C. Dhiman* Department of Botany, Department of zoology*, M.S. College, Saharanpur (U.P.) Email- [email protected] ABSTRACT Galls are the localized outgrowth of various plant organs in which host tissue stimulated by host parasite interaction. Galls act as nutritional sinks provide essential nutrients for insect growth and development. Gall former have the ability to alter the developmental process of plant tissue to cause the formation of tumour like growth that surround the insect to protect it from the environment and supply it with a source of food. Galls caused by insect Pauropsylla tuberculata on Alstonia scholaris tree were studied to examine the different changes resulting from the biotic stress caused by insect feeding. It is supposed that Ist nymphal instar initiates gall formation during feeding by injecting its proteins and lytic enzymes rich saliva. This leads to hypertrophy and hyperplasia in the localized area of feeding site causing mobilization of nutrients such as free amino acids, reducing sugars, total soluble sugars, total phenols and protein to the gall from the un-galled region of plant. Key words - Pauropsylla tuberculata, Alstonia scholaris, gall, nutritional sink. INTRODUCTION Alstonia scholaris R.Br. (Apocynaceae) commonly known as Saptaparni is an evergreen, tropical tree with white funnel-shaped creamish flower and milky sap. It grows upto 40m tall with a spread of 10m. It is well known remedy for the treatment of various types of disorders in the Ayurveda, homoeopathic and folklore system of medicine in India [1, 2]. Various parts of the A. scholaris tree have been widely used in treatment of several aliments. It barks is used for medicinal purpose, ranging from malaria and epilepsy to skin conditions and asthma [3]. In Ayurveda it is used as a bitter and as an astringent herb for treating skin disorders, malarial fever, urticarial, chronic dysentery, diarrhoea and in snake bite. The milky juice from the different part of tree is used for the treatment of ulcers [4]. A. Scholaris is also a beautiful foliage tree with large canopy and because of this; it has become a popular ornamental tree in landscapes and gardens. Now a days it is preferred as roadside plant in cities as it is highly affected to pollutants. However, this plant of great economic value suffers with the gall infested by Pauropsylla tuberculata (Homoptera: Psyllidae). Galls have been reported on the various parts of the tree and this causes serious damage to A. scholaris tree. Gall insects have developed highly specialized and nutritional relationship with their host plant because these insects spend a major part of their life within galls and interact with galls through modifications that range from simple tissue removal or damage to vascular tissue to complex manipulation of synthesis and transport of host-plant nutrients [5, 6, 7, 8]. Insects can use the galls as sinks for nutrient to employ in the growth and reproduction of larvae [9, 10]. Growth of gall tissue is associated with the changes in the levels of their cellular contents such as carbohydrates, proteins, nucleic acids phenols and IAA enzymes

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in [11]. This work was undertaken to find out the formation of nutritional sink by identifying the different chemicals level in the galled and ungalled leaf tissue. MATERIALS AND METHODS A. scholaris tree having galls of different stages were collected at Saharanpur district and adjacent areas. Comparisons of different phytochemicals among various stages of gall development namely young, mature and old gall were carried out. Total free amino acids were extracted and determined by following the method of Sugano et al. [12] with slight modification. Total protein was extracted by the Acetone-TCA precipitation method as described by Parida et al. [13] and estimated by following the method of Lowery et al. [14] using defatted bovine serum albumin (BSA) as standard. The amount of total phenolic in normal and galled tissue were determined with folin ciocalteu reagent according to the method of Singalton and Rossi [15] with slight modification using tannic acid as a standard. Estimation of reducing sugar was done by phenol sulphuric acid reagent method [16, 17]. The determination of different enzyme activity viz., peroxidase, IAA-oxidase, Invertase, α-amylase were estimated using the methods suggested by Birecka et al., [18], Mahadeven and Sridhar [19], Harris and Jaffcoat [20] and Bernfeld [21] respectively. RESULTS AND DISCUSSION Galls are specialized plant structures formed when a galling organism alters the development of normal plant tissue [22]. The development of gall is a function of cell inhibition differentiation, growth or suppression of host plant tissues, regardless of their induction agents. However, the initiation of galling and the subsequent manipulation mechanism of gall forming process, such as phenolic compounds, protein, plant hormone analogues and genetic manipulation are still unclear [23, 24, 25, 26]. The amount of total amino acid was recorded to be more in gall tissue as compared to normal leaf tissue (Table- 1). Since, insect derive their nutrition from gall tissue, the gall becomes a sink for different nutrients and energy that will be vital for the insect’s growth. Miles and Lloyd [27] and Miles [28] suggested that increase in quantity of amino acids in gall tissues may be due to break down of proteins into utilizable unites by the enzyme protease secreted by the salivary glands of the insects. Koyama et al. [29] observed high concentration of amino acids in aphid gall and this support the nutrition hypothesis for gall formation. Total protein show high values at the start of gall growth and decline progressively with ageing. Secondly quantity of total protein was low when compared with the values obtained from normal leaf (Table-1). These observations were corroborates with the observations of Arora & Patni [30], El-Akkad [31], ScareliSantos & Varanda [32], and Kumar [33]. The formation of gall requires mechanical and chemical stimulus. According to Rosenthal and Jenzen [34] an interaction between the offensive stimuli involving growth substances released by insects and defensive response by appears to be the hallmark of gall production. The action of stimulus leads to the formation of new tissues, which cover the nymph. Kinsella [35] suggested that proteins are the primarily building block sources for new tissue, in plants, animal and human beings. Thus stimulus of gall forming insects redirects the growth and differentiation of cells which act as a sink of nutritive substances [36]. On the other hand defensive importance of diverse plant proteins was also suggested by

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Reinbothe et al., [37]. Defensive proteins are to be believed that they block the action of proteolytic enzymes from herbivores. These proteins known as proteinase inhibitors [38]. So, it can be concluded that when plants are attacked by insects they generate signals and one of these signals is the initiator of expression of certain polypeptides that may be useful in providing the basis for new crop protection strategies. Total phenol was high in galled tissues as compared to healthy leaf (Table- 1). The increased quantity of total phenol might be attributed to defence mechanism. This resistance to disease caused by pathogen was attributed to the presence of high amount of phenol [39, 40, 41, 42]. Insects can stimulate plants to produce galls and secrete high levels of phenolic compounds [43, 44]. Kaur et al., [45] reported potent antioxidant activity of ethanolic extract in Quercus infectoria galls contained high levels of polyphenols. Hence, the increased quantity of phenols in the galled tissues may be contributing to the resistance against pathogen (P. tuberculata). According to Kraus and Spiteller [46] different classes of phenolics substances act as plant defensive agents against microorganisms and herbivores. The resistance to disease caused by the pathogens is due to the presence of high amount of phenol [47]. The increase in phenolics in relation to resistance was reported in Brassica [48]. Subsequent increase in phenol contents were general responses associated with resistance mechanism in plants as reported by Ghosal et al., [49]. Gupta [50] also found higher phenolic (total phenol and orthohydroxy phenols) as compared to their normal counterparts, in gall tissues of Dalbergia sissoo, Salvodora oleoides and S. persica. Thus, increased quantity of total phenolics in the galled tissue of A. scholaris is basically for providing resistance against insect infestation. The quantity of soluble sugar was considerably high in gall tissue as compared to normal tissue (Table1). Sugar has large numbers of stereo-isomers, because they contain several asymmetric carbon atoms [51]. Increase in sugar content might be due to accumulation of these substances. This accumulation may involve the translocation of soluble sugars from the neighboring healthy tissues to physiological sinks. Findings of Shaw and Samborski [52] also support this view. Sugars are very important biochemical nutrients for the plants. If abundantly existing, these are detected naturally by various micro-organisms like bacteria, fungi, insects etc. So once infected, inside the host tissue these organisms utilize the excess sugar present in the plant for their growth and substance [53, 54]. High sugar contents in young and mature galls may be due to increased metabolic activity under feeding stress of nymph, which in turn may be responsible for addition of sugar. In Alstonia scholaris, IAA- oxidase activity was higher in galled tissue as compared to normal leaves tissue (Table- 2). IAA (Indole Acetic Acid) the main auxin in higher plants has profound effects on plant growth and development. Both plants and some plant pathogens can produce IAA to modulate plant growth [55]. A higher level of phenol affects adversely the IAA- oxidase activity in plant tissue resulting in a higher level of IAA [50, 56], thus leading to hyperauxinity and gall formation. Kumar [57] also observed higher amount of free auxin concentration and IAA- oxidase in gall tissue. High level of IAA will support cell expansion (Hypertrophy) and Cell division (Hyperplasia). Gall tissues showed increased peroxidase activity as compared to normal leaves tissue (Table- 2). Increase in peroxidase activity is due to phenol concentration, which plays an important role in oxidating enzymes [30, 50, 57]. Peroxidases are responsible for oxidation of phenolics [58]. Increase activity of these

