Thin-layer identification method for folium artemisia argyi and vinegar folium artemisia argyi formula granules

Abstract

A thin-layer identification method for Folium Artemisia argyi and vinegar Folium Artemisia argyi formula granules, comprising: pre-treating Folium Artemisia argyi formula granules to obtain a test solution; pre-treating vinegar Folium Artemisia argyi formula granules to obtain a vinegar Folium Artemisia argyi formula granule solution; taking a scopoletin reference substance and adding methanol for dissolution to obtain a reference solution; performing thin-layer chromatographic testing on the test solution, the vinegar Folium Artemisia argyi formula granule solution, and the reference solution, a thin-layer plate being a silica gel G thin-layer plate, and a developer being n-hexane-ethyl acetate-toluene; placing the plate under an ultraviolet lamp for inspection, if the test solution and the reference solution have the same color fluorescent spots at the same Rf, and the inspected substance also has fluorescent spots at Rf=0.625-0.741, then the inspected substance is Folium Artemisia argyi formula granules; if there are no fluorescent spots, then the inspected substance is vinegar Folium Artemisiae argyi formula granules.

Description

Thin layer identification method of mugwort and mugwort vinegar granules

[0001] This application claims priority to a Chinese patent application filed with the Patent Office of China on October 9, 2023, with application number 202311301309.X and invention name “Thin layer identification method for mugwort and vinegar mugwort formula granules”, the entire contents of which are incorporated by reference into this application. Technical Field

[0002] The invention relates to the technical field of analysis and detection, in particular to a thin-layer identification method for mugwort leaves and mugwort vinegar formula granules. Background Art

[0003] Artemisia argyi Lévl. et Vant. is a dried leaf of the Asteraceae plant [1]. It has the effects of warming the meridians and stopping bleeding, dispelling cold and relieving pain, and externally removing dampness and relieving itching. Vinegar-treated wormwood is warm but not dry, focusing on warming the meridians and stopping bleeding, and can enhance the cold-dispelling and analgesic effects. There are differences in the physical and chemical properties and the dehumidifying and relieving itching effects of wormwood before and after processing. The effect of stopping bleeding and warming the interior is better after processing with vinegar. This shows that the chemical composition of wormwood and vinegar-treated wormwood has changed during the processing process.

[0004] The chemical components of mugwort mainly include volatile oils, flavonoids, terpenes, phenylpropanoids, organic acids, steroids, polysaccharides, and trace elements. Current research on mugwort focuses on the composition and pharmacological effects of different processed products. For example, Zhang Tiantian, Sun Lili, Zhou Qian, et al. conducted a GC-MS study on the volatile oil components of mugwort and its processed products, which showed that the chemical composition of mugwort changed significantly after vinegar processing. While properties can be used to distinguish mugwort and vinegar-processed mugwort slices, methods for distinguishing the two formula granules have not been reported.

[0005] Since mugwort and vinegar mugwort granules are made from mugwort through a series of processes, some of which destroy the original properties, the property identification method is not suitable for the identification of formula granules. Currently, there is no effective method for distinguishing mugwort granules from vinegar mugwort granules. Therefore, it is urgent to establish a method that can distinguish mugwort granules from vinegar mugwort granules.

[0006] Summary of the Invention

[0007] In view of this, the technical problem to be solved by the present invention is to provide a thin layer identification method for mugwort and vinegar mugwort formula granules.

[0008] The terms "include," "comprising," and "having" are used interchangeably herein and are intended to indicate the inclusiveness of a solution, meaning that other elements may be present in addition to the listed elements. It should also be understood that the use of "include," "comprising," and "having" in this document also provides a "consisting of" solution.

[0009] In this application, the term "and / or" describes the association relationship between associated objects, indicating that three relationships may exist. For example, A and / or B can mean: A exists alone, A and B exist at the same time, and B exists alone. A and B can be singular or plural.

[0010] In this application, "at least one" means one or more, and "more than one" means two or more. "At least one of the following" or similar expressions refers to any combination of these items, including any combination of single or plural items.

