Effects Of Fermented Soybean Meal On Carbon And Nitrogen Metabolism In The Large Intestine Of Piglets And Its Mechanism

This thesis aimed to study the mechanism of fermented soybean meal(FSM)promoted piglet growth performance from the perspective of microflora and its metabolism in the large intestine,then provided the theoretical basis for the application of FSM and the development of biological feed in piglet.Changes of the product composition of FSM under different fermented condition were studied,and the FSM was provided for the following feeding experiment.Effects of wet-fermented soybean meal(WFSM)and dry-fermented soybean meal(DFSM)on the fermentation,microflora structure and metabolism in the large intestine of piglets were studied.Then the effects of FSM on the fermentation parameters after it changed the microflora structure was confirmed by in vitro fermentation.Finally,the continuation effect of FSM on the ammonia production from piglet manure was studied by gas production experiment.This study included six parts,the main results were as follows:Part one: research on fermentation condition and components of soybean meal.Two experiments were conducted to study the changes of mannan oligosaccharides and ?-glucan content at different temperatures.First experiment with different microbial additions consisted of 8 treatments from combinations of three microorganisms(Bacillus subtilis KC 101,Saccharomyces cerevisiae JM 102 and Bacillus lactis RG 103).Fermentation broth was mixed with soybean meal in 18 L bucket incubated under normal room temperature(averaged 21 °C)for 10 days.Based on the results of first experiment,the highest level combination of the three microorganisms was selected and incubated with soybean meal in 3 L beaker at 25,30 and 35 °C for 7 days in the second experiment.Results of the first experiment showed that Bacillus subtilis and Bacillus lactis didn'taffect the production of Saccharomyces cerevisiae fermentation products when used for soybean meal fermentation.Results of the second experiment showed that 35 °C was more conducive to the accumulation of soybean meal fermentation products compared with 25 and 30 °C.The quantity of Bacillus subtilis,Saccharomyces cerevisiae and Bacillus lactis,as well as the content of the fermentation products tended to be stable at 35 °C until day 4.Therefore,FSM at 35 °C for 4 day was chosen as the material for the following feeding experiment.Part two: effects of FSM on fermentation parameters in the large intestine of piglets.This experiment was conducted to research the supplementing 7.33 % WFSM and 5 %DFSM(equivalent on a dry matter basis)in the diet on fermentation parameters in the large intestine of piglets.A total of seventy-five 41 days old Duroc × Landrace × Yorkshire piglets with an initial body weight of 13.14 ± 0.22 kg were used in a four-week feeding trial.At the end of the feeding trial,one pig from each pen was randomly selected for sampling of large intestinal contents.The results showed that the average daily gain(ADG)of WFSM and DFSM groups increased by 27.08 and 14.58 %,the feed conversion ratio(FCR)improved by 18.18 and7.27 %,butyric acid concentration increased by 38.35 and 17.10 %,while valeric acid concentration decreased by 32.00 and 17.58 %,isovaleric acid concentration decreased by32.24 and 18.25 %,respectively,compared with the control group.Therefore,FSM could improve carbohydrate fermentation and reduce protein fermentation in the large intestine of piglets.Part three: effects of FSM on microflora structure in the large intestine of piglets.To better understand the effects of FSM on microflora structure in the large intestine of piglets,second generation Illumina MiSeq technology was used to investigate the difference of bacterial component,and analysis the predictive functional profiling of microbial communities in the different part of large intestine from part two.Based on the ordination of distance matrix,clear demarcation were observed between both the WFSM and DFSM with the control,indicatd that WFSM and DFSM intervened the microbial communities in the large intestinal of piglets.The results showed that FSMincreased the microbial diversity in large intestine of piglets,increased the relative abundance of butyrate-producing bacteria,reduced the relative abundance of amino acid fermenting bacteria,improved the relative abundance of carbohydrate metabolism function families,reduced the relative abundance of amino acid metabolism function families,and the effects of WFSM was higher than DFSM.Part four: effects of FSM on changes of metabolites in the large intestine of piglets.To better understand the effects of FSM on metabolites in the large intestine of piglets,metabonomics technology was used to compare the differences of biochemical composition contents in different part of large intestine from part two.Qualitative and quantitative analysis for metabolites were conducted by using multivariate statistical analysis methods.The OPLS-DA model,which was established using multivariate data analysis,was used to excavate and identify a total of 135 significant discriminant metabolites(P<0.05).There were a certain degree of separation among the three treatments in each part of large intestine,indicating that metabolites could be affected by both WFSM and DFSM in the large intestine of piglets.Combined with metabolic pathway tracing,these results obtained using this model revealed that WFSM and DFSM increased the levels of carbohydrate-related metabolites,such as 4-aminobutanoate,lactic acid,mannitol,threitol and ?-alanine,while decreased the levels of those related to protein catabolism,such as1,3-diaminopropane,creatine,glycine and inosine.DFSM mainly improve carbohydrate metabolism in the large intestine of piglets through 4-aminobutanoate pathway,while WFSM could convert protein metabolism to carbohydrate metabolism in the large intestine of piglets through 4-aminobutanoate pathway,?-alanine pathway and 5-aminopentanoate pathway at the same time.Part five: effects of FSM on fermentation parameters in the large intestine of piglets with in vitro intestinal model.In order to verify the effects of changes in the large intestine microflora on fermentation parameters,the contents of large intestine from part two were used as the bacteria source,and the fermentation was carried out at 39 °C for 24 h with in vitro intestinal model.The results showed that the butyric acid concentration in large intestine of WFSM and DFSM groups increased by 18.39 and 15.15 %,valeric acid concentration decreased by15.63 and 8.33 %,respectively,compared with the control group.And these results were consistent with part two.In addition,the ammonia production of WFSM and DFSM groups decreased by 29.68% and 17.73%,respectively,compared with the control group.Therefore,FSM could improve carbohydrate fermentation and reduce protein fermentation by changing the microflora structure in the large intestine of piglets.Part six: effects of FSM on ammonia emissions from piglet manure.In order to study the continuation effect of FSM changed microflora and its metabolism on the ammonia production from piglet manure,the manure of piglets from part two was collected to a composting chamber to determine the ammonia production for 7days.The results showed that the ammonia production from piglet manure in WFSM group was significantly lower than that in DFSM group(P<0.05),and the ammonia production of manure in DFSM group was significantly lower than that in control group(P<0.05).The ammonia production from manure in WFSM group and DFSM group decreased by 33.46%and 13.39% in the first day,decreased by 23.53% and 13.26% in 7 days,respectively,compared with the control group.Therefore,adding FSM on the diet had a continuation effect on the reduction of ammonia production from piglet manure.In conclusion:(1)The results of feeding experiment and in vitro intestinal model showed that feeding diets supplemented with FSM could improve the carbohydrate metabolism and reduced the protein metabolism by improving the relative abundance of carbohydrate metabolism function families and reducing the relative abundance of amino acid metabolism function families,and WFSM was more effective than DFSM.(2)Through the metabonomic analysis found that DFSM improved carbohydrate metabolism through 4-aminobutanoate pathway,while WFSM converted protein metabolism to carbohydrate metabolism in the large intestine of piglets through4-aminobutanoate pathway,?-alanine pathway and 5-aminopentanoate pathway at the sametime.(3)Adding FSM on the diet had a continuation effect on the reduction of ammonia production from piglet manure.