Detection method of erigeron breviscapus formula granules

Abstract

The invention belongs to the technical field of medicine detection, and particularly relates to a method for detecting erigeron breviscapus formula granules, the erigeron breviscapus formula granules are prepared by decocting, concentrating, drying and granulating erigeron breviscapus decoction pieces, and the detection method comprises the items of character determination, identification, inspection, extract determination, characteristic spectrum determination and content determination. According to the identification method, thin-layer chromatography is adopted for identification; the characteristic chromatogram is characterized in that 11 characteristic peaks are marked by adopting a high performance liquid chromatography and taking a scutellarin reference substance peak as an S peak; and the content determination is to detect the content of scutellarin. According to the detection method established by the invention, the detection requirements on erigeron breviscapus medicinal materials, decoction pieces, standard decoction and formula granules are improved, establishment of a perfect quality control system for each product of erigeron breviscapus is facilitated, and the quality and clinical curative effect of the erigeron breviscapus formula granules are guaranteed.

Description

Technical Field

[0001] The invention belongs to the technical field of drug detection, and particularly relates to a detection method for breviscapus breviscapus formula granules. Background Art

[0002] Chinese medicine formula granules are made from single Chinese medicine pieces through one or more processes to extract and concentrate, and are used in Chinese medicine clinical prescriptions. They have the advantages of being stable and controllable, safe and efficient, no need for decoction, easy to prepare, convenient to carry, and ready to take.

[0003] Erigeron breviscapus (Vant.) Hand.-Mazz. is a dried herb of Erigeron breviscapus (Vant.) Hand.-Mazz., a plant of the Asteraceae family. It is mainly produced in Guizhou, Guangxi, Yunnan, Sichuan, Hunan, etc. It is pungent, slightly bitter, warm, and enters the heart and liver meridians. It has the effects of promoting blood circulation, dredging collaterals, relieving pain, and dispelling wind and cold. It is used for hemiplegia caused by stroke, chest pain, rheumatic pain, headache, toothache, etc.

[0004] At present, the only content determination and liquid chromatography identification of baicalin in the drug standard of Erigeron breviscapus is the content determination of baicalin. With the increasing clinical application of formula granules, more comprehensive, convenient and controllable detection methods are urgently needed to control the quality of Erigeron breviscapus formula granules. Summary of the invention

[0005] Aiming at the imperfect detection methods in the prior art of Erigeron Breviscapus formula granules, the present invention provides a quality detection method for Erigeron Breviscapus formula granules, which is specifically:

[0006] The testing items of the Radix Erigeron Chinensis formula granules include properties, identification, inspection, extracts, characteristic spectra, and content determination.

[0007] In some embodiments, the granules of the Radix Erigeron tetracycline are light brown granules with a slight odor and a slightly bitter and astringent taste.

[0008] In some embodiments, the granules of the formula of Erigeron breviscapus are brown granules; they have a slight odor and a slightly bitter and astringent taste.

[0009] The identification method is as follows: 0.5 g of the formula granules of Erigeron breviscapus are taken, ground, 20 ml of methanol is added, ultrasonic treatment is performed for 30 minutes, filtered, the solvent is recovered from the filtrate until dry, 20 ml of water is added to the residue, dissolved at a low temperature, extracted with ether 3 times, 20 ml each time, the ether layer is discarded, the pH value of the water layer is adjusted to 2-3 with dilute hydrochloric acid, extracted with ethyl acetate 3 times, 20 ml each time, the ethyl acetate is combined, evaporated to dryness, 1 ml of methanol is added to the residue to dissolve it, and the solution is used as the test solution; 2.0 g of Erigeron breviscapus reference medicinal material is taken separately, 60 ml of water is added, decocted for 30 minutes, filtered, the filtrate is evaporated to dryness, 20 ml of methanol is added to the residue, and the reference drug solution is prepared in the same way. According to the thin layer chromatography method (Chinese Pharmacopoeia 2020 General Rules 0502), 5-10 μl of each of the above solutions were taken and spotted in strips on the same polyamide film, developed with glacial acetic acid as the developing agent, taken out, dried, sprayed with 5% ferric chloride ethanol solution, and inspected under sunlight. In the chromatogram of the test sample, the main spot of the same color should appear at the corresponding position of the chromatogram of the control medicinal material.

[0010] The inspection method is as follows: it should comply with the relevant provisions of the General Rules for Granules in the Chinese Pharmacopoeia;

[0011] The extract determination method is as follows: take the formula granules of Erigeron breviscapus, and determine them according to the hot soaking method under the alcohol-soluble extract determination method (General Rules 2201 of the 2020 edition of the Chinese Pharmacopoeia), using ethanol as the solvent.

[0012] The characteristic spectrum detection method is: according to the high performance liquid chromatography method of the Chinese Pharmacopoeia, specifically including the following operations:

[0013] Chromatographic conditions and system suitability experiment: octadecylsilane bonded silica gel is used as filler, acetonitrile is used as mobile phase A, 0.1%-0.5% formic acid solution is used as mobile phase B for gradient elution, the flow rate is 0.2ml-1ml per minute; the column temperature is 35-45°C; the detection wavelength is 300-350nm, and the theoretical plate number calculated based on the scutellarin peak should be not less than 2000-5000; the gradient elution conditions are:

[0014] 0-5min, mobile phase A 2%, mobile phase B 98%;

[0015] 5-8min, mobile phase A 2→5%, mobile phase B 98→95%;

[0016] 8-24 min, mobile phase A 5→15%, mobile phase B 95→85%;

[0017] 24-55 min, mobile phase A 15→18%, mobile phase B 85→82%;

[0018] Preparation of reference solution: Weigh 0.5-2g of Erigeron breviscapus reference medicinal material, add 20-50ml of water, heat and reflux for 30-60 minutes, filter, evaporate the filtrate to dryness, add 25%-50% methanol 20-50ml of the residue, ultrasonically treat for 10-60 minutes, cool, filter, and take the filtrate as the reference solution of the reference medicinal material; take an appropriate amount of scutellaria baicalin reference substance, add methanol to make a solution containing 0.2mg per 1ml, as the reference solution of the reference substance;

[0019] Preparation of the test solution Take an appropriate amount of Erigeron breviscapus granules, grind them into powder, weigh 0.1-1g, add 25%-50% methanol 20-50ml, ultrasonically treat for 10-60min, cool, filter, and take the filtrate to obtain the solution;

[0020] Determination method: Accurately pipette 2 μl of reference solution and test solution respectively, inject into liquid chromatograph, measure according to HPLC working conditions, and record the chromatogram from 0 to 55 min.

