Construction method of UPLC specific chromatogram of sword bean and preparation thereof

Abstract

The invention provides a construction method of the UPLC specific chromatogram of sword bean and preparation thereof, which comprises the following steps: A) dissolving a test raw material with a solvent, and extracting to obtain a to-be-detected solution; b) determining the liquid to be detected by adopting a high performance liquid chromatography to obtain the UPLC characteristic chromatogram of the sword bean and the preparation thereof; chromatographic conditions of the high performance liquid chromatography are as follows: a chromatographic column is a C18 column; a mobile phase A is an acetonitrile solution, a mobile phase B is a 0.1% phosphoric acid aqueous solution, and gradient elution is carried out. According to the method, a high performance liquid chromatography is adopted, an acetonitrile-0. 1% phosphoric acid solution is selected as a mobile phase for gradient elution, quercetin-3-O-glucose-7-O-rhamnoside, myricetin-3-O-galactoside, lysimachin, rutin and isoquercitrin are taken as reference substances, and the UPLC characteristic spectrum method of the sword beans and the preparation thereof is established. And more scientific technical means are provided for controlling the medicinal quality of the sword beans and the preparation thereof.

Description

Technical Field

[0001] The present invention relates to the technical field of drug and preparation analysis and detection, and in particular to a method for constructing a UPLC characteristic spectrum of sword bean and its preparation. Background Art

[0002] The use of sword beans was documented as early as the Ming Dynasty's "Compendium of Materia Medica." Shi Zhen's "Compendium of Materia Medica" states: "The sword bean is named after its pod shape... When it matures, the seeds, which are as large as a thumb and light red, are collected." It is sweet and warm in nature and enters the stomach and kidney meridians. It has the effects of warming the middle, purging qi, and relieving hiccups, and is used for hiccups and vomiting caused by cold deficiency. Research has shown that sword beans contain 28.75% protein, 37.20% starch, 7.50% soluble sugars, 1.36% lipids, and 6.10% fiber. It also contains canavanine, concanavalin A, γ-guanidoxypropylamine, aminopropylconcanavalin A, and aminobutylconcanavalin A, as well as lectins. Currently, no UPLC characteristic profiles of sword beans have been reported, and existing methods make it difficult to effectively compare and analyze the differences and variations between sword beans and their preparations, making it difficult to comprehensively evaluate and control the quality of sword beans and their preparations. Therefore, establishing a unified method for determining the characteristic spectra of sword bean and its preparations is conducive to the overall evaluation of the scientificity and rationality of the sword bean-related process, and can better control the intrinsic quality of sword bean and its preparations as a whole, and ensure the clinical efficacy of sword bean formula granules. Summary of the Invention

[0003] In view of this, the technical problem to be solved by the present invention is to provide a method for constructing a UPLC characteristic spectrum of sword bean and its preparations, and the method of the present invention is accurate and reliable.

[0004] The method of the present invention can effectively separate the components of sword bean medicinal materials, standard decoctions, and formula granules, and successfully identifies quercetin-3-O-glucoside-7-O-rhamnoside, myricetin-3-O-galactoside, mauritius herbicide, rutin, and isoquercetin. The characteristic spectra of 21 batches of sword bean medicinal materials, 21 batches of standard decoctions, and 3 batches of sword bean formula granules were synthesized into a comparison spectrum using the Traditional Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System (2012 Edition). A comparison spectrum of the characteristic spectra of sword bean medicinal materials, standard decoctions, and sword bean granules was established. It is stipulated that 13 characteristic peaks should be present in the chromatogram of the test sample.

[0005] The present invention provides a method for constructing a UPLC characteristic spectrum of sword bean and its preparation, comprising:

[0006] A) Dissolve the test sample in a solvent and extract to obtain a test solution;

[0007] B) measuring the test solution by high performance liquid chromatography to obtain a UPLC characteristic spectrum of the sword bean and its preparation;

[0008] The HPLC conditions are as follows: the chromatographic column is a C18 column; the mobile phase A is an acetonitrile solution, the mobile phase B is a 0.1% phosphoric acid aqueous solution, and the elution is performed in a gradient manner;

[0009] The gradient elution is specifically as follows:

[0010] 0-12 min, phase A: 10→14%, phase B: 90→86%;

[0011] 12-22 min, phase A: 14% → 16%, phase B: 86% → 84%;

[0012] 22-25 min, phase A: 16% → 18%, phase B: 84% → 82%;

[0013] 25-45 min, phase A: 18% → 30%, phase B: 82% → 70%;

[0014] 45-50 min, phase A: 30%, phase B: 70%.

[0015] The test sample raw materials of the present invention are one or more of the following: Jack bean medicinal materials, Jack bean medicinal standard decoctions or Jack bean medicinal formula granules.

[0016] The present invention provides a method for constructing a UPLC characteristic spectrum of sword bean and its preparation. Firstly, a test sample raw material is dissolved in a solvent and extracted to obtain a test solution.

[0017] Specifically, the test sample raw materials are dissolved in a solvent, extracted, cooled, and filtered to obtain the product.

[0018] The extraction method of the present invention is heating reflux extraction or ultrasonic extraction, preferably ultrasonic extraction. The ultrasonic power is 600W, the frequency is 40kHz, and the ultrasonic time is 20 to 40 minutes, preferably 30 minutes. The extraction solvent is preferably 80% methanol.

[0019] The invention can completely extract the product under the above ultrasonic parameters, and has good chromatogram peak shape and separation degree.

[0020] The ratio of the mass g of the test sample raw material to the volume mL of the solvent is 1:25.

[0021] When the above extraction solvent and specific solvent addition amount are used, the peak shape and separation degree of each chromatographic peak are good, and the peak size is appropriate.

[0022] Specifically, the preparation of the test solution: take the sword bean medicinal material, standard decoction, and formula granules, accurately weigh them, place them in a stoppered conical flask, accurately add 80% methanol, seal it tightly, weigh the weight, ultrasonically treat it for 30 minutes, take it out, cool it, filter it, and take the filtrate to obtain it.

[0023] In some embodiments, the ratio of the mass g of the test sample raw material to the volume mL of the solvent is 0.5 to 1.5:25; specifically, it can be 0.5:25; 1:25 or 1.5:25.

[0024] The above raw materials of the present invention can all be quality controlled by the method of the present invention.

[0025] The present invention further comprises preparing a reference solution: taking quercetin-3-O-glucose-7-O-rhamnoside, myricetin-3-O-galactoside, herbicide-resistant mauritius, rutin, and isoquercetin reference substances, respectively, and dissolving them in 80% methanol to obtain a reference solution;

[0026] The control medicinal material of sword bean was decocted in water for 30 minutes, cooled, filtered, evaporated to dryness, and the residue was dissolved in 80% methanol and filtered to obtain a control medicinal material reference solution;

[0027] The reference substance solution and the reference medicinal material reference substance solution are respectively measured by high performance liquid chromatography to obtain chromatograms of the reference substance and the reference medicinal material reference substance; and the components of sword bean and its preparation are qualitatively analyzed based on the chromatograms of the reference substance.

