Method for quantitatively measuring concentrations of multiple compounds having same chromophore by using single standard

Abstract

A method for quantitatively measuring the concentrations of multiple compounds having a same chromophore by using a single standard. The measuring method comprises the following steps: using high-performance liquid chromatography to measure standard working solutions having different concentrations, and drawing a standard curve of mass concentrations and peak areas of a standard; further converting said curve into a standard curve of molar concentrations and peak areas of the standard; when the standard and a compound to be measured have the same chromophore, the same molar concentration and the same absorbance, obtaining a standard curve of mass concentrations and peak areas of said compound; and substituting a measured peak area of said compound into the curve, and calculating to obtain the mass concentration of said compound. The measuring method uses few standards, has accurate measurement results, and saves the time and costs of series methodological investigations such as standard curve formulation of each component, thus improving method development efficiency.

Description

[0002] Complex systems are research objects that often occur in the chemical analysis and research. Complex systems are widely present in various fields such as food and medicine. Complex samples often include many structural analogues (including isomers, homologues, etc.) with similar physical and chemical properties. When a complex sample is subjected to compositional analysis, especially quantitative evaluation, it is necessary to quantitatively analyze a plurality of components to comprehensively characterize the quality of the complex sample.

[0003] There are the following two methods for the quantification of multi-components: traditional multi-standard quantification (the multi-standard curve method) and single-standard quantification of multiple components. In practical applications, the multi-standard curve method presents challenges such as a large workload and difficulty in obtaining standards, which are related to the fact that some standards are difficult to prepare or / and have unstable structures and high prices. Therefore, the use of a single standard to quantify a plurality of components has gradually become a research hotspot. Currently, the common single-standard quantification method is the quantitative analysis of multi-components by single marker. The quantitative analysis of multi-components by single marker was first proposed in 2006, in which a plurality of components are simultaneously quantified with a single standard by establishing an intrinsic functional relationship among different components. The quantitative analysis of multicomponents by single marker is also known as single standard to determine multi-components (SSDMC). Currently, the SSDMC method based on liquid chromatography analysis is a major multi-component analysis method.

[0004] However, it has been found that the SSDMC method is still faced with many problems in applications. For example: 1) Error increase progressively: The quantitative analysis of a plurality of components by the SSDMC method involves many calculation steps, and an error is generated in each calculation step. Therefore, the overall error will be amplified step by step. 2) A relative correction factor (RCF) is required for the calculation of each component in a sample, but the RCF is influenced by many factors, and the calculated value has a large deviation, which makes it difficult to avoid errors. 3) Selection of internal reference substances: When an internal standard is a trace component in a sample, RCFs acquired by different laboratories are quite different; that is, the value of RCF fluctuates greatly, resulting in errors in the determination of the contents of a plurality of components. Therefore, how to simplify the multi-component quantification for a complex system, improve the efficiency of method development, and improve the accuracy of a method is a problem to be solved. SUMMARY

[0005] An objective of the present disclosure is to provide a method for quantifying a plurality of compounds with the same chromophore using a single standard. The method requires few standards, provides accurate detection results, and also saves time and costs in a series of methodological investigations such as establishment of individual standard curve for each component, thereby improving the efficiency of method development.

[0006] To achieve the objective of the present disclosure, the present disclosure provides the following technical solutions:

[0007] The present disclosure provides a method for quantifying a plurality of compounds with the same chromophore using a single standard, including the following steps:

[0008] 1) detecting standard working solutions with different concentrations by high-performance liquid chromatography, and with mass concentrations as x-coordinate and peak areas as y-coordinate, plotting a standard curve of the mass concentrations and the peak areas for a standard;

[0009] 2) according to a relationship between the mass concentrations and molar concentrations of the standard, plotting a standard curve of the molar concentrations and the peak areas for the standard; and

[0010] 3) under a condition that a compound to be tested shares the same chromophore as the standard, and that a molar concentration Cmeasured of the compound to be tested is the same as a molar concentration Cr of the standard, an absorbance ymeasured of the compound to be tested is the same as an absorbance yr of the standard, and basing on a relationship between mass concentrations and molar concentrations for the compound to be tested, plotting a standard curve of the mass concentrations and peak areas for the compound to be tested: y =------av ( / 77g / / / I ) + / ) measured measured v 1 J measured ($)

[0011] where ymeasured represents a peak area of the compound to be tested, xmeasured represents a mass concentration of the compound to be tested, Mr represents a molar mass of the standard, Mmeasured represents a molar mass of the compound to be tested, and a and b are parameters; and substituting the peak axea. ymeasured of the compound to be tested into the equation (5) to calculate the mass concentration Xmeasured of the compound to be tested.

[0012] In some embodiments, in step 1), the standard curve of the mass concentration and the peak area for the standard is: yr = axr(mg / mL) + b [0013 ] where yr represents a peak area of the standard, xr represents a mass concentration of the standard, and a and b are parameters.

[0014] In some embodiments, in step 2), the relationship between the mass concentration and the molar concentration of the standard is: yr(mg / mL) = 1000* ]\ / [ r* (J r(mmol / L) . . r r r (2)

[0015] where xr represents the mass concentration of the standard, Mr represents the molar mass of the standard, and Cr represents the molar concentration of the standard.

[0016] In some embodiments, in step 2), the standard curve of the molar concentration and the peak area for the standard is: yr = a*\QQQ*Mr*CSmmo^l + b p)

[0017] where jr represents the peak area of the standard, Cr represents the molar concentration of the standard, Mr represents the molar mass of the standard, and a and b are parameters.

[0018] In some embodiments, in step 3), the relationship between the mass concentration and the molar concentration for the compound to be tested is: C1 =------------r v—' measured 1000^ K / f * / Vmeasured Ivi measured (4)

[0019] where Cmeasured represents the molar concentration of the compound to be tested, Xmeasured represents the mass concentration of the compound to be tested, and Mmeasured represents the molar mass of the compound to be tested.

