阅读说明：本技术 姜竹茹中药配方颗粒中竹茹的鉴别方法 (Method for identifying bamboo shavings in ginger bamboo shavings traditional Chinese medicine formula granules ) 是由 吕渭升 杨文惠 潘晓君 侯栩轩 叶梅霞 李振雨 何民友 陈向东 孙冬梅 魏梅 于 2021-08-02 设计创作，主要内容包括：本发明涉及一种姜竹茹中药配方颗粒中竹茹的鉴别方法,所述鉴别方法包括如下步骤：分别取姜竹茹中药配方颗粒和竹茹对照药材,制备供试品溶液和对照药材溶液；将所述供试品溶液和所述对照药材溶液点样于薄层层析板上,以甲苯-三氯甲烷-丙酮-甲酸为展开剂,展开,检视。本发明对薄层层析工艺特别是展开剂配方进行调整,展开剂能针对性溶解竹茹成分从而将其从供试品溶液中分离并随后在层析板上展开,有效去除其他干扰成分的影响,斑点显色清晰,分离度好。(The invention relates to an identification method of bamboo shavings in ginger bamboo shavings traditional Chinese medicine formula granules, which comprises the following steps: respectively taking ginger-bamboo shavings traditional Chinese medicine formula granules and bamboo shavings reference medicinal materials, and preparing a test solution and a reference medicinal material solution; spotting the test solution and the control solution on a thin-layer chromatography plate, developing with toluene-chloroform-acetone-formic acid as developing agent, and inspecting. The invention adjusts the thin-layer chromatography process, particularly the formula of the developing agent, the developing agent can specifically dissolve the components of the bamboo shavings so as to separate the components from the test solution and then develop the components on the chromatography plate, thereby effectively removing the influence of other interference components, and the spots are clear in color development and good in separation degree.)

1. The method for identifying the bamboo shavings in the ginger bamboo shavings traditional Chinese medicine formula particles is characterized by comprising the following steps of:

respectively taking ginger-bamboo shavings traditional Chinese medicine formula granules and bamboo shavings reference medicinal materials, and preparing a test solution and a reference medicinal material solution;

spotting the test solution and the control solution on a thin-layer chromatography plate, developing with toluene-chloroform-acetone-formic acid as developing agent, and inspecting.

2. The method for identifying bamboo shavings in ginger-bamboo shavings traditional Chinese medicine formula granules according to claim 1, wherein the volume ratio of the toluene, the chloroform, the acetone and the formic acid is (7.5-8.5): (4.5-5.5): (2.5-3.5): (0.08-0.12).

3. The method for identifying bamboo shavings in ginger bamboo shavings traditional Chinese medicine formula granules according to claim 1 or 2, wherein the thin layer chromatography plate is silica gel GF₂₅₄And (3) a plate.

4. The method for identifying bamboo shavings in ginger-bamboo shavings traditional Chinese medicine formula granules according to claim 1 or 2, wherein the preparation step of the test solution comprises: dissolving rhizoma Zingiberis recens caulis Bambusae in Taenia Chinese medicinal granule in water, extracting the obtained water solution with diethyl ether, removing solvent from the obtained extractive solution, and dissolving the obtained residue with methanol.

5. The method for identifying bamboo shavings in ginger bamboo shavings traditional Chinese medicine formula granules according to claim 4, wherein the number of times of extraction is 1 to 3, and the amount of diethyl ether used in each 1g of ginger bamboo shavings traditional Chinese medicine formula granules is 15 to 25mL in each 1 time of extraction.

6. The method for identifying bamboo shavings in ginger-bamboo shavings traditional Chinese medicine formula granules according to claim 1 or 2, wherein the preparation step of the reference medicinal material solution comprises the following steps: decocting the caulis Bambusae in Taenia in water, filtering, collecting filtrate, removing water from the filtrate, dissolving the residue with water, extracting the water solution with diethyl ether, removing solvent from the extractive solution, and dissolving the residue with methanol.

7. The method for identifying bamboo shavings in ginger-bamboo shavings traditional Chinese medicine formula granules as claimed in claim 6, wherein the number of times of extraction is 1-3, and the amount of ethyl ether used in each 1g of bamboo shavings medicinal material is 3-5 mL.

8. The method for identifying bamboo shavings in ginger-bamboo shavings traditional Chinese medicine formula granules according to any one of claims 1, 2, 5 and 7, wherein the method for identifying bamboo shavings further comprises the following steps: and preparing a negative control solution from a negative sample of the bamboo shavings, spotting the negative control solution on the thin layer chromatography plate, developing in the developing agent, and inspecting.

9. The method for identifying bamboo shavings in ginger-bamboo shavings traditional Chinese medicine formula granules as recited in claim 8, wherein negative samples of the bamboo shavings are maltodextrin.

10. The method for identifying bamboo shavings in ginger-bamboo shavings traditional Chinese medicine prescription granule as claimed in any one of claims 1, 2, 5, 7 and 9, wherein the sample application mode adopts spray strip-like sample application.

11. The method for identifying caulis Bambusae in Taenia as in any one of claims 1, 2, 5, 7 and 9, wherein the inspection is performed under ultraviolet light.

12. The method for identifying bamboo shavings in ginger bamboo shavings traditional Chinese medicine formula granules according to any one of claims 1, 2, 5, 7 and 9, wherein the sample amount of the sample solution is 1.8 μ L-2.2 μ L; or/and the sample amount of the control medicinal material solution is 7.8-8.2 muL.

Technical Field

The invention relates to the technical field of traditional Chinese medicine detection, and particularly relates to a method for identifying bamboo shavings in ginger bamboo shavings traditional Chinese medicine formula granules.

