Liquid chromatography detection method for fingerprint spectrum of membranaceous beautyleaf root traditional Chinese medicinal material

Abstract

The invention discloses a liquid chromatography detection method for a fingerprint spectrum of a membranaceous beautyleaf root traditional Chinese medicinal material, which comprises the following steps: crushing membranaceous beautyleaf root to obtain membranaceous beautyleaf root powder, adding the membranaceous beautyleaf root powder into a solvent, shaking for 30 minutes, performing ultrasonic treatment for 30 minutes, and filtering to obtain filtrate; the liquid chromatography detection conditions are as follows: octadecylsilane chemically bonded silica is taken as a filler, a solution prepared by mixing acetonitrile, methanol and an ammonium acetate solution is taken as a mobile phase, the detection wavelength is 254nm, the flow velocity is 1mL / min, and the column temperature is 25-30 DEG C; collecting more than 10 kinds of membranaceous beautyleaf root traditional Chinese medicinal materials, detecting according to the steps, comparing the obtained detection results, and selecting common characteristic peaks to obtain the membranaceous beautyleaf root traditional Chinese medicinal material fingerprint spectrum. By detecting and analyzing the fingerprint spectrum of the membranaceous beautyleaf root traditional Chinese medicinal material, the authenticity and quality of the membranaceous beautyleaf root traditional Chinese medicinal material are simply and accurately identified, the quality control of the membranaceous beautyleaf root traditional Chinese medicinal material is improved, and the safety and stability of the membranaceous beautyleaf root traditional Chinese medicinal material are ensured.

Description

Technical Field

[0001] The invention belongs to the technical field of detection of Hengjingxi Chinese medicinal materials, and particularly relates to a liquid chromatography detection method for fingerprints of Hengjingxi Chinese medicinal materials. Background Art

[0002] Hengjingxi (Hengjingxi) is also known as the King of Combs, King of Combs, Thin-leaved Red-thick Shell, and Unicorn Wind. It belongs to the genus Calophyllum in the family Garciniaaceae. It is a shrub to small tree, 1 to 5 meters tall. Young branches are quadrangular with narrow wings. Leaves are opposite, thinly leathery, oblong or lanceolate. Flowers are bisexual, white with a reddish tinge. The drupe is ovate-oblong with a short pointed tip. It blooms in summer and fruit in autumn. Hengjingxi grows in sparse or dense mountain forests and is primarily distributed in Guangdong Province and Guangxi Zhuang Autonomous Region in my country. Hengjingxi has the properties of dispelling rheumatism, strengthening muscles and bones, promoting blood circulation, and relieving pain. It is used to treat rheumatic joint pain, low back pain, injuries from falls, icteric hepatitis, irregular menstruation, and dysmenorrhea. With the development of the Traditional Chinese Medicine (TCM) industry, the quality requirements for Hengjingxi medicinal materials have also increased significantly. The need exists to develop quality control methods for Hengjingxi medicinal materials that can simply and accurately detect their authenticity and quality, thereby ensuring the quality and stability of TCM.

[0003] Fingerprinting is a method that uses modern analytical techniques to comprehensively characterize the chemical components of traditional Chinese medicines and their preparations. It can reflect the overall chemical characteristics of the medicine, not just individual components. Different traditional Chinese medicines have different fingerprints, which can be used to distinguish different medicinal materials and control their quality. However, there are currently few reports on fingerprints related to Hengjingxi. There is still a need to establish a stable and reproducible fingerprint of Hengjingxi to assist in the quality control of Hengjingxi medicinal materials. Summary of the Invention

[0004] In response to the above-mentioned shortcomings, the present invention discloses a liquid chromatography detection method for the fingerprint spectrum of Hengjingxi Chinese medicinal materials. By detecting and analyzing the fingerprint spectrum of Hengjingxi Chinese medicinal materials, the authenticity and quality of Hengjingxi Chinese medicinal materials can be simply and accurately identified, the quality control of Hengjingxi Chinese medicinal materials can be improved, and the safety and stability of Hengjingxi Chinese medicinal materials can be ensured.

