A method for establishing a characteristic chromatogram of corn silk and its pharmaceutical preparation and application thereof

Abstract

The application belongs to the technical field of quality detection of traditional Chinese medicine preparation, and particularly relates to a characteristic spectrum of corn silk medicinal material and corn silk medicinal preparation, and further discloses a construction method of the characteristic spectrum and a quality detection method of the corn silk medicinal material and the corn silk medicinal preparation. The construction method of the characteristic spectrum of the corn silk medicinal material and the corn silk medicinal preparation takes the corn silk medicinal material and the corn silk medicinal preparation as a detection object, and establishes a characteristic spectrum method for the medicinal preparation based on high performance liquid chromatography. The method has high precision, good stability and repeatability, and can comprehensively and quickly detect effective components and contents in the corn silk formula granules and other medicinal preparations.

Description

Technical Field

[0001] This invention belongs to the field of quality testing technology for traditional Chinese medicine preparations, specifically relating to a characteristic spectrum of corn silk medicinal material and corn silk drug preparations, and further disclosing the method for constructing the characteristic spectrum, as well as the quality testing method for corn silk medicinal material and corn silk drug preparations. Background Technology

[0002] Traditional Chinese medicine (TCM) has complex components, and its effective parts are often not single components. Using a single component as a quality control indicator is increasingly inadequate for the quality control requirements of TCM. Therefore, TCM fingerprinting technology has emerged. Originating from fingerprint identification, TCM fingerprinting technology utilizes modern information technology and quality analysis methods to comprehensively reflect the types and quantities of chemical components contained in TCM. It obtains a characteristic spectrum of TCM by identifying the common peaks of various component groups. For raw TCM materials, the characteristic spectrum can be used to identify authenticity and assess quality; for prepared TCM products, it can identify product authenticity, assess the rationality of the preparation process, and effectively control product quality. Currently, the effective components of most TCMs are not yet clearly defined. The holistic and fuzzy nature of TCM characteristic spectrum is well-suited to this characteristic, making it more scientific and comprehensive than single-component quality control methods. Internationally, TCM characteristic spectrum is now recognized as a quality control model for TCM. High-performance liquid chromatography (HPLC), with its advantages of high separation efficiency and fast analysis speed, has become the main analytical method for characteristic spectrum analysis.

[0003] Corn silk is the dried style and stigma of Zea mays L., a plant belonging to the Poaceae family. It mainly contains flavonoids, organic acids, nucleosides, amino acids, and other components, and is primarily used for renal edema, dysuria, damp-heat jaundice, hypertension, and diabetes. Corn silk granules are obtained through the extraction, concentration, and granulation of this traditional Chinese medicine.

[0004] The 1977 edition of the Chinese Pharmacopoeia included quality control measures for corn silk, covering aspects such as the original plant variety, processing of the processed slices, characteristics of the slices, and physicochemical identification. Literature also described the chemical components of corn silk, including organic acids, nucleosides, and amino acids. However, on the one hand, determining or identifying the content of a single component in corn silk formulations cannot comprehensively detect and control their quality; on the other hand, combining the determination of a single component with the identification of other components is time-consuming and labor-intensive, making it difficult to widely apply in production practice. Furthermore, most existing quality control techniques for corn silk focus on processed corn silk slices, not on corn silk pharmaceutical preparations, and existing detection methods for processed corn silk slices are not applicable to corn silk pharmaceutical preparations.

[0005] Therefore, there is a need in the field to develop a method that is applicable to corn silk and its pharmaceutical preparations, and that can comprehensively and rapidly detect flavonoid components in corn silk pharmaceutical preparations. This is of great significance for comprehensive quality testing and overall quality control. Summary of the Invention

[0006] The technical problem to be solved by the present invention is to provide a method for constructing a characteristic spectrum of corn silk medicinal material and its pharmaceutical preparations, so as to comprehensively reflect the intrinsic quality and medication safety of corn silk medicinal material and its pharmaceutical preparations;

[0007] The second technical problem to be solved by the present invention is to provide a quality testing method for corn silk medicinal materials and their pharmaceutical preparations.

[0008] To solve the above-mentioned technical problems, the present invention provides a method for constructing characteristic spectra of corn silk and its pharmaceutical preparations, which includes the step of performing high-performance liquid chromatography detection on the test solution of corn silk medicinal material and / or corn silk pharmaceutical preparations.

[0009] Chromatographic conditions included: using octadecylsilane-bonded silica gel as the stationary phase, acetonitrile as mobile phase A, and 0.1% formic acid solution as mobile phase B, with gradient elution performed according to the following procedure:

[0010] 0-10 min, A:B ratio was 7%:93% → 10%:90%;

[0011] 10-22 min, A:B ratio changed from 10%:90% to 12%:88%;

[0012] 22-40 min, A:B ratio changed from 12%:88% to 14%:86%;

[0013] 40-45 min, A:B ratio is 14%:86%;

[0014] Preferably, the gradient elution further includes the following procedure:

[0015] 45-60 min, A:B ratio changed from 14%:86% to 16%:84%;

[0016] 60-65 min, A:B ratio was 16%:84%.

[0017] Specifically, in the method for constructing the characteristic chromatograms of corn silk and its pharmaceutical preparations, the chromatographic conditions in the high-performance liquid chromatography detection step further include:

[0018] The column temperature is 20-30℃, preferably 25℃; and / or,

[0019] The flow rate is 0.1-0.5 mL / min, preferably 0.29-0.31 mL / min, more preferably 0.3 mL / min; and / or,

[0020] The detection wavelength is 330 nm; and / or,

[0021] The injection volume is 1-5 μL, preferably 3 μL.

[0022] Specifically, the method for constructing the characteristic spectrum of corn silk and its pharmaceutical preparations includes the following steps for preparing the test solution: taking the test sample, adding an organic solvent for extraction, collecting the extract for solid-liquid separation, and collecting the filtrate to obtain the final product.

[0023] Specifically, the method for constructing the characteristic spectrum of corn silk and its pharmaceutical preparations further includes the step of preparing a reference solution of a control medicinal material, which specifically includes: taking corn silk as a control medicinal material, adding water and heating and refluxing for extraction, collecting the extract and separating the solid and liquid, collecting the residue and adding an organic solvent for extraction, collecting the extract and separating the solid and liquid, collecting the filtrate, and obtaining the reference solution of the control medicinal material.

[0024] Specifically, the method for constructing the characteristic chromatogram of corn silk and its pharmaceutical preparations further includes the step of preparing a reference solution and the step of constructing the characteristic chromatogram of the reference solution based on the high performance liquid chromatography method;

[0025] The reference standard includes ferulic acid;

[0026] The method for preparing the reference solution includes the step of adding the reference substance to an organic solvent and mixing.

[0027] The reference solution contains 1-20 μg of the reference substance per 1 ml.

[0028] Specifically, in the method for constructing the characteristic spectrum of corn silk and its pharmaceutical preparations, the organic solvent includes an aqueous methanol solution, preferably with a concentration of 30-80 v / v%, more preferably 50 v / v%.

