Thin-layer identification method for honeysuckle in Shuanghuanglian preparation

Abstract

The invention discloses a thin-layer identification method for honeysuckle in a Shuanghuanglian preparation. According to the method, an upper-layer solution of butyl acetate, formic acid and water with the volume ratio of 14: 5: 5 is used as a developing solvent, after two times of developing, the separation degree of each spot is good, and methodology verifies that the method is good in specificity and durability. The thin-layer identification of honeysuckle is carried out by adopting a test solution, a reference medicinal material solution, a chlorogenic acid reference solution and a negative reference solution, and an accurate identification result can be obtained. Single identification of only one component of chlorogenic acid is increased to simultaneous identification of multiple components of a reference medicinal material, so that exclusive identification of the Shuanghuanglian preparation can be increased, and the quality controllability of the Shuanghuanglian preparation is improved.

Description

Technical Field

[0001] The invention belongs to the technical field of traditional Chinese medicines, and particularly relates to a thin layer identification method for honeysuckle in a Shuanghuanglian preparation. Background Art

[0002] Existing TLC methods for Shuanghuanglian preparations, such as Shuanghuanglian for injection (lyophilized), Shuanghuanglian granules, Shuanghuanglian eye drops, and Shuanghuanglian granules, primarily use baicalin and chlorogenic acid as reference substances to identify Scutellaria baicalensis and honeysuckle in the preparations; and forsythiaside and forsythia suspensa as reference herbs to identify forsythia suspensa. Using chlorogenic acid as a reference substance for TLC identification of honeysuckle suffers from poor specificity. In the Chinese Pharmacopoeia alone, chlorogenic acid is used as a reference substance for TLC identification of varieties such as honeysuckle, Eucommia ulmoides leaves, Artemisia capillaris, and plum blossoms. Therefore, using chlorogenic acid as a single reference substance for specific identification of honeysuckle is difficult.

[0003] Shuanghuanglian Inhalation Solution is a traditional Chinese medicine aerosol inhalation preparation developed through process optimization based on Shuanghuanglian Injection. Its formulation is identical to Shuanghuanglian Injection, consisting of honeysuckle, scutellaria, and forsythia. Currently, a thin-layer chromatography (TLC) identification method for honeysuckle has not been established. If the original TLC method for Shuanghuanglian Injection is used, the specificity for honeysuckle will be poor. Therefore, further investigation is needed into the sample handling and development conditions for TLC identification.

[0004] The information disclosed in this background technology section is only intended to enhance understanding of the overall background of the invention and should not be regarded as an admission or any form of suggestion that the information constitutes the prior art already known to a person skilled in the art. Summary of the Invention

[0005] In order to solve the technical problems of poor specificity and poor separation in the thin layer identification method of honeysuckle in Shuanghuanglian preparations in the prior art, the present invention provides a thin layer identification method of honeysuckle in Shuanghuanglian preparations, which has good specificity, durability and high separation.

[0006] To achieve the above object, the present invention provides a thin layer identification method for honeysuckle in Shuanghuanglian preparation, comprising the following steps:

[0007] S1. Preparation of test solution: Take 2-5 ml of Shuanghuanglian liquid preparation, evaporate the solvent to dryness in a water bath or rotary evaporator, and dissolve the residue in methanol to prepare the test solution; or take Shuanghuanglian solid preparation and dissolve it in methanol to prepare the test solution;

[0008] S2. Preparation of negative sample solution: Take 2-5 ml of the negative sample solution lacking honeysuckle and prepare a negative sample solution according to the method in S1;

[0009] S3. Preparation of control medicinal material solution: taking honeysuckle control medicinal material, adding water, heating and refluxing extraction, filtering, evaporating to dryness, adding methanol to dissolve the residue to prepare a control medicinal material solution;

[0010] S4. Preparation of reference solution: Take chlorogenic acid reference and add methanol to make a solution containing 0.5 mg of chlorogenic acid per 1 ml of methanol to prepare a reference solution;

[0011] S5. Thin layer chromatography test: 1 to 2 μL of each of the above four solutions were taken and spotted on the same silica gel G thin layer plate. The upper layer solution of butyl acetate-formic acid-water with a volume ratio of 14:5:5 was used as the developing agent. After secondary development, the plate was taken out, dried with hot air, and examined under ultraviolet light (365 nm). In the chromatogram of the test sample, fluorescent spots of the same color were displayed at the corresponding positions in the chromatogram of the reference sample and the chromatogram of the reference medicinal material.

