METHOD OF PREPARING THE HIGH-PURIFIED MONOCOMPONENT INSULIN Russian patent published in 1998 - IPC

FIELD: medicine, biotechnology. SUBSTANCE: insulin-sodium containing 85% monofraction, not less, is fed on chromatography column filled with sulfocationite-exchange resin КУ-23И (particle size is 200-300 mcm). Insulin is eluted with 0.018-0.022 M phosphate linear buffer and insulin is isolated by isoelectric precipitation. Insulin is dissolved in 1 M acetic acid and fed on chromatography column filled with Sephadex G-50 Sf (Pharmacia firm) and insulin is eluted with 1 M acetic acid. Part of insulin-containing eluate that is eluted at the range from 0.6-0.7 to 0.9-1.0 packing volume of column is fed to column inlet for recirculation and elution is continued with 1 M acetic acid. Monopeak insulin obtained after this circulatory gel-chromatography stage is fed on column filled with anione-exchange silica gel PAE-300 (Amicon firm; particle size is from 35 to 70 mcm). Insulin is eluted at the rate from 20 to 45 ml/cm²×h^-1, using exponential gradient of sodium chloride concentration. The obtained insulin has 98.5% monofraction (not less), 0.5% deamidoinsulin (not above), 1.0% nonidentified impurities (by HPLC method) and 10 p.p.m. proinsulin and its derivatives (not above). The yield of product by mass is from 55% to 65% of initial insulin-sodium. EFFECT: improved method of insulin preparing, increased yield. 2 ex, 2 dwg