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in oxidative enzymes indicates a state of high catabolism induced during pathogenesis. Devnathan et al., [59] observed high peroxidase activity in bunchy top banana virus infected cultivars of banana. Meena et al., [42] suggested that increased peroxidase activity was associated with resistance reaction which would be due to increased phenol concentration, where phenols were cofactor of peroxidase and hence influenced. Increase activity of alpha-amylase was found in gall tissues (Table- 2). Due to larval feeding stress, metabolic activity increased galled tissues which in turn stimulate the synthesis of sugar. Carbohydrates may also accumulate by depletion of starch due to the activated alpha-amylase activity and other enzymes [60, 61, 62]. Shekhawat [61] and Purohit [63] also reported increased activity of alpha-amylase along with increased sugar contents. Invertase activity was also higher in Alstonia gall tissues ((Table- 1)). As in crown galls formed by Agrobacterium [64] and galls of aphid Hormaphis hamameludis [65]. Our results implicate elevated invertase activity as a means by which insect galls become sinks. Galls are known to act as a sinks for plant assimilates [66] and high level of soluble invertase are associated with the establishment of sink characteristics [67, 68, 69]. Within the tissue, vacuolar acid invertase may hydrolyze sucrose to provide the hexoses required for elevated metabolic activity [70, 71] and invertases in nutritive tissue are known to hydrolyse sucrose to glucose and fructose [66]. While gypsy moth wounding may also elicit invertase activity [72], the increase brought about by gallers was much greater than that caused by 1 week of gypsy moth feeding. CONCLUSION Plant and insects interact at various levels and insect-induced galls are the one of the most exciting and deepest relationship between them. Galling insects are unique in controlling the within plant movement of photo-assimilates and tissue morphogenesis. The nutritional status of host plant tissues favours the induction and the establishment of the insect gall. Our analyses led us to the conclusion that the gall inducing insect (Pauropsylla tuberculata) use host plant resources (Alstonia scholaris) to its own benefit ACKNOWLEDGEMENTS The authors are grateful to the Principal M. S. (P.G) College Saharanpur for providing the laboratory research facilities, and encouragement. REFERENCES 1.

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 5. Rohfritsch, O. Food supply mechanism related to gall structure, the example of Goocrypta galii. L.W. Cecidomydiidae, ologotrophini on Galium mollugo. Phytophaga, 2: 1-17, (1988) 6. Bronner, R. The role of nutritive cells in the nutrition of Cynipids and Cecidomyiids. In: Shorthouse, J.D., Rohfritsch, O. (eds). Biology Of Insect – Induced Galls. Oxford University Press, Oxford, 118-140, (1992) 7. Dreger-Jauffret, F. and Shorthouse, J.D. Diversity of gall – inducing insects and their galls. In: Shorthouse, J.D., Rohfritsch, O. (eds). Biology Of Insect – Induced Galls. Oxford University Press, Oxford, 8-33, (1992) 8. Raman, A. and Dhileepan, K. Qualitative evolution of damage by Epiblema strenuana (Lepidoptera: Tortricidae) to the weed Parthenium hysterophorus (Asteraceae). Ann. Entomol. Soc. Am. 92: 717723, (1999) 9. Larson, K.C. and Whitham, T.G. Manipulation of food resources by a gall-forming aphid the physiology of source-sink interactions. Oecologia, 88: 15-21, (1991) 10. Raman, A. and Abrahamson, W.G. Morphometric relationships and energy allocation in the apical rosette galls of Solidago altissima (Asteraceae) induced by Rhopalomyia solidaginis (Diptera: Cecidomyiidae), Environ. Entomol., 24: 635-639, (1995) 11. Chil-Woo Lee, Maria Efetova, Julia, C Engelmann, Kramell, K., Wasternack, C., Jutta Ludwig-Muller, Hedrich, R. and Deeken, R. Agrobacterium tumefaciens promotes tumor induction by modulating pathogen defence in Arabidopsis thaliana [W]. The Plant Cell, 9: 2948-2962, (2009) 12. Sugano, N., Tanaka, T., Yamamoto, E. and Nishi, A. Phenyl-alanine ammonialyase in carrot cells in suspension cultures, Phytochemistery, 14: 2435-2440, (1975) 13. Parida, A.K., Das, A.B., Mittra, B. and Mohanty, P. Salt stress induced alterations in protein profile and protease activity in the mangrove Bruguiera parviflora. Z Naturforsch, 59: 408-414, (2004) 14. Lowery, O.H., Rosebrough, N.J., Farr, A.L. and Randall, R.J. Protein measurement with the folin phenol reagent. J. Biol. Chem., 193-265, (1951) 15. Singleton, V.L. and Rossi, J.A. Colorimetry of total phenolics with phosphomolybdic- phosphotungstic acid reagents. Am. J. Enol. Vitic., 16: 144-158, (1965) 16. Miller, G.L. Use of dinitrosalicyclic acid reagent determination of reducing sugar. Annal. Chem., 31: 426428, (1972) 17. Dubios, M.K., Gilles, J.K., Robers, P.A. and Smith, F. Calorimetric determination of sugar and related substance. Analyt. Chem., 26: 351-356, (1951) 18. Birecka, H., Briber, K. A., and Catalfama, J. L. Comparative studies on tobacco pith and sweet potato root isoperoxidases in relation to injury, indole acetic acid and ethylene effects. Plant Physiology, 52: 4349, (1973) 19. Mahadevan, A. and Sridhar, R. Methods in physiological plant pathology. 3rd Edn. Sivakami Publication, Madras. India, p.316, (1986) 20. Harris, G.P. and Jaffcoat, B. Effects of temperature on the distribution C 14-labelled assimilates in the flowering shoots of Carnation. Ann. Bot., 38: 77-83, (1974) 21. Bernfeld, P. α and β amylase. Methods in Enzymol., 1: 149-158, (1955)

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 22. Felt, E.P. Plant galls and gall makers. Comstock. Ithaca, N.Y., (1940) 23. Hori, K. Insect secretions and their effect on plant growth, with special reference to hemipterans. In Biology of Insect-induced Galls (J.D. Shorthouse and O. Rohfritsch, eds.) pp.157-179. Oxford University Press, Oxford, England, (1992) 24. Price, P.W., Evolution and Ecology of gall-inducing sawflies. In Biology of Insect-induced Galls (J.D. Shorthouse and O. Rohfritsch, eds.) pp.208-224. Oxford University Press, Oxford, England, (1992) 25.

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 41. Prashar, A. and Lodha, P. Phenolics estimation in Foeniculum vulgare infected with Ramularia blight. Ann. Pl. Protec. Sci., 15: 396-398, (2007) 42. Meena, R.K., Patni, V. and Arora, D. K. Study on phenolics and their oxidative enzyme in Capsicum annuum L. infected with Geminivirus, Asian J. Exp. Sci., 22(3): 307-310, (2008) 43. Hartley, S.E. The Chemical composition of plant galls: are levels of nutrients and secondary compounds controlled by the gall-former? Oecologia. 113: 492-501, (1998) 44. Motta, L.B., Kraus, J.E., Salatino, A. and Salatino, M.L.F. Distribution of metabolites in galled and nongalled foliar tissues of Tibouchina pulchra. Biochem. Systematics. Ecol., 33: 971-981, (2005) 45. Kaur, G., Athar, M. and Alam, M.S. Quercus infectoria galls possess antioxidant activity and abrogates oxidative stress-induced functional alterations in murine macrophages. Chem. Biol. Interact. 171: 272-282, (2008) 46. Kraus, R. and Spiteller, G. Phenolic compounds from roots of Urtica dioica. Phytochemistery, 29(5): 16531659. (1990) 47. Mehrotra, R.S. and Aggrawal, A. Plant Pathology. Tata McGraw-Hill Pub., Co. Ltd., New Delhi, (2003) 48. Singh, H.V. Biochemical transformation in Brassica spp. Due to Peronospora parasitica infection. Ann. Pl. Proct. Sci., 12: 442-444, (2004) 49. Ghoshal, T.K., Dutta, S., Senapati, S.K. and Deb, D.C. Role of phenol contents in legume seeds and its effect on the biology of Collosbrchus chinensis. Ann. Pl. Protec. Sci., 12: 442-444, (2004) 50. Gupta, J.P. Enzyme involved in phenol metabolism of gall and normal tissues of insect induced leaf galls on economically important plants in Rajasthan India. Bioscience Discovery, 2 (3): 345-347 (2011) 51. Lindhorst, T.K. and Thisbe, K. Essentials of carbohydrate chemistry. New Delhi, Wiley Eastern Ltd., (2003) 52. Shaw, M. and Samborski, D.L. The physiology of host-parasite relations. The accumulation of radioactive substances at infection of facultative and obligate parasite including TMV., Canad. J. Bot., 34: 389405, (1956) 53. Sheen, S.J. and Anderson, R.A. Comparison of polyphenols and related enzyme in the capsule and nodal tumor of Nicotiana plants. Can. J.Bot, 52: 1379, (1974) 54. Srilakshmi, P., Sailaja, D., Bhanuteja, M., Kumar, D.R. and Madhuri, M. Quantitative estimation of carbohydrates in insect induced and fungal infected leaf galls of Pongamia pinnata. Int. J. Plant, Animal and Envi. Sci., 2(2): 203-205, (2012) 55. Zhao, Y. Auxin biosynthesis and its role in plant development. Annual Review of plant Biology, 61: 49-64, (2010) 56. Sisodia, R. and Panti, V. Histochemical localization of metabolites and enzyme in the leaf gall and normal leaf of Diospyros melanoaylon Roxb., J. Indian Bot. Soc. 85: 37-42, (2006) 57. Ramani, V. and Kant, U. Phenolics and enzymes involved in phenol metabolism of gall and normal tissues of Prosopsis cineraria (Linn.) Druce in vitro and in vivo. Proc. Indian Nat. Sci. Acad., B55 (5&6): 477-420, (1989) 58. Kosuge, T. The role of phenolics in host response to infection. Ann. Rev. Phytopath., 7: 195-22, (1969)

