Study On The Grading Method And Processing Mechanism Of Prepared Slices Of Curcumae Radix

Curcumae Radix,as a typical multi-source traditional Chinese medicine(TCM),is derived from the dried radix of Curcuma wenyujin,Curcuma longa,Curcuma kwangsiensis or Curcuma phaeocaulis.With the support of the National Project for standardization of Chinese Materia Medica and other projects,this paper took the deficiency of grading method of prepared slices of Curcumae Radix as the breakthrough point,deeply explored the grading method of prepared slices of Curcumae Radix,and provided the research basis for the development of prepared slices of Curcumae Radix with high quality,effective and controllable quality.Took the different efficacy of prepared slices of Curcumae Radix as the breakthrough point,combined with traditional medication method,water decoction,the rat model of blood stasis due to liver-Qi depression was established to study the scientificity of Curcumae Radix stir-frying with vinegar or wine,and the processing mechanism of stir-frying with vinegar into liver to strengthen the effect of dispersing stagnated hepatoqi to stop pain and stir-frying with wine strengthen the effect of promoting blood circulation for removing blood stasis.Objective:Through literature,market and clinical research,traditional grading method and modern scientific connotation integrated to establish the grading method of prepared slices of Curcumae Radix,which can control the quality of grading method of prepared slices of Curcumae Radix and ensure the clinical efficacy,and provide reference for the development of high-quality grading method of prepared slices of Curcumae Radix and grading method for prepared slices of multi-source TCM.By comparing the differences of chemical constituents of raw Curcumae Radix,Curcumae Radix stir-frying with vinegar and Curcumae Radix stir-frying with wine,as well as their pharmacodynamic indexes related to disperse stagnated hepatoqi to stop pain and promote blood circulation for removing blood stasis in rats with bloodstasis due to liver-Qi depression,and the differences of endogenous metabolites in plasma,the scientificity of stir-frying with vinegar or wine of Curcumae Radix and their processing mechanism were discussed,which provided experimental basis for the clinical selection of appropriate processed products in TCM.Method:1.Through the literature and market research of Curcumae Radix,we could understand its medication history,processing methods and intentions,quality evaluation system status,etc.At the same time,169 batches of prepared slices of Curcumae Radix were collected.Referring to literature,clinical,traditional experience and grading methods of raw medicinal materials,the internal components of prepared slices of Curcumae Radix were analyzed with modern technology,and the grading method of prepared slices of Curcumae Radix was established,which was unified with traditional and modern technology.2.Based on traditional form of TCM,the change of extracting rate and content of characteristic components of different processed products of Curcumae Radix were compared,UPLC-QTOF/MS and GC-MS were used to study the influence of different processing methods on the chemical composition of Curcumae Radix and its changing regularity.3.Combined closely with the difference of the effect of Curcumae Radix“stir-frying with vinegar into liver to strengthen the effect of dispersing stagnated hepatoqi to stop pain and stir-frying with wine strengthen the effect of promoting blood circulation for removing blood stasis”,the pharmacodynamic indexes related to disperse stagnated hepatoqi to stop pain and promote blood circulation for removing blood stasis in rats with blood stasis due to liver-Qi depression were investigated by means of hematoxylin-eosin staining(HE),Enzyme-Linked ImmunoSorbent Assay(ELISA),Western Blot(WB),Quantitative Real-time PCR and flow cytometer,revealing the processing mechanism of Curcumae Radix from the aspect of pharmacodynamics.4.Processing mechanism of Curcumae Radix based on metabonomics,GC-MS was used to study the changes of plasma metabolites in rats with blood stasis due to liver-Qi depression.PCA and PLS-DA were used to analyze and screen potential biomarkers.The effects of different processed products of Curcumae Radix on potential biomarkers were discussed.Metaboanalyst was used to construct and analyze related metabolic pathways.Result:1.Curcuma longa Radix(Huangsiyujin,HSYJ)has genuine medicinal material with the best quality.Processing