The effects of pepsin-assisted hydrolysis conditions on the antioxidant activity (AA) of porcine hemoglobin hydrolysate (PHH) were investigated in this study. Effects of temperature, pH and enzyme to substrate ratio on the AA were evaluated and compared using response surface methodology. Temperature and pH were the major factors affecting the AA. Enzyme to substrate ratio influenced AA to a lesser extent. The model for the AA of PHH was established and it showed a good fit with the experimental data. The PHH (1 mg/mL) showed the strongest AA (65.43%) at 40.4C, pH 1.6, enzyme to substrate ratio = 1.6% (w/w). The degree of hydrolysis (DH) and AA of pepsin-assisted hydrolyzed PHH were significant higher (P < 0.05) than chemical hydrolyzed PHH whatever the hydrolysis time. There was no correlation between DH and AA.