[0011] It should be understood that in the various embodiments of the present application, the size of the serial numbers of the following processes does not mean the order of execution. Some or all of the steps can be executed in parallel or sequentially. The execution order of each process should be determined by its function and internal logic, and should not constitute any limitation on the implementation process of the embodiments of the present application.

[0012] The present invention discloses the construction and application of a thin-layer chromatography (TLC) identification method for mugwort and vinegar mugwort granules. By observing the TLC of a sample, the presence of a red fluorescent spot at a ratio shift value of 0.556-0.752 is used as an identification point for the mugwort granules, while the absence of a red fluorescent spot at the same location is used as an identification point for the vinegar mugwort granules. The established TLC method can quickly and effectively identify mugwort and vinegar mugwort granules. This method has strong specificity, good stability, high precision, and is convenient and easy to master. This method enables more standardized quality control of mugwort and vinegar mugwort granules, ensuring the efficacy of the drugs.

[0013] A thin layer identification method for mugwort and mugwort vinegar granules, comprising:

[0014] A) Pre-treat the Artemisia argyi granules to obtain a test solution;

[0015] Pre-treating the vinegar wormwood formula granules to obtain a vinegar wormwood formula granule solution;

[0016] B) Take the scopolamine reference substance and dissolve it in methanol to obtain a reference solution;

[0017] C) The test solution, vinegar wormwood formula granule solution and reference solution were subjected to thin layer chromatography (TLC) using a silica gel G TLC plate and n-hexane-ethyl acetate-toluene as the developing solvent;

[0018] D) Check under ultraviolet light. If the test sample and control medicinal material solution have fluorescent spots of the same color at the same Rf, and there are fluorescent spots at Rf = 0.625-0.741, then they are mugwort formula granules; if there are no fluorescent spots at Rf = 0.625-0.741, then they are vinegar mugwort formula granules.

[0019] The invention provides wormwood leaf and wormwood leaf vinegar formula granules. First, the wormwood leaf formula granules are pretreated to obtain a test solution.

[0020] The pretreatment of the mugwort formula granules or vinegar mugwort formula granules of the present invention is specifically as follows:

[0021] a) adding methanol to the wormwood formula granules or vinegar wormwood formula granules, sonicating, filtering, evaporating, dissolving the residue in water, passing it through a D101 macroporous adsorption resin column, eluting with 30% methanol, discarding the eluate, and then eluting with ethyl acetate, and collecting the eluate;

[0022] b) Recover the solvent from the eluate until it is dry, and dissolve the residue in ethyl acetate to obtain the product.

[0023] The processing method of the mugwort formula granules or the vinegar mugwort formula granules provided by the present invention comprises the following steps: firstly adding methanol to the mugwort formula granules or the vinegar mugwort formula granules, ultrasonicating the granules, filtering, volatilizing, and dissolving the residue in water.

[0024] In some embodiments of the present invention, the ultrasonic time is 5 to 10 minutes.

[0025] In some embodiments of the present invention, the ultrasonic time is 10 minutes.

[0026] Then pass through D101 macroporous adsorption resin column, elute with 30% methanol, discard the eluate, elute with ethyl acetate, and collect the eluate; the inner diameter of the D101 macroporous adsorption resin is 1 cm, and the column height is 14 cm.

[0027] In some embodiments of the present invention, the mass volume ratio of the mugwort formula granules or vinegar mugwort formula granules, methanol, water, 30% methanol and ethyl acetate is 0.5g:10mL:5mL:20mL:20mL.

[0028] The present invention does not limit the specific steps of pre-treating the vinegar and wormwood formula granules, and the same steps may be used as above.

[0029] Take the scopolamine reference substance, add methanol to dissolve it, and obtain a reference substance solution.

[0030] The mass volume ratio of the scopolamine reference substance of the present invention and methanol is 40 μg:1 mL.

[0031] The test solution, vinegar wormwood formula granule solution and reference solution were subjected to thin layer chromatography.

[0032] Preferably, the test solution, vinegar wormwood formula granule solution and reference solution are aspirated and spotted on the same silica gel G thin layer plate, developed with a developing agent, taken out, dried and inspected under ultraviolet light. The ultraviolet light wavelength is 365nm.

