Abstract: Veterinary antibiotics are widely used as feed additives in domestic animal rearing either to improve growth performance or to prevent disease infection. A wide range of antibiotics is excreted unchanged in urine and feces of animals, posing potential human and ecological health risks after entering the environment. This is confirmed by the presence of widespread detectable antibiotic concentrations in surface waters and manure. Antibiotics originating in manure from livestock operations are a concern because they remain bioactive. Anaerobic digestion is one of the most promising biotechnologies for the conversion of diverse organic substrates, ranging from high solid feedstocks (animal manure, food waste and municipal solid waste) to municipal and industrial wastewaters and to energy-rich biogas. Most of the biogas plants in China are run on animal manure to reduce biological oxygen demand and to produce biogas. After ingestion by animals, some of the compounds are excreted along with the manure which may inhibit biogas process when manure is used as substrate in biogas plants. Most studies on antibiotic inhibition under anaerobic fermentation have focused on animal manure as substrate, showing significant anaerobic fermentation inhibition primarily at higher antibiotic concentrations typically found in manure. An experiment was therefore designed to study the effects of Enrofloxacin (ENR) addition to pig manure on hydrolytic enzyme activities and biogas production in anaerobic fermentation processes with pig manure and corn stalks as raw materials. The results showed that pig manure anaerobic fermentation added with 20 mg?kg1, 60 mg?kg1 and 120 mg.kg^-1 ENR inhibited cellulose and urease activities at the initial stage of anaerobic fermentation (P < 0.05). However, the addition of 20 mg.kg^-1 ENR promoted the activities of sucrose, while the addition of 60 mg.kg^-1 and 120 mg.kg^-1 ENR inhibited the activities of sucrase in the first 15 days (P < 0.05). The effects of ENR on cellulase, urease and sucrose activities were not statistically different in the late stage of anaerobic fermentation (P > 0.05). Pig manure-based anaerobic fermentation with 20 mg?kg^-1, 60 mg.kg^-1 and 120 mg.kg^-1 ENR significantly inhibited gas production rate during 511 days and 2131 days (P < 0.05) of anaerobic fermentation. Compared with control (no ENR application), the gas production rate had no obvious changes in 20 mg.kg^-1, 60 mg.kg^-1 and 120 mg.kg^-1 ENR treatments after 31 days of anaerobic fermentation. Furthermore, biogas yield decreased by 7.38%, 12.08% and 15.77%, respectively, in 20 mg.kg^-1, 60 mg.kg^-1 and 120 mg.kg^-1 ENR treatments during anaerobic fermentation for 50 days. It was obvious that ENR influenced hydrolytic enzyme activities, gas production rate and biogas production during anaerobic fermentation. The effects of ENR on different hydrolytic enzyme activities and gas production rate were also different at different stages of anaerobic fermentation. The results provided a reference for harmless disposal of animal manure containing ENR and for improvement of anaerobic fermentation efficiency.