Inhibition of rumen methanogenesis induced by Bioflavex® and its pure flavonoid components under in vitro fermentation using rumen fluid from steers fed high concentrate diets

Abstract Text: Four separated incubation series were performed in a complete randomized blocks design to determine the effect of a citrus extract rich in flavonoids (Bioflavex^®) and its main components on methane mitigation under in vitro condition. Rumen liquor from four rumen cannulated growing steers fed a high concentrate diet (90:10 commercial concentrate:barley straw) was used as inoculum. Bottles of 120 ml were prepared under the CO₂ stream and filled with incubation solution and a mixture of concentrate:barley straw (600:60 mg/bottle) was used as substrate. Bioflavex^® (BF) was added to the incubation media and its effect compared against its pure flavonoid components (Hesperidine [HS]; Isonaringine [IN]; Naringine [NG]; Neoeriocitrine [NE]; Neohesperidine [NH] and Poncirine [PC]) at 200µg/g dry matter (DM), and the substrate without flavonoids was also included as a control (CTR). Bottles were sealed and incubated at 39±1^◦C for 72 h. The head space pressure was measured at intervals of 2 hours, started from 2 to 12 h then 24, 48 and 72 h. Values (mbar) converted to volume (ml) by a linear regression. Since 12 h post incubation, a sample (0.1 ml) from head space gas was analyzed for methane concentration using GC. The pattern of cumulative gas production (y) was fitted to the model: y=a(1-e^-b(t-c)),  being a the potential cumulative gas production (ml); b the production rate (ml/h) and c is the lag time (h). The addition of BF or any of its components reduced the cumulative gas (P<0.01) and methane production (mmol/g DM) (P<0.01) except for NE and PC, that did not differ from the CTR values. However, reduction in CH₄ production was more pronounced than it was in gas production (P<0.01). No changes were observed in the gas (ml/h) and methane production rate (mmol/h) in relation to CTR. The addition of flavonoids in the in vitro culture media reduced gas production and it would reflect the activity of bioflavex and its main components against the fermentative activity of the rumen liquor although our result also showed specific activity against methanogenic archaea.

Keywords: in vitro incubation, Methanogenesis and Pure flavonoids