Paper Chromatographic Separation Class Notes

Chromatography comes from Latin for writing in color. Chromatography is the method used to separate molecules or ions based upon their structure and/or composition and their interactions with the mobile and stationary phases.

All forms of chromatography involves moving a preparation of the materials to be separated, called the "test preparation" over a stationary phase. The molecules or ions in the test preparation will have different interactions with the stationary phase leading to a separation.

Molecules or ions with a strong interaction with the stationary phase will tend to move more slowly through the stationary phase than those molecules or ions with weaker interactions. In this way, different types of molecules or ions can be separated from each other as they move at differing rates over the stationary phase.

How Is Paper Chromatography Used?

Paper Chromatography can separate minute amounts of substances. This makes it very useful for forensics investigators. Drugs from narcotics to aspirin can be identified in urine and blood by using chromatography. Detectives can analyse a sample of paint to find out the car it came from, including the model and year it was made. Ink can be separated into its components to help identify the pen used for ransom notes and forgeries, as well as the type of ink used for counterfeiting.

Terms To Know

The mobile phase in paper chromatography is the liquid mixture called the developing solution.

The stationary phase is the paper with a thin layer of adsorbed water. The water is not moving over the paper with the developing solution.

The developing solution moves up the paper by capillary action. The leading edge of the developing solution is called the solvent front. The mobile phase moves up past the spot of ion and carries it along the paper at a rate determined by the interaction of the ion with the paper. This flow process is called the development of the chromatogram. The chromatogram is the paper with the separated spots of ions or molecules.

Retention Factor, R_f

A spot of pure substance may move at the same rate as the solvent front (no affinity for the stationary phase), at a slower rate than the solvent front (some affinity for both the stationary and mobile phase), or the spot may not move at all (no affinity for the mobile phase).

The retention factor can be calculated by measuring the distance the spot traveled and the distance the solvent traveled.

Sample Calculation of R_f

The distance a spot moves is measured as 3.05 cm, and the distance the solvent front travels is measured as 5.33 cm.

R_f = 3.05 cm / 5.33 cm

R_f = 0.572

The units cancel out and there are no units for R_f.

R_f of mixtures

When you have a mixture of ions or molecules in your test unknown, there will be some interaction between the ions or molecules. This effects the calculated R_f for each spot of the mixture so that it will not be exactly the same as the R_f of the pure ion or molecule.

Calculation of Separation of Spots on the Chromatogram

If one spot has an R_f value of .732 and the second spot has an R_f value of .348. The solvent front moves 8.61 cm. Calculate the separation between the spots.

First spot:

x-y =6.30 cm - 3.00 cm = 3.30 cm

Colors You Will See

Once the chromatogram has been developed, Cu²⁺ should turn light blue, Fe³⁺ should be yellowish brown, and Ni²⁺ will be faint green, but it is so light that you probably will not be able to see it.

After the chromatogram is exposed to NH₃ the Cu²⁺ will turn a deep blue, Fe³⁺ turns a rust brown, and Ni²⁺ is not affected.

Exposure to dimethylglyoxime, DMG, turns the Cu²⁺ a black color, Fe³⁺ turns brown, and Ni²⁺ turns a strawberry red.

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last updated: October 1, 2011