[0021] There should be 11 characteristic peaks in the chromatogram of the test sample, and the retention time should correspond to the 11 characteristic peaks in the chromatogram of the reference medicinal material. Among them, Peak 5 should correspond to the retention time of the scutellarin reference peak; the scutellarin reference peak is taken as the S peak, and the relative retention time of each characteristic peak and the S peak is calculated, and the relative retention time should be within the range of ±10% of the specified value; the relative retention time of Peak 1 is 0.43, the relative retention time of Peak 2 is 0.59, the relative retention time of Peak 3 is 0.63, and the relative retention time of Peak 4 is The relative retention time of peak 10 was 1.26 and 1.33, respectively; peak 1 was neochlorogenic acid, peak 2 was chlorogenic acid, peak 3 was cryptochlorogenic acid, peak 5 (S) was scutellarin, peak 6 was isochlorogenic acid B, peak 7 was 3,5-O-dicaffeoylquinic acid and peak 11 was 4,5-di-O-caffeoylquinic acid.

[0022] Preferably, the mobile phase B in the characteristic spectrum detection is 0.1% formic acid solution;

[0023] Preferably, the flow rate in the characteristic spectrum detection is 0.2 ml per minute;

[0024] Preferably, the column temperature in the characteristic spectrum detection is 40°C;

[0025] Preferably, the detection wavelength in the characteristic spectrum detection is 326 nm;

[0026] Preferably, the number of theoretical plates in the characteristic spectrum detection should be no less than 3000 calculated based on the scutellarin peak;

[0027] Preferably, the chromatographic column in the characteristic spectrum detection is Waters ACQUITY UPLC HSS T3, C18, 2.1×100 mm, 1.8 μm;

[0028] Preferably, the reference solution in the characteristic spectrum detection is prepared by weighing 0.5 g of Erigeron breviscapus reference medicinal material, adding 50 ml of water, heating and reflux for 30 minutes, filtering, evaporating the filtrate to dryness, adding 25 ml of 50% methanol to the residue, ultrasonically treating for 30 minutes, cooling, filtering, and taking the subsequent filtrate as the reference medicinal material reference solution; taking an appropriate amount of scutellaria baicalin reference substance, accurately weighing, adding methanol to prepare a solution containing 0.2 mg per 1 ml, as the reference substance reference solution;

[0029] Preferably, the preparation of the test solution in the characteristic spectrum detection is to take an appropriate amount of the granules of Breviscapus asarum, grind them into powder, weigh 0.2 g, add 25 ml of 50% methanol, ultrasonically treat for 30 minutes, cool, filter, and take the filtrate to obtain;

[0030] In some embodiments, the test solution is prepared by taking an appropriate amount of the standard decoction of Erigeron breviscapus, weighing 0.2 g, adding 25 ml of 50% methanol, ultrasonically treating for 30 min, cooling, filtering, and taking the filtrate to obtain;

[0031] In some embodiments, the test solution is prepared by taking an appropriate amount of Erigeron breviscapus medicinal material, crushing it, weighing 1.0 g, adding 50 ml of 50% methanol, ultrasonically treating it for 30 min, cooling it, filtering it, and taking the filtrate to obtain the solution;

[0032] In some embodiments, the test solution is prepared by taking an appropriate amount of Erigeron breviscapus slices, crushing, weighing 0.5 g, adding 50 ml of 50% methanol, ultrasonically treating for 30 min, cooling, filtering, and taking the filtrate to obtain;

[0033] The content determination method is: determined according to high performance liquid chromatography (General Rules 0512 of the 2020 Edition of the Chinese Pharmacopoeia).

[0034] Chromatographic conditions and system suitability test: Octadecylsilane bonded silica gel was used as filler; methanol-0.1% phosphoric acid solution-tetrahydrofuran (13:74:13) was used as mobile phase; the detection wavelength was 335nm. The theoretical plate number calculated based on the scutellarin peak should be no less than 5000.

[0035] Preparation of reference solution: Take an appropriate amount of scutellaria baicalin reference substance, weigh it accurately, and add methanol to make a solution containing 0.2 mg per 1 ml.

[0036] Preparation of test solution: Take an appropriate amount of the product, grind it into powder, take about 0.2 g, weigh it accurately, put it in a stoppered conical flask, accurately add 25 ml of 80% methanol, weigh it, treat it with ultrasound (power 500 W, frequency 53 kHz) for 30 minutes, let it cool, weigh it again, make up the lost weight with 80% methanol, shake it well, filter it, and take the filtrate to obtain it.

[0037] Determination method: Accurately pipette 5μl of reference solution and test solution respectively, inject into liquid chromatograph, and determine.

[0038] Among them, the formula granules of Erigeron breviscapus are prepared by taking 4000g of Erigeron breviscapus decoction pieces, decocting with water, filtering, concentrating the filtrate, drying, crushing, adding appropriate amount of auxiliary materials, mixing, granulating, and making into 1000g.

[0039] In some embodiments, the formula granules of Erigeron breviscapus are prepared by taking 4000 g of Erigeron breviscapus slices, decocting with water, filtering, concentrating the filtrate, vacuum drying, crushing, adding appropriate amount of maltodextrin, mixing, granulating, and making 1000 g.

[0040] In some embodiments, the formula granules of Erigeron breviscapus are prepared by taking 4000 g of Erigeron breviscapus slices, decocting with water, filtering, concentrating the filtrate, vacuum drying, crushing, adding appropriate amount of starch, mixing, granulating, and making 1000 g.

[0041] Some embodiments are to prepare a standard decoction of Erigeron breviscapus, specifically, 4000 g of Erigeron breviscapus slices are taken, decocted with water, filtered, the filtrate is concentrated under reduced pressure to form a clear paste, which is packaged in vials and freeze-dried (-40° C.) to a dry state.

[0042] Beneficial effects:

[0043] 1. The present invention is a detection method developed for the formula granules of Erigeron breviscapus. The detection method has comprehensive detection items, simple operation method, high precision, strong specificity and good reproducibility.

