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Objective: To establish a biological potency assay for Xiaojin Pills against platelet aggregation in vitro, evaluate the quality consistency of Xiaojin Pills, and screen traditioanal Chinese medicines which play the role of promoting blood circulation in Xiaojin Pills. Methods: Xiaojin Pills and ten Chinese medicines [artificial musk, Momordica cochinchinensis, Aconitum kusnezoffii, Liquidambar formosana, Boswellia carterii, Commiphora myrrha, Faeces Trogopterori, Angelica sinensis, Pheretima aspergillum, Fragrant Ink] in its formula were extracted by ultrasound in 40% methanol. The antiplatelet aggregation rate of the extract was measured by platelet aggregation meter. The platelet-rich plasma (PRP) and platelet-poor plasma (PPP) were prepared from abdominal aorta of rats. The platelet aggregation was induced by adenosine diphosphate (ADP). With sodium ferulate as a standard reference material, the biological potency of antiplatelet aggregation of Xiaojin Pills was calculated by the simplified probit principle. Results: The results showed that the biological potency of Xiaojin Pills was between 0.598 and 1.338 U/mg among different manufacturers and batches. In Xiaojin Pills group, Pheretima, Faeces Trogopterori, and Momordicae Semen had stronger inhibitory effects on platelet aggregation with inhibition rates of 70.87%, 31.83% and 67.52%, respectively. Conclusion: The quality consistency of Xiaojin Pills from different manufacturers and batches is poor, and Pheretima, Faeces Trogopterori, and Momordicae Semen may be the key drugs for Xiaojin Pills to play the role of promoting blood circulation.
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OBJECTIVE： To investigate the best compatibility proportion of Mongolian medicine “Terminalia chebula decomposing the poison of Aconitum kusnezoffii”. METHODS： Totally 40 rats were randomly divided into blank control group （0.05％ CMC-Na）， raw A. kusnezoffii group （0.12 g/kg by crude drug） and raw A. kusnezoffii-T. chebula （1 ∶ 3，1 ∶ 1，3 ∶ 1， mass ratio） group （0.12 g/kg raw A. kusnezoffii by raw material）， 8 rats in each group. They were given relevant medicine intragastrically once a day， for consecutive 28 d. 0.5 h after last medication， serum contents of cTn-I and MB were determined， and the changes of cardiological structure were observed； the detoxification effects of T. chebula on cardiotoxicity induced by A. kusnezoffii were investigated. Binding rates of 3 kinds of aconitine （concentrations of aconitine， mesaconitine and hypaconitine were 5， 10， 20， 40， 80， 160， 400 ng/mL） to binding rate of plasma protein with normal human plasma were determined by equilibrium dialysis combined with liquid chromtography-mass spectrometry. The concentrations of 3 kinds of aconitine were fixed as 100 ng/mL. After adding main detoxification component tannic acid in different proportions of T. chebula （1 ∶ 0.025， 1 ∶ 0.075， 1 ∶ 0.1， 1 ∶ 0.5）， the effects of them on plasma protein binding rate of 3 kinds of aconitine were investigated； the possible mechanism of T. chebula decomposing the poison of A. kusnezoffii inducing cardiotoxicity were investigated. RESULTS： In detoxification experiment， compared with blank control group， serum contents of cTn-I and MB were increased significantly in raw A. kusnezoffii group （P＜0.05）. There were obvious pathological changes in myocardial tissue， such as disorder of cell arrangement， cell shrinkage and cytoplasm staining. Compared with raw A. kusnezoffii group， serum contents of cTn-I and MB in raw A. kusnezoffii-T. chebula （1 ∶ 3， 1 ∶ 1， 3 ∶ 1） groups were decreased significantly （P＜0.05）， and there was no significant difference between the raw A. kusnezoffii-T. chebula （3 ∶ 1， 1 ∶ 1） groups and blank control group （P＞0.05）； myocardial pathological changes were improved to varying degrees. The structure of myocardial tissue in raw A. kusnezoffii-T. chebula （3 ∶ 1） groups were similar to blank control group. In the mechanism investigation experiment under the condition of different concentrations， plasma protein binding rates of 3 kinds of aconitine with normal human plasma were about 30％， without statistical significance （P＞0.05） and significant concentration-dependent manner. After adding tannic acid， plasma protein binding rate of 3 kinds of aconitine in A. kusnezoffii was decreased to different extent； when 3 kinds of aconitine were combined with tannic acid in the ratio of 1 ∶ 0.1， 1 ∶ 0.075 and 1 ∶ 0.5， the plasma protein binding rate decreased significantly （P＜0.05）， in proportion-dependent manner. CONCLUSIONS： Compatible use of raw A. kusnezoffii-T. chebula （3 ∶ 1） show that best detoxification effect on A. kusnezoffii-induced cardiotoxicity. Under this compatibility ratio， the plasma protein binding rate of 3 kinds of aconitine in A. kusnezoffii with normal human plasma is relatively high and the free drug concentration is relatively low.
