Immobilisation of soybean seed coat peroxidase on polyaniline Synthesis optimisation and catalytic properties

M.L. MAGRI; M.V. MIRANDA; O. CASCONE

BIOCATALYSIS AND BIOTRANSFORMATION

Taylor and Francis

Año: 2005 vol. 23 p. 339 - 346

1024-2422

  Soybean seed coat peroxidase (SBP) was immobilised on various polyaniline-based polymers (PANI), activated with glutaraldehyde. The most reduced polymer (PANIG2) showed the highest immobilisation capacity (8.2 mg SBP/g PANIG2).  Optimum pH for immobilisation was 6.0 and the maximum retention was achieved after a 6-h reaction period. Efficiency of enzyme activity retention was 82 %. When stored at 4ºC, the immobilised enzyme retained 80% of its activity for 15 weeks as evidenced by tests performed at 2-week intervals. The immobilised SBP showed the same pH-activity profile as that of the free SBP for pyrogallol oxidation and optimum temperature (55ºC) was 10ºC below that of the free enzyme. Michaelis-Menten experiments proved that Km was conserved while specific Vmax dropped from 14.6 to 11.4 mmol min-1 µg-1, in agreement with the immobilisation efficiency. Substrate specificity was practically the same for both enzymes. Immobilised SBP showed a greatly improved tolerance to different organic solvents: while free SBP lost around 90% of its activity at a 50% organic solvent concentration, immobilised SBP underwent only 30% inactivation at a concentration of 70% acetonitrile. Taking into account that immobilised HRP lost more than 40% of its activity at a 20% organic solvent concentration, immobilised SBP performed much better than its widespread counterpart HRP.