[0044] 2. In addition, the present invention adds the detection of characteristic spectrum on the basis of conventional detection items, which can more comprehensively reflect the types and quantities of chemical components of medicinal materials contained in the product. In particular, the characteristic spectrum adopts UPLC detection method, which has the characteristics of short detection time, high precision, good reproducibility, etc. At the same time, the characteristic spectrum also clearly states that there are 11 characteristic peaks in the characteristic spectrum, among which peak 1 is neochlorogenic acid, peak 2 is chlorogenic acid, peak 3 is cryptochlorogenic acid, peak 5 (S) is scutellariae glycoside, peak 6 is isochlorogenic acid B, peak 7 is 3,5-O-dicaffeoylquinic acid, and peak 11 is 4,5-di-O-caffeoylquinic acid, which is more targeted for controlling product quality.

[0045] 3. Through experiments, the detection method of the present invention is also applicable to Radix Erigeron, decoction pieces, standard decoctions, etc., and has a wide range of applications. BRIEF DESCRIPTION OF THE DRAWINGS

[0046] Figure 1 Thin layer chromatography of Dengzhan Xixin formula granules

[0047] Figure 2 This is the reference characteristic spectrum of the standard decoction of Erigeron breviscapus, where peak 1: neochlorogenic acid, peak 2: chlorogenic acid, peak 3: cryptochlorogenic acid, peak 5 (S): scutellariae glycoside, peak 6: isochlorogenic acid B, peak 7: 3,5-O-dicaffeoylquinic acid, peak 11: 4,5-di-O-caffeoylquinic acid

[0048] Figure 3 This is the reference characteristic spectrum of the formula granules of Dengzhan Xixin, among which, peak 1: neochlorogenic acid, peak 2: chlorogenic acid, peak 3: cryptochlorogenic acid, peak 5 (S): scutellarin, peak 6: isochlorogenic acid B, peak 7: 3,5-O-dicaffeoylquinic acid, peak 11: 4,5-di-O-caffeoylquinic acid

[0049] Figure 4 Characteristic spectrum of Dengzhan Xixin formula granules - specific spectrum

[0050] Figure 5 Determination of baicalin content in Dengzhanxixin formula granules-specificity map

[0051] Figure 6 Comparative Example 1 Chromatograms of reference substance, test substance and corresponding spectra

[0052] Figure 7 Example 5 Reference substance, test substance chromatogram and corresponding spectrogram DETAILED DESCRIPTION

[0053] In order to make the technical scheme and technical effect of the present invention clearer, the technical scheme in the embodiment of the present invention is described clearly and completely. The embodiments described below are part of the embodiments of the present invention, rather than all of the embodiments. In combination with the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0054] Example 1 Establishment of TLC Identification Method for Erigeron Breviscapus Formula Granules

[0055] The present invention uses the Erigeron breviscapus control medicinal material as a positive control, uses the auxiliary materials and methanol used in the Erigeron breviscapus formula granules as a negative control, and uses glacial acetic acid as a developing agent to investigate the thin layer identification method.

[0056] 1.1 Solution preparation

[0057] (1) Preparation of test solution

[0058] Take 0.5g of the formula granules of Erigeron breviscapus, grind them into powder, add 20ml of methanol, ultrasonically treat for 30 minutes, filter, recover the solvent from the filtrate until dry, add 20ml of water to the residue, dissolve it at a slight temperature, extract it with ether 3 times, 20ml each time, discard the ether layer, adjust the pH value of the aqueous layer to 2-3 with dilute hydrochloric acid, extract it with ethyl acetate 3 times, 20ml each time, combine the ethyl acetate, evaporate to dryness, add 1ml of methanol to the residue to dissolve it, and use it as the test solution.

[0059] (2) Preparation of control medicinal material solution

[0060] Take 2.0 g of Erigeron breviscapus reference medicinal material, add 60 ml of water, boil for 30 minutes, filter, evaporate the filtrate to dryness, and prepare the reference medicinal material solution with the same method as the residue starting from "add 20 ml of methanol".

[0061] (3) Preparation of blank negative solution

[0062] A. Preparation of blank excipient solution Take 0.5 g of maltodextrin, an excipient used in the formula granules of Erigeron breviscapus, and prepare a negative excipient solution in the same way as the test solution.

[0063] B. Preparation of Blank Solvent Solution Methanol was used as the negative-solvent solution.

[0064] 1.2 Chromatographic conditions

[0065] Carrier: Polyamide film

[0066] Developing agent: glacial acetic acid

[0067] Inspection: Spray 5% ferric chloride ethanol solution and inspect under sunlight.

[0068] 1.3 Methodological considerations

[0069] 1.3.1 Specificity: Prepare solutions according to the above solution preparation method by taking the formula granules of Erigeron breviscapus, negative-solvent, negative-excipient and Erigeron breviscapus control medicinal materials respectively, take 10μl of each of the above solutions respectively, spot on the same polyamide film, develop according to the above thin layer chromatography conditions, take out, dry, spray 5% ferric chloride ethanol solution, and inspect under sunlight. As a result, in the chromatogram of the test sample, the main spot of the same color appears at the corresponding position of the chromatogram of the control medicinal material. Negative-solvent and negative-excipient have no interference. See Figure 1 .

[0070] 1.3.2 Spotting volume: Take 2μl, 3μl, 5μl, 15μl and 20μl of the test solution respectively, spot them on the same polyamide film, develop them according to the above-mentioned thin layer chromatography conditions, take them out, dry them, spray them with 5% ferric chloride ethanol solution, and inspect them under daylight. According to the spot separation and clarity, determine that the spotting volume of the test solution is 5-10μl.

[0071] 1.3.3 Durability:

[0072] 1.3.3.1 Investigation at different temperatures and humidities: 10 μl of each of negative-solvent, negative-excipient, control medicinal material and test solution were taken and spotted on the same polyamide film. According to the above-mentioned thin layer chromatography conditions, the film was developed at low temperature (18°C), normal temperature (25°C), high temperature (40°C), low humidity (18%), normal humidity (49%), and high humidity (72%). The film was taken out, dried, sprayed with 5% ferric chloride ethanol solution, and inspected under sunlight. Results In the chromatogram of the test product, the main spot of the same color appeared at the corresponding position of the chromatogram of the control medicinal material. Negative-solvent and negative-excipient had no interference. The durability at different temperatures and humidities was good.

[0073] 1.3.3.2 Inspection of thin layer plates from different manufacturers: 10 μl of negative-solvent, negative-excipient, control medicinal material and test solution were taken respectively and spotted on polyamide films from different manufacturers (Zhejiang Sijia, Qingdao Bangkai, Shandong Xiya), developed according to the above thin layer chromatography conditions, taken out, dried, sprayed with 5% ferric chloride ethanol solution, and inspected under sunlight. Results The use of polyamide films from different manufacturers had no significant effect on thin layer identification, and the thin layer identification method had good durability.

