Method for establishing fingerprint spectrum of xinsuning capsule and fingerprint spectrum

Abstract

To provide a method for establishing a fingerprint spectrum of a xinsuning capsule, and a fingerprint spectrum.SOLUTION: The present invention includes the steps of: collecting contents of xinsuning capsules with different batches, adding a methanol aqueous solution to perform ultrasonic extraction, and obtaining a test solution (S1); injecting the test solution into a rapid liquid chromatograph, performing gradient elution, performing chromatography analysis, and recording 0-140 min chromatogram (S2); introducing the chromatogram into the traditional Chinese medicine chromatographic Fingerprint similarity evaluation system for processing to obtain a fingerprint spectrum (S3); and performing mass spectrometry on the test solution and determining a chemical composition of chromatographic peaks in the fingerprint spectrum on the basis of mass spectrometry results (S4). A method for detecting a xinsuning capsule fingerprint spectrum is easy, stable, precise, and can be highly regenerated. Quality of xinsuning capsules can be evaluated comprehensively and scientifically and quality of a product and the effect of treatment can be guaranteed.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to the field of traditional Chinese medicine detection, and in particular to the fingerprint spectrum construction method and fingerprint spectrum of Xinshuoning Capsule. [Background technology]

[0002] Xin Su Ning Capsules are prescribed for the purpose of clearing heat and eliminating phlegm, calming the heart and stopping palpitations, which conforms to the theory of Lord, Minister, Assistant and Secretary in traditional Chinese medicine. The principal drugs in the prescription are Coptis Rhizome and Portulaca japonica, which are for clearing heat and eliminating phlegm, the subordinate drugs are Sweet Artemisia Herb, Dichroa febrifuga Lure, Clam, and Plumera nelumbinis, which help the principal drugs to clear heat, eliminate phlegm and detoxify, clear the mind and calm the spirit, and the auxiliary drug is Poria cocos. , Aurantii immatulus fructus, which regulates qi, expels dampness and dispels phlegm; ginseng and dwarf lily turf tubers are also auxiliary medicines that nourish the mind and restore the pulse; licorice is an emissary that harmonizes the various medicines; at the same time, Coptis Rhizome, Melilotus japonica and Dichroa febrifuga-rue release acid and reduce bitterness, which, together with Aurantii immatulus fructus, eliminate stagnation of qi and regulate the pulse.

[0003] "Shinsokunei Capsules" is a compound medicine consisting of 11 kinds of herbal medicines, namely, Coptis Rhizome, Melastoma Root, Poria Cocos, Aurantium Inmatulus Fructus, Dichroa Febrifuga Lure, Plumula Nelumbinis, Clam, Sweet Artemisia Herb, Ginseng, Dwarf Lily Turf Tubers, and Licorice. It is used to treat palpitations, chest tightness, heartburn, startling, dry mouth, bitter mouth, insomnia, hypersomnia, dizziness, and arrhythmia caused by phlegm heat, and is suitable for mild to moderate ventricular premature beats caused by coronary heart disease and viral myocarditis.

[0004] The quality of Chinese herbal medicine is directly related to its development. In recent years, Chinese herbal medicine has attracted more and more attention worldwide due to its reliable efficacy, few side effects, low toxicity, and low drug resistance, but the quality control of Chinese herbal medicine is becoming increasingly difficult due to the complexity of the chemical composition of Chinese herbal medicine and the interactions between ingredients. At present, the quality control of Xinshuning Capsules is relatively simple, but the testing method is complicated. The current national standard for Xinshuning Capsules only requires the content of berberine hydrochloride, which cannot reflect the overall composition of Xinshuning Capsules, and cannot provide a basis for quality control and evaluation. Summary of the Invention [Problem to be solved by the invention]

[0005] SUMMARY OF THE PRESENT EMBODIMENT In view of the technical shortcomings of the existing detection methods, the objective of the present invention is to provide a fingerprint spectrum construction method and fingerprint spectrum for cardiac speed-up capsule. [Means for solving the problem]

[0006] The present invention is achieved by the following technical solutions:

[0007] The fingerprint spectrum construction method of Xinshuning Capsule is as follows: S1, taking different batches of cardiac stimulating capsule contents, adding methanol aqueous solution, and ultrasonically extracting to obtain a test solution; S2, injecting the test solution into a high performance liquid chromatograph, performing gradient elution, performing chromatographic analysis, and recording a chromatogram from 0 to 140 min; S3, the chromatogram obtained in S2 is introduced into the herbal medicine chromatogram fingerprint spectrum similarity evaluation system, and the chromatogram peaks existing in the chromatograms of different batches of Xinshuning capsules are selected as common peaks, and the chromatograms of the test solution are respectively subjected to data introduction, multi-point correction and data matching to obtain a fingerprint spectrum; S4, performing mass spectrometry on the test solution, and determining the chemical composition of the chromatogram peaks in the fingerprint spectrum based on the mass spectrometry results.

[0008] Preferably, in S1, the volume concentration of methanol in the aqueous methanol solution is 80%.

[0009] Preferably, in S1, the ultrasonic extraction time is 20 to 40 minutes.

[0010] Preferably, in S2, the detection wavelength used for the chromatographic analysis is 210 nm.

[0011] Preferably, in S2, the chromatographic column used in the high performance liquid chromatograph is a Hedera ODS-2-C18 (250 mm×4.6 mm, 10 μm) chromatographic column.

[0012] Preferably, in S2, the mobile phase used for gradient elution is acetonitrile and 0.1% formic acid in water.

[0013] Preferably, in S2, the gradient elution procedure is as shown in the table below.

[0014] TIFF2025031696000001.tif42168

[0015] Preferably, in S2, the flow rate used for gradient elution is 0.8 mL / min.

[0016] Preferably, in S4, the chemical compositions of the chromatogram peaks in the fingerprint spectrum are determined to be: Peak 1 trifolylrhizin, Peak 2 naringin, Peak 3 kaempferide, Peak 4 valine, Peak 5 palmatine, Peak 6 epiberberine, Peak 7 glycyrrhizin / isoglycyrrhizin, Peak 8 limonin, Peak 9 liquiritin, Peak 10 picrotoxin, Peak 11 methylopiogonanone A, Peak 12 rotacin, Peak 13 sophoridine / sophocarpidine, Peak 14 sophoranol / sophocarpidine oxide, Peak 15 kurarinone, Peak 16 dehydropatiminic acid, Peak 17 polyporenic acid C, Peak 18 quercetin, and Peak 19 sophocarpine.

[0017] The present invention provides a fingerprint spectrum of a cardiac speed-up capsule constructed by the method according to any one of claims 1 to 9. Compared with the existing technology, the present invention has the following beneficial effects:

[0018] According to the structural characteristics of the active ingredients contained in Xinshuning capsules, the present invention establishes a method for detecting its fingerprint spectrum. Through the verification of many experiments, it has been found that the fingerprint spectrum detection method of Xinshuning capsules provided by the present invention has better separation of each chromatogram peak, smooth baseline, good peak shape, and can comprehensively respond to the type and amount of chemical ingredients contained in the capsules. Through the HPLC fingerprint spectrum of Xinshuning capsules constructed by the present invention, 19 common peaks and standard fingerprint spectra are obtained and recognized, and the obtained chromatogram similarity is high, and the stability, reproducibility and accuracy are calculated using the relative retention time and relative peak area, and the RSD values ​​of the relative retention time and relative peak area are both less than 5.0%, which shows that the method provided by the present invention has good reproducibility and the fingerprint spectrum constructed by the method provided by the present invention is reliable. The fingerprint spectrum detection method of Xinshuning capsules provided by the present invention has the advantages of simple method, stable, high accuracy and good reproducibility, which can more comprehensively and scientifically evaluate the quality of Xinshuning capsules and ensure the quality and therapeutic effect of the product. [Brief description of the drawings]