[0029] Preferably, the extraction step includes reflux extraction or ultrasonic extraction;

[0030] Preferably, the solid-liquid separation includes filtration or centrifugation.

[0031] Specifically, the method for constructing the characteristic spectrum of corn silk and its pharmaceutical preparations includes corn silk formula granules, corn silk slices, corn silk standard decoction, or corn silk standard decoction freeze-dried powder.

[0032] The present invention also discloses a characteristic spectrum and / or a control characteristic spectrum of corn silk medicinal material and its pharmaceutical preparation, wherein the characteristic spectrum or control characteristic spectrum of corn silk medicinal material and its pharmaceutical preparation is constructed by the method described above;

[0033] Preferably, the characteristic chromatogram shows 13 characteristic peaks, corresponding to the retention times of 13 characteristic peaks in the reference chromatogram of the reference medicinal material. Peak 3 corresponds to the retention time of the corresponding reference peak. The peak corresponding to the retention time of the ferulic acid reference peak is peak S. The relative retention times of each characteristic peak and peak S are calculated, and these relative retention times should be within ±10% of a specified value. The specified value is:

[0034] Peak 1: 0.76, Peak 2: 0.90, Peak 4: 1.37, Peak 5: 1.75, Peak 6: 1.92, Peak 7: 2.08, Peak 8: 2.13, Peak 9: 2.19, Peak 10: 2.66, Peak 11: 3.07, Peak 12: 3.15, Peak 13: 3.51.

[0035] This invention also discloses a method for constructing characteristic spectra of the corn silk medicinal material and its pharmaceutical preparations, and / or the application of the characteristic spectra and / or control characteristic spectra of the corn silk medicinal material and its pharmaceutical preparations in the field of quality testing of corn silk medicinal material and its pharmaceutical preparations.

[0036] The present invention also discloses a quality testing method for corn silk medicinal material and its pharmaceutical preparations, including the steps of constructing the characteristic spectrum and the control characteristic spectrum according to the method, and the step of comparing the characteristic spectrum with the control characteristic spectrum.

[0037] The method for constructing characteristic chromatograms of corn silk medicinal materials and their pharmaceutical preparations, as described in this invention, uses corn silk medicinal materials and their pharmaceutical preparations as the detection targets and establishes a characteristic chromatogram method for these pharmaceutical preparations based on high performance liquid chromatography. The method for constructing characteristic chromatograms described in this invention has good separation and can simultaneously determine the contents of 4-coumaric acid, veselene-2, ferulic acid, isohyperside 2"-O-rhamnoside, kojicin-7-O-glucose-2-O-apigenin, and N-trans-feruloyltyramine. Furthermore, this method has high precision, good stability, and good repeatability, enabling comprehensive and rapid detection of the active ingredients and their contents in corn silk granules and other pharmaceutical preparations.

[0038] The method for constructing the characteristic chromatogram of corn silk medicinal material and its pharmaceutical preparation described in this invention selects ferulic acid as the internal reference peak in the fingerprint spectrum, identifies 13 common characteristic peaks using traditional Chinese medicine chromatographic fingerprint spectrum, and calculates the relative retention time of each common characteristic peak based on ferulic acid in the S peak. This method is beneficial for comprehensive quality detection and overall quality control of corn silk formula granules, thereby helping to improve the safety and stability of the drug.

[0039] The method for constructing the characteristic spectrum of corn silk medicinal material and its pharmaceutical preparations described in this invention can be applied to the construction of characteristic spectrum of pharmaceutical preparations such as corn silk slices, corn silk standard decoction freeze-dried powder, and corn silk formula granules, and can realize the quality detection and quality control of the above preparations.

[0040] This invention establishes a characteristic chromatogram (high performance liquid chromatography) method for determining corn silk formulation granules. The sample preparation method is simple, the chromatographic conditions are easy to achieve, and the analytical method verification results show that the method is simple to operate, accurate, and has good reproducibility. Attached Figure Description

[0041] To make the content of this invention easier to understand, the invention will be further described in detail below with reference to specific embodiments and accompanying drawings, wherein...

[0042] Figure 1 Chromatograms of different elution programs in Example 2;

[0043] Figure 2 These are chromatograms at different detection wavelengths in Example 2;

[0044] Figure 3 These are chromatograms of different mobile phase systems in Example 2;

[0045] Figure 4 The chromatograms are shown at different column temperatures in Example 2.

[0046] Figure 5 The chromatograms are shown at different flow rates in Example 2.

[0047] Figure 6 The chromatograms are shown under different chromatographic columns in Example 2.

[0048] Figure 7 Chromatograms of different extraction solvents in Example 3;

[0049] Figure 8 Chromatograms of different extraction methods in Example 3;

[0050] Figure 9 Chromatograms of different extraction times in Example 3;

[0051] Figure 10 The chromatograms are shown for different extraction powers in Example 3.

[0052] Figure 11 The chromatograms are for different extraction solvent amounts in Example 3;

[0053] Figure 12 The chromatogram of ferulic acid reference standard in Example 4 is shown below.

[0054] Figure 13 The chromatogram of the corn silk formulation granules test sample in Example 4 is shown below.

[0055] Figure 14 This is a characteristic chromatogram of corn silk as a control medicinal material in Example 5;

[0056] Figure 15 Chromatograms of different reference standards in Example 5;

[0057] Figure 16 This is a comparative chromatogram of the characteristics of the corn silk formulation granules in Example 5;

[0058] Figure 17 This is the negative blank chromatogram from Example 6;

[0059] Figure 18 The chromatograms are shown under different chromatographic columns in Example 6.

[0060] Figure 19 These are chromatograms from different liquid chromatography devices used in Example 6;

[0061] Figure 20 The image shows the characteristic spectrum of 20 batches of corn silk formulation granules in Example 7. Detailed Implementation

[0062] In the following embodiments of the present invention, the instruments and reagents involved include:

[0063] High Performance Liquid Chromatography System 1: Waters ACQUITY UPLC I-Class Chromatography System, including a quaternary solvent manager (ACQ-QSM), an autosampler (ACQ-FTN), an original imported column oven (ACQ-CM), a diode array UV detector (ACQ-TUV), and an Empower chromatography management system;

[0064] High Performance Liquid Chromatograph 2: Thermo Fisher Vanquish Chromatography System, including LPG-3400A quaternary pump, WPS-3000TSL autosampler, PDA diode array detector, and chromatography workstation;

[0065] One ten-thousandth electronic balance (Shanghai Shunyu Hengping Scientific Instruments Co., Ltd., FA1004);

[0066] Electronic balance with a range of 1 / 100,000 (Sartorius Scientific Instruments Ltd., SQP);

[0067] Ultrasonic Cleaner: WB400US, Shanghai Wangbiao Instrument Co., Ltd.