[0074] 1.3.3.3 Durability test by different personnel: Different inspectors, at different times, used the same method to prepare the test sample, negative sample, and control medicinal material solution, and took 10 μl of each of the negative-solvent, negative-excipient, control medicinal material, and test sample solution, respectively, and spotted them on the same polyamide film, developed them according to the above-mentioned thin layer chromatography conditions, took them out, dried them, sprayed them with 5% ferric chloride ethanol solution, and inspected them under sunlight. Results Different inspectors used the same inspection method, which had no significant effect on the thin layer identification, and the thin layer identification method had good durability.

[0075] The thin-layer chromatography identification method of the present invention has good specificity and durability after methodological investigation, and can be used for thin-layer chromatography identification of the Radix Erigeron breviscapus formula granules.

[0076] Example 2 Establishment of characteristic spectrum detection method for standard decoction of Erigeron breviscapus

[0077] Determined by high performance liquid chromatography (General Rules 0512 of the Chinese Pharmacopoeia 2020 Edition).

[0078] Chromatographic conditions and system suitability test: Octadecylsilane bonded silica gel was used as filler (column length was 100 mm, inner diameter was 2.1 mm, particle size was 1.8 μm); acetonitrile was used as mobile phase A, 0.1% formic acid solution was used as mobile phase B, and gradient elution was performed according to the provisions in the following table; flow rate was 0.2 ml per minute; column temperature was 40°C; detection wavelength was 326 nm. The theoretical plate number calculated based on the scutellarin peak should be no less than 3000.

[0079]

[0080] Reference solution preparation

[0081] (1) Preparation of reference substance solution

[0082] Take an appropriate amount of scutellaria baicalin as a reference substance, add methanol to make a solution containing 0.2 mg per 1 ml, which is used as the reference substance solution.

[0083] (2) Preparation of reference solution of control medicinal materials

[0084] Take about 0.5 g of Erigeron breviscapus reference medicinal material, place it in a round-bottom flask, add 50 ml of water, heat and reflux for 30 minutes, cool, filter, evaporate the filtrate to dryness, add 25 ml of 50% methanol to the residue, transfer it to a stoppered conical flask, ultrasonically treat it for 30 minutes, cool, filter, and take the filtrate as the reference solution of the control medicinal material.

[0085] Preparation of test solution

[0086] Take about 0.2 g of the standard decoction of Erigeron breviscapus, add 25 ml of 50% methanol, ultrasonically treat for 30 minutes, cool, filter, and take the filtrate to obtain the product.

[0087] Determination method

[0088] Accurately pipette 2 μl of the reference solution and the test solution respectively, inject into the liquid chromatograph, measure, record and analyze the chromatogram.

[0089] Identification of characteristic peaks in characteristic spectra and selection of reference peaks (S peaks)

[0090] Take appropriate amount of neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, scutellariae glycoside, isochlorogenic acid B, 3,5-O-dicaffeoylquinic acid, and 4,5-di-O-caffeoylquinic acid reference substances, and add methanol to prepare peak reference substance solutions.

[0091] Accurately pipette 2 μl of each peak reference solution and test solution, inject into liquid chromatograph, measure, and record the chromatogram. The results are shown in the following table:

[0092]

[0093] By consulting the literature and comparing the retention times of the chromatograms, the retention times and absorption spectra of peaks 1, 2, 3, 5, 6, 7, and 11 in the characteristic spectrum were consistent with those of the reference peaks of neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, scutellarin, isochlorogenic acid B, 3,5-O-dicaffeoylquinic acid, and 4,5-di-O-caffeoylquinic acid, respectively. Combined with the literature, it was determined that peak 1 was neochlorogenic acid, peak 2 was chlorogenic acid, peak 3 was cryptochlorogenic acid, peak 5 was scutellarin, peak 6 was isochlorogenic acid B, peak 7 was 3,5-O-dicaffeoylquinic acid, and peak 11 was 4,5-di-O-caffeoylquinic acid.

[0094] Baicalin can be obtained through the legal unit China Food and Drug Inspection Institute. At the same time, this component is also an indicator component for content determination, and its response value is high in the characteristic spectrum chromatogram. Therefore, baicalin is set as the reference peak (S peak) to calculate the relative retention time of the characteristic peak.

[0095] Determination of characteristic peaks in characteristic spectra

[0096] The relative retention time of the characteristic spectrum is determined according to the research results. According to the analysis of the characteristic spectrum of 21 batches of standard decoctions of Erigeron breviscapus, 11 characteristic peaks are presented in the chromatogram of the test sample, and the retention time of the 11 characteristic peaks in the chromatogram of the reference medicinal material corresponds to that of the reference material. Peak 5 should correspond to the retention time of the reference material peak of scutellaria baicalin. The reference material peak of scutellaria baicalin is taken as the S peak, and the relative retention time of each peak and the S peak is calculated. The relative retention time should be within the range of ±10% of the specified value, and the specified value is: 0.43 (peak 1), 0.59 (peak 2), 0.63 (peak 3), 0.68 (peak 4), 1.11 (peak 6), 1.16 (peak 7), 1.21 (peak 8), 1.24 (peak 9), 1.26 (peak 10), 1.33 (peak 11). The relative peak areas are quite different, so the relative peak areas are not specified. The reference characteristic spectrum of the standard decoction of Erigeron breviscapus can be found in Figure 2 .

[0097] Example 3 Establishment of characteristic spectrum detection method of Erigeron breviscapus formula granules

[0098] Determined by high performance liquid chromatography (General Rules 0512 of the Chinese Pharmacopoeia 2020 Edition).

[0099] Chromatographic conditions and system suitability test: Octadecylsilane bonded silica gel was used as filler (column length was 100 mm, inner diameter was 2.1 mm, particle size was 1.8 μm); acetonitrile was used as mobile phase A, 0.1% formic acid solution was used as mobile phase B, and gradient elution was performed according to the provisions in the following table; flow rate was 0.2 ml per minute; column temperature was 40°C; detection wavelength was 326 nm. The theoretical plate number calculated based on the scutellarin peak should be no less than 3000.

[0100]

[0101] Reference solution preparation

[0102] (1) Preparation of reference substance solution

[0103] Take an appropriate amount of scutellaria baicalin as a reference substance, add methanol to make a solution containing 0.2 mg per 1 ml, which is used as the reference substance solution.

