Chicken eggs are the most commonly add protein to one's diet as well as other nutrients . They supply all essential amino acids for humans, and provide several vitamins and minerals. They are also a cheap single-food source of protein. Protein are the most abundant biological macromolecules occurring in all cells in our body. Proteins are compounds, which contain carbon, hydrogen, oxygen, nitrogen, and sometimes sulphur. Each protein molecules is made from a large number of molecules of amino acids, joined together by peptide bonds.
Proteins by themselves are usually colourless and therefore it is difficult to measure the presence of protein. An easy way to solve this problem is to stain the protein with biuret reagent and measure the amount of absorption of the Biuret reagent by a spectrophotometer. The spectrophotometer is an instrument that is used to measure the intensity of the colour of a solution. When Biuret reagent reacts with peptide bonds it produces a violet colour which will be detected by the spectrophotometer. The violet colour indicates a positive Biuret test for protein, the more protein presents the darker the colour of the solution.
So, how should we determine protein concentrations in eggs? There are a number of ways. In many lab experiments, for example, determining the amount of protein biomolecules in a given solution is essential. In addition, the experiment is perform to determine if absorbance levels correspond with protein concentrations in solutions. In addition, it is very important that the method by which the protein concentrations of various substances were determined apply universally. In other words, the method must work in every test in which it is used. The method chosen for this experiment was the Biuret Assay method, which requires that the solutions being tested be mixed with a Biuret reagent and run through a spectrophotometer.
In order to determine how much protein is represented by a particular absorbance reading it is necessary to construct a standard curve.This is done by performing the biuret reaction on a serial dilution using six tubes. The absorbance readings obtained from these samples are used to construct a graph of absorbance as a function of protein concentration. This graph is called the standard curve for the assay, and can be used to convert the absorbance readings for the experimental samples into a protein amount. One of the sample in the list with no protein added is blank that should be used to zero the spectrophotometer. Blanks are useful when there are other substances in the experimental tube besides the substance we are trying to measure. Since those other substances are not the chemical that we are trying to measure, they often interfere with the absorbance reading of the chemical of interest. The way to do that is to use a blank. A blank contains all the substances in the experimental tube except the substance that is being measured. Then, before reading our experimental tube, we place the blank tube in the spectrophotometer.
Record the data from the biuret reaction on the standard protein concentrations in a spreadsheet. Make an x-y line graph of the results. The protein concentration should be on the x axis, and the absorbance on the y axis. The line on the graph relates the protein concentration to the absorbance of the biuret reaction. We use this standard curve to convert the absorbance that we measured back to the amount of protein in the sample given. Using the data from six samples, a standard curve was graphed. The remaining four samples, A, B, C and X, the unknown solutions, were run through the spectrophotometer. The results are shown below:
Table above shows the average results of absorbance from four unknown solution protein concentration resulted from the equation we got at the standard curve graph above.
Graph above shows the standard curve of absorbance vs concentration of gelatin (protein).
Table above shows the average results of absorbance from six different protein concentration (gelatin).Because the data of this experiment was used to create a standard curve graph for protein concentration of known solutions, it is possible to accurately identify protein concentrations in any number of unknown solutions. One implication of these results is that protein concentration and absorbance are linked, and correspond to each other, as was initially predicted. But, there are no differences between the grade and the concentration of protein. The grade only make difference in the quantity and shape of the egg and not the concentration of protein, which based to the experiment result we got above. There also is no difference in nutritive value between the different grades. Because production and marketing methods have become very efficient, eggs move so rapidly from laying house to market that you will find very little difference in quality between.
By Siti Maimun Ali Al Tamas
(D20071029415)
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