Thin-layer identification method for honeysuckle medicinal material in phlegm-heat clearing preparation

Abstract

The invention discloses a thin-layer identification method for a honeysuckle medicinal material in a phlegm-heat-clearing preparation, which comprises the following steps: (1) pretreating the phlegm-heat-clearing preparation to obtain a test solution, and pretreating a reference medicinal material or a reference substance to obtain a reference medicinal material solution or a reference substance solution; and (2) carrying out thin-layer chromatography identification analysis on the pair of test sample solutions, the honeysuckle-lacking phlegm-heat clearing preparation negative blank sample solution, the reference medicinal material solution and / or the reference substance solution, wherein in the step of thin-layer chromatography identification analysis, an upper-layer solution of butyl acetate, formic acid and water is used as a developing solvent, and the volume ratio of butyl acetate to formic acid to water is (1-10): (1-10): (1-10). According to the thin-layer identification method disclosed by the invention, the honeysuckle medicinal material in the phlegm-heat-clearing preparation is detected, so that a theoretical basis is provided for the quality control of the phlegm-heat-clearing preparation and the deep research of a pharmacodynamic material basis.

Description

Technical Field

[0001] The invention relates to the technical field of medicine, and in particular to a thin-layer identification method for honeysuckle medicinal materials in a Tanreqing preparation. Background Art

[0002] Tanreqing preparation is made from five medicinal materials extracted and refined, including Scutellaria baicalensis, bear bile powder, goat horn, honeysuckle, and forsythia. It has the effects of clearing heat, detoxifying, and resolving phlegm. It is effective in treating upper respiratory tract infections, acute bronchitis, pneumonia, tuberculosis with lung infections, late-stage lung cancer with lung infections, and severe asthma with infections.

[0003] There is no report on the thin layer identification method of honeysuckle medicinal materials in Tanreqing preparations in the prior art. Summary of the Invention

[0004] Based on this, the present invention provides a thin layer chromatography identification method for honeysuckle medicinal materials in Tanreqing preparations, which comprises the following steps:

[0005] (1) pre-treating the Tanreqing preparation to obtain a test solution, and pre-treating a control medicinal material or a control substance to obtain a control medicinal material solution or a control substance solution; and

[0006] (2) performing thin layer chromatography identification analysis on the test sample solution, the negative blank sample solution of the Tanreqing preparation lacking honeysuckle, the control medicinal material solution and / or the reference sample solution;

[0007] In the thin layer chromatography identification analysis step, an upper layer solution of butyl acetate-formic acid-water is used as a developing agent, and the volume ratio of the butyl acetate, the formic acid and the water is (1-10):(1-10):(1-10).

[0008] Furthermore, the volume ratio of the butyl acetate, the formic acid and the water is (5-9):(1-4):(1-4).

[0009] Furthermore, the volume ratio of the butyl acetate, the formic acid, and the water is about 7:about 2.5:about 2.5.

[0010] Furthermore, the thin layer identification method further comprises a thin layer plate activation step.

[0011] Furthermore, the thin layer plate activation step includes activating the thin layer plate at 100° C. to 120° C. for 20 min to 60 min.

[0012] Furthermore, the thin layer plate is selected from at least one of a silica gel G prefabricated thin layer plate, a silica gel GF254 thin layer plate, a silica gel H prefabricated thin layer plate, a silica gel HF254 thin layer plate and a polyamide thin layer plate.

[0013] Furthermore, the thin layer plate is a prefabricated thin layer plate made of silica gel G.

[0014] Furthermore, the inspection condition in the thin layer chromatography identification analysis step is: inspection under ultraviolet light, wherein the ultraviolet wavelength used for inspection is 365nm.

[0015] Furthermore, the temperature condition in the thin layer chromatography identification analysis step is 1-30°C, such as about 4°C or about 25°C.

[0016] Furthermore, the humidity condition in the thin layer chromatography identification analysis step is 35% to 80%, for example, about 42%, about 50%, about 58%, or about 72%.

[0017] Furthermore, the sample solution is spotted in an amount of 1 to 10 μl, for example, about 5 μl.

[0018] Furthermore, the Tanreqing preparation is Tanreqing injection, Tanreqing capsule or Tanreqing oral liquid.