[0104] (2) Preparation of reference solution of control medicinal materials

[0105] Take about 0.5 g of Erigeron breviscapus reference medicinal material, place it in a round-bottom flask, add 50 ml of water, heat and reflux for 30 minutes, cool, filter, evaporate the filtrate to dryness, add 25 ml of 50% methanol to the residue, transfer it to a stoppered conical flask, ultrasonically treat it for 30 minutes, cool, filter, and take the filtrate as the reference solution of the control medicinal material.

[0106] Preparation of test solution

[0107] Take the formula granules of Erigeron breviscapus, grind them into powder, take about 0.2g, add 25ml of 50% methanol, ultrasonically treat for 30 minutes, cool, filter, and take the filtrate to obtain the product.

[0108] Determination method

[0109] Accurately pipette 2 μl of the reference solution and the test solution respectively, inject into the liquid chromatograph, measure, and record the chromatogram.

[0110] There are 11 characteristic peaks in the chromatogram of the test sample, and the retention times of the 11 characteristic peaks in the chromatogram of the reference medicinal material correspond to each other. Peak 5 corresponds to the retention time of the reference peak of the baicalin reference substance. The reference peak of the baicalin reference substance is taken as the S peak, and the relative retention time of each peak and the S peak is calculated. The relative retention time should be within the range of ±10% of the specified value, and the specified value is: 0.43 (peak 1), 0.59 (peak 2), 0.63 (peak 3), 0.68 (peak 4), 1.11 (peak 6), 1.16 (peak 7), 1.21 (peak 8), 1.24 (peak 9), 1.26 (peak 10), 1.33 (peak 11). The reference characteristic spectrum of the formula granules of Dengzhan Xixin is shown in Figure 3 .

[0111] Example 4: Methodological verification of characteristic spectrum detection method of Dengzhanxixin formula granules

[0112] The practicability and accuracy of the characteristic spectrum detection method of Example 3 were verified from the aspects of the specificity, linear range, precision, solution stability, durability, etc. of the detection method.

[0113] In the examples, all raw materials are commercially available, and the experimental methods without specific conditions are conventional methods and conditions known in the art, or according to the conditions recommended by the instrument manufacturer. The reagents, reagents and instruments used in the examples are detailed in the following table:

[0114]

[0115]

[0116] 4.1 Specificity test

[0117] Blank solvent for test sample solution: Take 50% methanol solution.

[0118] Blank excipient test solution: Take an appropriate amount of maltodextrin, an excipient used in the formula granules of Erigeron breviscapus, and prepare it according to the method of "Preparation of test solution" in Example 3.

[0119] Preparation of reference solution: Prepare according to the method of Example 3 “Preparation of reference solution”.

[0120] Preparation of test solution: Prepare according to the method of Example 3 "Preparation of test solution".

[0121] Each solution was sampled and tested, and the results showed that the blank solvent and blank excipient had no interference with the test, and the method had good specificity. Figure 4 .

[0122] 4.2 Instrument precision test

[0123] Take the reference material and prepare the reference solution according to the method of "Preparation of Reference Solution" in Example 3, and inject each sample once. Take the Dengzhan Xixin Formula Granules and prepare the test solution according to the method of "Preparation of Test Solution" in Example 3, and repeat the injection 6 times; use Peak 5 as the reference peak to calculate the relative retention time and relative peak area of ​​the remaining peaks. The results show that the relative retention time RSD of each characteristic peak is 0.03% to 0.10%, and the relative peak area RSD is 0.20% to 1.40%, both less than 2.0%, and the relative retention time of each peak is within the specified range, indicating that the instrument has good precision.

[0124] 4.3 Sample stability

[0125] Take the reference material and prepare the reference solution according to the method of "Preparation of Reference Solution" in Example 3, and inject each sample once; take the formula granules of Dengzhan Xixin and prepare the test solution according to the method of "Preparation of Test Solution" in Example 3. Inject and detect at 0h, 3h, 6h, and 12h respectively, and calculate the relative retention time and relative peak area of ​​the remaining peaks with peak No. 5 as the reference peak. The results show that the relative retention time RSD of each characteristic peak is 0.03% to 0.14%, and the relative peak area RSD is 0.41% to 1.63%, which are all less than 2%. The relative retention time of each peak is within the specified range, indicating that the test solution is stable within 12 hours.

[0126] 4.4 Repeatability test

[0127] Take the reference and prepare the reference solution according to the method of "Preparation of Reference Solution" in Example 3, and inject each sample once; take the Dengzhan Xixin Formula Granules and repeat the method of "Preparation of Test Solution" in Example 3 to prepare 6 portions of the test solution; use Peak No. 5 as the reference peak to calculate the relative retention time and relative peak area of ​​the remaining peaks, and the results are shown in the table below. The results show that the relative retention time RSD of each characteristic peak is 0.04% to 0.12%, and the relative peak area RSD is 0.32% to 2.48%, both less than 3.0%, and the relative retention time of each peak is within the specified range, indicating that the method has good repeatability.

[0128] 4.5 Intermediate precision

[0129] Different inspectors, at different times and using different instruments, prepared samples according to the following method and tested them using different instruments.

[0130] Take 50% methanol solution as blank solvent test solution; take an appropriate amount of maltodextrin, prepare blank auxiliary material test solution according to the method of "Preparation of test solution" in Example 3; take reference material and prepare reference material solution according to the method of "Preparation of reference material solution" in Example 3. Take Dengzhan Xixin formula granules, repeat the method of "Preparation of test solution" in Example 3 to prepare 6 portions of test solution, and inject each sample once; calculate the relative retention time and relative peak area of ​​the remaining peaks with peak No. 5 as the reference peak. The results show that different inspectors use different instruments for detection, and the relative retention time RSD of each characteristic peak is 0.76% to 0.95%, the relative peak area RSD is 4.52% to 18.89%, and the relative retention time of each peak is less than 2%, indicating that the intermediate precision of this method is good; but the relative peak area RSD is greater than 2%, indicating that different instruments have a greater impact on the relative peak area of ​​the test sample, so when formulating the standard, the relative peak area is not used as a detection indicator.

[0131] 4.6 Durability

[0132] 4.6.1 Investigation of different column temperatures

[0133] Take the reference material and prepare the reference solution according to the method of "Preparation of reference solution" in Example 3; take the formula granules of Dengzhan Xixin and prepare the test solution according to the method of "Preparation of test solution" in Example 3. Samples were injected and tested under column temperatures of 35°C, 40°C, and 45°C respectively, and the relative retention time and relative peak area of ​​the remaining peaks were calculated with peak No. 5 as the reference peak. The results showed that the RSD of the relative retention time of each characteristic peak was 0.21% to 2.58%, all less than 3%, and the relative retention time of each peak was within the specified range, indicating that the column temperature durability of this method was good.