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TABLE -1 Quantitative estimation of different metabolites PARTS

IAAOXIDASE

NL YG MG OG

2.44±0.52 2.90±0.30 3.6±0.42 2.50±0.02

αAMYLASE starch hydrolysed mg/min.dwt 2.3±0.2 3.1±0.1 3.5±0.3 2.8±0.3

PEROXIDASE ∆A/gm.dw/min

INVERTASE Sucrose/ hydrolysed mg/min. dwt

0.8±0.1 1.8±0.4 1.4±0.3 1.3±0.3

3.6±0.4 4.4±0.2 4.2±0.1 3.9±0.1

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TABLE - 2 Quantitative estimation of different enzymes *NL=Normal leaf, YG=Young gall, MG=Mature gall, OG=Old gall

PARTS

TOTAL SOLUBLE SUGARS (mg/gm dw)

TOTAL REDUCING SUGAR (mg/gm dw)

TOTAL PHENOL (mg/gm dw)

TOTAL PROTEIN (mg/gm dw)

NL

3.3 ± 0.08

1.3± 0.1

0.62± 0.02

1.8± 0.22

TOTAL FREE AMINO ACIDS (mg/gm dw) 3.0± 0.71

YG

4.2± 0.01

2.8± 0.2

1.8± 0.46

3.6± 0.35

4.6± 0.52

MG

3.7± 0.40

3.5± 0.2

1.02± 0.03

2.8± 0.34

5.3± 0.30

OG

2.5± 0.22

2.3±0.2

0.82± 0.02

2.5± 0.33

4.2± 0.21

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Perspective of Stem Cell Research in India Bindu Sharma Associate Professor, DN PG College, Meerut (UP) Email: [email protected] ABSTRACT Stem cell therapy has shown immense potential by treating diseases that were traditionally considered “degenerative and incurable” such as diabetes, Parkinson's, Alzheimer's disease. In India, stem cell programmes have been initiated with the aim of promoting both basic and translational research in view of its potential applications. Techniques have been developed for the in vitro culture of stem cells. As a result, scientists can now carry out experiments aimed at determining the mechanisms underlying the conversion of a single, undifferentiated cell, the fertilized egg, into the different cells comprising the organs and tissues of the human body. National agencies are pro-active in supporting and promoting this area. Over 30 institutions, hospitals and industry are involved in SCR in the country. However, there are many challenges in current stem cell research such as non-availability of human resources of adequate expertise; very few indigenous hESC lines generated; inter disciplinary structure is yet to be created; derivation of ES cells from early human embryos, and EG and fetal stem cells from aborted, fetal tissues raise ethical, legal, religious, and policy questions. Further, the potential use of stem cells for generating human tissues and, perhaps, organs, is a subject of ongoing public debate. Key words: Stem cell, India, embryonic stem cell research. INTRODUCTION Stem cells are one of the human body’s master cells with the ability to grow into any one of the body’s more than 200 cell types Stem cells are undifferentiated biological cells that can differentiate into specialized cells and can divide to produce more stem cells. In mammals, there are two broad types of stem cells: embryonic stem cells, which are isolated from the inner cell mass of blastocysts, and adult stem cells, which are found in various tissues. In adult organisms, stem cells and progenitor cells act as a repair system for the body, replenishing adult tissues Stem cells are precursor, unspecialized, undifferentiated cells capable of selfproliferation, migration and differentiation. In the simplest form, the stem cell is an immature cell that has the capability to differentiate into any possible mature cell. Much like the bone marrow, cord blood is one of the richest sources of stem cells. Cord blood stem cell research is being conducted for potential future use in the treatment of certain auto-immune disorders, neurological disorders, muscular/cartilage diseases, stroke, etc. Stem Cell Research in India Indians have also started to develop stem cell lines, including at least three human embryonic stem cell (hESC) lines to date (UK Stem Cell Bank and National Institutes of Health (NIH) Human Embryonic Stem Cell Registry).Two hESC lines derived by the Jawaharlal Nehru Centre for Advanced Scientific Research (JNCASR), in Bengaluru, have been accepted for deposition and distribution by the UK Stem Cell Bank. The cell lines, derived from low-quality embryos discarded post-IVF procedures, will be part of the International

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Stem Cell Initiative 2 (ISCI2) project to identify the common genetic changes that occur in hESC lines on prolonged culture. This will be the first time that India is represented on an ISCI project. India’s Department of Biotechnology (DBT), has listed more than 30 research institutes, hospitals, and firms involved in stem cell research in India [1-5]. Research Institutes Public research institutes are the largest group active in stem cell research with in India. Some institutes focus primarily on basic research. JNCASR studies ESC differentiation into cardiovascular cells. Other institutes balance basic research with applied activities such as animal modeling, clinical trials, or pilot treatments. NCCS has conducted animal and preclinical analyses of bone marrow stem cell injections for pancreatic regeneration. Research efforts from this institute succeeded in rescuing mice with experimentally induced diabetes after a 30 day follow up [5-7], and scientists at NCCS hope to extend this work to an autologous clinical trial in human diabetic patients. The institute is working to establish a team of clinicians, scientists, and patients to act as a platform for the trial, a process they estimate will take 3–4 years. Their general business model is to conduct the basic research and find a private partner for further development. This is a model they have successfully applied in the past, as illustrated in their transfer of collagen sheet wound dressing technology to a local company, Euchre Pharmaceutical Pvt. Ltd. (Chennai), for production. Hospitals and Clinics Unlike India’s biotechnology sector as a whole [4, 7], few Indian companies are involved in stem cell research. Instead, Indian hospitals and clinics are key players. India’s large research-intensive hospitals, such as AIIMS, conduct basic and applied research and have clinical trial and pilot treatment capabilities. This combination of resources creates a bridge between research and therapy and makes hospitals pivotal for Indian stem cell innovation. AIIMS works on a wide spectrum of clinical applications in cardiology, ophthalmology, neurology, and hematology and also carries out basic research on, for example, stem cells and biopolymers aimed at treatments for orthopedic, ocular, and skin diseases. Christian Medical College and Hospital (CMC) in Vellore has funded a center for stem cell research in collaboration with DBT to promote translational research with stem cells. Sankara Nethralaya, an eye hospital based in Chennai, has similarly built a research building to house the Kamalnayan Bajaj Institute of Research in Vision and Ophthalmology. Stempeutics, in Bengaluru, is the research arm of Manipal Education and Medical Group (MEMG). CryoStem Cell, in partnership with Sri Bhagwan Mahaveer Jain Hospital (in Bengaluru), conducted a pilot stem cell treatment for Berger’s disease, a severe form of peripheral vascular disease, which is relatively rare in the Western hemisphere but common in India If successful, CryoStemCell’s pilot therapy has the potential to address a very real health need in India. Reliance Life Sciences is also involved in treatment development and is investing in an animal facility to conduct toxicology and preclinical efficacy studies for cell based therapies. Universities Few Indian universities appear active in stem cell research at this time. The main exception is the University of Delhi, which, together with the Indian Institute of Nuclear Medicine and Allied Sciences, is examining basic mechanisms of stem cell function. However, many of the hospitals, firms, and research institutes active in stem

