Chromatography Lab Answers

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Purpose

The purpose of the experiment is to determine the specific types of pigments found in a beat leaf and in a spinach leaf by using paper chromatography and two solvents: water soluble solvent and lipid soluble solvent.

Hypothesis

If a water soluble solvent is present, then there will be the movement of only the water soluble pigments up the chromatography paper. This happens because as the water travels up the paper, the water soluble pigments’ bonds become attracted to the water molecules due to dipole-dipole forces. Through capillary action these pigments will travel up the paper until the bonds between the water and pigment become so weak that the pigment must break the attraction and leave itself imprinted at a certain height up the paper. On the other hand, Lipid soluble pigments won’t move due to the molecules’ lack of polarity. Therefore, they will remain at the concentrated area unless a lipid soluble solvent is present. If this lipid soluble solvent is present, as opposed to the water soluble solvent, then the lipid soluble pigments will move up the chromatography paper instead of the water soluble pigments. The same idea will happen. The lipid soluble pigments will travel up the paper until their bonds between the water are so weak that it must stop following the movement of the solvent, and get placed at a certain height above the original concentrated dot. Spinach will have mainly chlorophyll A and B because the leaf is completely green, compared to the beat leaf which consists of both a red and green shade showing that other pigments are present in this leaf.

Calculations:

Discussion

• In the beat leaf/petroleum ether acetone trial, the carotene traveled the fastest because it displaced the furthest distance in a period of time.
• In the beat leaf/petroleum ether acetone trial, the carotene pigment travelled the furthest with a displacement of 9.0 cm [up].
• Chlorophyll a and b are two common types of chlorophyll found on the thylakoid membrane in several photosynthetic units. Both have a similar purpose: to trap light to ultimately convert it into energy. The molecules do this by absorbing specific wavelenghts of light rays (red and blue-violet light) which excite electrons contained within the double bonds of the porphyrin ring of the chlorophyll molecule. Chlorophyll b passes its exited electrons to chlorophyll a, which passes its excited electrons to other molecules, which store the energy as chemical potential energy. This is the base step of photosynthesis. Cholorphyll b is an accessory pigmant, meaning it always passes its excited electrons to chlorophyll a. Chlorophyll a is the main absorber, meaning only it can pass the excited electrons to other molecules. Leaves appear green because chlorophyll  is a very poor absorber of green wavelengths, therefore it reflects green light the best. Because leaves are very rich in chloroplasts containing chlorophyll for photosynthesis (photosynthesis occurs mainly on leaves), they reflect a lot of green light wavelengths, which our eyes interpret as the colour green.
• As the fall comes so do the shorter days. This results in less light during the day for the plant to undergo photosynthesis. This is where the plant’s leaves begin to change colour. This allows the accessory pigments (carotene, xanthophyll) to take over the light capturing process because they are able to trap different wavelengths of light. The combined trapping ability of the accessory pigments and the regular pigments (chlorophyll A and B) allows for a greater combined absorption of light, becuase there are more pigments trapping a greater range of light from the visible light spectrum. This results in the plant being able to perform more photosynthesis because more photons of light are being absorbed which will result in more glucose being formed which will ultimately be stored as potential energy (in the form of sugars) during the winter months when photosynthesis doesnt occur.
• Paper chromatography is a process in which pigments are seperated, from an initial concentrated solution, through the process of capillary action. A solvent is placed at the bottom of the paper. As the solution travels up the paper, like soluble pigments will travel with the solvent until the bonds between the solvent and pigment become so weak that it must break the attraction and imprint itself a certain height up the paper.
• Some factors that affect paper chromatography are:

Solvent: the solvent is a majoy factor that plays in the outcome of the experiment. A certain solvent will only attract certain pigments up the paper. For example this experiment used water soluble and lipid soluble solvents. This means that when one of these solvents are present in the trial, only that type of pigment will travel with the movement of the solvent (lipid soluble solvent with lipid soluble pigment and water soluble solvent with water soluble pigment).

The beat leaf contained more pigments in the leaf compared to the spinach leaf. This could be because the beat has a root where it stores starch. This would require it to undergo photosynthesis more times in order for its to create more glucose to store. The increased number of accessory pigments allows for a wider range of light to be taken in and converted into energy (excitation of electrons on the chlorophyll) for the photosynthetic process. In comparison to the spinach leaf, this plant doesn’t have a massive storage “unit” at the base of the plant. Therefore it doesn’t need to undergo photosynthesis as rigerously so it doesn’t require as many accessory pigments.

Conclusion

The experiment was carried out and it was proven that:

Beat leafs contain: Carotene, Xanthophylls, Chlorophyll A and B, and Anthocynin as pigment in the leaf.

Spinach leaf contain: Chlorophyll A and B.

This was shown when the paper chromatography was done, the beat leaf trials had many different coloured pigments over the paper, vs the spinach leaf only a green pigment which represented that only chlorophyll was present.