Extraction and determination of trypsin enzyme from visceral organs of rainbow trout (Oncorhynchus mykiss) and characteristic's optimizations for obtained trypsin

This investigation aimed to extract, characterize and to study the effect of some parameters on trypsin proteinase recovery from visceral organs of Rainbow Trout (Oncorhynchus mykiss). Optimal temperature for the activity of extracted Trypsin was determined as 45 °C. Enzyme was active at temperature ranges of (20–60 °C), where maintained 85–90% of initial activity at 60 °C. At 80 °C enzyme lost 85% of its activity. The optimal pH for the trout trypsin determined to be 8, where 90% of activity was retained. Enzyme was stable between neutral and alkaline pH ranges in which 75–85% of relative activity maintained and maximum activity observed between pH 7–8. The enzyme was stable in a wide pH ranges of 6.0–11.0 maintaining 80% of its initial activity. But unstable at the temperatures greater than 55 °C, where enzyme gradually lost its activity at temperatures greater than 55 °C. The proteolytic activity was strongly inhibited by some trypsin inhibitors. In the presence of Benzamidine HCl, trypsin lost 38% of its relative activity. While Phenylmethylsulfonyl fluoride (PMSF) decreased activity by 67%, and at the existence of Ethylenediaminetetraacetic acid (EDTA), 81% of trypsin activity was lost. Both EDTA and PMSF were observed to have significant inhibiting impact on trypsin activity. Enzyme activity continuously decreased with increasing NaCl concentration. While at 0% NaCl, relative activity of trypsin was 100%, at 1% NaCl 53.31 activity was observed. Trypsin at %3, 5, 7.5, 10, 12.5, and 15 NaCl ranges, showed 47.55, 43.52, 40.98, 37.82, 44.33 and 37.16 relative activity respectively. At 40 °C, CaC performed as an activator on trypsin enzyme. The molecular weight of trout trypsin was determined to be 25 kDa based on SDS-PAGE Electrophoresis. Keywords: Trypsin enzyme, Fish internal organs, Trout, Extraction, Characteristics.