Today in class we did an experiment where we used thin layer chromatography.
Procedure:
1) Add a small spatula measure of anhydrous sodium sulphate(VI) to the bottom of a plastic eppendorf tube. This is to remove any moisture that may be present.
2) Place small pieces of a spinach in the bottom of the eppendorf tube. Add 5 drops of Solvent A. consisting of 2 parts ethyl ethanoate and 3 parts proponent.
3) Press/stir the spinach with the end of forceps to extract as much colour as possible.
4) Use a fine brush to place a small dot of the coloured solvent about 5mm from the bottom edge of the TLC plate. Mark this position on the side of the plate with a pencil.
5) Gently blow on the dot to dry it before pitting another dot of coloured solution on the top of it. Continue to do this until the dot is not to faint but not to strong.
6) Use a dropper pipette to place o.5cm^3 of Solvent B into a clean vial so that the depth is about 5mm. The level of the solvent must be below the level of the dot on the chromatography plate.
7) Put the TLC plate into the vial and replace the screw-top lid.
8) After a few minutes, when the 'solvent front' has moved 7/8ths of the way up the plate, remove the plate from the vial using forceps and let it dry. Mark the position of the solvent front.
What the results mean:
In order of line appearance from the top end of the TLC plate, where the solvent front is.
- First seen is the B-catorene
- Second is Pheophytin A
- Then is Pheophytin B
- Followed by the Chlorophyll A
- After is Chlorophyll B
- Then Lutein
- And lastly follows any other Xanthophyll's
Procedure:
1) Add a small spatula measure of anhydrous sodium sulphate(VI) to the bottom of a plastic eppendorf tube. This is to remove any moisture that may be present.
2) Place small pieces of a spinach in the bottom of the eppendorf tube. Add 5 drops of Solvent A. consisting of 2 parts ethyl ethanoate and 3 parts proponent.
3) Press/stir the spinach with the end of forceps to extract as much colour as possible.
4) Use a fine brush to place a small dot of the coloured solvent about 5mm from the bottom edge of the TLC plate. Mark this position on the side of the plate with a pencil.
5) Gently blow on the dot to dry it before pitting another dot of coloured solution on the top of it. Continue to do this until the dot is not to faint but not to strong.
6) Use a dropper pipette to place o.5cm^3 of Solvent B into a clean vial so that the depth is about 5mm. The level of the solvent must be below the level of the dot on the chromatography plate.
7) Put the TLC plate into the vial and replace the screw-top lid.
8) After a few minutes, when the 'solvent front' has moved 7/8ths of the way up the plate, remove the plate from the vial using forceps and let it dry. Mark the position of the solvent front.
What the results mean:
In order of line appearance from the top end of the TLC plate, where the solvent front is.
- First seen is the B-catorene
- Second is Pheophytin A
- Then is Pheophytin B
- Followed by the Chlorophyll A
- After is Chlorophyll B
- Then Lutein
- And lastly follows any other Xanthophyll's