Background

Bamboo shavings are dried middle layer of stems of gramineous green stalk bamboo Bambusa tuldoides Munro, large-headed bamboo Sinocalamus benheyanus McClure var. pubescens P.F.Li or light bamboo Phyllostachys nigra (Lodd.) Munro var. Henonis (Mitf.) Stapf ex Rendle, and are loaded from Jinkuiyao, the main components of the bamboo shavings are triterpenes and flavones, and the bamboo shavings also contain polysaccharides, amino acids, phenolic substances, resins and the like, and are commonly used for clearing lung and eliminating phlegm. The ginger bamboo shavings are prepared by processing bamboo shavings and are roasted or fried with ginger juice to be yellow. It is in the shape of bamboo shavings, has a yellow surface and slight ginger fragrance. Zhu Ru with ginger is good at resolving phlegm and stopping vomiting. Clinically, Zhu Ru is used for arresting vomiting and Zhu Ru is used for eliminating phlegm.

The traditional Chinese medicine formula particle is a particle prepared by carrying out water extraction, separation, concentration, drying and granulation on single traditional Chinese medicine decoction pieces. The traditional Chinese medicine formula granules are obtained from traditional Chinese medicine decoction pieces, and the drug effect substances are basically consistent with the decoction pieces of the traditional Chinese medicine decoction pieces. Traditionally, people can identify the types of medicinal materials by eye observation, nose smell, mouth taste and hand touch, and the traditional Chinese medicine formula granules are changed into finished products without the appearance shape of the medicinal pieces from shaped medicinal pieces through a series of processing, and are difficult to identify by traditional visual observation, nose smell and the like. Because the traditional Chinese medicine formula particle loses the characteristics of appearance, smell and the like which are recognized by the traditional decoction pieces, and plays an important role in the prescription as solid decoction preparation, the identification of the types of medicinal materials contained in the traditional Chinese medicine formula particle is particularly important by adopting a proper technical means. At present, the qualitative methods of traditional Chinese medicine components mainly comprise physicochemical tests, thin-layer chromatography, high performance liquid chromatography, gas-mass spectrometry, ultra-high performance liquid chromatography-mass spectrometry and the like. High performance liquid chromatography or liquid chromatography-mass spectrometry, etc., the required instruments are more precise, leading to greatly increased inspection cost. Thin-layer chromatography, also known as thin-layer chromatography, is a method in which a sample solution is spotted on a thin-layer plate, developed with a developing agent in a developing container, and the components contained in the sample are separated and used for identification, examination, and content measurement. The thin layer chromatography is one of the most widely applied methods in the plane chromatography, and has the advantages of easy mastering, cheap equipment and simple and flexible operation.

The traditional technology for detecting bamboo shavings based on thin-layer chromatography is as follows: after inspection, only one report is that thin-layer identification is carried out on bamboo shavings in a compound prescription, and the detected components have larger polarity, but spots are blurry and trailing is serious. The thin-layer chromatography identification research of rhizoma gastrodiae dizzy ning tablets published by Cao Rui, Cao Lin and the like relates to the thin-layer identification of bamboo shavings, and concretely comprises the steps of taking 10 rhizoma gastrodiae dizzy ning tablets, removing coatings, grinding, adding 30mL of 70% ethanol for dissolving, performing ultrasonic treatment for 40min, filtering, concentrating and evaporating filtrate, adding 2mL of 70% ethanol for dissolving, and preparing a bamboo shavings negative control solution by the same method as a test solution. Control solution: taking 1g of caulis Bambusae in Taenia as a reference material, adding 20mL of 70% ethanol, reflux extracting for 40min, filtering, concentrating the filtrate, evaporating to dryness, and dissolving with 2mL of 70% ethanol to obtain the final product. Sucking 10 μ L of the control solution, 15 μ L of the sample solution and 15 μ L of the negative control solution, dropping on the same silica gel G thin layer plate, developing with n-butanol-glacial acetic acid-water (4: 5), taking out, air drying, spraying ninhydrin solution, and heating at 105 deg.C until the spots are clearly developed. "

However, the thin-layer chromatography is not reported for identifying the ginger-bamboo shavings traditional Chinese medicine formula granules, and how to realize the identification of the ginger-bamboo shavings traditional Chinese medicine formula granules based on the thin-layer chromatography is a technical problem to be solved.

Disclosure of Invention

Based on the background technology, the invention mainly aims to provide a method for identifying bamboo shavings in ginger bamboo shavings traditional Chinese medicine formula granules. The identification method is mainly used for identifying the ginger-bamboo-shaving traditional Chinese medicine formula granules based on the thin-layer chromatography, and provides an effective means for quality detection of the ginger-bamboo-shaving traditional Chinese medicine formula granules.

The purpose of the invention can be realized by the following technical scheme:

a method for identifying bamboo shavings in ginger bamboo shavings traditional Chinese medicine formula granules comprises the following steps:

respectively taking ginger-bamboo shavings traditional Chinese medicine formula granules and bamboo shavings reference medicinal materials, and preparing a test solution and a reference medicinal material solution;

spotting the test solution and the control solution on a thin-layer chromatography plate, developing with toluene-chloroform-acetone-formic acid as developing agent, and inspecting.

In one embodiment, the volume ratio of the toluene, the chloroform, the acetone, and the formic acid is (7.5-8.5): (4.5-5.5): (2.5-3.5): (0.08-0.12).

In one embodiment, the thin layer chromatography plate is silica gel GF₂₅₄And (3) a plate.

In one embodiment, the step of preparing the test solution comprises: dissolving rhizoma Zingiberis recens caulis Bambusae in Taenia Chinese medicinal granule in water, extracting the obtained water solution with diethyl ether, removing solvent from the obtained extractive solution, and dissolving the obtained residue with methanol.