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in cell R&D are also involved in education. Both CMC and AIIMS are teaching hospitals that provide postgraduate degrees. Many of India’s research institutes are collocated with a university and informally train students within their research labs. Government Support The Indian government has been a key factor in encouraging stem cell R&D activity and growth. Stem cell engineering is seen as an important area for the government and is identified as a strategic biotechnology area in the DBT’s 2007 Biotechnology Strategy (available online). Four government departments are key supporters of stem cell R&D: DBT, the Indian Council for Medical Research (ICMR), the Department of Science and Technology, and the Council of Scientific and Industrial Research. Stem cell R&D promotion is driven largely by the DBT Stem Cell Taskforce and by ICMR through its affiliated institutes and regulatory capacity. DBT provides direct funding to targeted initiatives in this field and supports both infrastructure building and operational activities such as clinical trials. It has begun to support large-scale strategic programs, such as a phase III multicenter trial using bone marrow cells to treat myocardial infarctions, and sponsors stem cell research centers like the Centre for Stem Cell Research (CMC, in Vellore) and a soon to-be-established stem cell research center in Bengaluru, the Institute for Stem Cells and Regenerative Medicines. As has been shown elsewhere for the case of health biotechnology [10-14], government support is often key to capability building within emerging economies. Building Stem Cell R&D on India’s Strengths India’s capacity to participate in a cutting-edge field such as stem cell research is, in part, built on skills and infrastructure previously developed by the nation’s pharmaceutical and biotechnology sectors [17-19]. One such skill is the proven ability of Indian firms to develop process innovations in order to lower prices [20]. Innovations that lower process costs have already occurred in the stem cell field. Researchers do not necessarily copy blindly the techniques used in developed countries but create their own cost-efficient cell and tissue culturing and storage techniques that use fewer disposable devices. Indian companies have also begun to produce materials, such as growth factors, that are needed in stem cell research at significantly lower rates. In addition, India’s pharmaceutical and biotechnology sectors have helped India develop an expertise in conducting clinical trials. Bolstering this expertise, India’s large, diverse, and treatment-naïve population provides a valuable resource for clinical trials, especially for rare diseases where the Indian population could provide sufficient patients for trial groups. As a result, India has great potential to act as a clinical trial destination of choice for stem cell therapies. This could help India develop and test stem cell therapies for a variety of diseases. This strength is likely to encourage more international ties and joint ventures, a trend already exemplified in the collaborations of United States/Indian firm StemCyte India Therapeutics and the Japanese/Indian venture NCMR. Overcoming India’s Stem Cell R&D Challenges Promoting Linkages As a translational research field, stem cell development requires a high degree of linkage between basic and clinical expertise. India’s most successful research institutes and hospitals, such as AIIMS and LVPEI, are those that integrate the efforts of basic scientists with clinicians. Unfortunately, this integration has historically been weak at most research sites elsewhere in India. Increasing coherence and connectivity between these different

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in sectors is the cornerstone of India’s new biotechnology strategy (DBT Strategy, 2007), and specific institutions, as well as the government, have initiated steps to promote interactions between basic and clinical stem cell researchers. The construction of a joint research center between the academic Centre for Cellular and Molecular Biology and the hospital-based Nizam Institute of medical Sciences, both in Hyderabad, is an example of an Indian institutional initiative aiming to increase clinician/ scientist interaction. The DBT is promoting improved integration with its stem cell cluster initiative, which encourages publicly and privately funded stem cell research groups to share ideas, facilities, and research. Four clusters are already established: around the CMC in Vellore, the NCCS in Pune, in Bengaluru, and in Hyderabad. The project is expected to expand and promote an additional cluster in Delhi. Strengthening Training For India to increase its stem cell research capacity, it will need to strengthen the quality of its current scientific education. The DBT is helping Indian scientists gain expertise abroad by offering overseas stem cell fellowships and travel bursaries for conferences. Research institutes are increasing their involvement in training through postgraduate supervision and by coordinating workshops, such as the stem cell training workshops currently run by JNCASR in partnership with the NCBS, both in Bengaluru. Universities are also becoming more involved in stem cell training programs, such as Manipal University’s efforts to establish an Institute of Regenerative Medicine with a related graduate program in the summer of 2007. NCMR, in collaboration with Acharya Nagarjuna University (in Chennai), launched a stem cell PhD program in April 2008. The program will be focused on bringing clinicians together with scientists. Indians can also strengthen their capacity for stem cell research by attracting Indian experts who are currently active in this field in developed countries. Members of the Indian scientific diaspora (expatriates working in industrially developed countries) have begun to return to India in greater numbers, encouraged by economic prosperity and active recruitment initiatives from firms and research institutes. More proactive strategies could strengthen this flow of returning scientists, with targeted efforts designed to attract expatriate experts in stem cell research. Increasing Public Awareness While Indians who have heard of stem cell research and therapies are generally supportive, this field is still relatively unknown to the general public, except to a small subpopulation of educated urbanites. There is some risk that this lack of broad understanding may lead to uninformed mistrust of the field, particularly if scandals or slow results help to destroy public support and lead Indians to mobilize against new stem cell therapies. This mobilization against new technologies has already occurred in India with the introduction of genetically modified cotton [21]. While this risk is not unique to India, it is particularly challenging to educate the extensive Indian population about stem cells, due to the logistical difficulty of communicating the details of scientific advances to a vast population with typically low levels of general education. Another risk that is nonspecific to India, but that might be unusually challenging to overcome in this country, is that expectations for stem cellbased therapies may be overly inflated to the ultimate detriment of the field. There is considerable potential for the exploitation of patients who see stem cell therapy as a ‘‘magic bullet’’ to solve their health needs. This exploitation has already been described in anecdotes of Indian clinics offering stem cell therapies without a strong scientific basis or proper safety and efficacy tests.

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Developing the Regulatory Framework Until recently, India lacked comprehensive stem cell research and therapy guidelines, an omission that compounded the potential for patient exploitation. To address this barrier, ICMR led the development of stem cell research and therapy guidelines (see ICMR National Guidelines) with the participation of DBT and others. Newly finalized, the guidelines are relatively permissive, allowing hESC and adult stem cell research and cell line development under close monitoring. Experimental treatments, embryo creation for research, and chimera studies are permissible subject to approval. The guidelines create two levels of stem cell research review and monitoring: a National Apex Committee for Stem Cell Research and Therapy as well as institutional committees. Depending on the research topic, projects will be approved nationally or institutionally; research will be categorized into permissive, restricted, and prohibitive areas for research, and all projects will have to register nationally. India’s guidelines are relatively permissive when compared to other countries. Regarding hESC research, one study surveyed 50 countries and found that hESC research was allowed under strict conditions in 23 countries and banned in five, while the rest had no explicit policy [14-18]. The new Indian guidelines are not legally binding; however, many Indians remain optimistic that their existence will encourage researchers to begin working in the area of stem cells while simultaneously stopping unethical R&D. It’s unclear how successful the guidelines’ implementation will be. India has a poor track record at monitoring IVF clinics and enforcing its guidelines for biomedical research using human subjects [12-15]. Ethical issues The extraction of HESCs from inner cell mass for research purpose leads to the destruction of the embryo. The major source of human embryonic stem cell tissues are the spare or supernumerary embryos created during in vitro fertilization as a part of infertility treatment. The other source is creating embryos with somatic cell nuclear transfer techniques (SCNT). The legislation of most countries including India allows use of spare or supernumerary embryos either fresh or frozen created during in-vitro fertilization. Some countries with more liberal view have allowed creation of human embryos with SCNT as a source of embryonic tissues. Conclusion As India’s capacity in stem cell research continues to develop, it can draw upon the nation’s numerous strengths to actively expand its involvement in this field. Based on its track record in information technologies and biotechnology, it is likely that India will be successful in building its capacity for implementing stem cell therapies. All ethical principles applying to research must also be ensured in stem cell research: Principles of essentiality, of voluntariness, informed consent and community agreement, of non-exploitation, of privacy and confidentiality, of precaution and risk minimization, of professional competence, of accountability and transparency, of maximization of public interest and distributive justice, of public domain and the principle of totality of responsibility and compliance. References 1. Becker A. J., McCulloch E.A. and Till J. E. Cytological demonstration of the clonal nature of spleen colonies derived from transplanted mouse marrow cells. Nature. 197:452-4 (1963)

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 2. Siminovitch L, McCulloch E. A. and Till J. E. The distribution of colony-forming cells among spleen colonies. J Cell Physiol. 62:327-36 (1963) 3. Velu N. Stem cell transplantation. API medical update. 14:366-77(2004) 4. Friedenstein A. J., Gorskaja J. F. and Kulagina N. N. Fibroblast precursors in normal and irradiated mouse hematopoietic organs. Exp Hemato. l 4(5):267-74 (1971) 5. Murrell W., Feron F. and Wetzig A. Multipotent stem cells from adult olfactory mucosa. Dev Dyn. 233(2):496-515 (2005) 6. Caveleri F. and Scholar H. R. Nanog: a new recruit to embryonic stem cell orchestra. Cell. 113:551-2 (2003) 7. Wang X., Yang Y. J. and Jia Y. J. The best site of transplantation of neural stem cells into brain in treatment of hypoxic-ishemic damage: experiment with newborn rats. Zhonghua Yi Xue Za Zhi. 27;87(12):847-50 (2007) 8. Molofsky A. V., Pardal R. and Iwashita T. Bmi-1 dependence distinguishes neural stem cell self-renewal from progenitor proliferation. Nature. 425 (6961):962-7 (2003) 9. Park I. K., Qian D. and Kiel M. Bmi-1 is required for maintenance of adult self-renewing haematopoietic stem cells. Nature. 423(6967):302-5(2003) 10.

