Study On Drug Interaction And Bio-active Components Of The Herb Pair Danggui And Honghua

In the former work of our laboratory,we had discovered that Danggui(DG)was usually combined with Honghua(HH)in clinical application,which also exerted synergistic effects.Thus,our research team applied and achieved the National Natural Science Foundation of China(81274058)-"Study on dose-effect relationship and synergistic interation of nourishing and activating blood effect for the compatibility of Danggui and Honghua" We want to answer the following scientific questions:(1)To demonstrate the range,properties and extent of the pharmacodynamic interaction of activating blood effects and tonifying effects of Danggui-Honghua(GH)herb pair;(2)To demonstrate the synergistic effect mechanisms of GH herb pair.This dissertation was totally divided into four major chapters and the results were as follows:Chapter 1.Literature research(1)Data analysis of series of herb pairs containing DG in the TCM clinic applicationWe retrieved 16529 formulae which contains DG from the "Dictionary of Traditional Chinese Medicine Formulae",then built the database to analyze the relationship among their proportion,corresponding major disease and its category.The results showed that there were 170 herbs(compatibility frequency>200)and even 22 herbs(compatibility frequency>1500).It revealed that their compatibility proportion "1:1" appears to be most after further analysis of the 22 herb pairs containing DG,then 1:2,2:1,3:2.In addition,they were most frequently used for the treatment of the diseases in internal medicine and gynecology.(2)A review of herb pairs containing DGA review of bio-active constituents and compatibility effects of herb pairs containing Danggui based on data analysis and latest research was given.(3)A review of GH herb pairA review was given about the latest research of DG and HH compatibility including phytochemistry,pharmacological effects and mechanism of action.Chapter 2.Research on the interaction of DG and HH compatibility(1)Study on promoting blood circulation effects of the herb pair containing DG and HH by integrated effect and response surface methodBased on the acute blood stasis rats model,the effects of DG and HH with different proportions(1:0,4:1,2:1,3:2,1:1,2:3,1:2,1:4,0:1 w/w)and different concentrations on hemorheology were observed.Then all indexes were integrated to the total activating blood circulation effect value by comprehensive index method.The interaction was analyzed by response surface method.Model parameters were estimated with nonlinear regression.The three-dimensional response surfaces were constructed with Matlab software.The results showed that the proportion from 0.8:1 to 1.1:1 of DG and HH had synergetic effects,and when the dose of DG ranged from 0 g to 5 g,and HH from 8 g to 11 g,the herb pair also had synergetic effects(the value was between-0.2 and-0.8).In addition,when the dose of DG ranged from 0 g to 0.2 g,and HH from 0.2 g to 1 g,the herb pair had antagonism action(the value was 0.2).(2)Study on nourishing and tonifying blood effects of the herb pair containing DG and HH by integrated effect and response surface methodBased on the blood deficiency rats model,the effects of DG and HH with different proportions(1:0,4:1,2:1,3:2,1:1,2:3,1:2,1:4,0:1)and different concentrations on the peripheral blood indexes and organ indexes were observed.Then all indexes were integrated and analyzed by relative equation.The three-dimensional response surfaces were gained by operating Matlab software.The results showed the best dose of DG and HH is 7.8 g/kg and 0.52 g/kg,respectively,when they were used singly.Most dose of GH exhibited addition action(50%),followed by antagonism action,and little dose(0.5?1.2 g/kg of DG and 0?0.5 g/kg of HH)had synergetic effect.The range of synergetic effect on tonifying blood of GH is little while the range of antagonism action is obvious.Chapter 3.Study on main bio-active components of GH compatibility in vivo and in vitro(1)Comparative analysis of the mian bio-active