HPLC analysis is an interesting technique which uses stationary and solid phase with a mobile or liquid phase. A plastic, glass or even metal tube, called a column, is used to house the stationary phase, usually silica beads or a kind of porous solid laminated with specific compounds at different levels to attract the items needing separation. As the reactive solvent containing the compound moves through the column, separate molecules and even atoms can attach themselves to the stationary phase, and a separation of the compound into different parts occurs.
We have used many different techniques for a number of applications and one included the synthesis of particular proteins.
This all begins with the purification of amino acids and this can be fulfilled by using HPLC columns to separate the individual amino acids from different proteins. The stationary phase of the column separates the amino acids individually as the proteins pass through the column. Solvents are used to liquify and homogenize the proteins, and the solvent becomes a carrier, known as the mobile phase. The proteins are separated in an efficienct way as it is pumped through the system.
Pore size and particle size in the stationary phase of an high performance liquid chromatography column is very important to the speed at which the phase can separate the compound travelling through it. Silica is a common substance used in reverse-phase chromatography, and this can be used in a HPLC chromatography system. Silica is not the only substance, and great care should be taken when using certain acidic solvents. Temperature is also important, as silica can be damaged by excessive heat. The electrical make up, as well as pore size found on the beads are the main benefits of silica.