Lipid-membrane characteristics of large lipid-vesicles prepared by two-step emulsification technique and enzymatic NAD+-recycling in the vesicles.

Abstract

Large lipid-vesicles having a diameter of about 5–50 μm, consisting of Span8O (sorbitan monooleate), Tween80 (polyoxyethylene (20) sorbitan monooleate) and lecithin, were successfully prepared, with Span80 as the main constituent of the vesicle. The preparation was performed by applying the procedure of two-step emulsification for providing a W/O/W emulsion. The characteristics of the lipid membrane of the vesicle were experimentally examined and the main results can be described as follows.1) The vesicle could be observed with a phase-contrast microscope but not with an optical microscope. 2) With increase in the amount of an additive, soybean-lecithin, to Span80, the yield of the vesicle increased and the vesicle started to coagulate. 3) The vesicle yield increased with increase in weight ratio of water in water-lipid mixture, which was prepared by removing organic solvent from W/O emulsion. 4) The rupture percentage of the vesicle prepared with distilled water as the aqueous-phase solution was below 5% even after twenty days under an incubation temperature of 4°C.The applicability of the vesicle for encapsulation of enzyme was also examined. The model reaction in the vesicle was the enzymatic conversion of ethanol to acetaidehyde with NAD+-recycling catalyzed by malate dehydrogenase (MDH) and yeast alcohol dehydrogenase (ADH). It was found that enzymatic NAD+-recycling in the vesicle could occur successfully.