Study On The Quality Standard Of Burdock Root,Shiluozi And Hongyaozi Based On Spectrum-Effect Relationship

With the development of analytical and detection technology,more and more new techniques and methods have been used in the research of quality standards of Traditional Chinese Medicine(TCM).In recent years,the quality standard that can reflect the efficacy and comprehensively control the quality of TCM has become the focus of research.Burdock root,Shiluozi and Hongyaozi are local traditional medicinal materials in Gansu Province.At present,their quality standards have been included in the "Gansu Provincial Standard of Chinese Medicinal Materials"(2009 edition),which mainly includes the morphological,microscopic identification and some examination items.They can neither reflect the efficacy of medicinal materials nor the quality of medicinal materials.Fingerprint can reflect the chemical composition of TCM comprehensively.Spectrum-effect relationship based on fingerprint can be used to screen the active components of TCM,which is an important method for the quality evaluation of TCM.According to the results of the spectrum-effect relationship,the ingredients that can represent the activity of the medicinal materials were screened and their content was determined.Furthermore,the quality standards that could reflect the quality of the medicinal materials were established by combining the characteristics identification,TLC identification and inspection items.Based on the above,this paper studied the spectrum-effect relationship of the three medicinal materials,the content determination of index components,TLC identification and inspection items.Finally,this paper established the quality standards of the three medicinal materials.Specific research contents are as follows:1 Study on the spectrum-effect relationship of the three medicinal materials1.1 Relationship between Ultra Performance Liquid Chromatography(UPLC)fingerprint and α-glucosidase inhibitory activity of burdock root:Firstly,the UPLC-DAD method was used to build the fingerprints of 11 batches of burdock root,and 19 common peaks were matched,with the similarity between0.5976-0.9710.Secondly,the in vitro α-glucosidase inhibitory activity of 11 batches of ethanol extracts of burdock root was determined.Partial least squares(PLS)method was used to analyze the spectrum-effect relationship between the peak area of fingerprints and α-glucosidase inhibitory activity.It was found that 6 peaks(3,6,9,11,12 and 19)were positively correlated with α-glucosidase inhibitory activity,and the VIP value was all greater than 1.Finally,UHPLC-Q-TOF-MS analysis showed that the peaks 6,9,11,12 and 19 were Chlorogenic acid,1,5-dicaffeoyl-3-maloylqunic acid,3,5-dicaffeoyl-3-maloylqunic acid,1,3,5-tri-o-caffeoyl-4-maloylquinic acid,respectively.1.2 Relationship between High Performance Liquid Chromatography(HPLC)fingerprint and DPPH free radical scavenging activity of shiluozi:Firstly,the HPLC-DAD method was used to build the fingerprints of 11 batches of shiluozi,and 20 common peaks were matched,with the similarity between 0.653-0.964.Secondly,the DPPH free radical scavenging activity of 11 batches of ethanol extracts of shiluozi was determined.Partial least squares(PLS)method was used to analyze the spectrum-effect relationship between the peak area of fingerprints and DPPH free radical scavenging activity.It was found that 9 peaks(1,2,4,6,7,8,9,10 and17)were positively correlated with DPPH free radical scavenging activity,and the VIP value was all greater than 1.Peak 17 was identified as dextral carvone by comparison of reference substances.1.3 Relationship between High Performance Liquid Chromatography(HPLC)fingerprint and antioxidant activity of hongyaozi:Firstly,the HPLC-DAD method was used to build the fingerprints of 10 batches of hongyaozi,and 21 common peaks were matched,the similarity is all greater than0.90.Secondly,the in vitro DPPH radical scavenging activities,hydroxyl radical scavenging activities and superoxide anion scavenging activities of 10 batches of ethanol extracts of hongyaozi was determined.Partial least squares(PLS)method was used to analyze the spectrum-effect relationship between the peak area of fingerprints and antioxidant activity.It was found that 9 peaks(4,5,9,11,12,16,18,19,20)were positively correlated with antioxidant activity,and the VIP value was all greater than 1.3 peaks(2,10,14)were positively correlated with superoxide anion scavenging activity,and the VIP value was all greater than 1.8 peaks(3,10,14,15,16,17,19,20)were positively correlated with hydroxyl radical scavenging activity,and the VIP value was all greater than 1.Finally,HPLC-Q-TOF-MS analysis showed that the peaks 10,12 and14 were 3-caffeoylquinic acid,bergenin and 4-caffeoylquinic acid,respectively.2 Study on the HPLC content determination method of active-related components in three medicinal materialsBased on the study of spectrum-effect relationship,it was found that Chlorogenic acid is the important active component in burdock root that is positively correlated withα-glucosidase inhibitory activity,and dextral carvone is the important component in shiluozi that is positively correlated with DPPH free radical scavenging activity.Bergenin is the important active ingredient positively correlated with the antioxidant activity in hongyaozi.By the examination of the preparation method of the sample solution and the repeatability,stability and recovery of the chromatographic conditions,a sensitive and accurate HPLC methods for the detection of chlorogenic acid,dextral carvone and bergenin in burdock root,shiluozi and hongyaozi were constructed respectively.Finally,it was determined that the content of chlorogenic acid was 0.18-0.45%,the content of dextral carvone was 0.69-4.40 μL/g,and the content of bergenin was 3.31-9.57%.3 Thin Layer Chromatography(TLC)study of the three kinds of medicinal materialsTLC identification methods were established for the three medicinal materials of burdock root,shiluozi,and hongyaozi.In addition,the TLC of the three medicinal materials could clearly showed the spots of chlorogenic acid,dextral carvone,bergenin,respectively,and the established TLC could realize the qualitative identification of the three medicinal materials.4 Draft quality standard of three medicinal materialsBased on the above-mentioned research results and combined with the identification items,inspection items and extracts of the medicinal materials,the draft quality standards of the three medicinal materials were formulated.