Fingerprint spectrum of Xinlokang tablets, construction method and application of fingerprint spectrum, and multi-index component detection method of Xinlokang tablets

Abstract

The invention provides a fingerprint spectrum of a Xinlekang tablet, a construction method and application of the fingerprint spectrum and a multi-index component detection method of the Xinlekang tablet, and relates to the technical field of pharmaceutical analysis structures. According to the method, trichloromethane and ammonia water are used as extracting agents, a C18 chromatographic column is adopted, acetonitrile is used as a mobile phase A, a 0.38-0.42 v / v% phosphoric acid-0. 04-0.06 v / v% triethylamine aqueous solution is used as a mobile phase B, a gradient elution mode is adopted, a gradient elution procedure is controlled, the detection wavelength is controlled to be 202-206 nm, and the detection wavelength is controlled to be 206-206 nm. A test solution of the Xinlekang tablet and a mixed reference substance solution are subjected to high performance liquid chromatography detection respectively, liquid chromatograms of the test solution and the mixed reference substance solution are compared and analyzed, the constructed fingerprint spectrum of the Xinlekang tablet comprises 28 characteristic peaks including isorhynchophylline, rhynchophylline and reserpine, the chromatographic peaks are rich, the quality of the Xinlekang tablet can be effectively represented, and the quality of the Xinlekang tablet is improved. The quality of the medicine can be comprehensively monitored, and the application value is relatively high.

Description

Technical Field

[0001] The present invention relates to the technical field of drug analysis structure, and in particular to a fingerprint spectrum of Xinlekang tablets, a construction method and application thereof, and a multi-index component detection method of Xinlekang tablets. Background Art

[0002] Insomnia is a subjective experience that the quality or quantity of sleep does not meet normal physiological needs due to difficulty falling asleep and / or maintaining sleep, which affects daytime social functions. It is the most common sleep disorder. The main drugs used in clinical practice are phenobarbital drugs, benzodiazepine drugs, non-benzodiazepine drugs and Chinese patent medicines. Compared with Western medicines, Chinese patent medicines have fewer side effects and can fundamentally solve the problem of insomnia, and are accepted by the majority of patients.

[0003] Xinlekang tablets have the effects of calming the mind, nourishing the liver and calming the nerves. They are suitable for neurasthenia, insomnia, dreaminess, palpitations and dizziness. They are a Chinese medicine compound tablet composed of three medicinal materials: Uncaria rhynchophylla, Semen Ziziphi Spinosae and Rauwolfia odorata. Among them, the main medicine Uncaria rhynchophylla has the effects of protecting nerves, lowering blood pressure, protecting myocardium, anti-cancer, anti-inflammatory, and relieving asthma; the auxiliary medicine Semen Ziziphi Spinosae has the effects of sedation and hypnosis, anti-anxiety, anti-depression, protecting nerves, protecting cardiovascular and cerebrovascular vessels, protecting the liver, and anti-oxidation; the auxiliary medicine Rauwolfia odorata alkaloids have the effects of lowering blood pressure, anti-arrhythmia, clearing away heat and detoxifying. There are many Chinese medicinal materials in the formula of Xinlekang tablets, and it is of great significance to effectively characterize the quality of Chinese medicine as a whole. So far, there has been no report on the quality control of Xinlekang tablets using the fingerprint method. Therefore, it is of great significance to establish a fingerprint that can comprehensively reflect the overall quality of Xinlekang tablets. Summary of the invention

[0004] In view of this, the purpose of the present invention is to provide a fingerprint spectrum of Xinlekang tablets and a construction method and application thereof, and a multi-index component detection method of Xinlekang tablets. The fingerprint spectrum of Xinlekang tablets constructed by the present invention has a large number of characteristic peaks, which can comprehensively reflect the overall quality of Xinlekang tablets and is conducive to comprehensive monitoring of the quality of drugs.

[0005] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0006] The present invention provides a method for constructing a fingerprint of Xinlekang tablets, comprising the following steps:

[0007] The powder of Xinlekang tablets, ammonia water and chloroform are mixed and extracted to obtain an extract, and the extract is concentrated and dissolved in methanol to obtain a test solution;

[0008] The test solution and the mixed reference solution are subjected to high performance liquid chromatography detection respectively to obtain a sample liquid phase spectrum and a reference liquid phase spectrum respectively, and a fingerprint spectrum of Xinlekang tablets is obtained according to the sample liquid phase spectrum and the reference liquid phase spectrum;

[0009] The reference substances in the mixed reference solution include isorhynchophylline, rhynchophylline and reserpine;

[0010] The conditions for the high performance liquid chromatography detection include: the chromatographic column includes a C18 chromatographic column; the mobile phase A is acetonitrile, the mobile phase B is a phosphoric acid-triethylamine aqueous solution, the volume fraction of phosphoric acid in the mobile phase B is 0.38-0.42%, and the volume fraction of triethylamine is 0.04-0.06%; the elution method is gradient elution, and the procedure of the gradient elution includes: 0-15min, the volume fraction of the mobile phase A increases from 14-16% to 20-22%; 15-30min, the volume fraction of the mobile phase A increases from 20-22% to 24-26%; 30-65min, the volume fraction of the mobile phase A increases from 20-22% to 24-26% The volume fraction of mobile phase A increases from 24-26% to 39-41%; at 65-70min, the volume fraction of mobile phase A increases from 39-41% to 48-52%; at 70-71min, the volume fraction of mobile phase A increases from 48-52% to 88-92%; at 71-75min, the volume fraction of mobile phase A is 88-92%; at 75-76min, the volume fraction of mobile phase A decreases from 88-92% to 14-16%; at 76-81min, the volume fraction of mobile phase A is 14-16%; the detection wavelength is 202-206nm;

[0011] The fingerprint spectrum of Xinlekang tablets contains 28 characteristic peaks, including the characteristic peaks of isorhynchophylline, rhynchophylline and reserpine.

[0012] Preferably, the solid-liquid ratio of the powder of the Xinlekang tablets to the ammonia water is 1g:1.8-2.2mL; the mass fraction of the ammonia water is 25-28%;

[0013] The solid-liquid ratio of the powder of the Xinlekang tablets to chloroform is 1g:24-26mL.

[0014] Preferably, the extraction is reflux extraction, and the extraction time is 28 to 32 minutes.

[0015] Preferably, in the mixed reference solution, the concentration of isorhynchophylline is 7.6-7.8 μg / mL, the concentration of rhynchophylline is 7.9-8.1 μg / mL, and the concentration of reserpine is 4.7-4.9 μg / mL;

[0016] The solvent in the mixed reference solution includes methanol.