[0028] The concentrations of the reference substance solution of the present invention are preferably as follows: quercetin-3-O-glucoside 20 μg / mL, myricetin-3-O-galactoside 20 μg / mL, herbicide-resistant galactopyranoside 20 μg / mL, rutin 20 μg / mL, and isoquercetin 20 μg / mL.

[0029] The test solution is measured by high performance liquid chromatography to obtain a UPLC characteristic spectrum of the sword bean and its preparation.

[0030] The chromatographic conditions of the high performance liquid chromatography method are: the chromatographic column is a C18 column; the chromatographic column is C18, with specifications of 150×2.1mm 1.8μm; the present invention does not limit the specific chromatographic column model, and any C18 chromatographic column that meets the above model can be used.

[0031] In some embodiments, the chromatographic column can be the following chromatographic columns: DIKMAC18 2.1*150 mm, 1.8 μm; Shimadzu C18 2.1*150 mm, 1.8 μm; Waters C18 2.1×150 mm, 1.8 μm.

[0032] The column temperature of the present invention is 20-35° C., preferably 25° C. At the above column temperature, the chromatographic peaks of the present invention are symmetrical, the separation is good, and the peaks are more complete.

[0033] The mobile phase A of the present invention is acetonitrile solution, the mobile phase B is 0.1% phosphoric acid aqueous solution, and the elution is performed in a gradient manner.

[0034] The inventors found that when the mobile phase is acetonitrile-0.1% phosphoric acid solution, the chromatographic peak baseline is relatively stable and there are more chromatographic peaks. Therefore, acetonitrile-0.1% phosphoric acid solution gradient elution is used as the mobile phase for the characteristic spectrum determination method of sword bean medicinal material.

[0035] Specifically, the gradient elution is as follows:

[0036] 0-12 min, phase A: 10→14%, phase B: 90→86%;

[0037] 12-22 min, phase A: 14% → 16%, phase B: 86% → 84%;

[0038] 22-25 min, phase A: 16% → 18%, phase B: 84% → 82%;

[0039] 25-45 min, phase A: 18% → 30%, phase B: 82% → 70%;

[0040] 45-50 min, phase A: 30%, phase B: 70%.

[0041] The theoretical plate number calculated based on rutin should not be less than 5000.

[0042] The present invention has good baseline separation under the above elution gradient, good separation of each peak, uniform distribution and stable baseline.

[0043] The flow rate of the mobile phase of the present invention is 0.2-0.3 mL / min.

[0044] The present invention found that the chromatographic peaks were well separated, the separation was moderate, and the peaks were completely separated under the above flow rate, which was the most preferred solution.

[0045] The injection volume is 1 to 3 μL; specifically, it can be 1 μL, 2 μL, or 3 μL.

[0046] The detection wavelength is 340-360 nm, preferably 340 nm. The inventors have found that at the above wavelength, the chromatographic peak information volume is larger, the chromatogram baseline is more stable, and the peak area of ​​each peak is larger.

[0047] The present invention, for the first time, uses ultra-high performance liquid chromatography to establish a characteristic spectrum detection method for distinguishing jack bean and its preparations. This method facilitates the overall evaluation of the scientific and rationality of the relevant process steps for extracts and preparations of jack bean and its processed products, and can better comprehensively control the intrinsic quality of the extracts and preparations of jack bean and its processed products, thereby ensuring the clinical efficacy of the extracts and preparations of jack bean and its processed products.

[0048] The invention adopts a traditional Chinese medicine chromatographic fingerprint similarity evaluation system to evaluate the similarity of UPLC characteristic spectra of sword bean and its preparation, and obtains a UPLC standard characteristic spectrum consisting of 13 characteristic peaks, wherein 0.44 (peak 1), 0.48 (peak 2), 0.61 (peak 3), 0.73 (peak 4), 0.88 (peak 5), 1.08 (peak 7), 1.53 (peak 8), 1.57 (peak 9), 1.59 (peak 10), 1.62 (peak 11), 1.66 (peak 12), and 1.70 (peak 13); wherein, peak 3 is quercetin-3-O-glucose-7-O-rhamnoside; peak 4 is myricetin-3-O-galactoside; peak 5 is mauritius herbicide; peak 6 (S) is rutin; and peak 7 is isoquercetin.

[0049] Quality judgment standard: Take samples of the extracts and preparations of Jack Bean and its processed products, and operate according to the same method as above to obtain the above-mentioned characteristic spectrum. Use the "Similarity Evaluation System of Chromatographic Fingerprints of Traditional Chinese Medicine" (2012 edition) of the National Pharmacopoeia Committee to analyze the characteristic spectrum of the extracts and preparations of Jack Bean and its processed products and the sample characteristic spectrum, and the similarity is greater than 0.90.

[0050] The present invention also provides a method for identifying characteristic spectra of sword beans and their preparations, which uses the method described in any one of claims 1 to 9 to perform detection and analyze the detection results.

[0051] The present invention provides a method for constructing a UPLC characteristic spectrum of Jack Bean and its preparations, comprising: A) dissolving and extracting a test sample raw material with a solvent to obtain a test solution; B) determining the test solution by high performance liquid chromatography to obtain a UPLC characteristic spectrum of Jack Bean and its preparations; the HPLC chromatographic conditions are as follows: a C18 column; mobile phase A is an acetonitrile solution; mobile phase B is a 0.1% phosphoric acid aqueous solution; and gradient elution is performed. The present invention uses high performance liquid chromatography, selects acetonitrile-0.1% phosphoric acid solution as the mobile phase for gradient elution, and uses quercetin-3-O-glucoside-7-O-rhamnoside, myricetin-3-O-galactoside, mauritius herbicide, rutin, and isoquercetin as reference substances to establish a UPLC characteristic spectrum method for Jack Bean and its preparations, providing more scientific and technological means for controlling the medicinal quality of Jack Bean and its preparations.

[0052] This invention, for the first time, employs ultra-high performance liquid chromatography to establish a method for simultaneously determining the characteristic chromatographic profiles of sword bean medicinal materials, standard decoctions, and formulated granules. The chromatographic conditions for this method, newly developed, offer the advantage of a wide selection range: wavelength: 340-360nm; column temperature: 20-40°C; flow rate: 0.2ml / min-0.3ml / min. This method achieves effective separation across a wide range of wavelengths, column temperatures, and flow rates.

