Abstract: A UHPLC-MS/MS method was used to measure penicillin residues in beef and milk. The sample was extracted with NaH2PO4 solution. Zinc acetate was used to precipitate proteins,and hexane was used for defatting. The extract was then purified on a HLB solid phase extraction column. The penicillin was detected by UHPLC-MS/MS and quantified by external standard method. The linear ranges of 6 kinds of penicillin were within 25.0 μg·L-1,the detection limit (3S/N) was 2 μg·kg-1 and the lower limit of determination (10S/N) was 5 μg·kg-1. The recovery rates measured by standard addition method were in the range of 77.0%-99.8%,with RSD (n=6) was in the range of 3.7%-13%.