Optimised isolation, partial-purification and profiling of peroxidase extracted from saccharomyces cerevisiae of fresh palm wine

Abstract

Saccharomyces cerevisiae was obtained from palm wine and used for the extraction of peroxidase. Peroxidase isolated from Saccharomyces cerevisiae was partially purified and characterized. A twelve-day pilot study was carried out which gave the highest activity on day eight. The crude enzyme-specific activity –0.751 U/mg with 70% ammonium sulfate precipitation of peroxidase was achieved. Peroxidase was partially purified with ammonium sulfate precipitation, following gel filtration on Sephadex G-100. Two peaks connoted as fractions (A & B) on the elution profile of the gel filtration were obtained. The optimum operational conditions for peroxidase - substrate reactions were evaluated. A combined purification fold for both fractions yielded 0.657 with specific activity –0.494 U/mg. A purification fold of 2.470 with specific activity –1.855 U/mg was obtained for fraction A, while 2.269 fold with specific activity –1.704 U/mg was for fraction B. The optimum pH and temperature were 6.5 and 30°C for fraction A and 6.0 and 40 °C for fraction B. The peroxidase activity was stable at a pH range of 5.5-7.0 and below 40°C. The Michaelis-Menten constant (Km) for o-dianisidine and hydrogen peroxide were 0.2 mM, 0.1111 mM, and 0.0833 mM, 0.1428 mM for fractions A and B. The maximum velocity (Vmax) for o-dianisidine and hydrogen peroxide were 10 U/min, 7.1428 U/min, and 8.333 U/min, 9.0909 U/min for fraction A and B. considering these characteristics of peroxidase from Saccharomyces cerevisiae, it helps to understand how yeast cells operate during fermentation.