Experiment 11 - Digestion of Foodstuffs

DEPARTMENT OF PURE AND APPLIED CHEMISTRY Visayas State University, Baybay, Leyte CHEM31a Biochemistry Laboratory Report

Name : Mark Ryan R. Tripole Course/Yr : BS Chemistry II Group No : 6

Date Performed : 05/27/2015 Date Submitted : 06/01/2015 Score

Experiment No. 11 Digestion of Foodstuffs OBJECTIVES  

Identify digestion as a process of hydrolysis Determine the hydrolysis products of carbohydrate, fat and protein digestion

I. Results Shown below are the results as obtained from the experimental procedure performed:

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II. Discussion INTRODUCTION Digestion basically refers to the process by which large food molecules (which are insoluble) are broken down into their smaller components that can be absorbed by the bloodstream. This form of digestion can be broken down into two categories based on where they occur – one being the mechanical digestion that occurs in the mouth when it is physically broken through chewing and the chemical digestion that occurs once the food is broken down by the digestive enzymes in the body. This laboratory report will focus only on the latter, wherein the experimental procedures are simulations of the action of these digestive enzymes and the effect that they have on the different components that they were meant to break down. Three molecules of note that was tackled in the experiment were carbohydrates (broken down by amylase and other enzymes), fats (broken down by lipase) and proteins (broken down by proteases like pepsin for example). The following sections will talk about each and an interpretation of the data will be given as well. PART A – DIGESTION OF CARBOHYDRATES Carbohydrates are essential components for the nutrition for the body because it is the prime source of energy for one’s day-to-day activities as well as ensuring the functionality of the body systems. Carbohydrates can either be simple, such as the ones obtained from consumption of food such as fruits and starch sources, or complex, which can be obtained from food such as unrefined pasta and whole grains. Regardless of the source, carbohydrates undergo a unique process of digestion within the body, which starts at the mouth through the saliva. When food is consumed, the enzymes that are present in the saliva, known as salivary amylase, go to work on the food and break down the carbohydrates present. These are broken down into much simpler sugars (disaccharides and trisaccharides) which then travel through the stomach when swallowed. Progressing through, the partially broken down food then ends up in the stomach where they are broken down into more “manageable” proportions by the hydrochloric acid present in the system. The food then passes through into the small intestine where the key stages of carbohydrate digestion can be found. The bulk of the breakdown occurs in the small intestine, where pancreatic amylase breaks down any of the long carbohydrate chains into much simpler forms of sugar. Augmenting this effect is also the presence of other carbohydrate interacting enzymes such as Page 3 of 7