[0068] Column 1: Shim-pack GIST-HP C18-AQ (2.1×100mm, 1.9μm);

[0069] Column 2: ACQUITY UPLC HSS T3 (2.1×100mm, 1.8μm);

[0070] Column 3: Poroshell 120SB-C18 (2.1×100mm, 1.9μm);

[0071] 4-Coumaric acid reference standard (batch number: 112037-202102, provided by China National Institutes for Food and Drug Control);

[0072] Viclin-2 reference standard (batch number: 112096-202201, provided by the National Institutes for Food and Drug Control);

[0073] Ferulic acid reference standard (batch number: 110773-202316, provided by China National Institutes for Food and Drug Control);

[0074] Isorhizonidine 2"-O-rhamnoside reference standard (batch number: 005375-202402, provided by Jiangxi Baicaoyuan Biotechnology Co., Ltd.);

[0075] Koilin-7-O-glucose-2-O-apiosinoside: (Batch No.: 110085-202407, provided by Shanghai Hongyong Biotechnology Co., Ltd.);

[0076] N-trans-feruloyltyramine (batch number: 001151-202201, provided by Jiangxi Baicaoyuan Biotechnology Co., Ltd.);

[0077] Corn silk reference material (batch number: 121659-202103, provided by China National Institutes for Food and Drug Control);

[0078] Acetonitrile was of chromatographic grade, and water was ultrapure water; all other reagents were of analytical grade.

[0079] In the following embodiments of the present invention, the corn silk pharmaceutical preparation can be prepared according to processes known in the art, or it can be tested based on commercially available products.

[0080] As an exemplary preparation method, the corn silk pharmaceutical preparation can be prepared by the following method: Corn silk is extracted by heating and reflux at least once, with 10-15 times its weight of water added each time for at least 0.5 hours. The mixture is then filtered, the filtrates are combined, and the filtrate is concentrated to a relative density of 1.05-1.10 g / mL at 60°C. Conventional excipients are added, and the preparation is carried out according to conventional processes to produce clinically acceptable tablets, capsules, pills, granules, honey-processed pills, sustained-release preparations, immediate-release preparations, controlled-release preparations, oral liquid preparations, or injectable preparations. For example, the pharmaceutically acceptable excipients include: fillers, disintegrants, lubricants, suspending agents, binders, sweeteners, flavoring agents, preservatives, and matrices. Fillers include: starch, pregelatinized starch, lactose, mannitol, chitosan, microcrystalline cellulose, sucrose, etc.; disintegrants include: starch, pregelatinized starch, microcrystalline cellulose, sodium carboxymethyl starch, croscarmellose, low-substituted hydroxypropyl cellulose, croscarmellose sodium, etc.; lubricants include: magnesium stearate, sodium lauryl sulfate, talc, silica, etc.; suspending agents include: polyvinylpyrrolidone, microcrystalline cellulose, sucrose, agar, hydroxypropyl methylcellulose, etc.; binders include: starch paste, polyvinylpyrrolidone, hydroxypropyl methylcellulose, etc.; sweeteners include: sodium saccharin, aspartame, sucrose, cyclamate, glycyrrhetinic acid, etc.; flavoring agents include: sweeteners and various flavorings; preservatives include: parabens, benzoic acid, sodium benzoate, sorbic acid and its salts, benzalkonium bromide, chlorethidium acetate, eucalyptus oil, etc.; matrix includes: PEG6000, PEG4000, insect wax, etc.

[0081] As an exemplary embodiment, the test sample selected in the following examples of the present invention includes corn silk formulation granules. The specific preparation method of the corn silk formulation granules can be as follows: take corn silk, heat and reflux extract twice. For the first extraction, add 15 times the weight of water and soak for 30 min, heat and reflux extract for 0.5 h, filter, add 12 times the weight of water for the second extraction for 0.5 h, filter, combine the filtrates, concentrate the filtrate to a relative density of 1.05 g / mL at 60°C, spray dry, add maltodextrin as an excipient to the dry powder, mix evenly, and then dry granulate to make granules.

[0082] In the following embodiments of the present invention, information on corn silk formulation granules is shown in Table 1 below.

[0083] Table 1. Information on Corn Silk Granule Formula

[0084] batch number Origin 1903001W Guangyuan City, Sichuan Province 1906001S Guangyuan City, Sichuan Province 1909001W Guangyuan City, Sichuan Province 1912001W Shangluo City, Shaanxi Province 2001001S Shangluo City, Shaanxi Province 2002001W Shangluo City, Shaanxi Province 2005001S Xia Village, Jiefang Village, Qiubei County, Yunnan Province 2008001W Pojiao Town, Maguan County, Wenshan City, Yunnan Province 2011001S Xiangyang District, Xiangyang City, Hubei Province 2102001S Xiangyang District, Xiangyang City, Hubei Province 2105001S Xiangyang District, Xiangyang City, Hubei Province 2108001W Zhecheng County, Shangqiu City, Henan Province 2110001S Zhecheng County, Shangqiu City, Henan Province 2112001S Zhecheng County, Shangqiu City, Henan Province 2202001W Anqiu City, Weifang City, Shandong Province 2206001W Anqiu City, Weifang City, Shandong Province 2212001S Anqiu City, Weifang City, Shandong Province 2309001W Anqiu City, Weifang City, Shandong Province 2311001S Anqiu City, Weifang City, Shandong Province 2312001W Anqiu City, Weifang City, Shandong Province

[0085] Example 1

[0086] This embodiment constructs fingerprint spectra of flavonoids from corn silk and its pharmaceutical preparations.

[0087] Preparation of the test solution: Take about 0.5g of corn silk formula granule powder, place it in a stoppered conical flask, accurately add 10ml of 50% methanol, sonicate (power 250W, frequency 40kHz) for 30 minutes, remove, cool, shake well, filter, and collect the filtrate to obtain the test solution.

[0088] Preparation of reference solution: Take 4g of corn silk reference material, place it in a stoppered conical flask, add 100ml of water, heat under reflux for 1 hour, filter, evaporate the filtrate to dryness, add 10ml of 50% methanol to the residue, sonicate (power 250W, frequency 40kHz) for 30 minutes, remove, cool, shake well, filter, and take the filtrate as the reference solution of the reference material.

[0089] Take an appropriate amount of ferulic acid reference standard and add methanol to prepare a solution containing 10 μg per 1 ml, which will be used as the reference solution.

[0090] Example 2

[0091] This embodiment establishes chromatographic conditions based on different conditions.

[0092] (1) Gradient selection

[0093] In this embodiment, octadecylsilane-bonded silica gel was used as the packing material under different chromatographic conditions; acetonitrile was used as mobile phase A, and 0.1% formic acid solution was used as mobile phase B. Gradient elution was performed according to the specifications in Table 2 below; the flow rate was 0.3 ml per minute; the column temperature was 25°C; and the detection wavelength was 330 nm.

[0094] Table 2 Gradient Procedure

[0095]

[0096] In this embodiment, the spectra formed under different gradient elution programs (gradient 1 to gradient 3) are shown in the attached figures. Figure 1 As shown in (a)-(c), the system adaptability parameters under each condition are shown in Table 3 below.

