Abstract: In order to study the antimicrobial activity of horseradish leaf protein and its enzymatic hydrolysate,the protein in Moringa oleifera leaf was extracted by alkali extraction and acid precipitation,and its amino acid composition was analyzed. Using six single enzymes and three complex enzymes to enzymatic hydrolysis protein,the antimicrobial activity of Moringa oleifera leaves protein and its enzymatic hydrolysates was investigated to determine the best protease. The minimal inhibitory concentration(MIC)of effective antibacterial components on different bacteria was studied by double broth dilution method. The results showed that the extraction rate of protein was 32.67%±0.45%,and essential amino acids account for 39.27% of total amino acids. The research of antibacterial activity showed that the protein of Moringa oleifera leaves showed antibacterial activity only against Staphylococcus aureus and the antimicrobial zone diameter was(7.67±0.24) mm. The antimicrobial activity of the enzymatic hydrolysate obtained by sequential hydrolysis with pepsin followed by trypsin was higher than that of other proteases. The enzymatic hydrolysate had the strongest antibacterial activity for Streptococcus pyogenes and the antimicrobial zone diameter was(10.83±0.62) mm. The minimum inhibitory concentration(MIC)were 5,10 and 2.5 mg/mL,respectively. The results showed that both the protein and its enzymatic hydrolysate of Moringa oleifera leaf had antimicrobial activity,and the enzymatic hydrolysate prepared by double enzymatic hydrolysis had strong antimicrobial activity and wide antimicrobial spectrum.