[0019] Furthermore, the preparation method of the control medicinal material solution or the control substance solution includes: weighing an appropriate amount of the control medicinal material or the control substance, and adding an organic solvent to dissolve it to obtain the control medicinal material solution or the control substance solution.

[0020] Furthermore, the organic solvent is selected from at least one of methanol, ethanol, n-butanol and ethyl acetate.

[0021] Furthermore, the organic solvent is methanol and / or ethanol.

[0022] Furthermore, the volume percentage concentration of the methanol and / or ethanol is 10% to 100%, for example, 100%.

[0023] Furthermore, the concentration of the reference solution is 0.1-10 mg / ml, for example, about 1 mg / ml.

[0024] Furthermore, the spotting volume of the reference solution is 1 to 10 μl, for example, about 5 μl.

[0025] Furthermore, the spotting volume of the control medicinal material solution is 1 to 10 μl, for example, about 10 μl.

[0026] Furthermore, the reference substance is chlorogenic acid, caffeic acid and / or 4,5-dicaffeoylquinic acid.

[0027] Furthermore, the control medicinal material is a honeysuckle control medicinal material.

[0028] Furthermore, the thin layer identification criteria include: checking whether the thin layer plate shows characteristic spots of the same color at corresponding positions as the chlorogenic acid reference substance, the caffeic acid reference substance and / or the 4,5-dicaffeoylquinic acid reference substance, and whether there is interference from the blank sample; if the thin layer plate has the characteristic spots and the blank sample has no interference, then it is determined that the Tanreqing preparation being tested contains honeysuckle medicinal materials, and the identification is completed.

[0029] According to another aspect of the present invention, there is provided a use of the above-mentioned thin layer identification method in identifying honeysuckle medicinal materials or identifying Tanreqing preparations containing honeysuckle medicinal materials.

[0030] According to another aspect of the present invention, a kit for identifying honeysuckle medicinal materials or Tanreqing preparations containing honeysuckle medicinal materials is provided, which comprises a thin layer plate and a developing agent, wherein the developing agent is an upper layer solution of butyl acetate-formic acid-water, and the thin layer plate is a silica gel G prefabricated thin layer plate.

[0031] Furthermore, the volume ratio of the butyl acetate, the formic acid and the water is (1-10):(1-10):(1-10).

[0032] Furthermore, the volume ratio of the butyl acetate, the formic acid and the water is (5-9):(1-4):(1-4).

[0033] Furthermore, the volume ratio of the butyl acetate, the formic acid, and the water is about 7:about 2.5:about 2.5.

[0034] Beneficial effects of the present invention:

[0035] The thin-layer identification method of the present invention has appropriate developer selection, high identification efficiency, high identification accuracy and is easy to operate. By detecting the honeysuckle medicinal material in the Tanreqing preparation, a theoretical basis is provided for the quality control of the Tanreqing preparation and the in-depth study of the medicinal material basis of the efficacy. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the following briefly introduces the drawings required for use in the description of the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without exceeding the scope of protection required by the present invention.

[0037] Figure 1 Schematic diagram of the comparison of test solution preparation methods for honeysuckle identification. 1 represents chlorogenic acid; 2 represents caffeic acid; 3 represents 4,5-dicaffeoylquinic acid; 4 represents Tanreqing Injection method ①; 5 represents Tanreqing Injection honeysuckle negative blank method ①; 6 represents Tanreqing Injection method ②; 7 represents Tanreqing Injection method ③; and 8 represents the honeysuckle control medicinal material.

[0038] Figure 2 Schematic diagram of the comparison of sample quantity for identification of honeysuckle. 1 represents chlorogenic acid; 2 represents caffeic acid; 3 represents 4,5-dicaffeoylquinic acid; 4 represents Tanreqing Injection method ①; 5 represents Tanreqing Injection method ②; 6 represents Tanreqing Injection honeysuckle negative blank method ①; 7 represents Tanreqing Injection honeysuckle negative blank method ②; 8 represents the honeysuckle control herb.

[0039] Figure 3 Schematic diagram of the comparison of developing agents for identification of honeysuckle. A represents method ①; B represents method ②; C represents method ③; 1 represents chlorogenic acid; 2 represents caffeic acid; 3 represents 4,5-dicaffeoylquinic acid; 4 represents Tanreqing injection; 5 represents the negative blank of Tanreqing injection with honeysuckle; and 6 represents the control medicinal material of honeysuckle.

