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INTRODUCTION: Traditional methods to derive experimentally-generated relative correction factors (RCFs) for the quantitative analysis of herbal multi-components by single marker (QAMS) method require reference standards and multiple validations with different instruments and columns, which hampers high throughput implementation. OBJECTIVES: To effectively reduce the application amounts of raw material and provide higher and more stable accuracy, this study aimed to develop a method to computationally generate RCFs of herbal components. MATERIALS AND METHODS: This strategy included the published data collection, calibration curves screening, computer algorithm-based RCFs generation and accuracy validation. RESULTS: Using the in silico approach, we have successfully produced 133 RCFs for the multi-component quantitative analysis of 63 widely used herbs. CONCLUSION: Compared with conventional RCFs, this in silico method would be a low cost and highly efficient way to produce practical RCFs for the QAMS method.

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A simple, specific and selective quantitative analysis of multi-components by single marker(QAMS) method for simultaneous determination of anthraquinones and anthraquinone glycosides in Polygonum multiflorum was developed. Four main anthraquinones and its glycosides, emodin, emodin-8-O-ß-D-glucoside, physcion and physcion-8-O-ß-D-glucoside were selected as analytes to evaluate the quality of P. multiflorum. Emodin was used as the internal standard, and the relative correction factors(RCFs) between emodin and the other three anthraquinones were calculated. Comparison of the contents of the four components in 30 batches of P. multiflorum from different regions and 12 batches decoction pieces from different manufacturers by QAMS and external standard method(ESM) showed that there was no significant difference between QAMS and ESM for quantification of the four main components by using relative error results, and the QAMS method was accurate and reliable, and had a good repeatability. In addition, compared with the results calculated by the difference method between total anthraquinone and free anthraquinone in the content determination of P. multiflorum in Chinese Pharmacopoeia, the results of direct determination combined anthraquinone by QAMS were very close to that by measured the external standard method. Therefore, simultaneous quantification of four main anthraquinones by using QAMS is suitable to evaluate the quality of P. multiflorum. Then the optimized assay method of the combined anthraquinone contents showed simple and feasible, which could be replaced and improved the quantification method of the combined anthraquinone in the current Chinese Pharmacopeia.

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A simple, specific and selective quantitative analysis of multi-components by single marker(QAMS) method for simultaneous determination of anthraquinones and anthraquinone glycosides in Polygonum multiflorum was developed. Four main anthraquinones and its glycosides, emodin, emodin-8-O-β-D-glucoside, physcion and physcion-8-O-β-D-glucoside were selected as analytes to evaluate the quality of P. multiflorum. Emodin was used as the internal standard, and the relative correction factors(RCFs) between emodin and the other three anthraquinones were calculated. Comparison of the contents of the four components in 30 batches of P. multiflorum from different regions and 12 batches decoction pieces from different manufacturers by QAMS and external standard method(ESM) showed that there was no significant difference between QAMS and ESM for quantification of the four main components by using relative error results, and the QAMS method was accurate and reliable, and had a good repeatability. In addition, compared with the results calculated by the difference method between total anthraquinone and free anthraquinone in the content determination of P. multiflorum in Chinese Pharmacopoeia, the results of direct determination combined anthraquinone by QAMS were very close to that by measured the external standard method. Therefore, simultaneous quantification of four main anthraquinones by using QAMS is suitable to evaluate the quality of P. multiflorum. Then the optimized assay method of the combined anthraquinone contents showed simple and feasible, which could be replaced and improved the quantification method of the combined anthraquinone in the current Chinese Pharmacopeia.

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Bufadienolides are the main bioactive components of Venenum Bufonis (VB) and have been widely used to treat different types of human cancers for decades. The bufadienolide content in VB varies significantly in materials from different geographical origins. In this work, a new strategy for the quality assessment of VB was developed through quantitative analysis of multi-components by single marker (QAMS). Cinobufagin was selected as the internal reference substance; seven bufadienolides were separated and simultaneously determined based on relative correction factors. The correlation coefficient value (r ≥ 0.9936) between QAMS and the normal external standard method proved the consistency of the two methods. According to the outcomes of 30 batches of VB samples, the contents of the seven bufadienolides were used for further chemometric analysis. All of the samples of VB from various geographical origins were divided into three categories based on hierarchical cluster analysis and radar plot, which indicated the crucial influence of geographical origins on VB. This study showed that QAMS combined with chemometristry could be used to comprehensively evaluate and effectively control the quality of VB from different geographical origins.

