GEL-FILTERING (synonym gel chromatography) — a method of unmixing of substances with different molecular weights by its filtering through various cross and connected cellular gels.
- f. the pier is widely applied in various fields of chemistry, biochemistry, medicine, etc. to the analysis of mix of connections with various. scales. The method is simple on technology of performance, is carried out in soft conditions that allows to use it for work with very unstable connections, napr, enzymes. - f. the pier is used for determination of sizes. scales, concoction of solutions of polymers, studying some physical. - chemical properties of macromolecules, etc.
In G.'s medicine - f. serves as supportive application in diagnosis. So. e.g., fractionation of proteins of a blood plasma on G-200 sephadex is used for bystry diagnosis of the diseases which are followed by sharp disturbances of composition of proteins of plasma. By means of G. - f. on G-200 sephadex Valdenstrem's (macroglobulinemia) disease from a plasmacytoma is possible to otdifferentsirovat serums that cannot be made electrophoretically. - f. it is used for early diagnosis of pregnancy, for definition of qualitative composition of proteins in microamounts of cerebrospinal liquid etc.
Thanks to a set of modifications of G. - f. it is applied to works at the microlevel and in the preparative purposes.
For G. - f. use preferential hromatografichesky columns (see. Chromatography ). As a filler serves the granulated gel (see. Gels ); for a research of mix of the connections dissolved in water use hydrophilic gels (gels of sefadeks — dextrans, biogels, etc.); fractionation of hydrophobic polymers is carried out by means of filtering through lipophilic gels.
The column with the gel which bulked up in the corresponding solvent has two appearance of volumes of this solvent: internal and outside. Passing of substances with various a pier. is defined scales by pore sizes between granules of gel by a column. Substances which molecule it is much less than pore sizes of granules, will freely diffuse in these granules. Therefore large molecules will move on a column with greater speed, than small.
The distribution coefficient (Kr) of solute between internal and external volumes is defined from the equation of Kp = (Ve-Vn)/Vv where Ve — the volume of liquid following from a column from the moment of drawing on it the studied solution before emergence of fraction with the maximum concentration of the substance interesting the experimenter; Vv — the volume of solvent which is in granules of gel, a Vn — the volume of solvent out of granules of gel.
If Kr is equal 0, then substance is not capable to get in granules. At Kr = 1 substance is distributed evenly between external and internal volumes.
At a transmission through a column of the eluating solution first of all the substances which are in external volume begin to move. Thus, components of the studied mix are eluated from a column as reduction a pier. weight.
For elution (see) at G. - f. usually use the diluted saline or buffered solutions since at elution by pure water a number of connections is occluded by a matrix of gel. Also some proteins interact with a matrix (RNK-aza, a lysozyme, trypsin, seralbumin, etc.)
Efficiency of division at G. - f. does not depend on concentration of substances in solution. However, if viscosity of the studied solution is very high, then quality of division worsens.
For desalting or change of buffered solution the column is filled with the gel which bulked up in new buffered solution then put the studied test. Protein (or other high-molecular compound) is eluated in new buffered solution after an effluence of volume of Vn. The initial buffered solution and other ions which are contained in it appear after passing of the volume equal to Vn + Ve.
Definition pier. weight by method G. - f. it is based that nearly a volume of an exit of substance from a column is linear function of a logarithm of size a pier. weight. Before carrying out definition a pier. the weight of unknown polymer, it is necessary to calibrate a column by means of polymers with known a pier. weight (standards). Knowing the volume of an exit of the studied substance, determine required size by a standard curve a pier. weight.
Definition pier. the weight of enzymes it is possible to make in the crude drugs and in strongly divorced solutions; - f. allows to investigate also reversibly the dissociating systems which studying by means of other methods is rather labor-consuming.
See also Gels .
Bibliography: Determan G. Gel chromatography, the lane with it., M., 1970; T and-honenko T. I. Methodological fundamentals of biochemistry of viruses, M., 1973, bibliogr.