[0017] Preferably, the conditions for the HPLC detection also include: a mobile phase flow rate of 0.7 to 0.9 mL / min, a diode array detector, a column temperature of 28 to 32° C., and an injection volume of 9.5 to 10.5 μL.

[0018] Preferably, the liquid phase spectrum of the sample and the liquid phase spectrum of the reference substance are calculated by the "Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System" software, and the similarity is both greater than 0.95.

[0019] The present invention also provides a fingerprint of Xinlekang tablets obtained by the fingerprint construction method described in the above technical solution, which contains 28 characteristic peaks, including the characteristic peaks of isorhynchophylline, rhynchophylline and reserpine.

[0020] The present invention also provides the application of the fingerprint spectrum of Xinlekang tablets described in the above technical solution in the quality control of Xinlekang tablets.

[0021] The present invention also provides a multi-index component detection method for Xinlekang tablets, comprising the following steps:

[0022] The powder of the Xinlekang tablets to be tested, ammonia water and chloroform are mixed and extracted to obtain an extract solution, and the extract solution is concentrated and dissolved in methanol to obtain a sample solution to be tested;

[0023] The sample solution to be tested is subjected to high performance liquid chromatography to obtain a multi-index component test result of Xinlekang tablets; the multi-index components include at least two of isorhynchophylline, rhynchophylline and reserpine;

[0024] The conditions for the high performance liquid chromatography detection include: the chromatographic column includes a C18 chromatographic column; the mobile phase A is acetonitrile, the mobile phase B is a phosphoric acid-triethylamine aqueous solution, the volume fraction of phosphoric acid in the mobile phase B is 0.38-0.42%, and the volume fraction of triethylamine is 0.04-0.06%; the elution method is gradient elution, and the procedure of the gradient elution includes: 0-15min, the volume fraction of the mobile phase A increases from 14-16% to 20-22%; 15-30min, the volume fraction of the mobile phase A increases from 20-22% to 24-26%; 30-65min, the volume fraction of the mobile phase A increases from 20-22% to 24-26% The volume fraction of mobile phase A increases from 24-26% to 39-41%; from 65 to 70 min, the volume fraction of mobile phase A increases from 39-41% to 48-52%; from 70 to 71 min, the volume fraction of mobile phase A increases from 48-52% to 88-92%; from 71 to 75 min, the volume fraction of mobile phase A is 88-92%; from 75 to 76 min, the volume fraction of mobile phase A decreases from 88-92% to 14-16%; from 76 to 81 min, the volume fraction of mobile phase A is 14-16%; the detection wavelength is 202 to 206 nm.

[0025] Preferably, the solid-liquid ratio of the powder of the Xinlekang tablets to the ammonia water is 1g:1.8-2.2mL; the mass fraction of the ammonia water is 25-28%;

[0026] The solid-liquid ratio of the powder of the Xinlekang tablet to chloroform is 1g:24-26mL;

[0027] The extraction is reflux extraction, and the extraction time is 28 to 32 minutes;

[0028] The conditions for the high performance liquid chromatography detection also include: a mobile phase flow rate of 0.7 to 0.9 mL / min, a diode array detector, a column temperature of 28 to 32° C., and an injection volume of 9.5 to 10.5 μL.

[0029] Xinlekang tablets are a traditional Chinese medicine compound tablet composed of three medicinal materials, Uncaria rhynchophylla, Semen Ziziphi spinulosae and Rauwolfia ovata, and other auxiliary materials. The ingredients of the traditional Chinese medicine are numerous and complex, and the extraction and liquid chromatography separation are difficult. The present invention uses ammonia water and chloroform as extractants, extracts a large number of ingredients, can effectively characterize the quality of Xinlekang tablets, and is conducive to comprehensive monitoring of the quality of the drug.

[0030] Since the fingerprint spectrum is not for determining the precise content of a certain component, but for fully reflecting the information of the chemical composition, when the present invention uses 202-206nm as the detection wavelength, it has more peaks, reflects more complete information, has good absorption values ​​of each peak, and has a stable baseline, and also avoids the situation where the near-ultraviolet impurity peak has a large absorption. The present invention adopts a C18 chromatographic column, acetonitrile as mobile phase A, 0.38-0.42v / v% phosphoric acid-0.04-0.06v / v% triethylamine aqueous solution as mobile phase B, adopts a gradient elution mode and controls the gradient elution program, and the test solution and the mixed reference solution of the new Lekang tablets are respectively subjected to high performance liquid chromatography detection and the liquid chromatograms of the two are compared and analyzed, and there are 28 characteristic peaks in the obtained fingerprint spectrum of the new Lekang tablets, wherein the 28 characteristic peaks include the characteristic peaks of isorhynchophylline, rhynchophylline and reserpine. The fingerprint spectrum constructed by the present invention has rich chromatographic peaks, which is conducive to comprehensive monitoring of the quality of the drug, provides an effective means for comprehensively reflecting, monitoring and evaluating the overall quality control of Xinlekang tablets, guides standardized production, and thus better ensures the quality stability, consistency and controllability of Xinlekang tablets, improves the overly simple existing quality standards of Xinlekang tablets, and has high application value.

[0031] The high-performance liquid fingerprint method established in the present invention has realized the quality control of the whole formula of Xinlekang Tablets for the first time. It does not identify a single compound or medicinal material. It can more effectively guide the feeding, strictly regulate the production operation, and improve the safety and effectiveness of clinical drug use.

[0032] The present invention establishes a multi-index component synchronous detection method for Xinlekang tablets, which has the advantage of quantitative monitoring of multiple components in the same chromatographic system. As a quality control method for Xinlekang tablets, it can fully and accurately evaluate the effectiveness, safety and stability of Xinlekang tablets, control product quality raw materials more accurately and comprehensively, and ensure the quality and efficacy of the product. Moreover, the detection method of the present invention has the advantages of simple method, stability, high precision and good reproducibility. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] Figure 1 It is the HPLC chromatogram of the test solution in Example 1;

[0034] Figure 2 It is the fingerprint superposition of 12 batches of Xinlekang tablets in Example 1;

[0035] Figure 3 This is a graph showing the similarity evaluation results of 12 batches of Xinlekang tablets in Example 1;

[0036] Figure 4 The HPLC chromatogram of the test solution lacking Uncaria rhynchophylla in Example 2;

[0037] Figure 5 The HPLC chromatogram of the test solution lacking Ziziphus jujuba seeds in Example 2;

[0038] Figure 6 The HPLC chromatogram of the test solution lacking Rauwolfia in Example 2;

[0039] Figure 7 It is the HPLC chromatogram of the Uncaria rhynchophylla test solution in Example 3;

[0040] Figure 8 The HPLC chromatogram of the test solution of Semen Ziziphi Spinosae in Example 3 is shown in FIG.

