| 摘要 |
PURPOSE:To enable mass production of a novel heparitinase to act on heparitinase (HS) and heparin (Hep) to produce an oligosaccharide, by activating established Mphi with LPS. CONSTITUTION:Mphi (P388D1) is cultivated in medium RPMI164O containing 10% bovine fetal serum, transferred to a serum-free medium and cultivated under the same condition. LPS in an amount to make about 20mug/ml final concentration is added to the medium and further Mphi (P388D1) is cultivated under the same condition for 18 hours. Then a treated supernatant liquid is successively subjected to heparin Sepharose column and concanavalin A Sepharose column and heparitinase is eluted with a buffer solution containing acetylglucosamine. Then the prepared purified heparitinase acts on heparan sulfate (HS) and heparin (Hep) to produce an oligosaccharide, has optimum pH 7.0-7.5, 37-40 deg.C optimum temperature, about 98,000 molecular weight, stable pH 6-7 and <=40 deg.C stable temperature. |