[0053] The present invention, for the first time, uses ultra-high performance liquid chromatography to establish a method for simultaneously determining the characteristic spectrum of sword bean medicinal materials, standard decoctions, and formulated granules. The method also has the advantages of simple preparation and a wide range of selection for sample preparation. The method achieves effective separation using extraction solvents of methanol, 30% methanol, 50% methanol, and 80% methanol; extraction times of 20 minutes, 30 minutes, and 40 minutes; sample sizes of 0.5g, 1.0g, and 1.5g; and extraction methods of ultrasound, reflux, and decoction combined with ultrasound.

[0054] This study, for the first time, uses high-performance liquid chromatography to establish a characteristic chromatographic method for the simultaneous determination of Jack Bean medicinal materials, standard decoctions, and formulated granules. The method successfully identifies quercetin-3-O-glucoside-7-O-rhamnoside, myricetin-3-O-galactoside, mauritius herbicide, rutin, and isoquercetin, with quercetin-3-O-glucoside, myricetin-3-O-galactoside, and mauritius herbicide being identified for the first time.

[0055] The invention has the characteristics of strong operability, convenient analysis and stability. The invention uses ultra-high performance liquid phase fingerprint technology to control the quality of sword bean and its preparation, so that the product quality is effectively controlled.

[0056] The present invention can analyze the differences and changes of sword bean and its preparations, is conducive to the overall evaluation of the scientificity and rationality of sword bean-related process, can better control the intrinsic quality of sword bean and its preparations as a whole, and ensure the clinical efficacy of sword bean formula granules. BRIEF DESCRIPTION OF THE DRAWINGS

[0057] Figure 1 Mobile phase selection;

[0058] Figure 2 Chromatograms of sword bean medicinal materials at different wavelengths;

[0059] Figure 3 Column temperature investigation;

[0060] Figure 4 Flow rate investigation;

[0061] Figure 5 Injection volume inspection;

[0062] Figure 6 Delayed inspection;

[0063] Figure 7 Investigation of extraction solvents;

[0064] Figure 8 Extraction time investigation;

[0065] Figure 9 Sampling volume inspection;

[0066] Figure 10Investigation of extraction methods;

[0067] Figure 11 Chromatographic peak identification;

[0068] Figure 12 Surveys using different instruments;

[0069] Figure 13 Investigation of different chromatographic columns;

[0070] Figure 14 Characteristic spectrum of sword bean medicinal material;

[0071] Figure 15 Comparative characteristic spectrum of sword bean medicinal materials;

[0072] Figure 16 Characteristic spectrum of sword bean soup;

[0073] Figure 17 Comparative characteristic spectrum of standard decoction of sword bean;

[0074] Figure 18 Characteristic spectrum of sword bean formula particles;

[0075] Figure 19 Comparative characteristic spectrum of sword bean formula granules;

[0076] Figure 20 Comparison of characteristic spectra of sword bean medicinal material, standard soup, and granules;

[0077] Figure 21 UV spectrum of sword bean;

[0078] Figure 22 MS of myricetin-3-O-galactoside 1 and MS 2 Atlas;

[0079] Figure 23 MS of Mauritius herbicide 1 and MS 2 Atlas;

[0080] Figure 24 Rutin MS 1 and MS 2 Atlas;

[0081] Figure 25 MS of quercetin-3-O-glucoside-7-O-rhamnoside 1 and MS 2 Atlas;

[0082] Figure 26 MS of isoquercetin 1 and MS 2 Atlas;

[0083] Figure 27Gladiatoside A1 for MS 1 and MS 2 Atlas;

[0084] Figure 28 Gladiatoside A2 for MS 1 and MS 2 Atlas;

[0085] Figure 29 Gladiatoside A3 for MS 1 and MS 2 Atlas;

[0086] Figure 30 Gladiatoside B1 for MS 1 and MS 2 Atlas;

[0087] Figure 31 Gladiatoside B2 for MS 1 and MS 2 Atlas;

[0088] Figure 32 Gladiatoside B3 for MS 1 and MS 2 Atlas; Figure 33 Identification of chromatographic peaks of sword bean particles. DETAILED DESCRIPTION

[0089] The present invention provides a method for constructing a UPLC characteristic spectrum of sword bean and its preparation. Those skilled in the art can refer to the content of this article and appropriately improve the process parameters to achieve it. It should be noted in particular that all similar replacements and modifications are obvious to those skilled in the art and they all fall within the scope of protection of the present invention. The methods and applications of the present invention have been described through preferred embodiments. Relevant personnel can obviously modify or appropriately change and combine the methods and applications herein without departing from the content, spirit and scope of the present invention to implement and apply the technology of the present invention.

[0090] To further illustrate the present invention, the method for constructing a UPLC characteristic spectrum of sword bean and its preparation provided by the present invention is described in detail below in conjunction with the examples.

[0091] 1.1 Experimental instruments and materials

[0092] High performance liquid chromatograph: Agilent 1290 high performance liquid chromatograph, Thermo Fisher Vanquish F high performance liquid chromatograph, Waters-Class UPLC high performance liquid chromatograph;

[0093] Electronic balance: ME204E / 02, MS205DM, XP26 (Mettler-Toledo Instruments Co., Ltd.);

[0094] Ultrapure water machine: Cell type 1810A (Shanghai Moller Scientific Instrument Co., Ltd.);

[0095] Ultrasonic cleaner: KQ-600DB (600W, 40KHz; Kunshan Ultrasonic Instrument Co., Ltd.);

[0096] Chromatographic columns: DIKMAC18 2.1*150mm, 1.8μm; Shimadzu C18 2.1*150mm, 1.8μm; Waters C18 2.1×150mm, 1.8μm.

[0097] 1.2 Reagents and test drugs

[0098] Acetonitrile (Sigma-Aldrich, chromatographic grade); phosphoric acid (chromatographic grade); and ultrapure water.

[0099] Rutin (China Food and Drug Inspection Institute, batch number: 100080-202012, purity: 91.6%)

[0100] Quercetin-3-O-glucoside-7-O-rhamnoside (Tianjin Huaen Lilong Technology Co., Ltd., batch number: CFS202302, purity: ≥98%) Myricetin-3-O-galactoside (Chengdu Pusi Biotechnology Co., Ltd., batch number: PS013374, purity: 98.35%)

[0101] Mauritius herbicide (Sichuan Weiqiqi Biotechnology Co., Ltd., batch number: WP24081404, purity: 95.96%)

[0102] Isoquercetin (China Food and Drug Administration, batch number: 111809-202205, purity: 96.3%)

[0103] Jack bean control medicinal material (Chengdu Desite Biotechnology Co., Ltd., batch number: DSTYD001301)

[0104] Sword bean medicinal material batch number: DD01~DD21.