In one embodiment, the number of times of extraction is 1 to 3, and the amount of diethyl ether used in each 1g of the ginger-bamboo shavings traditional Chinese medicine formula granules is 15 to 25 mL.

In one embodiment, the preparation of the reference solution comprises: decocting the caulis Bambusae in Taenia in water, filtering, collecting filtrate, removing water from the filtrate, dissolving the residue with water, extracting the water solution with diethyl ether, removing solvent from the extractive solution, and dissolving the residue with methanol.

In one embodiment, the number of times of extraction is 1 to 3, and the amount of diethyl ether used in each 1g of the caulis bambusae in taeniam medicinal material in each 1 time of extraction is 3 to 5 mL.

In one embodiment, the authentication method further comprises the steps of: and preparing a negative control solution from a negative sample of the bamboo shavings, spotting the negative control solution on the thin layer chromatography plate, developing in the developing agent, and inspecting.

In one embodiment, the negative sample of mangosteen flabellatus is maltodextrin.

In one embodiment, the printing mode adopts spray stripe printing.

In one embodiment, the inspection is performed under ultraviolet light.

In one embodiment, the sample solution is applied in an amount of 1.8. mu.L to 2.2. mu.L.

In one embodiment, the sample amount of the control solution is 7.8 μ L-8.2 μ L.

Compared with the prior art, the invention has the following beneficial effects:

the invention applies the thin-layer chromatography to the identification of bamboo shavings in the ginger bamboo shavings traditional Chinese medicine formula particles, and in order to meet the identification requirements of the ginger bamboo shavings traditional Chinese medicine formula particles, the invention adjusts the thin-layer chromatography process, particularly the formula of a developing agent, and the developing agent can specifically dissolve the components of the bamboo shavings so as to separate the components from a test solution and then develop the components on a chromatographic plate, thereby effectively removing the influence of other interference components, having clear spot color development and good separation degree.

The identification method provided by the invention is simple and easy to operate, requires less equipment, is low in inspection cost, can present the detection result more accurately and clearly, and is suitable for popularization and application in the field of medicine inspection. In addition, the identification method is rapid, stable, strong in specificity and good in durability, and can accurately display the characteristics of the components of the bamboo shavings. The invention provides a scientific, rapid and low-cost new method for controlling the quality of the ginger-bamboo shavings traditional Chinese medicine formula granule product.

Drawings

In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and other drawings can be obtained by those skilled in the art without creative efforts.

FIG. 1 shows different sample amounts of thin layer chromatography of rhizoma Zingiberis recens, caulis Bambusae in Taenia, and Chinese medicinal granule; wherein: 1. caulis Bambusae in Taenia reference medicinal material 5 μ L; 2. caulis Bambusae in Taenia reference medicinal material 8 μ L; 3. caulis Bambusae in Taenia reference medicinal material 10 μ L; 4. 1 mu L of ginger bamboo shavings formula particles; 5. 2 mu L of ginger bamboo shavings formula particles; 6. 5 mu L of ginger bamboo shavings formula particles;

FIG. 2 is a thin layer chromatography specificity of a granule of a traditional Chinese medicine formula of rhizoma Zingiberis recens caulis Bambusae in Taenia; wherein: 1.2 mul of ginger bamboo shavings formula particle (CG 2006002); 2.2 mul of ginger bamboo shavings formula particle (CG 2006003); 3. 2 mul of ginger bamboo shavings formula particle (CG 2006004); 4. caulis Bambusae in Taenia reference medicinal material 8 μ L; 5. 2 μ L of negative sample;

FIG. 3 is a thin-layer chromatogram for identification of granule prepared from rhizoma Zingiberis recens, caulis Bambusae in Taenia, and Chinese medicinal materials at room temperature; wherein: 1. caulis Bambusae in Taenia reference medicinal material 8 μ L; 2.2 mul of ginger bamboo shavings formula particle (CG 2006002); 3. 2 mul of ginger bamboo shavings formula particle (CG 2006003); 4. 2 mul of ginger bamboo shavings formula particle (CG 2006004);

FIG. 4 is a thin-layer chromatogram for identification of granule of rhizoma Zingiberis recens, caulis Bambusae in Taenia, and Chinese medicinal materials at low temperature; wherein: 1. caulis Bambusae in Taenia reference medicinal material 8 μ L; 2.2 mul of ginger bamboo shavings formula particle (CG 2006002); 3. 2 mul of ginger bamboo shavings formula particle (CG 2006003); 4. 2 mul of ginger bamboo shavings formula particle (CG 2006004);

FIG. 5 is a thin-layer chromatogram for identification of formulation granules of caulis Bambusae in Taenia with ginger under high humidity; wherein: 1. caulis Bambusae in Taenia reference medicinal material 8 μ L; 2.2 mul of ginger bamboo shavings formula particle (CG 2006002); 3. 2 mul of ginger bamboo shavings formula particle (CG 2006003); 4. 2 mul of ginger bamboo shavings formula particle (CG 2006004);

FIG. 6 is a thin-layer chromatogram for identification of formulation granules of caulis Bambusae in Taenia with ginger under low humidity; wherein: 1. caulis Bambusae in Taenia reference medicinal material 8 μ L; 2.2 mul of ginger bamboo shavings formula particle (CG 2006002); 3. 2 mul of ginger bamboo shavings formula particle (CG 2006003); 4. 2 mul of ginger bamboo shavings formula particle (CG 2006004);

FIG. 7 is a chromatogram for investigation of silica gel G plates from different manufacturers; wherein: 1. caulis Bambusae in Taenia reference medicinal material 8 μ L; 2.2 mul of ginger bamboo shavings formula particle (CG 2006002); 3. 2 mul of ginger bamboo shavings formula particle (CG 2006003); 4. 2 mul of ginger bamboo shavings formula particle (CG 2006004);

