| Methane is a loss of feed energy for ruminants, and a pollution resource for atmosphere environment. So, the more attention were highlighted for modulating ruminal fermentation, reducing ruminal methane emission and prompting the ruminants performance. Saponin, as one of plant extracts, can inhibit the growth of protozoa and reduce the methane emission. The common interesting were attracted for exploitation of new natural plant saponin for reducing methane emission and mechanisms of reducing methane were investigated. The effects of gypenosides with dammarane structural triterpenes saponin were selected to investigate the effects on in vitro and in vivo methane production of goat rumen. Effects of gypenosides on community of rumen microbe and mechanisms of decreased methane were analysed by the shifts of community of microbes when saponin was usded to reduce methane. The reasons of reducing methane with short time effect were investigated by the experiment of microbial resistance to gypenosides. The theoretical evidences were explained for regulating ruminal fermentation and reducing methane emission.1 Effects of gypenosides on in vitro fermentation and fermentation kinetic parameters by mixed ruminal microorganismsEffects of Gypenosides on rumen methane prodcution were investigated by in vitro gas technique using mixed rumen microorganisms from goats as inoculum. The study includes two experiments. Experiment 1 was investigated to estimate its effects on fermentation characteristics of rumen microorganisms. Experiment 2 was conducted to assay its effects on fermentation kinetic parameters. The diet consisted of Leymus chinensis (70%), ground corn grain (21%) and soybean meal (9%)(1 mm screen). Five different gypenosides doses were used for each compound:0,5,10,20 and 40 mg/60 ml of the total culture medium. The results showed that as compared with the control, methane production and concentration were reduced by gypenosides addition (p<0.05) and there was a linear effects on methane production between control and treatments. The hydrogen utilization was lower than control at 10mg level. The TVFA concentration, acetate, propionate and butyrate acids proportion of TVFA were significantly increased (P<0.05) at 10mg level, and there was a significant quadratic effects on acetate, butyrate, isobutyrate, valerate, isovalerate, BCP proportion, acetate to propionate ratio and TVFA concentration with the gypenosides doses. There was a linear effect on butyrate with gypenosides addition. The protozoa counts for treatment groups decreased with a significantly linear and quadratic effects (P<0.05). Microbial protein was not changed abruptly, but with a increasing trend. Ammonia nitrogen concentration was significantly higher than control at highest saponin level and there was a linear effect on ammonia concentration with gypenosides addition. The theoretic and observed gas production decreased at high saponin level with a significant linear and quadratic effects. There was a significant linear effect on rate of gas production with saponin doses increasing. Above results indicated that gypenosides addition could modify the microorganisms fermentation pattern, reduce protozoa numbers and methane emission, abate pollution of methane derived from ruminants to environment. The VFA production and feed conversion efficiency were promoted.2 Effects of gypenosides on rumen fermentation, microorganisms population and blood parameters in goatsFour Boer crossbred with local goat wether male goats (30±2.3 kg) were used in a 4×4 Latin square experiment to determine effects of gypenosides from Gynostemma Pentaphyllum Makino on ruminal fermentation characteristics and ruminal microbial community. The goats fed a 70%Leymus chinensis:30%concentrate (corn:soybean meal=2:1) were intraruminally given with suspensioned gypenosides at 0800 and 1700 daily. Doses were 0 (control),2,4 and 8g/d.head-1. Ruminal contents were sampled at 0(immediately prior to feeding),2,4 and 8 h after feeding in the morning. The results showed that ruminal ammonium concentration in gypenosides treatments decreased (P=0.057), while there was no significant effect on microbial crude protein, methane emission and hydrogen recovery (P>0.05). But, there was a decreasing trend for methane and a increasing rate of hydrogen utilization Acetate, BCVFA concentration and ratio of acetate to propionate decreased slightly, whereas feed intake (low and high dose), propionate concentration increased than control group (P>0.05). There were no differences for blood parameters such as blood serum glucose, TG, TC, BUN and TP concentration between control and treatments (P>0.05). The numbers of total protozoa, Entodinium spp. and Diplodinium spp. in rumen with gypenosides addition were significantly lower (P<0.05) than those in control group, with a dose-dependent manner. Total genomic DNA were extracted from ruminal microbes, and populations of rumen microbes were determined by a real-time PCR. Populations of total methanogens, anaerobic fungi, Ruminococcus flavefaciens and