[0033] According to the present invention, the developing agent is n-hexane-ethyl acetate-toluene;

[0034] In some preferred embodiments of the present invention, the mass ratio of n-hexane-ethyl acetate-toluene is 1:3:2.

[0035] The thin layer plate described in the present invention is a silica gel G thin layer plate; preferably a GF254 thin layer plate. Prefabricated silica gel GF254 plates can be selected from Tianjin Silida, Merck, and Qingdao Ocean. Results demonstrate that this method is robust and meets identification requirements. Compared with the control herbal material atlas, the sample and control herbal material atlases exhibit spots of the same color at corresponding locations.

[0036] Check under ultraviolet light. If the test and reference solutions have fluorescent spots of the same color at the same Rf, and red fluorescent spots at Rf = 0.625-0.741, then they are mugwort formula granules.

[0037] If there is no fluorescent spot at Rf=0.625~0.741, it is vinegar and mugwort formula granules.

[0038] In a preferred embodiment of the present invention, if the test sample and the control medicinal material solution have fluorescent spots of the same color at the same Rf, and there is a fluorescent spot at Rf = 0.654, then it is the mugwort formula granules;

[0039] If there is no fluorescent spot at Rf=0.654, it is vinegar and mugwort formula granules.

[0040] The spotting amount of the thin layer chromatography test solution, vinegar wormwood formula granule solution and reference solution is 5 to 15 μL.

[0041] When 5-15 μl of the reference solution and the test solution are spotted, the spots are clearly colored and have good separation, and the spot volume is determined to be 5-15 μl.

[0042] The inspection temperature of the present invention is 4°C to 35°C.

[0043] This method has good durability at different temperatures. Compared with the reference sample spectrum, the test sample spectrum and the reference sample spectrum show spots of the same color at corresponding positions.

[0044] In some specific embodiments, at the ratio shift values ​​of 0.634, 0.633, and 0.648, the mugwort formula particles show red fluorescent spots, while the vinegar mugwort formula particles have no spots.

[0045] This method has good durability to different humidity. Compared with the reference sample, the test sample and the reference sample show spots of the same color at corresponding positions.

[0046] In some specific embodiments, the mugwort formula particles show red fluorescent spots at the ratio shift values ​​of 0.638, 0.610, and 0.667, while the vinegar mugwort formula particles have no spots.

[0047] The mugwort formula granules have red fluorescent spots at the above-mentioned ratio shift value (Rf), while the vinegar mugwort formula granules do not have such spots. The fluorescent spots can be used to distinguish the two.

[0048] The invention provides a thin layer chromatography identification method for mugwort and vinegar mugwort formula granules, comprising: A) pre-treating the mugwort formula granules to obtain a test solution; pre-treating the vinegar mugwort formula granules to obtain a vinegar mugwort formula granule solution; B) taking a scopolamine reference substance, adding methanol to dissolve it, and obtaining a reference substance solution;

[0049] C) Subjecting the test solution, vinegar wormwood formula granule solution, and reference solution to thin layer chromatography (TLC) using a silica gel G TLC plate and n-hexane-ethyl acetate-toluene as the developing solvent; D) Examining under ultraviolet light: If the test sample and reference medicinal material solutions exhibit fluorescent spots of the same color at locations where Rf is equal, and fluorescent spots are present at Rf = 0.625-0.741, the samples are wormwood formula granules; if no fluorescent spots are present at Rf = 0.625-0.741, the samples are vinegar wormwood formula granules. The present invention discloses the construction and application of a TLC identification method for wormwood and vinegar wormwood formula granules. By observing the TLC of the test sample, the presence of a red fluorescent spot at a shift value of 0.625-0.741 is used as the identification point for wormwood formula granules, while the absence of a red fluorescent spot at the same location is used as the identification point for vinegar wormwood formula granules. The established TLC method can quickly and effectively identify wormwood and vinegar wormwood formula granules. The method is highly specific, stable, precise, convenient, and easy to master. It allows for more standardized quality control of mugwort leaf and vinegar mugwort leaf granules, ensuring the efficacy of the drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0050] Figure 1 Spotting quantity investigation;