FIG. 8 is a silica gel G plate investigation chromatogram of different manufacturers (Yinlong Nicotiana); wherein: 1. caulis Bambusae in Taenia reference medicinal material 8 μ L; 2.2 mul of ginger bamboo shavings formula particle (CG 2006002); 3. 2 mul of ginger bamboo shavings formula particle (CG 2006003); 4. 2 mul of ginger bamboo shavings formula particle (CG 2006004);

FIG. 9 is a silica gel G plate investigation chromatogram from different manufacturers; wherein: 1. caulis Bambusae in Taenia reference medicinal material 8 μ L; 2.2 mul of ginger bamboo shavings formula particle (CG 2006002); 3. 2 mul of ginger bamboo shavings formula particle (CG 2006003); 4. 2 mul of ginger bamboo shavings formula particle (CG 2006004);

FIG. 10 is a thin layer chromatography of ginger bamboo shavings Chinese medicinal granule in different batches; wherein: 1. caulis Bambusae in Taenia reference medicinal material 8 μ L; 2.2 mul of ginger bamboo shavings formula particle (CG 2006002); 3. 2 mul of ginger bamboo shavings formula particle (CG 2006003); 4. 2 mul of ginger bamboo shavings formula particle (CG 2006004);

FIG. 11 is a thin layer chromatography of ginger bamboo shavings Chinese medicinal granule of different batches; wherein: 1. 7.8 mu L of caulis Bambusae in Taenia reference material; 2. 1.8 mu L of ginger bamboo shavings formula particle (CG 2006002); 3. 1.8 mu L of ginger bamboo shavings formula particle (CG 2006003); 4. 1.8 mu L of ginger bamboo shavings formula particle (CG 2006004);

FIG. 12 is a thin layer chromatography of different batches of rhizoma Zingiberis recens, caulis Bambusae in Taenia Chinese medicinal granule; wherein: 1. caulis Bambusae in Taenia reference medicinal material 8.2 μ L; 2. 2.2 mu L of ginger bamboo shavings formula particle (CG 2006002); 3. 2.2 mu L of ginger bamboo shavings formula particle (CG 2006003); 4. 2.2 mu L of ginger bamboo shavings formula particle (CG 2006004);

FIG. 13 is a thin layer chromatography of ginger bamboo shavings Chinese medicinal granule of different batches; wherein: 1. caulis Bambusae in Taenia reference medicinal material 8 μ L; 2.2 mul of ginger bamboo shavings formula particle (CG 2006002); 3. 2 mul of ginger bamboo shavings formula particle (CG 2006003); 4. ginger and bamboo shavings formula particle (CG2006004)2 μ L.

Detailed Description

In order to facilitate an understanding of the present invention, the present invention will be described in more detail below. It should be understood, however, that the present invention may be embodied in many different forms and should not be construed as being limited to the embodiments or examples set forth herein. Rather, these embodiments or examples are provided so that this disclosure will be thorough and complete.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The terminology used in the description of the invention herein is for the purpose of describing particular embodiments or examples only and is not intended to be limiting of the invention. As used herein, the term "and/or" includes any and all combinations of two or more of the associated listed items, including any and all combinations of two or more of the associated listed items, or all of the associated listed items.

In the present invention, the technical features described in the open type include a closed technical solution composed of the listed features, and also include an open technical solution including the listed features.

In the present invention, the numerical range is defined to include both end points of the numerical range unless otherwise specified.

The percentage contents referred to in the present invention mean, unless otherwise specified, mass percentages for solid-liquid mixing and solid-solid phase mixing, and volume percentages for liquid-liquid phase mixing.

The percentage concentrations referred to in the present invention refer to the final concentrations unless otherwise specified. The final concentration refers to the ratio of the additive component in the system to which the component is added.

The temperature parameter in the present invention is not particularly limited, and may be a constant temperature treatment or a treatment within a certain temperature range. The constant temperature process allows the temperature to fluctuate within the accuracy of the instrument control.

The ginger-bamboo shavings traditional Chinese medicine formula granules are obtained by extracting ginger-bamboo shavings decoction pieces with water, concentrating and drying, and compared with the ginger-bamboo shavings decoction pieces, the ginger-bamboo shavings decoction pieces lose the characteristics of the traditional decoction pieces such as appearance, smell and the like, and can only judge the truth and the standard processing by analyzing the components. The ginger bamboo shavings traditional Chinese medicine formula particle contains chemical components of two medicines of bamboo shavings and ginger and chemical components generated in the processing process, the whole components are complex in composition, the components in the bamboo shavings and the components of the ginger are mutually interfered in the identification process, and the exact judgment result cannot be clearly and intuitively obtained by simply using the traditional thin-layer identification method (such as the method mentioned in the background technology) of the bamboo shavings. Therefore, the key for identifying the bamboo shavings based on the thin-layer chromatography is to effectively remove the influence of other components in the ginger bamboo shavings traditional Chinese medicine formula particles, particularly the components from the ginger medicinal material. Therefore, the thin-layer chromatography process, particularly the formula of the developing agent is adjusted, the developing agent can specifically dissolve the components of the bamboo shavings so as to separate the components from the test solution and then develop the components on the chromatography plate, the influence of other interference components is effectively removed, the spots are clear in color development, and the separation degree is good. Specifically, the technical scheme of the invention comprises the following steps:

the invention provides a method for identifying bamboo shavings in ginger bamboo shavings traditional Chinese medicine formula granules, which comprises the following steps:

respectively taking ginger-bamboo shavings traditional Chinese medicine formula granules and bamboo shavings reference medicinal materials, and preparing a test solution and a reference medicinal material solution;

spotting the test solution and the control solution on a thin-layer chromatography plate, developing with toluene-chloroform-acetone-formic acid as developing agent, and inspecting.