[0012] In some preferred embodiments, in step S3, the time of the heating reflux extraction is no less than 1.5 h.

[0013] In some preferred embodiments, in step S5, the amount of the absorbed solution is 1 μL.

[0014] In some preferred embodiments, in step S5, the secondary expansion distance is 7 to 9 cm, more preferably 8 cm.

[0015] In some preferred embodiments, the Shuanghuanglian solid preparation is selected from one of tablets, granules, and freeze-dried powder; the Shuanghuanglian liquid preparation is selected from one of injection, eye drops, and nebulized inhalation solution; more preferably, the Shuanghuanglian preparation is a Shuanghuanglian nebulized inhalation solution.

[0016] The preparation method of the Shuanghuanglian aerosol inhalation solution comprises the following steps:

[0017] (1) Scutellaria baicalensis extract, honeysuckle extract and forsythia suspensa extract are extracted from Scutellaria baicalensis, honeysuckle and forsythia suspensa respectively;

[0018] (2) dissolving the scutellaria baicalensis extract in hot water and a strong base, adding the honeysuckle extract and the forsythia suspensa extract, stirring evenly, and filtering to obtain solution A;

[0019] (3) adding an osmotic pressure regulator to the solution A and filtering to obtain a Shuanghuanglian atomized inhalation solution;

[0020] Wherein, the temperature of the hot water in step (2) is 70-90° C.; and the strong alkali in step (2) is selected from one or both of sodium hydroxide and potassium hydroxide.

[0021] In some preferred embodiments, the method for preparing the Shuanghuanglian aerosol inhalation solution comprises the following steps:

[0022] (1) Preparation of Scutellaria baicalensis extract

[0023] 1) Take 250g of Scutellaria baicalensis root and decoct it twice in water for 1 hour each time. Filter the decocted decoctants in batches and combine the filtrates. Adjust the pH of the filtrates to 1.9 with hydrochloric acid solution, incubate at 80°C for 30 minutes, let stand for 24 hours, filter, and collect precipitate A.

[0024] 2) Add 8 times the amount of water to Precipitate A, stir, adjust the pH to 7.0 with 40% sodium hydroxide solution, add ethanol to 50% alcohol content, stir to dissolve, filter, adjust the pH of the filtrate to 2.0 with hydrochloric acid solution, incubate at 60°C for 30 minutes, let stand for 12 hours, filter, and collect Precipitate B;

[0025] 3) Precipitate B was washed with ethanol until the pH reached 4.0, 10 times the amount of water was added, stirred, and the pH was adjusted to 6.0 with 40% sodium hydroxide solution. 0.5% activated carbon was added and stirred thoroughly. The mixture was incubated at 50°C for 30 minutes, 1 time the amount of ethanol was added and stirred evenly. The mixture was immediately filtered. The pH of the filtrate was adjusted to 2.0 with hydrochloric acid solution, incubated at 60°C for 30 minutes, allowed to stand for 12 hours, filtered, and Precipitate C was collected.

[0026] 4) washing the precipitate C with a small amount of ethanol and drying it at 60°C to obtain the Scutellaria baicalensis extract;

[0027] (2) Preparation of honeysuckle extract and forsythia suspensa extract

[0028] Take 250g of honeysuckle and 500g of forsythia, soak them in water for 30 minutes, decoct them twice, each time for 1 hour, filter them in batches, combine the filtrates, concentrate them to a relative density of 1.20 at 70°C, cool them to 40°C, slowly add ethanol to make the alcohol content reach 75%, stir them thoroughly, let them stand for 12 hours, filter the supernatant, recover the ethanol until there is no alcohol taste, add 4 times the amount of water, adjust the pH to 7.0, stir them thoroughly and heat them to boiling, let them stand for 48 hours, filter the supernatant, concentrate them to a relative density of about 1.13 at 70°C, cool them to 40°C, add ethanol to make the alcohol content reach 85%, let them stand for 12 hours, filter the supernatant, recover the ethanol until there is no alcohol taste, and obtain the honeysuckle extract and forsythia extract;

[0029] (3) Preparation of Shuanghuanglian atomized inhalation solution

[0030] Take Scutellaria baicalensis extract, add appropriate amount of 80℃ hot water, add sodium hydroxide solution to dissolve, add honeysuckle extract and forsythia suspensa extract, add water to 1000ml, add appropriate amount of activated carbon, heat to a slight boil, cool, filter, refrigerate and let stand, filter, ultrafilter, add water to 1000ml, add appropriate amount of sodium chloride, filter, and obtain Shuanghuanglian nebulized inhalation solution.

