A method for detecting chemical components in a traditional Chinese medicine composition

Abstract

This invention provides a method for detecting chemical components in a traditional Chinese medicine composition. The method uses ultra-high performance liquid chromatography (UHPLC) to separate and detect 23 characteristic peaks. It can be used for the identification and content determination of a traditional Chinese medicine composition made from gardenia, forsythia, magnolia bark, pinellia, immature bitter orange, poria, perilla stem, and licorice, providing an effective detection means for the quality control and medication safety of the traditional Chinese medicine composition.

Description

Technical Field

[0001] This invention belongs to the field of traditional Chinese medicine technology, and specifically relates to a method for detecting chemical components in a traditional Chinese medicine composition. Background Technology

[0002] Traditional Chinese medicine has a long history and wide application, but due to its complex composition and unclear components, it is limited in many aspects such as mechanism research and quality control.

[0003] In countries without a history of using traditional Chinese medicine (TCM), strict quality standards are typically required for pharmaceuticals to ensure that the active ingredients and their content are clearly defined and controllable. This applies to TCM as well. For example, the U.S. *Industrial Guidelines for Herbal Drug Development* (2016 edition) stipulates that for drugs produced by mixing several medicinal herbs, optimal analytical techniques should be used to identify (qualitatively) and quantify each active ingredient or chemical component. In response to this situation, researchers have been continuously striving to develop quality control methods for TCM products, aiming to solve domestic quality control issues and support the export of TCM products.

[0004] Chinese patent CN100998710A discloses a traditional Chinese medicine composition for treating depression and its preparation method. This composition comprises gardenia, forsythia, magnolia bark, pinellia, immature bitter orange, poria, perilla stem, and licorice. Gardenia is the principal ingredient, with forsythia, magnolia bark, and pinellia as assistant ingredients, perilla stem, immature bitter orange, and poria as adjuvant ingredients, and licorice as the guiding ingredient. The combined effects of these herbs clear stagnant heat and regulate qi, making it suitable for treating depression. This traditional Chinese medicine composition is an innovative prepared medicine with broad market prospects; therefore, reliable testing methods are needed to ensure its safe and controllable quality.

[0005] Chinese patent CN115327018A discloses a method for identifying multiple components in a traditional Chinese medicine composition. This method uses thin-layer chromatography to identify *Citrus aurantium*, *Forsythia suspensa*, *Magnolia officinalis*, *Pinellia ternata*, *Citrus aurantium*, *Poria cocos*, *Perilla frutescens*, and *Glycyrrhiza uralensis* in a traditional Chinese medicine composition. However, this method can only perform qualitative analysis of the composition and cannot provide definitive conclusions. Furthermore, it can only identify three herbs in the composition, resulting in poor reliability for quality control. Summary of the Invention

[0006] The purpose of this invention is to provide a detection method for the above-mentioned traditional Chinese medicine composition, which can simultaneously detect multiple chemical components in the traditional Chinese medicine composition.

[0007] To achieve the above objectives, the inventors have provided the following technical solutions.

[0008] A method for detecting chemical components in a traditional Chinese medicine composition, wherein the raw materials of the traditional Chinese medicine composition are: gardenia, forsythia, magnolia bark, pinellia, immature bitter orange, poria, perilla stem, and licorice, and the detection method uses ultra-high performance liquid chromatography;

[0009] The extraction solvent used in the preparation of the test solution was 50% methanol;

[0010] Chromatographic conditions: Mobile phase A was acetonitrile, mobile phase B was 0.5% phosphoric acid aqueous solution, and gradient elution was used. The gradient elution system is as follows.

[0011] Time (min) Mobile phase A (%) Mobile phase B (%) 0 2 98 5 2 98 26 15 85 40 17 83 50 17 83 60 34 66 61 40 60 70 90 10 71 2 98 81 2 98 .

[0012] In the above detection method, the sample is extracted by ultrasound during the preparation of the sample solution for 30 minutes, and the concentration of the sample solution is 12 mg / mL.

[0013] In the above detection method, the chromatographic conditions are as follows: the chromatographic column is a C18 reversed-phase column with a particle size of 1.7 μm, and the flow rate of the mobile phase is 0.3 mL / min.