[0019] [Figure 1] 1 is a chromatogram obtained in Example 1 during the detection wavelength optimization process of the present invention. [Diagram 2] 1 is a chromatogram obtained in Example 2 during the flow rate optimization process of the present invention. [Diagram 3] 1 is a chromatogram obtained in Example 3 during the elution gradient optimization process of the present invention. [Figure 4] 1 is a chromatogram obtained in Example 4 during the elution gradient optimization process of the present invention. [Diagram 5] 1 is a chromatogram obtained in Example 5 during the elution gradient optimization process of the present invention. [Figure 6] 1 is a chromatogram obtained in Example 6 during the elution gradient optimization process of the present invention. [Figure 7] 1 is a chromatogram obtained in Example 7 of the optimal method of the present invention. [Figure 8] 1 is a mass spectrum of trifolirhizin of the present invention. [Figure 9] 1 is a mass spectrum of naringin of the present invention. [Figure 10] 1 is a mass spectrum of Kaempferide of the present invention. [Figure 11] 1 is a mass spectrum of valine of the present invention. [Figure 12] 1 is a mass spectrum of palmatine of the present invention. [Figure 13] 1 is a mass spectrum of epiberberine of the present invention. [Figure 14] 1 is a mass spectrum of glycyrrhizin / isoglycyrrhizin of the present invention. [Figure 15] 1 is a mass spectrum of limonin of the present invention. [Figure 16] 1 is a mass spectrum of liquiritin of the present invention. [Figure 17] 1 is a mass spectrum of picrotoxin of the present invention. [Figure 18]1 is a mass spectrum of methylopiogonanone A of the present invention. [Figure 19] 1 is a mass spectrum of rotacin of the present invention. [Figure 20] 1 is a mass spectrum of sophoridine / sophocarpidine of the present invention. [Figure 21] 1 is a mass spectrum of sophoranol / sophocarpidine oxide of the present invention. [Figure 22] 1 is a mass spectrum of the clarinone of the present invention. [Diagram 23] 1 is a mass spectrum of dehydropatiminic acid of the present invention. [Figure 24] 1 is a mass spectrum of polyboronic acid C of the present invention. [Diagram 25] 1 is a mass spectrum of quercetin of the present invention. [Figure 26] 1 is a mass spectrum of sophocarpine of the present invention. [Figure 27] 1 is a fingerprint spectrum of 15 batches of test samples based on the Xinxing capsule of the present invention. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0020] In order to better understand the present invention, the present invention will be described below in conjunction with examples, which are used to further interpret the features and advantages of the present invention and are not intended to limit the scope of the claims of the present invention.

[0021] 1. The fingerprint spectrum construction method of Xinshuning Capsule includes the following steps: S1. Preparation of test solutions: 0.3-1.0 g of the contents of Shinsokuning capsules from different batches were precisely weighed and placed in a stoppered conical flask, 20-30 mL of aqueous methanol solution was added, and ultrasonic extraction was performed for 20-40 min. The subsequent filtrate was filtered through a 0.45 μm microporous filtration membrane to obtain the test solution. S2: 15-30 μL of the test solution prepared in S1 is precisely aspirated and injected into the high performance liquid chromatograph, gradient elution is performed, and the chromatogram from 0 to 140 min is recorded. S3, the chromatograms obtained in S2 are derived and introduced into the Chinese medicine chromatogram fingerprint spectrum similarity evaluation system (2004 A version), and the chromatogram peaks existing in the chromatograms of different batches of Xinshuning capsules are selected as common peaks, and data introduction, multi-point correction and data matching are respectively performed on the chromatograms of the test solutions to obtain fingerprint spectra and perform similarity analysis, and the similarity is more than 0.90; S4, determine the chemical composition in the fingerprint spectrum, perform mass spectrometry on the above test solution to obtain a total ion flow diagram, introduce the detection data into Xcalibur software, input into the Qual Browser interface, and perform data analysis according to the chemical composition peak situation.

[0022] The preparation method of the test solution described in S1 above is as follows: 0.5 g of the contents of 15 batches of Shinsokuning capsules are precisely weighed and placed into a stoppered conical flask, 25 mL of 80% aqueous methanol solution is added, and ultrasonic extraction is performed for 30 minutes. The subsequent filtrate is filtered through a 0.45 μm microporous filtration membrane to obtain the test solution.

[0023] The liquid phase chromatogram conditions described in S2 above are as follows: chromatogram column: Hedera ODS-2-C18 (250 mm × 4.6 mm, 10 μm) chromatogram column, mobile phase: acetonitrile (A) and 0.1% formic acid aqueous solution (B), gradient elution, detection wavelength: 210 nm, column temperature 30 °C, flow rate 0.8 mL / min, sample injection volume: 20 μL, elution procedure: 0 min, 20% A, 0 ~ 35 min, 20% ~ 27% A, 35 ~ 45 min, 27% ~ 35% A, 45 ~ 85 min, 35 ~ 45% A, 85 ~ 90 min, 45 ~ 65% A, 90 ~ 140 min, 65 ~ 100% A.

