Barengo MP, Amerio NS, Bich GA, Castrillo ML, Zapata PD

Language: Spanish
References: 0
Page: 2201-2206
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ABSTRACT

Mycoparasitic fungi are capable of hydrolyzing the cell walls of their host by secreting hydrolytic enzymes, such as proteases. To quantify proteolytic activity, the methodology based on the use of the azocasein substrate is considered one of the most reliable methodologies. However, in the literature it has multiple modifications available according to its applicability and the type of microorganism. Therefore, this work was aimed to establish the optimal reaction conditions using the papain pure enzyme; in order to quantify the proteolytic activity of fungal enzyme supernatants from Trichoderma and Escovopsis. The effect of reaction time, pH, and temperature over enzymatic activity at different concentrations of papain was tested. The chromogenic substrate azocasein was used to measure the enzymatic activity. Enzyme supernatants were obtained by liquid fermentation of two fungal mycoparasites: Escovopsis HEP25 and Trichoderma POS7. According to the evaluated conditions, significant proteolytic activities were observed at 50 min of reaction, 37 °C and pH 7.4.The activity was expressed in terms of units equivalent to the activity of a given mass of papain (1 unit = 1 mg of papain). For the Trichoderma strain (POS7) an activity of 54.3 ± 2.8 U/L was obtained, and for the Escovopsis strain (HEP25) of 2 ± 0.5 U / L. The adjusted methodology and the enzyme activity calculation were determined to be appropriate for analyzing the crude supernatant samples. Particularly, for the Escovopsis genus, the determination of quantitative proteolytic activity is reported for the first time.