It is understood that the sequence of the steps involved in the identification method of the present invention is not particularly limited, and for example, the sample solution may be prepared first and then the control solution, or the control solution may be prepared first and then the sample solution may be prepared, the sample solution may be applied first and then the control solution, or the sample solution may be applied first and then the sample solution may be applied. Alternatively, it will be appreciated that the thin layer chromatography plate may be dried after being unfolded and then inspected.

In one example, the volume ratio of the toluene, the chloroform, the acetone, and the formic acid is (7.5-8.5): (4.5-5.5): (2.5-3.5): (0.08-0.12). For example, it may be 7.5:5.5:2.5:0.12, 8.5:4.5:3.5:0.08, or 8:5:3: 0.1.

In one example, the thin layer chromatography plate is silica gel GF₂₅₄And (3) a plate.

In one example, the step of preparing the test solution comprises: dissolving rhizoma Zingiberis recens caulis Bambusae in Taenia Chinese medicinal granule in water, extracting the obtained water solution with diethyl ether, removing solvent from the obtained extractive solution, and dissolving the obtained residue with methanol. The test solution is obtained by the preparation step, and the medium polar components of the bamboo shavings are enriched.

In one example, the number of extractions is 1 to 3 (e.g., 1, 2, 3), and the amount of diethyl ether per 1g of the ginger-processed bamboo shavings Chinese medicinal granule is 15mL to 25mL (e.g., 15mL, 20mL, 25mL) per 1 extraction. It can be understood that: when the extraction times are 2 times or 3 times, the obtained extracting solutions need to be combined; in the present invention, the method for removing diethyl ether from the obtained extract is not particularly limited, and for example, a method of evaporating diethyl ether; the amount of methanol used for dissolving the residue is not particularly limited, and for example, the amount of methanol used for dissolving the residue extracted from 1g of the bamboo shavings-ginger processed traditional Chinese medicine formula particles is 0.8mL to 1.2mL (e.g., 0.8mL, 1mL, 1.2 mL).

In one example, the preparation step of the control drug solution comprises: decocting the caulis Bambusae in Taenia in water, filtering, collecting filtrate, removing water from the filtrate, dissolving the residue with water, extracting the water solution with diethyl ether, removing solvent from the extractive solution, and dissolving the residue with methanol.

In one example, the number of times of extraction is 1 to 3 (e.g. 1, 2, 3), and the amount of ethyl ether used per 1g of the bamboo shavings material in each 1 extraction is 3mL to 5mL (e.g. 3mL, 4mL, 5 mL). It can be understood that: when the extraction times are 2 times or 3 times, the obtained extracting solutions need to be combined; in the present invention, the method for removing diethyl ether from the obtained extract is not particularly limited, and for example, a method of evaporating diethyl ether; the amount of methanol in the residue is not particularly limited, and is, for example, 0.18mL to 0.22mL (e.g., 0.18mL, 0.2mL, 0.22mL) per 1g of the control drug of caulis Bambusae in Taenia.

In one example, the authentication method further includes the steps of: and preparing a negative control solution from a negative sample of the bamboo shavings, spotting the negative control solution on the thin layer chromatography plate, developing in the developing agent, and inspecting.

In the present invention, the preparation of the negative control solution refers to the preparation of the test solution. It is understood that, in the identification method of the present invention, the preparation and spotting of the test solution, the control solution and the negative control solution are not sequentially defined.

It is understood that the present invention does not specifically limit the kind of the negative sample of bamboo shavings, and the negative sample of bamboo shavings does not contain bamboo shavings and is suitable for thin layer chromatography, and may be maltodextrin, for example.

In one example, the printing mode adopts spray strip printing.

In one example, the inspection is performed under ultraviolet light.

In one example, the wavelength of the ultraviolet light is 250nm to 260 nm. The invention takes 254nm as an example to illustrate the technical solution of the invention, but this is not a limitation to the wavelength of the ultraviolet light of the invention.

In one embodiment, the sample solution is applied in an amount of 1.8. mu.L to 2.2. mu.L. For example, 1.8. mu.L, 2. mu.L, 2.2. mu.L.

In one embodiment, the sample amount of the control solution is 7.8 μ L-8.2 μ L. For example, 7.8. mu.L, 8. mu.L, 8.2. mu.L.

The test methods described in the following examples are all conventional methods unless otherwise specified; the reagents and biomaterials, if not specifically mentioned, are commercially available.

Example 1

The embodiment relates to a method for identifying bamboo shavings in ginger bamboo shavings traditional Chinese medicine formula granules, which adopts thin-layer chromatography for identification and specifically comprises the following steps:

1.1 instruments, reagents and reagents

The instrument comprises the following steps: a thin layer auto imager (CAMAG REPROSTAR 3, Switzerland Karma); a double-tank deployment cylinder; fully AUTOMATIC thin layer spotter (AUTOMATIC TLC SAMPLER4, Switzerland Karma); a one in ten thousand balance (ME204E, mettler, switzerland); ultrapure water systems (Milli-Qdirect, Merck, Inc.); silica gel GF₂₅₄Thin layer plate (GF)₂₅₄The batch number is: 20160312, Qingdao ocean chemical Co., Ltd.); silica gel GF₂₅₄Thin layer plate (TLC Silica gel 60F)₂₅₄The batch number is: HX03452229, Merck GmbH); silica gel GF₂₅₄Thin layer plates (SGF254, batch No. 20210318, Nicoti chemical industry research institute).

Reagent: diethyl ether (batch number: SY22001010, Guangzhou chemical Co., Ltd.); toluene (batch number: SY22005016, Guangzhou chemical reagent works); trichloromethane (batch number: SY22001009, Guangzhou chemical Co., Ltd.); acetone (batch No. SY21810005, Kung san science, Inc.); the formic acid (batch number: SY22007013, Yongda chemical reagent Limited of Tianjin) is analytically pure; the water was ultrapure water (self-made in the laboratory).