[0005] The present invention is achieved by adopting the following technical solutions: A liquid chromatography detection method for the fingerprint of a traditional Chinese medicine of Hengjingxi, comprising the following steps: (1) Grinding the horizontal warp mat to obtain horizontal warp mat powder, adding the horizontal warp mat powder to a solvent, shaking for 30 minutes, ultrasonicating for 30 minutes, and then filtering to obtain a filtrate; the solvent is any one of methanol and petroleum ether; (2) The filtrate obtained in step (1) was subjected to liquid chromatography detection. The conditions for the liquid chromatography detection were as follows: octadecylsilane bonded silica gel (C18) was used as a filler, a solution prepared by mixing acetonitrile, methanol and ammonium acetate solution was used as a mobile phase, the detection wavelength was 254 nm, the flow rate was 1 mL / min, and the column temperature was 25 to 30°C; (3) Collect more than 10 kinds of Hengjingxi Chinese medicinal materials and test them according to the steps described in steps (1) to (2), compare the obtained test results, select common characteristic peaks to obtain the Hengjingxi Chinese medicinal materials fingerprint spectrum, the Hengjingxi Chinese medicinal materials fingerprint spectrum includes peaks 1 to 4, and the relative retention times of peaks 1 to 4 are 8.093 to 8.49 min, 11.549 to 12.246 min, 12.268 to 12.962 min, and 14.413 to 15.338 min, respectively.

[0006] Furthermore, in step (1), the horizontal warp mat is crushed and passed through an 80-100 mesh sieve to obtain horizontal warp mat powder.

[0007] Furthermore, in step (1), the mass ratio of the horizontal mat powder to the volume ratio of the solvent is 1 g:5 mL, and the solvent is methanol.

[0008] Furthermore, in step (1), the cross-corrugated mat powder is added to the solvent and then shaken at a speed of 2000-3000 r / min for 30 minutes, and then ultrasonicated for 30 minutes, and the shaking speed during the ultrasonication process is 2000-3000 r / min.

[0009] Furthermore, in step (2), the mass concentration of the ammonium acetate solution is 0.5%, and the volume ratio of the acetonitrile, methanol and ammonium acetate solution is 50:18:32.

[0010] The above-mentioned fingerprint of the Hengjingxi Chinese medicinal material is used to identify the Hengjingxi Chinese medicinal material. Specifically, the medicinal material to be identified is tested according to the method in steps (1) to (2), and then the obtained liquid chromatography results are compared with the fingerprint of the Hengjingxi Chinese medicinal material. If the peaks 1 to 4 are present, it can be preliminarily determined that the medicinal material to be tested is Hengjingxi.

[0011] Compared with the existing technology, this technical solution has the following beneficial effects: 1. The present invention adopts liquid chromatography method to detect and construct fingerprint spectrum of Hengjingxi Chinese medicinal materials, and then detects the corresponding liquid chromatogram of the purchased Hengjingxi medicinal materials according to the detection method of the present invention. By comparing with the constructed Hengjingxi fingerprint spectrum, the authenticity and quality of Hengjingxi Chinese medicinal materials can be accurately identified, and the counterfeit or substandard medicinal materials or medicinal materials that do not meet the quality requirements can be effectively screened out, thereby improving the quality control of Hengjingxi Chinese medicinal materials and ensuring the safety and stability of Hengjingxi medicinal materials.

[0012] 2. The method of the present invention is simple, has good accuracy, repeatability and durability, and the peak shape in the chromatogram obtained by detection is beautiful and the separation is appropriate, which is conducive to comparative analysis and improves the accuracy of analysis; at the same time, when the present invention is used for sample detection, the time required for single sample analysis is short, which is conducive to improving the efficiency of analysis work, and the operation is convenient, which reduces the workload of detection and analysis personnel. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Figure 1 This is a liquid chromatogram of the No. 1 Hengjingxi medicinal material sample detected in Example 1.

[0014] Figure 2 This is a liquid chromatogram of the No. 2 Hengjingxi medicinal material sample detected in Example 1.

[0015] Figure 3 This is a liquid chromatogram of the No. 3 Hengjingxi medicinal material sample detected in Example 1.

[0016] Figure 4 This is a liquid chromatogram of the No. 4 Hengjingxi medicinal material sample detected in Example 1.

[0017] Figure 5 This is a liquid chromatogram of the No. 5 Hengjingxi medicinal material sample detected in Example 1.

[0018] Figure 6 This is a liquid chromatogram of the No. 6 Hengjingxi medicinal material sample detected in Example 1.

[0019] Figure 7 This is a liquid chromatogram of the No. 7 Hengjingxi medicinal material sample detected in Example 1.

[0020] Figure 8 This is a liquid chromatogram of the No. 8 Hengjingxi medicinal material sample detected in Example 1.

[0021] Figure 9 This is a liquid chromatogram of the No. 9 Hengjingxi medicinal material sample detected in Example 1.