[0097] Table 3 System Adaptability Parameters

[0098]

[0099]

[0100] It is evident that the characteristic peaks under gradient 3 have more information and better separation, so we tentatively choose the mobile phase gradient elution program as condition 3.

[0101] (2) Selection of detection wavelength

[0102] The test solution of corn silk formula granules (batch number: 2202001W) was scanned at all wavelengths. The results showed that there was a lot of characteristic peak information at four different absorption wavelengths (270nm, 290nm, 310nm, and 330nm). Therefore, the chromatograms of the four wavelengths and the system suitability parameters were further evaluated.

[0103] In this embodiment, the chromatograms at different detection wavelengths (270nm, 290nm, 310nm, 330nm) are shown in the appendix. Figure 2 As shown in (a)-(d), the system adaptability parameters are shown in Table 4 below.

[0104] Table 4 System Adaptability Parameters

[0105]

[0106]

[0107]

[0108] The results showed that the peak responses of each characteristic peak were larger and the system adaptability parameters were relatively better when the wavelength was 330 nm. Therefore, the wavelength of 330 nm was determined as the detection wavelength of the characteristic spectrum of corn silk formulation particles.

[0109] (3) Selection of mobile phase

[0110] In this embodiment, the separation effects of three mobile phase systems—acetonitrile-0.1% formic acid, acetonitrile-0.1% acetic acid, and acetonitrile-0.1% phosphoric acid solution—on the characteristic chromatograms of the corn silk formulation granule test solution were compared.

[0111] In this embodiment, the chromatograms for different mobile phases (acetonitrile-0.1% formic acid, acetonitrile-0.1% acetic acid, acetonitrile-0.1% phosphoric acid solution) are shown in the appendix. Figure 3 As shown in (a)-(c).

[0112] The results showed that the acetonitrile-0.1% formic acid solution group had better separation performance and richer peak information in the mobile phase system. Therefore, the acetonitrile-0.1% formic acid solution was selected as the mobile phase of the system.

[0113] (4) Selection of different column temperatures

[0114] This embodiment compares the effects of different column temperatures on the robustness of the characteristic chromatograms of corn silk formulation granules. Column temperatures were set at 20℃, 25℃, and 30℃. An appropriate amount of corn silk formulation granules (batch number: 2202001W) was taken, and the effect of different column temperatures on the separation effect of the corn silk formulation granule sample was observed.

[0115] In this embodiment, the chromatograms at different column temperatures (20℃, 25℃, 30℃) are shown in the appendix. Figure 4 As shown in (a)-(c), the results of the investigation at different column temperatures are shown in Table 5 below.

[0116] Table 5 Results of investigation at different column temperatures

[0117]

[0118]

[0119]

[0120] The results showed that the separation of each chromatographic peak was relatively better at a column temperature of 25℃, so the column temperature was tentatively set at 25℃.

[0121] (5) Selection of different flow velocities

[0122] This embodiment compares the effects of different flow rates on the robustness of the characteristic chromatograms of corn silk formulation granules. The flow rates were set at 0.2 mL / min, 0.25 mL / min, and 0.3 mL / min. An appropriate amount of corn silk formulation granules (batch number: 2202001W) was taken and measured according to the method determined above to observe the effect of different flow rates on the separation effect of the corn silk formulation granule test sample.

[0123] In this embodiment, the chromatograms at different flow rates (0.2 mL / min, 0.25 mL / min, and 0.3 mL / min) are shown in the appendix. Figure 5 As shown in (a)-(c), the results of the investigation at different flow rates are shown in Table 6 below.

[0124] Table 6 Results of investigation at different flow velocities

[0125]

[0126]

[0127] The results showed that the separation of each chromatographic peak was relatively good when the flow rate was 0.3 ml / min, so the flow rate was tentatively set at 0.3 ml / min.

[0128] (6) Selection of different chromatographic columns

[0129] Take the same sample solution of corn silk granules (2202001W) and measure the characteristic chromatograms on different types of chromatographic columns (Column 1: Shim-pack GIST-HP C18-AQ, 2.1×100mm, 1.9μm; Column 2: ACQUITY UPLC HSS T3, 2.1×100mm, 1.8μm; Column 3: Poroshell 120SB-C18, 2.1×100mm, 1.9μm). Compare the chromatograms and relative retention times obtained from different types of chromatographic columns.

[0130] In this embodiment, the chromatograms under different chromatographic columns (column 1, column 2, and column 3) are shown in the appendix. Figure 6 The results of the investigation of different brands of chromatographic columns are shown in Table 7 below.

[0131] Table 7 Results of chromatographic column analysis for different brands

[0132]

[0133]

[0134]

[0135] The results showed that the Poroshell 120SB-C18 column had better separation performance. There were significant differences in the relative retention times between different brands of columns. Therefore, the Poroshell 120SB-C18 column with a diameter of 2.1*100mm and a diameter of 1.9μm was selected.

[0136] Chromatographic conditions determined

[0137] In summary, the chromatographic conditions and system suitability tests confirmed in this embodiment are as follows: Octadecylsilane-bonded silica gel was used as the packing material (column length 100 mm, inner diameter 2.1 mm, particle size 1.9 μm); acetonitrile was used as mobile phase A; 0.1% formic acid solution was used as mobile phase B; gradient elution was performed according to the procedure specified in Table 8; the column temperature was 25°C, the flow rate was 0.3 mL / min, and the detection wavelength was 330 nm. The theoretical plate number, calculated based on the ferulic acid peak, should not be less than 5000.

[0138] Table 8 Elution Procedure

[0139] Time (minutes) Mobile phase A (%) Mobile phase B (%) 0～10 7→10 93→90 10～22 10→12 90→88 22～40 12→14 88→86 40～45 14 86 45～60 14→16 86→84 60～65 16 84

[0140] Accurately pipette 3 μl of the reference solution and the test solution into the liquid chromatograph and measure them to obtain the final result.

[0141] Example 3

[0142] This embodiment further examines the preparation method of the test solution based on the chromatographic conditions determined in Example 2 above.

[0143] (1) Selection of extraction solvent

[0144] This embodiment refers to the test sample solution preparation method in Example 1 above, and investigates the extraction effect of different extraction solvents, methanol, 50% methanol and water, on corn silk formulation particles. By comparing the chromatograms of the test samples obtained with different solvents and the system adaptability parameters, a suitable extraction solvent is selected.

[0145] In this embodiment, the chromatograms for different extraction solvents (water, 50% methanol, methanol) are shown in the appendix. Figure 7 The results of the investigation under different solvents are shown in Table 9 below.

[0146] Table 9 Results of the investigation of different extraction methods

[0147]

[0148]

[0149]

[0150] The results showed that the system adaptability parameters of 50% methanol extraction of the test sample were relatively better, so 50% methanol was selected as the extraction solvent.