Beachy P. A., Karhadkar S. S. and Berman D. M. Tissue repair and stem cell renewal in

carcinogenesis. Nature. 432(7015):324-31 (2004) 11.

Rosenthal N. Prometheus’s vulture and the stem-cell promise. N Engl J Med. 349(3):267-74 (2003)

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Korbling M. and Estrove Z. Adult stem cells for tissue repair-a new therapeutic concept? N Engl J

Med. 349(6):570-82 (2003) 13.

Marshall G. P., Laywell E. D. and Zheng T. In vitro-derived "neural stem cells" function as neural

progenitors without the capacity for self-renewal. Stem Cells. 24 (3):731-8 (2006) 14.

Lavker R. M. and Sun T. T. Epidermal Stem cells: properties, markers, and location. Proc Natl Acad

Sci USA. 97(25):13473-5 (2000) 15.

Liu S., Dontu G. and Wicha M. S. Mammary stem cells, self-renewal pathways, and carcinogenesis.

Breast Cancer Res. 7(3):86-95 (2005) 16.

Shackleton M., Vaillant F. and Simpson K. J.. Generation of a functional mammary gland from a single

stem cell. Nature. 439:84-8 (2006) 17.

Tuch B. E. Stem cells--a clinical update. Aust Fam Physician. 35(9):719-21 (2006)

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Rideout W. M., Hochedlinger K. and Kyba M. Correction of a genetic defect by nuclear transplantation

and combined cell and gene therapy. Cell. 109(1):17-27 (2000) 19.

Evans M. J. and Kaufman M. H. Establishment in culture of pluripotential cells from mouse embryos.

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Reynolds B. A. and Weiss S. Generation of neurons and astrocytes from isolated cells of the adult

mammalian central nervous system. Science. 255(5052):1707-10 (1992) 21.

Vawda R., Woodbury J. and Covey M. Stem cell therapies for perinatal brain injuries. Semin Fetal

Neonatal Med. 12(4):259-72 (2007)

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Study of Medicinal Angiosperms of Rampur District (UP), India With Special Reference to Their Sustainable Use BEENA KUMARI Botany Department, Hindu College (PG),Moradabad.244001 Email: [email protected] ABSTRACT Sustainable development may be defined as development that is economically sound, socially relevant and environment friendly. There are more than two thousand five hundred plant species in India having documented medicinal value. These medicinal plants and their raw materials are used in the prevention, treatment and cure of health disorders since ancient time. The knowledge of plants and medicines has undergone various stages till today. Many of today’s drugs have been derived from plant sources. Rampur district is located at Longitude 78 0 54’ to 690 28’E and Latitude 280 25 to 29010’ N and spans an area of 2,367 km². It was incorporated into the state of U.P. in 1949. Due to rapid increase in industrial area and clearing of urban forest area for residential use green plants facing the danger of extinction. The maximum population of the district Rampur resides in rural areas. They use folk methods to cure different diseases through wild plants. But due to urbanization of the area, the traditional knowledge of wild plants is now disappearing day by day. In the present study 58 angiosperm plant species belonging to 32 families have been enumerated. According to one estimate more than 25% of medicinal plants used by pharmaceutical companies are collected from wild and much of this is illegal. 70% of this involves destructing harvesting. Therefore, in order to maintain a sustainable supply of the raw material from the forest, their cover exploitation should be stopped and strict law should be enforced by the government to converse these valuable medicinal plants. In this way we can save our plant wealth. Keywords: Angiosperms, Biodiversity, Medicine, Sustainable use, Rampur. Introduction Medicinal plants are considered very important in primary health care system. A large number of plants are known for their medicinal properties. There is a large demand for medicinal herbs due to increase in the use of herbal formulations. Herbal medicines are used by about 75-80% of the world’s population for primary health care because of better cultural acceptability, better compatibility with human body and lesser side effects [1,2]“Wild harvesting”- medicinal plants at risk. The negative impact of commercial collection of medicinal plants cause tremendous short and long term damage to plant resources, vegetation communities and ecosystem [3,4]. Earlier, medicinal plants were obtained from the forests. At that time in the forests they were in abundance and the consumption was in milligrams or grams. But now, the situation has reverseddue to deforestation, uprooting of the plants for fulfilling the requirements and the craze for herbalglobalization[5,2].So the medicinal plants have become endangered. Therefore, the rates have also increased and are unable to fulfil the requirement of the genuine material in the world. Study Area Rampur district is located at Longitude 780 54’ to 690 28’E and Latitude 280 25 to 29010’ N. Spread in area of 2,367 km2, Rampur is 192 meter above sea level in north and 166.4 meter in south. During summers the temperature is usually from 43 °C to 30 °C and during winters it is from 25 °C to 5 °C. The average rainfall varies between 800 to 900 mm.The relative humidity is up to 90% in monsoon season and in drier part of the

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in year it decreases to less than 20%. As per the 2011 Census of India Rampur had a population of 325,248 [compared to 281,549 in 2001] showing 16% growth in 2001-11.People living in the rural areas of Rampur district of Rohilkhand region of U.P. are greatly dependent on medicinal plants for variety of uses. It proved to be a good income source and cheap source of curing diseases at local level. Currently medicinal plants are under severe threat of extinction due to rapid deforestation, over and improper collection, over grazing etc. All the plant material comes from thewild and little effort has been made for their cultivation[6-8]. The present study is an effort to analyse the current status of medicinal angiosperms in the context of its value for the local people of Rampur district. Methodology The field studies were carried out in 2011 - 2012. To collect information on medicinal angiospermsand their uses for different ailments, local traders, Vaidyas and old women wereidentified and interviewed. Plant specimens were collected from study area and shown to them for authentic information. Available floras [9,10] were consulted for identification ofplants. The survey area map is enclosed for reference [fig.1]. Results &Discussion In the present study 58 angiosperm plant species belonging to 32 families have been enumerated [Table 1.]. Herbs [29 genera & 36 species] were collected from 21 families, shrub [9 genera & 10 species] from 6 families, trees [5 genera & 5 species] from 5 families and climbers [7 genera & 7 species] from 6 families. Asteraceae family is dominant with 6 species [fig.2& 3]. Total number of plant parts used in different ailments are shown in fig. 4. Medicinal plants collectors are untrained, and almost half of the material collected by untrained manpower is wasted. Therefore, there is a need to find ways to harvest medicinal plants sustainably from the wild. This includes training local collectors in proper collection techniques, training people to grow medicinal plants, and removing some of the middlemen from the trading chain [11,12]. More than 25% of the medicinal plants used by Indian Industry are collected from wild and much of this is illegal. 70% of the collection involves destructive harvesting practices like overexploitation, fragmentation of natural habitats and introduction of exotic species [13,4]. Medicinal plants can save lives, livelihoods and cultures until they themselves are saved. Therefore, in order to maintain a sustainable supply of the raw materials from the forests their overexploitation needs to be stopped and strict laws should be enforced by the Government for the conservation of forests [12,14].

The

conservation program can be strengthened by using the people who have knowledge and also respect for the Mother Nature. Women can be great contributors in the conservation program as they manage most of the plant resources that are used by humans [5,15,16]. Thus, with the involvement of the collectors, producers and traders including ultimate users, the motive of sustainable use of the resources can be achieved [fig. 5]. Acknowledgement Author is thankful to UGC for providing financial assistance (F.No.8-2 (302)/2011 (MRP/NRCB) and

local

people for providing necessary information regarding the uses of plant species for various ailments. References 1.

