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A continuous two-stage membrane (1st-SCMR, MWCO 10,000; 2nd-SCMR, MWCO 5,000) reactor was developed and optimized for the production of fish skin gelatin hydrolysate with different molecular size distribution profiles using trypsin and pronase E. The optimum operating conditions in the 1st-step membrane reactor using trypsin were: temperature, 55℃ ; pH 9.0; enzyme concentration, 0.1 ㎎/㎖; flux, 6.14 ㎖/min; reaction volume, 600 ㎖; and the ratio of substrate to trypsin, 100 (w/w). After operating for 1 hr under the above conditions, 79% of total amount of initial gelatin was hydrolysed. In the 2nd-step using pronase E under optimum operating conditions[temperature, 50℃ ; pH 8.0; enzyme concentration, 0.3 ㎎/㎖; flux, 6.14 ㎖/min; reaction volume, 600 ㎖; and the ratio of substrate to pronase E, 33 (w/w)], the 1st-step hydrolysate was hydrolysed above 80%. Total enzyme leakages in the 1st-step and 2nd-step membrane reactors were about 11.5% at 55℃ for 5 hrs and 9.0% at 50℃ for 4 hrs, respectively. However, there was no apparent correlation between enzyme leakage and substrate hydrolysis. The membrane has a significant effect on activity lose of trypsin and pronase E activity for 1 hr of the membrane reactors operation. The loss of initial activity of enzymes were 34% and 18% in the 1st-step and 2nd-step membrane reactor, whereas were 23% and 10% after operating time 3 hr in the 1st-step and 2nd-step membrane reactor lacking the membrane, respectively. The productivities of 1st-step and 2nd-step membrane reactor for 8 times of volume replacement were 334 ㎎ and 250 ㎎ per ㎎ enzyme, respectively.
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