[0151] (2) Selection of extraction method

[0152] This embodiment refers to the test sample solution preparation method in Example 1, and investigates the extraction effects of ultrasound and reflux on corn silk formulation particles. By comparing the chromatograms of the test samples obtained by different extraction methods and the system adaptability parameters, a suitable extraction method is selected.

[0153] In this embodiment, the chromatograms for different extraction methods (reflux, ultrasound) are shown in the appendix. Figure 8 The results of the investigation under different extraction methods are shown in Table 10 below.

[0154] Table 10 Results of the investigation of different extraction methods

[0155]

[0156]

[0157] The results showed that both ultrasonic and reflux extraction systems had good adaptability parameters. Considering the ease of operation of ultrasonic extraction, ultrasonic extraction was chosen as the extraction method.

[0158] (3) Selection of extraction time

[0159] This embodiment refers to the test sample solution preparation method in Example 1, and investigates the extraction effect of different extraction times (15 minutes, 30 minutes, and 45 minutes) on corn silk formula granules. By comparing the chromatograms of the test samples obtained at different extraction times and the system adaptability parameters, a suitable extraction time is selected.

[0160] In this embodiment, the chromatograms at different extraction times (15 minutes, 30 minutes, and 45 minutes) are shown in the appendix. Figure 9 The results of the investigation at different extraction times are shown in Table 11 below.

[0161] Table 11 Results of the investigation at different extraction times

[0162]

[0163]

[0164] The results showed that the system adaptability parameters of the test sample were relatively better when the extraction time was 30 minutes, so 30 minutes was selected as the extraction time.

[0165] (4) Selection of extraction power

[0166] This embodiment refers to the test sample solution preparation method in Example 1, and investigates the extraction effect of different ultrasonic extraction powers (100W, 250W, 500W) on corn silk formula granules. By comparing the chromatograms of the test samples obtained by different ultrasonic extraction powers and the system adaptability parameters, a suitable ultrasonic extraction power is selected.

[0167] In this embodiment, the chromatograms at different extraction powers (100W, 250W, 500W) are shown in the appendix. Figure 10 The results of the investigation under different extraction powers are shown in Table 12 below.

[0168] Table 12 Results of different extraction powers

[0169]

[0170]

[0171] The results showed that the ultrasonic extraction power of 250W was relatively better than that of the sample extraction system, so 250W was selected as the ultrasonic extraction power.

[0172] (5) Selection of extraction solvent amount

[0173] This embodiment refers to the test sample solution preparation method in Example 1, and investigates the extraction effect of different extraction solvent amounts (10ml, 25ml, 50ml) on the lyophilized powder. By comparing the chromatograms of the test samples obtained with different extraction solvent amounts and the system adaptability parameters, an appropriate sampling amount is selected.

[0174] In this embodiment, the chromatograms for different extraction solvent volumes (10 ml, 25 ml, 50 ml) are shown in the appendix. Figure 11 The results of the investigation under different amounts of extraction solvent are shown in Table 13 below.

[0175] Table 13 Results of the investigation of different extraction solvent amounts

[0176]

[0177]

[0178]

[0179] The results showed that the system adaptability parameters of the test sample were relatively better when the extraction solvent volume was 10 ml. Therefore, 10 ml was selected as the extraction solvent volume for different extraction methods.

[0180] Validation of test sample preparation method

[0181] In summary, the preparation method of the optimized corn silk formula granule test solution of the present invention is as follows: Take about 0.5g of the powder of this product, place it in a stoppered conical flask, accurately add 10ml of 50% methanol, sonicate (power 250W, frequency 40kHz) for 30 minutes, take it out, cool it, shake it well, filter it, and take the filtrate to obtain the product.

[0182] Example 4

[0183] This embodiment constructs fingerprint spectra of flavonoid compounds from corn silk and its pharmaceutical preparations based on the aforementioned determined conditions.

[0184] Preparation of the test solution: Take about 0.5g of corn silk formula granule powder, place it in a stoppered conical flask, accurately add 10ml of 50% methanol, sonicate (power 250W, frequency 40kHz) for 30 minutes, remove, cool, shake well, filter, and collect the filtrate to obtain the test solution.

[0185] Preparation of reference solution: Take 4g of corn silk reference material, place it in a stoppered conical flask, add 100ml of water, heat under reflux for 1 hour, filter, evaporate the filtrate to dryness, add 10ml of 50% methanol to the residue, sonicate (power 250W, frequency 40kHz) for 30 minutes, remove, cool, shake well, filter, and take the filtrate as the reference solution of the reference material.

[0186] Take an appropriate amount of ferulic acid reference standard and add methanol to prepare a solution containing 10 μg per 1 ml, which will be used as the reference solution.

[0187] High Performance Liquid Chromatography (UPLC) Analysis Conditions

[0188] Chromatographic column: Octadecylsilane-bonded silica gel was used as the packing material (column length 100 mm, inner diameter 2.1 mm, particle size 1.9 μm). Acetonitrile was used as mobile phase A, and 0.1% formic acid solution was used as mobile phase B. Gradient elution was performed according to the specifications in Table 8 above. The column temperature was 25℃, the flow rate was 0.3 mL / min, and the detection wavelength was 330 nm. The theoretical plate number, calculated based on the ferulic acid peak, should be no less than 5000.

[0189] Accurately pipette 3 μl of the reference solution and the test solution into the liquid chromatograph and measure them to obtain the final result.

[0190] This embodiment, based on the aforementioned method, uses the fingerprint chromatogram similarity evaluation software "Traditional Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System 2012 Edition" compiled by the Pharmacopoeia Commission to generate a chromatogram of ferulic acid reference standard, i.e., a reference characteristic fingerprint chromatogram, as shown below. Figure 12 The chromatogram of the corn silk formulation granule test sample is shown in the attached figure. Figure 13 As shown. The detection results of the characteristic fingerprint spectrum of the formulated granules can be analyzed and compared based on the aforementioned characteristic fingerprint spectrum, which is used for the quality control of the formulated granules.

[0191] In this embodiment, the results of the system's adaptability parameters are shown in Table 14 below.

[0192] Table 14 System Adaptability Parameters

[0193]

[0194] Selection of main chemical components in characteristic spectra

[0195] According to existing literature, the main active components of corn silk are flavonoids, and it also contains polysaccharides, organic acids, sterols, and amino acids. Corn silk is effective in promoting diuresis and reducing swelling, and lowering blood pressure. It is used for nephritis edema, difficulty urinating, damp-heat jaundice, and hypertension. Current research mainly focuses on total flavonoids, polysaccharides, and organic acids in corn silk. Analysis of the flavonoid components in corn silk formulation granules revealed the presence of 4-coumaric acid, ferulic acid, vincain-2, isopropanol 2”-O-rhamnoside, kojicin-7-O-glucose-2-O-apigenin, and N-trans-feruloyltyramine. 4-coumaric acid and ferulic acid showed high peak responses and good separation. Therefore, the main active components, flavonoids, were selected as the primary components for developing the characteristic chromatogram of corn silk formulation granules.