[0024] 2. Optimization of fingerprint spectrum detection S1. Optimization of extraction method The present invention carried out experimental comparisons through different extraction methods (ultrasonic 20min, ultrasonic 30min, reflux 60min) and different extraction solvents (ultrapure water, 80% methanol aqueous solution, methanol). As a result, it was found that the spectral composition obtained by 30min ultrasonic extraction was more comprehensive and the degree of separation was good, so the 30min ultrasonic extraction method was adopted. In the study of the extraction solvent, the chromatogram information volume of the 80% methanol aqueous solution extract was the largest and the composition content was the highest, so 80% methanol aqueous solution was used for extraction.

[0025] S2, Optimization of chromatographic conditions In the present invention, the detection wavelengths (210 nm, 254 nm, 275 nm, 300 nm) were examined using an ultraviolet-visible light absorption detector, and it was found that when the detection wavelength condition was 210 nm, the amount of information contained in the chromatogram was the most comprehensive and the baseline was the smoothest, so this method was selected as the detection wavelength condition.

[0026] In the present invention, flow rates (0.6 mL / min, 0.8 mL / min, 1.0 mL / min) were screened, and it was found that a flow rate of 0.8 mL / min provided the best separation effect for each substance and the highest degree of separation, and therefore a flow rate of 0.8 mL / min was adopted.

[0027] In the present invention, after screening temperatures (25°C, 30°C, 35°C), the effect of temperature on peak time and peak number is small, and the separation effect of each substance is good, so a temperature of 30°C is selected.

[0028] In the present invention, the elution effect under different gradients of several different elution systems, such as acetonitrile-water, methanol-water, acetonitrile-0.2% phosphoric acid aqueous solution, acetonitrile-0.1% formic acid, and acetonitrile-0.1% glacial acetic acid, is compared. As a result, when acetonitrile and 0.1% formic acid aqueous solution are used as mobile phase, the peak conditions of each component in the heart speed neem capsule are good, and the separation effect is high, so finally, acetonitrile and 0.1% formic acid aqueous solution are selected as mobile phase.

[0029] After determining the optimal mobile phase composition, the present invention screens the optimal gradient elution procedure through a large number of experiments. The experimental results show that by using 0 min acetonitrile volume 20%, 0-35 min acetonitrile volume 20%-27%, 35-45 min acetonitrile volume 27%-35%, 45-85 min acetonitrile volume 35%-45%, 85-90 min acetonitrile volume 45%-65%, and 90-140 min acetonitrile volume 65%-100%, good separation of each chromatogram peak in the fingerprint spectrum can be achieved.

[0030] The equipment used in the present invention is shown in Table 1 below, the reagents used in the present invention are shown in Table 2 below, and the drugs used in the present invention are shown in Table 3 below.

[0031] [Table 1]

[0032] [Table 2]

[0033] [Table 3]

[0034] Hereinafter, embodiments of the present invention will be described in detail with reference to specific examples.

[0035] Example 1 The method for constructing a fingerprint spectrum of the heart speed capsule includes the following steps: S1. Preparation of test solutions: 0.5 g of the contents of the Shinsokunei capsule was precisely weighed and placed in a stoppered conical flask, 25 mL of aqueous methanol solution was added, and ultrasonic extraction was performed for 30 minutes. The subsequent filtrate was filtered through a 0.45 μm microporous filtration membrane to obtain a test solution. S2, chromatographic conditions Chromatography column: Hedera ODS-2-C18 (250 mm × 4.6 mm, 5 μm) chromatogram column, mobile phase: acetonitrile and 0.1% formic acid aqueous solution, gradient elution, detection wavelength: 210 nm / 254 nm / 270 nm / 300 nm, column temperature 30 ° C, flow rate 0.8 mL / min, sample injection volume: 20 μL, elution procedure is shown in Table 4 below.

