Construction method of wine-fried radix berberidis fingerprint spectrum
By constructing a fingerprint spectrum of roasted ginseng using HPLC, the problem of the lack of a recorded quality standard for roasted ginseng was solved, enabling accurate quality control and batch difference identification of roasted ginseng, and providing a reliable quality control method.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-12-10
- Publication Date
- 2026-03-10
AI Technical Summary
In the existing technology, there is no quality standard for wine-processed Kejing medicinal slices, and there is a lack of effective quality control methods, which makes it difficult to guarantee the quality of medication.
A fingerprint chromatogram of roasted ginseng was constructed using HPLC. Multiple batches of roasted ginseng test solutions were detected using ellagic acid solution. A Shim-pack VP-ODS column and gradient elution technology were used, with acetonitrile and 0.1% phosphoric acid aqueous solution as the mobile phase. The detection wavelength was 254 nm. A quality control method for roasted ginseng was established.
It enables accurate quality control of processed herbal medicine, identifies differences between different batches, provides a reliable reference for the establishment and control of quality standards for processed herbal medicine, and ensures safe and effective use.
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Figure CN121633339A_ABST
Abstract
Description
TECHNICAL FIELD
[0001] The present application relates to the technical field of traditional Chinese medicine drug analysis, and particularly relates to a construction method of a wine-fried Kejing fingerprint spectrum. BACKGROUND
[0002] Goingz (Kejing or Alangium chinense (Lour.) Harms) is the dry lateral root and fibrous root of Alangium chinense (Lour.) Harms in Alangiaceae. Kejing is also used as a medicine by the Yao, Miao, Yi and Dong ethnic groups. The Chinese name of Kejing is Alangium chinense. Alangium chinense was first recorded in the Qing Dynasty Bencao Cong Xin. The Chinese Pharmacopoeia (1977 edition) recorded Alangium chinense. In Zhuang medicine, Alangium chinense is bitter, pungent, slightly warm, and toxic. It can remove wind and toxin, dissipate blood stasis and relieve pain. It is used for treating wind-damp arthralgia (rheumatic pain), numbness of limbs (limb numbness), Bangbayin (shoulder periarthritis), Huoyinyin (cervical spondylosis), Lindedingdingxiang (traumatic injury) and Heuyin (lumbago). In modern Chinese medicine, Alangium chinense is pungent, slightly warm, and slightly toxic, and belongs to the liver and kidney meridians. Alangium chinense is used in many prescription formulas. For example, in the Zhuang medicine Longzhuantongbi formula, Alangium chinense is used as an auxiliary medicine, and Alangium chinense has the effects of dispelling wind and dampness, removing blood stasis and relieving pain. The rheumatism-defining capsule, which uses Alangium chinense as the main medicine, has the effects of activating blood, unblocking collaterals, removing arthralgia and relieving pain. The “Xiaobuling” mixture, which uses Alangium chinense as the main medicine, can dispel wind, unblock collaterals, dissipate blood stasis and relieve pain.
[0003] The traditional Chinese medicine fingerprint spectrum is a quality evaluation mode of traditional Chinese medicine or natural medicine that is widely accepted at home and abroad. Its application and rapid development embodies the comprehensive quality evaluation of traditional Chinese medicine, and meets the characteristics of the overall characterization analysis of traditional Chinese medicine quality control. At present, the quality standard of wine-fried Kejing decoction pieces has not been recorded, and there is no literature report on the quality standard of wine-fried Kejing decoction pieces. In order to ensure the quality of the medicine, it is urgent to establish the quality standard of wine-fried Kejing. SUMMARY
[0004] The purpose of the present application is to provide a construction method of a wine-fried Kejing fingerprint spectrum to solve the problems existing in the prior art. The present application detects specific chemical components in wine-fried Kejing to construct a fingerprint spectrum, so as to better realize the quality control of wine-fried Kejing.
[0005] To achieve the above-mentioned purpose, the present application provides the following solutions:
[0006] The present application provides a construction method of a wine-fried Kejing fingerprint spectrum, which comprises the following steps: using tannic acid solution as a control solution, performing HPLC determination on multiple batches of wine-fried Kejing test sample solutions, and obtaining the wine-fried Kejing fingerprint spectrum.
[0007] The preparation method of the wine-fried Kejing comprises the following steps: mixing Kejing and yellow rice wine, and then sequentially performing dampening, frying and air-drying to obtain the wine-fried Kejing.
[0008] Preferably, the HPLC determination adopts a Shim-pack VP-ODS chromatographic column; gradient elution is carried out with acetonitrile as mobile phase A and 0.1% phosphoric acid aqueous solution as mobile phase B.