[0051] Figure 2 Specificity investigation;

[0052] Figure 3 shows the inspection of different thin layer plates - Tianjin Silida;

[0053] Figure 4 Investigation of different thin layer plates - Merck;

[0054] Figure 5 Survey of different thin layer plates - Qingdao Ocean;

[0055] Figure 6 Different temperatures -4°C;

[0056] Figure 7 Different temperatures -25°C;

[0057] Figure 8 Different temperatures 35°C;

[0058] Figure 9 Different humidity -32%;

[0059] Figure 10 Different humidity levels - 51%

[0060] Figure 11 Different humidity levels - 75%;

[0061] Figure 12 Different times of verification;

[0062] Figure 13 is a diagram showing the results of the method of Comparative Example 1;

[0063] FIG14 is a diagram showing the method results of Comparative Example 2;

[0064] FIG15 is a graph showing the results of comparative example 3 using a developing agent of n-hexane-ethyl acetate in a ratio of 9:1. DETAILED DESCRIPTION

[0065] To further illustrate the present invention, a thin layer identification method for mugwort and vinegar mugwort formula granules provided by the present invention is described in detail below in conjunction with examples.

[0066] Mortar, thin layer imaging system: CAMAG TLC Visualizer, silica gel G thin layer plate (Qingdao Ocean Chemical Plant, batch number: 20180527, Tianjin Silida Technology Co., Ltd., batch number: 191016, Merck, Germany, batch number: HX87183353)

[0067] Hexane, toluene, ethyl acetate, and methanol were all of analytical grade.

[0068] Ethanol, n-butanol, isopropanol, and glacial acetic acid were all analytical grade, and water was ultrapure water (prepared in the laboratory).

[0069] Scopoletin (China Food and Drug Inspection Institute, batch number: 110768-202105, purity 99.7%), mugwort formula granules (Sichuan New Green Pharmaceutical Technology Development Co., Ltd., batch numbers: 2208001; 2208002; 2208003) vinegar mugwort formula granules (Sichuan New Green Pharmaceutical Technology Development Co., Ltd., batch numbers: 2302081, 2302082, 2302083).

[0070] Example 1

[0071] Take 0.5g of this product, grind it finely, add 10ml of methanol and sonicate for 10 minutes, filter, evaporate the filtrate to dryness, dissolve the residue in 5ml of water, pass it through a D101 macroporous adsorption resin column (inner diameter 1cm, column height 14cm), elute with 20ml of 30% methanol, discard, and elute with 20ml of ethyl acetate. Collect the eluate, evaporate to dryness, and dissolve the residue in 1ml of ethyl acetate to prepare the test solution. For the reference solution of dongguan puer lactone, add methanol to prepare a solution containing 40μg per 1ml, which is used as the reference solution.

[0072] According to the thin layer chromatography method (Chinese Pharmacopoeia 2020 Edition General Chapter 0502), a 10 μl spot of the test solution and the reference solution was aspirated and spotted on the same silica gel G thin layer plate. The plate was developed with n-hexane-ethyl acetate-toluene (1:3:2) as the developing solvent. The plate was removed, dried, and examined under ultraviolet light (365 nm). In the chromatogram of the test sample, a main spot of the same color appeared at the corresponding position in the chromatogram of the reference sample.

[0073] Example 2 Methodological Investigation

[0074] 2.1 Sampling quantity inspection

[0075] Under the experimental conditions described above, 5μl, 10μl, and 15μl of the dongliangpolulide reference solution, mugwort granule solution, and vinegar mugwort granule solution, respectively, were spotted onto the same silica gel G thin-layer plate. The results are shown in Figure 1. The results show that when 5-15μl of the reference solution and test solution were spotted, the spots developed clearly and the resolution was good, establishing a spot size of 5-15μl. Furthermore, at a shift ratio of 0.674, the mugwort granules exhibited a red fluorescent spot, while the vinegar mugwort granules showed no spot, serving as a discriminant between the mugwort granules and the vinegar mugwort granules. The presence of a red fluorescent spot at a shift ratio of 0.674 can be used as a discriminant.

