The objectives of the present study were to investigate the effect of five chemical inhibitors on methane production of the pure cultures of methanogens. To examine the methanogenesis inhibition by 2-bromoethane sulfonic acid (BES), 3-bromopropane sulfonic acid (BPS), lumazine, propynoic acid and ethyl 2-butynoate, they were added into the pure cultures of Methanobacterium formicicum KOR-1 and KOR-12, Methanoculleus bourgensis KOR-2, Methanosarcina mazei KOR-3, KOR-7, KOR-9 and KOR-10, Methanosarcina vacuolata (DSM 1232), Methanobacterium bryantii (ATCC 33272), Methanosarcina mazei (ATCC BAA159) and Methanosarcina barkeri (ATCC 43240). Each methanogen was anaerobically incubated at 39℃ for 6 and 10 days, and then methane gas production was measured. All methanogens were sensitive to BES, lumazaine and ethyl 2-butynoate and as the addition levels of the chemicals increased, the methanogenesis was linearly reduced. BPS and propynoic acid failed to inhibit any of the methanogens. There were important differences among species of methanogens regarding their sensitivity to the different inhibitors. In general, the strains of Methanosarcina species were the most resistant to BES, lumazaine and ethyl 2-butynoate and the strains of Methanobacteriales order the least resistant. The different resistances to chemical inhibitors may be caused by different cell membrane structure, substrate utilization for methanogenesis and competition with other microorganisms for electron receptors. In conclusion, differences among methanogens habituating in anaerobic ecosystems, such as the rumen of ruminants regarding their resistance to chemical inhibitors should be considered when strategies of inhibition of methanogenesis are designed.