I then placed the double-sided sellotape onto the Gas Jar Lid and folded the other end of the chromatography paper and stuck it to the sellotape so that the saturated end of the chromatography paper was dangling just above the bottom of the Gas Jar, this is so that when I put the solvent into the Gas Jar it was just submerged into the solvent. So as I just mentioned I then put the solvent into the bottom of the Gas Jar, so that it could rise up the chromatography paper, put then lid on so that the vapours from the solvent were trapped in the Gas Jar and left it overnight.
I then removed the chromatography paper from the Gas Jar and sprayed it with Ninhydrin in a fume cupboard I have to do this because amino acids have no colour; therefore I have to spray on a revealing agent (Ninhydrin) so that we can see them. I then drew a line on it when it was thoroughly dry to mark where the solvent had run too (called the solvent front). I then measured the distance from the solvent front to the start line (where I had originally dropped the egg/protease mixture) this is the distance moved by the solvent front. I then measured the distance from each spot of amino acid to the start line, this is the distance moved by each by the spot. I will then work out the Rf Factor for each spot of amino acid and compare the Rf factors to the table and work out what amino acid each spot is.
Once the experiment had been carried out on the Chicken egg I then repeated the exact same process for the Duck egg so the that it was a fair test.
Prediction
My prediction is that the egg white’s will contain many different amino acids and my prediction is that they will have over 8 different amino acids present in them; I think this because an egg white contains approximately 40 different proteins so obviously when it contains that many proteins it will contain a lot of amino acids as they are what makes up the proteins but in different formations.
Diagram
Gas Jar Lid
Gas Jar
Chromatography Paper
Solvent
Results
Chicken egg
Rf = Retention Factor
Rf = x
y
x = Distance from starting point to spot
y = Distance to solvent front
Distance to solvent front/y = 166mm
Distance to amino acid/x1 = 86mm
Distance to amino acid/x2 = 132mm
Calculation for the Rf factor of x1
Rf factor of x1 = 0.158
166mm = 0.158
86mm x1 = Methionine
Calculation for the Rf factor of x2
Rf factor of x2 = 0.795
166mm = 0.795
132mm x2 = Leucine
Duck egg
Rf = Retention Factor
Rf = x
y
x = Distance from starting point to spot
y = Distance to solvent front
Distance to solvent front/y = 220mm
Distance to amino acid/x1 = 25mm
Distance to amino acid/x2 = 60mm
Distance to amino acid/x3 = 97mm
Distance to amino acid/x4 = 132mm
Calculation for the Rf factor of x1
Rf factor of x1 = 0.113
220mm = 0.113
25mm x1 = Lysine
Calculation for the Rf factor of x2
Rf factor of x2 = 0.272
220mm = 0.272
60mm x2 = Serine
Calculation for the Rf factor of x3
Rf factor of x3 = 0.440
220mm = 0.440
97mm x3 = Tyrosine
Calculation for the Rf factor of x4
Rf factor of x4 = 0.60
220mm = 0.60
132mm x4 = Valine
Analysis
The proteins in an egg white are globular proteins, which means that the long protein molecule is twisted and folded and curled up into a more or less spherical shape. A variety of weak chemical bonds keep the protein curled up tight as it drifts placidly in the water that surrounds it. When these proteins are broken up into amino acids by the protease the amino acids all become free and so we are able to perform the chromatography test on them. From my results I can gather that only two amino acids showed up on the chromatography paper for the chicken egg, the two amino acids that showed up were Methionine and Leucine the results from my test lead me to deduce that from the two Leucine was the lighter molecule this is because in chromatography the lighter molecules travel further than the heavier molecules. From the results of the Duck egg though I can clearly see that more amino acids showed up on the chromatography paper these were Lysine, Serine, Tyrosine and Valine out of these Valine had the lighter molecule because it travelled further along the chromatography paper, lysine therefore has the heaviest molecule because it moved the shortest distance along the chromatography paper, serine was the second heaviest molecule followed by tyrosine up o valine the lightest out of the four.
Evaluation
Overall I think my experiment was pretty fair although I think that a couple of things affected the results; I think that my prediction was correct in the fact that although there were only two amino acids in the chicken egg white and four amino acids in the duck egg white that showed on the chromatography paper I think that because the Rf factors of them were so close that they were indistinguishable on the paper and so I think that there were more amino acids present but they could not separate in the time scale that I did the experiment in. I think to correct this we could do the experiment for longer as this would let the amino acids have longer to separate out into there respective positions on the chromatography paper, I also think that I could have also done a two way paper chromatography where by which a chromatogram is turned by ninety degrees, and placed in a different solvent in the same way as before; some spots separate in the presence of more than one pigment. As before, the value is calculated, and the two pigments are identified. I could have also used a square of chromatography paper and done the experiment turning it 90 degrees each time until four results are shown on the paper.
From research, I have found that chicken eggs contain 17 different amino acids, this is shown in the table below and so in my prediciton I do not think I was entirely wrong as it may not have shown all of these on the chromatography paper but I have already discussed a possible reason for this.
Table from:
