A - Chromatography

Question 1

What can chromatography be used for?

Answer 1

To separate and identify the components in a mixture.

Question 2

There are different types of chromatography but they all use the same basic principle. Explain this.

Answer 2

1. The mobile phase moves through or over the stationary phase.
2. The distance each substance moves up the plate depends on its solubility in the mobile phase and its retention in the stationary phase.
3. Components that are more soluble in the mobile phase will travel further up the plate.
4. The differences in solubility and retention by the stationary phase separate out the different substances.

Question 3

What is the mobile phase?

Answer 3

Where the molecules can move. This is always a liquid or gas.

Question 4

What is the stationary phase?

Answer 4

Where the molecules can’t move. This much be a solid, or a liquid on a solid support.

Question 5

What does separation depend on?

Answer 5

The balance between solubility in the moving phase and retention by the stationary phase.

Question 6

Very briefly explain what thin-layer chromatography involves.

Answer 6

A plate is coated with a solid and a solvent moves up the plate.

Question 7

What is the stationary phase in thin-layer chromatography?

Answer 7

A thin layer of silica (silicon dioxide) or alumina (aluminium oxide) fixed to a glass or metal plate.

Question 8

Outline the method for carrying out thin-layer chromatography.

Answer 8

1. Draw a line in pencil near the bottom of the plate (the baseline) and put a very small drop of each mixture to be separated on the line.
2. Allow spots on the plate to dry.
3. Place the plate in a beaker with a small volume of solvent (the mobile phase). The solvent level must be below the baseline so it doesn’t dissolve the samples.
4. The solvent starts to move up the plate. As it moves, it carries the substances in the mixture with it (some move faster and therefore further up the plate).
5. Leave the beaker until the solvent had moved almost to the top of the plate. Then remove the plate from the beaker. Before it evaporates, use a pencil to mark how far the solvent travelled up the plate (this line is called the solvent front).
6. Place the plate in a fume cupboard and leave it to dry.
7. The result is the chromatogram where you can use the position of the spots on this to identify the chemicals.

Question 9

Why should you put the TLC plate into a fume cupboard when leaving it to dry?

Answer 9

It will prevent any toxic or flammable fumes from escaping into the room.

Question 10

What methods can you use to make colourless spots on chromatograms visible?

Answer 10

1. Many TLC plates have a special fluorescent dye added to the silica or alumina later that glows under UV light. You can place it under a UV lamp and draw around the patches to show where the spots of chemical are.
2. Expose the chromatogram to iodine vapour (leaving the plate in a sealed jar with a couple of iodine crystals). Iodine vapour is a locating agent - it sticks to chemicals on the plate and they’ll show up as brown/purple spots.

Question 11

How do you work out how many chemicals are present in a mixture using a chromatogram?

Answer 11

Count the number of spots that form on the plate.

Question 12

How do you identify what each spot is on a chromatogram?

Answer 12

By calculating the Rf value and looking this up in a table of standard Rf values.

Question 13

What is the formula for Rf? How do you measure each part?

Answer 13

Distance travelled by the spot divided by the distance travelled by the solvent.

To measure distance travelled by spot - measure from the baseline to the middle of the spot.

To measure distance travelled by solvent - measure from the baseline to the solvent front.

Question 14

What can alter Rf values?

Answer 14

A different composition of the TLC plate, a different solvent or different temperature.

Question 15

Very briefly explain what column chromatography is.

Answer 15

A column is packed with a solid and a solvent moves down the column.

Question 16

What is column chromatography mostly used for?

Answer 16

Purifying an organic product (this is done to separate the product from un reacted chemicals and by-products.

Question 17

Outline the method of column chromatography, including stating what the mobile and stationary phases are.

Answer 17

1. A glass column is packed with a slurry of absorbent material such as Al2O3, coated with water. This is the stationary phase.
2. The mixture to be separated is added to the top of the column and allowed to drain down into the slurry. A solvent is then run slowly and continually through the column. This solvent is the mobile phase.
3. As the mixture is washed through the column, it’s components separate out according to how soluble they are in the mobile phase and how strongly they are absorbed onto the stationary phase.

Question 18

Very briefly explain what is involved in gas chromatography.

Answer 18

A column is packed with a solid or with a solid coated by a liquid, and a gas is passed through the column under pressure at high temperature.

Question 19

When is gas chromatography used?

Answer 19

If you have a mixture of volatile liquids (ones that turn into gases easily) then GC is a good method of separating them for identification.

Question 20

What is the stationary phase in GC?

Answer 20

A solid or a solid coated by a vis our liquid, such as an oil, packed into a long tube. The tube is coiled to save space and built into an oven.

Question 21

What is the mobile phase in GC?

Answer 21

An unreactive carrier gas such as nitrogen.

Question 22

What is the retention time in GC and what does it depend on?

Answer 22

The time taken from when the component is injected into the tube, to being recorded at the other end.

It depends on how much time the component spends moving along with the carrier Gad, and how much time it spends stuck to the viscous liquid.

Question 23

How can you identify the components of a mixture using GC?

Answer 23

Using their unique retention times which is shown on the chromatogram. The area under each peak on the chromatogram tells you the relative amount of each component that’s present in the mixture.

Question 24

What is gas chromatography-mass spectrometry? What are the advantages of using this method?

Answer 24

Combines the two methods so the sample is separated using GC and then fed into a MS to produce a mass spectrum for each component, which can be used to identify each component.

The advantages are that the components can be positively identified, which is impossible from a chromatogram alone.

Computers can also be used to match up the mass spectrum for each component of the mixture against a database, so the whole process can be automated.