[0041] Fig. 9 The HPLC chromatogram of the Rauwolfia test solution in Example 3;

[0042] Fig.10 This is a full wavelength scan of the diode array detector of the test solution in Example 5;

[0043] Fig.11 The HPLC chromatogram of the test solution using the gradient elution program of Table 5 in Example 5;

[0044] Fig.12 The HPLC chromatogram of the test solution using the gradient elution program of Table 6 in Example 5;

[0045] Fig.13 The HPLC chromatogram of the test solution using the gradient elution program in Table 7 in Example 5. DETAILED DESCRIPTION

[0046] The present invention provides a method for constructing a fingerprint of Xinlekang tablets, comprising the following steps:

[0047] The powder of Xinlekang tablets, ammonia water and chloroform are mixed and extracted to obtain an extract, and the extract is concentrated and dissolved in methanol to obtain a test solution;

[0048] The test solution and the mixed reference solution are subjected to high performance liquid chromatography detection respectively to obtain a sample liquid phase spectrum and a reference liquid phase spectrum respectively, and a fingerprint spectrum of Xinlekang tablets is obtained according to the sample liquid phase spectrum and the reference liquid phase spectrum;

[0049] The reference substances in the mixed reference solution include isorhynchophylline, rhynchophylline and reserpine;

[0050] The conditions for the high performance liquid chromatography detection include: the chromatographic column includes a C18 chromatographic column; the mobile phase A is acetonitrile, the mobile phase B is a phosphoric acid-triethylamine aqueous solution, the volume fraction of phosphoric acid in the mobile phase B is 0.38-0.42%, and the volume fraction of triethylamine is 0.04-0.06%; the elution method is gradient elution, and the procedure of the gradient elution includes: 0-15min, the volume fraction of the mobile phase A increases from 14-16% to 20-22%; 15-30min, the volume fraction of the mobile phase A increases from 20-22% to 24-26%; 30-65min, the volume fraction of the mobile phase A increases from 20-22% to 24-26% The volume fraction of mobile phase A increases from 24-26% to 39-41%; at 65-70min, the volume fraction of mobile phase A increases from 39-41% to 48-52%; at 70-71min, the volume fraction of mobile phase A increases from 48-52% to 88-92%; at 71-75min, the volume fraction of mobile phase A is 88-92%; at 75-76min, the volume fraction of mobile phase A decreases from 88-92% to 14-16%; at 76-81min, the volume fraction of mobile phase A is 14-16%; the detection wavelength is 202-206nm;

[0051] The fingerprint spectrum of Xinlekang tablets contains 28 characteristic peaks, including the characteristic peaks of isorhynchophylline, rhynchophylline and reserpine.

[0052] Unless otherwise specified, the materials and equipment used in the present invention are all commercially available products in the art.

[0053] The invention comprises the following steps: mixing powder of Xinlekang tablets, ammonia water and chloroform for extraction to obtain an extract, concentrating the extract and then dissolving it in methanol to obtain a test solution.

[0054] In the present invention, the particle size of the powder of the Xinlekang tablets is preferably ≥ 60 meshes; the powder of the Xinlekang tablets is preferably obtained by removing the coating from the Xinlekang tablets and grinding them to less than 60 meshes.

[0055] In the present invention, the mass fraction of the ammonia water is 25-28%, and in a specific embodiment it can be 25%, 26%, 27% or 28%; the solid-liquid ratio of the powder of the Xinlekang tablets to the ammonia water is preferably 1g:1.8-2.2mL, and in a specific embodiment it can be 1g:1.8mL, 1g:1.9mL, 1g:2mL, 1g:2.1mL or 1g:2.2mL.

[0056] In the present invention, the solid-liquid ratio of the powder of the Xinlekang tablets to chloroform is preferably 1g:24-26mL, and in a specific embodiment it can be 1g:24mL, 1g:25mL or 1g:26mL.

[0057] In the present invention, the extraction is preferably reflux extraction, and the extraction time is preferably 28 to 32 minutes, and in a specific embodiment can be 28 minutes, 29 minutes, 30 minutes, 31 minutes or 32 minutes.

[0058] After the extraction, the present invention preferably further comprises: filtering the obtained extraction system to obtain an extract.

[0059] In the present invention, the solid-liquid ratio of the powder of the Xinlekang tablets to methanol is preferably 1 g:4.5-5.5 mL, and in a specific embodiment it can be 1 g:4.5 mL, 1 g:5 mL or 1 g:5.5 mL.

[0060] After the dissolution, the present invention preferably further comprises filtering the solution obtained by the dissolution through a 0.22 μm microporous filter membrane to obtain a test solution.

[0061] After obtaining the test solution, the present invention performs high performance liquid chromatography on the test solution and the mixed reference solution, respectively, to obtain a sample liquid phase spectrum and a reference liquid phase spectrum, respectively, and obtains a fingerprint spectrum of Xinlekang tablets according to the sample liquid phase spectrum and the reference liquid phase spectrum.

[0062] In the present invention, the reference substances in the mixed reference solution include isorhynchophylline, rhynchophylline and reserpine. In the present invention, the concentration of isorhynchophylline in the mixed reference solution is preferably 7.6-7.8 μg / mL, and in a specific embodiment it can be 7.6 μg / mL, 7.7 μg / mL or 7.8 μg / mL; the concentration of rhynchophylline is preferably 7.9-8.1 μg / mL, and in a specific embodiment it can be 7.9 μg / mL, 8 μg / mL or 8.1 μg / mL; the concentration of reserpine is preferably 4.7-4.9 μg / mL, and in a specific embodiment it can be 4.7 μg / mL, 4.8 μg / mL or 4.9 μg / mL; the solvent in the mixed reference solution preferably includes methanol.

[0063] In the present invention, the preparation method of the mixed reference solution preferably comprises the following steps: dissolving isorhynchophylline reference substance, rhynchophylline reference substance and reserpine reference substance in a solvent, filtering through a 0.22 μm microporous filter membrane to obtain a mixed reference solution.

