Studies On The Quality Control About Lipid-lowering Class Of Chinese Herbal Drugs Compound Preparations

In recent years, clinical pharmacology studies have shown that many Chinese herbal drugs, such as Obtuseleaf Senna Seed, rhubarb, Tuber Fleeceflower Root, Oriental Waterplantain Tuber, Ginggo Leaf, have the function of lowering blood lipids. Chinese herbal drugs compound preparation is used widely in current clinical. The lipid-lowering class of Chinese herbal drugs compound preparations based on research of natural plants and traditional chinese physician, which have the role of lowering blood lipids, can reduce blood fat, reduce the incidence and mortality of coronary heart disease and cerebrovascular disease greatly, improve the role of antioxidant capacity. Statins, niacin, phenoxy acids, polyunsaturated fatty acids and other reduced blood fat drugs ware used as western medicine to treat the cardiovascular and cerebrovascular diseases in the clinical, causing headaches, dizziness, nausea, diarrhea, abdominal distension, skin rashes and other adverse reactions. Though the reducing blood fat effect is slowly, there was safely, efficacy comprehensive, greatly effective, without side effects obviously of the Chinese herbal drugs. Compared with Western medicine, studies confirmed that Chinese herbal drugs preparations could slow down or terminate the process of atherosclerosis except reducing blood fat effect; making great therapeutic effect by long-term use.In order to use the Chinese herbal drugs preparations widely and safely, the quality control has become the main target of our study. Controllability of the quality of medicines is a fundamental property, and is a basis of the safety and effectiveness of drugs. Quality standards are the important means of controlling the quality of medicines, which can control the drug quality effectively and ensure the drug safely, and are the concrete embodiment of the quality control. Jiangzhining granules, Zhijiangning tablets and Xiaokeling tablets were selected as the research subjects to study; High performance liquid chromatography and high performance capillary electrophoresis methods were used to control the compound preparations quality, committed systematic methodology verification and determination. The method had good specificity, precision and stability. The experiment results were accurate and reliable. The test method could provide scientific and effective accordance with control the quality of lipid-lowering class of chinese herbal drugs compound preparations.Part OneDetermination of the stilbene glucoside, aurantio-obtusin, emodin and physcion in Lipid-lowering granules by RP-HPLCObjective: A method for the simultaneous separation and determination of stilbene glucoside, aurantio-obtusin, emodin and physcion in Lipid- lowering granules were developed using reverse phase high performance liquid chromatography. The method provided the basis for quality control of the lipid-lowering granules.Methods:1 The extraction method: different extraction solvents and extraction time were investigated to improve the index ingredients of lipid granules extraction efficiency.2 Optimization of the chromatographic conditions: chromatographic condi- tions such as analytical columns, detection method, mobile phase buffer sys- tems, gradient programs, flow rate, maximum absorption wavelength and column temperatures were tested for optimization of the chromatographic conditions, so that the chromatographic peaks of components to be measured were completely separated with other peaks.3 The system suitability test: under the determining extraction and chromatographic conditions, the resolution, theoretical plate number and symmetry factor of four chemical components meet the requirements.4 The study methodology: the limits of detection test, precision test, repeatability test, stability test and recovery test were study, and the calibration curve was constructed.5 The sample content: It was determined the content that the four index ingredients in four batches of granule samples.Results:1 The extraction method: Jiangzhining granules were extracted with 75% methanol as solvent, and 30 minutes was selected as the best ultrasound time for improving extraction rate of the four components.2 The chromatographic conditions: The chromatographic separation was performed on a DiamonsilTM C18 column (150 mm×4.6 mm, 5.0μm). The standards and samples were separated using a gradient mobile phase consisting of 0.1% phosphoric acid (A) and methanol (D). The gradient program was: 0~5 min 75% A, 5~15 min 60% A, 15~25 min 50% A, 25~30 min 20% A and 30~40 min 10% A; The column temperature was maintained at 25℃.The detection wavelengths was set at 286 nm. The flow rate was 1.0 mL/min.3 The system suitability test: in the above chromatographic conditions, stilbene