[0020] The present disclosure provides a method for quantifying a plurality of compounds with the same chromophore using a single standard. The method of the present disclosure requires few standards. Standards are mostly expensive. Thus, compared with the multi-component quantification method adopting a plurality of standards, the method of the present disclosure reduces the cost, saves time and costs in a series of methodological investigations such as establishment of individual standard curve for each component, and improves the efficiency of method development. With the method of the present disclosure, only a standard curve for a component with a known standard is required to deduce standard curves for the remaining components to be quantified. In the existing SSDMC method, RCFs need to be determined for the remaining components to be quantified relative to the component with the known standard, and the RCFs are greatly affected by component concentrations, experimental conditions, etc., resulting in large errors of content detection results. In addition, the method of the present disclosure does not require the determination of a relationship among different components, and also does not require the cumbersome correction factors to correct other components, which avoids the influence of external factors such as instruments and laboratories and internal factors such as purities of standard substances, reduces the measurement time and components, and improves the efficiency and accuracy of quantification method development. The standard curve conversion for quantification in the present disclosure completely depends on a molar mass among physical characteristics of a compound. Once a compound to be tested is determined, a molar mass of the component remains unchanged. Therefore, the method of the present disclosure reduces unnecessary errors, allows the free selection of internal reference substances, and does not need to consider a content of a compound in a sample. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] FIG. 1 shows a high-performance liquid chromatography spectrum of a ginseng decoction piece as a test sample (in FIG. 1, 1 represents ginsenoside Rgl; 2 represents ginsenoside Re; 3 represents ginsenoside Rf; 4 represents ginsenoside Rbl; 5 represents ginsenoside Rb2; and 6 represents ginsenoside RD);

[0022] FIG. 2 shows chemical structures of the ginsenosides;

[0023] FIG.3 shows a comparison of the measurement results of example 1 and comparative example 2;

[0024] FIG. 4 shows a high-performance liquid chromatography spectrum of a Scutellariae Radix decoction piece as a test sample (in FIG. 4, 1 represents baicalin; 2 represents wogonoside; and 3 represents baicalein);

[0025] FIG. 5 shows chemical structures of the baicalin, the wogonoside, and the baicalein; and

[0026] FIG. 6 is a comparison chart of detection results of Example 2 and Comparative Example 4. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0027] The present disclosure provides a method for quantifying a plurality of compounds with the same chromophore using a single standard, including the following steps:

[0028] 1) Standard working solutions with different concentrations are detected by high-performance liquid chromatography, and with mass concentrations as x-coordinate and peak areas 5 as y-coordinate, a standard curve of the mass concentrations and the peak areas for a standard is plotted.

[0029] 2) According to a relationship between the mass concentrations and molar concentrations of the standard, a standard curve of the molar concentrations and the peak areas for the standard is plotted.

[0030] 3) Under a condition that a compound to be tested shares the same chromophore as the standard, and that a molar concentration Cmeasured of the compound to be tested is the same as a molar concentration Cr of the standard, an absorbance ymeasured of the compound to be tested is the same as an absorbance yr of the standard, and based on a relationship between mass concentrations and molar concentrations for the compound to be tested, a standard curve of the mass concentrations and peak areas for the compound to be tested is plotted: =—M—     (mg / mL) + b ' measured measured v measured where ymeasured represents a peak area of the compound to be tested, xmeasured represents a mass concentration of the compound to be tested, Mr represents a molar mass of the standard, tel’measured represents a molar mass of the compound to be tested, and a and b are parameters.

[0031] A peak area ymeasured of the compound to be tested is substituted into the equation (5) to calculate the mass concentration ^measured of the compound to be tested.

[0032] In the present disclosure, in the step 1), standard working solutions with different concentrations is detected by high-performance liquid chromatography, and with mass concentrations as x-coordinate and peak areas as y-coordinate, a standard curve of the mass concentrations and the peak areas for a standard is plotted. Preferably, the standard curve of the mass concentrations and the peak areas for the standard is as follows: yr = aXr(niglmL) + b

[0033] where yr represents a peak area of the standard, xr represents a mass concentration of the standard, and a and b are parameters.

[0034] In the present disclosure, in step 2), according to a relationship between the mass concentrations and molar concentrations of the standard, a standard curve of the molar concentrations and the peak areas for the standard is plotted. Preferably, the relationship between the mass concentrations and the molar concentrations of the standard in step 2) is as follows: (mg / mL) = 1000(Jr(mmolIL) ( . r r t

[0035] where xr represents a mass concentration of the standard, Mr represents the molar mass of the standard, and Cr represents a molar concentration of the standard.

[0036] Preferably, the standard curve of the molar concentrations and the peak areas for the standard in step 2) is as follows: yr = a*\QQQ* Mr* CMnmol I,,,.

[0037] where yr represents a peak area of the standard, Cr represents a molar concentration of the standard, Mr represents the molar mass of the standard, and a and b are parameters.

[0038] In the present disclosure, in step 3), under a condition that a compound to be tested shares the same chromophore as the standard, and that a molar concentration Cmeasured of the compound to be tested is the same as a molar concentration Cr of the standard, an absorbance ymeasured of the compound to be tested is the same as an absorbance yr of the standard, and based on a relationship between mass concentrations and molar concentrations for the compound to be tested, a standard curve of the mass concentrations and peak areas for the compound to be tested is plotted: Mt y =------(mg / mL) + b measured *4 measur ed measured ($)

[0039] where ymeasured represents a peak area of the compound to be tested, xmeasUred represents a mass concentration of the compound to be tested, Mr represents a molar mass of the standard, Mmeasured represents a molar mass of the compound to be tested, and a and b are parameters.

[0040] A peak ymeasured of the compound to be tested is substituted into the equation (5) to calculate the mass concentration Xmeasured of the compound to be tested. Preferably, the relationship between the mass concentrations and the molar concentrations for the compound to be tested in step 3) is as follows: =------!------(mg / m£) measured 1000* JW” measured measured (4)

[0041] where Cmeasured represents a molar concentration of the compound to be tested, xmeaSured represents a mass concentration of the compound to be tested, and MmeaSured represents a molar mass of the compound to be tested.

[0042] According to the Lambert-Beer law, a molar absorption coefficient 8 refers to an absorbance of a 1 mol / L solution with a layer thickness of 1 cm, and compounds with the same concentration and a same chromophore should theoretically have the same absorbance. In high-performance liquid chromatography, the concentration is proportional to the peak area. For a plurality of compounds with a same chromophore, a molar concentration standard curve of a compound can be used to quantify other compounds.

[0043] A standard curve with a mass concentration as x and a peak area as y is established with a standard. According to the conversion of a mass concentration into a molar concentration, the standard curve is converted into a standard curve with a molar concentration as C and a peak area as y. Based on the principle that compounds with a same chromophore have a same absorbance when at a same molar concentration, a slope of the standard curve is converted to obtain mass concentration standard curves for other compounds, and then the converted standard curves are used to implement the quantification of other compounds. In this way, the compounds with the same chromophore are quantified by the single-standard curve method.