maltase, which aids in the complete breakdown of the carbohydrates into either glucose or fructose. These two sugars are of prime importance in this step because they are absorbed into the bloodstream and are ultimately sent to various parts of the body such as muscles, organs and tissues where they are used in the body’s metabolic processes. The hydrolysis or “digestion” of carbohydrates was observed in the laboratory through measuring the amount of time it took for a particular amount of saliva to partially hydrolyze a solution of starch, which was tested through with the use of iodine. Based on the results from the experiment, it took 8 to 10 minutes for there to be a noticeable change in the results. The sample with starch only remained dark blue-black for the whole of the first session because there are no enzymes to hydrolyze them into their smaller components. At around this mark, the sample with the saliva started getting tinges of brown, meaning that there is evidence of the breaking down of the long polysaccharide chains. A further test was performed to support this by performing the Benedict’s test on both the samples. The starch and saliva mixture produced a reddish—orange precipitate, which is an indication of the presence of reducing sugars which in this case would be either glucose or maltose. The starch only solution on the other hand would give a negative result because starch is a long polysaccharide and is not a reducing sugar. PART B – DIGESTION OF TRIACYLGLYCEROLS/FATS Fats have a variety of uses in the human body and are important substances in living organisms. It can act as an alternative source of energy when there is a scarcity of carbohydrates, and this energy can be used in the different bodily functions. They also ensure the proper functioning of the nerves, where myelin is a fatty material that wraps around the nerve cells and facilitates neural transmission. Moreoever, fats are transport carriers for the fat-soluble vitamins A, D, E and K and can carry them to the parts of the body that need them through the bloodstream. The digestion of fat follows the same pattern as carbohydrates save for a few differences. Fats are first digested in the mouth relatively simply by what are known as lingual lipase, and when the move along into the stomach, gastric lipase does it’s part in breaking down the fats further while the stomach does it’s part by mixing and churning it’s contents. By this point, the fats in the food are broken down into smaller fragments called globules so they can be distributed around evenly. The partially digested food then basically moves on to the small intestine where it is broken down further mechanically through interaction with the bile acids that are secreted by the gall bladder. There is also the action of what is known as pancreatic lipase on the food, which breaks the fats into smaller components of diglycerides, monoglycerides or free fatty acids. This interaction with bile salts and digestion by lipase are two of the processes that were looked at and simulated in the experimental procedure. The first test involved observing how bile aids in the digestion of fats in the body. One sample contained only water and oil, while the other contained a sample of oil and an aqueous solution of bile. After having left both samples to stand for 30 minutes, the sample of oil and water remained separate from with each other, while the sample with the bile showed the conversion of the oil into smaller globules that are miscible in the solution. Bile basically acts as a surfactant that emulsifies the fats in a process similar to that of soaps by the formation of micelles. They prevent the reaggregation of the globules back into larger portions. The other test involved observing the digestion of fats in milk through the use of lipase. A sample of condensed milk was obtained to which a part of the meat of an animal that contained the enzymes necessary for fat digestion was added. This was then placed in a water bath heated up to physiological body temperature (37°C), 10mL samples taken every 20 minutes for an hour, the pH of each sample roughly estimated using pH paper and each sample titrated with dilute sodium hydroxide (with the use of phenolphthalein indicator). Keep in mind that through the course of digestion, the products formed are fatty acids, and so it would be expected that there is a drop in Page 4 of 7

the pH level. The results contradict this though, because the pH paper showed the same colors for each sample throughout (where the milk rated at around pH 6). As for the volume of NaOH, the trend would be that the amount would increase because as time progresses, more and more of the fat is converted into the free fatty acids, which means there is more acid to neutralize. The results, however, show a decrease in the amount of base used. This can be attributed to not allowing for enough time for the enzyme to do its part, or could be just plainly due to human error in the measuring process. PART C – PROTEIN DIGESTION Proteins are essentially large, complex biomolecules that play many a critical role in the body. They do most of the work in the cells are components that are required for structure, function and regulation of the body’s tissues and organs. To further illustrate the importance of proteins in the body, they also function as antibodies to help fight off disease, as biological catalysts to chemical reactions that take place in the cells, as messengers in the coordination of biological processes between different cells, as structural components and as transport molecules. The pathway of digestion basically starts at the mouth, where a small amount of the proteins are broken down by the enzymes that are present in the saliva. The real show occurs at the stomach, where the acidic pH of the stomach allows for the denaturation of the proteins in the food causing them to unfold and deform. This unfolding is important because it allows the enzymes to perform much better at breaking down the proteins. This is done by a protease that is known as pepsin (which initially exists as pepsinogen until it is activated by the hydrochloric acid), which also relies on the low pH of the stomach for its activation. Pepsin breaks down the large, bulky and unfolded protein into small pieces of proteins that are known as peptides. After this portion of the digestion, the food moves out and the pancreas helps in neutralizing the acidic mixture and activate some of the other protein hydrolyzing enzymes. In the small intestine, the pancreatic enzymes function to break down any of the remaining proteins, where they are converted into the smaller amino acids and are absorbed through structures in the lining called villi, then from there enter the bloodstream for transportation to the cells as needed. This was observed in the experiment through the usage of the white part of a hard-boiled egg. Three small pieces were taken and then placed in three separate test tubes containing three different test tubes. One with water and hydrochloric acid, a second with only pepsin, and a third with pepsin and hydrochloric acid. Observing the results, the test tube with the pepsin and hydrochloric acid deformed the egg white to a great extent, indicating the presence of proteins in egg whites. The test tube with pepsin only showed little activity with only minute deformation of the egg white, while the water and HCl combination showed no reaction whatsoever. The main reason why the pepsin alone doesn’t work is because without the aid of HCl to provide the acidic pH level, pepsin is inactive and dormant, meaning it doesn’t function to break down proteins at all. The pepsin and HCl broke down the egg white because it is a simulation that closely resembles the conditions of the stomach.