[0009] Preferably, the gradient elution is as shown in the following table:
[0010] .
[0011] Preferably, the detection condition of the HPLC determination is as follows: column temperature 25 DEG C, flow rate 1.0 mL / min, injection volume 10 mu L, and detection wavelength 254 nm.
[0012] Preferably, the amount of the yellow rice wine is 10% of the amount of the Kejing quality.
[0013] Preferably, the time of the dampening is 24 h.
[0014] Preferably, the time of the frying is 16 min, the temperature is 150 DEG C, the rotation speed is 40 r / min, and the frequency of the frying is 120 times / min.
[0015] Preferably, the preparation method of the wine-fried Kejing test sample solution comprises the following steps: mixing the wine-fried Kejing and an ethanol solution, and then sequentially performing ultrasonic treatment and filtration to obtain the wine-fried Kejing test sample solution.
[0016] Preferably, the mass-volume ratio of the wine-fried Kejing and the ethanol solution is 1g:(20-25)mL; the volume percentage content of ethanol in the ethanol solution is 50%; the ultrasonic treatment is performed for 60 min, the power is 200W, and the frequency is 40Hz.
[0017] The application provides application of the wine-fried Kejing fingerprint spectrum obtained by the above-mentioned construction method in evaluation of quality of the wine-fried Kejing.
[0018] The application provides application of the wine-fried Kejing fingerprint spectrum obtained by the above-mentioned construction method in identification of authenticity of the wine-fried Kejing.
[0019] The application discloses the following technical effects:
[0020] The application detects specific chemical components in the wine-fried Kejing, thereby constructing a fingerprint spectrum, so that quality control of the wine-fried Kejing is better achieved.
[0021] The fingerprint spectrum of the wine-fried Kejing constructed by the construction method of the application is used as a quality evaluation standard, has the advantages of accuracy and reliability, and can provide reference and basis for wine-fried Kejing research. BRIEF DESCRIPTION OF DRAWINGS
[0022] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the drawings needed to be used in the embodiments will be briefly introduced below. Obviously, the drawings in the following description are only some embodiments of the present application, and other drawings can be obtained by those skilled in the art without creative effort on the basis of these drawings.
[0023] Figure 1 Fig. 14 is the HPLC characteristic spectrum of 14 batches of wine-fried Kejing; wherein, 3 is tannic acid;
[0024] Figure 2 Fig. 15 is the fingerprint spectrum of 14 batches of wine-fried Kejing and the control fingerprint spectrum; wherein, 3 is tannic acid. DETAILED DESCRIPTION
[0025] The various exemplary embodiments of the present application will now be described in detail, which should not be considered as limiting the present application, but should be understood as a more detailed description of certain aspects, characteristics and embodiments of the present application.
[0026] It should be understood that the terms described in the present application are only for describing the specific embodiments, and are not used to limit the present application. In addition, for the numerical range in the present application, it should be understood that each intermediate value between the upper limit and the lower limit of the range is also specifically disclosed. Each smaller range between any stated value or intermediate value in the stated range, and any other stated value or intermediate value in the stated range, is also included in the present application. The upper limit and the lower limit of these smaller ranges can be independently included or excluded from the range.
[0027] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the present application pertains. Although preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present application. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials in connection with which the documents are concerned. In the event of any conflict between the content of this specification and the documents incorporated by reference, the content of this specification will control.
[0028] Many modifications and variations of the present application specification can be made without departing from the scope or spirit of the present application, which will be apparent to those skilled in the art. Other embodiments resulting from the present application specification will be apparent to those skilled in the art. The present application specification and examples are only exemplary.
[0029] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.
[0030] The materials and instruments used in the specific embodiments of this invention are as follows:
[0031] instrument:
[0032] Waters Alliance e2695 UV-vis high-performance liquid chromatograph (Waters Corporation, USA); CAMAG semi-automatic spotter (CAMAG, Switzerland); Reprostar 3 CAMAG thin-layer chromatography scanning imaging system (CAMAG, Switzerland); Olympus BX53 microscope (Olympus (China) Co., Ltd., Japan); 0.0001 g electronic balance (Sartorius Scientific Instruments (Beijing) Co., Ltd.); Milli-Q Reference laboratory ultrapure water system (Millipore Corporation, USA); Type 25B electric roasting machine (Shenma Machinery Factory, Ruian, Zhejiang); non-contact infrared thermometer (Xima Instruments Co., Ltd., Hong Kong, China).
