CHROMATOGRAPHY

A process to separate molecules to purify them.

There are many kinds of chromatography; Column Paper Thin Layer Gas High Performance Liquid

We will concentrate on Column Chromatography

There are many kinds of column chromatography; size exclusion ion-exchange affinity

For this project we will use ion-exchange

Prepare the column

Run the column

Above pictures are from: emistry/courses/CC/index.htm

When running the column, the target molecule will stick to the charged beads. It must be eluted out with increasing ionic strength buffers.

In this demonstration we will separate the pigments of; Grape Kool-Aid. A procedure is as follows.

1. Make up solvent solutions 2. Wash Sep-Pak cartridges in 5 ml of 70% alcohol followed with 5 ml of water using a syringe which easily attaches to the column.

3. Add 2 ml of Kool-Aid contained in a syringe to the column. 4. Rinse the column with 5 ml of water as above.

5. Elute the firsts dye into a test tube with 14% isopropyl alcohol 6. Elute second dye into a test tube with 35% isopropyl alcohol. 7. Finally, rinse the column with 70% isopropyl alcohol.