Application of Chromatography in the isolation, purification and identification

• Explore the Application of Chromatography in the isolation, purification and identification for accurate compound detection and purity analysis.
• Application of Chromatography in the isolation, purification and identification enables separation of complex plant mixtures with precision.
• Chromatography is essential for separating, purifying, and identifying bioactive compounds from crude plant or microbial extracts.
• These techniques exploit differences in polarity, size, and other physicochemical properties.

1. Thin-Layer Chromatography (TLC)

   • Principle:

     • Separates compounds based on migration rates on a coated plate (silica, alumina).

   • Application:

     • Rapid, cost-effective screening of multiple components in crude extracts.
     • Rf value comparison with known standards.

2. Column Chromatography

   • Principle:

     • Uses a stationary phase (e.g., silica gel) and a mobile phase (solvent) for separation.

   • Application:

     • Scalable fractionation of plant extracts.
     • Pre-purification step before advanced techniques.

3. High-Performance Liquid Chromatography (HPLC)

   • Principle:

     • High-pressure pumps drive solvents through a column with small particles for high-resolution separation.

   • Application:

     • Quantitative analysis and quality control of phytochemicals.
     • Coupled with UV, fluorescence, or MS for improved identification (HPLC-UV, HPLC-FLD, LC-MS).
     • Standardization of herbal extracts in pharmaceuticals.

4. Gas Chromatography (GC)

   • Principle:

     • Separates volatile compounds based on boiling points and stationary phase interactions.

   • Application:

     • Analysis of essential oils and volatile compounds.
     • Commonly paired with MS (GC-MS) for compound identification.

5. Supercritical Fluid Chromatography (SFC)

   • Principle:

     • Uses supercritical fluids (e.g., CO₂) as the mobile phase.

   • Application:

     • Suitable for non-polar to moderately polar compounds.
     • Eco-friendly alternative to organic solvents.

6. Preparative Chromatography

   • Principle:

     • Scaled-up column or HPLC for bulk purification.

   • Application:

     • Produces milligram to gram quantities of pure compounds for pharmacological testing.

• These chromatographic methods complement spectroscopic techniques, ensuring comprehensive compound identification and purification.

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