Papain (1 g; obtained from J. E. Siebel Son's Co.) was well suspended in 2 ml of 3.0M potassium citrate buffer (pH 5.1), the suspension was then mixed with 2 ml of glycerol. To 100 ml of ethyl acetate, 80 mmole of L-phenylalanine methyl ester hydrochloride and 80 mmole of triethylamine were added and stirred at 25° C. for 40 min then 40 mmole of N-benzyloxycarbonyl-L-aspartic acid was added and stirred for 20 min. The whole ethyl acetate mixture, containing soluble L-phenylalanine methyl ester and N-benzyloxycarbonyl-L-aspartic acid and insoluble triethylamine hydrochloride, w-as subsequently added to the papain suspension. The reaction mixture was stirred at 25° C. for 22 hr then 200 ml of ethyl acetate was added and stirred at 55° C. for 15 min and filtered. As determined by HPLC analysis the yield of the N-benzyloxycarbonyl-α-L-aspartyl-L-phenylalanine methyl ester in the filtrate was 85% calculated on the basis of the initial amount of the N-benzyloxycarbonyl-L-aspartic acid. The ethyl acetate solution was then extracted twice with 150 ml of 0.01M phosphate buffer (pH 7.0), then twice with 150 ml of 0.3N hydrochloric acid. The final organic layer was dehydrated with 10 g of anhydrous Na2SO4, then evaporated and vacuum dried to give 17.4 partially purified N-benzyloxycarbonyl-α-L-aspartyl-L-phenylalanine methyl ester.