glucoside, aurantio-obtusin, emodin and physcion had good peak shape without interference of other peaks, the separation of adjacent peaks were more than 1.5, the theoretical plate number was more than 5000.4 Methodological study: The ranges of the stilbene glucoside, aurantio-obtusin, emodin and physcion showed good linear relationship, which were 0.580-4.06μg,0.219-1.32μg, 0.0540-0.320μg, 0.189-1.12μg respectively and the regres- sion equations were: A=2.06×103c-5.60(r=0.9998), A=1.36×103c-5.49 (r= 0.9998), A=7.24×102c-2.27(r=0.9995), A=8.53×103c-20.5(r=0.9997) respect- tively; The precision of the method of RSD values were 0.94%, 1.8%, 1.7%, 1.9%; The reproducibility of RSD values were 1.3%, 1.2% , 0.76%, 2.0%; The stability of the RSD within 12 hours were 1.9%, 2.7%, 1.4%, 2.6%; The average recoveries were 96.2%, 96.1%, 96.6%, 97.7%, and the RSD values were 0.74% , 1.0%, 3.1%, 2.6% (n=5).5 The sample Content: Under the established chromatographic conditions, the different batches of lipid granules in the four effective components were investigated, and the four effective components in the sample were calculated by the external standard method.Conclusions: The method was convenient, accurate and highly sensitive. It can be used to determine the contents of the stilbene glucoside, aurantio- obtusin, emodin and physcion in Lipid-lowering granules. The method provided a basis for quality control of lipid-lowering granules.Part TwoDetermination of Propanoid Acid, Chlorogenic Acid, Puerarin, Stilbene Glucoside and Aurantio-obtusin in Zhijiangning Tablets by HPLCObjective: An HPLC method was established for the determination of propanoic acid, chlorogenic acid, puerarin, stilbene glucoside and aurantio- obtusin in Zhijiangning tablets. The method provided a basis for quality control of Zhijiangning tablets.Methods:1 The extraction method: the test solution was abstracted by reflux extraction and ultrasonic extraction: the two extraction methods were compared with, and the best method were choosed to improve the extraction efficiency of the index ingredients.2 Optimization of the chromatographic conditions: chromatographic conditi- ons such as analytical columns, detection method, mobile phase systems, gradient programs, flow rate and column temperatures were tested for optimi- zation of the chromatographic conditions.3 The system suitability test: under the determining extraction and chromato- graphic conditions, the resolution, theoretical plate number and symmetry factor of five index components meet the requirements.4 The study methodology: the limits of detection test, precision test, repea- tability test, stability test and recovery test were study, and the calibration curve was constructed.5 The sample content: It was determined the content that the five index ingredients in three batches of Zhijiangning tables.Results:1 The extraction method: Zhijiangning tables were extracted with 75% methanol as solvent, and 30 minutes was selected as the best ultrasound time for improving extraction rate of five components.2 The chromatographic conditions: A C18 column was used with the mobile phase of 0.1% phosphoric acid-methanol-acetonitrile by gradient elution at the detection wavelength of 286 nm. The column temperature was maintained at 25℃. The flow rate was 1.0 mL/min. The sample size was 10μL.3 The system suitability test: in the above chromatographic conditions, propanoic acid, chlorogenic acid, puerarin, stilbene glucoside and aurantio- obtusin had good peak shape without interference of other peaks, the separation of adjacent peaks were more than 1.5, the theoretical plate number was more than 5000.4 Methodological verification: The calibration curves of propanoid acid, chlorogenic acid, puerarin, stilbene glucoside and aurantio-obtusin were linear in the concentration ranges of 4.93~1.58×102μg/mL, 1.39~4.44×102μg/mL, 8.08~2.59×102μg/mL, 4.53~1.45×102μg/mL and 1.10~3.52×102μg/mL res- pectively and the regression equations were: A=2.06×103c-5.60 (r=0.9998), A=1.36×103c-5.49(r=0.9998), A=7.24×102c-2.27(r=0.9995), and A=8.53×103c -20.5(r=0.9997) respectively; The precision of the method of RSD values were 0.41%, 0.74%, 0.66%, 0.83% and 1.3%; The reproducibility of RSD values were 0.56%, 0.71%, 0.96%, 1.2% and 1.8%; The stability of the RSD within 12 hours were 1.1%, 1.8%, 0.95%, 1.6% and 2.1%; The average recoveries were 96.5%, 98.2%, 97.2%, 95.9% and 96.0%, with RSDs of 0.94%, 1.6%, 0.68%, 2.0% and 0.72%, respectively.5 The sample content: Under the established chromatographic conditions, the different batches of samples in the five index components were measured, and then using the external standard method to calculate the five components in the samples.Conclusions: The method determined the contents of the propanoic acid, chlorogenic acid, puerarin, stilbene glucoside and