[0105] Sword bean standard decoction (prepared by Sichuan New Green Pharmaceutical Technology Development Co., Ltd., batch number: DDBT01~DDBT21)

[0106] Sword bean formula granules (batch number: DDKL1~DDKL3).

[0107] Example 1 Characteristic Spectrum Determination Method

[0108] Chromatographic conditions and system suitability testing were performed using octadecylsilane bonded silica gel as the filler (column length, 150 mm, inner diameter, 2.1 mm, particle size, 1.8 μm); acetonitrile as mobile phase A, 0.1% phosphoric acid solution as mobile phase B, with gradient elution as specified in the table below; flow rate, 0.2 ml / min; column temperature, 25°C; detection wavelength, 340 nm. The number of theoretical plates, calculated based on the rutin peak, must be no less than 5000.

[0109]

[0110] Preparation of Reference Solution: Take 1 g of the Cona bean reference medicinal material and place it in a stoppered Erlenmeyer flask. Add 50 ml of water and boil for 30 minutes. Let cool, filter, and evaporate the filtrate to dryness. Dissolve the residue in 25 ml of 80% methanol. Filter and use the filtrate as the reference medicinal material solution. Separately, accurately weigh an appropriate amount of rutin reference substance and add 80% methanol to a solution containing 20 μg per ml. This will serve as the reference substance solution.

[0111] Preparation of the test solution: Take 1.0 g of sword bean medicinal material powder (passed through No. 4 sieve), place it in a stoppered conical flask, add 25 ml of 80% methanol, stopper it tightly, and ultrasonically treat it (power 600 W, frequency 40 kHz) for 30 minutes. Let it cool, filter, and take the filtrate to obtain the product.

[0112] Determination method: Accurately aspirate 2μl of reference solution and test solution respectively, inject into liquid chromatograph, and determine.

[0113] 1.2.1 Chromatographic conditions and system suitability test

[0114] 1.2.1.1 Mobile phase selection

[0115] Based on the experimental conditions proposed above, the separation effect of acetonitrile-0.1% phosphoric acid solution mobile phase was investigated. Figure 1 . Figure 1 Selection of mobile phase: The results showed that when the mobile phase was acetonitrile-0.1% phosphoric acid solution, the chromatographic peak baseline was relatively stable and there were more chromatographic peaks. Therefore, acetonitrile-0.1% phosphoric acid solution gradient elution was used as the mobile phase for the characteristic spectrum determination method of sword bean medicinal material.

[0116] 1.2.1.2 Wavelength Selection

[0117] Based on the experimental conditions proposed above, the diode array detector was used to scan the entire wavelength range of the test solution, and the chromatograms of the test solution at wavelengths of 254 nm, 280 nm, 310 nm, 340 nm, and 360 nm were extracted respectively. Figure 2 . Figure 2 Chromatograms of sword bean medicinal material at different wavelengths; the results show that when the detection wavelength is 340nm-360nm, the chromatographic peak information volume is larger and the chromatogram baseline is more stable.

[0118] 1.2.1.3 Column temperature investigation

[0119] Based on the experimental conditions proposed above, the column temperatures of 20℃, 25℃, 30℃ and 35℃ were investigated respectively. Figure 3 , Table 1. Figure 3 Column temperature inspection.

[0120] Table 1-1 Column temperature investigation-relative retention time

[0121]

[0122] Table 1-2 Column temperature investigation-relative retention time

[0123]

[0124] The results showed that when the column temperature was 20℃, 25℃, 30℃ and 35℃, the chromatogram peak shape was relatively symmetrical, the separation was good and the peak extraction was more complete.

[0125] 1.2.1.4 Flow rate investigation

[0126] Based on the experimental conditions proposed above, the flow rates of 0.20ml / min, 0.25ml / min and 0.30ml / min were investigated respectively. Figure 4 , Table 2. Figure 4 Flow rate investigation.

[0127] Table 2-1 Flow rate investigation-relative retention time

[0128]

[0129] Table 2-2 Flow rate investigation-relative retention time

[0130]

[0131] The results showed that the chromatographic peak shapes were good and the separation was moderate when the flow rates were 0.20ml / min, 0.25ml / min and 0.30ml / min.

[0132] 1.2.1.5 Sample injection volume investigation

[0133] Based on the experimental conditions proposed above, the injection volumes of 1μl, 2μl and 3μl were investigated respectively. Figure 5 , Table 3. Figure 5 Injection volume inspection.

[0134] Table 3-1 Injection volume investigation-relative retention time

[0135]

[0136] Table 3-2 Injection volume investigation-relative retention time

[0137]

[0138]

[0139] The results showed that the chromatographic peak shape was good when the injection volume was 1-3 μl.

[0140] 1.2.1.6 Delay Investigation

[0141] Based on the experimental conditions proposed above, a delay test was conducted. The results are shown in Figure 6 . Figure 6 Delayed inspection.

[0142] The results showed that the sample had basically no chromatographic peak after 50 minutes, so the sample detection time was set to 50 minutes.

[0143] In summary, the chromatographic conditions and system suitability testing for the characteristic spectrum of Jack Bean (canavalia) are as follows: octadecylsilane bonded silica gel as the filler (column length, 150 mm, inner diameter, 2.1 mm, particle size, 1.8 μm); acetonitrile as mobile phase A, 0.1% phosphoric acid solution as mobile phase B, gradient elution as specified in the table below; flow rate, 0.2 ml / min; column temperature, 25°C; detection wavelength, 340 nm. The number of theoretical plates, calculated based on the rutin peak, must be no less than 5000.

[0144]

[0145] 1.2.2 Preparation of test solution

[0146] 1.2.2.1 Investigation of extraction solvent

[0147] Take 1.0 g of Jack Bean powder (passed through a No. 4 sieve) (batch number: 010646-2211001) and place it in a stoppered conical flask. 25 ml of each of the extraction solvents for the test sample were water, methanol, 80% ethanol, 80% methanol, 50% methanol, and 30% methanol. Seal the flask tightly and ultrasonically treat (power 600 W, frequency 40 kHz) for 30 minutes. Let it cool, shake well, filter, and take the filtrate to obtain the product. Figure 7 . Figure 7 The extraction solvent was investigated; the results showed that when the extraction solvent was methanol of different concentrations, the peak shapes of the characteristic peaks were good and the separation was moderate.

[0148] 1.2.2.2 Extraction time investigation

[0149] Take 1.0 g of Jack Bean powder (passed through a No. 4 sieve) (batch number: 010646-2211001) and place it in a stoppered conical flask. Add 25 ml of 80% methanol, stopper tightly, and sonicate (power 600 W, frequency 40 kHz) for 20 minutes, 30 minutes, and 40 minutes, respectively. Let cool, shake well, filter, and take the filtrate to obtain the product. Figure 8 . Figure 8 Extraction time investigation.

