| In recent years, toxicity reducing and efficacy enhancing of Chinese medicine compatibility are more and more paid attention by researchers, toxic Chinese medicine Fuzi is widely applied in the clinical because of its better effect, but Fuzi has significant toxic and side effects it has been widely concerned by people. The national basic research program of973-"Study on toxic Chinese medicine compatibility attenuation principle and method(2009CB522806)" was studied systematically from compatibility attenuated from the literature, theory, method and research on evaluation standards and other aspects, this project was a part of the research content of973program. The topic made SND prescription dismantlement analysis and compared the acute toxicity, toxic target organs-the heart toxicity and the Fuzi major component of SND prescriptions with different combination between the single Fuzi group, Fuzi combined with Gancao group, Fuzi combined with Ganjiang group, Fuzi combined with Gancao and Ganjiang group, and observed the efficacy influence of SND different prescription compatibility through the rat model of heart failure Cardiac function. Combining with aconitine constituent from this two aspects of toxicology and pharmacodynamics studied SND compatibility rationality, then provided experimental basis for traditional Chinese medicine compatibility attenuated and references for safe and reasonable clinical application of toxic traditional Chinese medicines.1Aconitine content determination of SND prescription compatibilityThe experiment was based on the selected samples drug, by HPLC determined the double ester type alkaloids and single ester type alkaloids constituents of several decotions about Fuzi and Fuzi combined different combinations(1g fuzi/ml) in SND. The results showed that four decotion of SND compatibility had not detected aconitine, Fuzi single and Fuzi combined with Gancao had not detected new aconitine. Compared with single ester type alkaloids constituents and the double ester type alkaloids constituents, three compatibility groups were also lower than Fuzi single; compared with Fuzi single, the double ester type alkaloids constituents of Fuzi compared with Gancao group(-40.1%) was lower30.9%than one of Fuzi compared with Ganjiang group(-9.2%), and was almost close to SND.2Toxicology study of SND prescription compatibility2.1Study on median lethal dose(LD50) in mice of Fuzi and Fuzi combined different combinations in SNDThis study used intragastric administration that was consistent with the clinical practice in mice, from the preliminary experiment we got0%(undead) and100%(full dead) lethal dose range and selected five different dose of four kinds of decotions about Fuzi and Fuzi combined different combinations to performed by multiple proportion dilute method. After being fed3days for adaptability,50mice of each solution were taken and randomly divided into5groups,10mice for each group, Mice were treated by intragastric at the dose of40ml/kg after being fasted but provided water for16h. Different doses of the solution were given only once in mice from the beginning with the middle dose, after taken observing the animal toxicity reaction. Every group mice deaths were statistical treated by Bliss program and calculated the median lethal dose (LD50) and95%confidence interval of LD50. The results showed that the animals behavior of the four groups of Fuzi, SND, Fuzi combined with Ganjiang, Fuzi combined with Gancao were identical with Fuzi single after the treatment, but versus to Fuzi single LD50, SND, Fuzi combined with Ganjiang, Fuzi combined with Gancao could increase23.6%,11.8%,32.8%than it, so Fuzi combined with Gancao increased21%than Fuzi combined with Ganjiang in LD50of Fuzi, increased9.2%than LD50of SND.2.2Study on median toxic dose (TD50) in rats of Fuzi and Fuzi combined different combinations in SNDAfter SD rats were fed adaptively for5days, anmials were fasted but provided water for16h before the experiment, then rats were anaesthetized by intraperitoneal injection of3%sodium pentobarbital solution, lied on its back and fixed. Rats electrocardiogram(ecg) status were recorded By16channel physiological recorder, positive electrode inserted into the animal’s right upper limb, negative electrode inserted into the animal’s left lower extremity, ground wire inserted into the animal’s right lower extremity, rats were abandoned whose electrocardiogram(ecg) was abnormal after continuous observation for10minutes. After disinfected anmial’s stomach skin with alcohol and cut animal abdominal cavity along the ventral white line, rats were injected by duodenum at the dose of2ml/100g. At first the experiment regarded1/2of Fuzi LD50as the initial dosage, rats were observated contiously60min by16channel physiological recorder, when whose hearts appeared all types of arrhythmia as positive outcome of heart cardiotoxicity.0.8group interval,6mice for each group, received all animal poison dosage and all animals disappear toxic dosage of the heart. After the experiment, counted the number of every group rats poisoned, statistical treated with by Bliss program and calculated the median toxic dose (TD50) and95%confidence interval of TD50. The results showed that the animals behavior of the four groups of Fuzi, SND, Fuzi combined with Ganjiang, Fuzi combined with Gancao were identical with Fuzi single after the treatment, heart mainly appeared all types of arrhythmia as positive outcome of heart cardiotoxicity. In contrast to LD50Fuzi single, SND, Fuzi combined with Ganjiang, Fuzi combined with Gancao could increase28.6%,-63.1%,45.2%than it, so Fuzi combined with Gancao increased16.6%thanSND in LD50of Fuzi, but TD50of Fuzi combined with Ganjiang was lower63.1%than Fuzi single.2.3Correlation analysis of aconitine ingredients and toxic effect of Fuzi and Fuzi combined different combinations in SNDUsed the SPSS13.0software, three kind of the double ester type alkaloids constituents as dependent variable, LD50and TD50as independent variables, the application of mult-variable