[0134] 4.6.2 Investigation of different flow rates

[0135] Take the reference and prepare the reference solution according to the method of "Preparation of Reference Solution" in Example 3; take the Radix Erigeron lanceolata granules and prepare the test solution according to the method of "Preparation of Test Solution" in Example 3. Samples were injected and tested at a flow rate of 0.2±0.02ml / min, and the relative retention time and relative peak area of ​​the remaining peaks were calculated with Peak 5 as the reference peak. The results showed that the RSD of the relative retention time of each characteristic peak was 0.34% to 1.63%, all less than 2%, and the relative retention time of each peak was within the specified range, indicating that the flow rate durability of this method was good.

[0136] 4.6.3 Investigation of different brands of chromatographic columns

[0137] Take the reference material and prepare the reference solution according to the method of "Preparation of reference solution" in Example 3; take the Dengzhan Xixin formula granules and prepare the test solution according to the method of "Preparation of test solution" in Example 3. Use Aglient ZORBAX SB-Aq RRHD, Aglient ZORBAX SB, and Waters ACQUITY UPLC HSS T3 chromatographic columns for sample injection and detection, and use peak No. 5 as the reference peak to calculate the relative retention time and relative peak area of ​​the remaining peaks. The results show that the relative retention time RSD of each characteristic peak is 0.43% to 19.35%, and the results show that different brands of chromatographic columns have obvious differences in the relative retention time of the measurement results. Taking into account the various chromatographic peaks, the Waters ACQUITY UPLC HSS T3 chromatographic column is fixed to measure the Dengzhan Xixin formula granules.

[0138] 4.6.4 Inspection of chromatographic columns from different batches of the same brand

[0139] The reference solution was prepared according to the method of "Preparation of Reference Solution" in Example 3 for the reference material; the test solution was prepared according to the method of "Preparation of Test Solution" in Example 3 for the Dengzhan Xixin Formula Granules. The chromatographic columns of different batches, Waters ACQUITY UPLC HSS T3 (No.: 0270313563) and Waters ACQUITY UPLC HSS T3 (No.: 0243302951), were used for sample injection and detection, and the relative retention time and relative peak area of ​​the remaining peaks were calculated with peak No. 5 as the reference peak. Results The relative retention time of each characteristic peak of different batches of chromatographic columns had a relative deviation of 0.01% to 0.03%, indicating that the use of chromatographic columns produced from different batches of the same brand Waters ACQUITY UPLC HSS T3 had no significant effect on the detection results of the relative retention time of the characteristic peaks.

[0140] Example 5 Establishment of the content determination method of Dengzhan Xixin formula granules

[0141] Determined by high performance liquid chromatography (General Rules 0512 of the Chinese Pharmacopoeia 2020 Edition).

[0142] Chromatographic conditions and system suitability test: Octadecylsilane bonded silica gel was used as filler; methanol-0.1% phosphoric acid solution-tetrahydrofuran (13:74:13) was used as mobile phase; the detection wavelength was 335nm. The theoretical plate number calculated based on the scutellarin peak should be no less than 5000.

[0143] Preparation of reference solution: Take an appropriate amount of scutellaria baicalin reference substance, weigh it accurately, and add methanol to make a solution containing 0.2 mg per 1 ml.

[0144] Preparation of test solution: Take an appropriate amount of Radix Glehniae granules, grind them into powder, take about 0.2 g, weigh it accurately, put it in a stoppered conical flask, accurately add 25 ml of 80% methanol, weigh it, treat it with ultrasound (power 500 W, frequency 53 kHz) for 30 minutes, let it cool, weigh it again, make up the lost weight with 80% methanol, shake it well, filter it, and take the filtrate to obtain the test solution.

[0145] Determination method: Accurately pipette 5 μl of reference solution and test solution respectively, inject into liquid chromatograph, and record the chromatogram.

[0146] Example 6 Methodological Verification of the Content Determination Method of Dengzhan Xixin Formula Granules

[0147] 6.1 Exclusivity

[0148] Take the formula granules of Erigeron breviscapus, and prepare the test solution according to the method of "Preparation of test solution" in Example 5; take about 0.2g of maltodextrin, and prepare the blank auxiliary material solution in the same way as "Preparation of test solution"; take 80% methanol as the blank solvent solution. The above solution was measured according to the chromatographic conditions of Example 5. The results show that the chromatographic peak of baicalin in the chromatogram of the test solution has good separation from the adjacent chromatographic peaks, and the blank and negative have no interference with the detection, indicating that the specificity of this method meets the requirements. See Figure 5 .

[0149] 6.2 Linearity and Range

[0150] Accurately weigh 13.180 mg of scutellarin reference substance, place it in a 25 ml volumetric flask, and add methanol to prepare a reference substance stock solution containing 848.39 μg of scutellarin per 1 ml.

[0151] A certain amount of the above stock solution was accurately drawn to prepare reference substance solutions with concentrations of 26.51 μg / ml, 53.02 μg / ml, 106.05 μg / ml, 212.10 μg / ml and 424.20 μg / ml, respectively. The above 5 reference substance solutions and reference stock solution with different concentrations were taken respectively, and the chromatographic conditions of Example 5 were measured, and the peak area of ​​the chromatogram was recorded. A standard curve was drawn with the peak area as the ordinate (Y) and the reference substance concentration as the abscissa (X). As a result, the linear regression equation of scutellaria baicalin was y=17692x+50005, the correlation coefficient r=0.9998, and SS=184, indicating that the injection concentration of scutellaria baicalin had a good linear relationship with the peak area within the range of 26.51 μg / ml to 848.39 μg / ml.

[0152] 6.3 Stability investigation

[0153] Take an appropriate amount of Dengzhan Xixin formula granules, grind them into powder, take about 0.2g, weigh them accurately, prepare the test solution according to the test solution preparation method of Example 5, and inject samples at 0, 3, 6, 9, and 12 hours according to the chromatographic conditions of Example 5, and record the baicalin peak area and RSD value in the sample. As a result, the RSD value of the baicalin peak area of ​​the same test solution within 12 hours was 0.27%, which was less than 2%, indicating that the test solution had good stability within 12 hours.