[0031] In some preferred embodiments, the thin layer identification method of honeysuckle in the Shuanghuanglian aerosol inhalation preparation comprises the following steps:

[0032] S1. Preparation of test solution: Take 4 ml of Shuanghuanglian nebulized inhalation solution, evaporate to dryness in a water bath, and dissolve the residue in 5 ml of methanol to prepare the test solution;

[0033] S2. Preparation of negative sample solution: Take another 4 ml of the negative sample solution lacking honeysuckle and prepare a negative sample solution according to the method in S1;

[0034] S3. Preparation of control medicinal material solution: 0.5 g of honeysuckle control medicinal material was taken, 30 ml of water was added, refluxed and extracted for 1.5 h, filtered, evaporated to dryness, and the residue was dissolved in 5 ml of methanol to prepare a control medicinal material solution;

[0035] S4. Preparation of reference solution: Take the chlorogenic acid reference substance and add methanol to make a solution containing 0.5 mg per 1 ml to prepare the reference solution;

[0036] S5. Detection by thin layer chromatography: 1 μL of each of the above four solutions was taken and spotted on the same silica gel G thin layer plate. A secondary development was performed using an upper layer solution of butyl acetate-formic acid-water with a volume ratio of 14:5:5 as the developing agent. The development distance was 8 cm. The plates were taken out, dried with hot air, and examined under an ultraviolet lamp (365 nm). In the chromatogram of the test sample, fluorescent spots of the same color were displayed at the corresponding positions in the chromatogram of the reference sample and the chromatogram of the reference medicinal material.

[0037] Compared with the prior art, the present invention has the following beneficial effects:

[0038] The thin layer chromatography identification method provided by the present invention uses a honeysuckle control medicinal material and a chlorogenic acid reference substance to perform thin layer chromatography identification on the honeysuckle in the Shuanghuanglian preparation, and uses an upper layer solution of butyl acetate-formic acid-water with a volume ratio of 14:5:5 as a developing agent, and performs two developments. The spots on the developed thin layer chromatogram are round and clear, the separation of each spot is high, the Rf value is within the range of 0.3 to 0.7, the negative is interference-free, and the specificity is strong. According to methodological verification, the method has good specificity and durability. The thin layer chromatography identification method provided by the present invention is increased from the single identification of only one component, chlorogenic acid, to the simultaneous identification of multiple components of the control medicinal material, which can add exclusive identification to the Shuanghuanglian preparation and improve the quality controllability of the Shuanghuanglian preparation. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1 This is the thin layer chromatogram of honeysuckle in Shuanghuanglian preparation;

[0040] Figure 2 This is the TLC of honeysuckle under different extraction methods of the reference herb;

[0041] Figure 3 This is a thin layer chromatogram of honeysuckle with a spread of 7 cm;

[0042] Figure 4 This is a thin layer chromatogram of honeysuckle with a spread of 8 cm;

[0043] Figure 5 This is a thin layer chromatogram of honeysuckle with a spread of 9 cm;

[0044] Figure 6 This is a thin layer chromatogram of honeysuckle developed on a thin layer silica gel G plate produced by the Qingdao Ocean Chemical Plant Branch;

[0045] Figure 7 This is a thin layer chromatogram of honeysuckle developed on a thin layer silica gel G plate produced by Merck Company in the United States;

[0046] Figure 8 This is a thin layer chromatogram of honeysuckle developed on a thin layer silica gel G plate produced by Yantai Chemical Industry Research Institute;

[0047] Figure 9 This is a thin layer chromatogram of honeysuckle developed under high humidity environment;

[0048] Figure 10 This is a thin layer chromatography of honeysuckle developed under low temperature and low humidity environment;

[0049] Figure 11 This is the thin layer chromatogram of honeysuckle when the development number is 1. DETAILED DESCRIPTION

[0050] The specific embodiments of the present invention are described in detail below with reference to the accompanying drawings, but it should be understood that the protection scope of the present invention is not limited by the specific embodiments.