[0150] The results showed that the peak shape and separation of the chromatograms were better when the extraction time was 20, 30 and 40 minutes.

[0151] 1.2.2.3 Sampling volume investigation

[0152] Take 0.5g, 1.0g and 1.5g of the jack bean powder (passed through a No. 4 sieve) (batch number: 010646-2211001) respectively, add 25ml of 80% methanol, place in a stoppered conical flask, seal tightly, and ultrasonically treat (power 600W, frequency 40kHz) for 30 minutes. Let cool, shake well, filter, and take the filtrate to obtain the product. Figure 9 . Figure 9 The results of sampling volume investigation showed that when the sampling volume was 0.5g, 1.0g and 1.5g, the peak shape and separation of each chromatographic peak were good.

[0153] 1.2.2.4 Investigation of extraction methods

[0154] Take 1.0g of sword bean powder (passed through No. 4 sieve) (batch number: 010646-2211001), place it in a stoppered conical flask, add 25ml of 80% methanol, seal it tightly, and ultrasonically treat it (power 600W, frequency 40kHz), reflux it for 30 minutes, let it cool, filter it, and take the filtrate to obtain the product; take another 1.0g of sword bean powder, place it in a stoppered conical flask, add 50ml of water, decoct it for 30 minutes, filter it, evaporate the filtrate to dryness, add 25ml of 80% methanol to the residue, seal it tightly, ultrasonically treat it (power 600W, frequency 40kHz) for 30 minutes, let it cool, shake it well, filter it, and take the filtrate to obtain the product. Figure 10 . Figure 10 The extraction methods were investigated; the results showed that there was little difference between the different extraction methods.

[0155] In summary, the preparation method of the test solution of the characteristic spectrum of the sword bean medicinal material is tentatively determined as follows: take 1.0 g of the sword bean medicinal material powder (passed through a No. 4 sieve), place it in a stoppered conical flask, add 25 ml of 80% methanol, seal it tightly, and ultrasonically treat it (power 600 W, frequency 40 kHz) for 30 minutes. Let it cool, shake it evenly, filter it, and take the filtrate to obtain it.

[0156] Example 2 Methodological Investigation

[0157] 1.2.3.1 Chromatographic peak identification

[0158] Preparation of test solution: Take 1.0 g of sword bean powder (passed through No. 4 sieve), place in a stoppered conical flask, add 25 ml of 80% methanol, stopper, ultrasonically treat (power 600 W, frequency 40 kHz) for 30 minutes, let cool, filter, and take the filtrate to obtain

[0159] Preparation of control medicinal material solution: Take 1 g of sword bean control medicinal material, place it in a stoppered conical flask, add 50 ml of water, boil for 30 minutes, cool, filter, evaporate the filtrate to dryness, add 25 ml of 80% methanol to dissolve the residue, filter, and take the filtrate as the control medicinal material reference solution.

[0160] Preparation of reference solution: Take appropriate amount of quercetin-3-O-glucoside-7-O-rhamnoside, myricetin-3-O-galactoside, mauritius herbicide, rutin, and isoquercetin reference substances, accurately weigh them, and add 80% methanol to make a solution containing 20 μg of each per 1 ml, which serves as the reference solution.

[0161] Preparation of negative control solution: According to the experimental conditions proposed above, prepare the negative control solution lacking the sword bean medicinal material.

[0162] Position the characteristic spectrum peaks of sword bean medicinal materials. Figure 11 , specifically including 11-a~11-k; Figure 11 Chromatographic peak identification, specifically, 11-a is the spectrum of quercetin-3-O-glucose-7-O-rhamnoside - reference substance, 11-b is the spectrum of quercetin-3-O-glucose-7-O-rhamnoside - medicinal material; 11-c is the spectrum of myricetin-3-O-galactoside - reference substance, 11-d is the spectrum of myricetin-3-O-galactoside - medicinal material; 11-e is the spectrum of mauritius herbicide - reference substance; 11-f is the spectrum of mauritius herbicide - medicinal material; 11-g is the spectrum of rutin - reference substance; 11-h is the spectrum of rutin - medicinal material; 11-i is the spectrum of isoquercetin - reference substance; 11-j is the spectrum of isoquercetin - medicinal material, and 11-k is chromatographic peak identification.

[0163] The results showed that peak 3 was quercetin-3-O-glucoside-7-O-rhamnoside, peak 4 was myricetin-3-O-galactoside, peak 5 was mauritius herbicide, peak 6 was rutin, and peak 7 was isoquercetin. In the following methodological investigation, 13 characteristic peaks in the sample were investigated.

[0164] 1.2.3.2 Precision test

[0165] Take the test solution of Jack Bean (Batch No. 010646-2211001) and inject 2 μl of the solution six times according to the proposed experimental method. Calculate the retention time of each characteristic peak. See Table 4.

[0166] Table 4-1 Precision Investigation-Retention Time

[0167]

[0168] Table 4-2 Precision Investigation-Retention Time

[0169]

[0170] The results showed that the RSD of the retention time of each characteristic peak of the samples was 0.05% to 0.32%.

[0171] 1.2.3.3 Repeatability Study

[0172] Six samples of Cona Bean (Batch No. 010646-2211001) were prepared and assayed according to the proposed experimental method (see Table 5).

[0173] Table 5-1 Repeatability Study - Relative Retention Time

[0174]

[0175] Table 5-2 Repeatability Study - Relative Retention Time

[0176]

[0177]

[0178] The results showed that the RSD of the relative retention time of the six samples was 0.04% to 0.29%, indicating that the method had good repeatability.

[0179] 1.2.3.4 Intermediate precision study

[0180] 1.2.3.4.1 Investigation of different instruments

[0181] Based on the experimental conditions proposed above, two portions of Jack Bean (Batch No.: 010646-2211001) were weighed to prepare test solutions, which were then analyzed on Waters Acquity UPLC H-Class, Thermo Fisher Scientific Vanquish F, and Agilent 1290 HPLC instruments, respectively. See Table 6. Figure 12 . Figure 12 Survey with different instruments.

[0182] Table 6-1 Instrument durability assessment - relative retention time

[0183]

[0184] Table 6-2 Instrument durability assessment - relative retention time

[0185]

[0186] The results showed that when the three instruments were used to detect the test samples, the RSDs of the relative retention times of the characteristic peaks were 0.19% to 2.57%, indicating that the instruments had good durability.