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Multi-component analysis is one of the key techniques for the overall quality control of traditional Chinese medicines. However, the shortage and high cost of reference substances are the greatest obstacles. The substitute method is an alternative solution. In the present study, 11 compounds of rhubarb were simultaneously determined by a method named "two reference substances for determination of multiple components", which includes a qualitative method with linear calibration using two reference substances and a quantitative method with a relative correction factor combined with ultra high performance liquid chromatography. Using aloe-emodin-8-O-ß-D-glucopyranoside and chrysophanol as reference compounds, chromatographic peak identification was performed. The results demonstrated that linear calibration using two reference substances method showed higher accuracy, less deviation, and better column adaptability compared to the relative retention time method. Using chrysophanol as a reference compound, the relative correction factors were determined and showed good reproducibility and stability in different laboratories with different instruments, columns, and wavelength fluctuations. The results had no significant difference compared with the external standard method. The strategy of two reference substances for determination of multiple components coupled with ultra high performance liquid chromatography is economical, efficient, accurate, reliable, and environmentally friendly and is suitable for the quality control of traditional Chinese medicines.

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Objective To establish a QAMS method for content determination of six compositions (chlorogenic acid, caffeic acid, cryptochlorogenin acid, isochlorogenic acid A, isochlorogenic acid C and loganin) from Lonicerae Japonicae Caulis; To verify the feasibility and applicability of this method in quality control of Lonicerae Japonicae Caulis. Methods Chlorogenic acid was set as internal reference substance. The HPLC analysis was performed on a Waters Symmetry C18 column (4.6 mm × 250 mm, 5 μm) with a mobile phase consisted of acetonitrile and 0.4% phosphoric acid solution in gradient elution manner at a flow rate of 1 mL/min. The column temperature was maintained at 35 ℃, and the detection wavelength was set at 327 nm for chlorogenic acid, caffeic acid, cryptochlorogenin acid, isochlorogenic acid A, isochlorogenic acid C and 236 nm for loganin. Results The relative correction factors of caffeic acid, cryptochlorogenin acid, isochlorogenic acid A, isochlorogenic acid C and loganin were established; there was no obvious difference between calculated value of QAMS and measured value of external standard method. Conclusion The quality control mode of QAMS can be used for multi-index synchronization quality evaluation of the six compositions from Lonicerae Japonicae Caulis.

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Objective:To develop a method of quantitative analysis of multi-components by single marker( QAMS) for nine kinds of alkaloids in Xiaohuoluo pills. Methods: An HPLC-QTOF-MS method with an Agilent ZORBAX Extend-C18 RRHT(2. 1 mm × 50 mm,1. 8 μm) column was applied. The flow rate was 0. 21 ml·min-1 . The column temperature was 30 ℃. The mobile phase was methanol (A)-water (B;containing 0. 1％ formic acid and 2. 5 mmol·L-1 ammonium acetate) with gradient elution. The aconitine was used as the internal standard, and the relative correction factor ( RCFs) of hypaconitine, mesaconitine, benzoylaconine, benzoyl-hypaconine, benzoylmesaconine, aconine, hypaconine and mesaconine was respectively established, and the reproducibility inspection on the RCF was performed. The contents of the other 8 kinds of aconitum alkaloids were calculated according to the RCF. At the same time, an external standard method ( ESM) was performed for the content determination of the nine alkaloids. The results of the two methods were compared. The feasibility and accuracy of the QAMS method were verified. Results:Within a certain range,the RCF of hypacontine,mesacontine, benzoylaconine, benzoylhypaconine, benzoyl mesaconine, aconine, hypaconine and mesaconine to aconitine was 1. 736,1. 979,1. 0471,0. 9242,1. 2901,1. 3078,1. 2859,and 1. 0948,respectively. The QAMS method was established for determi-ning alkaloids. There were no significant differences between the results of the QAMS method and those of the external standard method ( ESM) . Conclusion:With the validation of methodology, the method established in our study can be used for the content determina-tion of aconitine, hypaconitine, mesaconitine, benzoylaconine, benzoylhypaconine, benzoylmesaconine, aconine, hypaconine and mesaconine in xiaohuoluo pills.

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Objective To establish QAMS method to determine the contents of three alkaloids in bile processed Coptidis Rhizoma; To compare the results of QAMS with those from external standard method; To prove the feasibility of QAMS.Methods An HPLC method was developed. Berberine hydrochloride was selected as the internal reference substance. 2 relative correction factors (RCF) of berberine hydrochloride to palmatine hydrochloride and to jatrorrhizine hydrochloride were established. Obtained RCFs were used to conduct content calculation (calculated value) to complete QAMS method. At the same time, the contents (measured value) of the three components were also determined by external standard method. Calculated value and measured value were compared.Results The analysis results showed that there was no significant difference between the calculated values and the measured values of the three alkaloids in 10 batches of bile processed Coptidis Rhizoma.Conclusion The QAMS method can be applied in the determination of alkaloids in bile processed Coptidis Rhizoma.