[0154] 6.4 Intermediate precision study

[0155] Different personnel used different instruments to take an appropriate amount of Radix Erigeron breviscapus granules, grind them into powder, take about 0.2 g, accurately weigh them, and prepare 6 test solutions according to the test solution preparation method of Example 5. According to the chromatographic conditions of Example 5, the samples were injected respectively, and the baicalin content and RSD value in the samples were calculated. The same batch of samples were measured 6 times by different experimenters using different detection instruments at different times. The average baicalin content was 21.04 mg / ml and 20.99 mg / ml, respectively, and the RSD value was 0.61%, indicating that the method had good repeatability and intermediate precision.

[0156] 6.5 Accuracy test

[0157] Accurately weigh 22.312 mg of baicalin reference substance, place it in a 250 ml volumetric flask, add 80% methanol to dissolve it, and make a solution containing 0.08166 mg of baicalin per 1 ml.

[0158] Take an appropriate amount of Dengzhan Xixin formula granules (content 21.07mg / g), grind it, take about 0.1g, accurately weigh it, put it in a stoppered conical bottle, accurately add 25ml of the above-mentioned reference substance stock solution, weigh it, ultrasonically treat it for 30 minutes, take it out, let it cool, weigh it, make up the lost weight with 80% methanol, shake it well, filter it, and take the filtrate. Prepare 6 test sample solutions in the same way, detect it according to the chromatographic conditions of Example 5, calculate the content of scutellariae in the test sample, calculate the sample recovery rate, and the results show that the recovery rate of scutellariae is in the range of 100.08% to 100.94%, the average recovery rate is 100.49%, and the RSD value is 0.37%, indicating that the accuracy of the analytical method is good.

[0159] 6.6 Durability test

[0160] Take an appropriate amount of Erigeron breviscapus granules, grind them into powder, take about 0.2 g, weigh them accurately, prepare the test solution according to the test solution preparation method of Example 5, and use different chromatographic columns (Waters XBridge C18 chromatographic column (4.6×250mm, 5μm); GL InertSμstain C18 chromatographic column (4.6×250mm, 5μm); Aglient Eclipse XDB-C18 chromatographic column (4.6×250mm, 5μm)) for determination. The results show that the separation effect of each chromatographic column is good, and the content of scutellaria baicalin is 21.34mg / ml, 21.51mg / ml, and 21.07mg / ml, and the RSD value is 1.1%, indicating that the analytical method has good analytical durability under different chromatographic columns;

[0161] The results of the determination using different column temperatures (25℃, 30℃ and 35℃) showed good separation effect, and the RSD value of scutellarin content was 0.46%, indicating that the analytical method has good analytical durability within the column temperature range of ±5℃;

[0162] The formula granules of Erigeron breviscapus were measured using different flow rates (0.9ml / min, 1.0ml / min and 1.1ml / min). The results showed that the chromatographic peaks at different flow rates had good separation effects, and the RSD value of baicalin content was 0.20%, indicating that the analytical method had good analytical durability within the flow rate range of ±0.1ml / min.

[0163] Comparative Example 1

[0164] The difference from Example 5 is that the mobile phase in the chromatographic conditions is different. The chromatographic conditions of Comparative Example 1 are the chromatographic conditions in the method for determining the content of Erigeron breviscapus medicinal material in the Chinese Pharmacopoeia.

[0165] Chromatographic column: Aglient Eclipse XDB-C18

[0166] Mobile phase: methanol-0.1% phosphoric acid solution (40:60)

[0167] The detection wavelength is 335nm;

[0168] Accurately pipette 10 μl of the test sample and reference sample solution respectively, inject into the liquid chromatograph, and record the chromatogram. Figure 6

[0169] Example 5 The mobile phase was methanol-0.1% phosphoric acid solution-tetrahydrofuran (13:74:13);

[0170] Accurately pipette 10 μl of the test sample and reference sample solution respectively, inject into the liquid chromatograph, and record the chromatogram. Figure 7 .

[0171] As a result, Figure 6 It can be seen that under this condition, the separation effect of scutellarin in the test sample chromatogram is not good, and the separation degree does not meet the requirements; however, after changing the mobile phase in Example 5, scutellarin in the test sample chromatogram can achieve baseline separation, and the separation effect is good.

[0172] The above is only a preferred embodiment of the present invention and does not limit the present invention in any form. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention shall be included in the protection scope of the present invention.

Claims

1. A method for detecting granules of Erigeron breviscapus, wherein the granules are prepared by boiling, concentrating, drying, crushing and granulating Erigeron breviscapus pieces; the detection method includes properties, identification, inspection, extract, characteristic spectrum and content determination, characterized in that: The characteristic spectrum detection method is to use high performance liquid chromatography for detection. The working conditions of the high performance liquid chromatography are: the chromatographic column uses octadecylsilane bonded silica gel as a filler, acetonitrile as a mobile phase A, and 0.1%-0.5% formic acid solution as a mobile phase B for gradient elution, and the flow rate is 0.2ml-1ml per minute; the column temperature is 35-45°C; the detection wavelength is 300-350nm, and the theoretical plate number calculated based on the scutellarin peak should be not less than 2000-5000; the gradient elution conditions are: 0-5min, mobile phase A 2%, mobile phase B 98%; 5-8min, mobile phase A 2→5%, mobile phase B 98→95%; 8-24min, mobile phase A 5→15%, mobile phase B 95→85%; 24-55 min, mobile phase A 15→18%, mobile phase B 85→82%; The detection method comprises the following operations: Preparation of reference solution: Weigh 0.5-2g of Erigeron breviscapus reference medicinal material, add 20-50ml of water, heat and reflux for 30-60 minutes, filter, evaporate the filtrate to dryness, add 25%-50% methanol 20-50ml of the residue, ultrasonically treat for 10-60 minutes, cool, filter, and take the filtrate as the reference medicinal material reference solution; take an appropriate amount of scutellaria baicalin reference substance, add methanol to make a solution containing 0.2mg per 1ml, as the reference solution of the reference substance; Preparation of the test solution Take an appropriate amount of Erigeron breviscapus granules, grind them into powder, weigh 0.1-1g, add 25%-50% methanol 20-50ml, ultrasonically treat for 10-60min, cool, filter, and take the filtrate to obtain the solution; Determination method: Accurately pipette 1-4 μl of the reference solution and the test solution respectively, inject into the liquid chromatograph, measure according to the working conditions of the high performance liquid chromatography, and record the chromatogram from 0 to 55 min.

