The objective of this study was to determine the chemical compounds of Iranian propolis (IP) extracts and to show flavonoids and phenol effects on methane production, fermentation characteristics and rumen microbial population (in vitro). In this study two diets with different concentrate: forage ratios as (HC: high concentrate) and (MC: middle concentrate), respectively as non-supplemented or supplemented with different Iranian propolis (IP) extracts were used. The treatments were HC (control), HC + IP 25%, HC + IP 50%, HC + IP 75%, MC (control), MC + IP 25%, MC + IP 50% and MC + IP 75%, which means 25, 50 and 75 g of propolis in 100 mL ethanol 70%, respectively. The results showed that IP significantly increased gas production in IP 75% with different concentrate: forage ratios. Adding IP caused a decrease in pH, however this decrease was not significant in all treatments. There was a significant difference between the effect of HC + IP 50% on NH3-N compared to the HC + IP 25% and HC. Adding IP 75% significantly decreased CH4 production compared to the other treatments. In HC diet, there was no significant difference in total populations of protozoa and Ruminococcus albus between the individual IP 25%, 50%, 75% treatments, however when they were considered as the group, a significant difference was observed between them and the control group. The highest decrease and the highest increase in total populations of Prevotella bryantii was observed in IP 75% and in IP25%, respectively. In MC diet, methanogens were significantly reduced in IP 25%, 50% and 75%, compared to control treatment. The propolis extract caused improvement in fermentation and decreased methane and nitrogen ammonia. This may help the nitrogen retain longer in ruminants.

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