Metabonomics Study Based On Efficacy Of Polysaccharides From Suanzaoren Decoction

Suan-Zao-Ren decoction(SZRD),a classical traditional Chinese formula containing five Chinese herbs Ziziphi Spinosae Semen,Poria,Chuanxiong Rhizoma,Anemarrhenae Rhizoma and Glycyrrhizae Radix Et Rhizoma,was originally described in Jin Kui Yao Lue,written in ancient China Eastern Han Dynasty by Zhongjing Zhang,which has been used for treatment of yin-deficiency,blood-deficiency and insomnia.Polysaccharides were important components in the Suanzaoren decoction.High performance liquid chromatography with mass spectrometry technology was applied to systemically reveal its efficacy mechanism.1.Effects of SZR decoctions with different extraction solvents on sleeping number,sleeping latency and time in mice induced by sodium pentobarbital were studied.As a result,30%alcohol was selected as the optimal extraction reagent.Furthermore,SZRD polysaccharides effects of molecular weight ultra-filtrated on sleeping number,sleeping latency time in mice induced by sodium pentobarbital were investigated.Finally,molecular weights below 30 KDa were active effects in SZRD polysaccharides.2.The physical and chemical methods induced yin-deficiency,blood-deficiency and insomnia model was established in this research.Blood routine,weight,signs,the brain and skin slices examinations were observed.Compared with control group,there were the pathological changes of weight,signs,blood routine,skin and brain tissues.As a result,two complex models were successfully established and improved after the treatment of polysaccharides from SZRD.It has some prevention and treatment on the yin-deficiency,blood-deficiency and insomnia rats.3.A metabolomics method for rat serum and urine based on high-resolution mass spectrometry was established to find the biomarkers related to insomnia due to yin-deficiency and blood-deficiency,and to explore the mechanism of the effect of Suanzaoren decoction polysaccharides on intervention of yin deficiency and blood deficiency insomnia.High performance liquid chromatography coupled with time-of-flight mass spectrometry was used to analyze the endogenous substances in biological samples.Principal component analysis and least-squares-discriminative analysis were used to identify the endogenous substances.The serum and urine samples of the control,model,and polysaccharides group were well grouped,indicating that the metabolites have changed after the modeling and polysaccharides administration.21 potential biomarkers were identified in serum and 15 potential biomarkers were identified in urine.The results showed that the levels of sphingolipids and acylcarnitine metabolites in the serum of the model group were up-regulated and the levels of vitamins and cholic acid metabolites were down-regulated in the serum;the levels of amino acids and vitamin metabolites were increased and the cholic acid and tricarboxylic acid cycle Metabolite levels were down-regulated in the urine.After oral administration of Suanzaoren decoction polysacchrides to rats,the levels of biomarkers showed different levels of callbacks.Further metabolic pathways identified by the potential biomarkers were analyzed,and a metabolic network that was related to the treatment of insomnia due-to yin deficiency and blood deficiency was constructed.The results of metabonomics demonstrated the pharmacological effects and the mechanism of Suanzaoren decoction polysaccharides on interfering yin-deficiency,blood-deficiency and insomnia.4.An ultra-fast liquid chromatography and tandem mass spectrometry method was established for simultaneous determination of Glu,GABA,Tyr,NE,Ach and Try in rat serum and brain samples.Serum and brain samples were subjected to simple protein precipitation and separated on an ODS-EP column.The established method has a good linear relationship in the corresponding concentration range,and the specificity,precision,accuracy,stability,extraction recovery and matrix effect were all in accordance with the requirements of biological sample analysis.The established methods were used to determine the concentrations of related substances in the control group,the model group,and the administration group.Compared with the control group,the levels of Glu,Tyr,NE,Ach were diversely up and GABA,Try in brain and serum samples.The concentrations of Glu,GABA,Tyr,NE,Ach,and Try in rat serum and brain samples all recovered and went toward the control group with the treatment of SZRD polysaccharides.It suggested that SZRD polysaccharides could inhibit sleep arousal and improve sleep quality.Using a corresponding score plot derived from partial least squares discriminate analysis,the profiles of brain and serum samples developed an obvious tendency of intra-cluster aggregation and showed clear separation between the control and the model groups,and the total polysaccharides administration groups.It further proved that Suanzaoren decoction polysaccharides had certain prevention of physical and chemical methods induced yin-deficiency,blood-deficiency and insomnia.