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Objective: To investigate the positioning based on the relative retention time of fingerprinting and to establish a new quality evaluation method for traditional Chinese medicine preparations, using one chemical reference substance to calcutate multi- components simultaneously. Methods: Employed icariin as the maker component, icariin relative correction factors (RCF) of epimedin C to icariin, asperosaponin VI to icariin, psoralen to icariin and angelicin to icariin were ealeatated in the chromatographic conditions for determination of the four components in Xianling Gubao Capsule (XGC). The contents of icariin were determined by external standard method, and those of epimedin C, asperosaponin VI, psoralen and angelicin were calculated by icariin and their RCF. The accuracy of the new method was evaluated by comparing the relative retention times and calculating the content which using different brands columns for determination. Results: The analysis methods were established,the linearity was good when sample volume was in the range of at 8.2—328.0 ng for icariine(r = 0.999 5), 0.055 6—2.224 μg for epimedin C (r = 0.999 6), 0.144 1—5.764 μg for asperosaponin VI (r = 0.999 6), 5.4—215.2 ng (r = 0.998 0) for psoralen, 6.6—265.6 ng (r = 0.998 5) for angelicin. The average recoveries of asperosaponin VI, psoralen and psoralen were 97.59%, 98.58%, 98.11%, 97.86%, 98.22%, respectively. The RSDs of recovery were all less than 2.0%; There has been no significant difference between the calculated contents and the determined contents, according to the angle cosine value. Conclusion: The method can control the components without providing epimedin C, asperosaponin VI, psoralen, and angelicin reference. The method is not only save reference substance and medicine resources, but also suitable quality evaluation pattern for TCM preparation. This new method made fingerprinting more meaningful in TCM quality control.

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Objective: To establish a quantitative analysis of multi-components by single marker (QAMS) for the simultaneous determination of six alkaloids in crude and processed Coptidis Rhizoma. Methods: An HPLC method was established to determine the six alkaloids (jatrorrhizine hydrochloride, columbamine hydrochloride, epiberberine hydrochloride, coptisine hydrochloride, palmatine hydrochloride, and berberine hydrochloride) by the external standard method (ESM). With this HPLC method, the berberine hydrochloride was used as the internal standard (IS) to determine five relative correction factors (RCFs) of the five other alkaloids, and their contents in all samples were calculated by their RCFs at the same time. Compared with the content results determined by the ESM and QAMS, the feasibility and accuracy of QAMS method were verified. Results: Within a certain range, the RCFs of jatrorrhizine hydrochloride, columbamine hydrochloride, epiberberine hydrochloride, coptisine hydrochloride, and palmatine hydrochloride to berberine hydrochloride were 1.131, 0.999, 1.011, 1.076, and 1.025, respectively, with the good repeatability in different experimental conditions. There was no significant difference between the QAMS method and ESM method. Conclusion: The QAMS method is feasible and accurate for the simultaneous determination of the six alkaloids in crude and processed Coptidis Rhizoma.

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To control the quality of Rhizoma Chuanxiong, a method based on high-performance liquid chromatography method coupled with diode array detection was developed for the quantitative analysis of six active ingredients using a single standard to determine multi-components and chemical fingerprint analysis for the first time. The separation was performed on an Agilent Zorbax SB-C18 column by gradient elution with methanol and aqueous phase (containing 0.5% glacial acetic acid) at a flow rate of 1.0 mL/min. The UV wavelength was set at 274 nm. This assay was fully validated with respect to precision, repeatability, and accuracy. All calibration curves showed good linearity (R(2) > 0.9994) within test ranges. The limit of detection and limit of quantification were lower than 0.01 and 0.03 µg/mL, respectively. The relative standard deviation for repeatability and the intermediate precision of six analytes were less than 1.6 and 2.5%, respectively, the overall recovery was 96.1-103.1%. In addition, fingerprint chromatography using hierarchical clustering analysis and similarity analysis was performed to differentiate and classify the samples. The method described here could provide a more comprehensive and reasonable scientific assessment of the quality of Rhizoma Chuanxiong. Therefore, the strategy is feasible, credible, and is easily and effectively adapted for evaluating the quality control of Rhizoma Chuanxiong.

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OBJECTIVE: To establish a quantitative analysis of multi-components by single-marker (QAMS) method for determination of uridine, guanosine and adenosine in Cordyceps sinensis. METHODS: Adenosine was used as the marker to calculate the relative correction factors (RCF) for uridine and guanosine by using high performance liquid chromatography(HPLC). Both QAMS method and external standard method (ESM) were performed to determine the three kinds of nucleosides in Cordyceps sinensis. RESULTS: There were no significant differences between the two methods. CONCLUSION: The developed QAMS method is feasible to evaluate the quality of Cordyceps sinensis.