Study On Preparation Technology And Quality Evaluation Of Qushiqingfei Granules

Objective:This study takes Qushi Qingfei prescription,an empirical clinical prescription for the treatment of mycoplasma pneumoniae pneumonia,as the research object.Which will be developed into Qushi Qingfei granules through researches on extraction process,molding process,quality evaluation and stability,which will provide a new choice for the prevention and treatment of mycoplasma pneumoniae pneumonia and lay a research foundation for the transformation of Qushi Qingfei prescription as a new drug.Methods:1.Study on the extraction technology of Qushi Qingfei Formula:The contents of chlorogenic acid,polygonin,baicalin,wogonoside,baicalein,aloeemodin,wogonin and emodin were determined by high performance liquid chromatography(HPLC).The entropy weight method was used to calculate the comprehensive score based on the contents of eight components and the extract rate.The extraction process was optimized by star design-effect surface method,and the optimal extraction process parameters of Qushi Qingfeifang were determined.The contents of inflammatory cytokines IL-6,IL-1β,TNF-α and NO in the supernatant of rat peritoneal macrophages(RAW 264.7)induced by lipopolysaccharide(LPS)were determined by ELISA.Using Western blot method to detect cell p-IKK、p-NF-κB p65、IκBα protein expression level of change,through the evaluation of clearing damp clearing lung extraction liquid outer anti-inflammatory activity,validation of entropy method in combination with star designresponse surface method to predict the extraction technology parameters stable and feasible.2.Study on the forming process of Qushi Qingfei Granules:The contents of chlorogenic acid,polygonin,baicalin,wogonoside,baicalein,aloeemodin,wogonin and emodin were taken as indexes,and the single factor test combined with entropy weight method was used to optimize the optimal process parameters of decompressive concentration of Qushi Qingfei formula extract.The thickened extractives were dried under vacuum pressure to obtain dry paste powder.Single factor experiment was used to optimize the type and dosage of excipients for the preparation of granules by taking the primary forming rate,hygroscopic rate and solubility of granules as evaluation indexes.The single factor investigation combined with Box-Behnken response surface method was used to optimize the granule parameters of Qushi Qingfei granule dry process,with rolling wheel pressure,feeding speed and rolling wheel speed as the factors and the Primary molding rate as the index.3.Study on the quality evaluation of Qushi Qingfei Granules:According to the provisions of 0104 Granules in General Guidelines for Preparations of Chinese Pharmacopoeia(2020 Edition),the granules were examined.Thin layer chromatography(TLC)was used for the qualitative identification of Yinchen,Scutellaria baicalensis,Fritillaria thunbergii and Polygonum cuspidatum in the prescription.The contents of chlorogenic acid,baicalin and Han baicalin were determined by HPLC,which were included in the quality control standard of Qushi Qingfei Granules.HPLC fingerprints of 10 batches of Qushi Qingfei granules were established,and multivariate analysis including fingerprint similarity,cluster analysis,principal component analysis,partial least squaresdiscriminant analysis was performed to identify the main chromatographic peak components and establish the common fingerprint pattern of Qushi Qingfei granules.4.Stability test research of Qushi Qingfei Granules:According to the quality standard of preparation and the guiding principles of preparation stability experiment,the stability of 3 batches of Qushi Qingfei granules was investigated through accelerated stability test and long-term stability test.Results:1.The optimal extraction process of Qushi Qingfei formula was 13 times of water,two times of extraction,105 min each time.Clearing damp clearing lung party in the supernatant extract can reduce the content of IL-6,IL-1β,TNF-α and NO of cell supernatant,inhibite the protein expression of p-IKK、p-NF-κB p65,and promote the protein expression of IκBα.2.The optimal decompression concentration temperature for Qushi Qingfei Formula is 65-70℃(-0.06～-0.08 MPa),and the decompression drying condition for thickened extract after concentration is 70-75℃(-0.08～-0.1MPa).After optimization,the ratio of dextrin and lactose as mixed auxiliary materials and dry paste powder is dry paste powder:dextrin:lactose=7.5:1.25:1.25,dry granulating conditions are set as rolling wheel pressure of 9 Mpa,feeding speed of 17 Hz,rolling wheel speed of 13 Hz.3.The particle size,solubility,moisture content,loading difference and microbial limit of Qushi Qingfei granules were all in accordance with the provisions of 0104 Granules in the 2020 Edition of Chinese Pharmacopoeia.The quality standard of Qushi Qingfei granules was established,and the TLC identification method of Radix herbaceae,Radix scutellariae,Thunbergam and Polygonum cuspatum was determined.The quantity of chlorogenic acid(C16H18O9)in each bag of Radix herbaceae was not less than 0.5 mg.The amount of baicalin(C21H18O11)in each bag should not be less than 9.9 mg.The amount of baicalin(C22H20O11)in each bag should not be less than 2.5 mg.HPLC fingerprints of Qushi Qingfei Granules were established in 10 batches,among which 23 peaks were identified and the similarity was above 0.99.The 10 batches of Qushi Qingfei Granules could be divided into 3 categories,and 8 main markers that caused quality differences between different batches of samples could be used for quality evaluation of Qushi Qingfei Granules.4.In the accelerated stability test,all indexes of the samples in the 0,1,2,3 and 6 months were within the prescribed range,all indexes of the samples at 0,3,6,9 and 12 months were within the prescribed range.Conclusion:In this study,the anti-inflammatory activity evaluation in vitro verified that the extraction process parameters predicted by entropy weight method combined with star design-effect surface method were stable and feasible,and the selected preparation process was reasonable and feasible.The established quality standard of Qushi Qingfei granules is conducive to improving the controllability of Qushi Qingfei granules,and the prepared Qushi Qingfei granules have great stability,It can lay a research foundation for the transformation of Qushi Qingfei prescription as a new drug.