[0033] Drug trials:
[0034] Illicium verum alkaloids (MedChemExpress, USA, batch number 129513, mass fraction ≥98%); Illicium verum reference material (China National Institute for Food and Drug Control, batch number: R5G2-HRA7); acetonitrile and methanol were chromatographically pure (Thermo-Fisher, USA); phosphoric acid was chromatographically pure (Tianjin Damao Chemical Reagent Factory); sodium 1-heptanesulfonate was analytically pure (Shanghai Maclean Biochemical Technology Co., Ltd., batch number: C12673682); potassium dihydrogen phosphate was analytically pure (Chengdu Jinshan Chemical Reagent Co., Ltd., batch number: 20210302); silica gel G plate (Qingdao Ocean Chemical Co., Ltd., batch number: 20220703); water was self-made ultrapure water.
[0035] Medicinal materials:
[0036] Fourteen batches of Alangium chinense (Lour.) Harms dried lateral roots and fibrous roots were purchased and collected, and were identified by ZHU Yilin, a senior experimentalist of Guangxi University of Chinese Medicine, as A. chinense (Lour.) Harms of Alangiaceae, and the origin information is shown in Table 1. The A. chinense (Lour.) Harms was processed by wine-frying to obtain wine-fried A. chinense (Lour.) Harms; wherein the specific steps of wine-frying were as follows: 200 g of A. chinense (Lour.) Harms was taken, and yellow rice wine was sprayed onto the medicinal slices, the total amount of yellow rice wine was 10% of the mass of A. chinense (Lour.) Harms, and the medicinal slices were soaked at room temperature for 24 h, then were put into a temperature-controlled frying machine (rotation speed 40 r / min, 120 times / min), and were fried at 150 ℃ for 16 min, and were taken out and dried to obtain 14 batches of wine-fried A. chinense (Lour.) Harms, which were numbered as S1-S 14 .
[0037] Table 1 Origin information of 14 batches of A. chinense (Lour.) Harms
[0038]
[0039] Example 1 Establishment of HPLC common peak chromatogram
[0040] 1. Medicinal materials
[0041] The 14 batches of wine-fried A. chinense (Lour.) Harms in Table 1.
[0042] 2. Chromatographic conditions
[0043] The chromatographic column was Shim-pack VP-ODS (250 mm×4.6 mm, 5 μm), the mobile phase was acetonitrile (A)-0.1% phosphoric acid aqueous solution (B), the elution mode was gradient elution (0-30 min: 7%-23% B, 77%-93% A; 30-45 min: 23%-35% B, 65%-77% A), the volume flow rate was 1 mL / min, the column temperature was 25 ℃, the detection wavelength was 254 nm, and the injection volume was 10 μL.
[0044] 3. Preparation method of reference solution
[0045] An appropriate amount of gallic acid was accurately weighed and placed in a 25 mL volumetric flask, and the gallic acid was dissolved and diluted with 50% ethanol to the mark to prepare a reference solution containing 1.200 mg / mL of gallic acid, thereby obtaining the reference solution.
[0046] 4. Preparation method of test solution
[0047] The wine-fried A. chinense (Lour.) Harms powder (passed through a No. 4 sieve) was accurately weighed, 1 g was taken and placed in a 50 mL conical flask with a stopper, 25 mL of 50% (V / V) ethanol was accurately measured and weighed, and then was ultrasonically treated for 60 min (power 200 W, 40 Hz), and then was cooled, the weight was made up, and then was shaken uniformly, and then was filtered through a 0.22 μm microporous filter membrane, thereby obtaining the test solution.
[0048] 5. Linear range investigation
[0049] Five reference solutions of ellagic acid were prepared with the mass concentrations of 0.0300 mg / mL, 0.1200 mg / mL, 0.2400 mg / mL, 0.4800 mg / mL, 0.6000 mg / mL and 1.2000 mg / mL, respectively. The peak area was recorded by injecting the reference solutions under the chromatographic conditions in the present embodiment. The standard curve was plotted with the mass of each reference as the abscissa (X) and the peak area as the ordinate (Y), and the regression equation, correlation coefficient and linear range were obtained by regression analysis. The regression equation of ellagic acid was Y = 5634468X - 61268, the correlation coefficient R2=0.9999, and the linear range was 0.0375 mg / mL-1.2000 mg / mL.
[0050] 6. Precision test
[0051] The same test sample solution (the sample No. S1 of the wine-fried Kujing medicinal material in Table 1) was prepared according to the preparation method in the present embodiment, and was continuously injected 6 times under the chromatographic conditions in the present embodiment. The relative retention time and relative peak area of each common characteristic peak to the peak S of ellagic acid were calculated, and the RSD value was calculated. The RSD value was 0.74%. The results showed that the RSD values were all less than 3.0%, indicating that the instrument precision was good.