[0044] A specific equation deduction process is as follows:

[0045] Assuming that a sample includes i (i = 1, 2, 3, ...., m, ...., k), components and structures of these components all include a same chromophore, a compound r is selected as an internal reference substance to establish a standard curve of mass concentrations and contents: yr = axr (mg i m! ) ■ b

[0046] According to an equation for converting a mass concentration into a molar concentration: Cr(mmol / L) = = -............=---1---xr(mg / mL) v(Z) Mr*v(L) 1000 *M

[0047] the following equation is obtained: Xr(mg I mL) -1000 * Cr(mmol IL)

[0048] A standard curve of molar concentrations and peak areas for the internal reference substance is established: yr = a * 1000*Mr * Cr(mmol I L) + b

[0049] Compounds with a same chromophore have a same absorbance value at a same molar concentration, that is: When Cr(mmol IL) — ^measured (mmol IL) — * ^measured / mL) 1 vuv ivl measure(i y measured = Jr = a* 1000* Mr* G (mmo / / L) + b

[0050] A standard curve of mass concentrations and peak areas for a compound to be tested is established: y measured = 77~---^measured I mL) + b 1 measured where Mr and Mmeasured represent molar masses of the internal reference substance r and the compound to be tested, respectively.

[0051] Based on this equation, contents of other components can be calculated successively.

[0052] The technical solutions provided by the present disclosure are described in detail below with reference to examples, but the examples cannot be understood as limiting the protection scope of the present disclosure. Example 1

[0053] Table 1 Sources of raw materials adopted for Example 1 and experiments. No. Batch No. Producing area Collection time RSI 180707 Fusong, Jilin 2018.7 RS2 180708 Fusong, Jilin RS3 180709 Fusong, Jilin RS4 180710 Fusong, Jilin RS5 180906 Dunhua, Jilin 2018.9 RS6 180910 Dunhua, Jilin RS7 20190601 Changbai Mountain, Jilin 2019.6 RS8 20190602 Changbai Mountain, Jilin RS9 20190603 Changbai Mountain, Jilin RS10 20190605 Changbai Mountain, Jilin

[0054] Preparation of standard solutions: A ginsenoside Rgl was weighed accurately and dissolved in methanol to prepare a 1 mg / mL stock solution, and the stock solution was stored at 4°C for later use. The stock solution was diluted with methanol 125-fold, 100-fold, 20-fold, 10-fold, 5-fold, 2.5-fold, and 2-fold to produce Rgl solutions with concentrations of 0.008 mg / mL, 0.01 mg / mL, 0.05 mg / mL, 0.1 mg / mL, 0.2 mg / mL, 0.4 mg / mL, and 0.5 mg / mL, respectively.

[0055] Preparation of a test sample solution: A ginseng decoction piece was crushed and then sieved through a 50-mesh sieve to obtain a medicinal powder. 0.5 g of the medicinal powder was weighed out, and then added into 15 mL of methanol. The resulting material was subjected to ultrasonic extraction for 30 min to obtain an extraction system. The extraction system was filtered through a filter paper to obtain a filter residue and a filtrate. 15 mL of methanol was added to the filter residue again, and a resulting material was subjected to ultrasonic extraction for 30 min, and then filtered again, and the filter paper was rinsed with 10 mL of methanol 3 times. Two filtrates were combined and subjected to rotary evaporation at 50°C until a dry extract was obtained. The dry extract was dissolved in 10 mL of methanol to obtain an extract solution. The extract solution was centrifuged at 10,000 rpm for 10 min and then filtered through a 0.22 microporous filter membrane to obtain the test sample solution.

[0056] Chromatographic conditions: A chromatographic column was BEH C18 (100 mm *2.1 mm, 1.7 pm). Gradient elution was conducted with acetonitrile (A)-water (B) as a mobile phase: 0 min to 8 min: 19% of A; 8 min to 13 min: 19% to 29% of A; 13 min to 16 min: 29% of A; 16 min to 23 min: 29% to 40% of A; and 23 min to 26 min: 40% to 19% of A. A flow rate was 0.3 mL min’1. A column temperature was 30°C. A detection wavelength was 203 nm. An injection volume was 2 pL. A sample chamber temperature was 10°C.

[0057] 2 pL of both the test sample solution and each of the single-standard (Rgl) serial solutions were accurately pipetted and injected into a high-performance liquid chromatograph for chromatographic analysis. With the detection methods established by the present disclosure, contents of six ginsenosides in the ginseng decoction piece were calculated.

[0058] Results: A standard linear equation for Rgl was y = 1359.5 x + 1.9403. Calculation linear equation for the remaining components, established according to the following equation, were shown in Table 2. Quantitative results obtained using the molar absorption coefficient method were shown in Table 3. y measured = T,........................................X 13 59-^measured (mS / +1-9403 , measured Mr represents a molar mass of the ginsenoside Rgl and Mmeasured represents a molar mass of any of the other five ginsenosides (Re, Rf, Rbl, Rb2, and Rd).

[0059] Apeak areay, as measured by an instrument, was substituted into the equation y = 1359.5 x + 1.9403 to calculate the mass concentration x of Rgl (mg / mL). Then, according to the preparation method of the test sample solution, the mass concentration was multiplied by a volume of 10 mL for the final test sample solution and divided by the mass of 0.5 g for the medicinal powder to obtain a percentage content of Rgl in the medicinal powder, namely, the data in Table 3.

[0060] Table 2 Calculated standard curve equations for the five ginsenosides in Example 1. Component Calculated Standard curve equation Re y=l 149.7x+l.9403 Rf y=1359.5x+1.9403 Rbl y=981.7x+1.9403 Rb2 y=1009x+1.9403 Rd y=l 130.6x+l.9403

[0061] Table 3 Quantitative results of the detection method of the present disclosure in Example 1. Ginseng decoction piece sample No. RSI RS2 RS3 RS4 RS5 RS6 RS7 RS8 RS9 RS10 Rgl Content / % 0.192 0.144 0.102 0.151 0.161 0.172 0.25 0.206 0.21 0.245 Re % 0.147 0.096 0.087 0.121 0.113 0.154 0.129 0.196 0.179 0.183 Rf % 0.057 0.049 0.036 0.053 0.051 0.054 0.073 0.072 0.072 0.085 Rbl % 0.254 0.167 0.155 0.188 0.188 0.223 0.229 0.242 0.241 0.304 Rb2 % 0.117 0.08 0.062 0.101 0.083 0.122 0.086 0.113 0.097 0.131 Rd % 0.061 0.038 0.037 0.055 0.039 0.046 0.03 0.031 0.036 0.038 Comparative Example 1

[0062] Preparation of standard solutions: Ginsenosides Rgl, Re, Rf, Rbl, Rb2, and Rd were accurately weighed and dissolved in methanol to prepare a 1 mg / mL stock solution, and the stock solution was stored at 4°C for later use. The standard stock solutions of Rgl, Re, Rf, Rbl, Rb2, and Rd ginsenoside were mixed and then diluted 125-fold, 100-fold, 20-fold, 10-fold, 5-fold, 2.5-fold, and 2-fold.

