Quality Markers Study Of Gingseng-Fuzi Prescriptions

The lateral root of Aconitum carmichaelii Debx,a plant of Ranunculaceae,is named Fuzi,and is commonly used clinically for the treatment of chronic rheumatism,chronic heart failure,and chronic renal failure.However,this herbs often trigger Aconitine poisoning events due to their relatively narrow therapeutic window,resulting in significant regional loss of property and life.Ginseng,the root and rhizome of Panax ginseng C.A.Mey.,is often found in prescriptions together with Fuzi,called Ginseng-Fuzi herb pair,which can play a role in reducing the toxicity and increasing the effects of Fuzi.At present,there are certain researches based on the pharmacological activity,content determination,pharmacokinetic behavior,and quality markers of Ginseng-Fuzi herb pair at home and abroad.However,there is still a gap in the bioactive equivalent combinatorial components linking potency/toxicity.Compared to the Shenfu injections,oral drugs are simple to administer and easily accepted by patients,and have a promising market prospect in the treatment of chronic heart failure.However,the safety dose window for orally administered drugs,such as Shenfu Qiangxin Pills,is generally relatively narrow and does not effectively balance cardiac efficacy and cardiotoxicity.The safety risks results in low market share in the field,which also impacts new drug applications such as Shenfu Dropping Pills.Aiming at this subject,the group proposed a new strategy named"toxicity reducing and efficacy increasing research of oral Ginseng-Fuzi prescription based on the new model of integrated bioavailability equivalence",including in vitro screening of"toxicity reducing and efficacy increasing"substances,in vivo comparative pharmacokinetic/toxicokinetic study,screening of cardiotonic superior/equivalent combinatorial components,cardiotonic pharmacological confirmation,and development of new oral drugs against heart failure.As part of the above project,this topic is intended to guide the screening of cardiotonic superior/equivalent combinatorial components through two aspects:the content/ratio and the in vivo pharmacokinetic characteristics,to assist the development of safe and effective new oral drugs.This project investigated the content of the key components and in vivo pharmacokinetics of Ginseng-Fuzi medicines and achieved the following results.1.An HPLC-MS quantitative method was developed for the quantification of 12 aconitines（Aconitine,Mesaconitine,Hypaconitine,Benzoylaconine,Benzoylmesaconine,Benzoylhypaconine,Aconine,Mesaconine,Hypaconine,Songorine,indaconitine,Yunaconitine）and 26ginsenosides（Ginsenoside Rg1,Ginsenoside F5,Ginsenoside F3,Ginsenoside Rh1,Protopanaxatriol,Panaxatriol,Ginsenoside CK,Ginsenoside Rh2,Protopanoxadiol,Panoxadiol,Ginsenoside Re,Ginsenoside Ro,Ginsenoside Rf,Ginsenoside Rb1,Pseudoginsenoside F11,Ginsenoside Rc,Ginsenoside Rg2,Ginsenoside Rb2,5,6-Dehydro ginsenoside Rd,Ginsenoside Rd,Ginsenoside Rd2,Ginsenoside Rg6,Ginsenoside F4,Ginsenoside Rg3,Ginsenoside Rk1,Ginsenoside Rg5）in Chinese patent medicines.The method was tested for linearity,precision,and accuracy,and met the requirements of the analytical methods specified in the Chinese Pharmacopoeia.2.The method was used to determine the contents of 38 key components in 10 Gingseng-Fuzi Chinese patent medicines（Xinbao Pill,Renshen Zaizao Pill,Wumei Pill,Nvbao Capsule,Pingfeng Shengmai Capsule,Shenfu Qiangxin Pill,Jianbu Qiangshen Pill,Shengui Lizhong Pill,Qili Qiangxin Capsule,Nanbao Capsule）for the evaluation of safety and efficacy.The results showed that the contents of aconitines（total alkaloid contents were 1.09—267.07μg/g）and ginsenosides（total ginsenosides concent were 167.1—126404.55μg/g）were significantly various among different drugs.The contents of the diester type alkaloids ranged from 0.10–3.94μg/g,while the contents of the monoester type alkaloids and nonester type alkaloids ranged from0.82–225.90μg/g and 0.09-22.55μg/g,respectively.Besides,the contents of ginsenosides in Xinbao Pill（total ginsenosides concent was 126404.55μg/g）,Pingfeng Shengmai Capsule（total ginsenosides concent was 11305.05μg/g）,and Qili Qiangxin Capsule（total ginsenosides concent was 3256.91μg/g）were significantly higher than those of other medicines（total ginsenosides concent were 167.1—2109.24μg/g）.3.An HPLC-MS quantitative method was developed for the quantification of 12 aconitines in rat plasma.The method was tested to meet the requirements of pharmacokinetic analysis methods in terms of linearity,precision,and accuracy.4.The contents of the 12 aconitines in the plasma of SD rats which were orally administered with equal doses of Fuzi and Ginseng-Fuzi（in the raw material amount of Fuzi）for 24 h were determined,and it was found that the absorption and metabolism of the 12 aconitine alkaloids were fast(T_1/2 were 3.42—124.13 h,T_maxwere 0.50—1.50 h)in SD rats,and the elimination rate of the diester type alkaloids(T_1/2 were 3.42—5.20 h)was higher than that of the monoester type alkaloids(T_1/2 were4.65—9.28 h).For some diester type alkaloids such as AT and MAT,the Ginseng-Fuzi group(T_1/2 were 6.62—89.71 h,AUC_0-t were 0.46—0.63 ng/m L·min)was significantly slower in metabolism the elimination phases than the Fuzi group(T_1/2 were 5.20—21.74 h,AUC_0-t were 0.31—0.33 ng/m L·min);for the monoester type alkaloids such as BAC,BMA,and BHA,the metabolism of the Ginseng-Fuzi group(T_1/2 were 32.55—80.39 h,AUC_0-t were 4.42—14.55 ng/m L·min)was similarly slower than that of the other group(T_1/2 were 4.65—9.28 h,AUC_0-t were 3.11—7.56 ng/m L·min),which predicted that the increase in efficacy was also existed in vivo after ginseng was used as a compatible material.5.Five potential efficacy markers of Ginseng-Fuzi medicine were screened:diester type alkaloid AT,MAT and monoester type alkaloids BAC,BMA,BHA.In summary,this project combined in vitro content determination and in vivo comparative pharmacokinetic study to evaluate the quantitative analysis based on chemical composition of Ginseng-Fuzi Chinese potent medicines,and to evaluate the pharmacokinetic comparative study of Ginseng-Fuzi medicines in rats orally administered with Fuzi and Ginseng-Fuzi decoction.The results of both experiments were synthesized and five potential quality markers of Ginseng-Fuzi medicines were screened.The aim is to achieve clinical effecacy increase,and to lay the foundation for scientific design of the prescription design,dosage,and controlled medicative combinatorial components of Shenfu oral administration.