CLASSIFICATION OF CHROMATOGRAPHY

Gas ChromatographyUsed to determine the chemical composition of unknown substances, such as the different compounds in gasoline shown by each separate peak in the graph below.

Liquid ChromatographyUsed to identify unknown plant pigments & other compounds.

Thin-Layer ChromatographyUses thin plastic or glass trays to identify the composition of pigments, chemicals, and other unknown substances.

Paper ChromatographyCan be used to separate the components of inks, dyes, plant compounds (chlorophyll), make-up, and many other substances

Represented To: Dr. IU khan

Represented By: Farzeen Javaid 0188-BH-CHEM-11

Govt. College University Lahore

DEFINITION

It is a physical separation method in which the components of a mixture are separated by differences in their distribution between two phases, one of which is stationary (stationary phase) while the other (mobile phase) moves through it in a definite direction .

BASIC PRINCIPLE

The samples are subjected to flow by mobile phase onto or through the stable stationary phase. The sample components are separated into fractions based on their relative affinity towards the two phases during their travel.

CLASSIFICATION

Different methods were attempted for classification of chromatography:Mechanism of separationBased on Physical means (Methods

holding the stationary phaseTypes of Mobile phase

1. ACCORDING TO MECHANISM OF SEPARATION:

MECHANISM OF SEPARATION

Partition chromatography

Adsorption chromatography

Size-exclusion chromatography

Affinity chromatography

Ion-exchange chromatography

a. Partition chromatography

PRINCIPLE:• Partition - based on the relative solubility of

analyte in mobile and stationary phases

• Normal – analyte is nonpolar organic; stationary phase MORE polar than the mobile phase

• Reverse – analyte is polar organic; stationary phase LESS polar than the mobile phase

b. Adsorption chromatography

PRINCIPLE:Separation is based mainly on differences between the adsorption affinities of the sample components for the surface of an active solid.

c. Size-exclusion chromatography

PRINCIPLE:Separation is based mainly on exclusion effects, such as differences in molecular size and/or shape or in charge.

• The term Size-Exclusion Chromatography may also be used when separation is based on molecular size.

• The terms Gel Filtration and Gel-Permeation Chromatography (GPC) were used earlier to describe this process when the stationary phase is a swollen gel.

• The term Ion-Exclusion Chromatography is specifically used for the separation of ions in an aqueous phase.

d. Affinity chromatography

PRINCIPLE:The stationary phase is typically a gel matrix (often agarose) . The molecule of interest has a known and defined property. The process is an entrapment in which the target molecule becomes trapped on stationary phase. The Stationary phase can then be removed from the mixture, washed and then target molecule is released from the entrapment.

e. Ion-exchange chromatography

PRINCIPLE:

Ion - exchange chromatography retains sample

molecules on the column based on ionic

interactions. The surface of stationary phase displays

ionic functional groups (R-X) that interact with

analyte ions of opposite charge.

2. BASED ON PHYSICAL MEANS (METHODS HOLDING THE

STATIONARY PHASE

Stationary phase

Two dimensional chromatography

Paper chromatography

Thin layer chromatography

Three dimensional

chromatography

Column chromatography

a. Two dimensional chromatography (planar or plane chromatography)

Planar or Plane Chromatography:In this type of chromatography the stationary phase is used in the form of layer. Plane chromatography is further classified into:l. Paper Chromatography (PC):A specific type of papers is used as stationary phase in the form of sheets.ll. Thin Layer Chromatography (TLC): The stationary phase in the form of fine powder is spread on glass or plastic or aluminum sheets.

l. Paper Chromatography (PC): A method of partition chromatography using filter paper strips as carrier or inert support.

The factor affecting separation of mixtures of solutes on filter paper is the partition between two immiscible phases.

• One is ( stationary phase) usually water adsorbed on cellulose fibers in the paper.

• The second is ( mobile phase) the organic solvent flows past the sample on the paper (stationary phase).

Principle:

The certain solvent are used to separate a mixture e.g: water, alcohol. With capillary action the solvent will move up to filter paper. Movement of a solvent will bring together component that are separated from the mixture. Every component that are separated will move to several velocity

ll. Thin layer chromatography (TLC)

TLC separates dried liquid samples with a liquid solvent (mobile phase) and a glass plate covered with a thin layer of alumina or silica gel (stationary phase)

b. Three dimensional chromatography (Columnar or Column Chromatography):

The stationary phase is held in to a tube made of glass or metal.The mixture to be analyzed is applied to the top of the column. The liquid solvent (the eluent) is passed through the column by gravity or by the application of air pressure.

3. TYPES TO MOBILE PHASE

MOBILE PHASE

Liquid chromatography

Gas chromatography

Supercritical fluid chromatography (SFC)

1.Liquid Chromatography

Four Basic liquid chromatography modes are:

1.Liquid/Solid Chromatography (adsorption chromatography)

A. Normal Phase LSC

B. Reverse Phase LSC

2.Liquid/Liquid Chromatography (partition chromatography)

A. Normal Phase LLC

B. Reverse Phase LLC

3.Ion Exchange Chromatography

4.Gel Permeation Chromatography (exclusion chromatography)

A. Liquid-solid Chromatography (Adsorption chromatography )

Stationary phase: solid with adsorption power (silica, alumina). Mobile phase: Non-polar solvent (or a mixture)

Mechanism :Separation is based mainly on differences between the adsorption affinities of the sample components for the surface of an active solid.

B. Liquid-liquid Chromatography (Partition chromatography)

• Mechanism:Solutes are separated based on their partition between a liquid mobile phase and a liquid stationary phase

I. Normal-phase chromatography

• Stationary phase Polar• Mobile phase non-polar

Silanol radicals ( -Si-OH ) on the surface of silica gel act as the active site and the sample is separated.

ll. Reversed- phase chromatography

• Stationary phase non-polar• Mobile phase Polar

Packing materials are hydrocarbons having 18 carbon atoms (called the

Octadecyl radical) which are chemically bonded to silica

gel.

2. Gas chromatography

Two types of gas chromatography are:

• Gas-solid chromatography (GSC)Mobile phase: Gas Stationary phase: Solid

• Gas-liquid chromatography (GLC)Mobile phase: Gas Stationary phase: Liquid

3.Supercritical fluid chromatography (SFC)

Advantages:

1. Fast: rate of diffusion between sample matrix & extraction fluid 10-60 min vs. days.

2. Solvent strength can be varied by changes in P & T.3. Less Harmful solvent.4. 4. Many SCF are gases at RT, recovery of analytes is

easy.5. 5. Many SCF are cheap, inert, and non-toxic.6. 6. On-line extraction.

REFERENCES:

• https://satyapsingh.files.wordpress.com/2012/09/chromatography.pdf

• http://s3.amazonaws.com/ppt-download/chromatographyre-edited-111209152137-phpapp01

• http://s3.amazonaws.com/ppt-download/principlesandapplicationofchromatography-130121011302

• http://s3.amazonaws.com/ppt-download/chromotography-140516031516

• http://www.separationprocesses.com/Adsorption/AD_Figures/CG_Fig003.gif

• https://mnh20.files.wordpress.com/2010/04/chromatography.ppt