[0014] The detection method provided by this invention can be used to detect chemical components in a traditional Chinese medicine composition (JYCF) made from gardenia, forsythia, magnolia bark, pinellia, immature bitter orange, poria, perilla stem, and licorice. The traditional Chinese medicine composition can be in various dosage forms such as capsules, tablets, granules, pills, and powders.

[0015] The composition and preparation method of the traditional Chinese medicine composition (JYCF) described in this invention are the same as those in Chinese Patent CN100998710A. In that patent, Magnolia officinalis is ginger-processed Magnolia officinalis, and Pinellia ternata is ginger-processed Pinellia ternata.

[0016] The traditional Chinese medicine composition of the present invention comprises the following parts by weight: Gardenia 120-200 parts, Forsythia 300-500 parts, Magnolia officinalis 120-200 parts, Pinellia ternata 180-300 parts, Citrus aurantium 120-200 parts, Poria cocos 144-240 parts, Perilla frutescens stem 144-240 parts, and Glycyrrhiza uralensis 36-60 parts.

[0017] The preferred weight composition of the traditional Chinese medicine composition is as follows: 160 parts of gardenia, 400 parts of forsythia, 160 parts of ginger-processed magnolia bark, 240 parts of ginger-processed pinellia, 160 parts of immature bitter orange peel, 192 parts of poria cocos, 192 parts of perilla stem, and 48 parts of licorice.

[0018] Alternatively, the following ingredients may be preferred: 120 parts gardenia, 500 parts forsythia, 120 parts magnolia bark, 300 parts pinellia, 120 parts immature bitter orange, 240 parts poria, 144 parts perilla stem, and 60 parts licorice.

[0019] Alternatively, the following ingredients may be preferred: 200 parts gardenia, 300 parts forsythia, 200 parts magnolia bark, 180 parts pinellia, 200 parts immature bitter orange, 144 parts poria, 240 parts perilla stem, and 36 parts licorice.

[0020] Alternatively, the following ingredients may be preferred: 160 parts gardenia, 480 parts forsythia, 150 parts magnolia bark, 210 parts pinellia, 160 parts immature bitter orange, 200 parts poria, 240 parts perilla stem, and 60 parts licorice.

[0021] The method for preparing the capsule dosage form of the traditional Chinese medicine composition consists of the following steps:

[0022] a. Weigh out the selected and cleaned gardenia, immature bitter orange, ginger, magnolia bark, and forsythia according to the prescription. Crush the gardenia and extract it 1-3 times with 6-10 times the amount of 50-80% ethanol, each time for 1-3 hours. Filter the extract and concentrate it to a clear paste with a density of 1.01-1.06 each time. Combine the concentrates and set aside.

[0023] b. Weigh out the selected Poria cocos, Perilla stem, and Licorice according to the prescription amount, add water and decoct three times. Add 8-10 times the amount of water for the first time, and 6-8 times the amount of water for the second and third times. Each time, decoct for 1-3 hours. Combine the extracts and concentrate them to a clear paste with a density of 1.10-1.15 for later use.

[0024] c. Weigh out the selected and cleaned ginger and pinellia according to the prescription amount, crush them, and pass them through an 80-100 mesh sieve for later use;

[0025] d. Combine the medicinal liquid and extract obtained in steps a and b, use the ginger and pinellia powder obtained in step c as the base material, spray granulate, sort, and fill into capsules to obtain capsules.

[0026] The present invention also provides the application of the above-mentioned detection method in the quality control method of a traditional Chinese medicine composition made from gardenia, forsythia, magnolia bark, pinellia, immature bitter orange, poria, perilla stem, and licorice.

[0027] The quality control methods are the identification and content determination of traditional Chinese medicine composition preparations.

[0028] When the above-mentioned detection is used in the quality control method of the traditional Chinese medicine composition, it also includes the preparation of a reference solution.

[0029] The reference standards used in the preparation of the reference solution include geniposide, genipin disaccharide, hyperoside, forsythoside A, pinoresinin-β-D-glucoside, rutin, hesperidin, forsythoside, ammonium glycyrrhizate, and magnolol. The detection method provided by this invention can separate a total of 23 characteristic peaks, and the reference standards corresponding to the characteristic peaks can be selected according to specific quality control requirements.