[0036] [Table 4]

[0037] The final experimental results are shown in Figure 1. As can be seen from the chromatograms, the resolution was generally poor under the four detection wavelengths, with few peaks in the 0-255 min range. However, when the detection wavelength was 210 nm, the peak situation in the 25-40 min range was better, so 210 nm detection was used in the subsequent optimization process.

[0038] Example 2 The method for constructing a fingerprint spectrum of the heart speed capsule includes the following steps: S1. Preparation of test solutions: 0.5 g of the contents of the Shinsokunei capsule was precisely weighed and placed in a stoppered conical flask, 25 mL of aqueous methanol solution was added, and ultrasonic extraction was performed for 30 minutes. The subsequent filtrate was filtered through a 0.45 μm microporous filtration membrane to obtain a test solution. S2, chromatographic conditions Chromatography column: Hedera ODS-2-C18 (250 mm × 4.6 mm, 5 μm) chromatogram column, mobile phase: acetonitrile and 0.1% formic acid aqueous solution, gradient elution, detection wavelength: 210 nm, column temperature 30 °C, flow rate 0.6 mL / min, 0.8 mL / min, 1 mL / min, sample injection volume: 20 μL, elution procedure is shown in Table 5 below.

[0039] [Table 5]

[0040] The final experimental results are shown in Figure 2. As can be seen from the chromatogram, when the flow rate is 0.6 mL / min, there are many peaks, but the resolution between adjacent peaks is low, and the peaks after 125 min are incomplete; when the flow rate is 1 mL / min, there are few peaks, and there are no peaks at 75-100 min, which cannot meet the fingerprint spectrum requirements; when the flow rate is 0.8 mL / min, the chromatogram peak conditions and resolution are good, so a flow rate of 0.8 mL / min is selected in the subsequent examples.

[0041] Example 3 The method for constructing and detecting the fingerprint spectrum of the heart rate monitoring capsule includes the following steps: S1. Preparation of test solutions: 0.5 g of the contents of the Shinsokunei capsule was precisely weighed and placed in a stoppered conical flask, 25 mL of 80% aqueous methanol solution was added, and ultrasonic extraction was performed for 30 minutes. The subsequent filtrate was filtered through a 0.45 μm microporous filtration membrane to obtain a test solution. S2, chromatographic conditions Chromatography column: Hedera ODS-2-C18 (250 mm × 4.6 mm, 5 μm) chromatogram column, mobile phase: acetonitrile and 0.1% formic acid aqueous solution, gradient elution, detection wavelength: 210 nm, column temperature: 30 °C, flow rate: 0.8 mL / min, sample injection volume: 20 μL, elution procedure is shown in Table 6 below.

[0042] [Table 6]

[0043] The final experimental results are shown in Figure 3. As can be seen from the chromatogram, the overall resolution is poor, there are few peaks and low content in the 55-75 min period, and there are many peaks in the 30-45 min period, but the peak density is large. During the optimization process, we will focus on separating the peaks in this period, and consider the effect of the mobile phase organic phase concentration on the peak interval, and then continue to optimize the concentration gradient.

[0044] Example 4 The method for constructing a fingerprint spectrum of the heart speed capsule includes the following steps: S1. Preparation of test solutions: 0.5 g of the contents of the Shinsokunei capsule was precisely weighed and placed in a stoppered conical flask, 25 mL of 80% aqueous methanol solution was added, and ultrasonic extraction was performed for 30 minutes. The subsequent filtrate was filtered through a 0.45 μm microporous filtration membrane to obtain a test solution. S2, chromatographic conditions Chromatography column: Hedera ODS-2-C18 (250 mm × 4.6 mm, 5 μm) chromatogram column, mobile phase: acetonitrile and 0.1% formic acid aqueous solution, gradient elution, detection wavelength: 210 nm, column temperature: 30 °C, flow rate: 0.8 mL / min, sample injection volume: 20 μL, elution procedure is shown in Table 7 below.

[0045] [Table 7]

[0046] The final experimental results are shown in Figure 4. As can be seen from the chromatogram, the separation effect is improved to a certain extent in 30 to 50 minutes, but the overall resolution cannot meet the fingerprint spectrum requirements, so the optimization of the concentration gradient is continued.

