Direct Production of an Extracellular Tyrosinase from Rhizopus oryzae NRRL-1510 by Solid Substrate Fermentation

Tyrosinase (EC 1.14.18.1) belongs to copper containing metalloproteins essential in melanogenesis pathway. This enzyme finds biotechnological applications in L-dopa production, wound healing in higher organisms, detection and removal of phenolic compounds from waste water. In the present study, we report on the novel production of an extracellular tyrosinase from Rhizopus oryzae NRRL-1510 by solid substrate fermentation (SSF). A range of different agricultural by-products including sunflower meal (SFM), soybean meal (SBM), almond meal, canola meal, mustard seed cake and rape seed meal collected from various local markets were evaluated as basal substrate under SSF. Initial moisture content was determined using different diluents including distilled water (pH 7), sodium citrate buffer (pH 8.1), phosphate buffer (pH 7.2) and acetate buffer (pH 3.5). Sodium citrate buffer (100 ml) was used to extract enzyme from the fermented mash culture. A noticeable enhancement in enzyme activity (64.55 U/mg) was observed when the significant process parameters viz. SFM level (15 g), volume of distilled water (20 ml), time of incubation (72 h), temperature (30 degrees C) and level of sodium citrate buffer at pH 8.1 (100 ml) were determined after Plackett-Burman design. Thin layer chromatography (TLC) of tyrosinase catalysis-products confirmed the presence of an active tyrosinase in the reaction mixture. The value of tyrosinase correlation (0.135E+0025) depicted that the model terms are highly significant (HS, p <= 0.05) indicating commercial viability of the fungal culture (df = 3, LSD = 0.0385).