methods are diverse,mainly cutting,stir-frying with vinegar or wine,but also some areas use stir-frying without adjuvant material.This study combines the clinical efficacy of TCM,refers to the grading method of medicinal materials,establishes the traditional grading method of prepared slices of Curcumae Radix on the basis of defining the origin and processing methods.At the same time,on the basis of fully respecting and inheriting the traditional grading method,adds modern evaluation index(alcohol-soluble extract,volatile oil,curcumins,germacrone),divides prepared slices of Curcumae Radix into wholesale and selection.Selected prepared slices of Curcumae Radix are divided into three levels(superior,first and second).Formation of traditional and modern combination of prepared slices of Curcumae Radix grading method.2.After being stir-fried with vinegar or wine,the extract yield and curcumins content of water decoction of HSYJ increased.UPLC-Q-TOF-MS analysis showed that the content of curcumins,13 degradation products and amide in processed products increased significantly.Stir-frying HSYJ with vinegar was more obvious at vanillyl glycol,3-(4-hydroxyphenyl)propane-1,2-diol,4,5-dihydroxy-6-(4-hydroxy-3-methoxyphenyl)-3-oxohexanoic acid and3-(3,4-dimethoxyphenyl)propane-1,2-diol and stir-frying HSYJ with wine was more obvious at 4,5-dihydroxy-4-methylcyclohex-2-en-1-yl)hexan-3-one.However,the content of 4-(4-hydroxy-3-methoxyphenyl)-3-butene-2-one and bisacuronesdecreased significantly after processing.GC-MS analysis showed that there were 78 components in HSYJ and its processed products,19 components were added after stir-frying with vinegar,13 components disappeared,and 11 unique components;16components were added after stir-frying with wine,14 components disappeared,and 8unique components.Among the volatile oils of HSYJ and its processed products,11 potential markers were identified by PCA and OPLS-DA.3.The rat model was successfully reproduced by compound factors.The results showed that:(1)Organ changes: The organ coefficient of brain and kidney increased significantly(P < 0.05),the organ coefficient of liver,heart and lung increased significantly(P < 0.01).After administration,the raw HSYJ group(SHG)had a significant effect on the organ coefficient,and the stir-frying HSYJ with vinegar group(CHG)and the stir-frying HSYJ with wine group(JHG)had a significant effect on the organ coefficient of liver(P < 0.01).The heart-brain ratio and lung-brain ratio the model rats increased significantly(P < 0.01),there was a callback effect after administration,but there was no significant difference.Obvious pulmonary and hepatic blood stasis or ecchymosis could be seen in the of model rats,and pathological examination showed different degrees of liver injury and brain injury.HSYJ and its processed products could inhibit liver injury,but there was no significant change in brain injury.There was no significant difference between administration groups.(2)Liver function test showed that the liver of model group had functional damage,SHG had significant effects on DBiL,TBiL and TBA,CHG had significant effects on ALT,DBiL,TBiL and TBA,JHG had significant effects on DBiL and TBA.Compared with SHG,CHG could significantly reduce the level of ALT.(3)The content of ?-EP in plasma and brain tissue of model rats decreased significantly(P < 0.01).SHG and JHG had significant effects on ?-EP in brain tissue,CHG had significant effects on ?-EP in plasma and brain tissue,and JHG had significant difference compared with SHG.Immunohistochemical method showed that the content of c-fos protein in brain tissue of model rats increased significantly(P< 0.01).SHG,CHG and JHG all had significant effects(P < 0.01),but there was no significant difference between administration groups.(4)GAS,MLT,E2,T3 and T4 increased(P < 0.01),while Cor,TSH and NA decreased(P < 0.01).SHG has significant effects on E2,T3,T4 and NA in plasma.CHG had significant effects on MLT,E2,T3,T4,TSH and NA in plasma,and JSH had significant effects on T3,T4 and TSH,while there was no significant difference between administration groups.(5)vWF,ET-1,TXA 2,TXB 2,5-HT,cAMP,cGMP,CD62 p,Epo and EpoR of model rats increased significantly(P < 0.05 or P < 0.01),while NO,NOS,PGI 2,6-keto-PGF-1?,PGI 2/TXA 2 and 6-keto-PGF-1?/TXB 2 decreased significantly(P <0.01).SHG had significant effects on vWF,NOS,5-HT,Epo and EpoR,CHG had significant effects on NOS,5-HT,Epo and EpoR,and JHG had significant effects on NO,NOS,5-HT,PGI 2,PGI 2/TXA 2,6-keto-PGF 1?