Optimum Levels of Sulphate and Iron for the Cultivation of Pure Cultures of Methanogens in Synthetic Media*

Experiments on the effects, of sulphate and iron content of medium on the growth of Methanobacterium strain M.O.H., Methanospirllum hungatii GPI, and an unidentified strain similar in characteristics to Methanobacterium formicicum , indicated that the 4.0 mM of sulphate normally incorporated into laboratory media was above the optimum level, while the iron content normally incorporated (0.01–0.08 mM) was below the optimum requirement of these methanogens. The tests were made in synthetic media containing acetate, carbon dioxide and hydrogen as carbon and energy sources, and a cysteine‐sodium sulphide solution as a reducing agent. The optimum sulphate requirement for Methanobacterium strain M.O.H., M. hungatii , and the unidentified methanogen was in the range of 0.16–0.52 mM. An absence of sulphate in the medium decreased significantly (95% confidence level) the growth of these cultures. The optimum requirement of Fe ++ was in the range of 0.3–0.9 mM. Addition of Fe +++ greater than 0.10 mM to pre‐reduced medium, or the addition of greater than 0.15 mM during preparation of medium resulted in precipitation of iron. A method for the preparation of medium without formation of precipitate is described and its contribution to improving growth and methanogenesis discussed.