Interfacial versus homogeneous enzymatic cleavage of mandelonitrile by hydroxynitrile lyase in a biphasic system

Abstract

The question of an interfacial versus a homogeneous reaction is carefully addressed for the enzymatic biphasic cleavage of mandelonitrile to benzaldehyde by Prunus amygdalus hydroxynitrile lyase (pa-Hnl) (Hickel et al. [1999] Biotechnol Bioeng 36:425-436). Experimental evidence, including 1) the reaction ceases when the interface is populated by previously adsorbed denatured pa-Hnl, 2) the reaction continues even after washout of the bulk enzyme from the aqueous phase, 3) highly nonpolar organic solvents initially promote fast reaction kinetics that relatively quickly decay to zero product production, and 4) the reaction rate is nonlinear in the bulk enzyme concentration, provide robust grounds for an interfacial reaction. We also model enzymatic mandelonitrile cleavage assuming a homogeneous aqueous-phase reaction. The homogeneous reaction scheme does not simultaneously account for the experimental observations of a linear dependence of the reaction rate on organic/water interfacial area, no dependence on the aqueous-phase volume, and a nonlinear dependence on pa-Hnl aqueous concentration. Further, simple calculations demonstrate that the homogeneous reaction rate is at least three orders of magnitude slower than those observed by Hickel et al. (1999). We again conclude that enzyme adsorbed at the organic solvent/water interface primarily catalyzes the biphasic mandelonitrile cleavage reaction.

Enzymatic catalysis via liquid–liquid interfaces

“…Therefore, the latter two rates can be set equal, i.e., a steady state might occur for the aqueous phase (Cascão-Pereira et al, 2002b):…”

“…It was also applied successfully to convert more difficult substrates such as 3-butenal, 4-pentenal, 5-hexenal, trans-cinnamic aldehyde, 2-hydroxybenzaldehyde, and 4-hydroxybenzaldehyde (Gerrits et al, 2001a,b;Willeman et al, 2002c). Nevertheless, the question is raised why the adsorbed enzyme model was not preferred to the mass transfer model (Cascão-Pereira et al, 2002b).…”

C−C Bond-Forming Lyases in Organic Synthesis

“…Enzymes in the DIPE/water interface are more active than in EtOAc. , As enzyme activity in biphasic systems takes place at the interface rather than in one phase of the solvent system, it is possible to remove the enzyme present in the aqueous phase by a simple phase separation step, so the enzyme could be reused. ,, …”

Oscillating drop/bubble tensiometry: effect of viscous forces on the measurement of interfacial tension

“…Surface-active molecules adsorbed at fluid/fluid interfaces influence many phenomena: food emulsion and foam stability [1,2], two-phase enzyme catalysis [3,4], human lung function [5][6][7], cell-membrane mechanical properties [8], and contact-angle hysteresis [9]. Time scales important to these phenomena are broad, necessitating an understanding of the dynamics of adsorbed molecules at interfaces.…”