[0064] In the present invention, the conditions for the high performance liquid chromatography detection include: the chromatographic column includes a C18 chromatographic column; the mobile phase system includes a mobile phase A and a mobile phase B, the mobile phase A is acetonitrile, and the mobile phase B is a phosphoric acid-triethylamine aqueous solution; the volume fraction of phosphoric acid in the mobile phase B is 0.38-0.42%, and in a specific embodiment, it can be 0.38%, 0.39%, 0.4%, 0.41% or 0.42%; the volume fraction of triethylamine in the mobile phase B is 0.04-0.06%, and in a specific embodiment, it can be 0.04%, 0.05% or 0.06%; the mobile phase flow rate is preferably 0.7-0.9m L / min, in a specific embodiment, it can be 0.7mL / min, 0.8mL / min or 0.9mL / min; the detector is preferably a diode array detector, and the detection wavelength is 202-206nm, in a specific embodiment, it can be 202nm, 203nm, 204nm, 205nm or 206nm; the column temperature is preferably 28-32℃, in a specific embodiment, it can be 28℃, 29℃, 30℃, 31℃ or 32℃; the injection volume is preferably 9.5-10.5μL, in a specific embodiment, it can be 9.5μL, 9.8μL, 10μL, 10.2μL or 10.5μL; the theoretical plate number calculated according to the rhynchophylline peak should be not less than 20000; the elution method is gradient elution, and the gradient elution program includes: 0-15min, the volume fraction of mobile phase A increases from 14-16% to 20-22%, and in a specific embodiment, it can be increased from any value of 14%, 15% and 16% to any value of 20%, 21% and 22%; 15-30min, the volume fraction of mobile phase A increases from 20-22% to 24-26%, and in a specific embodiment, it can be increased from any value of 20%, 21% and 22%. From 20%, 21% and 22% to any one of 24%, 25% and 26%; 30-65min, the volume fraction of mobile phase A increases from 24-26% to 39-41%, and in a specific embodiment, it can be increased from any one of 24%, 25% and 26% to any one of 39%, 40% and 41%; 65-70min, the volume fraction of mobile phase A increases from 39-41% to 48-52%, and in a specific embodiment, it can be increased from 39 %, 40% and 41% to any one of 48%, 49%, 50%, 51% and 52%; 70-71min, the volume fraction of mobile phase A increases from 48-52% to 88-92%, and in a specific embodiment, it can increase from any one of 48%, 49%, 50%, 51% and 52% to any one of 88%, 89%, 90%, 91% and 92%; 71-75min, the volume fraction of mobile phase A is 88-92%, In a specific embodiment, it can be 88%, 89%, 90%, 91% or 92%; at 75-76 min, the volume fraction of the mobile phase A is reduced from 88-92% to 14-16%, and in a specific embodiment, it can be reduced from any value among 88%, 89%, 90%, 91% and 92% to any value among 14%, 15% and 16%; at 76-81 min, the volume fraction of the mobile phase A is 14-16%, and in a specific embodiment, it can be 14%, 15% or 16%. .

[0065] The present invention preferably imports the sample liquid phase spectrum into the Chinese medicine chromatographic fingerprint similarity evaluation system to generate a common mode of characteristic spectrum and obtain a fingerprint spectrum containing 28 characteristic peaks; by comparing the retention time of the fingerprint spectrum with the reference liquid phase spectrum (based on the chromatographic peak of the reference solution), the fingerprint spectrum includes characteristic peaks of isorhynchophylline, rhynchophylline and reserpine. In the present invention, the sample liquid phase spectrum is preferably calculated by the "Chinese medicine chromatographic fingerprint similarity evaluation system" software, and the common peaks with similarities greater than 0.95 are the fingerprints of Xinlekang tablets. In the present invention, the "Chinese medicine chromatographic fingerprint similarity evaluation system" software is preferably "Chinese medicine chromatographic fingerprint similarity evaluation system software (2012A version)". The present invention preferably uses the average method for similarity analysis, and the time window width is preferably set to 0.1min, and a total of 28 common peaks are calibrated. In an embodiment of the present invention, 12 batches of Xinlekang tablets were subjected to high performance liquid chromatography analysis, and the obtained test sample chromatograms were introduced into the Chinese medicine chromatographic fingerprint similarity evaluation system for similarity analysis. The similarities of each batch of Xinlekang tablets were greater than 0.95, and the similarities were more preferably greater than 0.99.

[0066] In the present invention, by comparing and analyzing the retention time of the reference substance in the fingerprint spectrum and the reference substance liquid phase spectrum, it is obtained that peak No. 10 in the fingerprint spectrum is isorhynchophylline, peak No. 15 is rhynchophylline, peak No. 28 is reserpine, peaks No. 2, 13, 16, 22 and 27 belong to Semen Ziziphi Spinosae, peaks No. 1 to 12 and No. 15 to 27 belong to Uncaria rhynchophylla, and peaks No. 4, 6, 8, 11, 14, 20, 27 and 28 belong to Rauwolfia oleifera.

[0067] Traditional Chinese medicine fingerprint is an internationally recognized research model and technical platform for controlling the quality of traditional Chinese medicine. It analyzes the types and content distribution information of the effective ingredients and ineffective ingredients of traditional Chinese medicine, so that the research methods and quality analysis methods of traditional Chinese medicine have been developed from the analysis of one or a few active ingredients to the comprehensive analysis of the chemical fingerprint of the entire traditional Chinese medicine, which is universal and economical. The traditional Chinese medicine fingerprint reflects the synergistic effect, multi-level complexity and integrity of traditional Chinese medicine. There are many kinds of traditional Chinese medicine materials in the formula of Xinlekang tablets, and its material group must be controlled as a whole. Therefore, in addition to "microscopic analysis", a certain "macroscopic analysis" method should also be used to effectively characterize the quality of traditional Chinese medicine as a whole. The present invention can obtain fingerprints of 28 characteristic peaks by controlling the extraction conditions and HPLC detection conditions, which can effectively characterize the quality of Xinlekang tablets and is conducive to comprehensive monitoring of the quality of drugs.

[0068] The present invention provides a fingerprint of Xinlekang tablets obtained by the fingerprint construction method described in the above technical solution, which contains 28 characteristic peaks, including the characteristic peaks of isorhynchophylline, rhynchophylline and reserpine.

[0069] The present invention also provides the application of the fingerprint spectrum of Xinlekang tablets described in the above technical solution in the quality control of Xinlekang tablets.

[0070] The present invention also provides a multi-index component detection method for Xinlekang tablets, comprising the following steps:

[0071] The powder of the Xinlekang tablets to be tested, ammonia water and chloroform are mixed and extracted to obtain an extract solution, and the extract solution is concentrated and dissolved in methanol to obtain a sample solution to be tested;

[0072] The sample solution to be tested is subjected to high performance liquid chromatography to obtain a multi-index component test result of Xinlekang tablets; the multi-index components include at least two of isorhynchophylline, rhynchophylline and reserpine;

[0073] The conditions for the high performance liquid chromatography detection include: the chromatographic column includes a C18 chromatographic column; the mobile phase A is acetonitrile, the mobile phase B is a phosphoric acid-triethylamine aqueous solution, the volume fraction of phosphoric acid in the mobile phase B is 0.38-0.42%, and the volume fraction of triethylamine is 0.04-0.06%; the elution method is gradient elution, and the procedure of the gradient elution includes: 0-15min, the volume fraction of the mobile phase A increases from 14-16% to 20-22%; 15-30min, the volume fraction of the mobile phase A increases from 20-22% to 24-26%; 30-65min, the volume fraction of the mobile phase A increases from 20-22% to 24-26% The volume fraction of mobile phase A increases from 24-26% to 39-41%; from 65 to 70 min, the volume fraction of mobile phase A increases from 39-41% to 48-52%; from 70 to 71 min, the volume fraction of mobile phase A increases from 48-52% to 88-92%; from 71 to 75 min, the volume fraction of mobile phase A is 88-92%; from 75 to 76 min, the volume fraction of mobile phase A decreases from 88-92% to 14-16%; from 76 to 81 min, the volume fraction of mobile phase A is 14-16%; the detection wavelength is 202 to 206 nm.

