EFFECTS OF SORBITOL ADDITION ON THE ACTION OF FREE AND IMMOBILIZED HYDROLYTIC ENZYMES IN ORGANIC MEDIA

The effect of the addition of sorbitol on the activity and stability o f enzymes was examined by monitoring transesterification reactions per formed in organic media at various water activities (a(w) = 0.08 to 0. 97). Lipases from Chromobacterium viscosum and Candida rugosa immobili zed on celite, and chymotrypsin, free or immobilized on celite, were u sed. When the sorbitol-containing enzymes were employed, higher reacti on rates and less hydrolysis were observed. Immobilization of chymotry psin resulted in high activity and operational stability, while the no nimmobilized enzyme was stable only in the presence of sorbitol. The a ctivity of all preparations diminished after washing them with pyridin e to remove sorbitol. Furthermore, severe stability problems occurred in the preparations lacking sorbitol. Sorbitol treatment, even after r emoval of the sorbitol itself, improved the activity of nonimmobilized chymotrypsin relative to the washed control. On the other hand, washi ng to remove sorbitol had a negative effect on the activity of both co immobilized lipase and coimmobilized chymotrypsin. Addition of a subst rate analogue, N-acetyl-L-phenylalanine, to chymotrypsin yielded a pre paration that exhibited higher activity than both the control and its sorbitol-containing counterpart. Differential scanning calorimetry mea surements revealed that the chymotrypsin-sorbitol complex was stable a gainst thermal denaturation, undergoing transition at a high temperatu re (89 degrees C). The transition temperatures of the substrate-contai ning chymotrypsin and of the control were identical (72 degrees C). (C ) 1995 John Wiley and Sons, Inc.