[0040] Figure 4 Schematic diagram of the comparison results of thin-layer plate inspection for honeysuckle identification. A is a silica gel H precast thin-layer plate (Shanghai Dongfang Pharmaceutical Technology Industry Co., Ltd.); B is a silica gel G precast thin-layer plate (Merck); C is a silica gel G precast thin-layer plate (MN); and D is a silica gel G precast thin-layer plate (Yantai Institute of Chemical Industry). 1 is chlorogenic acid; 2 is caffeic acid; 3 is 4,5-dicaffeoylquinic acid; 4 is Tanreqing injection; 5 is a negative blank for Tanreqing injection and honeysuckle; and 6 is a control medicinal material for honeysuckle.

[0041] Figure 5 This is the identification chromatogram of honeysuckle. 1 is chlorogenic acid; 2 is caffeic acid; 3 is 4,5-dicaffeoylquinic acid; 4 is the honeysuckle control medicinal material; 5 is the honeysuckle negative blank of Tanreqing injection; 6 is test solution 1; 7 is test solution 2; 8 is test solution 3; 9 is test solution 4; 10 is test solution 5; 11 is test solution 6; 12 is test solution 7; 13 is test solution 8; and 14 is test solution 9. DETAILED DESCRIPTION

[0042] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative work shall fall within the scope of protection of the present invention.

[0043] Unless otherwise indicated, all technical and scientific terms and abbreviations used herein have the meanings commonly understood by those skilled in the art in the field of the present invention or the field in which the terms are used. Although any methods, conditions, substances or materials similar or equivalent to those disclosed herein can be used in the practice of the present invention, preferred methods, conditions, substances or materials are described herein.

[0044] The present invention is intended to encompass all alternatives, modifications, and equivalents that may come within the scope of the present invention as defined by the claims. Those skilled in the art will recognize many methods and materials similar or equivalent to those described herein that could be used in the practice of the present invention. The present invention is in no way limited to the methods and materials described.

[0045] As used in the specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise.

[0046] In the present invention, the term "comprising" is synonymous with "including." As used herein, the terms "comprises," "including," "having," "containing," or any other variations thereof, are intended to cover a non-exclusive inclusion. For example, a composition, process, method, article, or apparatus that comprises a list of elements is not necessarily limited to only those elements but may include other elements not expressly listed or inherent to such composition, process, method, article, or apparatus.

[0047] As described in the background technology section, there are no reports in the prior art on a method for thin-layer chromatography identification of honeysuckle in Tanreqing preparations. To address the above problem, the present invention provides a method for thin-layer chromatography identification of honeysuckle in Tanreqing preparations, the method comprising the following steps:

[0048] (1) pre-treating the Tanreqing preparation to obtain a test solution, and pre-treating a control medicinal material or a control substance to obtain a control medicinal material solution or a control substance solution; and

[0049] (2) performing thin layer chromatography identification analysis on the test sample solution, the negative blank sample solution of the Tanreqing preparation lacking honeysuckle, the control medicinal material solution and / or the reference sample solution;

[0050] In the thin layer chromatography identification analysis step, an upper layer solution of butyl acetate-formic acid-water is used as a developing agent, and the volume ratio of the butyl acetate, the formic acid and the water is (1-10):(1-10):(1-10).

[0051] In the present invention, when volume ratio, temperature, humidity, rate, time, concentration, spot size, equivalent, concentration, or other values ​​or parameters are expressed as a range, a preferred range, or a range defined by a series of upper preferred values ​​and lower preferred values, this should be understood as specifically disclosing all ranges formed by any pairing of any range upper limit or preferred value with any range lower limit or preferred value, regardless of whether the range is disclosed alone. For example, when the range "1-10" is disclosed, the described range should be interpreted as including the range "1-10", "1-9", "1-8", "1-7", "1-6", "1-5", "1-4", "1-3", "1-2", "2-10", "2-9", "2-8", "2-7", "2-6", "2-5", "2-4", "2-3", "3-10", "3-9", "3-8", etc. When a numerical range is described herein, unless otherwise stated, the range is intended to include its end values ​​and all integers and fractions within the range.

[0052] In a preferred embodiment, the volume ratio of the butyl acetate, the formic acid and the water is (5-9):(1-4):(1-4).