[0052] 7. Reproducibility test
[0053] The sample No. S1 of the wine-fried Kujing medicinal material powder (passed through a No. 4 sieve) in Table 1 was prepared according to the preparation method in the present embodiment to prepare 6 test sample solutions, and was determined under the chromatographic conditions in the present embodiment. The results showed that the RSD value was 1.58%, and the RSD values were all less than 3.0%, indicating that the method had good reproducibility.
[0054] 8. Stability test
[0055] The test sample solution of the sample No. S1 of the wine-fried Kujing medicinal material in Table 1 was prepared according to the preparation method in the present embodiment, and was determined at 0, 2, 4, 8, 12 and 24 h under the chromatographic conditions in the present embodiment. The relative retention time and relative peak area of each common characteristic peak to the peak S of ellagic acid were calculated, and the RSD value was calculated. The RSD value was 0.77%. The results showed that the RSD values were all less than 3.0%, indicating that the test sample solution had good stability within 24 h.
[0056] 9. Establishment and analysis of HPLC characteristic peak spectrum
[0057] Take 14 batches of wine Zhi Kejing medicinal materials, according to the preparation method in this example, 14 batches of wine Zhi Kejing test solution was prepared, according to the chromatographic conditions in this example, the chromatogram of each batch of sample was recorded, the CDF format data file of 14 chromatograms was exported and imported into "Traditional Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System (2012.1 version)" software for common peak identification. The results showed that the characteristic chromatograms of 14 batches of wine Zhi Kejing medicinal materials had 7 common peaks Figure 1 ), of which 3 was tannic acid.
[0058] Example 2 Construction of wine Zhi Kejing fingerprint
[0059] 1. Medicinal materials
[0060] Table 1 14 batches of wine Zhi Kejing medicinal materials.
[0061] Chromatographic conditions
[0062] Shim-pack VP-ODS chromatographic column (250 mm x 4.6 mm, 5 μm), mobile phase acetonitrile (A)-0.1% phosphoric acid aqueous solution (B), elution mode: gradient elution (0~30 min, 7%~23% B; 30~45 min, 23%~35% B); volume flow rate 1 mL / min; column temperature 25℃; detection wavelength 254 nm; injection volume 10 μL.
[0063] 2. Preparation method of test solution
[0064] Take wine Zhi Kejing medicinal material powder (pass through No. 4 sieve) 1 g, accurately weigh, put into a conical flask with plug, accurately take 25 mL of 50% ethanol, weigh, ultrasonic for 60 min (power 200 W, 40 KHz), cool, make up the weight, shake well, pass through 0.22 μm microporous filter membrane, get it.
[0065] 3. Precision test
[0066] Take the test solution, according to the chromatographic conditions in this example, continuously sample for 6 times, take tannic acid peak (peak 3) as reference peak, calculate the relative retention time and relative peak area of each characteristic peak, the results show that the RSD value is less than 3.0% (n=6), which indicates that the instrument precision is good.
[0067] 4. Reproducibility test
[0068] Take 14 batches of wine Zhi Kejing medicinal material powder, 6 portions, prepare test solution according to the preparation method in this example, according to the chromatographic conditions in this example, take tannic acid peak (peak 3) as reference peak, calculate the relative retention time and relative peak area of each characteristic peak, the results show that the RSD value is less than 3.0% (n=6), which indicates that the method has good reproducibility.
[0069] 5. Stability test
[0070] The test sample solution (sample No. S1 in Table 1) was prepared according to the preparation method of this example, and was determined at 0, 2, 4, 8, 12, and 24 h, respectively, according to the chromatographic conditions in this example, with the peak of ellagic acid (peak No. 3) as the reference peak, to calculate the relative retention time and relative peak area of each characteristic peak. The results showed that the RSD values were all less than 3.0% (n = 6), indicating that the test sample solution had good stability within 24 h.
[0071] 6. Sample determination results
[0072] Fourteen batches of Jiaozhi Kejing medicinal material powders were taken, and test sample solutions were prepared according to the method (prepared according to the preparation method of this example) and determined (chromatographic conditions in this example) to obtain the chromatographic superimposition diagram of the fourteen batches of Jiaozhi Kejing, as shown in Figure 2 The chromatograms of the fourteen batches of samples were analyzed using the Traditional Chinese Medicine Chromatographic Characteristic Spectrum Similarity Evaluation System (2012.1 version), and the similarity was greater than 0.850. According to the principle of good relative retention time stability and detection in each batch of sample, seven common peaks were selected, of which No. 3 was ellagic acid. The Jiaozhi Kejing fingerprint spectrum obtained by this method and the control fingerprint spectrum are shown in Figure 2 .