[0076] Figure 1: Sample quantity survey. Note: 1-3 are scopoletin reference solutions; 4-6 are mugwort granule solutions; 7-9 are vinegar mugwort granule solutions.

[0077] 2.2 Specificity Investigation

[0078] Following the above sample preparation method, negative solutions, scopoletin reference solutions, mugwort granule solutions, and vinegar mugwort granule solutions were prepared and subjected to thin-layer chromatography (TLC) identification. The results are shown in Figure 2. As can be seen, the negative sample showed no interference with the mugwort and vinegar mugwort granule samples, demonstrating good specificity. Furthermore, at a ratio shift of 0.741, the mugwort granules exhibited red fluorescent spots, while the vinegar mugwort granules showed no spots. Figure 2: Specificity evaluation. Note: 1 is the negative solution; 2 is the scopoletin reference solution; 3 is the mugwort granule solution; 4 is the vinegar mugwort granule solution.

[0079] 2.3 Durability Investigation

[0080] 2.3.1 Comparison of different thin layer plates

[0081] Tests were conducted using tailorable thin-layer chromatography plates from Tianjin Silida Technology Co., Ltd., prefabricated silica gel GF254 plates from Merck, and a branch of Qingdao Ocean Chemical Plant, according to the proposed test method. (See Figures 3-5.) The results show that the relative shift values ​​(Rf) of the red fluorescent spots at 8.5-11 cm from the origin on the three thin-layer plates were 0.662, 0.659, and 0.655, respectively, all meeting the identification requirements. Furthermore, the mugwort formula granules exhibited red fluorescent spots at Rf values ​​of 0.613, 0.623, and 0.504, while the vinegar mugwort formula granules showed no spots.

[0082] The results are shown in Figure 3. Figure 3 shows the results of different thin-layer plate tests at Tianjin Silida. Note: 1 is a scopoletin reference solution; 2 is a solution of mugwort granules; 3 is a solution of vinegar-mugwort granules. Figure 4 shows the results of different thin-layer plate tests at Merck. Note: 1 is a scopoletin reference solution; 2 is a solution of mugwort granules; 3 is a solution of vinegar-mugwort granules. Figure 5 shows the results of different thin-layer plate tests at Qingdao Ocean. Note: 1 is a scopoletin reference solution; 2 is a solution of mugwort granules; 3 is a solution of vinegar-mugwort granules.

[0083] 2.3.2 Comparison of different temperatures

[0084] After spotting, the thin-layer plates were developed at 4°C, 25°C, and 35°C, respectively. Figures 6 to 8 show that this method is well adapted to different temperatures. At the ratio shift values ​​of 0.634, 0.633, and 0.648, the mugwort granules exhibited red fluorescent spots, while the vinegar mugwort granules showed no spots. Figure 6: Different temperatures at 4°C; Note: 1 is a scopoletin reference solution; 2 is a mugwort granule solution; 3 is a vinegar mugwort granule solution. Figure 7: Different temperatures at 25°C; Note: 1 is a scopoletin reference solution; 2 is a mugwort granule solution; 3 is a vinegar mugwort granule solution. Figure 8: Different temperatures at 35°C; Note: 1 is a scopoletin reference solution; 2 is a mugwort granule solution; 3 is a vinegar mugwort granule solution.

[0085] 2.3.3 Comparison of different humidity levels

[0086] After spotting, the thin-layer plates were developed at humidity levels of 32%, 51%, and 75%, respectively, as shown in Figures 9-11. As can be seen from the figures, this method exhibits good adaptability to varying humidity levels. The mugwort granules exhibited red fluorescent spots at ratio shifts of 0.638, 0.610, and 0.667, while the vinegar mugwort granules showed no spots. Figure 9: Different humidity levels - 32%, Note: 1: Scopoletin reference solution; 2: Mugwort granule solution; 3: Vinegar mugwort granule solution; Figure 10: Different humidity levels - 51%, Note: 1: Scopoletin reference solution; 2: Mugwort granule solution; 3: Vinegar mugwort granule solution; Figure 11: Different humidity levels - 75%, Note: 1: Scopoletin reference solution; 2: Mugwort granule solution; 3: Vinegar mugwort granule solution.