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Rat model of blood stasis syndrome was prepared by subcutaneous injecting of epinephrine hydrochlorid,then the model rats were administrated by Yunnan Baiyao for 15 days. Blood rheology,coagulation function and histopathology were chosen as indicators to evaluate the successful replication of blood stasis syndrome model and the treatment effect of Yunnan Baiyao. UPLC-Q-TOF-MS was used to rapidly analyze the serum samples of blood stasis syndrome rat after 15 days Yunnan Baiyao treatment,Progenesis QI software was employed to identify the alkaloids components. The results showed that Yunnan Baiyao reduced the plasma viscosity and whole blood viscosity of rats with blood stasis syndrome,prolonged thrombin and prothrombin time,reduced fibrinogen content,and effectively improved pathological state such as inflammatory cell infiltration,blood stasis,congestion and edema of various organs in rats with blood stasis syndrome. Seven alkaloids components from Aconitum kusnezoffii,including karacolidine,senbusine B,isotalatizidine,karakoline,denudatine,talatisamine and chasmanine were found in the rat serum after Yunnan Baiyao treatment. Based on the effectiveness of Yunnan Baiyao in the treatment of blood stasis syndrome induced by epinephrine hydrochloride in rats,alkaloids components from the root of A. kusnezoffii absorbed into blood after Yunnan Baiyao treatment were clarified rapidly and accurately with the help of UPLC-Q-TOF-MS. Karacolidine,senbusine B,isotalatizidine,karakoline,denudatine,talatisamine and chasmanine are the pharmacodynamic material basis of the root of A. kusnezoffii for activating blood circulation and removing blood stasis.
Subject(s)
Animals , Rats , Aconitum , Chemistry , Blood Circulation , Blood Viscosity , Drugs, Chinese Herbal , Pharmacology , Prothrombin Time , Thrombin TimeABSTRACT
This present study is to develop an HPLC method for simultaneous determination of four diester diterpenoid alkaloids, beiwutine, mesaconitine, hypaconitine and aconitine in the leaves of Aconitum kusnezoffii, so as to provide evidence of the quality control of this herb. The four constituents were measured on a Waters XBridge CC₁₈ column(4.6 mmχ250 mm, 5 μm). The mobile phase was acetonitrile-40 mmol·L⁻¹ ammonium acetate solution(adjusted pH to 10.5 with ammonia solution)(33:67) with isocratic elution at a flow rate of 1.0 mL·min⁻¹; the detection wavelength was 235 nm; the column temperature was 30 °C, and the injection volume was 10 μL. Next, this contents of the four diester diterpenoid alkaloids in 12 samples were 0.025 5-0.088 5, 0.039 1-0.071 5, 0.026 6-0.081 0 and 0.008 12-0.031 2 mg·g⁻¹, respectively. Next, this method has been successfully applied to the analysis of A. kusnezoffii folium in different harvest periods. The contents of the four alkaloids decreased primarily, and then increased with the postponing of harvest. The established method is proved to be accurate and sensitive for the determination of alkaloids in A. kusnezoffii folium, and may be useful for the quality improvement of this herbal medicine. Moreover, these results indicated the scientific significance for the toxicity and the suitable harvest time of this herb.