[0187] 1.2.3.4.2 Inspection by different personnel and time

[0188] Based on the experimental conditions proposed above, two samples of Jack Bean (Batch No. 010646-2211001) were weighed by different individuals (A and B) at different times (T1 and T2) to prepare test samples for determination (see Table 7).

[0189] Table 7-1 Personnel and Time Inspection - Relative Retention Time

[0190]

[0191] Table 7-2 Personnel and Time Inspection - Relative Retention Time

[0192]

[0193] The results showed that when different people measured the same sample at different times, the RSDs of the relative retention times of the characteristic peaks were 0.06% to 0.38%, indicating that the method was stable.

[0194] 1.2.3.5 Durability inspection

[0195] 1.2.3.5.1 Column durability assessment

[0196] Based on the experimental conditions proposed above, the chromatographic columns DIKMA C18 2.1*150mm, 1.8μm; Shimadzu C18 2.1*150mm, 1.8μm; and Waters C18 2.1×150mm, 1.8μm were analyzed and investigated. The results are shown in Table 8. Figure 13 . Figure 13 Investigation of different chromatographic columns.

[0197] Table 8-1 Chromatographic column durability inspection - relative retention time

[0198]

[0199] Table 8-2 Chromatographic column durability inspection - relative retention time

[0200]

[0201] 8 The results showed that when the samples were detected using the above three chromatographic columns, the RSDs of the relative retention times of the characteristic peaks were between 1.39% and 4.98%, indicating that the chromatographic columns had good durability.

[0202] 1.2.3.5.2 Stability assessment

[0203] Based on the experimental conditions proposed above, the same test solution was taken and measured at 0h, 4h, 8h, 12h, 16h, and 24h. See Table 9.

[0204] Table 9-1 Stability Study - Retention Time

[0205]

[0206] Table 9-2 Stability Study - Retention Time

[0207]

[0208] 9 The results showed that the RSD of the characteristic peak retention time was between 0.03% and 0.34%, and the sample solution was stable within 24 hours.

[0209] In summary, the RSDs of the relative retention times of the characteristic peaks met the requirements in all the above investigations, indicating that the method was good. The above 13 characteristic peaks were included in the subsequent investigation.

[0210] 1.2.4 Verification of the characteristic spectrum of sword bean medicinal materials

[0211] The sword bean medicinal material was tested and the relative retention time was calculated. The results are shown in Figure 14 , Table 10. Figure 14 Characteristic spectrum of sword bean medicinal material; (S1-S21 are: DD01~DD21)

[0212] Peak 3: Quercetin-3-O-glucoside-7-O-rhamnoside; Peak 4: Myricetin-3-O-galactoside; Peak 5: Mauritius herbicide; Peak 6 (S):

[0213] Rutin; Peak 7: Isoquercetin

[0214] Table 10-1 Relative retention time of characteristic spectrum of sword bean medicinal material

[0215]

[0216] Table 10-2 Relative retention time of characteristic spectrum of sword bean medicinal material

[0217]

[0218]

[0219] Based on the principle of stable relative retention times and the fact that peaks can be detected in all batches of samples at relatively high levels, a total of 13 peaks with good reproducibility were selected as characteristic peaks. The relative retention time RSDs of the 13 characteristic peaks in 21 batches of sword bean medicinal materials were all less than 1.0%.

[0220] To ensure the relevance of the sword bean medicinal material and the sword bean formula granules, the specified relative retention time values ​​for the sword bean medicinal material characteristic spectrum are specified to be consistent with those for the sword bean formula granules. The final regulations stipulate that the test sample chromatogram should show 13 characteristic peaks, and the retention times should correspond to the 13 characteristic peaks in the reference medicinal material chromatogram. The peak corresponding to the rutin reference peak is the S peak, and the relative retention times of the remaining peaks and the S peak are calculated; their relative retention times should be within ±10% of the specified values. The specified values ​​are: 0.44 (peak 1), 0.48 (peak 2), 0.61 (peak 3), 0.73 (peak 4), 0.88 (peak 5), 1.08 (peak 7), 1.53 (peak 8), 1.57 (peak 9), 1.59 (peak 10), 1.62 (peak 11), 1.66 (peak 12), and 1.70 (peak 13).

[0221] The Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System (2012 Edition) was used to synthesize 21 batches of Jack Bean medicinal materials, and a reference characteristic spectrum of Jack Bean medicinal materials was established. Figure 15 . Figure 15 Comparative characteristic spectrum of Jack Bean medicinal materials. Among them, quercetin-3-O-glucoside-7-O-rhamnoside; Peak 4: myricetin-3-O-galactoside; Peak 5: mauritius herbicide; Peak 6 (S): rutin; Peak 7: isoquercetin.

[0222] Example 3 Sword Bean Standard Decoction

[0223] 2.1. Materials, reagents, and instruments are the same as those in Example 1;

[0224] 2.2 Characteristic spectrum determination method is the same as the characteristic spectrum determination method in Example 1.

[0225] 2.3 Determination of characteristic peaks and establishment of reference maps

[0226] The proposed method was used to determine the characteristic spectra of 21 batches of standard decoction samples of sword bean and calculate the relative retention time. Figure 16 , Table 11. Figure 16 Characteristic spectrum of sword bean soup (Note: 1 to 21 are DDBT01 to DDBT21 respectively)

[0227] Peak 3: quercetin-3-O-glucoside-7-O-rhamnoside; Peak 4: myricetin-3-O-galactoside; Peak 5: mauritius herbicide; Peak 6 (S): rutin; Peak 7: isoquercetin.

[0228] Table 11-1 Relative retention time of characteristic spectrum of standard decoction of sword bean

[0229]

[0230]

[0231] Table 11-2 Relative retention time of characteristic spectrum of standard decoction of sword bean

[0232]

[0233] 1 Based on the principle of stable relative retention times, detectable peaks across all batches of samples, and relatively high peaks, 13 peaks with good reproducibility were selected as characteristic peaks. The RSDs of the relative retention times of peaks 1 to 13 were 0.07% to 0.30%.

[0234] The relative retention time specifications for the standard decoction of sword bean are consistent with those for the sword bean formula granules. The final regulations stipulate that the test sample should present 13 characteristic peaks in the chromatogram, and the retention times should correspond to the 13 characteristic peaks in the chromatogram of the reference medicinal material. The peak corresponding to the peak of the rutin reference material is the S peak, and the relative retention times of the remaining peaks and the S peak are calculated; their relative retention times should be within ±10% of the specified values. The specified values ​​are: 0.44 (peak 1), 0.48 (peak 2), 0.61 (peak 3), 0.73 (peak 4), 0.88 (peak 5), 1.08 (peak 7), 1.53 (peak 8), 1.57 (peak 9), 1.59 (peak 10), 1.62 (peak 11), 1.66 (peak 12), and 1.70 (peak 13).