[0051] The following describes the embodiments of the present invention through specific examples. Those skilled in the art will readily understand the other advantages and benefits of the present invention from the disclosure herein. The present invention may also be implemented or applied through various other specific embodiments, and the details in this specification may be modified or altered based on different viewpoints and applications without departing from the spirit of the present invention.

[0052] The instruments, reagents and their sources used in the following examples are as follows: dark box UV projection reflectometer, WFH-201B, Shanghai Jingke Industrial Co., Ltd.; electronic analytical balance: Sartorius Scientific Instruments (Beijing) Co., Ltd. SQP (one ten-thousandth); silica gel G thin layer plate, purchased from Yantai Chemical Industry Research Institute, Qingdao Ocean Chemical Plant, and Merck, Germany; polyamide film, purchased from Luqiao Sijia Biochemical Plastic Factory, Taizhou City, Zhejiang Province and Wuhan Xinsrui Technology Co., Ltd.

[0053] Methanol, butyl acetate, and formic acid were purchased from Guangzhou Chemical Reagent Factory and Tianjin Damao Chemical Reagent Factory.

[0054] Chlorogenic acid reference substance (batch number: 110753-201817, purity: 96.8%, China Food and Drug Inspection Institute), honeysuckle reference medicinal material (batch number: 121060-201608, China Food and Drug Inspection Institute), Shuanghuanglian nebulized inhalation solution (homemade, batch number: CP-201025-01), and negative sample lacking honeysuckle (homemade, batch number CP-201105-02).

[0055] Example 1: Establishment of a Thin Layer Identification Method for Honeysuckle in Shuanghuanglian Atomized Inhalation Preparation

[0056] Take 4 ml of Shuanghuanglian nebulized inhalation solution, evaporate to dryness, and dissolve the residue in 5 ml of methanol to prepare the test solution;

[0057] Take another 4 ml of the negative sample solution lacking honeysuckle and prepare the negative sample solution in the same way;

[0058] Take 0.5 g of honeysuckle as a control medicinal material, add 30 ml of water, reflux extract for 1.5 hours, filter, evaporate the filtrate to dryness, add 5 ml of methanol to dissolve the residue, and prepare a control medicinal material solution;

[0059] Prepare a reference solution by adding methanol to a 0.5 mg chlorogenic acid reference solution per ml.

[0060] Thin layer chromatography detection: aspirate 1μl, 1.5μl, 2μl of the test solution, 1μl of the honeysuckle negative sample solution, the control medicinal material solution, and the reference solution, respectively, and spot them on the same silica gel G thin layer plate, use the upper layer solution of butyl acetate-formic acid-water with a volume ratio of 14:5:5 as the developing solvent, and develop twice, with the development distance of 8cm. Take out, blow dry with hot air, and examine under ultraviolet light (365nm). The results are as follows Figure 1 . Figure 1 The thin layer chromatogram of honeysuckle shown in the figure is a Qingdao silica gel G thin layer plate with a specification of 10×10 cm. The thin layer chromatogram is developed at 20.5° C. and RH 51%. Figure 1 In the table, 1 is 1 μl of chlorogenic acid reference solution, 2 is 1 μl of honeysuckle reference medicinal material solution, 3 is 1 μl of test solution, 4 is 1.5 μl of test solution, 5 is 2 μl of test solution, and 6 is 1 μl of honeysuckle-deficient negative sample solution.

[0061] Figure 1 It shows that in the thin layer chromatogram of the test sample, fluorescent spots of the same color appear at the corresponding positions in the chromatogram of the reference sample and the chromatogram of the reference medicinal material. The spots are round, clear, and highly separated. The Rf value is in the range of 0.3 to 0.7, and is negative without interference and has strong specificity.

[0062] Through the investigation of different sample volumes, the results showed that when the sample volume was 1μl, the spot separation was the best; when the sample volume was 1.5μl or 2μl, the spot separation was poor.