[0030] The reference solution was prepared by accurately weighing appropriate amounts of geniposide, and / or genipin disaccharide, and / or hyperoside, and / or forsythoside A, and / or pinoresinin-β-D-glucoside, and / or rutin, and / or hesperidin, and / or forsythoside, and / or ammonium glycyrrhizate, and / or magnolol, and / or honokiol reference standards, mixing them, and dissolving them in an appropriate amount of methanol to prepare solutions with concentrations of 0.106 mg / mL, 0.041 mg / mL, 0.013 mg / mL, 0.311 mg / mL, 0.025 mg / mL, 0.021 mg / mL, 0.007 mg / mL, 0.095 mg / mL, 0.011 mg / mL, 0.009 mg / mL, and 0.015 mg / mL.

[0031] Considering the amount and activity of the chemical components contained in the traditional Chinese medicine composition, the inventors determined the reference standards to be used in the identification and content determination of the detection method, totaling 11 types, namely geniposide, genipin disaccharide, hyperoside, forsythoside A, pinoresinin-β-D glucoside, rutin, hesperidin, forsythoside, ammonium glycyrrhizate, magnolol, and honokiol.

[0032] The detection method provided by this invention can effectively separate 23 characteristic peaks. By comparing the characteristic peaks, it can be used for the identification and content determination of traditional Chinese medicine compositions composed of gardenia, forsythia, magnolia bark, pinellia, immature bitter orange, poria, perilla stem, and licorice. This method can meet higher quality control requirements for the traditional Chinese medicine composition and better ensure the safety of drug use.

[0033] During the research process, the inventors examined multiple conditions involved in the detection method provided by this invention and finally determined the preferred technical solution. The examination included the flow system and detection wavelength in the chromatographic conditions, and the extraction solvent, extraction method, extraction time, and concentration of the test solution in the preparation method of the test sample solution. Some of the examined conditions are shown below.

[0034] Investigation of chromatographic conditions

[0035] (I) Investigation of the mobile phase system

[0036] 1. Sample solution preparation

[0037] Take 0.3g of the contents of JYCFZ capsules, accurately weigh it, place it in a stoppered conical flask, accurately add 25mL of methanol, seal tightly, weigh, sonicate (power 250W, frequency 40kHz) for 25min, cool, weigh again, and then filter through a 0.45μm microporous membrane to obtain the product.

[0038] 2 Initial chromatographic conditions

[0039] The chromatographic column is an ACQUITY UPLC BEH C18, 1.7 μm, 2.1 x 100 mm.

[0040] Flow rate: 0.3 mL / min; Injection volume: 2 μL;

[0041] Detection wavelength: Under the above conditions, the detection spectra of the terminal absorption at 230 nm, 254 nm, and 365 nm were compared. The results showed that, compared with the spectra at 254 nm and 365 nm, the 230 nm spectrum had the most detectable peaks, and the intensity was generally greater. Therefore, the detection wavelength was determined to be 230 nm.

[0042] 3. Mobile phase system

[0043] Different mobile phase systems were investigated under the above conditions.

[0044] The inventors initially set up the mobile phase system 1 shown in Table 1.

[0045] Table 1

[0046] Time (min) Mobile phase A (methanol, %) Mobile phase B (0.5% aqueous phosphoric acid solution, %) 0 2 98 21 17 83 30 17 83 54 34 66 56 40 60 68 90 10 73 90 10

[0047] The chromatogram obtained using mobile phase system 1 showed that the chromatographic peaks were mainly concentrated after 65 min, indicating that the elution ability of the methanol-0.5% phosphoric acid mobile phase system was too weak and unsuitable.

[0048] The mobile phase system 1 shown in Table 1 is improved and adjusted to the mobile phase system 2 shown in Table 2.

