The use of a poly(allyl carbonate) for the preparation of active, water-insoluble derivatives of enzymes

Abstract

The optimum pH conditions have been determined for the covalent binding of the soluble enzymes α-amylase (E.C.3.2.1.1), β-D-glucosidase (E.C.3.2.1.21), and trypsin (E.C.3.4.4.4) to an insoluble poly(allyl carbonate). The resultant activity of these solid-phase enzymes varied considerably with the pH of the coupling media and could be increased at a given pH by raising the concentration of the soluble enzyme and extending the duration of the coupling. It was found that the pH at which the maximum amount of protein is bound is not the same as that at which protein is bound with the greatest retention of enzyme activity. The factors influencing the extent of the coupling reaction are discussed. The extent of coupling of enzyme as a function of pH for poly(allyl carbonate) is compared with that for cellulose trans-2,3-carbonate, and it is concluded that the optimum conditions for coupling are unique for each enzyme and each insoluble matrix employed. The instability observed on storage of the insolubilised enzyme may be prevented by lyophilisation in the presence of a polyol, the latter presumably supplying a hydrophilic network in the solid state.