Structure and properties of enzyme graft copolymers: effects of using dissolved agarose on horseradish peroxidase immobilization.
The immobilization of horseradish peroxidase (HRP) onto dissolved agarose by a photochemically initiated graft copolymerization reaction, carried out at room temperature, was studied. Enzyme immobilization parameters such as the catalyst (FeCl3) and the enzyme concentration were considered. Using hexhydro-1,3,5-triacryloyl-s-triazine (HTsT) as vinyl monomer, the agarose/HTsT ratio was the main reaction parameter controlling the copolymer characteristics. By increasing the polymer content of the sample better stability properties were obtained. For the samples with agarose/HTsT ratios of 20/40 and 40/20 (S 20-40, S 40-20) the residual activities after 240 min at 60 degrees C were respectively 47 and 18%. The residual activity in continuous working was 33% for S 40-20 (after 20 h) and 64% for S 20-40 (after 70 h). For both the synthesized copolymers no limitation to substrate diffusion was found but the flexibility of immobilized enzyme decreased with the increase of polymer content as indicated by the Km values that were 0.90 X 10(-4) mol/liter for the sample S 40-20, and 1.50 X 10(-4) mol/liter for the sample S 20-40. Other enzymes (glucose oxidase, alpha-chymotrypsin, and lipoxidase), besides HRP, were immobilized with good yields, showing the wide applicability of the proposed methodology for the preparation of a solid biocatalyst which can be conveniently stored in water suspension or as lyophilized material.
D002918:Chymotrypsin / Q000378:metabolism; D004800:Enzymes, Immobilized / Q000378:metabolism*; D005949:Glucose Oxidase / Q000378:metabolism; D006735:Horseradish Peroxidase / Q000378:metabolism*; D007202:Indicators and Reagents; D007700:Kinetics; D008084:Lipoxygenase / Q000378:metabolism; D010544:Peroxidases / Q000378:metabolism*; D012685:Sepharose
D004800:Enzymes, Immobilized; D007202:Indicators and Reagents; D012685:Sepharose; D005949:Glucose Oxidase; D006735:Horseradish Peroxidase; D010544:Peroxidases; D008084:Lipoxygenase; D002918:Chymotrypsin