尼龙固定化木瓜蛋白酶及其应用研究

 尼龙经CaCl_2和H_2O的甲醇溶液处理,稀HCl水解用戊二醛交联以制备固定化木瓜蛋白酶。在溶液酶浓度为1mg/mL pH7.5—8.0、4—15℃条件下固定3h,活力回收42.5％,相对活力46％,偶联效率52％,半衰期72天。溶液酶Km值和固定化酶K_m~(aPP)值(底物酪蛋白W/V,％)分别为0.28％和0.35％。溶液酶和固定化酶分别在pH6.5和pH8.0以下活力稳定;最适pH分别为7.0和8.0;在65℃处理30min活力分别为原有活力的89％和66％。当酪蛋白浓度为1.5％和2.5％以上活力分别受到抑制。固定化酶在6mol/L脲中连续浸洗5次共6h其活力稳定,仍有原活力的44.4％;用以处理啤酒浊度比对照下降了2-11倍;蛋白质含量下降了55％;冷藏(4℃)120天,无冷混浊发生;同时各项理化指标和风味不变。

Study on Nylon Immobilized Papain and Its Application

Immobilized papain is prepared through the following procedure, nylon was first treated with CaCl2 solution containing methanol, and hydrolysed with dilute HC1 solution Finally papain was immobilized on the treated nylon with glutaraldehyde.Immobilization efficiency was best when the enzyme was at the concentration of 1 mg/ mL in solution immobilized for 3 hours, and pH7.5-8.0 at 4-15*******C.The rate of recovery of enzyme activity was 52 % with half life of 72 days.The Km value of the soluble enzyme and the apparent Km value of the immobilized enzyme (the substrate is casein, w/v %) were 0.28 % and 0.35 % respectively.The soluble enzyme was stable under pH6.5 and the immobilized enzyme under pH8.0 for 2 hours.Their optimal pH's were 7.0 and 8.0 respectively.When treated 30 min at 65*********C, the activity of the soluble enzyme was 89 % of the original activity and that of the immobilized enzyme was 66 % When the substrate (casein) concentration was above 1.5% and 2.0%, the activies of soluble enzyme and the immobilized enzyme were inhibited respectively.After the immobilized enzyme was eluted five times in 6 mol/L urea for about 6 hours, the activity of the enzyme tended to be stable, and retained 44.4 % of the original activity.When beer is treated with immobilized enzyme, its turbidity was reduced by 2-11 times in comparison with the control.The content of protein in treated beer was reduced 55 percent, after the treated beer was stored at 4********C for about 120 days, no chill-haze was observed,while the control had been apparently turbid.Physical and chemical indices were virtually unchanged in accord with the standards, and the flavor of the treated beer was comparable to the untreated one.