III. Conclusion Digestion is an essential process in living organisms, because it is by this process that these living organisms get all the nutrients that are needed in order to ensure survival. The experiment has helped in shaping the definition of what digestion really is and just how important it really is. In the most basic of context, all digestion really is is the breaking down of larger molecules into the smaller and more essential molecules that can be easily absorbed by the body. And all of this “breaking down” mainly occurs through the action of enzymes that catalyze the hydrolysis of these larger

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molecules. Moreoever, gaining the knowledge of the different products that come about as a result of digestion has been important into seeing exactly which of these biological molecules have the most impact on the human body, as well as how a deficiency of them can potentially affect the physiological conditions of the organism. Carbohydrates produce glucose that is essential for energy, fats are essential for neural transmission, and proteins are diverse and have a lot of functions.

IV. Answers to Questions PART A – DIGESTION OF CARBOHYDRATES 1. Where does starch digestion begin? The hydrolysis of starch begins in the mouth which is done by salivary amylase, but this is only small in comparison to the extent performed by the pancreatic amylase in the small intestine. 2. What carbohydrate digestion occurs in the small intestine? The breakdown of the longer carbohydrate chains facilitated by the pancreatic amylase and the supporting role of maltase in the breakdown of the shorter chains already broken down from the preceding processes. 3. What are the end products of carbohydrate digestion? Glucose and fructose are the end products of carbohydrate digestion. 4. Why do we need a digestive process? Having a digestive process is essential because it is the only pathway available for obtaining the important biomolecules that the body needs in order to survive. As has been discussed in this laboratory report, the different processes observed make products that are essential for human life, carbohydrates for energy, fats for proper nerve function, and proteins for functions such as in antibodies. PART B – DIGESTION OF TRIACYLGLYCEROLS/FATS 5. What is the effect of bile salts on an oil and water mixture? The bile emulsifies the oil into smaller globules and allows both of them to mix. This occurs in a mechanism similar to the concept of soap micelles. 6. What is the function of bile salts in the digestion of fats and oils (triacylglycerols)? Bile/bile salts act as a surfactant that aid in breaking down larger pieces of fat into smaller manageable pieces which are more readily observed than when they are larger in size. 7. What products of lipase action would change the pH of a mixture containing triacylglycerol? The production of free fatty acids that come about from lipase action on fats tend to change the pH levels of the mixture by virtue of them being acidic in nature because of the carboxylic acid group (-COOH) Page 6 of 7

PART C – PROTEIN DIGESTION 8. Why does a person with low protein production of stomach HCl have difficulty with protein digestion? This is because the lack or diminished amount of stomach HCl means that there are no suitable conditions for the activation of pepsinogen into pepsin, which is the major protease in the breakdown of proteins in the body. This leads to little to no protein digestion and could lead to various bodily abnormalities. 9. What are the products of protein hydrolysis? The end products of the protein hydrolysis are the amino acids that made them up.

V. References       

"What is Digestion?" http://www.news-medical.net/health/What-is-Digestion.aspx "Digestion of Carbohydrates" http://www.livestrong.com/article/417962-how-does-the-body-digest-carbohydrates/ "Bile" http://en.wikipedia.org/wiki/Bile "Digestive Enzymes" http://www.nutritionexpress.com/showarticle.aspx?articleID=1209 "Pancreatin" http://www.drugs.com/mtm/pancreatin.html "Secretion of Bile and the Role of Bile Acids in Digestion" http://www.vivo.colostate.edu/hbooks/pathphys/digestion/liver/bile.html "Digestion 101" http://www.functionalfitmag.com/blog/2011/12/21/digestion-101

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