[0063] Preparation of test sample solutions: ginseng decoction pieces was crushed and then sieved through a 50-mesh sieve to obtain medicinal powder. 0.5 g of the medicinal powder was weighed out, and then added into 15 mL of methanol. The resulting material was subjected to ultrasonic extraction for 30 min to obtain an extraction system. The extraction system was filtered to obtain the filter residue and the filtrate. 15 mL of methanol was added to the filter residue again, and a resulting material was subjected to ultrasonic extraction for 30 min, and then filtered again. A filter paper was rinsed with 10 mL of methanol 3 times. Two filtrates were combined and subjected to rotary evaporation at 50°C until a dry extract was obtained. The dry extract was dissolved in 10 mL of methanol to obtain an extract solution. The extract solution was centrifuged at 10,000 rpm for 10 min and then filtered through a 0.22 microporous filter membrane to obtain the test sample solution.

[0064] Chromatographic conditions: A chromatographic column was BEH C18 (100 mm x 2.1 mm, 1.7 pm). Gradient elution was conducted with acetonitrile (A)-water (B) as a mobile phase: 0 min to 8 min: 19% of A; 8 min to 13 min: 19% to 29% of A; 13 min to 16 min: 29% of A; 16 min to 23 min: 29% to 40% of A; and 23 min to 26 min: 40% to 19% of A. The flow rate was 0.3 mLmin’1. A column temperature was 30°C. A detection wavelength was 203 nm. An injection volume was 2 pL. A sample chamber temperature was 10°C.

[0065] 2 pL of both the test sample solution and each of the mixed standard (Rgl, Re, Rf, Rbl, Rb2, and Rd) serial solutions were accurately pipetted and injected into a high-performance liquid chromatograph for chromatographic analysis. The contents of six ginsenosides in the ginseng decoction piece were determined by a multi-standard curve method.

[0066] Results: A standard curve equation for each component was shown in Table 4. The comparison between results of the multi-standard curve determination in Comparative Example 1 and results of the single-standard curve determination in Example 1 is shown in Table 5. A peak area y, as measured by an instrument, was substituted into the equation y=1359.5x+1.9403 to calculate the mass concentration x (mg / mL) of Rgl. Then, according to the preparation method of the test sample solution, the mass concentration was multiplied by a volume of 10 mL for the final test sample solution and divided by the mass of 0.5 g for the medicinal powder to obtain a percentage content of Rgl in the medicinal powder. Percentage contents of other ginsenosides in the medicinal powder were calculated by the same method as for Rgl. Content data for Comparative Example 1 is shown in Table 5. Differences between the results of the single-standard curve determination in Example 1 and the results of the multi-standard curve determination in Comparative Example I were reflected by relative standard deviations (RSDs) / %.

[0067] Table 4 Standard curve equations for the six ginsenosides determined in Comparative Example 1. Component Linear equation R2 Linear range (mg / mL) Rgl y=1359.5x+1.9403 0.9998 0.0078872-0.49295 Re y=1096.8x+5.821 0.9991 0.0079264-0.4954 Rf y=1402.7x+1.1814 0.9993 0.0078824-0.49265 Rbl y=1009x-0.4386 0.9997 0.0079104-0.4944 Rb2 y=1025.3x+1.7361 0.9992 0.00788-0.4925 Rd y=1053.4x+3.9688 0.9998 0.0078832-0.4927

[0068] Table 5 Comparison between results of the multi-standard curve determination in Comparative Example 1 and results of the single-standard curve determination in Example 1. Ginseng Decoction Piece Sample No. RSI RS2 RS3 RS4 RS5 RS6 RS7 RS8 RS9 RS10 Rgl Content / % 0.192 0.144 0.102 0.151 0.161 0.172 0.25 0.206 0.21 0.245 Re Example 1 / % 0.147 0.096 0.087 0.121 0.113 0.154 0.129 0.196 0.179 0.183 Comparative 0.147 0.092 0.084 0.119 0.111 0.153 0.127 0.197 0.179 0.184 Example 1 / % RSD / % 0.078 2.667 2.398 1.478 1.393 0.103 1.263 0.184 0.301 0.307 Rf Example 1 / % 0.057 0.049 0.036 0.053 0.051 0.054 0.073 0.072 0.072 0.085 Comparative Example 1 / % 0.056 0.048 0.036 0.053 0.05 0.053 0.072 0.071 0.07 0.083 RSD / % 0.86 0.628 0.083 0.76 0.695 0.787 1.151 1.131 1.122 1.302 Rbl Example 1 / % 0.254 0.167 0.155 0.188 0.188 0.223 0.229 0.242 0.241 0.304 Comparative Example 1 / % 0.251 0.167 0.155 0.187 0.188 0.222 0.227 0.24 0.239 0.301 RSD / % 0.627 0.043 0.201 0.172 0.176 0.451 0.485 0.563 0.557 0.843 Rb2 Example 1 / % 0.117 0.08 0.062 0.101 0.083 0.122 0.086 0.113 0.097 0.131 Comparative Example 1 / % 0.115 0.08 0.061 0.1 0.082 0.12 0.085 0.111 0.096 0.129 RSD / % 0.893 0.785 0.679 0.856 0.797 0.903 0.807 0.885 0.844 0.919 Rd Example 1 / % 0.061 0.038 0.037 0.055 0.039 0.046 0.03 0.031 0.036 0.038 Comparative Example 1 / % 0.061 0.037 0.036 0.055 0.038 0.046 0.029 0.029 0.035 0.037 RSD / % 0.778 1.842 0.036 0.288 1.737 0.632 3.752 3.525 2.186 1.958

[0069] It can be seen from the above results that, compared with the traditional multi-standard curve detection method in Comparative Example 1, the detection method in Example 1 of the present disclosure can simultaneously detect the contents of six chemical components (ginsenosides Rgl, Re, Rf, Rbl, Rb2, and Rd) with the same chromophore in ginseng, and can lead to similar results to the traditional multi-standard curve detection method in Comparative Example 1, being able to be used for the accurate determination of the six components in ginseng. Comparative Example 2

[0070] Preparation of standard solutions: Ginsenosides Rgl, Re, Rf, Rbl, Rb2, and Rd were accurately weighed and dissolved in methanol to prepare a 1 mg / mL stock solution, and the stock solution was stored at 4°C for later use. The standard stock solutions of Rgl, Re, Rf, Rbl, Rb2, and Rd ginsenoside were mixed and then diluted 125-fold, 100-fold, 20-fold, 10-fold, 5-fold, 2.5-fold, and 2-fold.