[0087] 2.3.4 Verification

[0088] Scopoletin reference solution, mugwort granule solution, and vinegar mugwort granule solution were spotted onto the same silica gel GF254 thin layer plate and analyzed according to the established method. The experimental results are shown in Figure 12. The results show that the mugwort granule exhibited a red fluorescent spot at a ratio shift (Rf) of 0.625, while the vinegar mugwort granule did not. This fluorescent spot can be used to distinguish the two. Figure 12: Verification of different batches. Note: 1 is the scopoletin reference solution; 2-4 are the mugwort granule solutions; 5-7 are the vinegar mugwort granule solutions.

[0089] 2.4 Determination of the Rf value of the identification point of mugwort formula granules and vinegar mugwort formula granules

[0090] The data of various ratio shift values ​​in the thin layer chromatography methodology investigation are summarized and the results are shown in Table 1. Based on the summary results, it is determined that the ratio shift value (Rf) of the identification spot of the Artemisia argyi formula granules should be within the range of 15%, and the specified value is 0.654.

[0091] Table 1 Summary of methodological investigation ratios

[0092] The results showed that at the ratio shift value of 0.654, there were no fluorescent spots in the vinegar and mugwort formula granules, while the other mugwort formula granules had obvious red fluorescent spots, which could be used as an identification point to distinguish between the mugwort formula granules and the vinegar and mugwort formula granules.

[0093] Comparative Example 1

[0094] Add 4g of this product to 10mL of chloroform and sonicate for 15 minutes. Allow to stand for another hour. Filter the filtrate to dryness and dissolve in 1mL of chloroform to prepare the test solution. Separately, prepare a scopolamine reference solution with methanol to create a solution containing 40μg per 1mL. This serves as the reference solution. Aspirate 4μL of this solution. Spot each sample onto a silica gel G thin-layer plate, develop with cyclohexane-ether-ethyl acetate (20:5.5:2.5), remove, air dry, spray with 10% ethanolic sulfuric acid solution, and bake at 100°C for approximately 10 minutes for color development. Fluorescent spots of the same color will appear in the chromatogram of the test sample at the corresponding position in the chromatogram of the reference sample. The experimental results are shown in Figure 13. Figure 13 shows the results of the method in Comparative Example 1. Note: 1 is the scopolamine reference solution; 2-4 are the solutions of the wormwood formula granules; 5-8 are the solutions of the vinegar wormwood formula granules.

[0095] The results showed that using the above-mentioned thin layer identification method, there were no obvious spots on the mugwort leaves and the vinegar mugwort formula granules, and the two could not be identified.

[0096] Comparative Example 2

[0097] Grind 1g of this product, add 30ml of 70% ethanol and 2ml of hydrochloric acid, heat under reflux for 1 hour, filter, and dry the filtrate. Dissolve the residue in 20ml of water. Shake and extract twice with 20ml of acid and ester. Combine the ethyl acetate solutions, evaporate to dryness, and dissolve the residue in 1ml of methanol to prepare the test solution. Separately, take 3g of Artemisia argyi as a control medicinal material, add 50ml of water, boil for 30 minutes, filter, and evaporate the filtrate to dryness. Add 30ml of 70% ethanol and 2ml of hydrochloric acid to the residue and prepare a control medicinal material solution in the same manner. Also, take the dongliang lactone reference material and add methanol to a solution containing 40μg per 1ml. This is used as the reference solution. According to the thin layer chromatography method (General Rules 0502 of the 2020 edition of the Chinese Pharmacopoeia), 5ul of each of the above solutions was taken and spotted on the same silica gel G thin layer plate, and chloroformane-methanol-formic acid (20:3.5:2) was used as the developing agent. It was developed, taken out, dried, sprayed with aluminum chloride test solution, blown dry with hot air, and inspected under ultraviolet light (365nm). In the chromatogram of the test sample, fluorescent spots of the same color appeared at the corresponding positions of the chromatograms of the reference sample and the reference medicinal material. The experimental results are shown in Figure 14. Figure 14 is a diagram of the method results of Comparative Example 2. Note: 1 is a scopoletin reference solution; 2 is an argyi reference medicinal material solution; 3 is an argyi formula granule solution; 4 is a vinegar argyi formula granule solution.