Reagent testing: ginger bamboo shavings formula granules (batch number: CG2006002, CG2006003, CG 2006004; source: Guangdong one-party pharmaceutical Co., Ltd.); caulis Bambusae in Taenia reference drug (lot number: 180630, Chengdu Philippine biotechnology, Inc.); maltodextrin (batch number: F2004001, Biochemical energy of grain in Jilin (princess Ling) Co., Ltd.).

1.2 preparation of the solution

1.2.1 preparation of test solutions

Taking about 1g of rhizoma Zingiberis recens caulis Bambusae in Taenia traditional Chinese medicine formula granule, adding 20mL of water for dissolving, extracting with diethyl ether under shaking for 2 times, each time 20mL, mixing diethyl ether extractive solutions, volatilizing, and dissolving residue with 1mL of methanol to obtain sample solution.

1.2.2 preparation of reference drug solution

Taking 5g of caulis Bambusae in Taenia as a control medicinal material, adding 150mL of water, boiling for 30 min, filtering, adding 150mL of water again to the residue, boiling for 25min, filtering, combining the two filtrates, evaporating to dryness, adding 20mL of water to the residue for dissolving, shaking with diethyl ether for 2 times, extracting 20mL each time, combining the diethyl ether extract, volatilizing, adding 1mL of methanol to the residue for dissolving, and taking as a control medicinal material solution.

1.2.3 preparation of negative sample solution

Collecting negative (i.e. maltodextrin) sample of rhizoma Zingiberis recens caulis Bambusae in Taenia 1g, dissolving in water 20mL, extracting with diethyl ether for 2 times with shaking, each time 20mL, mixing diethyl ether extractive solutions, volatilizing, and dissolving the residue in methanol 1mL to obtain negative control solution.

1.3 thin layer chromatography conditions

Thin-layer plate: silica gel GF₂₅₄Thin layer plate (TLC Silica gel 60F)₂₅₄The batch number is: HX03452229, Merck GmbH);

developing agent: toluene-chloroform-acetone-formic acid (8: 5:3: 0.1);

sample application mode: spraying strip-shaped sample application;

the unfolding mode is as follows: adopting a double-groove unfolding cylinder, wherein the pre-saturation time of the unfolding cylinder is 15 minutes, and the unfolding distance is about 8 cm;

and (6) inspection: inspecting under UV light (254 nm).

1.4 investigation of different sample quantities of ginger bamboo shavings formula granules

Respectively sucking sample solution of rhizoma Zingiberis recens caulis Bambusae in Taenia formula granule (CG2006002) and caulis Bambusae in Taenia reference medicinal material solution with different volumes, and spotting on the same silica gel GF₂₅₄Spreading with toluene-chloroform-acetone-formic acid (8: 5:3: 0.1) as developing agent, taking out, air drying, and inspecting under ultraviolet light (254 nm). The results of the experiment are shown in FIG. 1.

As can be seen from fig. 1, when the sample amount of the sample solution of the ginger-bamboo shavings traditional Chinese medicine formula granule is 2 muL and the sample amount of the bamboo shavings reference medicinal material solution is 8 muL, the main spots of the sample chromatogram and the reference medicinal material chromatogram are clear in color development, the separation degree is good, no tailing phenomenon exists, the background is free of interference, the spots of the sample chromatogram and the reference medicinal material chromatogram can be in one-to-one correspondence, so that the sample amount of the sample solution is 2 muL and the sample amount of the reference medicinal material solution is 8 muL.

1.5 investigation of specificity of granule prepared from rhizoma Zingiberis recens, caulis Bambusae in Taenia and Chinese medicinal materials

Respectively sampling the test solution of the ginger-bamboo shavings Chinese medicinal formula granule, the bamboo shavings control medicinal material solution and the negative sample solution in the same silica gel GF₂₅₄Spreading with toluene-chloroform-acetone-formic acid (8: 5:3: 0.1) as developing agent, taking out, air drying, and inspecting under ultraviolet light (254 nm). The results of the experiment are shown in FIG. 2.

As can be seen from fig. 2, the chromatogram of the test sample of the ginger-bamboo shavings traditional Chinese medicine formula granule shows spots with the same color at the position corresponding to the chromatogram of the bamboo shavings reference medicinal material, and the negative sample has no interference. Indicating that the thin layer approach is well specified.

1.6 investigation of different temperatures

Respectively sampling the test solution of the ginger-bamboo shavings Chinese medicinal granule and the control solution of bamboo shavings on the same silica gel GF₂₅₄Developing the thin layer plate with toluene-chloroform-acetone-formic acid (8: 5:3: 0.1) as developing agent at normal temperature and low temperature, taking out, air drying, and inspecting under ultraviolet light (254 nm). The results of the experiment are shown in FIGS. 3 and 4.

As can be seen from fig. 3 and 4, under normal temperature and low temperature conditions, the main spots of the chromatogram of the test sample of the ginger-bamboo shavings traditional Chinese medicine formula particle and the chromatogram of the bamboo shavings reference medicinal material are clear in color, better in separation degree, free from tailing phenomenon and free from interference on background, the spots of the chromatogram of the test sample and the chromatogram of the reference medicinal material can be in one-to-one correspondence, which shows that the temperature reduction has no obvious influence on the thin-layer identification of the ginger-bamboo shavings traditional Chinese medicine formula particle, and the thin-layer identification method has good durability on the normal temperature and low temperature conditions.