[0074] In the present invention, the preparation method of the sample solution to be tested is the same as the preparation method of the test solution; the conditions of the HPLC detection are the same as the conditions of the HPLC detection in the fingerprint spectrum construction method, and will not be repeated here.

[0075] In the present invention, the multi-index component detection results include qualitative detection results and quantitative detection results. In the present invention, the qualitative detection results are preferably obtained using the Xinlekang tablet fingerprint spectrum described in the above technical solution.

[0076] The present invention has no particular limitation on the method for obtaining the quantitative detection results, and any quantitative method well known to those skilled in the art may be used.

[0077] The detection method of the present invention has the advantages of being simple, stable, high in precision and good in reproducibility.

[0078] In order to further illustrate the present invention, the fingerprint construction method of Xinlekang tablets, its preparation method and application, and the multi-index component detection method are described in detail below in combination with the embodiments, but they should not be understood as limiting the scope of protection of the present invention.

[0079] In the following examples, the instrument used was Agilent 1260 high performance liquid chromatograph (Agilent, USA); the reagents and medicinal materials used were as follows: isorhynchophylline reference substance and rhynchophylline reference substance were purchased from Zhuhai Anzhe Biotechnology Co., Ltd., and reserpine reference substance was purchased from China Food and Drug Inspection Institute. The batch number of Xinlekang tablets (purchased from Taiyuan Pharmaceutical Co., Ltd.) is shown in Table 1, wherein batch 12 of Xinlekang tablets was used in Example 1, and the batch number of Xinlekang tablets used in other examples was S1.

[0080] Table 112 Batch of Lecon Tablets

[0081] serial number batch number serial number batch number serial number batch number serial number batch number S1 231101 S4 231201 S7 240101 S10 240301 S2 231102 S5 231202 S8 240102 S11 240302 S3 231103 S6 231203 S9 240103 S12 240303

[0082] Example 1

[0083] Construction of fingerprint of Xinlekang tablets

[0084] Preparation of test solution: Take 10 tablets of Xinlekang tablets (S1), remove the coating, grind finely (below 60 mesh), accurately weigh 1.0g, put it in a 100mL round-bottom flask, add 2mL ammonia water (mass concentration 25-28%), add 25mL chloroform, heat and reflux to extract for 30min, filter, concentrate the filtrate to dryness, add methanol to the residue to dissolve, make up to 5mL volumetric flask, filter through 0.22μm microporous membrane, and the filtrate is the test solution. Prepare the test solutions of another 11 batches of Xinlekang tablets shown in Table 1 according to the above method.

[0085] Preparation of reference solution: Take isorhynchophylline reference, rhynchophylline reference, and reserpine reference, add methanol to dissolve and make up to volume to obtain a mixed reference solution with an isorhynchophylline concentration of 7.70 μg / mL, a rhynchophylline concentration of 8.00 μg / mL, and a reserpine concentration of 4.78 μg / mL, and filter through a 0.22 μm microporous filter membrane before entering HPLC.

[0086] The mixed reference solution and the test solution were subjected to HPLC detection respectively, and the liquid chromatograms were recorded to obtain the reference liquid chromatogram and the sample liquid chromatogram respectively. The sample liquid chromatogram was imported into the "Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System (2012 Edition)" to perform similarity analysis on the fingerprints of 12 batches of Xinlekang Tablets. With S1 as the reference spectrum, after cutting the first 5min solvent peak, the chromatograms of the 12 batches of samples (S1-S12) were matched, and the average method was used to generate the reference spectrum (R), and the time window width was set to 0.1min. The results are shown in Figures 1 to 3 and Tables 3 to 4, Figure 1 This is the HPLC chromatogram of the test solution (Xinlekang Tablets S1). Figure 2 This is the HPLC superimposed chromatogram of the test solution of 12 batches of Xinlekang tablets. Figure 3 This is the similarity evaluation result diagram of 12 batches of Xinlekang tablets.

[0087] Among them, the HPLC detection conditions are as follows: the chromatographic column is a C18 chromatographic column (250mm×4.6mm, 5μm), acetonitrile is used as mobile phase A, 0.4v / v% phosphoric acid-0.05v / v% triethylamine aqueous solution is used as mobile phase B, gradient elution, the flow rate is 0.8mL / min, the column temperature is 30°C, the detector is a diode array detector, the detection wavelength is 204nm, and the injection volume is 10μL; the gradient elution program is shown in Table 2, and the theoretical plate number calculated based on the rhynchophylline peak should be not less than 20000.

[0088] Table 2 Gradient elution program

[0089] Time / min Mobile phase A volume fraction / % Mobile phase B volume fraction / % 0 15 85 15 21 79 30 25 75 65 40 60 70 50 50 71 90 10 75 90 10 76 15 85 81 15 85

[0090] Table 3 Similarity evaluation results of 12 batches of Xinlekang tablets samples

[0091]

[0092]

[0093]

[0094] Table 4 Similarity evaluation results of 12 batches of Xinlekang tablets samples

[0095]

[0096] The common fingerprint peak was calibrated by selecting the reference substance rhynchophylline (S) as the reference substance, and its retention time was set as 1 to calculate the relative retention time of the fingerprint peak. Figures 1 to 3and Tables 3-4), the chromatographic peaks with good stability, strong absorption and obvious characteristics were selected as common peaks. As a result, a total of 28 common fingerprint peaks were calibrated, and the results showed that the relative retention time RSD of the common peaks of the test samples was <0.7%. The similarity of the 12 batches of test samples was between 0.988 and 0.999, indicating that the 12 batches of test samples had a high degree of similarity.