Singh, Akanksha, Singh, B. N., Singh, S.P., Chaudhary, Anita and Sharma, B.K. Sustainable use of

medicinal plant biodiversity for poverty alleviation. National Conference on Biodiversity, Development and Poverty Alleviation, U. P. State Biodiversity Board, Lucknow, 24- 29, (2010) 2.

Uniyal, R.C., Uniyal, M.R. and Jain, P. Cultivation of Medicinal Plants in India: A Reference Book.

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in TRAFFIC India, Delhi, India, (2000) 3.

Astaw, D., Abebe, D. and Urga, K. Traditional medicine in Ethiopia: Perspective and development efforts, J Ethiop MedPrac, 1, 114-117, (1999)

4.

Tewari, D. N. Report of the task force on conservation and sustainable use of medicinal plants. Govt. of India, Planning Commission, New Delhi, 175, (2008)

5.

Pandey,S.S., Srivastava, B. K., Pandey, V. S., Diwedi, S. and Shriya. Medicinal plant resources of Uttar Pradesh : An urgent need for conservation,Plant Archives,10 (2),861-864,(2010)

6.

Schippmann, U., Leaman, D.J. and Cunningham, A.B. Impact of cultivation and gathering of medicinal plants on biodiversity: global trends and issues. In: Biodiversity andthe Ecosystem Approach in Agriculture,Forestry and Fisheries. Ninth Regular session of the commission on Genetic Resources for Food and Agriculture. FAO, Rome, Italy, 1-21, (2002)

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Sharma, J., Painuli, R.M. and Gaur, R.D. Plants used by the rural communities of district Shahjahanpur, Uttar Pradesh, Indian Journal of Traditional Knowledge, 9(4), 798-803, (2010)

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Siddiqui, M.B. & Husain, W. Medicinal plants of wide use in India with special reference to Sitapur district (Uttar Pradesh), Fitoterapia, 3, 65-67, (1994)

9.

Duthie, J.F. Flora of Upper Gangetic Plain and of the Adjacent Siwalik and Sub Himalayan Tracts, 3 vols, (Botanical Survey of India, Calcutta), (1903-1929)

10. Maheswari, J. K. Flora of Delhi, CSIR Publication, India, (1963) 11. Chang, L., Hua, Y. and Chen, Shi Lin. Framework for Sustainable Use of Medicinal Plants in China. Plant Diversity and Resources, 33 (1), 65-68, (2011) 12. U., Manjkhola, S. and Joshi, M. Current status and future strategy for development of medicinal plants sector in Uttaranchal, India, Curr. Sci., 83, 956-964, (2002) 13. Mullikem, T. Sustainable use of medicinal plants -A multi – sectoral challenge and opportunity, TRAFFIC International, Cambridge, U. K.(www.traffic.org), (2012) 14. Hamilton, A. C. Medicinal plants conservation and livelihoods, Biodiversity Conservation, 13, 14771517. (2004) 15. Khanna, K.K. Unreported ethnomedicinal uses of plants from the tribal and rural folklore of Gonda district, Uttar Pradesh, Ethnobotany, 14, 52-56, (2002) 16. Taylor, J.L.S., Rabe, T., McGaw, L.J., Jager, A.K. and Staden, J. Van. Towards the scientific validation of traditional medicinal plants, PlantGrowth Regul., 34, 23–37, (2001)

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Fig.1. Map of Rampur district

Lamiaceae Chenopodiaceae Amaranthaceae Solanaceae Malvaceae Euphorbiaceae Asteraceae 0

1

2

3

4

Fig.2 Dominant families with species of the study area

40 30 20 10 0

Family Genera species

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5

6

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Fig.3. Total medicinal plants family, genera and species in different taxa. Latex Fruit Tuber 3 6 1 Seed 5 Root 16

Bark 3 Leaf 19

whole plant 19

Flower 3

Fig.4. Number of plant parts utilized for medicinal purpose.

Export middle man

Traders

Vaidyas

collectors

Drug shop keepers

consumers

manufacturer

Fig. 5. Chain of people involved in sustainable use of medicinal resources

Table-1. Medicinal plant diversity of Rampur district of Rohilkhand region of U.P.

S.no

Family & Botanical name

Local Name

Part used

Uses

Kalmegh

Leaf

Diabetes & snake bite

Chirchit

Whole

Wounds, gout & rheumatism

. Acanthaceae 1.

Andrographis

paniculata

(Burm. f.) Wallich ex Nees 2.

Peristrophe paniculata Burm

plant Aizoaceae 3.

Trianthema portulacastrum L.

Santhi

Root

Constipation & asthma

(Santhi) Amaranthaceae 4.

Achyranthes aspera L.

Latjeera

Leaf

Cuts & Wounds

5.

Amaranthus spinosus L.

Katelichaula

Root

Eczema

Whole

Vermifuge

i 6.

Amaranthus viridis L.

Chaulai

plant

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Calotropis procera (Willd)

Akua

Flowers

Constipation

Madar

Root bark

Dysentery

Gondhichedi

Whole

Skin disease& fever

Drey. 8.

C. gigantia (L.) R. Br.

Asteraceae 9.

Acanthospermum

hispidum

DC. 10.

plant

Ageratum conyzoides L.

Gundrya

Flower,

Cuts &sores,kidney

seed

stone,diarrhoea,leprosy & uterine disorders

11.

12.

EcliptaprostrataL.

SphaeranthusindicusL.

Bhangra

Mundi

Whole

Constipation,Livercomplaints,asthma&H

plant

air growth

Whole

Skin diseases & piles

plant 13.

TridaxprocumbensL.

Kanphuli

Leaf

Cuts & Wounds.

14.

Xanthium strumarium L.

Gokhuriya

Whole

Malaria, piles, ulcer & rheumatism

plant Bombacaceae 15.

Adansonia digitata L.

Balamkheera

Fruit

Kidney stones

Kachnar

Bark&

Dysentery & diarrhoea

Caesalpiniaceae 16.

Bauhinia racemose Lamk.

Fruit 17.

Cassia toraL.

Chakunda

Leaf&

Eczema, cuts & jaundice

Root Commelinaceae 18.

Commelina diffusa Burm.f.

Kansura

Root

Snake bite

Bathua

Leaf

Constipation& Urinary problems

Chenopodiaceae 19.

Chenopodium album L.

&Seed 20.

Chenopodium

ambrosioides

Bathua

L. 21.

Chenopodium murale L.

Whole

Dysentery, pneumonia and piles

plant Bathu

Leaf

Asthma

Shankhpuspi

Whole

Fever, cough and cold, stomach ache,

plant

ulcer, dysentery, asthma

Convolvulaceae 22.

Evolvulusalsinoides L.

bronchitis & brain tonic Cucurbitaceae 23.

Momordica dioica Roxb. ex

Jangli-

Willd.

Karela

Root

Fever, piles, asthma, bronchitis, head ache,

126

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in dysentery& Diphtheria.

24.

Coccinia grandis (L.) J.

Kanduri

Leaf

Skin disease

Ratalu

Tuber

Dysentery, abdominal pain, jaundice &

Voigt. Dioscoreaceae 25.

Dioscorea bulbiferaL.

boils, piles Crassulaceae 26.

Bryophyllum

pinnatum

Ajuba

Leaf

Swelling & Kidney stone

(Lamk) Oken Euphorbiaceae 27.

Euphorbia hirta L.

Duthi

Latex

Wounds & lip crakes

28.

Euphorbia tirucalli L.

Kharsani

Latex

Eczema, wounds, toothache, earache, scabies, rheumatism & warts.

29.

30.

Phyllanthus fraternus Web.

Ricinus communis L.

Bhuiamla

Arandi

Whole

Allergy, diarrhoea, dysentery, dropsy

plant

&jaundice.

Leaf, root,

Skin

seed

cholera, boils, constipation, dysentery,

diseases,

sores,

gum

trouble,

joint pain, muscular pain, head ache & burns Fabaceae 31.

Abrus precatorius L.

Ratti

Root

scorpion sting & snakebite

Pitpapra

Whole

blood purifier

Fumariaceae 32.

Fumaria indica Pugsley

plant Lamiaceae 33.

Leucas aspera (Willd.) Link.

Gubba

Leaf

34.

Ocimum basilicum L.

Ramatulsi

Leaf

35.

Ocimum sanctum L.

Shymatulsi

head ache & fever. &

Cold & cough, fever,

seed

stone complaints, dropsy & cholera

Whole

Antiseptic, cold & cough, fever, urinary

plant

troubles, vomiting, bronchitis, chickenpox, cholera, constipation, headache, diarrhoea, dropsy, ear complaints, malaria, colitis gastric complaints & antidote to poison leprosy

Malvaceae 36.