[0049] Table 2

[0050] Time (min) Mobile phase A (acetonitrile, %) Mobile phase B (0.5% aqueous phosphoric acid solution, %) 0 2 98 21 17 83 30 17 83 54 34 66 56 40 60 68 90 10 73 90 10

[0051] The spectra obtained using mobile phase system 2 show that the number of peaks and separation effect are significantly improved compared with mobile phase system 1, but there is obvious stacking of peaks in the 15-35 min range.

[0052] The mobile phase system 2 shown in Table 2 is improved and adjusted to the mobile phase system 3 shown in Table 3.

[0053] Table 3

[0054] Time (min) Mobile phase A (acetonitrile, %) Mobile phase B (0.5% aqueous phosphoric acid solution, %) 0 2 98 21 15 85 35 17 83 45 17 83 55 34 66 56 40 60 75 90 10 80 90 10 81 2 98 91 2 98

[0055] The spectrum obtained using mobile phase system 3 showed no significant difference in the number of peaks compared to mobile phase system 2. Peak separation was significantly improved between 15 and 35 min, but the peak separation effect was poor around 10 min.

[0056] The mobile phase system 3 shown in Table 3 is improved and adjusted to the mobile phase system 4 shown in Table 4.

[0057] Table 4

[0058]

[0059]

[0060] The chromatogram obtained using mobile phase system 4 showed no significant difference in the number of peaks compared to mobile phase system 3, indicating a significant improvement in separation efficiency that met the requirements for subsequent analysis. However, no obvious chromatographic peaks were observed after 70 minutes, requiring further adjustments.

[0061] The mobile phase system 4 shown in Table 4 is improved and adjusted to the mobile phase system 5 shown in Table 5.

[0062] Table 5

[0063] Time (min) Mobile phase A (acetonitrile, %) Mobile phase B (0.5% aqueous phosphoric acid solution, %) 0 2 98 5 2 98 26 15 85 40 17 83 50 17 83 60 34 66 61 40 60 70 90 10 71 2 98 81 2 98

[0064] The chromatograms obtained using mobile phase system 5 show that the chromatographic peaks are well separated under these conditions, meeting the requirements for subsequent analysis.

[0065] Investigation of the preparation method of the two test sample solutions

[0066] (I) Investigation of the extraction solvent

[0067] Chromatograms of water, 30% methanol, 50% methanol, 70% methanol and 100% methanol as extraction solvents were compared.

[0068] Take 0.3g of the contents of JYCF capsules, five portions, accurately weigh them, and place them in stoppered conical flasks. Accurately add water, 30% methanol, 50% methanol, 70% methanol, and 25mL of methanol as extraction solvents, respectively. Seal the flasks tightly, weigh them, and extract ultrasonically (power 250W, frequency 40kHz) for 25min. Cool the flasks, re-weigh them, and then filter them through a 0.45μm microporous membrane to obtain the test solution. Then, perform the detection according to the conditions determined under the chromatographic conditions section to obtain the chromatogram.

[0069] The results showed that the chromatogram obtained by extraction with 50% methanol had the most chromatographic peaks, relatively large peak intensities (especially at 10-15 min), and the least interference from solvent peaks. Therefore, 50% methanol was selected as the extraction solvent for the test sample.

[0070] (II) Examination of Extraction Methods

[0071] Chromatograms of ultrasonic extraction and reflux extraction were compared.

[0072] Accurately weigh two portions (0.3g each) of the contents of JYCF capsules and place them in stoppered conical flasks. Accurately add 25mL of 50% methanol as the extraction solvent, seal tightly, and weigh. Extract one sample ultrasonically (250W, 40kHz) for 25 min, and extract the other sample by reflux for 25 min. Cool, re-weigh, and filter through a 0.45μm microporous membrane to obtain the test solution. Then, perform the analysis according to the conditions determined under the chromatographic conditions section to obtain the chromatogram.

[0073] The results showed that there was no significant difference in the number and intensity of chromatographic peaks obtained by ultrasonic extraction and reflux extraction. Considering that ultrasonic extraction is simpler, ultrasonic extraction was selected as the extraction method.

[0074] (III) Examination of extraction time

[0075] The spectra obtained by ultrasonic extraction of the test sample for 15 min, 30 min, 45 min and 60 min were compared.