[0047] Example 5 The method for constructing a fingerprint spectrum of the heart speed capsule includes the following steps: S1. Preparation of test solutions: 0.5 g of the contents of the Shinsokunei capsule was precisely weighed and placed in a stoppered conical flask, 25 mL of 80% aqueous methanol solution was added, and ultrasonic extraction was performed for 30 minutes. The subsequent filtrate was filtered through a 0.45 μm microporous filtration membrane to obtain a test solution. S2, chromatographic conditions Chromatography column: Hedera ODS-2-C18 (250 mm × 4.6 mm, 5 μm) chromatogram column, mobile phase: acetonitrile and 0.1% formic acid aqueous solution, gradient elution, detection wavelength: 210 nm, column temperature: 30 ° C, flow rate: 0.8 mL / min, sample injection volume: 20 μL, elution procedure is shown in Table 8 below.

[0048] [Table 8]

[0049] The final experimental results are shown in Figure 5. As can be seen from the chromatogram, the overall peak condition is good, and the resolution of the chromatogram peaks is slightly improved. However, after 120 minutes, the peaks become incomplete, so we continue to optimize the concentration gradient and extend the elution time.

[0050] Example 6 The method for constructing a fingerprint spectrum of the heart speed capsule includes the following steps: S1. Preparation of test solutions: 0.5 g of the contents of the Shinsokunei capsule was precisely weighed and placed in a stoppered conical flask, 25 mL of 80% aqueous methanol solution was added, and ultrasonic extraction was performed for 30 minutes. The subsequent filtrate was filtered through a 0.45 μm microporous filtration membrane to obtain a test solution. S2, chromatographic conditions Chromatography column: Hedera ODS-2-C18 (250 mm × 4.6 mm, 5 μm) chromatogram column, mobile phase: acetonitrile and 0.1% formic acid aqueous solution, gradient elution, detection wavelength: 210 nm, column temperature: 30 ° C, flow rate: 0.8 mL / min, sample injection volume: 20 μL, elution procedure is shown in Table 9 below.

[0051] [Table 9]

[0052] The final experimental results are shown in Figure 6. As can be seen from the chromatogram, the peak situation after 100 min has improved to some extent, but the peaks are dense and the resolution is poor. Considering the effect of the mobile phase organic phase concentration on the peak interval, we will continue to optimize the concentration gradient.

[0053] Example 7 The method for constructing a fingerprint spectrum of the heart speed capsule includes the following steps: S1. Preparation of test solutions 15 batches of Xinsuo Ning capsule contents were weighed to a mass of 0.5g and placed in a stoppered conical flask, and 25mL of 80% methanol aqueous solution was added. Ultrasonic extraction was performed for 30 minutes, and the subsequent filtrate was filtered through a 0.45μm microporous filter membrane to obtain the test solution. S2, chromatographic conditions Chromatographic column: Hedera ODS-2-C18 (250 mm x 4.6 mm, 5 μm) chromatographic column, mobile phase: acetonitrile and 0.1% formic acid in water, gradient elution, 210 nm, column temperature 30°C, flow rate 0.8 mL / min, sample injection volume: 20 μL, elution procedure is shown in Table 10 below.

[0054] [Table 10]

[0055] The final chromatogram is shown in Figure 7. In this embodiment, the chromatogram obtained according to the elution procedure has good peak conditions and good separation, which meets the requirements for fingerprint spectrum construction. Therefore, this method is finally selected as the method for constructing fingerprint spectrum of Xinshuning Capsule, and the subsequent peak identification, similarity analysis and methodological review are carried out.

[0056] The chromatograms of the Xinshuning Capsule test solution obtained in S3 and S2 are derived and introduced into the Chinese medicine chromatogram fingerprint spectrum similarity evaluation system (2004 A version), and data input, multi-point correction and data matching are respectively performed on the chromatogram of the test solution to obtain the fingerprint spectrum (Figure 27) and perform similarity analysis (Table 11).

