PROCESS FOR PREPARING A SERINE PROTEASE INHIBITOR FROM CELL CULTURES INDUCED FROM LINUM USITATISSIMUM

摘要

The present invention relates to a process for preparing a serine protease inhibitor from cell cultures induced from flax (Linum usitatissimum) by sterilizing the biological material formed of cell cultures by using explants as a source, after sterilization, the explants are placed into Petri dishes, on the surface of a Murashige-Skoog growth medium, and are incubated in the growth chamber. Periodically, at an interval of 3 weeks, there takes place the transfer to a fresh growth medium, in compliance with the aseptic technique measures with the view of preventing accidental contamination with microorganims. The obtained somatic explants are processed in order to isolate the protease inhibitor by delipidation with acetone, extraction with acetate buffer solution 0.1M, pH=5 and fractional precipitation with ethanol, there resulting a protein precipitate with a specific protease inhibitory activity of 3129 IU/mg of protein, which is purified in phosphate buffer 0.05M, pH=7.5, on affinity chromatography column, on immobilized trypsin equilibrated with phosphate buffer. The non-bonded proteins are washed with the same buffer and by elution with phosphate buffer pH =7.5, in a concentration gradient from 0 to 0.3 M, in equal volumes, there resulting a serine protease inhibitor with a specific activity of 9681 IU/mg of protein.