Chromatography Techniques

Question 1

Introduction to the chromatography techniques

Answer 1

▪ Chromatography is a widely used method that allows the separation, identification, and determination of the chemical compounds in complex mixtures.

▪ Chromatography is a physical method of separation in which the components to be separated are distributed between 2 phases, one of which is a stationary phase while the other is a mobile phase that moves in a definite direction.

▪ The mixtures are also separated based on the differences in the migration rates.

▪ The mobile phase may only involve a gas or liquid phase.

Question 2

Chromatography techniques (mechanism)

Answer 2

▪ In column chromatography, the stationary phase is held in a narrow tube, while the mobile phase is forced through the tube under pressure or gravity.

• The mobile phase moves through the stationary phase by capillary action or under the influence of gravity.

• The mobile phase moves over the stationary phase, carrying with it the analyte mixture.

• In this case, it may be in the form of gas, a liquid, or a supercritical fluid.

Question 3

Principles of Chromatographic Separations

Answer 3

• Chromatographic separation is based on the balanced state among the components to be separated , an adsorbent agent in the column (stationery phase), and a solvent flowing through it (mobile phase).

• When component settles on the stationery phase, this is defined as adsorption.

• In comparison, a detachment by mobile phase is defined as desorption.

• A high adsorption capacity between the components and stationary phase means there is a high retention of these components, and hence, resulting the elution to be slowed down.

• The separation is only possible if the individual components have different adsorption/desorption properties.

Question 4

CLASSIFICATION OF CHROMATOGRAPHIC TECHNIQUES

Answer 4

1. Adsorption chromatography- the stationary phase on which the sample components are adsorbed. The mobile phase may be liquid or gas. Eg: Thin layer chromatography.
2. Partition chromatography – the stationary phase is a liquid supported on an inert solid. The mobile phase may be liquid or gas. Eg: GLC.
3. Ion exchange – the separation is based on the ion exchange equilibria. The stationary phase is an ion exchange resin.
4. Size exclusion chromatography or pore penetration- solvated molecules are separated according to their size by their by their ability to penetrate a sieve-like structure (the stationary phase)

Question 5

Chromatographic Techniques

Answer 5

1) Thin layer chromatography

2) Column Chromatography (CC)

3) Gas Chromatography (GC)

4) High Performance Liquid Chromatography (HPLC)

Question 6

Gas Chromatography (GC)

Answer 6

▪ GC is widely used in determining organic compounds by separating the complex mixture.

▪ There are two types of GC: solid-solid (adsorption) chromatography and gas-liquid (partition) chromatography.

▪ In GC, the sample is converted a vapour state and the eluent is a gas (the carrier gas)

▪ The stationary phase is generally a non-volatile liquid supported on a capillary wall or in an inert solid particle such as silica.

▪ The most important factor in GC is the selection of proper (stationary phase) for the particular separation to be attempted

Question 7

Mechanism of Gas Chromatography (GC)

Answer 7

• A sample is injected into a heated port where it is vaporized and distributed between the stationary phase and the carrier gas.

• Since the equilibria in gas phase is rapid, the resolution of separation can be high. Subsequently, the analytes are automatically recorded as they emerge from the column makes measurements rapid and convenient.