Immobilization of Pseudomonas fluorescens Lipase on Hollow Poly(o-phenylenediamine) Microspheres and Its Application in the Preparation of Citronellyl Acetate

Abstract

In order to address the challenges associated with free lipase in organic solvents, including aggregation, poor stability, and low catalytic efficiency, this study developed two types of poly(o-phenylenediamine) microspheres (solid and hollow) as supports for immobilizing lipase. The immobilization process utilized an adsorption method, with the poly(o-phenylenediamine) hollow microspheres being identified as the optimal support in a 2:5 enzyme-to-support ratio. On this basis, the lipase was immobilized by the covalent binding method. The immobilization conditions consisted of treating the support with 2% glutaraldehyde and immobilization at 40 °C for 2 h in pH 7.0 buffer. The specific activity of the immobilized enzyme was 5.3 times higher than that of the free enzyme. Covalent-binding immobilized lipase was also used for the preparation of citronellyl acetate by transesterification reaction, and, in optimized reaction conditions where the amount of immobilized enzyme was 0.1 g/mL, the reaction temperature was 50 °C and the shaking speed was 200 r/min during the reaction. Under these conditions, the citronellyl acetate yields can exceed 99% after 2 h. Furthermore, the stability of the immobilized lipase was investigated, and the residual activity of the immobilized enzyme was 95% after seven repetitions, while that of the free enzyme was only 70%. After 56 days of storage at room temperature, the immobilized enzyme retained 60% of its original viability, while the free enzyme retained only 31%.