[0235] The Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System (2012 Edition) was used to synthesize 21 batches of standard decoctions of Jack Bean, and a comparison of the characteristic spectra of standard decoctions of Jack Bean was established. Figure 17 . Figure 17 Comparative characteristic spectrum of standard decoction of sword bean. Peak 3: Quercetin-3-O-glucoside-7-O-rhamnoside; Peak 4: Myricetin-3-O-galactoside; Peak 5: Mauritius herbicide; Peak 6(S): Rutin; Peak 7: Isoquercetin.

[0236] Example 4 Sword Bean Formula Granules

[0237] 3.1 Materials, reagents and instruments The materials, reagents and instruments are the same as those in Example 1.

[0238] 3.2 Characteristic spectrum determination method: Same as the characteristic spectrum determination method in Example 1.

[0239] 3.3 Determination of characteristic peaks and establishment of reference maps

[0240] The proposed method was used to determine the characteristic spectra of three batches of sword bean formula granule samples and calculate the relative retention time. Figure 18 , Table 12. Figure 18 Characteristic spectrum of sword bean formula granules (S1 to S3 are: DDKL1, DDKL2, DDKL3); Peak 3: Quercetin-3-O-glucoside-7-O-rhamnoside; Peak 4: Myricetin-3-O-galactoside; Peak 5: Mauritius herbicide; Peak 6 (S): Rutin; Peak 7: Isoquercetin.

[0241] Table 12-1 Sword Bean Formula Granules - Relative Retention Time

[0242]

[0243] Table 12-2 Sword bean formula granules - relative retention time

[0244]

[0245] Based on the principles of stable relative retention times, consistent detection across all batches of samples, and relatively high peak heights, 13 peaks with good durability were selected as characteristic peaks. The relative retention times of each characteristic peak were stable and within ±10% of the mean value, so the specified range for the relative retention time of each peak was temporarily set at ±10%.

[0246] Final regulations: The test sample chromatogram should show 13 characteristic peaks, and the retention times should correspond to the 13 characteristic peaks in the chromatogram of the reference medicinal material. The peak corresponding to the rutin reference peak is the S peak, and the relative retention times of the remaining peaks and the S peak are calculated; their relative retention times should be within ±10% of the specified values. The specified values ​​are: 0.44 (peak 1), 0.48 (peak 2), 0.61 (peak 3), 0.73 (peak 4), 0.88 (peak 5), 1.08 (peak 7), 1.53 (peak 8), 1.57 (peak 9), 1.59 (peak 10), 1.62 (peak 11), 1.66 (peak 12), and 1.70 (peak 13).

[0247] Three batches of sword bean formula granules were synthesized using the Chinese medicine chromatographic fingerprint similarity evaluation system (2012 version), and a comparison spectrum of the sword bean formula granules characteristic spectrum was established. Figure 19 . Figure 19 Comparative characteristic spectrum of sword bean formula granules; Peak 3: Quercetin-3-O-glucoside-7-O-rhamnoside; Peak 4: Myricetin-3-O-galactoside; Peak 5: Mauritius herbicide; Peak 6(S): Rutin; Peak 7: Isoquercetin.

[0248] The Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System (2012 Edition) was used to combine the reference patterns of Jack Bean Herbal Material, Jack Bean Soup, and Jack Bean Formula Granules on one graph for comparison. Figure 23 Figure 23 Comparison of characteristic spectra of sword bean medicinal material, standard soup, and granules; Peak 3: Quercetin-3-O-glucoside-7-O-rhamnoside; Peak 4: Myricetin-3-O-galactoside; Peak 5: Mauritius herbicide; Peak 6(S): Rutin; Peak 7: Isoquercetin.

[0249] Example 5: Mass Spectrometry Identification of Sword Beans

[0250] 4.1 High-resolution mass spectrometry (Q-TOF-MS)

[0251] Instrument: Thermo Fisher Ultra-High Performance Liquid Chromatograph

[0252] AB Sciex Triple 5600 high-resolution mass spectrometer, SCIEX;

[0253] Chromatographic column: DIKMAC18 2.1*150mm, 1.8μm

[0254] Preparation of test solution: Take an appropriate amount of sword bean formula granules, grind them into powder, take 1g, place it in a stoppered conical flask, add 25ml of 80% methanol, stopper it tightly, and ultrasonically treat it (power 600W, frequency 40kHz) for 30 minutes. Let it cool, shake it well, filter it, and take the filtrate to obtain the product.

[0255] The mass spectrometry conditions are shown in Table 13.

[0256] Table 13 Mass spectrometry conditions

[0257]

[0258]

[0259] Chromatographic conditions and system suitability testing were performed using octadecylsilane bonded silica gel as the filler (column length, 150 mm, inner diameter, 2.1 mm, particle size, 1.8 μm); acetonitrile as mobile phase A, 0.1% phosphoric acid solution as mobile phase B, with gradient elution as specified in the table below; flow rate, 0.2 ml / min; column temperature, 25°C; detection wavelength, 340 nm. The number of theoretical plates, calculated based on the myricetin-3-O-galactoside peak, must be no less than 5000.

[0260]

[0261] 4.2 Experimental Results

[0262] 4.2.1 Related Graphs

[0263] Based on the experimental conditions proposed above, the test solution of sword bean formula granules was analyzed, and the results are shown in Figure 21 .

[0264] Figure 21 UV spectrum of sword bean.

[0265] 4.2.2 Mass spectrometry results

[0266] UPLC-Q-TOF / MS was used to analyze the sword bean formula granules. The mass spectrum peaks were identified based on the multi-level mass spectrum information of the samples, combined with the natural product high-resolution mass spectrum database and related literature. The results are shown in Table 14. Figure 22-32 . Figure 22 MS of myricetin-3-O-galactoside 1 and MS 2 Atlas; Figure 23 MS of Mauritius herbicide 1 and MS 2 Atlas; Figure 24 Rutin MS 1 and MS 2 Atlas; Figure 25 MS of quercetin-3-O-glucoside-7-O-rhamnoside 1 and MS 2 Atlas;

[0267] Figure 26 MS of isoquercetin 1 and MS 2 Atlas; Figure 27 Gladiatoside A1 for MS 1 and MS 2 Atlas; Figure 28 Gladiatoside A2 for MS 1 and MS 2 Atlas; Figure 29 Gladiatoside A3 for MS 1 and MS 2 Atlas; Figure 30 Gladiatoside B1 for MS 1 and MS 2 Atlas; Figure 31 Gladiatoside B2 for MS 1 and MS 2 Atlas; Figure 32 Gladiatoside B3 for MS 1 and MS 2 Atlas.

