A complex of trypsin and chymotrypsin effectively inhibited growth of pathogenic bacteria inducing cow mastitis and showed synergistic antibacterial activity with antibiotics

• The complex of trypsin and chymotrypsin at C/2C showed wide antimicrobial spectrum.
• The complex of tested enzymes had synergistic antibacterial effect with antibiotic.
• The complex of tested enzymes destroyed the external structure of bacterial cells.
• The complex of tested enzymes caused leakage of bacterial intracellular materials.
• Effect of the enzymes complex was verified in therapeutic test of diseased animals.

The objective of this research was to assess the inhibitory effects of a complex of trypsin and chymotrypsin alone and in combination with antibiotics, on pathogenic bacteria involved in cow mastitis in vitro. We also conducted investigations of the antimicrobial mechanism and primary treatment trials on infected cattle. The presence of trypsin (0.16 mg/mL) alone only clearly hindered the growth of Streptococcus agalactiae (CVCC586) and Streptococcus dysgalactiae (ATCC12388). Chymotrypsin (0.16 mg/mL) alone, and the complex of trypsin and chymotrypsin at the dose C (0.16 mg/mL trypsin + 0.16 mg/mL chymotrypsin) or 2 C showed varying inhibitory effects on all the tested bacterial strains, including Escherichia coli (ATCC8739), Pasteurella (C51-3), Staphylococcus aureus (ATCC25923). The experiments also indicated that the complex of trypsin and chymotrypsin could hydrolyze bacterial outer-membrane proteins, damage the integrity of surface structures, and lead to leakage of intracellular material such as alkaline phosphatase, glucose and DNA. The complex of Trypsin and chymotrypsin showed well synergistic antibacterial effects when combined with specific antibiotics. In the field trial on 20 cows naturally suffering from clinical mastitis, the complex of trypsin and chymotrypsin reduced effectively Streptococcus, E. coli, S. aureus, and some other Enterobacteriaceae as well as the somatic cell counts in milk samples of treated cattle compared to controls. The complex of trypsin and chymotrypsin, and its combination with antibiotics, seems to have a potential in clinical veterinary medicine to treat mastitis.