[0076] Take 0.3g of the contents of JYCF capsules, four portions, accurately weigh them, place them in a stoppered conical flask, accurately add 25mL of 50% methanol as the extraction solvent, seal tightly, weigh, and ultrasonically extract for 15min, 30min, 45min and 60min respectively (power 250W, frequency 40kHz), cool, re-weigh, and then filter through a 0.45μm microporous membrane to obtain the test solution. Then, detect according to the conditions determined under the chromatographic conditions section to obtain the chromatogram.

[0077] The results showed that there was no significant difference in the number of chromatographic peaks under the four conditions. The intensity of the chromatographic peaks increased slightly when the ultrasonic extraction time was increased to 30 min. Moreover, there was no significant difference in the peak intensity of the chromatograms when the ultrasonic time was increased to 45 min and 60 min compared with the ultrasonic time of 30 min. Therefore, the ultrasonic extraction time was determined to be 30 min.

[0078] (iv) Investigation of the concentration of the test solution

[0079] Chromatograms of test solutions with concentrations of 12 mg / mL, 6 mg / mL, 4 mg / mL, and 3 mg / mL were compared.

[0080] Take 0.3g of the contents of JYCF capsules, weigh 4 portions accurately, and place them in stoppered conical flasks. Accurately add 25mL, 50mL, 75mL, and 100mL of 50% methanol respectively, seal tightly, weigh, and extract by ultrasonication for 30min (power 250W, frequency 40kHz). Cool, re-weigh, and then filter through a 0.45μm microporous membrane to obtain test solutions with concentrations of 12mg / mL, 6mg / mL, 4mg / mL, and 3mg / mL, respectively.

[0081] The results showed that the number of chromatographic peaks did not change significantly after the concentration of the test sample decreased. On the contrary, some chromatographic peaks could not be detected due to the decrease in sample concentration. Therefore, the concentration of the test sample solution was determined to be 12 mg / mL. Attached Figure Description

[0082] Figure 1 The sample chromatogram obtained in the example is shown.

[0083] Figure 2 The image shows the chromatogram of the control sample obtained in the example. Detailed Implementation

[0084] The present invention will be further described in detail below with reference to specific embodiments.

[0085] Example 1

[0086] 1. Instruments and Materials

[0087] 1.1 Instruments

[0088] Waters ACQUITY UPLC H-CLASS ultra-high performance liquid chromatograph (PDA detector, Empower 3.0 chromatography workstation), circulating multi-purpose vacuum pump (Zhengzhou Changcheng Science & Industry Trade Co., Ltd.), KQ250DB ultrasonic cleaner (Kunshan Ultrasonic Instrument Co., Ltd.), and Swiss METTLER TOLEDOAL electronic analytical balance.

[0089] Table 6

[0090]

[0091]

[0092] 1.2.2 Sample Information

[0093] Sample: JYCF

[0094] Manufacturer: Shijiazhuang Yiling Pharmaceutical Co., Ltd.

[0095] Batch numbers: A1811001, A1812001, A1812002, A2106001, A2106002, A2106003, A2205001, A2205002, A2301001, A2302005, A2302006, A2303002, A2303006, A2401003, A2405005, A2405006.

[0096] 1.2.3 Reagents

[0097] Acetonitrile (chromatographic grade, Merck, batch number JA094930)

[0098] Methanol (chromatographic grade, Merck, batch number I1074607006)

[0099] Phosphoric acid (analytical grade, Tianjin Kemeo Chemical Reagent Co., Ltd., batch number 20180115)

[0100] 2. Preparation of the test solution

[0101] Take 0.3g of JYCF sample (batch number: A2401003), weigh it accurately, place it in an Erlenmeyer flask, add 25mL of 50% methanol, weigh it accurately, extract it by sonication for 30min, cool it, weigh it again, add 50% methanol to make up the lost weight, shake it well, let it stand, and filter the supernatant to obtain the sample.

