| Objective: Based on "You Gu Wu Yun" theory of TCM drugs,with effective/poison duality of Mahuang Xixin Fuzi decoction(the MXF)as the research object,uses the normal and the composite model of joint application,the correspondence analysis model in mice,given different doses of the drug intervention,the MXF explore disease and drug dosage of the MXF effect/factors the influence of selective expression,clarifying "effect/disease and poison" relationship of objective authenticity and scientific connotation.Reference research methods of “syndrome and treatment pharmacokinetics”,the normal and the composite model of mice given the MXF after the intervention,the target effect/toxic ingredients in the study of the “time-concentration” change of the plasma,to discuss disease factors effect on the MXF target plasma pharmacokinetic/toxic ingredients,provide experimental basis for clinical rational drug use safety the MXF.Methods: 1.Using LC and LC-MS technology,to establish qualitative quantitative analysis method of typical chemical composition(ephedrine and pseudoephedrine hydrochloride,hydrochloric acid methyl ephedrine hydrochloride,low alcohol,methyl eugenol,sesame fat fat element,asarum,benzoyl new monkshood alkali,benzoyl monkshood alkali,benzoyl times monkshood alkali,the new aconitine,aconitine,aconitine)which has the duality of effective/poison in Mahuang Xixin Fuzi decoction.With the content of constituents in the decoction for indicators,the study investigates different decocting methods(the party-ephedra decoction method first decoct method,clinical decoction methods-decoction methods commonly used aconite decoction method first,lab-long the medicine decoction method)and specifications of different processed products of lateral root of aconite(gun attached sheet,light pills,black,white,fry with pills attached)impact on the quality of the MXF decoction.One-way analysis of variance(One-way ANOVA)and multivariate statistical analysis are used to evaluate the quality of the decoction,and PLS regression mathematical model is used to extract the representative components that could characterize the quality of the decoction.2.To copy as the composite model of kidney yang deficiency diseases mice,normal mice and mice kidney Yang deficiency diseases different doses of the MXF,to general growth,mortality,myocardial enzymes,liver function,kidney,Viscera index,respiratory metabolism monitoring,lung expression,tissue section as indicators of pharmacology study and observation of normal mice under different dose the MXF intervention and pharmacological indexes of kidney Yang deficiency diseases in mice,to explore the effect of disease syndrome and dose on the selective expression of MXF effect/toxicity.3.One-way analysis of variance and multivariate pattern recognition are used to comprehensively evaluate and analyze the results of pharmacological experiments,so as to explore the effect of MXF at different doses on the expression of toxic effects in normal and extrasensory mice with kidney yang deficiency.Based on normal mice and mice with kidney yang deficiency diseases pharmacological experimental results of PLS regression mathematical model is set up,respectively extract can characterize the MXF after intervention "poison" and "effective" express the representation of the pharmacological index,from tolerance differences,function target organ effect,dose-effect/toxicity relations,etc.comprehensive analysis of the MXF function under normal and disease states the difference of pharmacological effects,and clarify the MXF poison selective expression of objective existence.4.LC/QQQ technique is used to establish an analytical method for the target effective/toxicity components(ephedrine hydrochloride,pseudoephedrine hydrochloride,methylephedrine hydrochloride,benzoyl aconitine,benzoyl aconitine,benzoyl aconitine)of MXF in plasma biological samples.5.The study adopts single oral method,measures target of effect/toxic after giving the MXF/toxic ingredients in normal model,model of disease and the study of the "time –concentration” change of the plasma,by comparing effect/toxic ingredients in normal mice and mice with kidney yang deficiency diseases model of blood drug concentration in the body after the MXF intervention,pharmacokinetic parameters is obtained by using the statistical moment method,examine the body disease and state of the MXF target effect/toxic ingredients in the plasma pharmacokinetic characteristics.Result: 1.LC and LC-MS test methods are used to establish the chromatographic quantitative analysis method for the target effective/toxic components in MXF decoction,and the methodological verification is carried out.The results showed that the analytical method is accurate,reliable and repeatable,meeting the requirements for the content determination of the above components in the decoction.The traditional decoction time of aconite is the shortest,but the dissolution rate of MA and AC is the lowest,and the dissolution rate of mono-ester alkaloid is higher than that of aconite.Compared with the MXF decoction prepared by other aconite preparations,the content of double-ester alkaloids is the lowest,the content of single-ester alkaloids is the highest,and the content of ephedra alkaloids is the lowest.Double-ester alkaloids MA and AC can be used as representative components to characterize the preparation of MXF decoction by different decoction methods,and carcuanol,methyl eugenol,asarin,MA,HA and AC can be used as representative components to characterize the preparation of MXF decoction by different prepared products of aconite.2.Compared with the normal group,the body mass of each group is significantly decreased on the 3rd to 4th day after administration,and the "transient" increase of anal temperature was observed on the 1st day after administration.Myocardial enzyme profiles(CK,ck-mb,AST),liver function(AST)and renal function(urea)are significantly different in each group.Lung index,kidney index and testis index shows significant difference in some groups.The expression trend of Mc P-1 and il-12p70 in the lungs of each group is negatively correlated with the dose.3.After the success of the external sensation model of kidney-yang deficiency,mice in the model group dies on the third day,and their body mass and anal temperature shows an "inflection point" on the fourth day,and then begins to decline continuously.The myocardial enzyme profiles(CK,ck-mb,-hbdh,LDH,AST),liver function(ALT,AST,ALP)and renal function(urea,creatinine,cystatin C,2 microglobulin)of the model group are significantly different from those of the normal group.The indexes of lung,kidney,thymus,adrenal gland and seminal