[0053] In a preferred embodiment, the volume ratio of the butyl acetate, the formic acid and the water is about 7: about 2.5: about 2.5.

[0054] In the present invention, "about" refers to a value within a range of ±5% of a specific value. For example, "about 7" includes ±5% of 7, or from 6.65 to 7.35; "about 2.5" includes ±5% of 2.5, or from 2.375 to 2.625.

[0055] In a preferred embodiment, the thin layer identification method further comprises a thin layer plate activation step.

[0056] In a preferred embodiment, the thin layer plate activation step comprises activating the thin layer plate at 100° C. to 120° C. for 20 min to 60 min.

[0057] In a preferred embodiment, the thin layer plate is selected from at least one of silica gel G prefabricated thin layer plate, silica gel GF254 thin layer plate, silica gel H prefabricated thin layer plate, silica gel HF254 thin layer plate and polyamide thin layer plate.

[0058] In a preferred embodiment, the thin layer plate is a silica gel G prefabricated thin layer plate.

[0059] In a preferred embodiment, the inspection condition in the thin layer chromatography identification analysis step is: inspection under ultraviolet light, wherein the ultraviolet wavelength used for inspection is 365nm.

[0060] In a preferred embodiment, the temperature condition in the thin layer chromatography identification analysis step is 1-30°C, such as about 4°C or about 25°C.

[0061] In the present invention, "about" refers to a value within a range of ±5% of a specific value. For example, "about 4" includes ±5% of 4, or from 3.8 to 4.2; "about 25" includes ±5% of 25, or from 23.75 to 26.25.

[0062] In a preferred embodiment, the humidity condition in the thin layer chromatography identification analysis step is 35% to 80%, such as about 42%, about 50%, about 58%, or about 72%.

[0063] In the present invention, "about" refers to a value within a range of ±5% of a particular value. For example, "about 42" includes ±5% of 42, or from 39.9 to 44.1; "about 50" includes ±5% of 50, or from 47.5 to 52.5; "about 58" includes ±5% of 58, or from 55.1 to 60.9; "about 72" includes ±5% of 72, or from 68.4 to 75.6.

[0064] In a preferred embodiment, the spotting volume of the test solution is 1-10 μl, for example, about 5 μl.

[0065] In the present invention, "about" refers to a value within a range of ±5% of a specific value. For example, "about 5" includes ±5% of 5, or from 4.75 to 5.25.

[0066] In a preferred embodiment, the Tanreqing preparation is Tanreqing injection, Tanreqing capsule or Tanreqing oral liquid.

[0067] In a preferred embodiment, the preparation method of the control medicinal material solution or the control substance solution comprises: weighing an appropriate amount of the control medicinal material or the control substance, and adding an organic solvent to dissolve the control medicinal material solution or the control substance solution.

[0068] In a preferred embodiment, the organic solvent is selected from at least one of methanol, ethanol, n-butanol and ethyl acetate.

[0069] In a preferred embodiment, the organic solvent is methanol and / or ethanol.

[0070] In a preferred embodiment, the volume percentage concentration of the methanol and / or ethanol is 10% to 100%, for example, 100%.

[0071] In a preferred embodiment, the concentration of the reference solution is 0.1-10 mg / ml, for example, about 1 mg / ml.

[0072] In the present invention, "about" refers to a value within a range of ±5% of a specific value. For example, "about 1" includes ±5% of 1, or from 0.95 to 1.05.

[0073] In a preferred embodiment, the spotting volume of the reference solution is 1-10 μl, for example, about 5 μl.

[0074] In the present invention, "about" refers to a value within a range of ±5% of a specific value. For example, "about 5" includes ±5% of 5, or from 4.75 to 5.25.

[0075] In a preferred embodiment, the spotting volume of the control medicinal material solution is 1-10 μl, for example, about 10 μl.

[0076] In the present invention, "about" refers to a value within a range of ±5% of a specific value. For example, "about 10" includes ±5% of 10, or from 9.5 to 10.5.

[0077] In a preferred embodiment, the reference substance is chlorogenic acid, caffeic acid and / or 4,5-dicaffeoylquinic acid.

[0078] In a preferred embodiment, the control medicinal material is honeysuckle control medicinal material.

