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Polyphenols, like flavonoids, have been investigated when present in intact plants or in extracts as methane mitigating dietary supplements in ruminants. The aim of the present study was to examine pure compounds in a short-term in vitro experiment using the Hohenheim Gas Test method. We focused on the group of the flavonoids and tested which of them had the potential to mitigate methane without negatively affecting ruminal fermentation. Eight flavonoids were tested: epicatechin, luteolin-7-glucoside, quercetin, and isoquercetin in Experiment 1; catechin, gallocatechin, epigallocatechin, and epigallocatechin gallate in Experiment 2. Tannic acid, no flavonoid but a phenolic acid with known methane mitigating properties, served as positive control, and the unsupplemented basal diet as negative control. In both experiments, each of these compounds (including tannic acid) was tested at dosages of 0.5, 5.0, and 50.0 mg/g basal diet dry matter (DM) in four runs each. Gallocatechin, tannic acid, and epigallocatechin gallate (50 mg/g DM) lowered fermentation gas formation and in vitro organic matter digestibility relative to the negative control (Experiment 2). Apart from tannic acid, epicatechin, quercetin, isoquercetin, and luteolin-7-glucoside (5 and 50 mg/g DM) reduced the amount of CH₄ produced in relation to total gas produced (Experiment 1). The incubation fluid ammonia concentration was decreased with luteolin-7-glucoside and tannic acid (50 mg/g DM). From the flavonoids tested especially luteolin-7-glucoside seems to have a similar potential as tannic acid to mitigate methane and ammonia formation during ruminal fermentation in vitro, both favourable in environmental respect. These results need to be confirmed in live animals.

Objective The methane mitigating potential of various plant-based polyphenol sources is known, but effects of combinations have rarely been tested. The aim of the present study was to determine whether binary and 3-way combinations of such phenol sources affect ruminal fermentation less, similar or more intensively than separate applications. Methods The extracts used were from Acacia mearnsii bark (acacia), Vitis vinifera (grape) seed, Camellia sinensis leaves (green tea), Uncaria gambir leaves (gambier), Vaccinium macrocarpon berries (cranberry), Fagopyrum esculentum seed (buckwheat), and Ginkgo biloba leaves (ginkgo). All extracts were tested using the Hohenheim gas test. This was done alone at 5% of dry matter (DM). Acacia was also combined with all other single extracts at 5% of DM each, and with two other phenol sources (all possible combinations) at 2.5%+2.5% of DM. Results Methane formation was reduced by 7% to 9% by acacia, grape seed and green tea and, in addition, by most extract combinations with acacia. Grape seed and green tea alone and in combination with acacia also reduced methane proportion of total gas to the same degree. The extracts of buckwheat and gingko were poor in phenols and promoted ruminal fermentation. All treatments except green tea alone lowered ammonia concentration by up to 23%, and the binary combinations were more effective as acacia alone. With three extracts, linear effects were found with total gas and methane formation, while with ammonia and other traits linear effects were rare. Conclusion The study identified methane and ammonia mitigating potential of various phenolic plant extracts and showed a number of additive and some non-linear effects of combinations of extracts. Further studies, especially in live animals, should concentrate on combinations of extracts from grape seed, green tea leaves Land acacia bark and determine the ideal dosages of such combinations for the purpose of methane mitigation.

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