This study was conducted to investigate the effect of sulfate loading on methane production and organic matter degradation during the mesophilic anaerobic co-digestion of corn stover and bio-kerosene production wastewater (BKPW). The highest methane production of 192.04 mL/gVS was obtained at a sulfate concentration of 86 mg/L. This was 46.80% higher than that achieved by a sulfate concentration of 113 mg/L. Additional degradation of organic matter was obtained at a sulfate concentration of 113 mg/L because organic matter in the corn stover and BKPW was oxidized by sulfate-reducing bacteria (SRB). The concentration of sulfate declined by approximately 23% after 29 days of anaerobic co-digestion, and this reduction in sulfate was enhanced when the soluble chemical oxygen demand (SCOD)/sulfate ratio was less than 15. The results of a mass balance analysis showed that 34.87% of C element and 10.04% of S element in substrate, respectively, were converted to biogas during anaerobic co-digestion of corn stover and BKPW at a sulfate concentration of 86 mg/L. The microbial community was analysed using 16S rDNA sequencing technology, and the results showed that the relative abundance of Synergistes (related to methane production with acetic acid) at a sulfate concentration of 86 mg/L had obviously increased and was approximately 287% higher than the abundance achieved at a sulfate concentration of 113 mg/L. (C) 2018, The Society for Biotechnology, Japan. All rights reserved.