Bio 120 exercise 4

In Figure 4.1, paper chromatography was used to separate the different photosynthetic pigments from San Francisco leaves. In this part of the experiment, 9:! Petroleum:ether was used as the mobile phase where the mobility due to solubility of the extracted photosynthetic pigments were based. Results showed 4 photosynthetic pigments namely: yellow green, green, yellow and orange. From these results, we can say that the pgments yellow green and green were the least soluble and the pigments yellow and orange were the more soluble ones. Also, we can now say the identity of the pigments. Yellow green and green pigments are most likely chlorophyll pigments while the other two are carotenoids or xanthophylls. Based from the data gathered in the spectroscopy of the photosynthetic pigments, the absorption spectrum was made. The relationship betwee the absorbance of the different pigments and wavelength was determined to be able to visualize their absorbance peaks. Results showed that the yellow green pigment peaked at 400 nm with absorbance of 0.135. for the green pigment, it peaked at 425 nm at 0.047. for the orange pigment, it peaked at 650 nm with absorbance of 0.19. lastly, the yellow pigment peaked at 475 nm with absorbance of 0.107. with these given absorbance values, we can now determine the identities of the said pigments. In order to identify the pigments, the wavelength for the thighest absorbance of each pigment. The peak of the spectrum would indicate the maximum absorbance of the pigment in that certain wavelength. According to published literature, most carotenoids has a maximum absorbance value of 450 – 500 nm. In this case, we conclude that the yellow pigment was a carotenoid since it peaked at 475 nm. The yellow and the orange pigment along with the yellow green and green pigments have small difference in the absorbance values and we ca say that the identity of these pigments are the same. In this experiment, the values suggest that the yellow green and green pigments are most likely chlophylls. Specifically, the yellow green pigment would be chl b and the green pigment id chl a. Figure 4.2 shows the standard curve made from the data gathered in the absorption of different concentrations of Methylene Blue. The calculation of the concentration of the unknown substance was done using the Lambert beer law. With given concentrations and absorbance values, we were able to compute for the concentration of the unknown using the formula for c. the calculation shown in the worksheet is near to the interpolated value, this supports that the computed concentration from the Lambert Beer law is correct. Based from these results we can now say that the absorption and the concentrations of given samples have a direct relationship. As the concentration of the sample increases, the value of absorbance also increases. At higher concentration, there is more electrons, therefore the higher the amount of light the pigments are able to absorb. Hence, higher concentrations of the sample result to higher absorbance value. In order to compute for the concentration of a certain substance given that it initially registers at a high value and have a reading of 100%, one should dilute the sample first. With this, the number of particles detected by the spectrophotometer can be reduced and the initial reading would decrease. Also, the dilution should be

considered and should have an absorbance value that falls under the Lambert-Beer plot that has been constructed for comparison. After getting the absorbance of the diluted sample, we can now multiply the dilution factor from one of the points in the plot to be able to get the exact concentration of the unknown.