Abutilon indicumL

Kanghi

Leaf, root

Dental problems

37.

Abelmoschus manihot

Janglibhindi

Root

Pneumonia

(L.) Med.

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Malvastrum

Suchi

Leaf

Wounds & sores

Chatkura

Leaf,

Diarrhoea, Dysentery,

Flower,

expectorant, body pain &rheumatism

coromandelianum (L) Garcke 39.

Urena lobata L.

Root

&

Bark Menispermaceae 40.

Cissampelos pareiera L.

Parha

Leaf

Antidote to snake & scorpion bite

Safed siris

Bark

Piles

Fagu

Latex,

Sores, constipation &

fruit

Stomachache

Mimosaceae 41.

Albizia lebbeck (L.) Benth

Moraceae 42.

Ficus palmate Forsk.

Nyctaginaceae 43.

Boerhaavia diffusa L.

Pundra

Root

&

Jaundice & constipation

whole plant Oleaceae 44.

Jasminum multiflorum

Chameli

Leaves

Pimples & eczema

Khatibuti

Whole

Diarrhoea, dysentery

plant

epilepsy, piles, fever & jaundice

Pili kateli

Root

Snake bite

Gharibel

Seed,

Tonic, headache,

leaves&

cold & cough, wounds, Inflammation,

fruit

itching& asthma

Root

Baldness

Whole

Snake bite

(Burm. f.) Aners Oxalidaceae 45.

Oxalis corniculata L.

Papaveraceae 46.

Argemone maxicana L.

Passifloraceae 47.

Passiflorafoetida L.

Polygonaceae 48.

Polygonum plebejum R.Br.

49.

Polygonum barbatumL.

Jayanti

plant Portulacaceae 50.

Portulaca oleracea L.

Kulfa

Whole

Constipation

plant 51.

Portulaca grandiflora L.

Luaniya

Leaf

Scrophulariaceae

128

Eczema

International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in 52.

Veronica anagallis-aquatica

Sadevi

L.

Whole

Healing burns

plant

Solanaceae 53.

Datura metel L.

Dathura

Leaf

Ear-ache

54.

Physalis minima L.

Damphu

Fruit

Dropsy

55.

Solanum nigrumL.

Makoi

Whole

Cough & cold

plant 56.

S. surratense Burm.f

Barkatali

Root

Gum trouble & tooth decay.

Jal booti

Whole

Menstrual complaints.

Verbenaceae 57.

Phyla nodiflora (L.) A. Rich

plant Zygophyllaceae 58.

Tribulus terrestris L.

ChotoGokhru

Fruit Root

129

&

Urinary diseases

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Water Quality Control of Industrial Effluents Using Crosslinked Chitosan Hydrogel Beads. Anuja Agarwal*& Vaishali** *Associate Professor, Department of Chemistry, J.V. Jain College, Saharanpur, India. E mail: [email protected] Mobile no.: +919411484839 **Research Scholar, Department of Chemistry, J.V. Jain College, Saharanpur. ABSTRACT Polymers from natural resources have been studied in the recent past as an important material for biotechnological application owing to their unique characteristics such as biological compatibility with natural environment, non-toxicity and biodegradability. Chitosan, deacetylated product of chitin obtained from skin of crustacean fish is(1→ 4) 2- amino 2-deoxy β – D glucan and is one of the well known biodegradable polymers metabolized by human enzymes. It can be prepared as hydrogel micro and nano sized beads, having a positive charge at wide pH range. These beads are known to adsorb a number of metals and some of other pollutants. Chitosan beads crosslinked with glutaraldehyde have been prepared by us to study their removal efficiency for dyes and beads have been characterized by SEM, FTIR, XRD and DSC analytical methods. Swelling behavior of beads has also been studied at pH 2.0, 7.4 and 10.0 which proves that swelling is more in acidic medium than in basic. Percent Color removal efficiency for dye effluents has been determined and found to be dependent of dye initial concentration, pH and temperature. The results concluded that chitosan beads can be used to remove color in effluents containing dyes and is able to improve water quality of effluents from industries. Key words: Dye, Effluent, FTIR, SEM, XRD, CRE Introduction Polymers from natural resources have been studied in the recent past as an important material for biotechnological application owing to their unique characteristics such as biological compatibility with natural environment, non-toxicity and biodegradability1,2 and also possess gel forming ability at low pH3. Deacetylated product of chitin obtained from crustacean fish is(1→ 4) 2- amino 2-deoxy β – D glucan and is known as the chitosan and is one of the well known biodegradable polymers metabolized by human enzymes 4. Chitosan can be prepared as hydrogel beads, having a positive charge at wide pH range5. Three dimensional hydrophilic polymer network of hydrogel beads are capable of retaining large amount of water for an extended time period. Hydrogels are thermodynamically compatible with water and exhibit swelling in aqueous media. Cross linked hydrogel polymer network can be obtained by cross linking chitosan using a cross linker like glutaraldehyde. Their properties depend mainly on the cross linked density(the ratio of moles of cross linking agent to the moles of polymer repeating units). Formation of hydrogel network requires a critical number of cross links per chain and it forms porous structure whose pore size depends upon swelling of beads which in turn depend on external environment like pH, temperature etc. Chitosan is a highly basic polysaccharide. Beads obtained from chitosan after crosslinking with glutaraldehyde are solid, spherical, micron or nano sized constituting a matrix type of structure. The beads obtained from hydrophobic polymers have found to be higher uptake as compared to the

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in beads prepared from more hydrophilic surfaces6. So nano/micro beads surface charges increased hydrophobicity of polymeric matrix have been found effective in a positive sense to utilize them as an adsorbent for removing pollutants from water effluents. Our study is an attempt to develop chitosan beads cross linked with glutaraldehyde for adsorbingdyes as a model adsorbent to investigate itsmodeling color removal efficiencyand adsorbing properties. We planned a study to obtain beads of chitosan crosslinked with glutaraldehyde which may be fruitful for further studies. We made an effort to study characteristics, swelling behavior and adsorbing capacity of the cross linked chitosan beads for dyes. Experimental Chitosan, a natural polymer of animal origin was purchased by India Sea Food, Kerala, and was used as received. Its percentage of deacetylation after drying was 89%.Glutaraldehyde was procured from Loba Chemie Pvt.Ltd ,India and used as a crosslinking agent between chitosanchain units of polymer. All other chemicals like acetic acid, methanol, NaOH, HCl, KCl, KH2PO4 etc. were used of analytical grade. Double distilled water was used throughout the studies. Preparation of chitosan beads Chitosan (1.0 g) was dissolved in 40 ml of 2% acetic acid under stirring condition for 3h at room temperature. The homogeneous mixture was extruded in the form of droplets using a syringe into NaOH-methanol solution (1:20 (w/w)) under stirring condition at 400 rpm. The resultant beads were then placed in a water jacket containing glutaraldehyde maintained at 50oC for about 10 minutes. Finally the beads were washed with hot and cold water successively and then vacuum dried 7. Swelling studies Swelling behavior of chitosan beads was studied in different pH (2.0, 7.4 and 10.0) solutions. A known weight (2.0 g) of the prepared beads was placed separately in the conical flask containing media for required period of time. After predecided time period the swollen beads were collected and their net weight were determined by first blotting the beads with filter paper to remove adsorbed water on the surface. The percentage of swelling for each sample at time t was calculated using the following formulaPercentage of swelling = {(Wt–Wo)/Wo} x 100 Where, Wt = weight of the beads at time t after emersion in the solution. Wo= weight of the dried beads. Color removal efficiency (CRE) 50 ml dye solution of known concentration with 1.0 g of beads was taken in conical flask at desired temperature and pH. It was shaken for 10 min and then kept for 24 h and lastly the remaining concentration of dye was estimated spectrophotometrically at λ max of used dye. The CRE (%) for CS beads was calculated by given formula (9)-