[0097] Example 2

[0098] Negative control experiment

[0099] Preparation of the test solution of Uncaria rhynchophylla: weigh 7 g of Ziziphus jujuba seeds, add 120 mL of water and soak for 24 h, then decoct twice, the first time for 6 h and the second time for 2 h, combine the decoctions and concentrate to dryness, add 0.04 g of total alkaloids of Rauwolfia ovata, add 2 mL of ammonia water (mass concentration 25-28%), add 25 mL of chloroform, heat and reflux for 30 min, filter, concentrate the filtrate to dryness, add methanol to the residue to dissolve, make up to 5 mL volumetric flask, filter the filtrate through a 0.22 μm microporous filter membrane to obtain the test solution of Uncaria rhynchophylla.

[0100] Preparation of the test solution lacking Ziziphus jujuba seed: weigh 14 g of Uncaria rhynchophylla, add 120 mL of water and soak for 24 h, then decoct twice, the first time for 6 h and the second time for 2 h, combine the decoctions and concentrate to dryness, add 0.04 g of total alkaloids of Rauwolfia oxyphylla, add 2 mL of ammonia water (mass concentration 25-28%), add 25 mL of chloroform, heat and reflux for 30 min, filter, concentrate the filtrate to dryness, add methanol to the residue to dissolve, make up to 5 mL volumetric flask, filter the filtrate through a 0.22 μm microporous filter membrane to obtain the test solution lacking Ziziphus jujuba seed.

[0101] Preparation of the test solution lacking Rauwolfia: weigh 14 g of Uncaria rhynchophylla and 7 g of Semen Ziziphi Spinosae, add 120 mL of water and soak for 24 h, then decoct twice, the first time for 6 h and the second time for 2 h, combine the decoctions and concentrate to dryness, add 2 mL of ammonia water (mass concentration 25-28%), add 25 mL of chloroform, heat and reflux for 30 min, filter, concentrate the filtrate to dryness, add methanol to the residue to dissolve, make up to a 5 mL volumetric flask, and filter the filtrate through a 0.22 μm microporous filter membrane to obtain the test solution lacking Rauwolfia.

[0102] According to the HPLC detection conditions of Example 1, HPLC detection was performed on each negative test solution.

[0103] Figure 4 This is the HPLC chromatogram of the solution of the test product lacking Uncaria rhynchophylla. Figure 5 This is the HPLC chromatogram of the test solution lacking Ziziphus jujuba seeds. Figure 6 This is the HPLC chromatogram of the solution lacking Rauwolfia test sample. It can be seen that the negative test samples have no interference and the chromatographic peak separation of each component is greater than 1.5.

[0104] Example 3

[0105] Identification and attribution of common peaks

[0106] Preparation of Uncaria rhynchophylla test solution: weigh 14 g of Uncaria rhynchophylla, add 120 mL of water and soak for 24 h, then decoct twice, the first time for 6 h and the second time for 2 h, combine the decoctions and concentrate to dryness, add 2 mL of ammonia water (mass concentration 25-28%), add 25 mL of chloroform, heat and reflux for 30 min, filter, concentrate the filtrate to dryness, add methanol to the residue to dissolve, make up to 5 mL volumetric flask, filter the filtrate through a 0.22 μm microporous filter membrane to obtain the Uncaria rhynchophylla test solution.

[0107] Preparation of the test solution of Ziziphus jujuba seed: weigh 7 g of Ziziphus jujuba seed, add 120 mL of water and soak for 24 h, then decoct twice, the first time for 6 h and the second time for 2 h, combine the decoctions and concentrate to dryness, add 2 mL of ammonia water (mass concentration 25-28%), add 25 mL of chloroform, heat and reflux for 30 min, filter, concentrate the filtrate to dryness, add methanol to the residue to dissolve, make up to 5 mL volumetric flask, filter the filtrate through a 0.22 μm microporous filter membrane to obtain the Ziziphus jujuba seed test solution.

[0108] Preparation of Rauwolfia test solution: weigh 0.04 g of total alkaloids of Rauwolfia, add 120 mL of water and soak for 24 h, then decoct twice, the first time for 6 h and the second time for 2 h, combine the decoctions and concentrate to dryness, add 2 mL of ammonia water (mass concentration 25-28%), add 25 mL of chloroform, heat and reflux for 30 min, filter, concentrate the filtrate to dryness, add methanol to the residue to dissolve, make up to 5 mL volumetric flask, filter the filtrate through a 0.22 μm microporous filter membrane to obtain the Rauwolfia test solution.

[0109] According to the HPLC detection conditions of Example 1, HPLC detection was performed on the test solution of each single medicinal material.

[0110] Figure 7 This is the HPLC chromatogram of the Uncaria rhynchophylla test solution. Figure 8 This is the HPLC chromatogram of the test solution of Ziziphus jujuba seed. Fig. 9 This is the HPLC chromatogram of the Rauwolfia test solution. Figures 1-2 and Figures 7 to 9 By comparison, the fingerprint of Xinlekang tablets was obtained, and three components were identified, namely isorhynchophylline (peak 10), rhynchophylline (peak 15), and reserpine (peak 28). Figure 1 By comparison, peaks 2, 13, 16, 22 and 27 belong to Semen Ziziphi Spinosae, peaks 1-12 and 15-27 belong to Uncaria rhynchophylla, and peaks 4, 6, 8, 11, 14, 20, 27 and 28 belong to Rauwolfia ovata.

[0111] The present invention established HPLC fingerprints of 12 batches of Xinlekang tablets, with similarities greater than 0.988. Through systematic component identification and attribution of single medicinal materials with common peaks in different batches of drugs, the chemical information contained in Xinlekang tablets and the current status of each component are comprehensively reflected, which can more comprehensively reflect the quality of Xinlekang tablets and provide a reference basis for quality control of Xinlekang tablets.

[0112] Example 4

[0113] Investigation of the preparation method of test solution

[0114] The HPLC detection conditions are the same as those in Example 1.

[0115] (1) Investigation of extraction methods

[0116] Common extraction methods include reflux extraction, maceration extraction, ultrasonic extraction, Soxhlet extraction, etc. Since Soxhlet extraction is cumbersome, reflux extraction, maceration extraction and ultrasonic extraction were investigated.

[0117] Reflux extraction: Preparation method of the test solution in Example 1.

[0118] Immersion extraction: The 30 min heating reflux extraction in the preparation process of the test solution in Example 1 was replaced by immersion extraction at room temperature for 12 h.

[0119] Ultrasonic extraction: The heating reflux extraction for 30 min in the preparation process of the test solution in Example 1 was replaced by ultrasonic extraction for 30 min at room temperature, power 220 W, and frequency 40 kHz.

[0120] HPLC detection was performed on each test solution. The results showed that the chromatographic area of ​​reflux extraction was significantly larger than that of immersion extraction and ultrasonic extraction. Therefore, reflux extraction was selected.