[0057] [Table 11]

[0058] S4, to determine the chemical composition in the fingerprint spectrum, mass spectrometry was performed on the above test solution. The equipment used was Thermo Scientific Ultimate 3000 RSLC ultra-high performance liquid chromatograph - Thermo Scientific Q Exactive Focus benchtop high resolution mass spectrometer. The mass spectrometry conditions were as follows: electrospray ionization (ESI), spray voltage 3800 V, sheath gas flow rate 45 arb, auxiliary gas flow rate 10 arb, capillary temperature 350 °C, mass charge ratio scan range m / z is 100-1200, scan mode is full scan mode, and all ion flow diagrams and mass spectrometry result diagrams of 19 chemical compositions were obtained (Figures 8-26). The detection data was introduced into the Xcalibur software, entered into the Qual Browser interface, and data analysis was performed according to the chemical composition peak situation. Peak 1 is trifolylrhizin at 14.73 min, peak 2 is naringin at 17.31 min, peak 3 is kaempferide at 22.85 min, peak 4 is valine at 24.98 min, peak 5 is palmatine at 26.89 min, peak 6 is epiberberine at 28.03 min, peak 7 is glycyrrhizin / isoglycyrrhizin at 32.76 min, peak 8 is limonin at 35.62 min, peak 9 is liquiritin at 37.68 min, peak 10 is picrotoxin at 44.75 min, and peak 11 is methylopiogonanone A. 50.83 min, peak 12 is rotacin 54.08 min, peak 13 is sophoridine / sophocarpidine 63.19 min, peak 14 is sophoranol / oxidized sophocarpidine 69.17 min, peak 15 is kurarinone 87.14 min, peak 16 is dehydropatiminic acid 96.23 min, peak 17 is polyporenic acid C 108.77 min, peak 18 is quercetin 113.61 min, and peak 19 is sophocarpine 118.39 min.

[0059] S5, Methodological considerations

[0060] 1. Stability Experiment The test samples obtained in S1 are taken, and the samples are injected and analyzed at 0h, 2h, 4h, 8h, 12h and 24h according to the S2 chromatogram conditions, respectively. Trifolyrhizin, naringin, kempferide, valine, palmatine, epiberberine, glycyrrhizin, isoglycyrrhizin, limonin, liquiritin, picrotoxin, methylopiogonanone, rotacin, sophoridine, sophocarpidine, sophoranol, oxidized sophocarpidine, kurarinone, dehydropatiminic acid, poric acid, quercetin, sophocarpine are used as reference peaks, and the peak area and retention time of the common peaks of the HPLC fingerprint spectrum of the samples are analyzed, and the chromatogram is recorded by calculating the RSD value, and the results are shown in Table 12. As can be seen from Table 12, the ratio of the relative retention time and its relative peak area of ​​each major chromatographic peak did not change significantly, and the RSD was 0.32%-3.07% and 0.45%-3.37%, respectively, with RSD<5.0%, which means that the composition of the test solution was stable within 24 h.

[0061] 2.Accuracy experiment The test sample obtained in S1 is taken and measured six times in succession according to the chromatographic conditions of S2. Trifolyrhizin, Naringin, Kaempferide, Valine, Palmatine, Epiberberine, Glycyrrhizin, Isoglycyrrhizin, Limonin, Liquiritin, Picrotoxin, Methylopiogonanone, Rotasin, Sophoridine, Sophocarpidine, Sophoranol, Oxidized Sophocarpidine, Curarinone, Dehydropatiminic Acid, Poric Acid, Quercetin, Sophocarpine are used as reference peaks to analyze the peak area and retention time of the common peaks in the HPLC fingerprint spectrum of the sample and calculate the RSD value, and the chromatogram is recorded, and the results are shown in Table 12. As can be seen, the ratio of each major chromatographic peak relative retention time and its relative peak area does not change significantly, and the RSD is 0.55%-3.40% and 0.27%-3.22%, respectively, with RSD<5.0%, which means that the precision of the instrument is good.

[0062] 3. Reproducibility Experiment Six test samples of Xinshuoning Capsules were taken, and samples were injected and analyzed according to the above chromatogram conditions. Trifolyrhizin, naringin, kaempferide, valine, palmatine, epiberberine, glycyrrhizin, isoglycyrrhizin, limonin, liquiritin, picrotoxin, methyropiogonanone, rotacin, sophoridine, sophocarpidine, sophoranol, oxidized sophocarpidine, kurarinone, dehydropatiminic acid, porinic acid, quercetin, and sophocarpine were used as reference peaks, and the peak area and retention time of the common peaks in the HPLC fingerprint spectrum of the samples were analyzed to calculate the RSD value, and the chromatograms were recorded, and the results are shown in Table 12. The ratios of the relative retention times of each major chromatogram peak and their relative peak areas did not change significantly, with RSDs ranging from 0.34% to 3.78% and 0.41% to 3.91%, respectively, and RSDs < 5.0%, which means that the reproducibility of this experimental method is good.