[0022] Figure 10 This is a liquid chromatogram of the No. 10 Hengjingxi medicinal material sample detected in Example 1.

[0023] Figure 11This is a liquid chromatogram of the No. 11 Hengjingxi medicinal material sample detected in Example 1.

[0024] Figure 12 This is the liquid chromatogram obtained when methanol solvent was used for detection in Experimental Example 1.

[0025] Figure 13 This is the liquid chromatogram obtained when petroleum ether solvent was used for detection in Experimental Example 1.

[0026] Figure 14 This is the liquid chromatogram obtained when the mobile phase ratio in Experimental Example 2 was 50:18:32.

[0027] Figure 15 This is the liquid chromatogram obtained when the mobile phase ratio in Experimental Example 2 was 67:12:21.

[0028] Figure 16 This is the liquid chromatogram obtained when the mobile phase ratio in Experimental Example 2 was 30:25:45.

[0029] Figure 17 This is the liquid chromatogram obtained when a C18 column was used for detection in Experimental Example 3.

[0030] Figure 18 This is the liquid chromatogram obtained when using a C8 chromatographic column for detection in Experimental Example 3.

[0031] Figure 19 This is the liquid chromatogram obtained when the chromatographic column produced by Shiseido was used for detection in Experimental Example 4.

[0032] Figure 20 This is the liquid chromatogram obtained when the chromatographic column produced by Napu was used for detection in Experimental Example 4.

[0033] Figure 21 This is the liquid chromatogram obtained when the chromatographic column produced by Shimadzu was used for detection in Experimental Example 4.

[0034] Figure 22 This is the liquid chromatogram obtained when the chromatographic column produced by the Phenylephrine manufacturer was used for detection in Experimental Example 4.

[0035] Figure 23 This is the liquid chromatogram obtained when the column temperature was 25°C in Experimental Example 5.

[0036] Figure 24 This is the liquid chromatogram obtained when the column temperature was 30°C in Experimental Example 5.

[0037] Figure 25 This is the liquid chromatogram obtained when the column temperature was 35°C in Experimental Example 5.

[0038] Figure 26This is the liquid chromatogram obtained when the column temperature was 40°C in Experimental Example 5.

[0039] Figure 27 This is the liquid chromatogram obtained during the blank control test in Experimental Example 6.

[0040] Figure 28 This is the liquid chromatogram obtained when parallel sample 1 was detected in Experimental Example 6.

[0041] Figure 29 This is the liquid chromatogram obtained when parallel sample 2 was detected in Experimental Example 6.

[0042] Figure 30 This is the liquid chromatogram obtained when parallel sample 3 was detected in Experimental Example 6.

[0043] Figure 31 This is the liquid chromatogram obtained when parallel sample 4 was detected in Experimental Example 6.

[0044] Figure 32 This is the liquid chromatogram obtained when parallel sample 5 was tested in Experimental Example 6.

[0045] Figure 33 This is the liquid chromatogram obtained when parallel sample 6 was detected in Experimental Example 6.

[0046] Figure 34 This is the liquid chromatogram obtained when parallel sample 7 was tested in Experimental Example 6.

[0047] Figure 35 This is the liquid chromatogram obtained by experimenter 1 during the test in Experimental Example 7.

[0048] Figure 36 This is the liquid chromatogram obtained by experimenter 2 during the test in Experimental Example 7. DETAILED DESCRIPTION

[0049] The present invention is further illustrated by the following examples, which are not intended to limit the present invention. Specific experimental conditions and methods not specified in the following examples are conventional methods well known to those skilled in the art.

[0050] Example 1: A liquid chromatography method for detecting the fingerprint of a Chinese medicinal material of Hengjingxi, comprising the following steps: (1) Grind the horizontal mat and pass it through a 100-mesh sieve to obtain horizontal mat powder, add the horizontal mat powder to a solvent, and then shake it at a speed of 3000 r / min for 30 minutes, then ultrasonicate it for 30 minutes, and shake it at a speed of 2000 r / min during the ultrasonication process, and then filter it to obtain a filtrate; the solvent is methanol; the mass ratio of the horizontal mat powder to the volume of the solvent is 1 g:5 mL; (2) The filtrate obtained in step (1) was subjected to liquid chromatography detection. The conditions of the liquid chromatography detection were as follows: octadecylsilane bonded silica gel was used as a filler, a solution prepared by mixing acetonitrile, methanol and ammonium acetate solution was used as a mobile phase, the detection wavelength was 254 nm, the flow rate was 1 mL / min, and the column temperature was 25°C; the mass concentration of the ammonium acetate solution was 0.5%, and the volume ratio of the acetonitrile, methanol and ammonium acetate solution was 50:18:32; (3) Collect 11 kinds of Chinese medicinal materials (see Table 1) and test them according to the steps (1) to (2). Figures 1 to 11 ) were compared, and the common characteristic peaks were selected to obtain the fingerprint of Hengjingxi Chinese medicinal materials. The fingerprint of Hengjingxi Chinese medicinal materials includes peaks 1 to 4, and the relative retention times of peaks 1 to 4 are 8.093-8.49 min, 11.549-12.246 min, 12.268-12.962 min, and 14.413-15.338 min, respectively.