aurantio-obtusin in Zhi- jiangning tables by HPLC, and provided a strong basis for quality control of Zhijiangning tables, which was convenient, accurate and highly sensitive.Part ThreeDetermination of the Three Constituents in Zhijiangning Tables by Micellar Electrokinetic Capillary ChromatographyObjective: A micellar electrokinetic capillary chromatography method with ultraviolet detection was developed for the simultaneous determination of puerarin, salvianolic acid B and propanoid acid, in Zhijiangning tables. The three components (puerarin, salvianolic acid B and propanoid acid) were systematically investigated and the influential factors were also tested.Methods:1 The extraction method: The sample solution was got out by reflux extraction and ultrasonic extraction, the two extraction methods were compared with, and the best method were choosed to improve the extraction efficiency of the index ingredients.2 Electrophoresis conditions: The separation was performed on an uncoated fused silica capillary column (50.2 cm×75μm×40 cm), with the running buffer contained 80 mmoL/L sodium phosphate monobasic (pH6.30), 8 mmoL/L sodium dodecylsulfate (SDS) and 10% acetonitrile, at the applied voltage of 20 kV, and the detection wavelength of 214 nm. Puerarin, salvianolic acid B and propanoid acid of Zhijiangning tables ware separated at equal pace, and examined influencing factors of the separation effect.3 The system suitability test: under the determining extraction and electrophoresis conditions, the study index components of the resolution and migration time had been inspected.4 The study methodology: the limits of detection test, precision test, repeatability test, stability test and recovery test were study, and the calibration curve was constructed.5 The sample content: The different batches of Zhijiangning tables were determined the contents of the three index ingredients in tables.Results: The linear ranges of puerarin, salvianolic acid B and propanoid acid were 18.0~2.89×102μg/mL, 3.23~51.7μg/mL and 19.7~3.14×102μg/mL, respectively. And the precision of the method of RSD values were 2.3%, 1.4% and 3.2%; The stability of the RSD within 24 hours were 1.2%, 2.6% and 2.5%; The average recoveries were 97.5%, 97.8% and 97.1%, with RSDs of 1.5%, 2.2% and 1.9%, respectively.Conclusions: This simple and rapid method provided a new basis for assessment on quality of Zhijiangning tables, which was low cost test for quality control traditional Chinese medicine and for determination of capillary electrophoresis to provide a reference. Part FourDetermination of Berberine Hydrochloride in Xiaokeling tablets by RP-HPLCObjective: A RP-HPLC method was established for the determination of berberine hydrochloride in Xiaokeling tablets. The method provided a basis for quality control of Xiaokeling tablets.Methods:1 The extraction method: Xiaokeling tables were extracted with 75% methanol as solvent, and 25 minutes was selected as the best ultrasound time for improving extraction rate of two components in samples.2 The chromatographic conditions: A C18 column was used with the mobile phase of 0.1% phosphoric acid and methanol by gradient elution at the detection wavelength of 265 nm. The column temperature was maintained at 25℃. The sample size was 10μL. The flow rate was 1.0 mL/min.3 The system suitability test: under the determining extraction and chromato- graphic conditions, the resolution, theoretical plate number and symmetry factor of two chemical components meet the requirements.4 The study methodology: the limits of detection test, precision test, repeatability test, stability test and recovery test were study, and the calibration curve was constructed.5 The sample content: It was determined the content that the index ingredient in three batches of Xiaokeling tables.Results:1 The system suitability test: Under the above chromatographic conditions, berberine hydrochloride had good peak shape without interference of other peaks, the separation of adjacent peaks were more than 1.5, the theoretical plate number was more than 5000.2 Methodological study: The calibration curves of berberine hydrochloride were linear in the concentration ranges of 0.0268～0.429 mg/mL. The regret- ssion equations were: A=9.70×103c+3.14(r=0.9999); The precision of the method of RSD value was 0.76%; The reproducibility of RSD value was 0.38%; The stability of the RSD value within 24 hours was 0.62%; The average recovery was 98.2% with RSD value of 1.4%(n=5), respectively.3 The sample content: Three batches of the average content of berberine hydrochloride were 4.78 mg/g, 4.47 mg/g and 4.86 mg/g.Conclusions: The method determined the content of berberine hydrochloride in Xiaokeling tables by HPLC, and which provided a basis for quality control of Xiaokeling tables, and was convenient, accurate and highly sensitive.