[0102] 3. Preparation of reference solution

[0103] Gardenoside, genipin disaccharide, hyperoside, forsythoside A, pinoresinin-β-D-glucoside, rutin, hesperidin, forsythoside, ammonium glycyrrhizate, magnolol, and honokiol were accurately weighed and placed in 50 mL volumetric flasks. Methanol was added to bring the volume to 50 mL to obtain reference solutions with concentrations of 0.106 mg / mL, 0.041 mg / mL, 0.013 mg / mL, 0.311 mg / mL, 0.025 mg / mL, 0.021 mg / mL, 0.007 mg / mL, 0.095 mg / mL, 0.011 mg / mL, 0.009 mg / mL, and 0.015 mg / mL, respectively.

[0104] 4 Chromatographic conditions

[0105] Chromatographic column: ACQUITY UPLC BEH C18 1.7μm, 2.1*100mm,

[0106] Mobile phase A is acetonitrile, mobile phase B is 0.5% phosphoric acid aqueous solution, and the gradient elution system is shown in Table 7 below.

[0107] Table 7

[0108] Time (min) Mobile phase A (%) Mobile phase B (%) 0 2 98 5 2 98 26 15 85 40 17 83 50 17 83 60 34 66 61 40 60 70 90 10 71 2 98 81 2 98

[0109] Flow rate: 0.3 mL / min.

[0110] Detection wavelength: 230nm.

[0111] Injection volume: 2 μL.

[0112] 4 Results

[0113] Appendix Figure 1 The obtained chromatogram of the test sample is attached. Figure 2The chromatogram of the control sample obtained from the detection is shown.

[0114] A total of 23 characteristic peaks were separated from the chromatogram of the test sample, with retention times of 11.45 min, 17.46 min, 18.78 min, 20.31 min, 29.40 min, 29.99 min, 31.47 min, 32.98 min, 33.14 min, 33.87 min, 34.23 min, 35.91 min, 39.09 min, 44.28 min, 53.81 min, 54.91 min, 59.86 min, 62.99 min, 63.11 min, 64.29 min, 65.97 min, 66.72 min, and 66.84 min.

[0115] Eleven characteristic peaks were isolated from the reference standard chromatogram, with retention times of 18.78 min, 20.31 min, 29.40 min, 32.98 min, 33.14 min, 33.87 min, 35.91 min, 44.28 min, 62.99 min, 65.97 min, and 66.83 min, respectively. The corresponding chemical components were genipin disaccharide, geniposide, hyperoside, forsythoside A, pinoresinin-β-D-glucoside, rutin, hesperidin, forsythoside, ammonium glycyrrhizate, magnolol, and honokiol.

[0116] By comparing the chromatogram of the test sample with that of the reference sample, the chemical components in the traditional Chinese medicine composition JYCF can be identified.

[0117] The content of each chemical component in the traditional Chinese medicine composition JYCF can be calculated by using the high performance liquid chromatography method.

[0118] Experimental Example

[0119] The inventors also conducted a methodological investigation of the method provided by this invention, the details of which are as follows.

[0120] 1. Preparation of reference solution

[0121] Gardenoside, genipin disaccharide, hyperoside, forsythoside A, pinoresinin-β-D-glucoside, rutin, hesperidin, forsythoside, ammonium glycyrrhizate, magnolol, and honokiol were accurately weighed and placed in 50 mL volumetric flasks. Methanol was added to bring the volume to 50 mL to obtain reference solutions with concentrations of 0.106 mg / mL, 0.041 mg / mL, 0.013 mg / mL, 0.311 mg / mL, 0.025 mg / mL, 0.021 mg / mL, 0.007 mg / mL, 0.095 mg / mL, 0.011 mg / mL, 0.009 mg / mL, and 0.015 mg / mL, respectively.

[0122] 2. Preparation of the test solution

[0123] Take 0.3g of JYCF sample (batch number: A2401003), weigh accurately, place it in an Erlenmeyer flask, add 25mL of 50% methanol, weigh accurately, extract by sonication for 30min, cool, weigh again, add 50% methanol to make up the lost weight, shake well, let stand, and filter the supernatant to obtain the sample.

[0124] 3 Chromatographic conditions

[0125] Chromatographic column: ACQUITY UPLC BEH C18 1.7μm, 2.1*100mm,

[0126] Mobile phase A is acetonitrile, mobile phase B is 0.5% phosphoric acid aqueous solution, and the gradient elution system is shown in Table 8 below.