[0079] In a preferred embodiment, the thin layer identification criteria include: checking whether the thin layer plate shows characteristic spots of the same color at corresponding positions as the chlorogenic acid reference substance, the caffeic acid reference substance and / or the 4,5-dicaffeoylquinic acid reference substance, and whether there is interference from the blank sample; if the thin layer plate has the characteristic spots and the blank sample has no interference, it is determined that the Tanreqing preparation being tested contains the honeysuckle medicinal material, and the identification is completed.

[0080] According to another aspect of the present invention, there is provided a use of the above-mentioned thin layer identification method in identifying honeysuckle medicinal materials or identifying Tanreqing preparations containing honeysuckle medicinal materials.

[0081] According to another aspect of the present invention, a kit for identifying honeysuckle medicinal materials or Tanreqing preparations containing honeysuckle medicinal materials is provided, which comprises a thin layer plate and a developing agent, wherein the developing agent is an upper layer solution of butyl acetate-formic acid-water, and the thin layer plate is a silica gel G prefabricated thin layer plate.

[0082] In a preferred embodiment, the volume ratio of the butyl acetate, the formic acid and the water is (1-10):(1-10):(1-10).

[0083] In a preferred embodiment, the volume ratio of the butyl acetate, the formic acid and the water is (5-9):(1-4):(1-4).

[0084] In a preferred embodiment, the volume ratio of the butyl acetate, the formic acid and the water is about 7: about 2.5: about 2.5.

[0085] In the present invention, "about" refers to a value within a range of ±5% of a specific value. For example, "about 7" includes ±5% of 7, or from 6.65 to 7.35; "about 2.5" includes ±5% of 2.5, or from 2.375 to 2.625.

[0086] The present invention will be further described below in conjunction with specific examples. It should be understood that these examples are intended to illustrate the present invention only and are not intended to limit the scope of the invention. The experimental methods in the following examples, for which specific conditions are not specified, are generally based on conventional conditions or conditions recommended by the manufacturer.

[0087] Unless otherwise defined, all technical and scientific terms used herein have the same meanings as those familiar to those skilled in the art. Furthermore, any methods and materials similar or equivalent to those described herein can be applied to the methods of the present invention. The preferred embodiments and materials described herein are for illustrative purposes only.

[0088] The above-mentioned features of the present invention or the features described in the embodiments may be combined in any combination. All features disclosed in this patent specification may be used in any combination, and each feature disclosed in the specification may be replaced by any alternative feature that can provide the same, equal, or similar purpose. Therefore, unless otherwise specified, the features disclosed are only general examples of equal or similar features.

[0089] Example

[0090] 1. Equipment and Materials

[0091] Fully automatic thin-layer sample analyzer, all reagents were of analytical grade, chlorogenic acid reference substance (provided by China Food and Drug Inspection Institute, batch number 110753-201415, purity 96.2%), caffeic acid reference substance (provided by China Food and Drug Inspection Institute, batch number 110885-200102), 4,5-dicaffeoylquinic acid reference substance (provided by Tongtian Biochemical, batch number 14080622, purity 98.0%), honeysuckle reference medicinal material (provided by China Food and Drug Inspection Institute, batch number 1060-200002), and Tanreqing injection (provided by Shanghai Kaibao Pharmaceutical Co., Ltd.).

[0092] 2. Investigation of thin layer method

[0093] The present invention investigates five aspects: preparation of test solution, sample spotting amount, developing agent, thin layer plate, temperature and humidity.

[0094] 2.1. Investigation of the preparation method of test solution

[0095] Tanreqing Injection (Batch No. 1803220) and a negative blank of Tanreqing Injection (Jin Yinhua) were used for the experiment. The following pretreatment methods were compared for the Tanreqing preparations: ① Tanreqing Injection was used as the test solution; ② 15 mg of graphitized carbon black was added to 1 ml of Tanreqing Injection, the mixture was shaken for 5 minutes, and the supernatant was collected as the test solution; ③ 5 ml of Tanreqing Injection was passed through a macroporous adsorption resin column (1 cm inner diameter, 5 cm height), eluting with 50 ml of water and then ethanol, sequentially. The ethanol eluate was collected and evaporated to dryness, and the residue was dissolved in 1 ml of methanol to serve as the test solution.