[0098] The results showed that using the above-mentioned thin layer identification method, there were no obvious different spots between mugwort leaves and vinegar mugwort formula granules, and the two could not be distinguished.

[0099] Comparative Example 3

[0100] Take 1g of this product, grind it finely, add 50ml of ether, sonicate for 30 minutes, filter, evaporate the filtrate, and dissolve the residue in 1ml of ethyl acetate to prepare the test solution. According to the thin layer chromatography method (General Method 0502), take 10μl of the above solution and spot it on a silica gel G thin layer plate. Use n-hexane-ethyl acetate (9:1) as the developing solvent. Develop the plate, remove it, air dry, and use 10% ethanolic sulfuric acid solution as the color developer. After heating at 105°C, examine it under ultraviolet light (365nm). The experimental results are shown in Figure 15. Figure 15 shows the results of the developing solvent n-hexane-ethyl acetate (9:1). Note: 1 is the scopoletin reference solution; 2 is the solution of the wormwood formula granules; 3 is the solution of the vinegar wormwood formula granules.

[0101] The results showed that using the above-mentioned thin layer identification method, there were no obvious different spots between mugwort leaves and vinegar mugwort formula granules, and the two could not be distinguished.

[0102] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as within the scope of protection of the present invention.

Claims

1. A thin layer identification method for wormwood and wormwood vinegar formula particles, comprising: A) taking the wormwood formula granules for pretreatment to obtain a test solution; Pre-treating the vinegar wormwood formula granules to obtain a vinegar wormwood formula granule solution; B) Take the scopolamine reference substance, add methanol to dissolve it, and obtain a reference substance solution; C) The test solution, vinegar wormwood formula granule solution and reference solution were subjected to thin layer chromatography, the thin layer plate was a silica gel G thin layer plate; the developing solvent was n-hexane-ethyl acetate-toluene; D) Check under ultraviolet light. If the test sample and the control medicinal material solution have fluorescent spots of the same color at the same Rf, and there are fluorescent spots at Rf = 0.625-0.741, then it is the mugwort formula granules; If there is no fluorescent spot at Rf=0.625~0.741, it is vinegar and wormwood formula granules.

2. The method according to claim 1, characterized in that The step A) of pre-treating the wormwood formula granules or vinegar wormwood formula granules is specifically as follows: a) adding methanol to the wormwood formula granules or vinegar wormwood formula granules, ultrasonicating, filtering, evaporating, dissolving the residue in water, passing through a D101 macroporous adsorption resin column, eluting with 30% methanol, discarding the eluent, eluting with ethyl acetate, and collecting the eluent; b) Recover the solvent from the eluate until it is dry, and dissolve the residue in ethyl acetate to obtain the product.

3. The method according to claim 1, characterized in that Step a) The ultrasonic time is 5 to 10 minutes; the mass volume ratio of the wormwood formula granules or vinegar wormwood formula granules, methanol, water, 30% methanol and ethyl acetate is 0.5g:10mL:5mL:20mL:20mL; The inner diameter of the D101 macroporous adsorption resin is 1 cm, and the column height is 14 cm.

4. The method according to claim 1, characterized in that: In the step B), the mass volume ratio of the scopolamine reference substance and methanol is 40 μg:1 mL.

5. The method according to claim 1, characterized in that The sample spotting volume of the thin layer chromatography test solution, vinegar wormwood formula granule solution and reference solution is 5 to 15 μL.

6. The method according to claim 1, characterized in that Step C) The mass ratio of n-hexane-ethyl acetate-toluene is 1:3:

2.

7. The method according to claim 1, characterized in that The wavelength of the ultraviolet light in step D) is 365 nm.

8. The method according to claim 1, characterized in that In step D), the silica gel G thin layer plate is a GF254 thin layer plate.

9. The method according to claim 8, characterized in that Step D) checking the temperature to be 4°C to 35°C; the checking humidity to be 32% rh to 75% rh.

10. The method according to claim 1, characterized in that The Rf value in step D) is 0.654.

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