Antibiotics are used in dairy farms for the treatment and prevention of diseases in dairy cattle, such as mastitis, lameness, respiratory infections and diarrhea. Antibiotic residues in milk can generate problems in the population in the form of allergies and antimicrobial resistance, as well as in industry and the environment. There are national and international regulations that prohibit the processing of milk with antibiotics, so it is important to study alternatives for their disposal. The aim of this work was to study the effect of treatment with β-lactamase (BL) on the degradation of penicillin G (PG) in milk. The trials were performed on skimmed milk powder contaminated with three antibiotic concentrations: 6, 12 and 30 ppb. Three concentrations of BL (0.1, 0.5 and 1.0 mU/mL) were tested. Samples were taken at 3, 6, and 9 hours for detection and quantification of antibiotics. The results obtained indicate that in three hours the PG was degraded using 0.5 mU/mL of β-lactamase, and in six hours with a lower concentration of enzime (0.1 mU/mL) in all the antibiotic concentrations analyzed. The β-lactamase studied have a high potential to degrade penicillin in milk in dairy farms before final disposal. Key words: antibiotic degradation, microbial enzymes, milk