| This study was conducted to investigate the effects of glycinin and β-conglycinin on growth performance,gene express of intestinal healthy(tight junction protein,apoptosis signal,inflammatory cytokines and signaling pathways)and metabolomics of Songpu mirror carp,to investigate the effect mechanism of glycinin and β-conglycinin on the intestinal health of fish.In this study,soybean protein utilization and intestinal health were studied from a new perspective of intestinal metabolic signal pathway,and the alleviating mechanism of dietary AKG supplementation on the antinutritional effect of soybean antigen protein was tentatively explored,which has important theoretical value for improving the application of soybean in aquatic products.The first chapter was aimed to study the effects of glycinin and β-conglycinin on growth performance of Songpu mirror carp.A total of 1080 juvenile carps(4±0.5g)were randomly divided into 12 groups with 3 replicates,and 30 fish in each replicate.4different levels of glycinin(4%,8%,12% and 16%)as 11S1,11S2,11S3 and 11S4,4different levels of β-conglycinin(4%,8%,12% and 16%)as 7S1,7S2,7S3 and 7S4,fish meal group(CON),50% soybean meal as soybean meal group(SBM),glycinin & β-conglycinin(The same amount of purified glycinin and β-conglycinin as 50% soybean meal)as 11+7S group,added 1%AKG base on 11+7S group as AKG group,glycinin,β-conglycinin and soybean meal were added respectively as the substitutes of fish meal in every group diet.The results shows that the specific growth rate(SGR),weight growth rate(WGR)and protein efficiency(PE)of SBM group were significantly decreased(P<0.05),the feed conversion rate(FCR)of soybean meal group was significantly increased(P<0.05),the performance of others weren’t significant different with soybean meal group(P>0.05).The Condition factor hasn’t changed in every group.As the level of glycinin increased,the crude protein contents of body was decreased first and then increased,in which,the crude protein of 11S1 group was significantly lower than soybean meal group(P<0.05),the crude protein of 11+7S group,AKG group and different levels of β-conglycinin haven’t significantly changed.As the β-conglycinin increased,the crude lipid contents of body decreased,other groups haven’t significantly changed.As the level of β-conglycinin increased,the water contents of body was showed a downward trend,other group haven’t significantly changed(P>0.05).In the second chapter,RT-PCR technology was used to test the gene expression in each sectional intestine of common carp,such as signal pathway genes(AMPK/ ACC and TOR/4E-BP),apoptosis signaling pathway genes(caspase3 and caspase9),tight junction protein genes(occluding,claudin3 c,claudin11 and claudin7)and Inflammatory cytokines genes(TNF-a,IL-1β and TGFβ1).The results show that with the increasing of glycinin and β-conglycinin,the gene expressions of ACC and TOR were gradually decreased in MI(middle intestine)and DI(distal intestine),the expression of apoptotic signaling pathway genes was increased in each intestine of common carp.The expression of proinflammatory cytokines increased first and then decreased in each intestine,in which DI was the most serious section,what’s more the gene expression levels of two important tight junction proteins(claudin-3c and occludin)were decreased in DI.In the third chapter,LC-QS untargeted metabolomics technology was used to study the effect of glycinin,β-conglycinin and AKG on intestinal mucosal metabolome of carp.Pairwise comparison was made among CON,11S3,7S3,11+7S and AKG.The results shows that there were 69 different known metabolites and two metabolic pathways(glycerophospholipid metabolism and choline metabolism)changed among 11S3 and CON group,54 different known metabolites and 7 metabolic pathways(apoptosis,sphingolipid signaling pathway,sphingolipid metabolism,arginine biosynthesis,linoleic acid metabolism,glycerophospholipid metabolism and choline metabolism)changed among 7S3 and CON group,69 different known metabolites and 5 metabolic pathways(PPAR signaling pathway,apoptosis,sphingolipid metabolism,linoleic acid metabolism,glycerophospholipid metabolism)changed among 11+7S and CON group,51 different known metabolites and three metabolic pathways(choline metabolism in cancer,arginine biosynthesis,pantothenate and Co A biosynthesis,glycerophospholipid metabolism and pyrimidine metabolism)changed among 11S3 and CON group.In summary:1.Supplementation of 11 S and 7S with different levels(0～16%)did not significantly affect the growth performance of carp,while affected the nutrient content of body components of carps.2.With the glycinin and β-conglycinin increased,the expression of ACC and TOR decreased in MI and DI,the expression of inflammatory factors in each segment showed a trend of increased first and then decreased,the expression of apoptosis in each segment were increased,the expression of tight junction genes were decreased in DI.In conclusion,glycinin and β-conglycinin caused damage to the fish intestine.3.Glycinin and β-conglycinin increased the consumption of lipids and phenylpropanoids and activated glycerol phospholipid metabolism in the intestinal mucosa of carp to provide energy for the intestine.4.Alpha-ketoglutarate(AKG)can alleviate the damage of glycinin and β-conglycinin to the intestine of fish and recover genes expression of intestinal healthy,in addition,it increased the metabolic pathway of lipids to provide energy. |
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