Stability assay of Candida rugosa lipase in miniemulsion system to synthesis of biodegradable polymers

摘要

Polymers represent one of the most versatile classes of materials available for a variety of applications. Biodegradable polymers such as polyesters have been used in orthopedic devices, tissue engineering, adhesion barriers, control drug delivery, and pharmaceutical formulations [1]. Biocatalysis in miniemulsion systems presents an alternative and environment-friendly way to synthesize polyesters in mild reaction conditions [2]. Miniemulsions are heterophasic systems consisting of small, stable and narrowly distributed droplets in a continuous phase. For a typical oil-in-water miniemulsion, an oil phase (e.g. substrates), a hydrophobic agent (e.g. hexadecane), an emulsifier (usually nonionic surfactant), and water are homogenized to obtain monodisperse droplets [3]. In order to establish a reproducible synthesis of biopolymers, different dicarboxylic acids and dialcohols were tested and their influence on the enzymatic stability was observed. The Candida rugosa lipase s (CRL) stability was characterized by Circular Dichroism (CD), Fluorescence Spectroscopy (FS) and Estereolytic Activity (EA). Structural changes of CRL showed that reactions systems with longer carbon chain dicarboxylic acids (suberic acid C8 and adipic acid C6) in combination with the short carbon chain dialcohol (1,4-butanediol) showed a good stability of a lipases secondary and tertiary structures in a miniemulsion system during 32 h. However, the opposite combination (succinic acid C4 with 1,6-hexanediol and 1, 8-octanediol) showed an enzymatic denaturation immediately in the beginning of the assay, which indicates the negative impact of short carbon chain dicarboxylic acids on CRL stability in miniemulsion system.