Phosphatase active cross-flow microfiltration poly(vinylidene difluoride) bioreactor

Alkaline phosphatase from human placenta has been chemically immobilized on a hydrophilic cross-flow microfiltration membrane made from poly(vinylidene difluoride) (PVDF) derivatized with 1,1'-carbonyldiimidazole. The physicochemical characterization of the immobilized biocatalyst paid special attention to the irreversibility of the bonding of the enzyme to the support, the effects of pH, temperature and ionic strength on this activity, the existence of limitations of internal and external diffusion for H+, substrate and/or products, and the kinetic behaviour (intrinsic and/or effective) of the immobilized enzyme. With respect to enzyme stability, patterns of hysteresis or memory are proposed, to account for a catalytic activity affected by previous experimental events and situations. The intrinsic kinetic behaviour, rate versus substrate concentration in the absence of diffusional restrictions, was analysed graphically and numerically (by non-linear regression and by utilizing the F statistical test for model discrimination), postulating a minimum rational rate equation of 2 : 2 degree in substrate concentration. In concordance, a mechanistic kinetic scheme for the catalytic enzyme action has been postulated.