vesicle show significant difference.The value of energy H1 and H2 decreases significantly.The expression of il-6,il-10,Mc P-1,IFN-and TNF-cytokines show significant difference in the lung.Compared with mice in the model group,the behavior and mental state of mice with external sensation of kidney-yang deficiency are improved after different doses of MXF,and the body mass and anal temperature show an increasing trend of callback to different degrees.The above abnormal biochemical indexes of mice in the model group shows different degree of callback.Among the viscera index,the lung index show a significant decline,while the kidney index and thymus index show a different degree of correction.RER,H1 and H2 show a callback trend,and showed a significant positive correlation with the dose.The expressions of il-6,il-10,Mc P-1,IFN-and TNF-in the lungs decrease,and the expression of il-12p70 increase.No obvious pathological changes are found in the heart and kidney tissues of the normal group,model group and each drug administration group.Compared with the normal group,the lung tissue of the model group show obvious inflammatory injury,and the liver tissue show obvious water-like lesion.After the intervention of MXF with different dosages,the lesion situation can be improved to different degrees,with the group receiving 40 times as much improvement.In the model group,the red and white pith boundaries are blurred and the white pith volume increase.The improvement effect is not obvious in the 10-fold group,but is greater in the 30-fold and 40-fold groups.However,the white pith part of the spleen of the 60-fold group begins to show irregular changes.4.According to the normal mice and mice kidney Yang deficiency diseases pharmacological experimental results of PLS regression mathematical model is set up,to extract the characterization of the MXF for normal mice "poison" effect evaluation indexes: weight(3,4 days),anal temperature(1 day),myocardial enzymes(CK,CK-MB),liver(AST,ALT),kidney(urea),viscera index(liver,lung index),TSE(RER,H1,H2)were,lung cytokines,IL-12 p70(MCP-1);extract the characterization of the MXF mice with kidney Yang deficiency diseases "effect" effect evaluation indexes: weight(1,2,3,4,5,6,7 days),anal temperature(1,2,3,5 days),liver meritorious service is(alkaline phosphatase),viscera index(kidney index,the index of seminal vesicle gland),TSE(RER,H1,H2)are,lung cytokines(MCP-1,TNF alpha).5.The PLS regression mathematical model is established based on the results of normal mouse pharmacological experiment.The results of comprehensive pharmacological score predicted by each administration group are the lowest in the 30-fold administration group.The PLS regression mathematical model is established based on the results of the pharmacological experiment on mice with exogenous sense of kidney-yang deficiency.The results of the comprehensive pharmacological score predicted by each administration group shows that the 40 times administration group has the highest score of the comprehensive pharmacological effect.6.According to the guiding principle of quantitative analysis method for biological samples of FDA,the established analytical and testing method is systematically verified by methodology,and the established analytical method is accurate,reliable and repeatable,meeting the requirements of biological sample content determination.The pharmacokinetic parameters of three ephedrine alkaloids,E,PE,and ME,were significantly higher than that of the normal group in auc 0-t,auc 0-infinity,t1/2,MRT 0-t,and MRT 0-infinity,while the values of CLz/F were the opposite.The pharmacokinetic parameters of Tmax and Cmax of three annexed alkaloids(BMA,BAC and BHA)in the mice with exogenous sense of kidneyyang deficiency are significantly higher than those in the normal group.The drug-time curves of E,PE,ME,BMA,BAC and BHA 6 components to be tested in mice with external sensation of kidney-yang deficiency all showed obvious double peaks.Conclusion: 1.Different decocting methods and specifications of different processed products of the preparation of the lateral root of aconite decoction in effect the content of toxic components exist obvious difference,the party uses decoction method and the processed products specifications preparation of lateral root of aconite decoction,can not only reduce the ephedra alkaloids and double ester lateral root of aconite alkaloid content,in order to reduce the side effects of ephedra and lateral root,and monkshood monoester type alkaloid content highest instead,does it produce attenuated without failure,which confirmed by the parties select aconite decoction methods and the scientific nature of the processed products specifications,party in the past to provide the reference.2.Disease and syndrome factors can affect the selective expression of MXF effect/toxicity.After different doses of MXF were given to normal mice,the "toxicity" effect was selectively expressed as the dosage increased.After MXF was given to mice with external syndrome of kidney Yang deficiency,the "low" state of body function affected the metabolism and absorption process of dual chemical components in vivo,and then affected the body function and metabolism level of disease and syndrome,and on the contrary selectively expressed the effect of "effect".3.The "dose-toxicity" relationship of MXF in normal mice showed a significant positive correlation.The "dose-effect" relationship of MXF in the intervention of mice with exogenous kidney Yang deficiency was nonlinear.In this study,40 times of the dose-effect became the dose inflection point for the selective expression of "efficacy" and "toxicity" of MXF.4.The most obvious "time inflection point" of toxic and side effects of MXF on normal mice was 3-4 days after drug administration,and the survival "inflection point" was 3-4 days after virus inoculation of mice with kidney Yang deficiency,and the "inflection point" was 3-4 days after MXF was given to mice with kidney Yang deficiency.5.Normal mice in the 80-fold dose group all died on the day,and mice with exogenous renal Yang deficiency did not die on the day of administration,and all died on the second day of administration,suggesting that the syndrome state of exogenous renal Yang deficiency increased the tolerance of the body to toxic ingredients.6.E,PE and ME are well absorbed in the body state of kidney Yang deficiency,and their metabolic rate in the body is slowed down and bioavailability is improved.Absorption rates of BMA,BAC and BHA decreased in the body state of renal Yang deficiency,indicating that toxicity was reduced by delayed absorption. |
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