Hydrazide-Functionalized Poly(2-hydroxyethyl methacrylate) Microspheres for Immobilization of Horseradish Peroxidase

Nonporous cross-linked poly(2-hydroxyethyl methacrylate-co-ethylene dimethacrylate) (poly(HEMA-co-EDMA)) microspheres were prepared by dispersion polymerization of HEMA and EDMA. The polymerization was performed in toluene/2-methylpropan-lol in the presence of cellulose acetate butyrate as a steric stabilizer and dibenzoyl peroxide initiator. The particle size may be increased by decreasing the toluene/2methylpropan-l-ol ratio and by increasing polymerization temperature. Adipohydrazide was attached to the microspheres activated with 2,4,6-trichloro-l,3,5-triazine. After periodate oxidation of its carbohydrate moieties, horseradish peroxidase was coupled to the hydrazide-functionalized poly(HEMA-co-EDMA) microparticles up to 7.3 (mu)g of enzyme/g of carrier without a significant loss of its activity. Immobilized peroxidase was found to be stable, retaining more than 97% of its initial activity when stored for 23 days after the preparation.