Immobilization of Invertase in DAD Type Polymers: Combination of Benzothiadiazole Acceptor Unit and 3,4-Ethylenedioxythiophene and Thiophene Donor Units

Abstract

Conductive polymers with donor-acceptor-donor (DAD) type units; benzothiadiazole acceptor unit and 3,4-ethylenedioxythiophene (EDOT) and thiophene (Th) donor units, were investigated for immobilization of invertase. The polymers were prepared potentiodynamically from their monomers, M1 (Th-benzothiadiazole-Th), M2 (EDOT-benzothiadiazole-EDOT), M3 (Th-benzothiadiazole-EDOT) and a copolymer, which is a homolog to homopolymer of M3, was prepared from M1 and M2. Invertase was trapped between a two layer-composite; DAD polymer and polypyrrole. The characterization of immobilized invertase was performed using a spectrophotometric method at 540 nm. Kinetic parameters, V max, maximum reaction rate and Km , Michaelis-Menten constant values of immobilized invertase in DAD type homopolymer (P3), 0.95 μmol min−1 and 22.7 mM, respectively, are found between those of homopolymers P1 and P2 and are better than the copolymer. Polymers and copolymers exhibited a broad optimal temperature profile between 30 and 50°C compared to the free enzyme. Optimum pH (5.0) is the same as for free invertase. The electrodes were found to be stable with 100% activity during one day for 40 consecutive measurements. As to the self-life, 25% of the initial activity was lost in the first ten days, then the electrodes were stable with 75% activity for a 40 day storage at 4°C.