1.7 differential humidity investigation

Respectively sampling the test solution of the ginger-bamboo shavings Chinese medicinal granule and the control solution of bamboo shavings on the same silica gel GF₂₅₄Spreading with toluene-chloroform-acetone-formic acid (8: 5:3: 0.1) as developing agent under high humidity and low humidity conditions, taking out, air drying, and inspecting under ultraviolet light (254 nm). The results of the experiment are shown in FIGS. 5 and 6:

as can be seen from fig. 5 and 6, under high humidity and low humidity conditions, main spots of the chromatogram of the test sample of the ginger-containing bamboo shavings formula particles and the chromatogram of the bamboo shavings reference medicinal material are clear in color, good in separation degree, free of tailing phenomenon and free of interference on background, the spots of the chromatogram of the test sample and the chromatogram of the reference medicinal material can be in one-to-one correspondence, and therefore it is shown that humidity has no obvious influence on thin-layer identification of the ginger-containing bamboo shavings formula particles, and the thin-layer identification method is good in durability to different humidity.

1.8 investigation of thin-layer plates from different manufacturers

Respectively sucking the test solution of the ginger-bamboo shavings Chinese medicinal granule and the control solution of bamboo shavings and spotting on the silica gel GF of different manufacturers₂₅₄Prefabricated thin layer plate (sea silica gel GF)₂₅₄Silver-dragon silica gel GF for board and cigarette platform₂₅₄Plate, Merck siliconGlue GF₂₅₄Plate), developing with toluene-chloroform-acetone-formic acid (8: 5:3: 0.1) as developing agent under the same temperature and humidity conditions, taking out, air drying, and inspecting under ultraviolet light (254 nm). The experimental results are shown in fig. 7, fig. 8 and fig. 9.

As can be seen from FIGS. 7, 8 and 9, the silica gels GF from different manufacturers were used₂₅₄Thin layer plate (sea silica gel GF)₂₅₄Silver-dragon silica gel GF for board and cigarette platform₂₅₄Plate, merck silica gel GF₂₅₄Plate), the main spots of the chromatogram of the test sample of ginger bamboo shavings and the chromatogram of the reference medicinal material of bamboo shavings are clear in color, the separation degree is better, and the spots of the chromatogram of the test sample and the chromatogram of the reference medicinal material can be in one-to-one correspondence. The durability of the thin layer identification method on silica gel G thin layer plates of different manufacturers is good.

1.9 summary

The Rf value of each spot of the chromatogram obtained by the method is moderate, the integral separation effect is good, the spots are clear, the negative is free from interference, and the durability of the method is proved to be good through examination of thin-layer plates of different brands and different humiture. Comparing the chromatogram of the test solution with the chromatogram spots of the reference solution, wherein the spots with the same color can correspond to each other. The method is proved to be well used for identifying the thin-layer chromatography of the ginger bamboo shavings formula particles by taking the reference medicinal material as the reference.

Example 2

The embodiment relates to a thin-layer identification of bamboo shavings in ginger bamboo shavings traditional Chinese medicine formula granules, which comprises the following steps:

1.1 preparation of test solutions

Respectively taking about 1g of rhizoma Zingiberis recens caulis Bambusae in Taenia traditional Chinese medicine formula granule (CG2006002, CG2006003, CG2006004), adding 20mL of water for dissolving, extracting with diethyl ether under shaking for 2 times, each time 20mL, mixing diethyl ether extractive solutions, volatilizing, adding 1mL of methanol into residue for dissolving, and using as sample solution.

1.2 preparation of reference drug solution

Taking 5g of caulis Bambusae in Taenia as a control medicinal material, adding 150mL of water, boiling for 30 min, filtering, adding 150mL of water again to the residue, boiling for 25min, filtering, combining the two filtrates, evaporating to dryness, adding 20mL of water to the residue for dissolving, shaking with diethyl ether for 2 times, extracting 20mL each time, combining the diethyl ether extract, volatilizing, adding 1mL of methanol to the residue for dissolving, and taking as a control medicinal material solution.

1.3 unfolding

Performing thin layer chromatography (China pharmacopoeia 2020 edition general rule 0502) experiment by sucking 2 μ L of test sample solution and 8 μ L of reference medicinal material solution, and respectively dropping on the same silica gel GF₂₅₄Thin layer plate (TLC Silica gel 60F)₂₅₄The batch number is: HX03452229, Merck Co., Ltd.) was developed with toluene-chloroform-acetone-formic acid (8: 5:3: 0.1) as developing agent at a development distance of about 8 cm.

1.4 color development

Inspecting under UV light (254 nm).

1.5 determination of results

The result is shown in figure 10, and spots with the same color appear on the chromatogram of the three test samples at the positions corresponding to the chromatogram of the bamboo shavings reference medicinal material, which indicates that the three batches of ginger bamboo shavings traditional Chinese medicine formula granules contain the same components as the bamboo shavings reference medicinal material and meet the regulations.

Example 3

This embodiment is a modification of embodiment 2, and includes the following steps:

1.1 preparation of test solutions

Respectively taking about 1g of rhizoma Zingiberis recens caulis Bambusae in Taenia traditional Chinese medicine formula granule (CG2006002), adding 20mL of water for dissolving, extracting with diethyl ether under shaking for 1 time, with the amount of diethyl ether being 15mL, volatilizing, adding 0.8mL of methanol for dissolving, and using as sample solution.

1.2 preparation of reference drug solution

Taking 5g of bamboo shavings as a reference medicinal material, adding 150mL of water, boiling for 30 minutes, filtering, adding 150mL of water again to filter residues, boiling for 25min, filtering, combining the two filtrates, evaporating to dryness, adding 20mL of water to residues for dissolving, shaking and extracting with diethyl ether for 1 time, using 15mL of diethyl ether, combining the diethyl ether extract, volatilizing, adding 0.8mL of methanol to residues for dissolving, and taking the residue as a reference medicinal material solution.