[0051] The application of the fingerprint spectrum of the Hengjingxi Chinese medicinal material described in this embodiment is used for the identification of the Hengjingxi Chinese medicinal material. Specifically, the medicinal material to be identified is tested according to the method in steps (1) to (2), and then the obtained liquid chromatography results are compared with the fingerprint spectrum of the Hengjingxi Chinese medicinal material. If the peaks 1 to 4 are present, it can be preliminarily determined that the medicinal material to be tested is Hengjingxi.

[0052] Table 1 Collection of Chinese medicinal materials for different horizontal mats

[0053] Table 2 Retention time results of different horizontal Chinese medicinal materials (min)

[0054] Experimental Example 1: The same sample of the medicinal material of Hengjingxi was detected according to the method described in step (1) and step (2) of Example 1, wherein a solution prepared by mixing acetonitrile, methanol and ammonium acetate solution was used as the mobile phase, the volume ratio of acetonitrile, methanol and ammonium acetate solution was 67:12:21, and methanol and petroleum ether were used as solvents respectively. The detection results are shown in Tables 3 and Figures 12-13 .

[0055] Table 3 Retention time results using different solvents (min)

[0056] According to Table 3 and Figures 12-13 The results show that both methanol and petroleum ether can be used as solvents in the method of the present invention, and the obtained chromatogram can clearly obtain the characteristic peaks corresponding to peaks 1 to 4.

[0057] Experimental Example 2: The same sample of the medicinal material Hengjingxi was tested according to the method described in steps (1) and (2) of Example 1, wherein different mobile phases were used for testing, i.e., the volume ratios of acetonitrile, methanol and ammonium acetate solution in the mobile phase were 50:18:32, 67:12:21 and 30:25:45, respectively. The test results are shown in Tables 4 and Figures 14-16 .

[0058] Table 4 Retention time results using different mobile phases (min)

[0059] According to Table 4 and Figures 14-16 The results show that the mobile phase ratio (50:18:32) used in the present invention is relatively reasonable, the peak elution time is appropriate, the peak shape is beautiful, and the separation degree is appropriate.

[0060] Experimental Example 3: The same sample of the medicinal material of Hengjingxi was tested according to the method described in step (1) and step (2) of Example 1, wherein different chromatographic columns (C18 and C8) were used for testing. The test results are shown in Tables 5 and Figures 17-18 .

[0061] Table 5 Retention time results using different chromatographic columns (min)

[0062] According to Table 5 and Figures 17-18 The results show that the C18 chromatographic column used in the present invention has good separation and good detection effect.

[0063] Experimental Example 4: The same sample of the medicinal material of Hengjingxi was tested according to the method described in steps (1) and (2) of Example 1, wherein C18 chromatographic columns produced by different manufacturers were used for testing. The test results are shown in Tables 6 and Figures 19-22 .

[0064] Table 6 Retention time results (min) using C18 columns produced by different manufacturers

[0065] According to Table 6 and Figures 19-22 The results show that the C18 column produced by Napu was selected, the peak elution time was appropriate, the amount of solvent used was appropriate, the peak shape was beautiful, and the separation was good.

[0066] Experimental Example 5: The same sample of the medicinal material Hengjingxi was tested according to the method described in step (1) and step (2) of Example 1, wherein different column temperatures were used, namely, column temperatures of 25°C, 30°C, 35°C, and 40°C, respectively. The test results are shown in Tables 7 and Figures 23-26 .

[0067] Table 7 Retention time results using different column temperatures (min)

[0068] According to Table 7 and Figures 23-26 The results show that only when the column temperature is 25-30°C as described in the present invention can a good peak elution effect be obtained, and too high a temperature will affect the peak elution effect.