[0096] According to the thin layer chromatography conditions of the present invention (such as Figure 1 The results show that method ② did not remove the interference of the main spot, and although method ③ removed part of the interference to the main spot, it also weakened the fluorescence of the main spot. Therefore, method ② and method ③ were not used, and method ① was still used.

[0097] 2.2. Investigation of different sample quantities

[0098] The test solution was prepared according to the method of the present invention, with ① 5 μl of reference solution, 10 μl of reference medicinal material solution, and 5 μl of Tanreqing injection as the spotting amount; ② 5 μl of reference solution, 10 μl of reference medicinal material solution, and 10 μl of Tanreqing injection as the spotting amount, and the results (such as Figure 2 (As shown in the chromatogram, the spots of method ① are clear and the separation effect is better, so method ① is adopted.

[0099] 2.3. Investigation of different developing agents

[0100] like Figure 3 As shown, the test solution was prepared according to the method of the present invention, using ① butyl acetate-formic acid-water (7:2.5:2.5) upper layer solution as the developing solvent, the spots of chlorogenic acid, caffeic acid and 4,5-dicaffeoylquinic acid had moderate Rf values, and the main spots of the same color were shown at the corresponding positions of the chromatogram of the honeysuckle control medicinal material, and there were 2 main spots between caffeic acid and 4,5-dicaffeoylquinic acid, 1 main spot between 4,5-dicaffeoylquinic acid and chlorogenic acid, and 1 main spot below chlorogenic acid; the separation effect of each spot was good; using ② butyl acetate-formic acid-water (2:1:1) upper layer solution as the developing solvent, there was interference at the 4,5-dicaffeoylquinic acid spot; using ③ butyl acetate-formic acid-water (1:1:1) upper layer solution as the developing solvent, the separation was poor compared with method ② and method ③. Therefore, the upper layer solution of method ① butyl acetate-formic acid-water (7:2.5:2.5) was used as the developing solvent.

[0101] 2.4. Investigation of different thin layer plates

[0102] like Figure 4 As shown, according to the method of the present invention, under the same experimental conditions, four types of thin layer plates, namely, silica gel H prefabricated thin layer plate (Shanghai Oriental Pharmaceutical Technology Industry Co., Ltd.), silica gel G prefabricated thin layer plate (Merck), silica gel G prefabricated thin layer plate (MN) and silica gel G prefabricated thin layer plate (Yantai Chemical Industry Research Institute), were compared. The results showed that the spots on the H plate were not clear and had tailing phenomenon, while the chromatographic separation effect of the G plate was better, so the silica gel G prefabricated thin layer plate was used.

[0103] 2.5. Investigation at different temperatures and humidity

[0104] According to the method of the present invention, four dosage forms were investigated using silica gel G prefabricated thin layer plates (Merck) at ① room temperature (temperature: 25°C, humidity 50%); ② low humidity (temperature: 25°C, humidity 42%); ③ medium humidity (temperature: 25°C, humidity 58%); ④ high humidity (temperature: 25°C, humidity 72%) and ⑤ low temperature (temperature: 4°C, humidity 50%). The results showed that the chromatographic separation effects were good.

[0105] 3. Sample determination

[0106] like Figure 5 As shown, 33 batches of Tanreqing injection samples were tested according to the method of the present invention. The results showed that in the chromatogram of the test solution, fluorescent spots of the same color were displayed at the corresponding positions of the chlorogenic acid reference substance, caffeic acid, 4,5-dicaffeoylquinic acid reference substance, and honeysuckle reference medicinal material.

[0107] The embodiments of the present invention are described in detail above. Specific examples are used herein to illustrate the principles and implementation methods of the present invention. The description of the above embodiments is only intended to help understand the method of the present invention and its core concept. At the same time, changes or modifications made by those skilled in the art based on the concept of the present invention, the specific implementation methods of the present invention, and the scope of application are all within the scope of protection of the present invention. In summary, the contents of this specification should not be understood as limiting the present invention.

Claims

1. A thin layer identification method for honeysuckle medicinal materials in Tanreqing preparations, characterized in that: The thin layer identification method comprises the following steps: (1) pre-treating the Tanreqing preparation to obtain a test solution, and pre-treating a control medicinal material or a control substance to obtain a control medicinal material solution or a control substance solution; and (2) performing thin layer chromatography identification analysis on the test sample solution, the negative blank sample solution of the Tanreqing preparation lacking honeysuckle, the control medicinal material solution and / or the reference sample solution; Wherein, in the step of thin layer chromatography identification analysis, an upper layer solution of butyl acetate-formic acid-water is used as a developing agent, and the volume ratio of the butyl acetate, the formic acid and the water is (1-10):(1-10):(1-10).

