The experiments which are required to directly assess the operational stability of thermostable biocatalysts can be time-consuming, troublesome, and, in the context of industry, expensive. In the present work, we develop and validate two methods for quickly estimating the total turnover number (a useful indicator of lifetime productivity) of a biocatalyst for any desired operating temperature. The first method is a heuristic approach, built upon a complete mathematical derivation from first principles, in which the total turnover number can be calculated from two simple biochemical measurements. The second method relies on a single non-isothermal, continuous-mode experiment in conjunction with mathematical modeling to determine the intrinsic deactivation parameters of the biocatalyst. Both methods provide estimates of the total turnover number which are well within one order of magnitude of the values measured directly via isothermal aging tests and therefore are extremely valuable tools in terms of the amount of experimental time eliminated.

Biocatalytic membrane reactors combine the highly efficient biotransformation capability of enzymes with the selective filtration performance of membrane filters. Common strategies to immobilize enzymes on polymeric membranes are based on chemical coupling reactions. Still, they are associated with drawbacks such as long reaction times, high costs, and the use of potentially toxic or hazardous reagents. In this study, a reagent-free immobilization method based on electron beam irradiation was investigated, which allows much faster, cleaner, and cheaper fabrication of enzyme membrane reactors. Two industrial lipase enzymes were coupled onto a polyvinylidene fluoride (PVDF) flat sheet membrane to create self-cleaning surfaces. The response surface methodology (RSM) in the design-of-experiments approach was applied to investigate the effects of three numerical factors on enzyme activity, yielding a maximum activity of 823 118 U m2 (enzyme concentration: 8.4 g L1, impregnation time: 5 min, irradiation dose: 80 kGy). The lipolytic membranes were used in fouling tests with olive oil (1 g L1 in 2 mM sodium dodecyl sulfate), resulting in 100% regeneration of filtration performance after 3 h of self-cleaning in an aqueous buffer (pH 8, 37 C). Reusability with three consecutive cycles demonstrates regeneration of 95%. Comprehensive membrane characterization was performed by determining enzyme kinetic parameters, permeance monitoring, X-ray photoelectron spectroscopy, FTIR spectroscopy, scanning electron microscopy, and zeta potential, as well as water contact angle measurements.