[0071] Preparation of a test sample solution: A ginseng decoction piece was crushed and then sieved through a 50-mesh sieve to obtain a medicinal powder. 0.5 g of the medicinal powder was weighed out and then added into 15 mL of methanol. The resulting material was subjected to ultrasonic extraction for 30 min to obtain an extraction system. The extraction system was filtered to obtain a filter residue and a filtrate, 15 mL of methanol was added to the filter residue again, a resulting material was subjected to ultrasonic extraction for 30 minutes and then filtered again. A filter paper was rinsed with 10 mL of methanol 3 times. Two filtrates were combined and subj ected to rotary evaporation at 50°C until a dry extract was obtained. The dry extract was dissolved in 10 mL of methanol to obtain an extract solution. The extract solution was centrifuged at 10,000 rpm for 10 min and then filtered through a 0.22 microporous filter membrane to obtain the test sample solution.

[0072] Chromatographic conditions: A chromatographic column was BEH C18 (100 mm x 2.1 mm, 1.7 pm). Gradient elution was conducted with acetonitrile (A)-water (B) as a mobile phase: 0 min to 8 min: 19% of A; 8 min to 13 min: 19% to 29% of A; 13 min to 16 min: 29% of A; 16 min to 23 min: 29% to 40% of A; and 23 min to 26 min: 40% to 19% of A. The flow rate was 0.3 mLmin’1. A column temperature was 30°C. A detection wavelength was 203 nm. An injection volume was 2 pL. A sample chamber temperature was 10°C.

[0073] 2 pL of both the test sample solutions and a mixed standard (RgL Re, Rf, Rbl, Rb2 and Rd) serial solutions were accurately pipetted. Contents of six ginsenosides in the Ginseng decoction piece were calculated by the SSDMC method.

[0074] Determination of relative correction factor: With Rgi as a single reference standard, the correction factor (RCF) of five components Re, Rf, Rbl, Rb2 and RD at each concentration point were calculated according to formula @, and then the relative correction factor of each component (NRCF) were obtained by taking the average value according to formula RCFx=--------(x=l~n) Anmredxiy-measuKax XjX

[0075] NRCF = %RCFX n

[0076] A peak area measured by an instrument, the relative correction factor in Table 6, and a mass concentration x (mg / mL) calculated for Rgi were substituted into the following equation: ^measured NRCF X Ameasured

[0077] where Ar and xr represent a peak area and a mass concentration of Rgi, respectively, Ameasured and Xmeaswed represent a peak area and a mass concentration of other five ginsenosides, respectively, and NRCF represents the relative correction factor of each of other five ginsenosides (Table 6). Then, the mass concentrations ^measured of other five ginsenosides was obtained by the above steps. And, according to the preparation method of the test sample solution, the mass concentration was multiplied by a volume of 10 mL for the final test sample solution and divided by the mass of 0.5 g for the medicinal powder to obtain percentage contents of other five ginsenosides in the medicinal powder, namely, showing in the data of Table 7.

[0078] Table 6 Relative correction factor of Re RK Rb L Rb2andRd. Component Re Rf Rbl Rb2 Rd Relative correction factor 1.15 0.97 1.37 1.32 1.22

[0079] Table 7 Comparison between detection results of the multi-standard curve method in Comparative Example 1 and detection results of the SSDMC method in Comparative Example 2. Ginseng Decoction Piece Sample No. RSI RS2 RS3 RS4 RS5 RS6 RS7 RS8 RS9 RS10 Rgi Content% 0.192 0.144 0.102 0.151 0.161 0.172 0.250 0.206 0.210 0.245 Re SSDMC method% 0.144 0.094 0.085 0.119 0.111 0.150 0.127 0.191 0.175 0.179 Multi-standard curve method% 0.147 0.093 0.084 0.120 0.112 0.154 0.128 0.199 0.181 0.185 RSD% 1.590 0.798 0.988 0.765 0.272 1.896 0.606 2.710 2.333 2.304 Rf SSDMC method% 0.057 0.049 0.036 0.053 0.051 0.054 0.073 0.071 0.071 0.084 Multi-standard curve method% 0.056 0.048 0.036 0.053 0.050 0.053 0.072 0.071 0.070 0.083 RSD% 0.876 0.866 1.129 0.735 0.918 0.862 0.669 0.534 0.561 0.430 Rbl SSDMC method% 0.252 0.168 0.153 0.189 0.188 0.223 0.231 0.243 0.243 0.304 Multi-standard curve method% 0.251 0.169 0.155 0.189 0.189 0.223 0.230 0.242 0.242 0.302 RSD% 0.079 0.396 0.956 0.292 0.205 0.076 0.279 0.128 0.141 0.333 Rb2 SSDMC method% 0.116 0.082 0.062 0.102 0.084 0.121 0.088 0.114 0.099 0.131 Multi-standard curve method% 0.115 0.081 0.061 0.101 0.083 0.121 0.086 0.112 0.097 0.130 RSD% 0.657 1.183 1.508 0.673 1.242 0.451 1.582 0.784 1.131 0.655 Rd SSDMC method% 0.064 0.042 0.040 0.058 0.042 0.049 0.034 0.035 0.040 0.041 Multi-standard curve method% 0.061 0.038 0.036 0.055 0.038 0.046 0.029 0.030 0.036 0.037 RSD% 2.760 7.183 7.035 3.376 7.145 5.228 11.145 10.582 8.226 7.956

[0080] It can be seen from the results of Comparative Example 2 (see Table 7) that compared with the detection results of the existing SSDMC method (see Table 5 and FIG. 3), the method for simultaneously detecting sixth chemical components (Rgl Re^ RE Rb K Rb2, and RDRd) in ginseng according to example 1 of the present disclosure shows more similar detection results to those of the conventional multi-standard curve method. That is, the RSD% between the detection results of the detection method of the present disclosure and those of the multi-standard curve method is equal to or less than the RSD% between the detection results of the SSDMC method and those of the multi-standard curve method (see Table 5, Table 7, and FIG. 3), indicating that the method in Example 1 of the present disclosure provides more accurate quantitative results, with small errors. Further, the detection method according to the present disclosure does not require the determination of the relative correction factor, saving workload and improving efficiency. Example 2

[0081] Table 8 Sources of raw materials adopted for Example 2 and experiments. No. Batch No. Producing area Collection time SI 180705 Lingchuan, Shanxi 2018.7 S2 180706 Lingchuan, Shanxi S3 180707 Lingchuan, Shanxi S4 180908 Changzhi, Shanxi S5 180909 Changzhi, Shanxi 2018.9 S6 180910 Changzhi, Shanxi S7 20190601 Chengde, Hebei 2019.6 S8 20190602 Chengde, Hebei S9 20190604 Chengde, Hebei S10 20190605 Chengde, Hebei

[0082] Preparation of standard solutions: Wogonoside was weighed accurately and dissolved in dimethyl sulfoxide (DMSO) to prepare a 1 mg / mL stock solution, and the stock solution was stored at 4°C for later use. The stock solution was diluted with DMSO 1,000-fold, 200-fold, 100-fold, 50-fold, 25-fold, 20-fold, 12.5-fold, 10-fold, 5-fold, 3.33-fold, and 2.5-fold to produce wogonoside solutions with concentrations of 1 pg / mL, 5 pg / mL, 10 pg / mL, 20 pg / mL, 40 pg / mL, 50 pg / mL, 80 pg / mL, 100 pg / mL, 200 pg / mL, 300 pg / mL, and 400 pg / mL, respectively.

