The Alleviative Effect Of Chicory Polysaccharides On The Non-alcoholic Fatty Liver Disease Induced By High-fat Diet In Rats

The aim of this study was to investigate the effect of chicory polysaccharides(extracted from the root of chicory)on nonalcoholic fatty liver disease in male rats,and to clarify the mechanism of this effect.The study consists of the following four parts:1 Extraction,purification and characteristic analysis of chicory polysaccharidesOptimal extraction means of chicory polysaccharide was selected via ultrasound assisted fibrin enzyme hydrolysis.The crude chicory polysaccharide was then purified and the properties of the purified chicory polysaccharide were studied.To study the effect of ultrasonic temperature,ultrasonic time,enzyme dosage,enzyme solution time and the liquid to solid ratio on chicory roots polysaccharide yield,quadratic regression models were established using central composite design.The extracted crude polysaccharide was deproteinized with four methods.The crude polysaccharide after deproteinization was purified by DEAE-52 cellulose ion exchange chromatography and Sephadex G-200 gel chromatography.The uniformity and molecular weight of purified polysaccharide were determined using high performance gel permeation chromatography(HPGPC).UV scan was performed and the protein content was also determined.The monosaccharide components of purified polysaccharides were analyzed with GC-MS.The structure of polysaccharide was identified using infrared spectroscopy.Experiment established that the optimal extraction conditions of polysaccharide from roots of Cichorium intybus L.were ultrasonic temperature 45?,ultrasonic time 35 min,enzyme dosage 1.6%,enzymolysis time 1.8 h and liquid-to-solid ratio of 44:1(mL/g).Under these conditions,the extraction yield of chicory roots polysaccharide was 46.75%,closed to the predicted yield of 47.71%.The results showed that the enzyme-sevag method had the best result on protein removal,the protein removal rate being 90.1%.Two polysaccharide fragments were obtained through a chromatography column and named CP-1 and CP-2 separately.The relative molecular weight of CP-1 is 8511.4 Da.CP-1 was determined to contain no protein and was mainly composed of sorbitol,glucose,fructose and saccharol.The molar ratio was 1.00:5.58:13.97:10.32.Infrared spectra showed that CP-1 was a heteropolysaccharide containing ?-glucoside bonds.The regression model was significant,could be used as the chicory roots polysaccharide extraction process of regression analysis and parameter optimization.CP-1 was a neutral heteropolysaccharide.2 The alleviative effect of chicory polysaccharides on high-fat diet induced non-alcoholic fatty liver disease in ratsIn this experiment,we study the alleviative effect of chicory polysaccharides on high-fat diet(HFD)induced non-alcoholic fatty liver disease(NAFLD)in rats.40 NAFLD rats were randomly divided into 5 groups and were treated for a duration of 8 weeks as follows:1)model group(HFD-feed,oral gavage of purified water),2)low dose group(HFD-feed,oral gavage of 50 mg/kg CP-1),3)middle does group(HFD-feed,oral gavage of 100 mg/kg CP-1),4)high dose group(HFD-feed,oral gavage of 200 mg/kg CP-1),5)drug group(HFD-feed,oral gavage of 50 mg/kg lovastatin).The rats in the control group were fed with a regular diet and were given oral gavage of purified water(RD+PW)for 8 weeks.8 weeks later,all rats were sacrificed,blood samples were collected and the levels of total cholesterol(TC),triglyceride(TG),low density lipoprotein cholesterol(LDL-C),high density lipoprotein cholesterol(HDL-C),malondialdehyde(MDA),superoxide dismutase(SOD),glucose(GLU),alanine aminotransferase(ALT),aspartate aminotransferase(AST),total bile acid(TBA),alkaline phosphatase(ALP)and lactate dehydrogenase(LDH)in the serum were determined.Liver organs were taken.The liver coefficient was determined.Also determined was the level of MDA,SOD,TC,TG and total antioxidant capacity(T-AOC).Meanwhile,the histopathological changes of the liver were observed.The results showed that CP-1 could significantly decrease body weight and liver index in NAFLD rats.CP-1 also significantly increased the levels of SOD and HDL-C and decreased the level of