1.3 unfolding

Performing thin layer chromatography (China pharmacopoeia 2020 edition general rule 0502) experiment, sucking 1.8 μ L of test solution and 7.8 μ L of reference solution, respectively dropping on the same silica gel GF₂₅₄Thin layer plate (TLC Silica gel60F₂₅₄The batch number is: HX03452229, Merck Co., Ltd.) was developed with toluene-chloroform-acetone-formic acid (7.5:5.5:2.5:0.12) as developing agent at a development distance of about 8 cm.

1.4 color development

Inspecting under UV light (254 nm).

1.5 determination of results

As a result: the result is shown in fig. 11, spots with the same color appear on the chromatogram of the three test samples at the positions corresponding to the chromatogram of the bamboo shavings reference medicinal material, which indicates that the three batches of ginger bamboo shavings traditional Chinese medicine formula granules contain the same components as the bamboo shavings reference medicinal material and meet the regulations.

Example 4

This embodiment is a modification of embodiment 2, and includes the following steps:

1.1 preparation of test solutions

Respectively taking about 1g of rhizoma Zingiberis recens caulis Bambusae in Taenia traditional Chinese medicine formula granule (CG2006002), adding water 20mL for dissolving, extracting with diethyl ether with shaking for 3 times, with diethyl ether amount of 25mL, volatilizing, adding methanol 1.2mL for dissolving, and making into sample solution.

1.2 preparation of reference drug solution

Taking 5g of bamboo shavings as a reference medicinal material, adding 150mL of water, boiling for 30 minutes, filtering, adding 150mL of water again to filter residues, boiling for 25min, filtering, combining the two filtrates, evaporating to dryness, adding 20mL of water to residues for dissolving, shaking and extracting with diethyl ether for 1 time with the dosage of 25mL of diethyl ether, combining the diethyl ether extract, volatilizing, adding 1.2mL of methanol to residues for dissolving, and taking the residue as a reference medicinal material solution.

1.3 unfolding

Performing thin layer chromatography (China pharmacopoeia 2020 edition general rule 0502) experiment, sucking 2.2 μ L of test sample solution and 8.2 μ L of reference medicinal material solution, respectively dropping on the same silica gel GF₂₅₄Thin layer plate (TLC Silica gel 60F)₂₅₄The batch number is: HX03452229, Merck Co., Ltd.) was developed with toluene-chloroform-acetone-formic acid (8.5:4.5:3.5:0.08) as a developing solvent at a development distance of about 8 cm.

1.4 color development

Inspecting under UV light (254 nm).

1.5 determination of results

As a result: the result is shown in figure 12, and spots with the same color appear on the chromatogram of the three test samples at the positions corresponding to the chromatogram of the bamboo shavings reference medicinal material, which indicates that the three batches of ginger bamboo shavings traditional Chinese medicine formula granules contain the same components as the bamboo shavings reference medicinal material and meet the regulations.

Example 5

This embodiment is a modification of embodiment 2, and includes the following steps:

1.1 preparation of test solutions

Respectively taking about 1g of rhizoma Zingiberis recens caulis Bambusae in Taenia traditional Chinese medicine formula granule (CG2006002), adding 20mL of water for dissolving, shaking and extracting with diethyl ether for 2 times, 20mL each time, mixing diethyl ether extractive solutions, volatilizing, and adding 1mL of methanol to dissolve the residue to obtain test solution.

1.2 preparation of reference drug solution

Taking 5g of caulis Bambusae in Taenia as a control medicinal material, adding 150mL of water, boiling for 30 min, filtering, adding 150mL of water again to the residue, boiling for 25min, filtering, combining the two filtrates, evaporating to dryness, adding 20mL of water to the residue for dissolving, shaking with diethyl ether for 2 times, extracting 20mL each time, combining the diethyl ether extract, volatilizing, adding 1mL of methanol to the residue for dissolving, and taking as a control medicinal material solution.

1.3 unfolding

Performing thin layer chromatography (China pharmacopoeia 2020 edition general rule 0502) experiment by sucking 2 μ L of test sample solution and 8 μ L of reference medicinal material solution, and respectively dropping on the same silica gel GF₂₅₄Thin layer plate (TLC Silica gel 60F)₂₅₄The batch number is: HX03452229, Merck Co., Ltd.) was developed with toluene-chloroform-acetone-formic acid (9: 6: 1: 0.1) as developing agent at a development distance of about 8 cm.

1.4 color development

Inspecting under UV light (254 nm).

1.5 determination of results

As a result: the result is shown in FIG. 13, and spots of the same color appear on the chromatogram of the three test samples at the positions corresponding to the chromatogram of the caulis Bambusae in Taenia control drug. However, the Rf value of the corresponding spot is lower, which is not in accordance with the specific shift Rf of the thin-layer chromatography item of Chinese pharmacopoeia 0502 and is preferably between 0.2 and 0.8, and the result may be misjudged, therefore, the chromatogram is not in accordance with the development condition.

The technical features of the embodiments described above may be arbitrarily combined, and for the sake of brevity, all possible combinations of the technical features in the embodiments described above are not described, but should be considered as being within the scope of the present specification as long as there is no contradiction between the combinations of the technical features.

The above-mentioned embodiments only express several embodiments of the present invention, so as to understand the technical solutions of the present invention specifically and in detail, but not to be understood as the limitation of the protection scope of the invention. It should be noted that, for a person skilled in the art, several variations and modifications can be made without departing from the inventive concept, which falls within the scope of the present invention. It should be understood that the technical solutions provided by the present invention, which are obtained by logical analysis, reasoning or limited experiments, are within the scope of the present invention as set forth in the appended claims. Therefore, the protection scope of the present invention should be subject to the content of the appended claims, and the description and the drawings can be used for explaining the content of the claims.