[0063] Example 2: Investigation of the treatment methods of control medicinal materials

[0064] Take 4 ml of Shuanghuanglian nebulized inhalation solution, evaporate to dryness, and dissolve the residue in 5 ml of methanol to prepare the test solution;

[0065] Take another 4 ml of the negative sample solution lacking honeysuckle and prepare the negative sample solution in the same way;

[0066] Take 0.5 g of honeysuckle control medicinal material, add 30 ml of water, reflux extraction for 1.5 hours, filter, evaporate the filtrate to dryness, add 5 ml of methanol to dissolve the residue, and prepare control medicinal material solution ① (water boiling method);

[0067] Take 0.5 g of honeysuckle control medicinal material, add 5 ml of methanol and ultrasonicate for 10 minutes to prepare control medicinal material solution ② (ultrasonic method);

[0068] Prepare a reference solution by adding methanol to a 0.5 mg chlorogenic acid reference solution per ml.

[0069] Thin layer chromatography detection: Pipette 1μl of the test solution, 1μl of the negative sample solution lacking honeysuckle, 1μl of the control medicinal material solution ①, 1μl of the control medicinal material solution ②, and 1μl of the reference solution, respectively, and spot them on the same silica gel G thin layer plate. Use the upper layer solution of butyl acetate-formic acid-water with a volume ratio of 14:5:5 as the developing solvent, and perform secondary development with a development distance of 8cm. Take out, blow dry with hot air, and examine under ultraviolet light (365nm). The results are as follows Figure 2 . Figure 2 The thin layer chromatogram of honeysuckle shown here is a Qingdao silica gel G thin layer plate with a specification of 10×10 cm. The thin layer chromatogram is developed at 20.1° C. and RH 51%. Figure 2 In the table, 1 is 1 μl of chlorogenic acid reference solution; 2 is 1 μl of honeysuckle reference medicinal material solution extracted by boiling method ①; 3 is 1 μl of honeysuckle reference medicinal material solution extracted by ultrasonic method ②; 4-6 are 1 μl of test solution; 7 is 1 μl of honeysuckle negative sample solution.

[0070] Figure 2The results showed that the water-boiling method for extracting the control medicinal material produced more spot information and higher spot clarity than direct ultrasonic extraction of the control medicinal material. Ultrasonic extraction lacked one fluorescent spot, confirming the use of the water-boiling method for the control medicinal material. Furthermore, experiments showed that if the extraction time for the honeysuckle water-boiling method for extracting the gold and silver control medicinal material was less than 1.5 hours, such as 0.5 hours, the spot clarity was also insufficient, and one fluorescent spot was also missing. Therefore, the water-boiling method was determined to be used for extracting the control medicinal material, with a reflux time of at least 1.5 hours.

[0071] Implementation Case 3: Distance Investigation

[0072] Take 4 ml of Shuanghuanglian nebulized inhalation solution, evaporate to dryness, and dissolve the residue in 5 ml of methanol to prepare the test solution;

[0073] Take another 4 ml of the negative sample solution lacking honeysuckle and prepare the negative sample solution in the same way;

[0074] Take 0.5 g of honeysuckle control medicinal material, add 30 ml of water, reflux extraction for 1.5 hours, filter, evaporate the filtrate to dryness, add 5 ml of methanol to dissolve the residue, and prepare a control medicinal material solution;

[0075] Prepare a reference solution by adding methanol to a 0.5 mg chlorogenic acid reference solution per ml.

[0076] Detection by thin layer chromatography: aspirate 1 μl of the test solution, take 1 μl of the honeysuckle negative sample solution, the control medicinal material solution, and the reference substance solution, respectively, and spot them on the same silica gel G thin layer plate, use the upper layer solution of butyl acetate-formic acid-water solution with a volume ratio of 14:5:5 as the developing solvent, and develop twice. The two development distances are the same, namely 7 cm, 8 cm, and 9 cm. Take out, blow dry with hot air, and examine under ultraviolet light (365 nm). The thin layer chromatogram when the development distance is 7 cm is as follows Figure 3 The thin layer chromatogram when the development distance is 8 cm is as follows Figure 4 The thin layer chromatogram when the development distance is 9 cm is as follows Figure 5 . Figures 3-5 The thin layer chromatograms of honeysuckle shown were developed using Qingdao silica gel G thin layer plates with a specification of 10×10 cm. The thin layer chromatograms were developed at 20.8° C. and RH 53%. Figures 3-5 In the table, 1 is 1 μl of chlorogenic acid reference solution; 2 is 1 μl of honeysuckle reference medicinal material solution; 3-5 are 1 μl of test solution; and 6 is 1 μl of honeysuckle-deficient negative sample solution.

