Investigation Of Quality Control Methods Of Sanhuang Tablet

Sanhuang tablet(SHT),derived from San-Huang-Xie-Xin-Tang(a famous prescription by Zhang Zhong-Jing),is a relatively simple herbal preparation that mainly consists of Rhei Radix et Rhizoma,Scutellariae Radix extract and berberine hydrochloride and has been officially adopted by the Chinese Pharmacopoeia.SHT,which has multiple pharmacological activities,including anti-inflammatory,antimicrobial,antioxidant and antineoplastic activities and performs the functions of clearing heat,detoxification,and purging pathogenic fire,has been commonly used for the clinical treatment of sore throat,gingival bleeding,intense heat in the body,reddish urine,and constipation,etc.To date,the quality control of TCM has conventionally been based upon the analysis of single or a few known marker compounds,however,this approach is certainly unsufficient since it is not yet available to characterize the properties of total chemical components of herbal products,nor veritably reflect the discipline of effective component groups in vivo.On the basis of overall concept is deemed as the guiding ideology,SHT is selected in the present paper and multiple analytical technologies are employed for the investigation of the quality control methods.Mainly research ideas are as follows:at first,multiple analytical methods and multi-angle integration modes are used establishing the qualitative and quantitative fingerprint for the purpose of studying the quality evaluation methods,moreover,chemical composition fingerprinting is coupled with chemometrics to explore the fingerprint-effect relationship of the complex herbal preparations,finally the chemical composition identification in vitro associated with prototype components primary investigation in the hope of providing scientific basis for further pharmacokinetics,pharmacodynamic material foundation.1.Study on the digitized HPLC fingerprints of SHT under 254 nmThe super information digitized fingerprints of 30 batches of SHTs under 254 nm were established after the principle of linear quantitative fingerprint method being expounded in this article.46 parameters were together to effectively excavate the potential chemical information and visually reflect sample inherent characteristics.The quality of the selected samples was evaluated comprehensively using linear quantitative fingerprint method(based on SL,ML and a values)to reveal the similarities and/or differences among the quantity and content levels of chemical fingerprints,in addition,the origins of the peaks were assigned by comparing the chromatograms of the extracts of the individual crude drug in SHT formula.2.Combinative method using multi-wavelength fingerprints and the DPPH radical-scavenging assay for the investigation of fingerprint—efficacy relationshipA HPLC method with DAD was developed and validated for multi-wavelength fingerprint profiling to evaluate the quality of 30 batches of SHTs using linear quantitative fingerprint method,the mean square-square root method was also successfully applied to integrate the chemical information at multiple wavelengths,then quantitative analysis of eight marker compounds(aloe-emodin,chrysophanol,rhein,emodin,physcion,berberine hydrochloride,baicalin and wogonoside)in each collected sample was performed using the wavelength where the marker compound showed the maximum absorbance.A combination of multi-wavelength fingerprints and simultaneous quantification of eight analytes may make it possible to achieve batch-to-batch and/or manufacturer-to-manufacturer quality differentiation assessment,meanwhile,a measurement of DPPH scavenging activity was taken in vitro.Moreover,the component-activity relationship was established by partial least squares regression(PLSR),and the constructed PLSR model not only produced satisfactory results for predicting the total antioxidant activity of the examined and new samples by determining the peak areas of chemical components from the HPLC fingerprints,but also provided a valuable clue for the quantification of multi-ingredient that were highly related with the biological activity.3.Study on capillary electrophoresis fingerprints and the hydroxyl radical-scavenging assay of SHTA capillary electrophoresis fingerprint was constructed for SHT,where the isosceles trapezoid method was first applied for the optimization of background electrolyte solution,in addition,the quality consistency of 30 batches of SHTs from different sources was evaluated using capillary electrophoresis fingerprint combined with two-component quantitative determination and the quality of the drug medicine was differentiated according to the linear quantitative fingerprint method from both the qualitative and quantitative aspects.At the same time,the classical Fenton reaction was performed to determine the hydroxyl radicals scavenging activity in vitro.A PLS model was successfully constructed with excellent performance to investigate the fingerprint-efficacy relationship and predict the antioxidant activity of the collected samples from their fingerprint pro-files.The results elucidated that the integrated use of fingerprint coupled with antioxidant activity may offer a reliable and efficient approach to the quality control of TCM and herbal preparations from both the chemical and bioactive profiles.4.The integrated use of ultraviolet spectrum fingerprint and infrared spectrum fingerprint coupled with combustion heat to comprehensively assess the quality grades of SHTUltraviolet spectrum fingerprint and infrared spectrum fingerprint were developed,where a measurement of combustion heat was taken for SHT.To evaluate the quality of the drug samples,SL,ML and a values were computed for each analytical method,and the integrated SL,ML and a values were also calculated by taking the average value of the above-established method.Mutiple analytical techniques were tentatively employed to monitor aturated and unsaturated bond of chemical composition.Attempts were made to elucidat the overall content characteristics of all types of chemical ingredients and display the quality profiles from multiple perspectives,which provided a simple,fast,comprehensive and objective approach for the quality control of TCM and herbal preparations from both the qualitative and quantitative aspects,in the case of complex components are not yet completely separated.5.The chemical components identification in vitro and prototype components preliminary studyUPLC-Q-TOF and UPLC-MS/MS were used to authenticate the chemical components of SHT and a total of 28 compounds were identified in vitro,where eight compounds(aloe-emodin,chrysophanol,rhein,emodin,physcion,berberine hydrochloride,baicalin and wogonoside)were confirmed by comparison with the reference standards and the others were tentatively speculated by means of precise molecular weight,MS/MS fragmental information and literature,and then some prototype components in rat plasma were primary investigated using multiple reaction-monitoring mode under UPLC-MS/MS analytical conditions,after oral administration of SHT extract,the dynamic trends of prototype components at different time points were also studied.It was therefore possible to conclude that the studies laid solid foundations for further researches about pharmacokinetics and pharmacodynamic material foundation.