| The purpose of this study was to determine the requirements of leucine,isoleucine and valine of juvenile tiger puffer(Takifugu rubripes)by growth experiments.The effects of three different levels of amino acids in the feed on the growth capacity,feed utilization capacity,body composition,muscle texture,amino acid content and enzyme activity in the metabolic process of experimental subjects were discussed,in order to provide evidence for the obtained demands.The effects of dietary branched amino acids(BCAA)on TOR and several amino acid transporters genes were investigated.This study consists of four parts:1.Nutrient requirement of isoleucine in juvenile tiger puffer(Takifugu rubripes)A 10-week feeding trial was conducted to evaluate the effects of dietary isoleucine level on growth performance,feed utilization,body composition and free amino acids contents of juvenile tiger puffer.Six isonitrogenous and isoenergetic diets were formulated to contain graded levels of isoleucine(1.23%,1.47%,1.61%,1.98%,2.17%,2.31%diet,respectively)from dietary ingredients and crystalline isoleucine.Juvenile tiger puffer initial body weight(29.00±0.01)g were randomly assigned to 18 tanks(35fish per tank and 3 replicate tanks per group in indoor aerated cages).The results indicated that protein productive values(PPV),body crude protein content of tiger puffer were significantly influenced by dietary isoleucine level and present first increased and then declined tendency.Higher PPV were obtained at dietary isoleucine levels of 1.98%and 2.17%compared to fish fed 1.23%isoleucine(P<0.05),and higher body protein contents were obtained at dietary isoleucine levels of 1.61%,1.98%and2.17%compared to fish fed 1.23%isoleucine(P<0.05).Free isoleucine contents in muscle decreased at first and climbed thereafter(P<0.05).The muscle free isoleucine contents declined first and then increased as the dietary isoleucine levels increased.There are no significant different appeared for growth and feed utilization parameters(P>0.05).Based on the quadratic regression analysis of the PPV and body crude protein content data,the optimum requirement of juvenile tiger puffer Takifugu rubripes for isoleucine was estimated to be in the range of 1.83%～1.96%dry diet.2.Effects of dietary valine on growth performance,biochemical indexes and relative gene expression of juvenile tiger puffer(Takifugu rubripes)The experimental diets were formulated with fixing graded levels crystalline valine into basal diets(1.09%,1.52%,2.05%,2.52%,3.14%and 3.63%dry weight,respectively).The juvenile tiger puffers(24.7±0.3g)were fed for 8 weeks.Results indicated that with the increase of dietary valine,weight gain rate(WGR),specific growth rate(SGR),feed efficiency(FE),protein efficiency ratio(PER)and protein productive value(PPV)showed a trend of first increasing and then decreasing,and reached the maximum value in 2.52%group.Dietary valine levels had a significant effect on Hepatosomatic index(HSI),condition factor(CF)and ash(P<0.05).HSI and CF climbed with the valine levels increased and ash showed a reverse tendency.With the valine levels increased,the valine contents in muscle showed a tend of increasing first and decreasing thereafter.The activities branched-chain amino acids aminotransferase(BCAT)in muscle and branched-chainα-keto acid dehydrogenase(BCKDH)complex in liver reached the peak values when the valine levels was 2.52%,and declined then.The activities of branched-chainα-keto acid dehydrogenase kinase(BCKDHK)in liver reached the maximum at the valine levels of 3.14%.Adequate valine down-regulated the expression of intestinal transporter genes B~0AT1,PAT1,y~+LAT2,EAAT3 and pep T1 significantly(P<0.05).On the basis of SGR,FE,PER,dietary valine requirements of juvenile tiger puffer were estimated to be 2.42%～2.90%of dry diet,corresponding to 5.41%～6.49%of dietary protein.3.Effects of dietary leucine on growth performance,amino acid contents and relative gene expression of juvenile tiger puffer(Takifugu rubripes)A 10-week feeding trial was conducted to evaluate the effects of dietary leucine level on growth performance,amino acid contents and relative gene influence of juvenile tiger puffer.Six isonitrogenous and isoenergetic diets were formulated to contain graded levels of leucine(1.93%、2.49%、2.86%、3.26%、3.63%、4.07%dry weight,respectively)from dietary ingredients and crystalline leucine.Juvenile tiger puffer initial body weight(28.97±0.02g)were randomly assigned to 18 tanks.Dietary leucine did not influence growth performance and feed parameters significantly(P>0.05).Dietary leucine did not influenced fish chemical composition,condition factor(CF),hepatosomatic index(HSI)and viscerosomatic index(VSI)(P>0.05).Free threonine,serine,glycine,valine,isoleucine,leucine,phenylalanine,lysine,histidine and proline contents in muscle increased significantly in 2.86%group(P<0.05).Free aspartic acid,valine,methionine,isoleucine and leucine contents in serum rose significantly in 4.07%group.Excessive dietary leucine inhibit the TOR-related genes and peptide transporter gene expression.In conclusion,although dietary leucine do not affect the growth performance of juvenile tiger puffer,dietary excessive leucine can increase free leucine in muscle and serum,and inhibit the TOR-related signaling genes expression.4.Effects of dietary imbalanced branched-chain amino acid on growth performance,quality,physiology indexes and relative gene expression in juvenile tiger puffer(Takifugu rubripes)The aim of this study was to investigate the dietary imbalanced branched-chain amino acid(BCAA)effects on tiger puffer.Five isonitrogenous and isoenergetic control diets were formulated with fish meal,peanut meal,gelatin,fish oil,soybean oil and lecithin and each amino acid of control diet reached the fish requirements.Five experimental diets were prepared by supplementing the control diet with a deficiency of leucine(Leu-L),an excess of leucine(Leu-H),an excess of isoleucine(Ile-H)and an excess of valine(Val-H).Juvenile tiger puffer(initial weight was 24.2±0.3g)were randomly assigned to 15 tanks.The results showed the leucine levels did not influence growth performance and feed utilization capacity significantly(P>0.05).Dietary imbalanced BCAAs significantly decreased ash contents(P<0.05).The condition factor(CF)and hepatosomatic index(HSI)significantly improved on Leu-H group(P<0.05).Dietary excess valine significantly decreased Hardness,resilience,gumminess and chewiness in muscle(P<0.05).The free isoleucine and valine contents in muscle and serum deceased with the dietary leucine level increasing.Branched-chainα-keto acid dehydrogenase(BCKDH)and branched-chainα-keto acid dehydrogenase kinase(BCKDHK)activities were influenced significantly in liver.Dietary excess BCAAs improved the transcriptional levels in muscle and down-regulated the intestinal transporter genes expression.In conclusion,dietary leucine have no significant effect on the growth of juvenile tiger puffer,and there is a high tolerance to leucine.Meanwhile,results of branched chain amino acid interaction shows the antagonism of branched chain amino acids is not obvious. |
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