linear return analysis statistical methods made the correlation analysis for the toxic effects and the double ester type alkaloids constituents. The results showed that double ester aconitine total content was negatively correlated with LD50and TD50, and was positively correlated with toxic effects. It Showed double ester aconitine total content is higher, so toxic effects is also stronger, further illustrated the content of Fuzi combined with Gancao was the lowest and the toxicity was the lowest.According to the above results showed:In SND prescription compatibility Gancao played a main role in attenuating Fuzi’s toxicity.3Pharmacodynamics studies of SND prescription compatibility3.1Pharmacodynamics studies on cardiac function aspect in heart failure models rats of Fuzi and Fuzi combined different combinations of SND prescription compatibilitySD rats were randomly divided into11groups by weight after they were fed adaptively for5days, they were:Normal control group(N group),Model control group(M group),positive control group(digoxin), SND high and low dose group, Fuzi high and low dose group, Fuzi combined with Gancao high and low dose group, Fuzi combined with Ganjiang high and low dose group,8rats for each group. Except for N group, the other animals heart failure models were induced by the twice injections of Doxorubicin Hydrochloride Inj via coccygeal vein (4mg/kg,0.2ml/100g weight),7days once, N group was injected equal volume of saline via coccygeal vein. From the modeling day on the rats were treated by intragastric corresponding medicine at the dose of lml/100g weight, N group and M group were treated equal volume distilled water. After the last dose each animal was weighed, then were anaesthetized by intraperitoneal injection of3%chloral hydrate, lied on its back. Then animals were cut open in the midline of the neck, exposed and blunt dissected right common carotid artery, connected BFM(blood flowmeter) to measure blood flow and recorded the data, then removed and ligatured telecentric end, clamped proximal end with the artery clamp, using a eye scissor cut a small oblique opening under ligation offline, Millar cather was pushed slowly into the left ventricle from arterial wall incision to the direction of the heart and fixed, connectted16channel physiological recorder,3minutes for stabilization later tied off catheter and observed contiously10min and recorded the following indicators:Heart rate (HR), Left Ventricular Systolic Pressure(LVSP), Left Ventricular End Diastolic Presssure(LVEDP), the maximum rate of rise of left ventricular pressure (+dp/dtmax), the largest decline rate of left ventricular pressure (-dp/dtmin).The results showed that compared with Normal control group(N group), LVSP, LVEDP,+dp/dtmax, HR and Carotid artery blood flow of Model control group(M group) were decreased significantly,-dp/dtmin was increased significantly(P<0.05,P＜0.01), this suggested the model was established. Compared with Model control group(M group), both groups of SND and Fuzi combined with Ganjiang could increase significantly LVSP,+dp/dtmax, HR and Carotid artery blood flow, could decrease significantly-dp/dtmin (P<0.05, P<0.01). but both groups of Fuzi single and Fuzi combined Gancao could only significantly improved HR and carotid artery blood flow (P＜0.05, P＜0.01). Compared with Fuzi single, both groups of SND and Fuzi combined Ganjiang high groups could improved significantly LVSP and+dp/dtmax, but Fuzi combined Gancao and Fuzi single groups had no effect.Compared with N group the rats weight results showed that the weight of M group animals were decreased significantly, both had significant difference (P<0.05, P<0.01). Compared with M group the weight of each treated group animals had increased trend, but no significant difference.3.2Correlation analysis of aconitine ingredients, toxic effect and therapeutic effect of Fuzi and Fuzi combined different combinations in SNDDue to the double ester aconitine was the main therapeutic effects ingredients of Fuzi subclasses medicinal materials, therapeutic effects directly were affected by aconitine ingredients. So comprehensive analytical SND prescription compatibility double ester alkaloids content determination results and phannacodynamics test results(Selected LVSP and+dp/dtmax two indicators which closely related to the cardiac function), for the double ester type aconitine total content and therapeutic effects had made comprehensive analysis.Under this test conditions the results showed, double ester aconitine total content of Fuzi single was the highest, followed by Fuzi combined with Ganjiang, SND, Fuzi combined with Gancao was the lowest; to pharmacodynamics index, Fuzi single was the worst, followed by Fuzi combined with Gancao, the therapeutic effects of SND and Fuzi combined with Ganjiang high groups were the most obvious. The therapeutic effects of Fuzi combined with Ganjiang was stronger than one of Fuzi combined with Gancao, but the therapeutic effects of SND which content was lower than Fuzi combined with Ganjiang were stronger Fuzi combined with Ganjiang, it illustrated the therapeutic effects of Fuzi combined with Ganjiang were the best under Fuzi combined with Gancao controlled the toxicity. All above analysis showed Fuzi combined with Ganjiang with stronger therapeutic effects has just simplicity promoted the therapeutic effects. So in SND prescription compatibility Ganjiang played a main role in enhancing efficacy.In summary, this study showed that SND in prescription compatibility had thought fully about the efficacy and safety of the prescription, Fuzi combined with Ganjiang played a main role in promoting the therapeutic effects, but Fuzi combined with Gancao played a main role in decreasing the toxicity of the prescription. It reflected the rationality of compatibility and also gave full reflections to the complexity of Chinese compound compatibility. As 《Zhengzhiyaojue》 said "Fuzi is not hot without Ganjiang, but combined with Gangcao is mild". |
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