| Objective: To optimize and improve the compound Shi-Lin-Tong(CSLT) capsule quality control method by content determination and qualitative identification, and study on the extraction technology and the spray drying process parameters of CSLT capsule, which lay the foundation of the quality control and moistureproof technology research. Method: 1) The thin layer plate of silica gel of HSGF254 was used to identify chlorogenic acid and schaftoside of CSLT, and the developing agent were formulated by ethyl acetate, butanone, formic acid and water in the proportion of 3:3:1:1, the aluminium trichloride was selected as the chromogenic agent. The thin layer plate of silica gel was used to identify loganin of CSLT, D101 macroporous adsorption resin was used to separate and enrich loganin, and the 10% sulfuric acid ethanol was selected as chromogenic agent. 2) Reversed-phase high performance liquid chromatography(RP-HPLC) was used to establish the methods of fingerprint and content determination. The RP-HPLC analysis was carried out on Boston C18 column(250×4.6mm, 5μm), gradient elution was performed by mobile phase of acetonitrile-0.1%formic acid solution, the flow rate was 1.0ml/min and the detection wavelength was set at 335 nm, the column temperature was kept at 30℃. 3) Different batches of CSLT were detected to establish fingerprint. The whole fingerprint similarity was evaluated by the Similarity Evaluation System for Chromatographic Fingerprint of Traditional Chinese medicines(Version 2004 A, National Committee of Pharmacopoeia, China). The MVSP 3.1(Multivariate statistical package) and SIMCA 13.0 were used to hierarchical cluster analysis(HCA) and principal component analysis(PCA), respectively. By comparing the retention time, ultraviolet absorption spectrum of CSLT, single herb and control substance, characteristic peaks of fingerprints were identified and herbs belonging were explained. Compared the contents of efficacy index components(chlorogenic acid, schaftoside and isoschaftoside) from 20 different batches, the quality of capsules/tablets were evaluated. 4) Two evaluation indexs were used to optimize the extraction process of CSLT, the indexⅠinclude the contents of chlorogentic acid and schaftoside, and the extract rate, the indexⅡinclude HPLC characteristic fingerprint peak area and the extract rate. The orthogonal experiment was used to investigate the influence of three factors on extraction process, including the solvent ratio, extraction time and alcohol concentration, and the single factor experiment was used to investigate the effect of extraction times on the transfer rate of active ingredients. The analytic hierarchy process(AHP) was used to determine weight coefficients of each indexs and to evaluate the results of the experiments. 5) To compare the transfer rates of chlorogenic acid and schaftoside, and the differences of two extraction processes was investigated. The sulfuric acid-phenol method was used to determine the polysaccharide content of extract powder, which obtained by two kinds of extraction technologies. To debase the moisture absorption rate of extract powders, the alcohol concentration of water extract-alcohol extraction method was improved and the polysaccharide content was reduced. 6) According to the moisture rate, the single factor experiment was used to optimize the dosage of main medicine and coating powder of CSLT. According to the sample of yield, moisture content and moisture absorption rate, instrument parameters of spray drying were optimized, which include pump speed, inlet temperature and dry air flow. The nebulization pressure was screened by the sample of yield, bulk density and particle size. Result: 1) The new thin layer chromatography of chlorogenic acid, schaftoside and loganin, can evaluate the quality of CSLT capsules. 2) Thirteen common peaks of fingerprints were identified, and eleven common peaks matching could be traced back to its origins. Compared with reference substance and ultraviolet absorption spectra, No.7 peak was chlorogenic acid, No.12 peak was schaftoside and No.13 peak was isoschaftoside. The similarity degree of the capsule was found to be higher than 0.8, including Tianqizhongmeng and Dekang manufacturer, and the fingerprint peak of tables were not complete, including Jixiangchun and Luofushan manufacturer. Through the HCA and PCA analysis, the samples were divided into 4 categories, and among of No.1, 9, 10, 11, 13 peaks had significant effect on the assessment of the sample quality. 3) The linear ranges of chlorogenic acid, schaftoside and isoschaftoside were 1.49~115.98μg/ml(r=0.9997), 1.34~104.76μg/ml(r=0.9999), 1.31~54.72μg/ml(r=0.9999), respectively. The recoveries of chlorogenic acid, schaftoside and isoschaftoside were 97.44%, 100.5%, 101.9%, respectively. The RSD of precision, stability and repeatability were less than 3%. The each capsules or tables content of chlorogenic acid, schaftoside and isoschaftoside were 0.36~1.66 mg, 0.06~1.52 mg, 0.02~1.10 mg, respectively. There were great differences in the contents of chlorogenic acid, schaftoside and isoschaftoside in different batches. The chlorogenic acid of two batch samples of Luofushan manufacturer, and the schaftoside and isoschaftoside of one batch sample of Jixiangchun manufacturer were not detected. 4) The results obtained from the two evaluation indexs were consistent. The optimized extraction process of CSLT capsule was as follow: the medicinal materials were extracted two times with ten times and six times amount of water, one hour for each time, and the concentrated solution was added anhydrous ethanol until the ethanol concentration reaching 80% for ethanol precipitation. 5) The transfer rates of chlorogenic acid and schaftoside were used to optimize the new extraction process, which were significantly higher than the original extraction process. Compared with original technology, polysaccharide content was lower than the original technology, and the new technology of moisture absorption rate was slightly lower than the original technology. 6) The optimum instrument parameters of spray drying were as follows: the pump speed was 6.72ml/min, the inlet temperature was 150℃, the nebulization pressure was 2.3bar, and the dry air flow was 4.1m/s. The moisture absorption rate of the coating sample was significantly lower than the sample without coating. Conclusion: 1) On the basis of the original quality control index of chlorogenic acid, the study established new methods of the thin layer identification(schaftoside and loganin), content determination(schaftoside and isoschaftoside) and fingerprint, which had a better repeatability. The quality of CSLT capsules can be control effectively from two aspects of qualitative and quantitative, which provide the reference for the improvement and perfecting of the quality standard of CSLT capsules. 2) Compared with the original extraction technology, the extracting time of the optimal extraction technology was shortened, the content of index components was increased, and the results was stable, repeatable. It could be used to extract the effective components of CSLT capsules and is suitable for industrial production. 3) On the basis of polysaccharide hygroscopic hypothesis, the alcohol concentration was improved, and the content of polysaccharide was reduced, and moisture absorption rate of powders was slightly lower. In addition, polymer film was formed on the sample surface at the aid of spray drying technique, and hindered the outside water. To a certain extent, the effect of moisture absorption was reduced. The results of the study lay the foundation of the moistureproof technology research. |
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