[0196] As can be seen from the above characteristic chromatograms, the UPLC characteristic fingerprint of corn silk formulation granules contains 13 chromatographic peaks, of which 6 peaks (peaks 1, 2, 3, 4, 11, and 13) are known component peaks. The peak corresponding to the ferulic acid reference peak is selected as the S peak. The relative retention times of each characteristic peak and the S peak are calculated, and the relative retention times should be within ±10% of the specified values. The specified values ​​are: 0.76 (peak 1), 0.90 (peak 2), 1.37 (peak 4), 1.75 (peak 5), 1.92 (peak 6), 2.08 (peak 7), 2.13 (peak 8), 2.19 (peak 9), 2.66 (peak 10), 3.07 (peak 11), 3.15 (peak 12), and 3.51 (peak 13).

[0197] As can be seen, the detection method in this embodiment can effectively obtain fingerprint spectra with good separation of each characteristic peak, and can simultaneously determine the contents of 4-coumaric acid, vezin-2, ferulic acid, isohyperidin 2"-O-rhamnoside, kojicin-7-O-glucose-2-O-apiside, and N-trans-feruloyltyramine. Furthermore, by selecting ferulic acid as the internal reference peak in the fingerprint spectrum, the relative retention times of the common characteristic peaks 1, 2, and peaks 4-13 of the corn silk formulation granules can be determined. Therefore, it can comprehensively and rapidly detect corn silk formulation granules, which is beneficial for comprehensive quality testing and overall quality control of corn silk formulation granules, thereby helping to improve the safety and stability of the drug.

[0198] Example 5

[0199] Characteristic peak identification and character peak assignment in the characteristic spectrum

[0200] This embodiment is based on the aforementioned method. It analyzes the detection results of fingerprint spectra of multiple batches of test samples and uses the fingerprint spectra similarity evaluation software "Traditional Chinese Medicine Chromatographic Fingerprint Spectra Similarity Evaluation System 2012 Edition" compiled by the Pharmacopoeia Commission to generate a reference feature spectrum.

[0201] In this embodiment, through the identification and assignment of characteristic peaks, the UPLC characteristic chromatogram of the corn silk formulation granules yielded 13 chromatographic peaks. The remaining chromatographic peaks were either low in response or had poor resolution. Therefore, only the 13 peaks with the largest responses were selected as characteristic peaks, and the peaks were rearranged according to their order. The characteristic chromatogram of the corn silk reference medicinal material is attached. Figure 14 .

[0202] Based on the peak identification and reference standard localization results, six known peaks were identified, including 4-coumaric acid (peak 1), virzalin-2 (peak 2), ferulic acid (peak 3), isoharonidine 2”-O-rhamnoside (peak 4), koilin-7-O-glucose-2-O-apiside (peak 11), and N-trans-feruloyltyramine (peak 13). These peaks are flavonoids and organic acids, and the characteristic spectrum contains the main chemical components of corn silk.

[0203] Selection criteria for characteristic peak S peak in characteristic spectrum

[0204] Based on the peak identification and selection results of characteristic peaks in corn silk formulation granules, flavonoids showed a low response in the characteristic spectrum of corn silk formulation granules, while ferulic acid showed a high response and a moderate peak time. It belongs to organic acid components and is one of the main active ingredients of corn silk. The peak corresponding to the ferulic acid reference standard is temporarily designated as the S peak, and the relative retention times of peaks 1-2 and peaks 4-13 are calculated.

[0205] Confirmation of characteristic peaks in the characteristic spectrum

[0206] UPLC localization studies were conducted using reference standards on corn silk formulation granules (as shown in the attached figure). Figure 13 In the analysis, peaks 1-14 were analyzed for molecular weight and molecular structure by LC / MS / MS to provide a reference for the subsequent qualitative identification of characteristic component peaks. Four peaks were identified as known chromatographic peaks using reference standards: peak 1 (4-coumaric acid), peak 2 (virzalin-2), peak 3 (ferulic acid), peak 4 (isosericin 2”-O-rhamnoside), peak 11 (cuilelin-7-O-glucose-2-O-apigenin), and peak 13 (N-trans-feruloyltyramine). The structures analyzed by LC / MS / MS are shown in Table 15 below.

[0207] The chromatograms of 4-coumaric acid, virzinin-2, ferulic acid, isoharonidin 2”-O-rhamnoside, koilin-7-O-glucose-2-O-apiside, and N-trans-feruloyltyramine are shown in the appendix. Figure 15 As shown in (a)-(f).

[0208] Table 15 LC / MS / MS Analysis Results of Corn Silk Granules

[0209]

[0210]

[0211] Characteristic peak values ​​of characteristic spectrum

[0212] The relative retention times of the above characteristic spectra were determined based on the research results: the characteristic spectra of corn silk formula granules should show 13 characteristic peaks, which should correspond to the retention times of the corresponding characteristic peaks of the control medicinal materials. The results are shown in Table 16-17 below.

[0213] In this embodiment, the reference chart of the corn silk formula granules is attached. Figure 16 As shown, peak 1: 4-coumaric acid; peak 2: virzidin-2; peak 3(S): ferulic acid; peak 4: isoharrenin 2"-O-rhamnoside; peak 11: kojicin-7-O-glucose-2-O-apiside; peak 13: N-trans-feruloyltyramine. Column: Agilent Poroshell 120SB-C18, 2.1*100mm, 1.9μm.

[0214] Table 16. Comparison chart of corn silk formulation granules and their relative retention times.

[0215] peak 1 2 3(S) 4 5 6 7 Retention time 12.752 15.203 16.874 23.081 29.548 32.436 35.107 Relative retention time 0.76 0.90 1.00 1.37 1.75 1.92 2.08 peak 8 9 10 11 12 13 Retention time 35.951 36.932 44.817 51.751 53.149 59.245 Relative retention time 2.13 2.19 2.66 3.07 3.15 3.51

[0216] Table 17. Comparison of relative peak areas in corn silk formulation granules.

[0217] peak 1 2 3(S) 4 5 6 7 Peak area 494.286 80.432 899.421 43.711 602.122 244.615 146.996 relative peak area 0.55 0.09 1.00 0.05 0.67 0.27 0.16 peak 8 9 10 11 12 13 Peak area 69.774 94.547 94.808 75.97 253.849 33.219 relative peak area 0.08 0.11 0.11 0.08 0.28 0.04

[0218] Example 6

[0219] This embodiment examines the feature map methodology based on the aforementioned construction method.

[0220] (1) Verification of specificity, integrity and system suitability

[0221] The test solution and negative blank solution were prepared according to the aforementioned method. UPLC analysis was performed using the characteristic chromatogram method for corn silk formula granules to verify whether the chromatographic conditions and system suitability were suitable for corn silk formula granules, and to investigate whether negative samples of corn silk formula granules would cause interference. The results are shown in the appendix. Figure 17 .

[0222] The results show that the negative blank control did not interfere with the characteristic chromatogram, and the chromatographic method has good system adaptability and specificity, and can be used for the detection of characteristic chromatograms of corn silk formulation granules.

