New inhibitors of methane production by rumen micro-organisms. Development and testing of inhibitors in vitro.

1. A procedure is described for assaying in vitro the activity of various inhibitors of methane production by rumen micro-organisms. 2. Methods of preparation of various inhibitors are described together with attempts to characterize these compounds by determining their physical properties (physical state, density, chromatographic behaviour), their hydrolysis by rumen contents and their relative potency as inhibitors. 3. The results of preliminary studies with trichloroethanol and its ester with pivalic acid are given. 4. The inhibitory activities of several groups of related compounds are reported. These include the polyhalogenated alcohols and their esters with pivalic acid, the esters of trihalogenated alcohols and monobasic fatty acids from C2 and C16 and the trihalogenated alcohol esters of dibasic acids. The results of experiments with esters of alcohols and polyhalogenated carboxylic and sulphonic acids are also given. 5. It is concluded that the mechanism of action of the inhibitors might be similar to that of known polyhalogenated methane analogues (e.g. chloroform). The relative activity of various compounds might be partly governed by the ease of their absorption into the microbial cells and by the extent to which the esters can be hydrolysed by rumen contents. 6. The results show that some substances are very poor inhibitors, unless they are esterified (e.g. trichloroacetic acid) but on the whole the esters in which the polyhalogen grouping is on the alcohol portion of the molecule are better inhibitors than those in which it is on the acid portion of the molecule.