Research And Development On Substance Benchmarks Of Classic Formula Xiaochengqi Decoction

Xiaochengqi Decoction is derived from "Treatise on Febrile Diseases" compiled by Zhang Zhongjing in the Eastern Han Dynasty.The prescription consists 4 Chinese ounces of Rhubarb(wine wash),2 Chinese ounces of Magnoliae Officinalis Cortex(sunburned,peeled),and 3 pieces of Aurantii Immaturus Fructus(larger,roasted),which are used to treat the syndrome of Yang Ming Fushi with light syndrome.This decoction is included in the catalogue of "Ancient Classic Famous Prescriptions(First Batch)".This research was carried out from four aspects:textual research of ancient books,substance benchmarks preparation procudure,quality control method and chemical composition analysis,which laid a foundation for the development of classical prescription Xiaochengqi Decoction.A textual research was carried out on Xiaochengqi Decoction prescription,herbal research and decoction pieces processing through consulting Chinese ancient book materials and modern books.According to the research of the herbal medicines,Rheum palmatum L.,Citrus aurantium L.and Magnolia officinalis Rehd.Et Wils.were finally selected to produce the preparation.Through the processing research of the corresponding pieces of various herbs,it was found that the pieces used in the prescription should be wine-processed Rhubarb,bran-fried Fructus Aurantii Immaturus and ginger-processed Magnoliae Officinalis Cortex.One Chinese ounce in Han Dynasty was equal to 13.8 g,and the prescription composition is 55.2 g of wine rhubarb,41.4 g of bran fried Aurantii Immaturus Fructus and 27.6 g of Magnolia Officinalis.However,in clinics,one Chinese ounce is equal to 3 g.Given the above,the prescription should be 12 g of wineprocessed Rhubarb,9 g of bran-processed Aurantii Immaturus Fructus and 6 g of ginger-processed Magnoliae Officinalis Cortex.The preparation procedure of Xiaochengqi decoction was studied with soaking time,amount of water,decocting time,decocting times,concentration temperature and drying method by comparing the indexes of extraction rate,fingerprint and transfer rate(rhein and neohesperidin).Finally,the substance benchmarks preparation process of Xiaochengqi decoction was determined as follows:take 48 g of wine-processed Rhubarb,36 g of bran-processed Aurantii Immaturus Fructus and 24 g of ginger-processed Magnoliae Officinalis Cortex in 3500 ml electric casserole,add 1080 ml water and immerse 30 min,heat to boiling,simmer for 1 h,filter.Then add 864 ml of water to the dregs,heat to boiling,simmer for 0.5 h,and filter.The filtrates were combined and concentrated under reduced pressure at 50℃,and then freeze-dried.The quality standard of Xiaochengqi decoction was established,including TLC,fingerprint and assay.The TLC conditions of Xiaochengqi decoction were determined by investigating the developing solvent,temperature and humidity.The results of methodology validation showed that the three TLC conditions were specific,stable and durable.By optimizing the preparation conditions and chromatographic conditions of the substance benchmarks,the fingerprint method of Xiaochengqi decoction were developed,and the method validation on the specificity,precision,repeatability and stability was carried out.Four reference peaks were determined to be naringin,neohesperidin,rhein and magnolol.Through the optimization of preparation conditions and chromatographic conditions,the HPLC methods for the determination of rhein and neohesperidin were developed.The results of methodology validation showed that the two methods had good specificity,linearity,accuracy,precision,repeatability,intermediate precision,durability and stability.Based on the combination of ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry(UPLC-QTOF)and Waters UNIFI software,the chemical constituents of the classic prescription Xiaochengqi decoction were qualitatively analyzed and identified.A total of 233 components in Xiaochengqi decoction were identified or characterized,including 93 components from wine-processed Rhubarb,104 from bran-processed Aurantii Immaturus Fructus,and 36 from ginger-processed Magnoliae Officinalis Cortex.These 233 components include anthraquinones,flavonoids,alkaloids,coumarins,phenylethanol glycosides and other compounds.The result provided experimental evidence for the further study on establishment of quality standard and product development of the formula.In conclusion,the textual research of ancient books,the technological research of substance benchmarks,the research on quality control method and the comprehensive characterization of chemical composition of Xiaochengqi decoction were completed in this study,which provided a solid theoretical basis for further study and quality control of the preparation,and also provided a new scientific basis and reference for the overall herbal quality evaluation.