The principle of gel chromatography is that the stationary phase of gel chromatography is porous gel substance, and the mobile phase is aqueous solution or organic solvent, which is separated according to the size of different molecular volumes. Small molecules can diffuse into the gel gap and pass through it, with the slowest peak; Medium-sized molecules can only pass through part of the gel gap and pass at a medium speed, while macromolecules are excluded and peak at the fastest; Solvent molecules are small, so there are peaks at the end, and there are peaks before the dead volume; Compounds with relative molecular mass between 100 and 105 can be separated by mass.

Gel chromatography, also called molecular exclusion chromatography, is based on the principle of molecular sieve, and is often used to separate organic macromolecules, such as protein, peptides and polysaccharides. It has a high separation effect on polymer materials. Macromolecules are difficult to enter the micropores of gel particles because of their large diameter, and can only be distributed between particles, so the downward movement speed is faster when eluting.

Small molecular substances can not only diffuse in the gaps between gel particles, but also enter the micropores of gel particles, that is, into the gel phase. During the downward movement, they diffuse from one gel to the gaps between particles, and then enter another gel particle. So they continue to enter and diffuse, and the downward movement speed of small molecular substances lags behind that of large molecular substances, so that the large molecules in the sample flow out of the chromatographic column first, then the medium molecules flow out, and finally the smallest molecules flow out.

Molecular sieve effect: when the sample solution containing various molecules slowly flows through the gel column, each molecule moves in two different ways at the same time, namely vertical downward movement and non-directional diffusion movement. Macromolecules are not easy to enter the micropores of gel particles because of their large diameter, and can only be distributed between particles, so they move down quickly when eluting.

Small molecular substances can not only diffuse in the gaps between gel particles, but also enter the micropores of gel particles, that is, into the gel phase. During the downward movement, they diffuse from one gel to the gaps between particles, and then enter another gel particle. So they continue to enter and diffuse, and the downward movement speed of small molecular substances lags behind that of large molecular substances, so that the large molecules in the sample flow out of the chromatographic column first, then the medium molecules flow out, and finally the smallest molecules flow out. This phenomenon is called molecular sieve effect.