[0073] Example 3. Identification of the authenticity of Jiaozhi Kejing medicinal material
[0074] 1. Medicinal material
[0075] The medicinal material to be detected.
[0076] 2. Chromatographic conditions
[0077] Ellagic acid conditions: the chromatographic column was Shim-pack VP-ODS (250 mm x 4.6 mm, 5 μm), the mobile phase was acetonitrile (A) - 0.1% phosphoric acid aqueous solution (B), the elution mode was gradient elution (0-30 min: 7%-23% B; 30-45 min: 23%-35% B), the volume flow rate was 1 mL / min, the column temperature was 25°C, the detection wavelength was 254 nm, and the injection volume was 10 μL.
[0078] 3. Preparation method of the control solution
[0079] An appropriate amount of ellagic acid was accurately weighed and placed in a 25 mL volumetric flask, and the ellagic acid was dissolved and diluted with 50% ethanol to the mark to prepare a control solution containing 1.200 mg / mL of ellagic acid, thereby obtaining the control solution.
[0080] 4. Preparation method of the test sample solution to be detected
[0081] Take the powder of the wine-fried Kejing medicinal material to be detected (pass through No. 4 sieve), take 1 g and accurately weigh, place in a 50 mL conical flask with a plug, accurately weigh 25 mL of 50% ethanol, weigh, ultrasonic for 60 min (power 200 W, 40 Hz), cool, make up the weight, shake well, pass through 0.22 μm microporous filter membrane, and thus the sample solution is obtained.
[0082] 5. Detection
[0083] Take the sample solution of the medicinal material to be detected, determine according to the chromatographic conditions in the example, record the chromatograms of each batch of sample, export the CDF format data file of the chromatogram and import into the “Traditional Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System (2012.1 version)” software for common peak identification.
[0084] 6. Judgment
[0085] Compare the obtained chromatogram with the wine-fried Kejing fingerprint chromatogram constructed in Example 2, if consistent, it is judged as true wine-fried Kejing.
[0086] The above examples only describe the preferred modes of the present application, and do not limit the scope of the present application. Without departing from the design spirit of the present application, various modifications and improvements of the technical solutions of the present application made by those skilled in the art shall fall within the protection scope determined by the claims of the present application.
Claims
1. A method for constructing a fingerprint of a wine, characterized in that, The steps of obtaining the fingerprint of the wine-fried Kejing by HPLC determination of multiple batches of wine-fried Kejing sample solutions with a solution of ellagic acid as a control solution are included. The preparation method of the wine-fried Kejing comprises the steps of mixing Kejing and yellow rice wine, then carrying out moistening, frying and air-drying in sequence to obtain the wine-fried Kejing.
2. The construction method of claim 1, wherein, The HPLC determination employs a Shim-pack VP-ODS chromatographic column, and gradient elution is carried out with acetonitrile as mobile phase A and 0.1% phosphoric acid aqueous solution as mobile phase B.
3. The construction method of claim 2, wherein, The gradient elution is shown in the following table: 。 4. The construction method of claim 1, wherein, The detection conditions of the HPLC determination are as follows: column temperature 25℃, flow rate 1.0 mL / min, injection volume 10 μL, and detection wavelength 254 nm.
5. The construction method of claim 1, wherein, The amount of the yellow rice wine is 10% of the mass of the Kejing. And / or, the moistening time is 24 h. And / or, the frying time is 16 min, the temperature is 150℃, the rotation speed is 40 r / min, and the frequency of turning and frying is 120 times / min.
6. The construction method of claim 1, wherein, The preparation method of the wine-fried Kejing sample solution comprises the steps of mixing the wine-fried Kejing and an ethanol solution, then carrying out ultrasonic treatment and filtration in sequence to obtain the wine-fried Kejing sample solution.
7. The construction method of claim 6, wherein, The mass-volume ratio of the wine-fried Kejing to the ethanol solution is 1 g: (20-25) mL, the volume percentage content of ethanol in the ethanol solution is 50%, the ultrasonic treatment time is 60 min, the power is 200 W, and the frequency is 40 Hz.
8. The application of the fingerprint of the wine-fried Kejing obtained by the construction method of any one of claims 1-7 in the evaluation of the quality of the wine-fried Kejing.
9. The application of the fingerprint of the wine-fried Kejing obtained by the construction method of any one of claims 1-7 in the identification of the authenticity of the wine-fried Kejing.