[0121] (2) Investigation of extraction solvent

[0122] The effects of methanol, ethanol, chloroform and dichloromethane extraction were investigated respectively. The chloroform in the preparation process of the test solution in Example 1 was replaced with ethanol, chloroform and dichloromethane respectively. Each test solution was tested by HPLC, and the results showed that the chromatographic peaks of the chloroform extract were the most, the chromatographic peaks of methanol and ethanol extraction were significantly less than those of chloroform, and the chromatographic peaks of dichloromethane extraction were less than those of chloroform. Therefore, chloroform with the most extraction peaks was selected as the extraction solvent.

[0123] (3) Investigation of extraction time

[0124] The test solution was prepared according to the preparation method of Example 1, and the extraction time was 20min, 30min, and 40min respectively. HPLC detection was performed on each test solution, and the results showed that the extraction effects of the three were relatively similar, and the total peak area was the largest when the reflux extraction was performed for 30min, so the reflux time was selected to be 30min.

[0125] Example 5

[0126] Investigation of test methods for test products

[0127] (1) Investigation of detection wavelength

[0128] The test solution prepared in Example 1 was scanned at full wavelength using a diode array detector and the spectrum was analyzed. Fig.10 It can be seen that the HPLC spectrum at a wavelength of 204nm has the most chromatographic peaks and the separation effect is the best, so 204nm is selected as the measurement wavelength of the fingerprint spectrum.

[0129] (2) Investigation of mobile phase composition

[0130] The test solution was subjected to HPLC detection according to the method of Example 1, and the only difference from Example 1 was that the mobile phase B was 0.2 v / v% phosphoric acid aqueous solution and 0.4 v / v% phosphoric acid aqueous solution. The results showed that when the acetonitrile-0.4 v / v% phosphoric acid aqueous solution elution system was used, the separation degree of each component in the test sample chromatogram was better.

[0131] The effect of the amount of triethylamine added in mobile phase B (0.05 v / v%, 0.06 v / v% and 0.07 v / v%) on peak shape and separation effect was further investigated. The results showed that the elution effect was best when 0.05 v / v% triethylamine was added. Therefore, acetonitrile was selected as mobile phase A and 0.4 v / v% phosphoric acid-0.05 v / v% triethylamine-aqueous solution was selected as mobile phase B for gradient elution.

[0132] (3) Investigation of gradient elution procedure

[0133] The test solution was subjected to HPLC detection according to the method of Example 1. The only difference from Example 1 is that the gradient elution procedures are shown in Tables 5 to 7, and the HPLC spectra of the test solution are shown in Tables 5 to 7. Fig.11 (Table 5), Fig.12 (Table 6) and Fig.13 (Table 7).

[0134] Table 5 Gradient elution program

[0135] Time / min Mobile phase A volume fraction / % Mobile phase B volume fraction / % 0 20 80 20 25 75 55 40 60 70 70 30 71 90 10 75 90 10 80 20 80 90 20 80

[0136] Table 6 Gradient elution program

[0137] Time / min Mobile phase A volume fraction / % Mobile phase B volume fraction / % 0 15 85 30 25 75 65 40 60 80 70 30 81 90 10 85 90 10 86 15 85 91 15 85

[0138] Table 7 Gradient elution program

[0139] Time / min Mobile phase A volume fraction / % Mobile phase B volume fraction / % 0 20 80 10 25 75 20 25 75 25 40 60 35 40 60 50 70 30 51 90 10 55 90 10 60 20 80 65 20 80

[0140] By comparison Figure 1 and Figures 11 to 13 It can be seen that, compared with the gradient elution programs in Tables 5 to 7, the gradient elution program shown in Table 2 has more chromatographic peaks and better separation effect of each peak.

[0141] Example 6

[0142] Methodological investigation

[0143] Stability test: Prepare the test solution according to the method of Example 1, and perform HPLC determination at 0h, 2h, 6h, 10h, 24h, and 48h after preparation (the detection conditions are the same as those in Example 1), record the fingerprint, and calculate the RSD values ​​of the relative peak area and relative retention time of each common peak. The results show that the relative retention time RSD of the common peak with a single peak area greater than 5% of the total peak area is less than 1%, and the RSD of the relative peak area is less than 3%. The chromatographic fingerprints of the five test samples measured were visually observed without obvious changes in the overall picture. The data file was imported into the "Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System (2012 Edition)" for similarity evaluation. The results show that the similarity of the fingerprints of each test sample is greater than 0.98, indicating that the sample has good stability within 48h and meets the requirements of fingerprint research technology.

[0144] Precision test: 15 μL of the same test solution was accurately aspirated, and the measurement was performed 5 times in a row, and the chromatogram was recorded. With rhynchophylline as the reference peak, the relative retention time and peak area ratio of each characteristic peak were calculated, and the RSD value was calculated. The results showed that the relative retention time RSD of the common peak with a single peak area greater than 5% of the total peak area was less than 1%, and the relative peak area RSD was less than 3%. The chromatographic fingerprints of the five test samples measured showed no obvious changes in the overall picture. The data file was imported into the "Similarity Evaluation System for Chromatographic Fingerprints of Traditional Chinese Medicine (2012 Edition)" for similarity evaluation. The results showed that the similarity of the fingerprints of each test sample was greater than 0.99, indicating that the instrument had good precision and met the requirements of fingerprint research technology.

[0145] Reproducibility test: According to the method of Example 1, the same batch of test sample powder was weighed and 5 test sample solutions were prepared in parallel. The solution was measured by HPLC and the chromatogram was recorded. With rhynchophylline as the reference peak, the relative retention time and peak area ratio of each characteristic peak were calculated, and its RSD value was calculated. The results showed that the relative retention time RSD of the common peak with a single peak area greater than 5% of the total peak area was less than 1%, and the RSD of the relative peak area was less than 3%. There was no obvious change in the intuitive overall picture of the chromatographic fingerprints of the 5 test samples measured. The data file was imported into the "Similarity Evaluation System of Chromatographic Fingerprints of Traditional Chinese Medicine (2012 Edition)" for similarity evaluation. The results showed that the similarity of the fingerprints of each test sample was greater than 0.98, indicating that the method had good repeatability and met the requirements of fingerprint research technology.