[0127] Table 8

[0128]

[0129]

[0130] Flow rate: 0.3 mL / min.

[0131] Detection wavelength: 230nm.

[0132] Injection volume: 2 μL.

[0133] 4 Exclusivity

[0134] Using 50% methanol as a blank solution, the chromatographic determination was performed under the determined chromatographic conditions. The results showed that the retention times of each index component in the sample corresponded to those in the reference solution, and the blank solvent did not interfere, indicating that the method has good specificity.

[0135] 5 Precision

[0136] The test solution was taken and measured under the determined chromatographic conditions. The sample was injected 6 times consecutively. The results showed that the RSD of the relative retention time and relative peak area of ​​each major chromatographic peak was less than 2%, and the similarity of the 6 samples was 1.000, indicating that the method has good precision.

[0137] 6. Stability

[0138] The test solution was taken and analyzed under the determined chromatographic conditions at 0h, 2h, 4h, 8h, 12h, and 24h. The results showed that the RSD of the retention time of each major chromatographic peak was less than 2%, and the RSD of the peak area was less than 3%. The similarity of the fingerprint chromatogram obtained after the test solution was placed at room temperature for 24h was 1.000, indicating that the method has good stability.

[0139] 7 Repeatability

[0140] Six JYCF samples (batch number: A2401003) were prepared according to the test solution preparation method and analyzed under the determined chromatographic conditions. The results showed that the RSD of the retention time of each major chromatographic peak was less than 2%, the RSD of the peak area was less than 3%, and the similarity was 1.000, indicating that the method has good repeatability.

Claims

1. A method for detecting chemical components in a traditional Chinese medicine composition, wherein the raw materials of the traditional Chinese medicine composition are: gardenia, forsythia, magnolia bark (processed with ginger), pinellia (processed with ginger), immature bitter orange peel, poria cocos, perilla stem, and licorice, characterized in that, The detection method employs an ultra-high performance liquid chromatograph. The extraction solvent used in the preparation of the test solution was 50% methanol; Chromatographic conditions: Mobile phase A was acetonitrile, mobile phase B was 0.5% phosphoric acid aqueous solution, and gradient elution was used. The gradient elution system is as follows. 。 2. The detection method according to claim 1, characterized in that, The test sample solution was prepared by ultrasonic extraction for 30 minutes, and the concentration of the test sample solution was 12 mg / mL.

3. The detection method according to claim 1, characterized in that, The chromatographic conditions specified are as follows: the chromatographic column is a C18 reversed-phase column with a particle size of 1.7 μm, and the flow rate of the mobile phase is 0.3 mL / min.

4. The application of the detection method as described in claim 1 in the quality control method of traditional Chinese medicine composition preparations made from gardenia, forsythia, magnolia bark, pinellia, immature bitter orange, poria, perilla stem, and licorice.

5. The application according to claim 4, characterized in that, The quality control methods are the identification and content determination of traditional Chinese medicine composition preparations.

6. The application according to claim 4, characterized in that, The quality control method also includes the preparation of a reference solution.

7. The application according to claim 6, characterized in that, The reference standards used in the preparation of the reference solution include geniposide, genipin disaccharide, hyperoside, forsythoside A, pinoresinin-β-D-glucoside, rutin, hesperidin, forsythoside, ammonium glycyrrhizate, magnolol, and honokiol.

8. The application according to claim 6, characterized in that, The reference solution was prepared by accurately weighing appropriate amounts of geniposide, and / or genipin disaccharide, and / or hyperoside, and / or forsythoside A, and / or pinoresinin-β-D-glucoside, and / or rutin, and / or hesperidin, and / or forsythoside, and / or ammonium glycyrrhizate, and / or magnolol, and / or honokiol reference standards, mixing them, and dissolving them in an appropriate amount of methanol to prepare solutions with concentrations of 0.106 mg / mL, 0.041 mg / mL, 0.013 mg / mL, 0.311 mg / mL, 0.025 mg / mL, 0.021 mg / mL, 0.007 mg / mL, 0.095 mg / mL, 0.011 mg / mL, 0.009 mg / mL, and 0.015 mg / mL.

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