[0083] Preparation of a test sample solution: A Scutellariae Radix decoction piece was crushed and then sieved through a 60-mesh sieve to obtain a medicinal powder. 0.1 g of the medicinal powder was weighed out, 50 mL of 70% methanol was added, and ultrasonic extraction was conducted for 15 min (room temperature, 40 kHz) to obtain an extraction system. The extraction system was centrifuged at 10,000 rpm for 10 min to obtain a supernatant, and then the supernatant was filtered through a 0.22 microporous filter membrane to obtain the test sample solution.

[0084] Chromatographic conditions: A chromatographic column was HSS T3 (100 mm * 2.1 mm, 1.8 pm). Gradient elution was conducted with 0.2% phosphoric acid aqueous solution (A)-acetonitrile (B) as a mobile phase: 0 min to 2 min: 78% to 75% of A; 2 min to 4 min: 75% of A; 4 min to 6 min: 75% to 68% of A; 6 min to 7 min: 68% to 60% of A; 7 min to 8 min: 60% of A; 8 min to 10 min: 60% to 50% of A; 10 min to 13 min: 50% to 5% of A; and 13 min to 17 min: 5% to 78% of A. A flow rate was 0.35 mL min’1. A column temperature was 30°C. A detection wavelength was 280 nm. An inj ection volume was 1 pL. A sample chamber temperature was 10°C.

[0085] 1 pL of both the test sample solution and each of the single-standard (wogonoside) serial solutions were accurately pipetted and injected into a high-performance liquid chromatograph for chromatographic analysis. Contents of three flavonoids in the Scutellariae Radix decoction piece were calculated by the single-standard curve method established by the present disclosure.

[0086] Results: A standard linear equation for wogonoside was y = 5.5879 x - 5.2928. Standard curves for the remaining components that were established according to the following equation are shown in Table 9. Quantitative results of contents are shown in Table 10: y—ed = —-----x—ed ! mL) - 5.2928 (6) 1 measured

[0087] Mr represents a molar mass of wogonoside and Mmeasured represents a molar mass of each of the other two flavonoids (baicalin and baicalein).

[0088] A content of wogonoside was calculated by a peak area of wogonoside: A peak area y, as measured by an instrument, was substituted into the equation y = 5.5879 x - 5.2928 to calculate the mass concentration x (mg / mL) of wogonoside. Then, according to the preparation method of the test sample solution, the mass concentration was multiplied by a volume of 50 mL for the final test sample solution and divided by the mass of 0.1 g for the medicinal powder to obtain a percentage content of wogonoside in the medicinal powder.

[0089] Contents of baicalin and baicalein were calculated as follows: Firstly, referring to the standard linear equation y = 5.5879 x - 5.2928 for wogonoside, according to the equation (6), the standard curve equation for the remaining components was converted (as shown in Table 9). Peak areas y, as measured by an instrument for baicalin and baicalein, were substituted into respective curves in Table 9 to calculate mass concentrations x of baicalin and baicalein (mg / mL), respectively. Then, according to the preparation method of the test sample solution, the mass concentrations each were multiplied by a volume of 50 mL for the final test sample solution and divided by the mass of 0.1 g for the medicinal powder to obtain percentage contents of baicalin and baicalein in the medicinal powder.

[0090] Table 9 Calculated standard curve equations for the three flavonoids in Example 2. Component Calculated standard curve equation Wogonoside y=5.5879x-5.2928 Baicalin y=5.7637x-5.2928 Baicalein y=9.5199x-5.2928 [0091 ] Table 10 Quantitative results of the detection method of the present disclosure in Example 2. Ginseng Decoction Piece Sample No. SI S2 S3 S4 S5 S6 S7 S8 S9 S10 Wogon oside Content / % 3.274 3.044 3.282 2.825 3.036 3.561 3.111 2.917 3.110 3.149 Baicali Quantitati 13.314 12.678 13.609 12.902 14.975 15.970 13.531 11.966 13.212 13.970 n ve content / % Bai cal e in Quantitati ve content / % 0.558 0.448 0.554 0.723 0.415 0.300 0.265 0.311 0.225 0.424 Comparative Example 3

[0092] Preparation of standard solutions: Wogonoside, baicalin, and baicalein each were accurately weighed. Wogonoside was dissolved in DMSO and baicalin and baicalein each were dissolved in methanol to prepare 1 mg / mL stock solutions, and the stock solutions were stored at 4°C for later use. The stock solutions of wogonoside, baicalin, and baicalein were diluted separately. The wogonoside stock solution was diluted 1,000-fold, 200-fold, 100-fold, 50-fold, 25-fold, 20-fold, 12.5-fold, 10-fold, 5-fold, 3.33-fold, and 2.5-fold. The baicalin stock solution was diluted 200-fold, 100-fold, 12.5-fold, 5-fold, 3.33-fold, and 2.5-fold. The baicalein stock solution was diluted 1,000-fold, 500-fold, 250-fold, 100-fold, 66.67-fold, and 50-fold.

[0093] Preparation of a test sample solution: A Scutellariae Radix decoction piece was crushed and then sieved through a 60-mesh sieve to obtain a medicinal powder. 0.1 g of the medicinal powder was weighed out and then added into 50 mL of 70% methanol. The resulting material was subjected to ultrasonic extraction for 15 min (room temperature, 40 kHz) to obtain an extraction system. The extraction system was centrifuged at 10,000 rpm for 10 min to obtain a supernatant. The supernatant was filtered through a 0.22 microporous filter membrane to obtain the test sample solution.

[0094] Chromatographic conditions: A chromatographic column was HSS T3 (100 mm x 2.1 mm, 1.8 pm). Gradient elution was conducted with 0.2% phosphoric acid aqueous solution (A)-acetonitrile (B) as a mobile phase: 0 min to 2 min: 78% to 75% of A; 2 min to 4 min: 75% of A; 4 min to 6 min: 75% to 68% of A; 6 min to 7 min: 68% to 60% of A; 7 min to 8 min: 60% of A; 8 min to 10 min: 60% to 50% of A; 10 min to 13 min: 50% to 5% of A; and 13 min to 17 min: 5% to 78% of A. A flow rate was 0.35 mL min'1. A column temperature was 30 °C. A detection wavelength was 280 nm. An injection volume was 1 pL. A sample chamber temperature was 10°C.