[0146] Traditional Chinese medicine fingerprint is an internationally recognized research model and technical platform for controlling the quality of traditional Chinese medicine. It analyzes the types and content distribution information of the effective ingredients and ineffective ingredients of traditional Chinese medicine, so that the research methods and quality analysis methods of traditional Chinese medicine have been developed from the analysis of one or a few active ingredients to the comprehensive analysis of the entire chemical fingerprint of traditional Chinese medicine, which is universal and economical. The fingerprint of traditional Chinese medicine reflects the synergistic effect, multi-level complexity and integrity of traditional Chinese medicine. There are many kinds of traditional Chinese medicine materials in the formula of Xinlekang tablets, and its material group must be controlled as a whole. In addition to "microscopic analysis", a certain "macroscopic analysis" method should also be used to effectively characterize the quality of traditional Chinese medicine as a whole. The present invention can obtain fingerprints of 28 characteristic peaks by controlling the extraction conditions and HPLC detection conditions, which can effectively characterize the quality of Xinlekang tablets and is conducive to comprehensive monitoring of the quality of drugs. And the method of the present invention has the advantages of simple method, stability, high precision and good reproducibility.

[0147] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A method for constructing a fingerprint of Xinlekang tablets, comprising the following steps: The powder of Xinlekang tablets, ammonia water and chloroform are mixed and extracted to obtain an extract, and the extract is concentrated and dissolved in methanol to obtain a test solution; The test solution and the mixed reference solution are subjected to high performance liquid chromatography detection respectively to obtain a sample liquid phase spectrum and a reference liquid phase spectrum respectively, and a fingerprint spectrum of Xinlekang tablets is obtained according to the sample liquid phase spectrum and the reference liquid phase spectrum; The reference substances in the mixed reference solution include isorhynchophylline, rhynchophylline and reserpine; The conditions for the high performance liquid chromatography detection include: the chromatographic column includes a C18 chromatographic column; the mobile phase A is acetonitrile, the mobile phase B is a phosphoric acid-triethylamine aqueous solution, the volume fraction of phosphoric acid in the mobile phase B is 0.38-0.42%, and the volume fraction of triethylamine is 0.04-0.06%; the elution method is gradient elution, and the procedure of the gradient elution includes: 0-15min, the volume fraction of the mobile phase A increases from 14-16% to 20-22%; 15-30min, the volume fraction of the mobile phase A increases from 20-22% to 24-26%; 30-65min, the volume fraction of the mobile phase A increases from 20-22% to 24-26% The volume fraction of mobile phase A increases from 24-26% to 39-41%; at 65-70min, the volume fraction of mobile phase A increases from 39-41% to 48-52%; at 70-71min, the volume fraction of mobile phase A increases from 48-52% to 88-92%; at 71-75min, the volume fraction of mobile phase A is 88-92%; at 75-76min, the volume fraction of mobile phase A decreases from 88-92% to 14-16%; at 76-81min, the volume fraction of mobile phase A is 14-16%; the detection wavelength is 202-206nm; The fingerprint spectrum of Xinlekang tablets contains 28 characteristic peaks, including the characteristic peaks of isorhynchophylline, rhynchophylline and reserpine.

2. The fingerprint construction method according to claim 1, characterized in that: The solid-liquid ratio of the powder of the Xinlekang tablet to the ammonia water is 1g:1.8-2.2mL; the mass fraction of the ammonia water is 25-28%; The solid-liquid ratio of the powder of the Xinlekang tablets to chloroform is 1g:24-26mL.

3. The fingerprint construction method according to claim 1 or 2, characterized in that: The extraction is reflux extraction, and the extraction time is 28 to 32 minutes.

4. The fingerprint construction method according to claim 1, characterized in that: In the mixed reference solution, the concentration of isorhynchophylline is 7.6-7.8 μg / mL, the concentration of rhynchophylline is 7.9-8.1 μg / mL, and the concentration of reserpine is 4.7-4.9 μg / mL; The solvent in the mixed reference solution includes methanol.

5. The fingerprint construction method according to claim 1, characterized in that: The conditions for the high performance liquid chromatography detection also include: a mobile phase flow rate of 0.7 to 0.9 mL / min, a diode array detector, a column temperature of 28 to 32° C., and an injection volume of 9.5 to 10.5 μL.

6. The fingerprint construction method according to claim 1, characterized in that: The liquid phase spectrum of the sample and the liquid phase spectrum of the reference substance were calculated by the software "Similarity Evaluation System of Traditional Chinese Medicine Chromatographic Fingerprint Spectrum", and the similarities were both greater than 0.

95.

7. The fingerprint of Xinlekang tablets obtained by the fingerprint construction method according to any one of claims 1 to 6 contains 28 characteristic peaks, including the characteristic peaks of isorhynchophylline, rhynchophylline and reserpine.

8. Application of the fingerprint of Xinlekang tablets as claimed in claim 7 in the quality control of Xinlekang tablets.

9. A multi-index component detection method for Xinlekang tablets, comprising the following steps: The powder of the Xinlekang tablets to be tested, ammonia water and chloroform are mixed and extracted to obtain an extract solution, and the extract solution is concentrated and dissolved in methanol to obtain a sample solution to be tested; The sample solution to be tested is subjected to high performance liquid chromatography to obtain a multi-index component test result of Xinlekang tablets; the multi-index components include at least two of isorhynchophylline, rhynchophylline and reserpine; The conditions for the high performance liquid chromatography detection include: the chromatographic column includes a C18 chromatographic column; the mobile phase A is acetonitrile, the mobile phase B is a phosphoric acid-triethylamine aqueous solution, the volume fraction of phosphoric acid in the mobile phase B is 0.38-0.42%, and the volume fraction of triethylamine is 0.04-0.06%; the elution method is gradient elution, and the procedure of the gradient elution includes: 0-15min, the volume fraction of the mobile phase A increases from 14-16% to 20-22%; 15-30min, the volume fraction of the mobile phase A increases from 20-22% to 24-26%; 30-65min, the volume fraction of the mobile phase A increases from 20-22% to 24-26% The volume fraction of mobile phase A increases from 24-26% to 39-41%; from 65 to 70 min, the volume fraction of mobile phase A increases from 39-41% to 48-52%; from 70 to 71 min, the volume fraction of mobile phase A increases from 48-52% to 88-92%; from 71 to 75 min, the volume fraction of mobile phase A is 88-92%; from 75 to 76 min, the volume fraction of mobile phase A decreases from 88-92% to 14-16%; from 76 to 81 min, the volume fraction of mobile phase A is 14-16%; the detection wavelength is 202 to 206 nm.

10. The multi-index component detection method according to claim 9, characterized in that: The solid-liquid ratio of the powder of the Xinlekang tablet to the ammonia water is 1g:1.8-2.2mL; the mass fraction of the ammonia water is 25-28%; The solid-liquid ratio of the powder of the Xinlekang tablet to chloroform is 1g:24-26mL; The extraction is reflux extraction, and the extraction time is 28 to 32 minutes; The conditions for the high performance liquid chromatography detection also include: a mobile phase flow rate of 0.7 to 0.9 mL / min, a diode array detector, a column temperature of 28 to 32° C., and an injection volume of 9.5 to 10.5 μL.

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