2. The thin layer identification method according to claim 1, characterized in that: The volume ratio of the butyl acetate, the formic acid and the water is (5-9):(1-4):(1-4); Preferably, the volume ratio of the butyl acetate, the formic acid, and the water is about 7: about 2.5: about 2.

5.

3. The thin layer identification method according to claim 1, characterized in that: The thin layer identification method further comprises a thin layer plate activation step; Preferably, the thin layer plate activation step comprises activating the thin layer plate at 100° C. to 120° C. for 20 min to 60 min; More preferably, the thin layer plate is selected from at least one of silica gel G prefabricated thin layer plate, silica gel GF254 thin layer plate, silica gel H prefabricated thin layer plate, silica gel HF254 thin layer plate and polyamide thin layer plate; More preferably, the thin layer plate is a silica gel G prefabricated thin layer plate.

4. The thin layer identification method according to claim 1, characterized in that: The inspection conditions in the thin layer chromatography identification analysis step are: inspection under ultraviolet light, wherein the ultraviolet wavelength used for inspection is 365nm; Particularly preferably, the temperature condition in the thin layer chromatography identification analysis step is 1 to 30°C, such as about 4°C or about 25°C; Particularly preferably, the humidity condition in the thin layer chromatography identification analysis step is 35% to 80%, such as about 42%, about 50%, about 58%, or about 72%.

5. The thin layer identification method according to claim 1, characterized in that: The sample volume of the test solution is 1 to 10 μl, for example, about 5 μl; Preferably, the Tanreqing preparation is Tanreqing injection, Tanreqing capsule or Tanreqing oral liquid.

6. The thin layer identification method according to claim 1, characterized in that: The preparation method of the control medicinal material solution or the control substance solution comprises: weighing an appropriate amount of the control medicinal material or the control substance, and adding an organic solvent to dissolve the control medicinal material solution or the control substance solution; Preferably, the organic solvent is selected from at least one of methanol, ethanol, n-butanol and ethyl acetate; More preferably, the organic solvent is methanol and / or ethanol; More preferably, the volume percentage concentration of the methanol and / or ethanol is 10% to 100%, for example 100%; Particularly preferably, the concentration of the reference solution is 0.1 to 10 mg / ml, for example about 1 mg / ml; Particularly preferably, the spotting volume of the reference solution is 1 to 10 μl, for example, about 5 μl; Particularly preferably, the spotting volume of the control medicinal material solution is 1 to 10 μl, for example, about 10 μl.

7. The thin layer identification method according to claim 1, characterized in that: The reference substances are chlorogenic acid, caffeic acid and / or 4,5-dicaffeoylquinic acid; Preferably, the control medicinal material is a honeysuckle control medicinal material.

8. The thin layer identification method according to claim 1, characterized in that: The thin layer identification criteria include: checking whether the thin layer plate shows characteristic spots of the same color at corresponding positions as the chlorogenic acid reference substance, the caffeic acid reference substance and / or the 4,5-dicaffeoylquinic acid reference substance, and whether there is interference from the blank sample; if the thin layer plate has the characteristic spots and the blank sample has no interference, it is determined that the Tanreqing preparation being tested contains honeysuckle medicinal materials, and the identification is completed.

9. Use of the thin-layer identification method according to any one of claims 1 to 8 in identifying honeysuckle or a Tanreqing preparation containing honeysuckle.

10. A kit for identifying honeysuckle or a Tanreqing preparation containing honeysuckle, comprising a thin layer plate and a developing agent, wherein the developing agent is an upper layer solution of butyl acetate-formic acid-water, and the thin layer plate is a silica gel G prefabricated thin layer plate; Preferably, the volume ratio of the butyl acetate, the formic acid and the water is (1-10):(1-10):(1-10); More preferably, the volume ratio of the butyl acetate, the formic acid and the water is (5-9):(1-4):(1-4); Preferably, the volume ratio of the butyl acetate, the formic acid and the water is about 7: about 2.5: About 2.5.

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