High-performance liquid chromatography (HPLC)

High-performance liquid chromatography (HPLC) formerly referred to as high-pressure liquid chromatography is used to separate the chemical components in a mixture, to identify and quantify each component of the mixture based upon the retention time. HPLC contains pumps, temperature module, injection loop, sampler, column, photo diode array detector and fraction collector. The pumps are used to pass a pressurized solvent (water, acetonitrile and/or methanol) containing the sample mixture and runs through a column filled with a solid adsorbent material (granular material made of silica, polymers, etc.).  Each component in the sample interacts differently with the adsorbent material, causing different retention time for the different components and leading to the separation of the chemical components as they flow out the column and detected by a detector. The interactions observed between the component and adsorbent material may be hydrophobic, dipole–dipole and ionic, most often a combination. The information from the detector is sent to the computer; each component will be observed as a peak with retention time and can be collected by fraction collector for further use. By comparing the retention time with reference we can identify the specific chemical compound. The various applications of HPLC include medical, legal, research and manufacturing purposes.