[0223] (2) Instrument precision

[0224] Take the corn silk formula granules (2202001W) test solution and inject it 6 times according to the method under the characteristic spectrum section. The RSD of the relative retention time of the characteristic peak should be less than 2.0%. The results are shown in Table 18-19 below.

[0225] Table 18 Results of Relative Retention Time in Instrument Precision Test

[0226]

[0227] Table 19 Results of relative peak area in instrument precision test

[0228]

[0229]

[0230] The results show that the instrument has good precision.

[0231] (3) Repeatability

[0232] Take a sample of corn silk formula granules (batch number: 2202001W), prepare 6 test solutions in parallel according to the method under the characteristic spectrum section, and determine them. The RSD of the relative retention time of each characteristic peak should be less than 2.0%. The results are shown in Table 20-21 below.

[0233] Table 20 Results of Relative Retention Time in Repeatability Tests

[0234]

[0235] Table 21 Results of Relative Peak Area in Repeatability Tests

[0236]

[0237]

[0238] The results show that the method has good reproducibility.

[0239] (4) Intermediate precision (personnel)

[0240] Three researchers prepared corn silk formula granules (2202001W) test solution according to the characteristic spectrum method and measured it. The RSD of the relative retention time of each characteristic peak should be less than 2.0%. The results are shown in Table 22-23 below.

[0241] Table 22 Intermediate Precision (Personnel) Relative Retention Time

[0242]

[0243] Table 23 Intermediate Precision (Personnel) Relative Peak Area

[0244]

[0245] The results showed that the intermediate precision was good.

[0246] (5) Durability-stability assessment

[0247] Take the test solution of corn silk formula granules (2202001W) and determine it according to the method under the characteristic spectrum at 0h, 4h, 8h, 12h, 18h and 24h respectively. The RSD of the relative retention time of each characteristic peak should be less than 2.0%. The results are shown in Table 24-25 below.

[0248] Table 24 Results of relative retention times in stability tests

[0249]

[0250] Table 25 Results of relative peak area in stability test

[0251]

[0252] The results showed that the test solution was stable within 24 hours, meeting the requirements for the assay.

[0253] (6) Durability - Test at different column temperatures

[0254] The effects of different column temperatures on the characteristic chromatograms of corn silk formulation granules (2202001W) were compared. Column temperatures were set at 24℃, 25℃, and 26℃. An appropriate amount of corn silk formulation granules was taken and measured according to the method determined above. The effect of different column temperatures on the separation effect of the corn silk formulation granules was observed. The results are shown in Tables 26-27 below.

[0255] Table 26 Results of relative retention times at different column temperatures

[0256]

[0257] Table 27 Results of relative peak areas at different column temperatures

[0258]

[0259] The results show that small fluctuations in column temperature have little effect on the specified values ​​of the relative retention times of each characteristic peak, indicating that the method has good robustness to small temperature fluctuations.

[0260] (7) Durability - Test at different flow rates

[0261] The effect of different flow rates on the robustness of the characteristic spectrum of corn silk formulation granules (2202001W) was compared. The flow rates were set at 0.29 mL / min, 0.30 mL / min, and 0.31 mL / min. An appropriate amount of corn silk formulation granules was taken and measured according to the method determined above. The effect of different flow rates on the separation effect of the corn silk formulation granules sample was observed. The results are shown in Tables 28-29 below.

[0262] Table 28 Results of relative retention time for different flow velocities

[0263]

[0264] Table 29 Results of relative peak area at different flow velocities

[0265]

[0266] The results show that small fluctuations in flow velocity have little impact on the specified values ​​of the relative retention time of each characteristic peak, indicating that the method has good durability for small fluctuations in flow velocity.

[0267] (8) Durability - Examination of different batches of columns

[0268] The same corn silk granule formulation (2202001W) test solution was analyzed using different batches of the same type of chromatographic column (brand and model: Agilent Poroshell 120SB-C18 (100mm×2.1mm, 1.9μm; column 1: S / N: USKEC01239; column 2: S / N: USKEC01312; column 3: S / N: USKEC01527) manufactured by the same company. The results are shown in Tables 30-31 below. The chromatograms under different columns (columns 1, 2, and 3) are shown in the appendix. Figure 18 As shown in (a)-(c).

[0269] Table 30 Results of relative retention times for different brands of chromatographic columns

[0270]

[0271] Table 31 Results of the determination of the specified values ​​of relative peak area for different brands of chromatographic columns

[0272]

[0273] The results showed that the specified values ​​of the relative retention times of each characteristic peak of the chromatographic columns from different batches of this manufacturer varied little, and the method had good durability for different batches of this model of chromatographic columns.

[0274] (9) Durability - Examination of different liquid phase equipment

[0275] The effects of different instrument brands on the robustness of the characteristic chromatogram of corn silk formulation granules (2202001W) were compared. Instrument brands included Waters Arc and Thermo Fisher Vanquish. An appropriate amount of corn silk formulation granules was taken and analyzed according to the method determined above. The effect of different instruments on the separation effect of the corn silk formulation granule sample was observed. The results are shown in Tables 32-33 below. Chromatograms under different liquid chromatography equipment (Waters Arc and Thermo Fisher Vanquish) are shown in the appendix. Figure 19 As shown in (a)-(b).

[0276] Table 32 Relative retention times of instruments from different brands

[0277]

[0278] Table 33 Relative peak area results for instruments of different brands

[0279]

[0280] The results show that different brands of liquid have little impact on the specified values ​​of the relative retention time of each characteristic peak.

[0281] In summary, the results of the methodological investigation in this embodiment show that among the 13 common peaks of the corn silk formulation granules [characteristic chromatogram] established by this invention, different high-performance liquid chromatography, column temperature, flow rate, chromatographic column and other factors have a certain degree of influence on each characteristic chromatogram. The relative retention time values ​​are within ±10%. In order to adapt to its robustness, it is recommended to control the specified value range within ±10%. The other chromatographic conditions have little effect.

[0282] Example 7

[0283] Based on the aforementioned method, this embodiment measures the characteristic spectra of 20 batches of corn silk formulation granules.

[0284] Twenty batches of corn silk formula granules were tested according to the proposed method for determining the characteristic chromatograms of corn silk formula granules. The results are shown in Tables 34-37 below. The characteristic chromatograms of the 20 batches of corn silk formula granules are attached. Figure 20 Among them, S1~20: 1903001W, 1906001S, 1909001W, 1912001W, 2001001S, 2002001W, 2005001S, 2008001W, 2011001S, 2102001S, 2105001S, 2108001W, 2110001S, 2112001S, 2202001W, 2206001W, 2212001S, 2309001W, 2311001S, 2312001W; R: reference spectrum.