[0063] [Table 12]

[0064] As can be seen from the above experimental results, the Heart Speed ​​Neck Capsule fingerprint spectrum construction method provided by the present invention has the characteristics of good stability, high accuracy and good reproducibility, and can comprehensively and objectively evaluate the quality of Heart Speed ​​Neck Capsule and provide quality assurance of clinical efficacy.

[0065] The above embodiments are merely illustrative examples of the present invention, and do not limit the present invention, and the protection scope of the present invention is limited by the claims. Those skilled in the art can make various modifications and equivalent replacements to the present invention within the substantial protection scope of the present invention, and such modifications and equivalent replacements shall also be considered to be included in the protection scope of the present invention.

Claims

1. S1: taking different batches of cardiac stimulating capsule contents, adding methanol aqueous solution, and ultrasonically extracting to obtain test solutions; S2: injecting the test solution into a high performance liquid chromatograph, performing gradient elution, performing chromatographic analysis, and recording a chromatogram from 0 to 140 min; S3, the chromatogram obtained in S2 is introduced into the herbal medicine chromatogram fingerprint spectrum similarity evaluation system, and the chromatogram peaks existing in the chromatograms of different batches of Xinshuning capsules are selected as common peaks, and the chromatograms of the test solution are respectively subjected to data introduction, multi-point correction and data matching to obtain a fingerprint spectrum; S4: performing mass analysis on the test solution and determining the chemical composition of the chromatogram peaks in the fingerprint spectrum based on the mass analysis results.

2. The method for constructing a fingerprint spectrum of Heart Rapid Necessity Capsule as described in claim 1, characterized in that in S1, the volume concentration of methanol in the methanol aqueous solution is 80%.

3. The fingerprint spectrum construction method for heart speeding capsules as claimed in claim 1, characterized in that in S1, the ultrasonic extraction time is 20-40 min.

4. The fingerprint spectrum construction method for Xin Shuning Capsules according to claim 1, characterized in that in S2, the detection wavelength used for the chromatographic analysis is 210 nm.

5. The fingerprint spectrum construction method for Xinshuning Capsules as described in claim 1, characterized in that in S2, the chromatographic column used in the high performance liquid chromatograph is a Hedera ODS-2-C18 (250 mm x 4.6 mm, 10 μm) chromatographic column.

6. The method for constructing a fingerprint spectrum of Xinxing Capsules according to claim 1, characterized in that in S2, the mobile phase used for gradient elution is acetonitrile and 0.1% formic acid aqueous solution.

7. In S2, the gradient elution procedure is shown in the table below: The method for constructing a fingerprint spectrum of a heart speeding capsule as claimed in claim 6.

8. The method for constructing a fingerprint spectrum of Xinxing Capsule as described in claim 1, characterized in that in S2, the flow rate used for gradient elution is 0.8 mL / min.

9. In S4, the chemical compositions of the chromatogram peaks in the fingerprint spectrum are determined as follows: Peak 1 is trifolirhizin, Peak 2 is naringin, Peak 3 is kaempferide, Peak 4 is valine, Peak 5 is palmatine, Peak 6 is epiberberine, Peak 7 is glycyrrhizin / isoglycyrrhizin, Peak 8 is limonin, Peak 9 is liquiritin, Peak 10 is picrotoxin, Peak 11 is methylopiogonanone A, Peak 12 is rotasine, Peak 13 is sophoridine / sophocarpidine, Peak 14 is sophoranol / sophocarpidine oxide, Peak 15 is kurarinone, Peak 16 is dehydropatiminic acid, Peak 17 is polyporenic acid C, Peak 18 is quercetin, and Peak 19 is sophocarpine.

10. 10. The fingerprint spectrum of the heart speed capsule constructed by the method according to any one of claims 1 to 9.

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