[0268] Table 14 Characteristic peak identification results

[0269]

[0270]

[0271] According to the results of high-resolution mass spectrometry (Q-TOF-MS), a total of quercetin-3-O-glucoside-7-O-rhamnoside, myricetin-3-O-galactoside, mauritius herbicide, rutin, isoquercetin, GladiatosideA1, GladiatosideA2, GladiatosideA3, GladiatosideB1, GladiatosideB2, and GladiatosideB3 were identified.

[0272] Peak identification by UPLC reference standard ( Figure 11 Peak identification by HPLC of quercetin-3-O-gluco-7-O-rhamnoside, myricetin-3-O-galactoside, herbamoyl benzoate, rutin, and isoquercetin were consistent with the spectral confirmation results. Peak 3 was identified as quercetin-3-O-gluco-7-O-rhamnoside, peak 4 as myricetin-3-O-galactoside, peak 5 as herbamoyl benzoate, peak 6 as rutin, and peak 7 as isoquercetin. Among them, Gladiatoside A1 (peak 8), Gladiatoside A2 (peak 9), and Gladiatoside A3 (peak 12) are isomers, and Gladiatoside B1 (peak 10), Gladiatoside B2 (peak 11), and Gladiatoside B3 (peak 13) are isomers.

[0273] Comparative Example 1

[0274] 1.2 Chromatographic conditions and system suitability test

[0275] Octadecylsilane bonded silica gel was used as the filler (column length: 250 mm, inner diameter: 4.6 mm, particle size: 5.0 μm);

[0276] Use methanol as mobile phase A and 0.1% phosphoric acid solution as mobile phase B, and perform gradient elution as specified in the table below; flow rate: 1.0 mL / min; column temperature: 30°C; detection wavelength: 330 nm. The number of theoretical plates, calculated based on the rutin peak, must be no less than 5000.

[0277] Table 15 Proposed mobile phase gradient

[0278] Time (minutes) Mobile phase A (%) Mobile phase B (%) 0～3 0→15 100→85 3～18 15→35 85→65 18～31 35→40 65→60 31～45 40→55 60→45 45～55 55→60 45→40

[0279] Take 1.0 g of sword bean granule powder, place it in a stoppered conical flask, add 10 mL of 80% methanol, and ultrasonically treat (power 600 W, frequency 40 kHz) for 30 minutes. Cool, filter, and take the filtrate to obtain the product.

[0280] Determination method: Accurately aspirate 10 μL of reference solution and test solution respectively, inject into liquid chromatograph, and determine.

[0281] The results are as follows Figure 33 As shown, Figure 33 The results showed that the retention times of peaks 3, 4, and 5 were consistent with those of mauritius herbicide, rutin, and isoquercetin, respectively, but quercetin-3-O-glucoside-7-O-rhamnoside could not be identified under these chromatographic conditions.

[0282] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as within the scope of protection of the present invention.

Claims

1. A method for constructing a UPLC characteristic spectrum of sword bean and its preparation, comprising: A) Dissolve the test sample in a solvent and extract to obtain a test solution; B) measuring the test solution by high performance liquid chromatography to obtain a UPLC characteristic spectrum of the sword bean and its preparation; The HPLC conditions are as follows: the chromatographic column is a C18 column; the mobile phase A is an acetonitrile solution, the mobile phase B is a 0.1% phosphoric acid aqueous solution, and the elution is performed in a gradient manner; The gradient elution is specifically as follows: 0-12 min, phase A: 10→14%, phase B: 90→86%; 12-22 min, phase A: 14% → 16%, phase B: 86% → 84%; 22-25 min, phase A: 16% → 18%, phase B: 84% → 82%; 25-45 min, phase A: 18% → 30%, phase B: 82% → 70%; 45-50 min, phase A: 30%, phase B: 70%.

2. The method according to claim 1, characterized in that The method further includes preparing a reference solution: taking quercetin-3-O-glucoside-7-O-rhamnoside, myricetin-3-O-galactoside, herbicide-resistant mauritius, rutin, and isoquercetin reference substances, dissolving them in 80% methanol to obtain reference solution; The control medicinal material of sword bean was decocted in water, cooled, filtered, evaporated to dryness, and the residue was dissolved in 80% methanol and filtered to obtain a control medicinal material reference solution; The reference substance solution and the reference medicinal material reference substance solution are respectively measured by high performance liquid chromatography to obtain chromatograms of the reference substance and the reference medicinal material reference substance; and the components of sword bean and its preparation are qualitatively analyzed based on the chromatograms of the reference substance.

3. The method according to claim 2, characterized in that The concentrations of the reference solution are specifically as follows: 20 μg / mL of quercetin-3-O-glucoside-7-O-rhamnoside, 20 μg / mL of myricetin-3-O-galactoside, 20 μg / mL of galactopyranoside, 20 μg / mL of rutin, and 20 μg / mL of isoquercetin.

4. The method according to claim 1, wherein The chromatographic column is C 18 150×2.1mm 1.8μm; column temperature 20-35℃; theoretical plate number calculated based on rutin should be no less than 5000.

5. The method according to claim 4, characterized in that The flow rate of the mobile phase is 0.2-0.3 mL / min; the injection volume is 1-3 μL.

6. The method according to claim 4, characterized in that The detection wavelength is 340-360 nm.

7. The method according to claim 4, characterized in that In step A), the solvent is 80% methanol; the extraction is ultrasonic extraction; the ultrasonic power is 600W, the frequency is 40kHz; and the ultrasonic time is 20 to 40 minutes.

8. The method according to claim 1, characterized in that The ratio of the mass g of the test material to the volume mL of the solvent is (0.5-1.5):25; The test sample raw materials are one or more of sword bean medicinal materials, sword bean standard decoction or sword bean formula granules.

9. The method according to claim 1, characterized in that The similarity of the UPLC characteristic spectra of Jack Bean and its preparations was evaluated using the traditional Chinese medicine chromatographic fingerprint similarity evaluation system, and a UPLC standard characteristic spectrum consisting of 13 characteristic peaks was obtained, including 0.44 (peak 1), 0.48 (peak 2), 0.61 (peak 3), 0.73 (peak 4), 0.88 (peak 5), 1.08 (peak 7), 1.53 (peak 8), 1.57 (peak 9), 1.59 (peak 10), 1.62 (peak 11), 1.66 (peak 12), and 1.70 (peak 13); among them, Peak 3: Quercetin-3-O-glucoside-7-O-rhamnoside; Peak 4: Myricetin-3-O-galactoside; Peak 5: Mauritius herbicide; Peak 6 (S): Rutin; Peak 7: Isoquercitrin.

10. A method for identifying characteristic patterns of sword beans and their preparations, characterized in that: The method according to any one of claims 1 to 9 is used for detection, and the detection results are analyzed.

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