what is hplc

Basic Principle of Thin Layer Chromatography

In Thin layer chromatography (TLC) the analytes or sample mixture is applied as a minute dot or spot at the bottom of the stationary phase it holds up on a glass, metal or plastic plate. A solvent of the sample is allowed to evaporate from the plate, which is after that, positioned in a closed chamber, in which at the bottom there is a shallow pool of the mobile phase. Mobile phase goes through capillary forces through the stationary phase.

Components of the analyte mixture travel at the different rates throughout the movement through the stationary phase of the mobile phase While the mobile phase has taken a suitable distance, from the chamber plate is removed and the marked the solvent front and through drying at room temperature the mobile phase is evaporated from the plate or by placing an oven. If the mixture components are not colored or fluorescent naturally, an identification reagent is applied to see the bands.

To ensure, the use of one or more identification techniques to detect all the components in the mixture is done. Rf value is a suitable method of expressing the position of an analyte on a establish chromatogram and it is calculated as, Rf ¼ Distance of the analyte from the origin/solvent distance from the origin.Different kinds of solvents are used as stationary phases in TLC; consist of silica gel, chemically modified silica gel, ion exchangers, polyamides, alumina, and cellulose.

Now, pre-coated high-performance TLC plates with fine particles are generally used in the pharmaceutical industry for reproducible, fast, and efficient separations. The options for the mobile phase are from a solitary component solvent system of many component solvent systems, and the multiple component solvent systems are the most common ones. The movement of every analyte in the mixture of mobile phase throughout TLC is the result of multi opposite forces, for example, the mobile phase capillary action and the stationary phase retardation action.

Both forces give towards achieving the migration of each analyte.


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