As the typical quarternary ammonium compounds (QACs), benzalammonium bromide (BK) are frequently detected in sewage sludge. This study explored the responses of methane production from digestion sludge at present different BK addition. Experimental results indicated that BK stress seriously inhibited the methane production at the pre-anaerobic digestion stage, but the final cumulative methane production (∼ 1124.0 ± 40.2 mL/g VSS) was similar in these digesters. While BK obviously promoted the release and transformation of organic substances, but severely inhibited the rate of methane production. In addition, the relative abundances of hydrogenotrophic methanogens were significantly decreased from 69.38% to 38.57%, but the acetotrophic methanogens enhanced from 26.75% to 58.86% with BK exposure from 0 to 8.69 mg/g VSS. Correspondingly, BK addition could impact the key functional genes involved in the major methanogenic metabolic pathways and energy metabolism. This work shed light on the effect of BK on methane production under anaerobic sludge digestion system, and provide useful information to understand the relevant mechanism.