A One-time Exercise Combined With GC-MS Method To Explore The Metabolic Mechanism Of Epimedium Polysaccharides In Improving Chronic Fatigue Syndrome

ObjectiveThis study established an animal model of chronic fatigue syndrome(CFS),investigating the effects of Herbal Epimedii Polysaccharide(HEP)on the expression of CFS rats by metabolomics combined with exercise stimulation.And to explore the mechanism of the improvement of CFS,this study identified differential metabolites biomarkers and metabolic pathways after intervention providing experimental evidence for the prevention and alleviation of CFS.Methods1.The crude HEP polysaccharide was prepared by centrifugation,purification,alcohol extraction and Sevage deproteinization and etc.The purified HEP polysaccharide was purified by cellulose(DEAE-52)and dextran gel,and a series of physicochemical identification was done including:sugar content,heterozygous content,UV spectrum,infrared spectrum,monosaccharide composition analysis,amino acid content,environmental scanning electron microscopy,and atomic force electron microscopy.2.After adjusting,the rats were randomly divided into 5 groups:normal group,CFS group,CFS + HEP(50mg/kg)group,CFS + HEP(100mg/kg)group and CFS + HEP(200mg/kg)group.The CFS rat model was established using multi-factor:the tethered tube,runway,sleep deprivation,and crowded environment.The effects of different doses of HEP on CFS were evaluated by observing the changes of body weight,food intake,drinking water and behavioral experiment(water maze,open field,tailing test).The urinary metabolic markers induced by tested rats were all analyzed by GC-MS and GC-MS after a high intensive training.The potential biomarkers were found by the principal component analysis,orthogonal partial least squares.The biomarkers were identified and analyzed by the MetPA.Results1.Herba Epimedii was purified by DEAE-52 cellulose column and Sephadex G-150 column to obtain a polysaccharide HEP,whose total sugar content is of 89.5%(w/w),with 10.26%(w/w)galacturonic acid,and the relative molecular weight was 13.6 × 104Da,and the total protein content of HEP was 4.39%.And the HEP was composed of mannose,rhamnose,glucose,galactose,arabinose and galacturonic acid.While,it contains of 3.37mol glutamic acid,8.79mol cystine,0.46mol leucine,9.07mol tyrosine 0.36mol lysine,1.13mol histidine.The structure of HEP was observed by SEM and AFM.The results showed that HEP molecules consisted of randomly distributed single spherical particles.2.Compared with NC group,the data of CFS rats were significantly higher on body weight,daily food intake,water-intake,suspended test time of tailing test,upright times of open field experiment,and the time of latent period at first two days in water maze test.While the number of swimming through the quadrant of the platform was lower than NC group.And the CFS group were depressed irritable and aggressive,with loose skin and unshapen excrement.Compared with the CFS group,the appearance and behavioral indexes of the rats were significantly improvedof high dose of HEP group,while the low dose HEP group the indicators did not significantly improve.The results of metabolomics analysis showed that 10 biomarkers were identified in CFS group and NC group,which were malonic acid,fumaric acid,dimethyl fumarate,malic acid,a-ketoglutaric acid,Tartaric acid,gluconic acid,palmitic acid,sphingosine,21-hydroxy and pregnenolone.The result was found that CFS affect the TCA cycle,alanine,aspartic acid and glutamic acid metabolism,sphingolipid metabolism,the synthesis of steroid hormones and other metabolic pathways,which can cause energy metabolism disorders,immune dysfunction,andhypoactivity of HPA and other symptoms.Compared with CFS group,there are five significant differences in metabolites of high dose group of HEP:benzoic acid,mandelic acid,creatine,pimelic acid,and norepinephrineConclusionThe model of CFS rats was successfully established by using the tethered tube,runway,sleep deprivation,and crowded environment.The HEP has eased the CFS rats of their fatigue state and depression,improved spatial exploration and memory capacity,which indicating that HEP has good preventive effect on CFS.Through the metabolomics analysis,it was found that the symptoms of CFS may be caused by the body's energy metabolism disorders,immune dysfunction,hypoactivity of HPA and other causes,involving TCA cycle,alanine,aspartic acid and glutamic acid metabolism,sphingolipid metabolism,synthesis of steroid hormones and other metabolic pathways..It was found that HEP mainly affect the tyrosine metabolic pathway and arginine and proline metabolism metabolic pathway to improve the symptoms of CFS,which may be the mechanism of prevention of CFS.