[0077] The test results show that when the expansion distance is 8 cm, the thin layer spots are clear and the separation is good. When the expansion distance is 7 cm, the spot separation is poor. When the expansion distance is 9 cm, the expansion time is long and the separation is not good. Therefore, the expansion distance is determined to be 8 cm.

[0078] Implementation Case 4: Thin Layer Methodology Durability Investigation

[0079] According to the TLC identification method of Example 1, a durability test was conducted, and thin-layer silica gel G plates from different manufacturers (Qingdao Ocean Chemical Plant Branch, Merck, USA, Yantai Chemical Industry Research Institute) and TLCs under different temperature, relative humidity and other influencing factors were examined. Figure 6 、 Figure 7 、 Figure 8 These are thin layer chromatograms developed using thin layer silica gel G plates produced by Qingdao Ocean Chemical Plant Branch, Merck, USA, and Yantai Chemical Industry Research Institute. Figures 6-8 All the developments were carried out in an environment of 20°C and RH46%; Figure 9 This is a thin layer chromatogram developed under the environmental conditions of 20.0℃ and RH84%. Figure 10 This is a thin layer chromatogram developed under the environmental conditions of 3.5℃ and RH32%. Figure 9-10 The thin layer silica gel G plates used were all produced by Qingdao Ocean Chemical Plant Branch. Figure 6-10 In the figure, 1 is 1 μL of chlorogenic acid reference solution, 2 is 1 μL of honeysuckle reference medicinal material solution, 3-5 are 1 μL of test solution, and 6 is 1 μL of honeysuckle-deficient negative sample solution.

[0080] Figure 6-10 It is shown that by investigating the durability of thin layer plates from different manufacturers and in low temperature and low humidity, normal temperature and high humidity environments, the results show that under different influencing factors, this thin layer identification method has good durability and clear spots.

[0081] Comparative Example 1: The number of expansions is one

[0082] Take 4 ml of Shuanghuanglian nebulized inhalation solution, evaporate to dryness, and dissolve the residue in 5 ml of methanol to prepare the test solution;

[0083] Take another 4 ml of the negative sample solution lacking honeysuckle and prepare the negative sample solution in the same way;

[0084] Take 0.5 g of honeysuckle as a control medicinal material, add 30 ml of water, reflux extract for 1.5 hours, filter, evaporate the filtrate to dryness, add 5 ml of methanol to dissolve the residue, and prepare a control medicinal material solution;

[0085] Prepare a reference solution by adding methanol to a 0.5 mg chlorogenic acid reference solution per ml.

[0086] Thin layer chromatography detection: aspirate 1μl, 1.5μl, 2μl of the test solution, 1μl of the honeysuckle negative sample solution, the control medicinal material solution, and the reference solution, respectively, and spot them on the same silica gel G thin layer plate, use the upper layer solution of butyl acetate-formic acid-water with a volume ratio of 14:5:5 as the developing solvent, develop only once, and the development distance is 8cm. Take out, blow dry with hot air, and examine under ultraviolet light (365nm). The results are as follows Figure 11 . Figure 11 The thin layer chromatogram of honeysuckle shown in the figure is a Qingdao silica gel G thin layer plate with a specification of 10×10 cm. The thin layer chromatogram is developed at 20.5° C. and RH 51%. Figure 11 In the table, 1 is 1 μl of chlorogenic acid reference solution, 2 is 1 μl of honeysuckle reference medicinal material solution, 3 is 1 μl of test solution, 4 is 1.5 μl of test solution, 5 is 2 μl of test solution, and 6 is 1 μl of honeysuckle-deficient negative sample solution.

[0087] Figure 11 The results showed that the separation in the chromatogram of the test sample was poor, especially the chlorogenic acid spot was superimposed on the adjacent spots, and the spot information of the honeysuckle control medicinal material was incomplete.