Subject(s)
Aconitine , Aconitum , Chemistry , Chromatography, High Pressure Liquid , Diterpene Alkaloids , Drugs, Chinese Herbal , Chemistry , Phytochemicals , Plant Leaves , Chemistry , Plants, Medicinal , ChemistryABSTRACT
The radix,leaf,flower and bud of raw medicinal materials and extraction of total alkaloids of Aconitum kusnezoffii Reichb.were all involved in this investigation.All the compositions from the samples were analyzed through fourier transform infrared spectroscopy (FTIR) combined with second derivative IR spectroscopy and two-dimensional IR correlation spectroscopy (2D-IR).It was found that the spectra of raw medicinal materials showed that the radix of A.kusnezoffii Reichb.featuring a large quantity of starch was the same as starch with the characteristic peaks at 1,155,1,070 and 1,019.The leaf,flower and bud contained the similar aromatic hydrocarbons (1,600),glycosides (1,050-1,070),while lipids were not clear.The characteristic peaks of the buds,flowers and leaves were all at 1,595 cm-1 (vibration of phenyl framework) and 1,262 cm-1 (=C-O).Therefore,it was suggested that the common compound of the three parts be diterpenoid alkaloids.Second derivative IR spectroscopy showed that the characteristic peaks of radix was stronger than those of the flower,leaf and bud at 1,712 cm-1 (C=O),which proved that the quantity of characteristic peaks in the radix was larger than those in the flower,leaf and bud.In addition,six autopeaks at 1,745,1,650,1,560 (the most strong),1,465,1,400,1,300 were detected from the radix.The similar autopeaks at 1,745,1,650,1,560 (the most strong),1,465,1,400,1,300 were found in the leaf,bud and flower.In conclusion,it was demonstrated that the macro-fingerprint infrared spectroscopic identification method provided a large quantity of the comprehensive information and entirely grasped the quality of A.kusnezoffii Reichb.Besides,FTIR and 2D-IR provided massive information of the integral structures of the radix,leaf,flower and bud of A.kusnezoffii Reichb.and verified the differences between the four parts of the herb in physical structure and the contents,laying a foundation for further systematic work.
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OBJECTIVE:To optimize the purification technology of total alkaloid from the flos of Aconitum kusnezoffii. METH-ODS:The content of total alkaloid from the flos of A. kusnezoffii was determined by acid-base titration. The purification technology of total alkaloid from the flos of A. kusnezoffii was optimized by ion resin with resin type,mass concentration of loading liquid and exchange speed as factors,maximum adsorption quantity,desorption rate and mass fraction of total alkaloid as index,and verifica-tion test was conducted. RESULTS:The optimal purification technology was as follows as type 732 cation exchange resin,mass concentration of loading liquid 0.32 g/L,exchange speed of 7 column volume(BV)/h. In validation test,the content of total alka-loid was 86.88%(RSD=0.52%,n=3),and desorption rate was 92.81%(RSD=0.40%,n=3)averagely. The extraction trans-port rate of total alkaloid from 3 batches of the flos of A. kusnezoffii was 81.76% and purification transport rate was 89.47% in av-erage. CONCLUSIONS:The established method is stable and feasible,and shows high transport rate.
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Objective To observe the poisoning components in plasma and histological changes of rabbits with acute toxicity of aconitum kusnezoffii (草乌).Methods Eight rabbits were garaged with aconitum kusnezoffii liquor,aconitum poisoning model was reproduced,electrocardiogram (ECG) and blood pressure were recorded,the concentrations of aconitine,hypaconitine and mesaconitine in plasma after 0.5,1, 2,3 and 6 hours were measured,and the pathological changes of heart,liver and cerebral cortex were observed.Results After garage with poisoning liquor,arrhythmias and the declination of blood pressure, presenting a tendency of progressive aggravation [before garage:(121.98?16.77)/(110.66?8.78) mm Hg, 1 hour after garage:(102.98?8.34)/(90.22?5.85) mm Hg,2 hours after garage:(66.81?9.13)/ (53.40?6.32) mm Hg,1 mm Hg=0.133 kPa,all P
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Objective To study how and why the lipo-alkaloids changed in decocting the flowers of Aconitum kusnezoffii (FAK). [WT5HZ]Methods The alkaloids in ethanol extract of FAK, FAK decoction, and the residues of decocted FAK were analyzed and compared by electrospray ionization tandem mass spectrometry (ESI-MSn). Results No lipo-alkaloids were detected in FAK decoction, however, triester-lipo-alkaloids become the dominant components in the residues of FAK decoction. Conclusion Besides hydrolysis reactions, ester-exchange reactions happen for diester-aconitines and triester-aconitines in the decocting of FAK. It is the first time to report that C_8-acetyl is displaced by long chain fatty acyl for triester-aconitines in the ester-exchange reactions.