components of GH with different proportions by UHPLC-TQ/MSA highly sensitive and rapid UHPLC-TQ/MS method was established and validated to compare thirteen main bio-active components(6-hydroxykaempferol-3,6,7-tri-O-glucoside,hydroxysafflor yellow A,chlorogenic acid,6-hydroxykaempferol-3-O-rutinoside-6-O-glucoside,p-coumaric acid,anhydrosafflor yellow B,ferulic acid,kaempferol-3-O-rutinoside,senkyunolide I,senkyunolide A,3-n-butylphthalide,ligustilide,3-n-butylidenephthalide)of DG and HH with 9 different proportions(1:0,4:1,2:1,3:2,1:1,2:3,1:2,1:4,0:1).With the added amount of DG,the contents of the detected components of HH had a total increasing tendency.With the added amount of HH,the contents of the detected components of DG had a total decreasing tendency.(2)Pharmacokinetic characteristcs of the main bio-active components in normal and blood stasis rats after oral administration of GH by UHPLC-TQ/MSA highly sensitive and rapid UHPLC-TQ/MS method was established and validated to compare seven main bio-active components(hydroxysafflor yellow A,caffeic acid,p-coumaric acid,kaempferol-3-O-rutinoside,ferulic acid,3-n-butylphthalide and ligustilide)in normal and blood stasis rats after oral administration of GH extracts.It was found that the Cmax and AUC0-1 of tnese componets in blood stasis rats had increasing tenaency compared witn normal rats.And most components in model and normal rats had significant difference in some pharmacokinetic parameters,suggesting that the rate and extent of drug metabolism were altered in blood stasis rats.(3)UFLC-Q-TOF/MS based screening and identification of the metabolites in plasma,bile,urine and feces of normal and blood stasis rats after oral administration of hydroxysafflor yellow AA UFLC-Q-TOF/MS method was established and successfully applied to the detection and identification of the metabolites in bile,urine,plasma and feces samples of normal and model rats with orally administrated HSYA.A total of 8 metabolites were observed in normal rats,while 7 metabolites were detected in model rats.The distribution of metabolites in the plasma,bile,urine and feces of normal and model rats had obvious differences.The major in vivo metabolic pathways for HSYA included hydroxylation,hydroxylation + methylation,acetylation and glucuronidation,and there were also dehydration,hydrogenation,hydration,and hydroxylation + glucuronidation.All of these metabolites were reported for the first time,and these results are valuable and important for the understanding of the metabolic process and therapeutic mechanism of HSYA and some other pigments in HH.Chapter 4.Study on activating blood action mechanism of GH herb pair in blood stasis rats based on metabolomics strategyActivating blood circulation effects of GH and its combination herbs were evaluated by testing hemorheological parameters indexes.Fifteen potential biomarkers including 8-isoprostaglandin F2a?3-methoxy-4-hydroxyphenylglycol glucuronide,indoleacrylic acid,LysoPC(22:6(4Z,7Z,10Z,13Z,16Z,19Z)),LysoPC(16:0),LysoPC(18:0),L-palmitoylcarnitine,dihydroceramide in positive mode,and 3-dehydro-L-gulonate,dihydrouracil,taurohyocholate,deoxycholic acid,LysoPC(P-18:1(9Z)),LysoPC(O-18:0),psychosine sulfate in negative mode were tentatively identified.Compared with control group,L-palmitoylcarnitine,taurohyocholate,deoxycholic acid,LysoPC(O-18:0)and indoleacrylic acid in blood stasis rats decreased significantly while other biomarkers increased significantly.They primarily involved in pentose and glucuronate interconversions,ether lipid metabolism,pantothenate and CoA biosynthesis,?-alanine metabolism,sphingolipid metabolism,starch and sucrose metabolism,glycerophospholipid metabolism,fatty acid metabolism,tryptophan metabolism,pyrimidine metabolism.DG,HH and GH had potential pharmacological effects by regulating multiple perturbed pathways close to normal state.The compatibility of DG and HH had synergetic activating blood effect because the potential metabolites after GH intervention were closer to the control group than the two single herbs.