Abstract: The hydrolysis of spore’s cortical peptidoglycan under HPTS treatment was a critical reason for spore inactivation. However,the hydrolysis principle was still not clear. The activity change of spore’s cortex-lytic enzyme had direct effects on hydrolysis of cortical peptidoglycan. The cortex-lytic enzyme was extracted from Bacillus subtilis spores,and the effects of sodium phosphate concentration,pH values,temperature and pressure on the activity of extracted enzyme were studied firstly,which was a basis for further research on HPTS’s effects on the activity. It was found that the optimal concentration of sodium phosphate was 0.05 mol/L,and when the concentration was 0.35 mol/L,the enzyme activity was the lowest. The optimal pH for the enzyme was 7,and at acidic or alkaline conditions its activity decreased. At 4~44 ℃ the enzyme had activity with no obvious change rule,but when the temperature increased to 68 ℃ the enzyme lost basically its activity. At 150~530 MPa,the pressure had almost no effects on activity of the enzyme,but HPTS treatment(530 MPa,68 ℃,15 min)inactivated the cortex-lytic enzyme basically,and it was the only enzyme in the spore that could hydrolyze cortical peptidoglycan specially,which suggested that non-enzymatic hydrolysis of spore’s cortical peptidoglycan occurred under HPTS treatment.