Porcine pancreatic lipase (PPL) was used to degrade the mycotoxin patulin (PAT) in pear juice. The dosage of PPL, the initial concentration of PAT, reaction temperature and time were investigated by batch experiments to study the optimal degradation condition. The concentration of PAT in pear juice was determined by high performance liquid chromatography with an ultraviolet detector (HPLC-UV). The results showed that the optimal condition was 0.02 g PPL/mL pear juice at 40 °C for 24 h. The content of organic acids, volatile flavor components, polyphenols, ascorbic acid and the degree of browning reaction in pear juice, relating to the quality of juice, changed insignificantly. Although the initial PAT concentration was very high, the degradation product was confirmed nontoxic by cytotoxicity test of Caco-2 cells. It suggested that PPL could be further considered to be applied in the degradation of PAT in pear juice.