COMPARISON OF HYDROLYSIS AND ESTERIFICATION BEHAVIOR OF HUMICOLA LANUGINOSA AND RHIZOMUCOR-MIEHEI LIPASES IN AOT-STABILIZED WATER-IN-OIL MICROEMULSIONS .1. EFFECT OF PH AND WATER-CONTENT ON REACTION-KINETICS

Lipolase and Lipozyme are produced in large quantities (as a result of genetic engineering and overexpression) for the detergents market and provide a cheap source of highly active biocatalysts. Humicola lanugi nosa lipase (HIL) and Rhizomucor miehei lipase (RmL) have been isolate d in partially purified form from commercial preparations of Lipolase and Lipozyme, respectively. These lipases were solubilized in Aerosol- OT (AOT)-stabilized water-in-oil (w/o) microemulsions in n-heptane. HI L and RmL activity in these microemulsions was assayed by spectrophoto metric measurement of the initial rate of p-nitrophenyl butyrate hydro lysis, and by chromatographic determination of the initial rate of oct yl decanoate synthesis from l-octanol and decanoic acid. The hydrolyti c activity of RmL in microemulsions measured as a function of buffer p H prior to dispersal, followed a sigmoidal profile with the highest ac tivities observed at alkaline pi-is. This broadly matches the pH-activ ity profile for tributyrin hydrolysis by Lipolase in an aqueous emulsi on assay. The hydrolytic activity of RmL in the same microemulsions, m easured as a function of pH, gave a bell-shaped profile with a maximum activity at pH 7.5. Again, the observed pH-activity profile was simil ar to that reported for a purified RmL in a tributyrin-based aqueous e mulsion assay. In contrast, the esterification activity exhibited by b oth HIL and RmL in AOT microemulsions over the available range pH 6.1 to 10.4, decreases as the pH increases, most likely reflecting the eff ect of substrate ionization. The dependence of the hydrolytic and cond ensation activity of HIL on R, the mole ratio of water to surfactant, were similar with both profiles exhibiting a maximum at R = 5. The hyd rolytic and esterification activities of RmL followed similar R-depend ent profiles, but the profiles in this case exhibited a maximum at R = 10. The water activities at these R values were directly measured as 0.78 and 0.9, respectively. Measured water activities were unperturbed by the presence of lipase at the concentrations used in these studies . (C) 1995 John Wiley & Sons, Inc.