Claims

1. A thin layer identification method for honeysuckle in Shuanghuanglian preparation, characterized in that: The method comprises the following steps: S1. Preparation of test solution: Take 2-5 ml of Shuanghuanglian liquid preparation, evaporate the solvent in a water bath or rotary evaporator, and dissolve the residue in methanol to prepare the test solution; or take Shuanghuanglian solid preparation, dissolve it in methanol to prepare the test solution; S2. Preparation of negative sample solution: Take 2-5 ml of the negative sample solution lacking honeysuckle and prepare a negative sample solution according to the method in S1; S3. Preparation of control medicinal material solution: taking honeysuckle control medicinal material, adding water, heating and refluxing extraction, filtering, evaporating to dryness, adding methanol to the residue to dissolve, and preparing control medicinal material solution; S4. Preparation of reference solution: Take chlorogenic acid reference substance and add methanol to prepare a solution containing 0.5 mg chlorogenic acid per 1 ml of methanol as the reference solution; S5. Thin layer chromatography detection: Pipette 1-2 μL of each of the above four solutions and spot them on the same silica gel G thin layer plate respectively. Use the upper layer solution of butyl acetate-formic acid-water with a volume ratio of 14:5:5 as the developing agent. After secondary development, take out, blow dry with hot air, and examine under a 365nm ultraviolet lamp. In the test sample chromatogram, fluorescent spots of the same color will be shown at the corresponding positions of the reference sample chromatogram and the reference medicinal material chromatogram.

2. The thin layer identification method according to claim 1, characterized in that: In step S3, the heating reflux extraction time is not less than 1.5 hours.

3. The thin layer identification method according to claim 1, characterized in that: In step S5, the amount of the absorbed solution is 1 μL.

4. The thin layer identification method according to claim 1, characterized in that: In step S5, the extension distance of the secondary expansion is 7-9 cm, preferably 8 cm.

5. The thin layer identification method according to claim 1, characterized in that: The Shuanghuanglian solid preparation is selected from any one of tablets, granules, and freeze-dried powder; the Shuanghuanglian liquid preparation is selected from any one of injection, eye drops, and nebulized inhalation solution, preferably a Shuanghuanglian nebulized inhalation solution.

6. A thin layer identification method for multiple components of honeysuckle extract in Shuanghuanglian preparation, characterized in that: The method comprises the following steps: S1. Preparation of the test solution: Take 4 ml of Shuanghuanglian atomized inhalation solution, evaporate it to dryness in a water bath, add 5 ml of methanol to the residue to dissolve it, and prepare the test solution; S2. Preparation of negative sample solution: Take another 4 ml of the negative sample solution lacking honeysuckle and prepare a negative sample solution according to the method in S1; S3. Preparation of control medicinal material solution: Take 0.5 g of honeysuckle control medicinal material, add 30 ml of water, reflux and extract for 1.5 h, filter, evaporate to dryness, add 5 ml of methanol to the residue to dissolve, and prepare a control medicinal material solution; S4. Preparation of reference solution: Take chlorogenic acid reference substance again, add methanol to make a solution containing 0.5 mg per 1 ml as the reference solution; S5. Detection by thin layer chromatography: 1 μL of each of the above four solutions were taken and spotted on the same silica gel G thin layer plate respectively. The upper layer solution of butyl acetate-formic acid-water with a volume ratio of 14:5:5 was used as the developing agent. Secondary development was performed with a development distance of 8 cm. The plates were taken out, dried with hot air, and examined under a 365nm ultraviolet lamp. In the chromatogram of the test sample, fluorescent spots of the same color were displayed at the corresponding positions of the chromatogram of the reference sample and the chromatogram of the reference medicinal material.

7. The thin layer identification method according to claim 5 or 6, characterized in that: The preparation method of the Shuanghuanglian atomized inhalation solution comprises the following steps: (1) extracting from Scutellaria baicalensis, honeysuckle and Forsythia suspensa to obtain Scutellaria baicalensis extract, honeysuckle extract and Forsythia suspensa extract respectively; (2) taking the scutellaria baicalensis extract, adding hot water and a strong base to dissolve it, then adding the honeysuckle extract and the forsythia suspensa extract, stirring evenly, and filtering to obtain a solution A; (3) adding an osmotic pressure regulator to the solution A and filtering to obtain a Shuanghuanglian atomized inhalation solution; Wherein, the temperature of the hot water in step (2) is 70-90° C.; the strong alkali in step (2) is selected from one or both of sodium hydroxide and potassium hydroxide.

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