/TXB 2,CD62 p,Epo and EpoR,while there was no significant difference between administration groups.4.Sixteen compounds related to the rat model of blood stasis due to liver-Qi depression were screened out.Stearic acid and doconexent in model group decreased significantly,and L-alanine,lactic acid,glycolic acid,L-leucine,(R)-3-Hydroxybutyric acid,L-valine,L-threonine,glycine,malic acid,L-phenylalanine,citric acid,myo-inositol,arachidonic acid and cholesterol increased significantly.They involved the Valine,leucine and isoleucine biosynthesis,Phenylalanine,tyrosine and tryptophan biosynthesis,Glyoxylate and dicarboxylate metabolism,Phenylalanine metabolism,Glycine,serine and threonine metabolism,Arachidonic acid and Inositol phosphate metabolism.After administration,it was significant effects on L-alanine,lactic acid,(R)-3-Hydroxybutyric acid,citric acid,arachidonic acid and cholesterol.Among them,raw HSYJ had significant effects on arachidonic acid only.After stir-frying with wine,it was significant effects on lactic acid,citric acid and arachidonic acid.After stir-frying with vinegar,it was significant effects on lactic acid and arachidonic acid.Compared with SHG,stir-frying with wine has significant differences in citric acid.Conclusion:1.Traditional grading method of prepared slices of Curcumae Radix is based on single source,which was not conducive to highlighting high-quality.In this study,a comprehensive collection of prepared slices of Curcumae Radix,traditional and modern combination,clinical efficacy and quality-oriented,established a grading method of prepared slices of Curcumae Radix which was different from the existing grade according to basic sources,to provide technical support for high-quality,low-cost and rational clinical dispensing of prepared slices of Curcumae Radix,promote the development of high-quality prepared slices of Curcumae Radix,ensure the safety and effectiveness of clinical medication,and also provide an idea for the grading method of multi-source TCM.2.Through the analysis of extract rate,index component content,integral chemical composition and volatile oil composition of different processed products of Curcumae Radix,the material differences were revealed,and the material basis of Curcumae Radix “stir-frying with vinegar into liver to strengthen the effect of dispersing stagnated hepatoqi to stop pain and stir-frying with wine strengthen the effect of promoting blood circulation for removing blood stasis” was discussed.3.HSYJ was a good effect on liver function damage,neuroendocrine-immune network disorder,vascular endothelial function damage,platelet activation and agglutination caused by blood stasis due to liver-Qi depression,which proved that Curcumae Radix was good at soothing liver and regulate Qi to resolve depression and promoting blood circulation and removing blood stasis to relieve pain.After being stir-fried with vinegar could enhance the regulation of ALT to improve hepatocyte injury,enhance the analgesic effect by promoting the release and blood entry of ?-EP,inhibiting the expression of c-fos,and enhance the improvement of gastrointestinal hormones and endocrine.After being stir-fried with wine,the stability of internal environment could be maintained by increasing the release of ?-EP,inhibiting the expression of c-fos,promoting NO and PGI 2,dilating blood vessels,inhibiting platelet activation and aggregation,improving the abnormalities of PGI 2/TXA 2 and6-keto-PGF 1?/TXB 2,so as to strengthen the effect of promoting blood circulation for removing blood stasis.It was proved the processing theory of Curcumae Radix“stir-frying with vinegar into liver to strengthen the effect of dispersing stagnated hepatoqi to stop pain and stir-frying with wine strengthen the effect of promoting blood circulation for removing blood stasis”.4.The disorders of amino acid metabolism,lipid metabolism,energy metabolism,arachidonic acid metabolism and steroid biosynthesis in rats with blood stasis due to liver-Qi depression.HSYJ could interfered with amino acid metabolism,energy metabolism,lipid metabolism,arachidonic acid metabolism and steroid biosynthesis by regulating different compounds,especially L-alanine,lactic acid,3-hydroxybutyric acid,citric acid,arachidonic acid and cholesterol.The effect of stir-frying with vinegar on L-alanine,lactic acid and cholesterol was better,and the effect of stir-frying with wine on lactic acid,citric acid and arachidonic acid.The processing mechanism of Curcumae Radix was explained based on metabonomics.