Anticancer and antifungal efficiencies of purified chitinase produced from Trichoderma viride under submerged fermentation

Abstract

Trichoderma viride AUMC 13021 isolated from Mangrove soil of Ras Mohammed protected area at Sharm El-Sheikh, Egypt, was optimized to pro-mote chitinase activity under submerged fermen-tation. The maximum enzyme yield (38.33 U/mg protein) was obtained at 1.4% of colloidal chitin, 96 h of incubation, 35∞C, pH 6.5 and 125, rpm and using maltose (1%) and yeast extract (1%) as sup-plementation of salt basal medium. The enzyme has been purified with an overall yield of 73.1% and 5.48 purification fold, and a specific activity of 210.16 U/mg protein. The molecular mass of the purified chitinase was 62 kDa. Maximal activity of chitinase was recorded at pH 6.5 and 40∞C. The highest activity was recorded in the case of colloi-dal chitin, with an apparent Km value of 6.66 mg/ ml and Vmax of 90.8 U/ml. The purified chitinase was activated by Ca2+ and Mn2+ while the activity was inhibited by Hg2+, Zn2+, Cu2+, Co2+, dodecyl sulphate and EDTA. In vivo, the median lethal dose (LD50) was approximately 18.43 mg/kg body weight of Sprague Dawley rats. MTT assay showed that the purified chitinase has a toxic effect to MCF7 with an IC50 value 20 mg/ml, and HCT-116 cell lines with an IC50 value 44 mg/ml. Moreover, the purified enzyme showed significant antifungal activity against Fusarium oxysporum f. sp. lycopersici race 3 the causal agent of tomato wilt.