The results of our study established that in vitro methanogenesis was not essentially related to the density of protozoa population. Tropical tree leaves containing tannins such as Autocarpus integrifolia, Jatropha curcus and Sesbania grandiflora have the potential to suppress methanogenesis. Therefore tannins contained in these plants could be of interest in the development of new additives in ruminant nutrition.

Ficus bengalensis, Au. integrifolis and Az. indica leaves are of interest in the enteric methane ameliorative strategies. Total mixed ration containing 10-15% ground F. bengalensis or Au. integrifolis or Az. indica leaves could be fed to ruminants to reduce enteric methanogenesis. However, in vivo trials need to be carried out to validate these in vitro results.

The potential of tannins from 21 medicinal and aromatic plant leaves as antimethanogenic additives in ruminant feeds was investigated. The effect of tannin from these leaves on rumen fermentation parameters, protozoa population and methanogenesis was studied by incubating the samples [200 mg dry matter (DM)] without and with polyethylene glycol (PEG)-6000 (400 mg DM) as a tannin binder during 24-h incubation in the in vitro Hohenheim gas method. Based on the methane percentage estimated in the total gas produced, methane production in millilitre was calculated [methane volume (ml) = methane % × total gas produced (ml) in 24 h]. In the samples, crude protein and neutral detergent fibre (g/kg DM) ranged from 113 to 172 and from 352 to 444 respectively. The total phenol (TP; g/kg DM) content was highest in Terminalia chebula (274) followed by Hemigraphis colorata (71) and Sapindus laurifolia (51) respectively. In the remaining samples, it was <43 g/kg DM. Activity of tannins, as represented by the increase in gas volume on addition of PEG, ranged from 0 to 133%, with the highest being recorded in T. chebula. The per cent increase in methane on PEG addition was 0 for Ammi majus, Aristolochia indica, Cascabela thevetia, Ipomea nil and Lantana camara, illustrating that tannins present in these samples had no effect on methane concentration. The PEG addition increased the total protozoa count by >50% in A. indica and C. thevetica. One of the important findings of our study was that of the 21 samples screened, Entodinia population increased in 12 with PEG as compared to 7 where Holotricha increased, indicating higher susceptibility of Entodinia to tannin. There was no increase in the protozoa population with PEG when incubating Cardiospermum halicacabum, Clerodendrum inerme, Dioscorea floribunda, Nerium oleander and Selastras paniculatus, which strongly suggested that methane suppression recorded in these samples was not because of a defaunating effect of their tannins per se. The fermentation pattern reflected increased total volatile fatty acid (TVFA) concentration from 0 to 28.3% with PEG addition among the leaves. Our results confirmed further observations that methanogenesis in vitro is not essentially related to density of protozoa population. Secondly, medicinal and aromatic plants such as C. inerme, Gymnema sylvestre and Sapindus laurifolia containing tannins appear to have a potential to suppress in vitro methanogenesis.

Aims: 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. Methods and Results: 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 À1 DM) was maximum in Pimenta officinalis (312) followed by Oenothera lamarckiana (185) and Lawsonia inermis (105). Of the 38 samples, condensed tannins exceeded 4Á0 g kg À1 in only Alpinia galanga (7Á50), Cinnamomum verum (4Á58), Pelargonium graveolens (18Á7) and Pimenta officinalis (23Á2) 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 (131Á8) leaves, followed by Indigofera tinctoria (16Á8) and Withania somnifera (10Á2) leaves. Total and differential protozoal counts increased with added PEG in twentytwo 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 NH 3 -N with added PEG.Conclusions: 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. Significance and Impact of the Study: 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.

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