Saccharomyces cerevisiae alcohol dehydrogenase (SCAD) was effectively immobilized on hybrid alginate-chitosan beads which are hardened with glutaraldehyde. Immobilization conditions and characterization of the immobilized enzyme were investigated. Orthogonal test design and intuitive analysis method were employed to evaluate the effects of immobilization parameters such as Na-alginate concentration, glutaraldehyde concentration, CaCl(2) concentration and immobilization time. Under optimized working conditions (3.0% Na-alginate, 0.5% chitosan, 2.0% CaCl(2), 0.5% glutaraldehyde and 6h), the SCAD activity was 339.25 U/mL. For the reduction of phenylglyoxylic acid, the immobilization process changed the enzyme's optimum temperature from 30 to 40 degrees C, the enzyme's optimum pH from 6.8 to 6.0, and the immobilized SCAD retained 62.76% of its original activity. The maximum reaction rate (V(max)) was 358.63 nmol min(-1) and the Michaelis-Menten constant (K(m)) was 37.33 mmol L(-1). Compared to the free SCAD, the immobilization of the enzyme showed higher thermal stability and operational stability.
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