| Objective:In this thesis,the normal rats,cold syndrome rats and heat syndrome rats were taken as research objects to confirm the nature characteristic of Semen Pharbitidis and its split compositions(polysaccharide composition,phenolic acid composition,resin glycoside composition and fatty oil composition)and the internal fine structure of Semen Pharbitidis’nature characteristic.Methods:1.Testing the content of pharmacological indexes,including DAS 5-HT,NE,AchE,17-OHCS-1,T3,T4,TSH,TRH,Na+K+-ATP enzyme,SDH,pyruvate acid,PDHα,hepatic glycogen,lactate,LDH,cAMP and cGMP,was used to study the changes of the nervous system,material metabolism,energy metabolism and endocrine system in the normal rats,cold syndrome rats and heat syndrome rats,that they were impacted by Semen Pharbitidis and its 4 split compositions.2.Combined with the pattern recognition method,ultra-performance liquid chromatography with quadrupole-time-of-flight mass spectrometry(UPLC-QTOF/MS)was used to study the changes of small molecule metabolites in the urine of the normal rats,cold syndrome rats and heat syndrome rats after the influence of Semen Pharbitidis and its 4 split compositions and to investigate the related metabolic pathway and its mechanism.3.Combined with the isobaric tag for relative and absolute quantitation(iTRAQ),the 2D-LCMS/MS analysis method was used to research the changes of protein expression in the liver tissue of 3 kinds of model rats after the influence of Semen Pharbitidis and its 4 split compositions,and to study the related metabolic pathway and its mechanism.4.The nextgeneration sequencing technology was applied to research the variations of mRNA expression in the liver tissue of 3 kinds of model rats after the influence of Semen Pharbitidis and its 4 split compositions,and to explore the related metabolic pathway and its mechanism.5.Using bioinformatics approach,the differential metabolites,proteins,mRNA in the 3 kinds of model rats after the influence of Semen Pharbitidis and its 4 split compositions were integrated and analyzed to further find out the prominent metabolic pathways.Combining the results of pharmacological evaluation,the the nature characteristic of Semen Pharbitidis and its split compositions were confirmed.Results:1.The Semen Pharbitidis decoction has cold characteristic,and it can reduce the conten of DA,AchE,T3,T4,Na+K+-ATP enzyme,SDH,pyruvate acid,lactate acid,LDH,cAMP in the 3 kinds of model rats.The polysaccharide composition and fatty oil composition have the similar effects to the decoction,they can reduce the conten of DA,AchE,T3,T4,Na+K+-ATP enzyme,SDH,pyruvate acid,PDHα,LDH,cAMP,however,the phenolic acid composition and the resin glycoside composition have no obvious effect.2.The Semen Pharbitidis and split compositions can significantly change the content of 29 metabolites in rat urine.So,Semen Pharbitidis,polysaccharide composition and fatty oil composition can affect the substance and energy metabolism in biological systems by restraining metabolism pathway such as glycerophospholipid metabolism,purine metabolism,pyrimidine metabolism,starch and sucrose metabolism,galactose metabolism,glycolysis or gluconeogenesis,citrate cycle(TCA cycle),amino sugar and nucleotide sugar metabolism,pyruvate metabolism,fructose and mannose metabolism etc.3.The Semen Pharbitidis and split compositions can significantly affect 25 protein spots that related to the energy metabolism and substances metabolism.So,Semen Pharbitidis,polysaccharide composition and fatty oil composition can affect the substance and energy metabolism in biological systems by restraining metabolism pathway such as glycolysis or gluconeogenesis,citrate cycle(TCA cycle),fatty acid degradation,pyruvate metabolism,PPAR signaling pathway,pentose phosphate pathway and steroid hormone biosynthesis etc.4.The Semen Pharbitidis and split compositions can significantly affect the mRNA that related to the energy metabolism and substances metabolism.So,Semen Pharbitidis,polysaccharide composition and fatty oil composition can affect the substance and energy metabolism in biological systems by promoting the biosynthesis of unsaturated fatty acids,and restraining metabolism pathway such as fatty acid degradation,glycerophospholipid metabolism,glycerolipid metabolism,PPAR signaling pathway,glycolysis or gluconeogenesis,purine metabolism,pyrimidine metabolism etc.5.After the influnce with Semen Pharbitidis,polysaccharide composition and fatty oil composition,the metabolic pathway of the normal rats,cold syndrome rats and heat syndrome rats were further enriched by integrating multinomial data.It showed that Semen Pharbitidis decoction can decrease the ATP generation and affect the use of substance and energy metabolism in biological systems by regulating metabolism pathway such as amino sugar and nucleotide sugar metabolism,glycolysis or gluconeogenesis,citrate cycle(TCA cycle),purine metabolism,pyrimidine metabolism,pyruvate metabolism,glycerophospholipid metabolism,steroid hormone biosynthesis,PPAR signaling pathway etc.Similarly,the polysaccharide composition and fatty oil composition can decrease the ATP generation and influence the use of substance and energy metabolism in biological systems by restraining glycerophospholipid metabolism,amino sugar and nucleotide sugar metabolism,glycolysis or gluconeogenesis,fatty acid degradation,citrate cycle(TCA cycle),purine metabolism,pyrimidine metabolism,pyruvate metabolism etc.Conclusion:The Semen Pharbitidis decoction and its composition can reduce the conten of DA,AchE,T3,T4,Na+K+-ATP enzyme,SDH,pyruvate acid,lactate acid,LDH,cAMP in the 3 kinds of model rats.The polysaccharide composition and fatty oil composition have the similar regulation effects to the decoction.On system biology,the Semen Pharbitidis and split compositions can significantly affect the metabolites,protein spots and mRNA that related to the energy metabolism and substances metabolism pathways.Semen Pharbitidis,polysaccharide composition and fatty oil composition can reduce the substance and energymetabolism in biological systems by promoting the amino sugar and nucleotide sugar metabolism,glycolysis or gluconeogenesis,citrate cycle(TCA cycle),purine metabolism,pyrimidine metabolism,pyruvate metabolism,glycerophospholipid metabolism,steroid hormone biosynthesis.In summary,the cold characteristic of Semen Pharbitidis decoction has been proved,Semen Pharbitidis,polysaccharide composition and fatty oil composition has cold characteristic,and the nature characteristic of phenolic acid composition and the resin glycoside composition are flat.The internal fine structure of Semen Pharbitidis’ nature characteristic is that,the decoction with " bitter,cold",polysaccharide composition with" bitter,cold",phenolic acid composition with "pungent,flat",resin glycoside composition with "pungent,flat",fatty oil composition with " bitter,cold".The traditional Chinese medicine and Chinese herbal resolving components can be effectively evaluated in normal rat model,heat syndrome rat model and cold syndrome rat model by the methods of the macrophysiological biochemical index system method and the methods of metabolomics,proteomics and transcriptomics.The results of physiological and biochemical indicators are consistent with the results of metabolomics,proteomics and transcriptomics,which confirm each other.It is fully demonstrated that these methods are scientific,reasonable,reproducible and feasible as methods for evaluating cold and hot medicines of Chinese medicines.The results of this study indicate that the application of macroscopic physiological and biochemical indicators or one of the three major groups alone,or the combined application of these methods can accurately appraise the cold and hot drug properties of traditional Chinese medicines.Therefore,the results of this study confirmed the scientificity and objectivity of the new hypothesis of the connotation of Chinese herbal(gas)flavor as proposed by the professor Haixue Kuang,and also provided the experimental basis for the traditional Chinese medicine for constructing Chinese medicine cold and hot medicinal property evaluation methods. |
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