ALT,AST,TG,TC,LDL-C,GLU,TBA,ALP,LDH,and MDA in NAFLD rats' serum.In addition,CP-1 effectively decreased the level of MDA,TC,and TG and increased the level of SOD and T-AOC in the livers of NAFLD rats.The histopathological and ultra microstructure examination of the livers showed that CP-1 alleviated the pathological injury of NAFLD rats.In conclusion,CP-1 has significant alleviative effect on the high-fat diet induced NAFLD in rats.3 Chicory polysaccharide alleviates high-fat diet induced non-alcoholic fatty liver disease in rats via the AMPK signaling pathway This part studied the mechanism of chicory polysaccharide's alleviative effect on high-fat diet induced non-alcoholic fatty liver in rats.The mRNA expression level of lipidmetabolism related genes in rats' livers was determined by real-time fluorescence quantitative PCR(RT-PCR).These genes were peroxisome proliferator-activated receptora(PPARa),sterol regulatory element binding protein-lc(SREBP-lc),carnitine palmityl transferase I(CPT-1),fatty acid synthase(FAS),and steary 1-coenzyme A desaturase(SCD-l).The expression level of lipid-metabolism related proteins in rats5 livers was determined by Western Blot(WB).These proteins were adenylate activated protein kinase(AMPK),acetyl CoA carboxylase(ACC),phosphorylation level,adipose triacylglyceride lipase(ATGL),FAS,CPT-land SCD-1.The results showed that the CP-1could significantly up-regulate the mRNA expression level of PPARa and CPT-1 and down-regulate the mRNA expression level of SREBP-lc,FAS and SCD-1 in the liver of NAFLD rats.CP-1 could also up-regulate the protein expression of p-AMPK,p-ACC,ATGL and CPT-1 in the liver of NAPLD rats,and down-reguiate the protein expression of ACC,FAS and SCD-1.In conclusion,CP-1 alleviate HFD-induced NAFLD by activating the AMPK-related signaling pathway.4 Metabolomics study of chicory polysaccharide9 s alleviative effect on high-fat diet induced non-alcoholic fatty liver disease in rats Study the metabolomics of chicory polysaccharide alleviative effect on high-fat diet induced non-alcoholic fatty liver in rats.Liver metabolic markers in different groups of rats were detected using ultra high performance liquid tandem chromatography with quadruple time of flight mass spectrometry(UHPLC-QTOFMS).Principal component analysis(PCA)and orthogonal projections to latent structures-discriminant analysis(OPLS-DA)were used to identify the metabolic markers that showed different levels among groups of rats.The identified markers were annotated using Kyoto gene and Kyoto encyclopedia of genes and genomes(KEGG).The metabolic pathways of these metabolites were further analyzed using MetaboAnalyst.The results of PCA showed that,expect in low dose group,the metabolic spectrum dispersion in blank group,drug group,middle dose group and high dose group was all significantly different from that in model group.The results of OPLS-DA showed that there were significant differences between the blank group and the model group and between the high-dose group and the model group as well.487 metabolic markers were screened out between the blank group and the model group,among which223 metabolic markers were up-regulated and 264 metabolic markers were down-regulated.373 metabolic markers were screened out between the high-dose group and the model group,among which 157 were up-regulated and 216 were down-regulated.The pathway analysis showed that the metabolites which showed different levels between the blank group and the model group participated in 16 metabolic pathways,while those between the high-dose group and the model group participated in 13 metabolic pathways.Further studies showed that there were 7 key metabolic pathways involved with the differently leveled metabolites between the blank group and the model group,and 7 key metabolic pathways involved with the differently leveled metabolites between the high-dose group and the model group.Our findings thus provide a systematic theoretical basis to the understanding of the pathogenesis of NAFLD and to the use of CP-1 as a prevention and treatment for NAFLD.In conclusion,CP-1 can alleviate the NAFLD induced by high fat diet in rats,which provides a theoretical basis for the prevention and treatment of NAFLD.