PSEUDOMONAS-FLUORESCENS LIPASE ADSORPTION AND THE KINETICS OF HYDROLYSIS IN A DYNAMIC EMULSION SYSTEM

To elucidate the adsorption characteristics of lipases and to study th e influence of the reaction conditions on the catalytic properties of lipases, the hydrolysis of decylchloroacetate by Pseudomonas fluoresce ns lipase in an emulsion reactor was studied as a model system. During the reaction the droplet size distribution of the emulsion was measur ed on-line using a particle sizer based on light scattering. Desorptio n experiments revealed that, at low surface coverage, the initial rate of reaction was not influenced by either the stirring speed or the or ganic volume fraction. Dilution of the reaction mixture during hydroly sis did not result in a decrease in activity. Based on these results, it is assumed that under the specified conditions adsorption of Pseudo monas fluorescens lipase is quantitative and probably irreversible. Ba sed on activity measurements and assuming that only a monolayer of lip ase is active, it is calculated that at saturation the emulsion interf ace is covered with 3 mg lipase per m(2). From these data the average interfacial area covered by one lipase molecule at saturation was calc ulated to be 1700-2100 Angstrom(2) per molecule. The emulsion was show n to be dynamic, e.g., during hydrolysis a significant increase in int erfacial area was observed as a result of a shift in droplet size dist ribution to smaller diameters. Experiments indicated that both the for mation of decanol and the emulgating effect of the lipase account for these observations. The formation of decanol also resulted in a dramat ic decrease in hydrolytic activity. Taking interfacial tension measure ments into account, it is shown that decanol accumulates at the liquid -liquid interface. As a result of this high local decanol concentratio n at the interface, a reversed reaction, alcoholysis, occurred.