2. The detection method according to claim 1, characterized in that: The working conditions of the high performance liquid chromatography method are as follows: the high performance liquid chromatograph is an ultra high performance liquid chromatograph, the chromatographic column is filled with octadecylsilane bonded silica gel, acetonitrile is used as mobile phase A, 0.1% formic acid solution is used as mobile phase B for gradient elution, and the flow rate is 0.2 ml per minute; the column temperature is 40°C; the detection wavelength is 326nm, and the number of theoretical plates calculated based on the scutellarin peak should be no less than 3000; Preparation of the reference solution: Weigh 0.5 g of Erigeron breviscapus reference medicinal material, add 50 ml of water, heat and reflux for 30 minutes, filter, evaporate the filtrate to dryness, add 25 ml of 50% methanol to the residue, ultrasonically treat for 30 minutes, cool, filter, and take the filtrate as the reference medicinal material reference solution; take an appropriate amount of scutellaria baicalin reference substance, accurately weigh it, add methanol to make a solution containing 0.2 mg per 1 ml, as the reference substance solution; Preparation of the test solution Take an appropriate amount of Erigeron breviscapus granules, grind them into powder, weigh 0.2g, add 25ml of 50% methanol, ultrasonically treat for 30min, cool, filter, and take the filtrate to obtain the solution; Determination method: Accurately pipette 2 μl of reference solution and test solution respectively, inject into liquid chromatograph, measure according to HPLC working conditions, and record the chromatogram from 0 to 55 min.

3. The detection method according to any one of claims 1 or 2, characterized in that The chromatographic column is Waters ACQUITY UPLC HSS T3, C18, 2.1×100 mm, 1.8 μm.

4. The detection method according to any one of claims 1 or 2, characterized in that The test sample chromatogram of the characteristic spectrum should present 11 characteristic peaks, and the retention times should correspond to the 11 characteristic peaks in the chromatogram of the reference medicinal material, among which Peak 5 should correspond to the retention time of the scutellarin reference peak; the scutellarin reference peak is taken as Peak S, and the relative retention time of each characteristic peak and Peak S is calculated, and the relative retention time should be within ±10% of the specified value; the relative retention time of Peak 1 is 0.43, the relative retention time of Peak 2 is 0.59, the relative retention time of Peak 3 is 0.63, and the relative retention time of Peak 4 is 0.

64. The retention time was 0.68, the relative retention time of peak 6 was 1.11, the relative retention time of peak 7 was 1.16, the relative retention time of peak 8 was 1.21, the relative retention time of peak 9 was 1.24, the relative retention time of peak 10 was 1.26, and the relative retention time of peak 11 was 1.33; among them, peak 1 was neochlorogenic acid, peak 2 was chlorogenic acid, peak 3 was cryptochlorogenic acid, peak 5 (S) was baicalin, peak 6 was isochlorogenic acid B, peak 7 was 3,5-O-dicaffeoylquinic acid, and peak 11 was 4,5-di-O-caffeoylquinic acid.

5. The detection method according to any one of claims 1 or 2, characterized in that: The identification method is as follows: take 0.2-1g of the formula granules of Erigeron breviscapus, grind them into powder, add 20-50ml of methanol, ultrasonically treat for 30-60 minutes, filter, recover the solvent from the filtrate until dry, add 20ml of water to the residue, dissolve it at a low temperature, extract it with ether for 3 times, 20ml each time, discard the ether layer, adjust the pH value of the water layer to 2-3 with dilute hydrochloric acid, extract it with ethyl acetate for 3 times, 20ml each time, combine the ethyl acetate, evaporate to dryness, add 1ml of methanol to the residue to dissolve it, and use it as the test solution; take another 2.0g of Erigeron breviscapus reference medicinal material, add 60ml of water, decoct for 30 minutes, filter, evaporate the filtrate, add 20ml of methanol to the residue, and prepare the reference drug solution in the same way; according to the thin layer chromatography test of the Chinese Pharmacopoeia, take 5-10μl of each of the above solutions, spot them in strips, and spot them on the same polyamide film respectively, use glacial acetic acid as the developing agent, develop, take out, dry, spray with 5% ferric chloride ethanol solution, and inspect it under sunlight. In the chromatogram of the test sample, a main spot of the same color should appear at the corresponding position in the chromatogram of the control medicinal material.

6. The detection method according to any one of claims 1 or 2, characterized in that: The content determination is to determine the scutellarin content by high performance liquid chromatography, and the detection conditions of the high performance liquid chromatography are: octadecylsilane bonded silica gel is used as a filler; methanol: 0.1%~0.5% phosphoric acid solution: tetrahydrofuran = 13:74:13 is used as a mobile phase; the detection wavelength is 335nm, and the theoretical plate number calculated according to the scutellarin peak should be not less than 2000-8000.

7. The detection method according to claim 6, characterized in that: The content determination method comprises the following operations: Preparation of reference solution Take an appropriate amount of scutellariae baicalin reference substance, weigh accurately, and add methanol to make a solution containing 0.2 mg per 1 ml. Preparation of the test solution Take an appropriate amount of Dengzhan Xixin granules, grind them into powder, take about 0.2g, weigh accurately, put it in a stoppered conical flask, accurately add 25ml of 80% methanol, weigh it, treat it with ultrasound for 30 minutes, let it cool, weigh it again, make up the lost weight with 80% methanol, shake it well, filter it, and take the filtrate to obtain; The determination method is to take 5ul of the reference solution and the test solution respectively, inject them into the liquid chromatograph, determine, record the chromatogram, and calculate the scutellarin content by the external standard method.

8. The detection method according to any one of claims 1 to 2, characterized in that: The extract detection method is: take the formula granules of Erigeron breviscapus, and determine them according to the hot leaching method under the alcohol-soluble extract determination method of the Chinese Pharmacopoeia, using ethanol as the solvent.

9. The detection method according to any one of claims 1 to 8, characterized in that: The formula granules of Erigeron breviscapus are prepared by taking Erigeron breviscapus decoction pieces, decocting with water, concentrating, drying, crushing, adding pharmaceutically acceptable excipients, mixing and granulating. Each 1g of the formula granules is equivalent to 4.0g of Erigeron breviscapus decoction pieces.

10. Use of the detection method according to any one of claims 1 to 9, characterized in that: The detection method is used in the quality detection of Erigeron breviscapus medicinal materials, Erigeron breviscapus decoction pieces, Erigeron breviscapus standard decoction, and Erigeron breviscapus formula granules.

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