Separation of lipids using thin layer chromatography.
Virtual experiment.

Samples 1 and 2 for this experiment are lipid mixtures extracted from several tissues, which composition we wish to analyse.

The code for this particular sample set is

To guide identification, 5 standards are available: solutions of a pure lipid in a 1_1 mixture of chloroform and methanol:

• C: cholesterol
• CE: a cholesterol ester (cholesteryl linoleate)
• FA: a fatty acid (oleic acid)
• TAG: a triacylglycerol (trioleoylglycerol)
• PL: a phospholipid (a phosphatidylcholine)

Step of the experiment

1: Samples and standards are applied on the plate covered with silica gel (stationary phase for the chromatography)

2: The plate is introduced into the tank containing the mobile phase (a mixture of solvents), the tank is covered and we wait for the mobile phase to ascend; it is stopped by switching to step 3.

3: The plate is extracted, we wait for the solvent to evaporate and we proceed to reveal the lipids by exposing to iodine vapours.

Instructions

1. To move the capillary, hold it with the mouse pointer near its lower end.
2. Drag the capillary to one of the tubes containing either a sample or a standard, and release it.
3. Drag the capillary to the plate with stationary phase, positioning its lower end on one of the application targets (a, b, c, d, e, f), at the level of the lateral reference lines.
4. Drag the capillary to the waste container (a new capillary will later appear at the left-hand side)
5. Repeat the process with all samples and standards, applying each one on a different position. Write down what you have applied on each of the 7 positions in the plate.
   You may apply samples in any order and position.
6. Press the ▶ button to go to next step (nº2), where the chromatography will be automatically run.
7. Before the mobile phase reaches the top of the plate, press the ▶ button to extract the plate and proceed to the next step (nº3).
8. Press on the iodine bottle, wait for the colour to develop and press again on the bottle to remove the iodine.
9. You may press on the photo camera to capture an image that may be copied or saved to disk using the context menu of the web browser (the photo will also have stamped the code of the experiment, date and time).

In your lab notebook: write down the code assigned to your experiment (in this case it is and what you have applied on each position in the plate. Add the photo of the result. Justify the different mobility of each standard relating them to their structure. By comparing the result of samples to that of standards, explain which type of lipids each sample contains.