[0285] Table 34. Results of relative retention time of 20 batches of corn silk formulation granules

[0286]

[0287]

[0288] Table 35. Results of relative peak area of ​​20 batches of corn silk formulation granules

[0289]

[0290] Table 36. Similarity Results of 20 Batches of Corn Silk Formula Granules

[0291] batch number Similarity batch number Similarity batch number Similarity R 1.000 2005001S 0.907 2112001S 0.989 1903001W 0.988 2008001W 0.981 2202001W 0.950 1906001S 0.959 2011001S 0.983 2206001W 0.961 1909001W 0.960 2102001S 0.919 2212001S 0.978 1912001W 0.980 2105001S 0.987 2309001W 0.984 2001001S 0.993 2108001W 0.990 2311001S 0.984 2002001W 0.902 2110001S 0.994 2312001W 0.983

[0292] Table 37 Common Pattern Matching Data for Corn Silk Formulation Granules

[0293]

[0294] As can be seen, the test results show that the characteristic chromatograms of 20 batches of corn silk formula granules have 13 characteristic peaks, which correspond to the 13 characteristic peaks in the chromatogram of the reference medicinal material. The relative retention times of each characteristic peak and the S peak are within ±10% of the specified value.

[0295] It is evident that the construction method of this invention meets the requirements.

[0296] In summary, the detection method described in this embodiment can effectively obtain fingerprint spectra with good separation of various characteristic peaks. It can also simultaneously determine the contents of 4-coumaric acid, vezin-2, ferulic acid, isohyperidin 2"-O-rhamnoside, kojicin-7-O-glucose-2-O-apiside, and N-trans-feruloyltyramine. Furthermore, by selecting ferulic acid as the internal reference peak in the fingerprint spectrum, the relative retention times of the common characteristic peaks 1, 2, and peaks 4-13 of the corn silk formulation granules can be determined. Therefore, it can comprehensively and rapidly detect corn silk formulation granules, which is beneficial for comprehensive quality testing and overall quality control of corn silk formulation granules, thereby helping to improve the safety and stability of the drug.

[0297] Obviously, the above embodiments are merely illustrative examples for clear explanation and are not intended to limit the implementation. Those skilled in the art will recognize that other variations or modifications can be made based on the above description. It is neither necessary nor possible to exhaustively list all possible implementations here. However, obvious variations or modifications derived therefrom are still within the scope of protection of this invention.

Claims

1. A method for constructing a characteristic spectrum of corn silk and its pharmaceutical preparations, characterized in that, This includes the step of performing high-performance liquid chromatography (HPLC) detection on the test solution of corn silk medicinal material and / or corn silk pharmaceutical preparation; Chromatographic conditions included: octadecylsilane-bonded silica gel as the stationary phase, acetonitrile as mobile phase A, 0.1% formic acid solution as mobile phase B, and a detection wavelength of 330 nm; gradient elution was performed according to the following procedure: From 0 to 10 minutes, the ratio of A to B changed from 7% to 93% to 10% to 90%. 10-22 min, A:B ratio changed from 10%:90% to 12%:88%; 22-40 min, A:B ratio changed from 12%:88% to 14%:86%; 40-45 min, A:B ratio is 14%:86%; 45-60 min, A:B ratio changed from 14%:86% to 16%:84%; 60-65 min, A:B ratio 16%:84%; The preparation method of the test solution includes: taking the test sample, adding an organic solvent for extraction, collecting the extract for solid-liquid separation, and collecting the filtrate to obtain the solution; the organic solvent includes an aqueous methanol solution; the extraction step includes reflux extraction or ultrasonic extraction.

2. The method for constructing the characteristic spectrum of corn silk and its pharmaceutical preparations according to claim 1, characterized in that, The chromatographic conditions in the high-performance liquid chromatography detection step also include: The column temperature is 20-30℃; and / or, The flow rate is 0.1-0.5 mL / min; and / or, The injection volume is 1-5 µL.

3. The method for constructing the characteristic spectrum of corn silk and its pharmaceutical preparations according to claim 2, characterized in that, The chromatographic conditions in the high-performance liquid chromatography detection step also include: The column temperature is 25°C; and / or, The flow rate is 0.29-0.31 mL / min, more preferably 0.3 mL / min; and / or, The injection volume was 3 µL.

4. The method for constructing the characteristic spectrum of corn silk and its pharmaceutical preparations according to claim 3, characterized in that, The chromatographic conditions in the high-performance liquid chromatography detection step also include: The flow rate was 0.3 mL / min.

5. The method for constructing the characteristic spectrum of corn silk and its pharmaceutical preparations according to any one of claims 1-4, characterized in that, The method further includes the step of preparing a reference solution of the control medicinal material, specifically including: taking corn silk as a reference medicinal material, adding water and heating and refluxing for extraction, collecting the extract and separating the solid and liquid, collecting the residue and adding an organic solvent for extraction, collecting the extract and separating the solid and liquid, collecting the filtrate, and obtaining the reference solution of the control medicinal material.

6. The method for constructing the characteristic spectrum of corn silk and its pharmaceutical preparations according to any one of claims 1-4, characterized in that, The method further includes the steps of preparing a reference solution and constructing a characteristic chromatogram of the reference solution based on the high performance liquid chromatography. The reference standard includes ferulic acid; The method for preparing the reference solution includes the step of adding the reference substance to an organic solvent and mixing. The reference solution contains 1-20 µg of the reference standard per 1 ml.

7. The method for constructing the characteristic spectrum of corn silk and its pharmaceutical preparations according to claim 1, characterized in that, The organic solvent comprises an aqueous methanol solution with a concentration of 30-80 v / v, more preferably 50 v / v; And / or, the solid-liquid separation includes filtration or centrifugation.

8. The method for constructing the characteristic spectrum of corn silk and its pharmaceutical preparations according to any one of claims 1-4, characterized in that, The corn silk pharmaceutical preparations include corn silk formula granules, corn silk slices, corn silk standard decoction, or corn silk standard decoction freeze-dried powder.

9. The application of the method for constructing characteristic spectra of corn silk and its pharmaceutical preparations according to any one of claims 1-8 in the field of quality testing of corn silk and its pharmaceutical preparations.

10. A method for quality testing of corn silk and its pharmaceutical preparations, characterized in that, The method includes the steps of constructing a feature map and a reference feature map according to any one of claims 1-8, and the step of comparing the feature map with the reference feature map.

11. The quality testing method for corn silk and its pharmaceutical preparations according to claim 10, characterized in that, The characteristic chromatogram shows 13 characteristic peaks, corresponding to the retention times of the 13 characteristic peaks in the reference chromatogram of the medicinal material. Peak 3 corresponds to the retention time of the corresponding reference peak. The peak corresponding to the retention time of the ferulic acid reference peak is peak S. The relative retention times of each characteristic peak and peak S are calculated, and these relative retention times should be within ±10% of the specified value. The specified value is: Peak 1: 0.76, Peak 2: 0.90, Peak 4: 1.37, Peak 5: 1.75, Peak 6: 1.92, Peak 7: 2.08, Peak 8: 2.13, Peak 9; 2.19, Peak 10: 2.66, Peak 11: 3.07, Peak 12: 3.15, Peak 13: 3.51.

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