[0095] 1 pL of both the test sample solution and each of the standard (wogonoside, baicalin, and baicalein) serial solutions were accurately pipetted and injected into a high-performance liquid chromatograph for chromatographic analysis. Contents of three flavonoids in the Scutellariae Radix decoction piece were calculated by the multi-standard curve method.

[0096] Results: A standard curve equation for each component is shown in Table 11. The comparison between results of the multi-standard curve determination in Comparative Example 3 and results of the detection method in Example 2 of the present disclosure is shown in Table 12.

[0097] Table 11 Standard curve equations for the three flavonoids determined in Comparative Example 3. Component Linear equation R2 Linear range (ug / mL) Wogonoside y=5.5879x-5.2928 0.9999 0.985-394 Baicalin y=5.8682x-17.19 0.9991 4.77-381.6 Baicalein y=9.0546x-2.7957 0.9990 0.985-19.7

[0098] Table 12 Comparisons between results of the multi-standard curve determination in Comparative Example 3 and results of the detection method in Example 3 of the present disclosure. Scutellariae Radix decoction piece Sample No. SI S2 S3 S4 S5 S6 S7 S8 S9 S10 Wogonoside Content / % 3.274 3.044 3.282 2.825 3.036 3.561 3.111 2.917 3.110 3.149 Baicalin Example 2 / % 13.314 12.678 13.609 12.902 14.975 15.970 13.531 11.966 13.212 13.970 Comparative Example 3 / % 13.177 12.553 13.467 12.772 14.809 15.786 13.391 11.854 13.077 13.821 RSD / % 0.728 0.702 0.740 0.713 0.789 0.816 0.736 0.664 0.726 0.754 Baicalein Example 2 / % 0.558 0.448 0.554 0.723 0.415 0.300 0.265 0.311 0.225 0.424 Comparative Example 3 / % 0.573 0.458 0.569 0.747 0.422 0.302 0.265 0.313 0.223 0.432 RSD / % 1.873 1.464 1.863 2.262 1.288 0.394 0.024 0.499 0.651 1.337

[0099] It can be seen from Comparative Example 3 that, compared with the conventional multi- standard curve method, the detection method provided in Example 2 of the present disclosure can simultaneously detect the contents of three chemical components (wogonoside, baicalin, and baicalein) with the same chromophore in Scutellariae Radix, and results all have RSDs / % of less than 3%, indicating that the method of the present disclosure can lead to accurate results when used in the detection of three flavonoids with a same chromophore in Scutellariae Radix. Comparative Example 4

[0100] Preparation of standard solutions: Wogonoside, baicalin, and baicalein each were accurately weighed. Wogonoside was dissolved in DMSO and baicalin and baicalein each were dissolved in methanol to prepare 1 mg / mL stock solutions, and the stock solutions were stored at 4°C for later use. The standard stock solutions of wogonoside, baicalin, and baicalein were diluted separately. The wogonoside stock solution was diluted 1,000-fold, 200-fold, 100-fold, 50-fold, 25-fold, 20-fold, 12.5-fold, 10-fold, 5-fold, 3.33-fold, and 2.5-fold. The baicalin stock solution was diluted 200-fold, 100-fold, 12.5-fold, 5-fold, 3.33-fold, and 2.5-fold. The baicalein stock solution was diluted 1,000-fold, 500-fold, 250-fold, 100-fold, 66.67-fold, and 50-fold.

[0101] Preparation of a test sample solution: A Scutellariae Radix decoction piece was crushed and then sieved through a 60-mesh sieve to obtain a medicinal powder. 0.1 g of the medicinal powder was weighed out and then added into 50 mL of 70% methanol. The resulting material was subjected to ultrasonic extraction for 15 min (room temperature, 40 kHz) to obtain an extraction system. The extraction system was centrifuged at 10,000 rpm for 10 min to obtain a supernatant. The supernatant was filtered through a 0.22 microporous filter membrane to obtain the test sample solution.

[0102] Chromatographic conditions: A chromatographic column was HSS T3 (100 mm * 2.1 mm, 1.8 um). Gradient elution was conducted with 0.2% phosphoric acid aqueous solution (A)-acetonitrile (B) as a mobile phase: 0 min to 2 min: 78% to 75% of A; 2 min to 4 min: 75% of A; 4 min to 6 min: 75% to 68% of A; 6 min to 7 min: 68% to 60% of A; 7 min to 8 min: 60% of A; 8 min to 10 min: 60% to 50% of A; 10 min to 13 min: 50% to 5% of A; and 13 min to 17 min: 5% to 78% of A. A flow rate was 0.35 mL min’1. A column temperature was 30°C. A detection wavelength was 280 nm. An inj ection volume was 1 pL. A sample chamber temperature was 10°C.

[0103] 1 pL of both the test sample solution and each of the standard (wogonoside, baicalin, and baicalein) serial solutions were accurately pipetted and injected into a high-performance liquid chromatograph for chromatographic analysis. Contents of three flavonoids in the Scutellariae 24 Radix decoction piece were calculated by the multi-standard curve method.

[0104] Determination of RCFs: With wogonoside as a single reference standard, the RCFs for baicalin and baicalein were calculated according to the following equation: kr ppp _ 1V1 k x measured

[0105] where kr represents a slope of a standard curve for wogonoside and kmeasured represents a slope of a standard curve for each of baicalin and baicalein. Results are shown in Table 13.

[0106] A peak area measured by an instrument, RCF in Table 13, and a mass concentration x (mg / mL) calculated for wogonoside were substituted into the following equation: RCFxA X _measured measured * / Ar / Xr where Ar and xr represent a peak area and a mass concentration of wogonoside, respectively, A measured and Xmeasured represent a peak area and a mass concentration of baicalin or baicalein, respectively, and RCF represents the relative correction factor of baicalin or baicalein (Table 13).

[0107] Then, according to the preparation method of the test sample solution, the mass concentrations xmeasure<i of baicalin and baicalein obtained by the above steps each were multiplied by a volume of 50 mL for the final test sample solution and divided by the mass of 0.1 g for the medicinal powder to obtain percentage contents of baicalin and baicalein in the medicinal powder, namely, showing in the data of Table 14.

