Chromatography is a very useful technique that is extensively used across various scientific disciplines like chemistry, biology, medicine, forensic science and many more.
What is Chromatography?
Chromatography refers to a set of laboratory techniques that are used to separate the components or solutes of a mixture. It relies on differential partitioning between a mobile fluid phase and a stationary phase. The components of the mixture that needs to be separated are dissolved in a mobile fluid phase which is then passed over a stationary phase. The components interact differently with the stationary phase based on their chemical properties which results in their separation.
Fundamentals of Chromatography
There are a few key components that make chromatography work:
- Mobile Phase: This refers to the fluid component that carries the sample mixture through the chromatography system. It can be a gas or liquid depending on the type of chromatography used.
- Stationary Phase: This component remains stationary in the system and interacts with the components in the mobile phase leading to separation. It can be a solid (for column chromatography) or liquid (for paper chromatography).
- Partition Coefficient: Chromatography refers to the affinity of each component for the mobile versus stationary phase. Components with high affinity for the stationary phase move slower through the column.
- Separation: Differences in the partition coefficients of various components results in their separation as they travel through the chromatography system at different rates.
Types of Chromatography
There are various chromatography techniques based on the type of mobile and stationary phases used:
Gas Chromatography (GC)
- Mobile Phase: Inert gas like helium or nitrogen
- Stationary Phase: Non-polar liquid coated on an inert support inside a column
- Used for separation of volatile components
Liquid Chromatography (LC)
- High Performance Liquid Chromatography (HPLC): Used for separation of both volatile and non-volatile components. Stationary phase includes silica or polymer beads inside a column.
- Ion-Exchange Chromatography: Separation based on ionic interactions.
- Size-Exclusion Chromatography: Separation based on molecular size.
Thin Layer Chromatography (TLC)
- Mobile Phase: Non-polar solvent
- Stationary Phase: Thin coating of adsorbent like silica on a plate
- Used for separation of minor components in complex mixtures
Paper Chromatography
- Mobile Phase: Polar (like water) or non-polar (like acetone) solvent
- Stationary Phase: Cellulose fibers of filter paper
- Used as a simple and inexpensive technique
Applications of Chromatography
Some key applications of chromatography include:
Biochemistry & Proteomics
- Separate and analyze amino acids, proteins, enzymes etc.
Environmental Analysis
- Identify pollutants and other compounds present in air, water and soil samples.
Forensic Science
- Fingerprint drugs, poisons, explosives and other compounds from crime scene samples.
Food Science & Quality Control
- Purity testing of food samples, identification of artificial flavors and preservatives.
Pharmaceutical Industry
- Analyze drug compounds, check purity of formulations and stability of drugs.
Metabolomics & Biomarker Discovery
- Separate and profile complex mixtures of metabolites in biological fluids and tissues.
Petrochemical Industry
- Separate hydrocarbon mixtures from crude oil for refining purposes.
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