Effects of extracts of Perilla frutescens (seeds) on in vitro rumen fermentation, methanogenesis and microbial population

Abstract Text: Seeds of Perilla frutescens are rich in linolenic and linoleic acid. An in vitro incubation was performed to investigate the effects of extracts from Perilla frutescens seeds on rumen fermentation, methanogenesis and microbial population. Five-hundred milligrams of substrate (mixture of Chinese wild rye and corn meals at 70:30, w/w) were incubated in 120 ml serum bottles. The fermentation medium was 5 ml of sheep rumen fluid and 45 ml of buffer medium. The seeds of Perilla frutescens were extracted by 70% ethanol. To accurately add 20 mg of extracts, the freeze-dried extracts were dissolved into dimethyl sulfoxide (DMSO) to a concentration of 40 mg/ml, and then 500 ml of the DMSO solution was added to the incubation system. In the control, 500ul of DMSO was used without extracts. After 24 h of incubation at 39°C, rumen fermentation parameters were measured and absolute abundance of total bacteria, protozoa, fungi and methanogens were quantified using real-time PCR based on the 16S, 18S rDNA and mcrA gene, respectively. Total volatile fatty acids and ammonia nitrogen concentrations were not affected (P>0.05) by extracts, while methane production was decreased by 63.6% (P<0.01), from 0.723 to 0.263 mmol/g DM substrate. Addition of extracts induced a dramatic shift in the population of rumen microbota (Table 1), indicating the decreased protozoa (99.5%, P<0.01) and fungi numbers (99.5%, P<0.01) and reduced methanogens (62.2%, P<0.01). These data indicate that ethanol extracts from seeds of Perilla frutescens have potential to reduce rumen methanogenesis by inhibiting protozoa and activity of methanogens without adversely affecting rumen fermentation.

Table 1. Effects of extracts of Perilla frutescens(seeds) on microbial population (copies/g centrifuged fermentation mixtures)

	Control	Extracts	SEM	P-value
Total bacteria (×1011)	7.12	6.81	0.472	>0.05
Protozoa (×107)	666.9	3.4	98.7	<0.01
Fungi (×105)	1684.3	8.3	239.5	<0.01
Methanogens (×108)	11.8	4.5	1.39	<0.01

Keywords: methanogenesis, Perilla frutescens, rumen fermentation