[0108] Table 13 Relative correction factor (RCF) of baicalin and baicalein. Component Baicalin Baicalein RCF 1.03 0.62

[0109] Table 12 Comparison between detection results of the multi-standard curve method in Comparative Example 3 and detection results of the SSDMC method in Comparative Example 4 Scutellariae Radix decoction piece Sample No. RSI RS2 RS3 RS4 RS5 RS6 RS7 RS8 RS9 RS10 Wogonosid e Content / % 3.274 3.044 3.282 2.825 3.036 3.561 3.111 2.917 3.110 3.149 Baicalin SSDMC method / % 14.28 8 13.61 8 14.60 5 13.87 6 16.09 4 17.12 9 14.53 3 12.86 0 14.18 9 15.00 3 Multistandard curve method / % 13.17 7 12.55 3 13.46 7 12.77 2 14.80 9 15.78 6 13.39 1 11.85 4 13.07 7 13.82 1 RSD / % 5.718 5.742 5.741 5.836 5.885 5.809 5.788 5.764 5.765 5.782 Baicalein SSDMC method / % 0.566 0.449 0.562 0.744 0.413 0.290 0.254 0.302 0.211 0.423 Multistandard curve method / % 0.573 0.458 0.569 0.747 0.422 0.302 0.265 0.313 0.223 0.432 RSD / % 0.894 1.167 0.995 0.390 1.511 2.345 3.004 2.510 3.936 2.308

[0110] It can be seen from Comparative Example 4 that, compared with the detection results of the conventional multi-standard curve method, detection results obtained by the existing SSDMC method have larger RSD / % than those by the detection method in Example 2 of the present disclosure (as shown in Table 12, Table 14, and FIG. 6). The method for simultaneously detecting three chemical components (wogonoside, baicalin, and baicalein) with identical chromophore in Scutellariae Radix provided by the present disclosure has detection results that are more similar to those of the multi-standard curve method, indicating that the method provides accurate quantitative results with small errors.

[0111] It can be seen from the above examples that the detection method of Example 2 provided in the present disclosure simultaneously detects the contents of three chemical components (wogonoside, baicalin, and baicalein) with the same chromophore in Scutellariae Radix. The comparison of the results with the multi-standard curve method (Table 12) proves that the detection 26 method according to the present disclosure is accurate in detecting the three components in Scutellariae Radix. Compared with the existing SSDMC method, the detection method of the present disclosure yield results that are closer to the quantitative results by the multi-standard curve method, indicating that the quantitative results of the detection method of the present disclosure are more accurate. As shown in Tables 12 and 14, and FIG. 6, it further illustrates that the method established in the present disclosure shows higher accuracy than the existing method and does not require determination of RCFs or multiple standard curves, saving workload and improving efficiency.

[0112] Table 15 Peak area values for components in Examples 1 and 2. Peak area Sam pie No. Wogonos ide Baicalin Baicale in pie No. Rgl Re Rf Rbl Rb2 RD SI 362.8 1538.6 101.7 RSI 135.1 88.3 41.3 128.9 61.9 36.9 S2 338.0 1469.3 80.9 RS2 100.6 57.4 35.4 85.6 43.3 23.9 S3 364.4 1576.0 101.0 RS3 72.9 52.6 26.8 79.3 33.6 23.3 S4 313.6 1496.8 133.8 RS4 105.5 72.3 38.5 95.9 53.9 33.4 S5 336.8 1734.7 74.3 RS5 112.9 67.9 36.9 96.1 44.7 24.3 S6 393.9 1841.1 52.0 RS6 120.7 91.6 39.2 113.8 64.5 28.4 S7 344.2 1562.3 45.5 RS7 172.9 76.7 52.1 116.4 46.2 19.4 S8 321.4 1376.9 54.0 RS8 143.5 115.9 51.2 123.0 60.0 19.8 S9 345.8 1533.0 38.1 RS9 145.6 105.7 50.9 122.5 51.8 22.9 S10 349.1 1616.3 76.0 RSI 0 170.5 108.6 60.5 154.2 69.3 23.6

[0113] The above are merely preferred embodiments of the present disclosure. It should be noted that a person of ordinary skill in the art may further make several improvements and modifications without departing from the principles of the present disclosure, but such improvements and modifications should be deemed as falling within the protection scope of the present disclosure.

Claims

1. A method for quantifying a plurality of compounds with a same chromophore using a single standard, comprising the following steps:a) detecting standard working solutions with different concentrations by high-performance liquid chromatography, and with mass concentrations as x-coordinate and peak areas as y-coordinate, plotting a standard curve of the mass concentrations and the peak areas for a standard;b) according to a relationship between the mass concentrations and molar concentrations of the standard, plotting a standard curve of the molar concentrations and the peak areas for the standard; andc) under a condition that a compound to be tested shares the same chromophore as the standard, and that a molar concentration Cmeasured of the compound to be tested is equal to a molar concentration Cr of the standard, an absorbance ymeasured of the compound to be tested is equal to an absorbance yr of the standard, and basing on a relationship between mass concentrations and molar concentrations for the compound to be tested, plot a standard curve of the mass concentrations and peak areas for the compound to be tested:A' / v I T \ 1y =-------ax (mg / mL) + by measured measuredmeasured (5)wherein ymeasured represents a peak area of the compound to be tested, xmeasured represents a mass concentration of the compound to be tested, Mr represents a molar mass of the standard, Mmeasured represents a molar mass of the compound to be tested, and a and b are parameters; andsubstituting the peak area ymeasured of the compound to be tested into the equation (5) to calculate the mass concentration ^measured of the compound to be tested.

2. The method for quantifying the plurality of compounds with the same chromophore using the single standard according to claim 1, wherein the standard curve of the mass concentrations and the peak areas for the standard in step 1) is as follows:yr = axr{mglmL) + bwherein yr represents a peak area of the standard, xr represents a mass concentration of the standard, and the a and the b are the parameters.

3. The method for quantifying the plurality of compounds with the same chromophore using the single standard according to claim 1, wherein the relationship between the mass concentrations28and the molar concentrations of the standard in step 2) is as follows:xSmg I mL) = WQ0* * (J (mmol IL) . r r rwherein xr represents a mass concentration of the standard, Mr represents the molar mass of the standard, and Cr represents a molar concentration of the standard.

4. The method for quantifying the plurality of compounds with the same chromophore using the single standard according to claim 1, wherein the standard curve of the molar concentrations and the peak areas for the standard in the step 2) is as follows:y r = a*M r* C rdnm°l! L) + b q)wherein yr represents a peak area of the standard, Cr represents a molar concentration of the standard, Mr represents the molar mass of the standard, and the a and the b are the parameters.

5. The method for quantifying the plurality of compounds with the same chromophore using the single standard according to claim 1, wherein the relationship between the mass concentrations and the molar concentrations for the compound to be tested in the step 3) is as follows:=--------!--------(nig / measured 1000 * measuredmeasured (4^)wherein Cmeasured represents a molar concentration of the compound to be tested, xmeasured represents the mass concentration of the compound to be tested, and Mmeasured represents a molar mass of the compound to be tested.

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