[0069] Experimental Example 6: According to the method described in steps (1) and (2) of Example 1, the Hengjingxi medicinal material sample was tested and divided into 7 samples for parallel testing. At the same time, the test solution without the Hengjingxi medicinal material sample was used as a blank control for testing. The test results are shown in Tables 8 and Figures 27-34 .

[0070] Table 8 Retention time results of parallel tests (min)

[0071] According to Table 8 and Figures 27-34 The results show that the method of the present invention has good specificity, and the method is stable and has good reproducibility.

[0072] Experimental Example 7: Arrange two experimenters to use two instruments of the same model to test the same Hengjingxi medicinal material sample according to the method described in step (1) and step (2) in Example 1. The test results are shown in Tables 9 and Figures 35-36 .

[0073] Table 9 Retention time results detected by different experimenters (min)

[0074] According to Table 9 and Figures 35-36 It can be seen from the results that the results obtained by different experimenters according to the method of the present invention are reproducible and accurate.

[0075] In addition, it should be understood that although this specification is described in terms of implementation methods, not every implementation method contains only one independent technical solution. This narrative method of the specification is only for the sake of clarity. Those skilled in the art should regard the specification as a whole. The technical solutions in each embodiment can also be appropriately combined to form other implementation methods that can be understood by those skilled in the art.

Claims

1. A liquid chromatography detection method for the fingerprint of Hengjingxi Chinese medicinal materials, characterized in that: The following steps are involved: (1) crushing the horizontal warp mat to obtain horizontal warp mat powder, adding the horizontal warp mat powder to a solvent, shaking for 30 minutes, ultrasonicating for 30 minutes, and filtering to obtain a filtrate; the solvent is any one of methanol and petroleum ether; (2) taking the filtrate obtained in step (1) for liquid chromatography detection, the conditions of the liquid chromatography detection are: using octadecylsilane bonded silica gel as a filler, a solution prepared by mixing acetonitrile, methanol and ammonium acetate solution as a mobile phase, a detection wavelength of 254 nm, a flow rate of 1 mL / min, and a column temperature of 25 to 30°C; (3) Collect more than 10 kinds of Hengjingxi Chinese medicinal materials and perform tests according to the steps described in steps (1) to (2), compare the test results, select common characteristic peaks to obtain a Hengjingxi Chinese medicinal material fingerprint spectrum, and the Hengjingxi Chinese medicinal material fingerprint spectrum includes peaks 1 to 4. The relative retention times of peaks 1 to 4 are 8.093-8.49 min, 11.549-12.246 min, 12.268-12.962 min, and 14.413-15.338 min, respectively.

2. The liquid chromatography detection method for the fingerprint of the traditional Chinese medicine Hengjingxi according to claim 1, characterized in that: In step (1), the horizontal warp mat is crushed and passed through a 80-100 mesh sieve to obtain horizontal warp mat powder.

3. The liquid chromatography detection method for the fingerprint of the traditional Chinese medicine of Hengjingxi according to claim 1, characterized in that: In step (1), the mass ratio of the horizontal warp mat powder to the volume ratio of the solvent is 1 g:5 mL, and the solvent is methanol.

4. The liquid chromatography detection method for the fingerprint of the traditional Chinese medicine of Hengjingxi according to claim 1, characterized in that: In step (1), the powder of the horizontal warp mat is added to the solvent and then shaken at a speed of 2000-3000 r / min for 30 minutes, and then ultrasonicated for 30 minutes, and the shaking speed during the ultrasonication process is 2000-3000 r / min.

5. The liquid chromatography detection method for the fingerprint of the traditional Chinese medicine of Hengjingxi according to claim 1, characterized in that: In step (2), the mass concentration of the ammonium acetate solution is 0.5%, and the volume ratio of acetonitrile, methanol and ammonium acetate solution is 50:18:

32.

6. Application of the fingerprint of the traditional Chinese medicine Hengjingxi obtained by the method according to any one of claims 1 to 5, characterized in that: The Hengjingxi Chinese medicinal material fingerprint spectrum is used for the identification of the Hengjingxi Chinese medicinal materials. Specifically, the medicinal materials to be identified are tested according to the methods in steps (1) to (2), and then the obtained liquid chromatography results are compared with the Hengjingxi Chinese medicinal material fingerprint spectrum. If the medicinal materials have the peaks 1 to 4, it can be preliminarily determined that the medicinal materials to be tested are Hengjingxi.

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