Fibrobacter succinogenes were expressed as a proportion of total ruminal bacterial 16S rDNA. The number of methanogens sharply reduced. While fungi, fibrolytic bacteria percentage of total bacteria in rumen markedly increased at 2 g/dh-1. The numbers of all microbial groups decreased obviously at high dose of gypenosides addition. DGGE was used to monitor the shifts of protozoal, methanogenic and bacterial population after gypenosides treatment. The similarity varied from 48%to 75%for protozoa,66 to 87%for methanogens and 40 to 82%for bacteria. The community assay of methanogens from seven protozoal species found that there were difference for the different protozoa. The shift of methanogenic population from single protozoal species was observed before and after treatment with gypenosides. The common dominant methanogen in all protozoa collected was Methanobrevibacter sp. after treat or untreat by gypenosides. These results suggest that the gypenosides supplementation could influence on the rumen fermentation, and reduce methanogens, total protozoal and single protozoal species number Gypenosides could promote the growth of two fibrolytic bacteria and fungi, change the microorganism community and the methanogens community associated with protozoa. 3 The study of ruminal microbial resistance to gypenosidesConsecutive batch cultures, involving six serial transfer were established to investigate the effects of gypenosides on ruminal microbial fermentation properties, and the communities of bacteria and methanogens from rumen digesta. The results showed that pH value increased after gypenosides treatment. Compared with the control group, the gas production of different gypenosides groups in different transfers was low excepted for the 5mg and 10mg groups of first transfer with higher gas production. Gas production of some treatment groups decreased with significant linear and quadratic effects. There was a significant reducing with linear and quadratic effects between methane production and gypenosides doses (P<0.05 or P<0.01). Ammonia nitrogen concentration of treatment were higher than that of control. MCP concentration of most of treatment groups decreased than that of control excepted for first transfer with gypenosides increasing. There were significant decrease for some treatments and different transfers(P<0.05). Protozoal numbers reduced significantly with a linear effect for first transfer and second transfer after treatment by gypenosides. Protozoa were not detected under microscopy for third transfer. There were significantly decreasing for TVFA, acetate, propionate and butyrate concentration after gypenosides addition. The shifts of communities of bacteria and methanogen were observed after treatment. Some predominant bands such as Butyrivibrio fibrisolvens and Fibrobacter succinogenes were gradually disappeared and a few new predominant bands such as Erwinia alni and uncultured bacterium were observed after treatment by gypenosides. These data indicated that the growth of some microbes were inhibited, and others bacteria with capability of adaptation to gypenosides and degradation of gypenosides became the predominant bacteria in fermentation systems. The results suggested that gypenosides affected the ruminal microbial fermentation. In turn, the community of bacteria were gradually formed to adapt to the survival environment with gypenosides.4 Effects of gypenosides on fungi co-culture with methanogens methane production and fermentation characteristicsThis study were conducted to investigate the effects of gypenosides on methane production and fermentation property under fungi co-culture methanogens (Ⅰ) and fungi as dominant species (Ⅱ). The formation of two culture system of fungi associated with methanogens and fungi as main species were treated by penicilline & streptomycin (I) and chloramphenicol (Ⅱ), respectively. The results showed that, in system I after treatment by gypenosdies, there were significantly linearly decrease for methane production and methanogenic numbers (P<0.01). Gas production and fungi numbers were siginificantly reduced with a linear or quadratic effects (P<0.01). pH value increased with significant quadratic effect (P<0.05). TVFA and acetate concentration decreased significantly with a significant quadratic effect. In system II after treatment by gypenosides, methane production were not detected for minor production. Gas production decreased with a linear or quadratic effects (P<0.01). Fungi and methanogens numbers were significantly reduced with a linear effect. TVFA (P<0.05) and acetate concentration (p<0.01) decreased with linear or quadratic effects. pH value increased (P<0.05) with linear or quadratic effects (P<0.01). These results indicated that the pH value increasing, gas and methane production decreasing were induced by the decreased numbers of fungi or methanogens, and inhibitation of fungi and methanogenic activity. Growth of fungi and methanogens in two fermentation systems were depressed by gypenosides. |
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