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in CRE (%) = (1 – Ac/Ai) X 100 Where, Ai and Ac are the absorbance of the dye solution before and after the adsorption process. Fourier transformed infra red (FTIR) spectroscopy Dried samples were ground into powder and crushed with KBr for homogenization. A Nicolet economy sample press was used to obtain optically clear pellets. Pellets were analyzed using transmission FTIR using a Thermo Nicolet Avatar 370 FT-IR spectrometer system. Dry air was used as the chamber purge stream for all samples. The FTIR spectra were obtained at room temperature over a spectral frequency range of 400-4000 cm-1. IR bands are expressed in terms of frequency (cm-1). The background was obtained against a pure KBr pellet and the data was analyzed by Omnic software. Functional groups present in the raw material and products were determined. Scanning electron microscopy (SEM) The shape and surface morphology of the beads were examined using FESEM QUANTA 200 FEG model “(FEI, The Netherlands make)” with operating voltage ranging from 200 V to 30 kV. FESEM micrographs were taken after coating the surfaces of bead samples with a thin layer of gold by using BAL-TEC-SCD-005 Sputter Coater (BAL-TEC AG, Balzers, Liechtenstein Company, Germany) under argon atmosphere to make the sample conducting. The surface appearance, shape and size of scanning electron micrograms were used to perform textural characterization of full and cross sectioned IPN beads. Magnifications were applied to each sample in order to estimate the morphology and interior of the bead. Thermal analysis Thermal gravimetric analysis (TGA), Differential thermal gravimetric (DTG) and Differential thermal analysis (DTA) were carried out simultaneously by using a (PYRIS Diamond). TG/DTA thermal analyzer model DSC-7, supplied by Perkin Elmer and the data was processed and analyzed by PYRIS muse measure and standard analysis software (V. 3.3U; #. 2002 Seiko instruments inc). The sample was kept in Alumina pan, the reference material was Alumina powder and study was carried out at heating rate 10°C/min under 200 ml/min flow rate of air or nitrogen atmosphere. Indium and gallium were used as standards for temperature calibration. The measurements were run from room temperature to 600oC. The thermal stability of produced beads was assessed. X-ray diffraction (XRD) X-ray diffraction studies were performed by using Bruker AXS D8 Advance using CuKα Nickel filter and Copper as target at wavelength of 1.54 Å with goniometer and speed was kept at 2°/min. Wide angle X-Ray scattering patterns of the samples were obtained using DIFFRAC plus XRD commander software and analysis was done by DIFFRAC plus (version 8.0) software. The range of scanning angle for the sample was kept in the range 2θ of 10 – 60o. Results and Discussion

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in The CS beads are almost round in shape when freshly prepared but drying results them in uneven shape and also decreased their volume. The original shape of beads was restored after swelling. The color of uncrosslinked beads was creamy which was changed into yellow after crosslinking with glutaraldehyde. The yellow colour of crosslinked beads was turned into brown after air drying or oven drying.

(a)

(b)

Figure 1–Photographs of freshly prepared (a) and dried (b) chitosan beads Fourier transform infra red spectroscopy (FTIR) FTIR curve for chitosan exhibited a broad peak at 3450 cm-1 which was assigned to –NH– stretching vibration which might be due to deacetylation of chitosan. The peak at around 3500 cm-1 due to hydrogen bonded O-H vibrational frequencies and O-H bond stretch of gluco pyranose units. Peaks at 1639 cm-1 and 1319 cm-1 were observed due to >C=O stretching of amide bond. The peak at 1613 cm-1 was assigned to strong N-H bending vibrations of secondary amide8. Bands at 2919 cm-1 and 2810 cm-1 represent the aliphatic C-H stretching vibrations. The observed sharp peak at 1384 cm-1 is due to CH3 symmetrical deformation mode9-10.Two peaks around 894 cm-1and 1171 cm-1 appeared in spectra corresponding to saccharide structure11. A broad band appearing near 1083 cm-1 indicated the >CO-CH3 stretching vibration of chitosan.

CHITOSAN Sun Oct 20 22:16:42 2002 (GMT-05:00)

BD

CS

3500

3000

2500 2000 Wavenumbers (cm-1)

133

1500

1000

500

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Figure 2-FTIR of pure chitosan powder (CS) and chitosan Beads (BD) The structural changes in the beads were investigated using infra-red spectroscopy. It was concluded that during crosslinking, formation of the C=N group of the imine involved a reaction between the amino group of chitosan chain with aldehyde group of glutaraldehyde. Thermal analysis TGA curve for chitosan indicates that it decomposed in two steps. Chitosan has shown approximately 10% weight loss below 100oC. This step involves a smaller weight loss, may be due to initial loss of water molecules, after this no weight loss occurs upto 249oC. A sudden weight loss is observed after 249 oC and the total weight loss at 400oC is about 60 %. This destruction of chitosan moiety occurs in the second step. TGA curves of beads clearly shows that pure chitosan crosslinked beads lost weight after 2000C and total weight loss at 4000C is approximately 46%. This indicates that crosslinking of chitosan with glutaraldehyde increases its thermal stability. 290Cel 1.425mg/m in

0.000

100Cel 89. 06 %

200Cel 88.04 % 275Cel 82.05 %

80.00 300Cel 57.05 %

-80.0 50

100

150

200

250

300 350 Temp Cel

400

450

500

60.00

uV D TA

68.6 mJ/mg

120 mJ/mg

-5.0

-10.0

2 0 0C e l 8 9 .6 8 %

5 0 4 Ce l - 7 .8 u V

4 1 2 Ce l - 8 .5 u V

1 0 0C e l 9 9 .5 3 %

100.00

-1.500

2 6 4C e l 7 2 .0 9 %

-2.000

3 9 8 Ce l 5 3 .7 0 %

-8.00 -20.0

5 9 7 Ce l 2 2 .6 2 %

50.00 -2.500

5 0 0 Ce l 2 8 .8 1 %

40.00 -25.0

550

Figure 3– Thermal analysis of chitosan.

-1.000

2 3 Ce l 1 0 0 .0 0 %

-6.00

-10.00

-0.500

150.00 -57.4 mJ/mg 1 41 C el - 2. 4 u V

D T G m g /m i n

0.0

-15.0

589Cel 32.47 % 350Cel 44.09 % 400Cel 450Cel 40.21 % 37.68 % 500Cel 550Cel 35.62 % 33.81 % 325Cel 47.88 %

-60.0

-4.00

97.5 mJ/mg

D T G m g /m i n

100.00

249Cel 87.14 %

%

-152 mJ/mg

120.00

2 4 0 Ce l 2 .6 u V

-2.00

TG

140.00

65Cel -7.4 uV 17Cel 99. 95 %

-40.0

200.00

5.0

268 mJ/mg

0.500

1 59 C el 0 .1 5 0m g / m in

0.00

160.00

296Cel 12.9 uV

TG

uV D TA

-20.0

2 4 4 Ce l 0 .3 9 5 m g / m i n

10.0

20.0

0.0

4 1 9 Ce l 0 .6 1 0 m g / m i n

15.0

2.00 180.00

63Cel 0.169mg/ min

%

40.0

50

100

150

200

250

300 350 Temp Cel

400

450

500

550

Figure 4– Thermal analysis of chitosanbeads

DTG thermograms of pure chitosan indites one peak at 290 oC corresponding to higher rate of weight loss about 1.4 mg/ min. In DTG thermograms in case of crosslinked beads showed lesser rate of weight loss at 2440C. DTA thermograms for chitosan powder showed endothermic peak at 65oC due to loss of free water and one exothermic peak at 296oC due to chemical transformation.In case of crosslinked beads only one exothermic peak is observed and no endothermic peak is observed by chitosan powder. X-Ray diffraction The diffraction pattern of pure chitosan has the characteristic peaks at 2θ of 12 to 16, 20 and 29.Xray diffractograms of beads clearly show the similarly peaks as chitosan powder

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BD

CS

Figure 5- X-ray diffractograms of chitosan (CS)& chitosan beads (BD) Scanning electron microscopy The surface appearance, shape and size of scanning electron micrograms were used to perform textural characterization of full and cross sectioned IPN beads. Magnifications were applied to each sample in order to estimate the morphology and interior of the bead.

(a)

(b)

(c)

(d)

Figure 6– SEM micrographs of full chitosan bead (a), magnified bead (b), cross sectioned bead (c) and magnified cross sectioned bead (d)

SEM micrographs of full dried beads with their magnified (2000 X) surface morphology are shown in Figure. It was concluded from these Figures that the beads were nearly spherical or some what oval in shape. The approximate size of beads is 164 µm. They had rough, rubbery fibrous and folded surfaces with wrinkles. The micrographs showing internal structure of beads with their magnification are obtained by half cut beads which are presented in Figure. Interior of the beads appeared to have micropores and micro tube like interior spaces which confirms the highly porous structures of polymeric beads, although they appeared solid externally.

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International Science Congress Association www.isca.in , www.isca.co.in , www.isca.net.co , www.isca.net.in Swelling studies The percentage swelling of pure chitosan beads cross linked with glutaraldehyde in solution of pH 2.0, 7.4 and 10.0 is shown in Figure 7. It was observed that swelling rate increases with increasing pH. When the cross linked beads were placed in the solution, the solution penetrates into the bead and the bead subsequently tries to swell. Generally, the swelling process of the beads in pH