1. Hands-on Chromatography
Sheryl K. Hoffmann
Concordia College
CONASTA 60
Wednesday then LUNCH

3. Definitions Definition 1:
Chromatography is a technique for separating similar compounds from one another by using differences in the strength of their adsorption on an inert material.
Adsorb: hold (molecules of a gas or liquid or solute) to its surface
Absorb: swallow up, incorporate, take in
Definition 2:
Chromatography is a technique which can be used to separate the components of a mixture. The method depends upon differences in the abilities of the components to adhere to a porus solid such as silica gel, called the stationary phase differences in the abilities of the components to be dissolved in a stream of liquid or gas, called the mobile phase, which is passed over the solid.

4. Main Types Paper Chromatography Thin Layer Chromatography (TLC)
Column Chromatography
High Performance Liquid Chromatography (HPLC)
Gas-Liquid Chromatography

5. Basic Method Basic Method for Paper Chromatography
Take a piece of chromatography paper and mark a pencil line 2 cm from the bottom.
Mark the positions of the samples evenly across the paper with a cross, leaving at least 2cm at each side.
Spot the samples on the positions marked, and allow to dry.
Pour < 1cm of solvent into a beaker, test tube or chromatography tank.
Lower the chromatography paper into the solvent in a beaker & support.
Allow the run until the solvent nearly reaches the top of the paper.
Mark final solvent height if Rf values are to be calculated.

6. Spotting Technique Spotting Methods
To get good separation of bands, it is important to keep the spot as small as possible & as concentrated as possible. Capillary Tubes, Toothpick & Pasteur Pipettes can all be used to spot chromatography paper

7. Tips
If the spot is large, components start from different origins and will thus blur any separation that would normally occur.
Try a quick dabbing motion, the longer the liquid is in contact with the paper the larger the spot
If the spot is not very concentrated the resulting bands, once spread over a wider area, will be even fainter.
If the solution to be spotted is not very strong, spot several times, being sure to allow the spot to dry in-between, a hair drier will speed up the drying process.

8. 2. Comparison of 1M NaCl & Water in Separating Food Colour
Hands On Activities
1. Practice Spotting
2. Comparison of 1M NaCl & Water in Separating Food Colour
3. Sticks of Chalk
4. Counterfeit Smarties?
5. Comparing Black Inks
Go through & explain each practical (because they never read the notes)
We will be using a plastic ice-cream container as a Chromatography tank
FIRST - everyone
practice spotting technique on a filter paper
look at the TLC plate & try spotting on it
NOTES:
4 activities to try – don’t all start at #1
At least try washing a Smartie with 2 drops water
Towards the end we will look at everyones’ results + Activity 5 & 6 + watch a short video

9. Activity 2 – Food Dyes LEFT RIGHT 1M NaCl vs water
(Blue & yellow) and (blue & red) separate more completely in 1M NaCl

10. Activity 3 - Chalk

11. Activity 4 - Smarties

12. Activity 5 – Black Pen Ink
Clearly the pen third from the right matches the ink sample on the right hand side

13. Activity 6 – Geranium TLC Paper LEFT RIGHT TLC vs Paper
Clear TLC gives better separation than paper
TLC Paper

14. Activity 6A – Frozen Spinach

15. Activity 6A –Spinach

16. Activity 7 – 2 Columns
Large column in a Quickfit separating funnel & a small column in a Pasteur pipette