Candida rugosa Lipase Supported on High Crystallinity Chitosan as Biocatalyst for the Synthesis of 1-Butyl Oleate

Abstract

Lipase from Candida rugosa was immobilized onto chitosan using four different protocols. The variation of crystallinity (5.57–92.86%), which was a result of thermal treatments and crosslinking of the chitosan, influenced the protein load (7.46–25.15 mg g⁻¹ chitosan) and protein load efficiency (21.67–41.68%) for immobilization assays made with identical lipase solution concentration (1.3 mg of protein/mL). The effects of protein load (10, 30, 50 and 70 mg of lipase), reaction temperature (30, 40, 50, 60, 70 °C) and substrates molar ratio (0.05–0.30 M) have been studied in the butyl oleate synthesis in iso-octane when water activity of the free and immobilized enzymes were fixed around 0.53 ± 0.04. The catalytic activity of the immobilized lipase has also been tested. The Ping–Pong bi–bi mechanism with dead end complex of n-butanol was found to fit the initial rate data. The values of the apparent kinetic parameters were determined by graphic and parametric method as: V _max = 18.2–19.0 mmol min⁻¹ g⁻¹; K _{M; Acid} = 0.599–0.640 mol L⁻¹; K _{M; Alcohol} = 0.128–0.149 mol L⁻¹; and K _{i; Alcohol} = 1.933 mol L⁻¹.