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J Appl Microbiol. 2013 Aug;115(2):455-65. doi: 10.1111/jam.12238. Epub 2013 May 16.

Effect of plant secondary compounds on in vitro methane, ammonia production and ruminal protozoa population.

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Energy Metabolism Laboratory, Division of Bioenergetics and Environmental Sciences, National Institute of Animal Nutrition and Physiology, Bangalore, India.



The objective of this study was to evaluate the potential of secondary plant metabolites from 38 sources to serve as antimethanogenic additives in ruminant diets. The effect of leaf tannins from these different plant sources on rumen fermentation, protozoal populations and methanogenesis was also studied.


Samples (200 mg dry matter, DM) were incubated without and with polyethylene glycol (PEG)-6000 (400 mg DM) as a tannin binder during 24-h incubation in the in vitro Hohenheim gas system. In the leaf samples, total phenol (g kg(-1) DM) was maximum in Pimenta officinalis (312) followed by Oenothera lamarckiana (185) and Lawsonia inermis (105). Of the 38 samples, condensed tannins exceeded 4.0 g kg(-1) in only Alpinia galanga (7.50), Cinnamomum verum (4.58), Pelargonium graveolens (18.7) and Pimenta officinalis (23.2) and were not detected in seven samples. When the bioactivity of the leaf samples was assessed using the tannin bioassay, the percentage increase in the amount of gas produced during incubation of samples with the tannin-binding agent PEG-6000 over the amount produced during incubation without the tannin binder ranged from nil (zero) to 367%, with the highest being recorded with A. galanga leaves. The ratio of methane reduction per ml of total gas reduction was maximum with Rauvolfia serpentina (131.8) leaves, followed by Indigofera tinctoria (16.8) and Withania somnifera (10.2) leaves. Total and differential protozoal counts increased with added PEG in twenty-two samples, maximum being in Pimenta officinalis. Increased accumulation of total volatile fatty acids during incubation with added PEG-6000 was recorded, and the values ranged from zero to 61%. However, the increase was significant in only 11 of the 38 tannin sources tested indicating noninterference of tannin on in vitro fermentation of carbohydrates by the majority of samples tested. Conversely, in 26 of 38 plant sources, the leaf tannins reduced N-digestibility as evidenced by increased accumulation of NH3 -N with added PEG.


Our study unequivocally demonstrated that plants containing secondary metabolites such as Rauvolfia serpentine, Indigofera tinctoria and Withania somnifera have great potential to suppress methanogenesis with minimal adverse effect of feedstuff fermentation.


It was established that methanogenesis was not essentially related to the density of protozoa population in vitro. The tannins contained in these plants could be of interest in the development of new additives in ruminant nutrition.


methane; plant secondary compounds; protozoa; rumen fermentation

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