Abstract

ENTRAPMENT OF INVERTASE AND GLUCOSE OXIDASE IN ACRYLAMIDE MATRIX

Michele Vitolo*

Abstract

Invertase and glucose oxidase were immobilized by entrapment in polyacrylamide hydrogel. The support was electrically neutral as pHoptimum shifting was not observed for both enzymes. The concentration of the cross-linking agent (EDMA) played a crucial role in the immobilization coefficient (IC), which was over 30% for both enzymes. The concentrations of IC and EDMA were linearly correlated. Invertase and glucose oxidase (GO), either soluble or immobilized, were highly stable when used in a membrane bioreactor operated continuously for at least 30 h. All enzymes forms were thermodynamically characterized and the following kinetic constants were calculated: a) Invertase: a.1) soluble form: Vmax = 84 g RS/min. g protein; KM = 22.6 mM; Ea = 37.0 kJ/mol; G = -10.5 kJ/mol; H = 34.4 kJ/mol; S = 0.141 kJ/mol.K; a.2) immobilized form: Vmax = 65 g RS/min. g protein; KM = 17.0 mM; Ea = 24.3 kJ/mol; G = - 10.6 kJ/mol; H = 21.7 kJ/mol; S = 0.102 kJ/mol.K; b) Glucose oxidase: b.1) soluble form: Vmax = 0.218 A/min; KM = 7.89 mM; Ea = 28.0 kJ/mol; G = - 11.4 kJ/mol; H = 25.4 kJ/mol; S = 0.180 kJ/mol.K; b.2) immobilized form: Vmax = 0.144 A/min; KM = 7.69 mM; Ea = 53.5 kJ/mol; G = - 11.0 kJ/mol; H = 50.9 kJ/mol; S = 0.20 kJ/mol.K.

Keywords: Entrapment, acrylamide, glucose oxidase, invertase.