CN112326864A - Thin-layer chromatography method for identifying medicinal component of sculellaria barbata in zilongjin tablets and application - Google Patents
Thin-layer chromatography method for identifying medicinal component of sculellaria barbata in zilongjin tablets and application Download PDFInfo
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Abstract
The invention discloses a thin-layer chromatography method for identifying the medicinal material components of sculellaria barbata in zilongjin tablets and application thereof, belonging to the technical field of traditional Chinese medicine material identification and comprising the following steps: providing a Zilongjin tablet solution as a test solution, providing a scutellarin solution as a reference solution, and providing a preparation solution prepared from the rest medicinal ingredients except the sculellaria barbata in the Zilongjin tablet as a negative sample solution; analyzing the sample solution, the reference solution and the negative sample solution by thin layer chromatography, wherein the developing agent is mixed solution of ethyl acetate-butanone-formic acid-water. The method has the advantages of high accuracy and small identification error, and also has the advantages of strong specificity, strong durability, good reproducibility and simple operation.
Description
Technical Field
The invention relates to the technical field of traditional Chinese medicine identification, in particular to a thin-layer chromatography method for identifying a barbat skullcap medicinal component in a zilongjin tablet and application thereof.
Background
The Zilongjin tablets are unique varieties produced by Longshun banyan pharmaceutical factory, a member of the Tianjin pharmaceutical industry group, and the quality standard is recorded in the first part of the 2010 edition of Chinese pharmacopoeia. At present, partial medicinal materials in the prescription have no legal quality standard, the quality of the raw materials is difficult to control, and the detection of the components of the eight medicinal materials in the Zilongjin tablets cannot be covered by the pharmacopeia standard which only comprises three identifications and one content measurement.
In view of the above, the present invention is particularly proposed.
Disclosure of Invention
One of the purposes of the invention is to provide a thin-layer chromatography method for identifying the medicinal component of the sculellaria barbata in the zilongjin tablets, which can accurately identify the medicinal component of the sculellaria barbata in the zilongjin tablets and has the advantages of high accuracy, small identification error, strong specificity, strong durability and the like.
The second purpose of the invention is to apply the thin-layer chromatography method for identifying the barbed skullcap herb in the quality evaluation of the Zilongjin tablets.
In order to achieve the above purpose of the present invention, the following technical solutions are adopted:
in a first aspect, the invention provides a thin-layer chromatography method for identifying the medicinal material components of sculellaria barbata in zilongjin tablets, which comprises the following steps:
providing a Zilongjin tablet solution as a test solution, providing a scutellarin solution as a reference solution, and providing a preparation solution prepared from the rest medicines except the sculellaria barbata in the Zilongjin tablet as a negative sample solution;
analyzing the test solution, the reference solution and the negative sample solution by adopting a thin-layer chromatography to identify the sculellaria barbata medicinal material in the zilongjin tablets;
the developing agent of the thin-layer chromatography is a mixed solution of ethyl acetate, butanone, formic acid and water, and the volume ratio is 8-9: 3-4: 2-3: 2-3.
Preferably, the silica gel plate of the thin layer chromatography is a silica gel G plate.
Further, the silica gel G board includes cigarette platform silica gel G board and MERCK silica gel G board.
Preferably, the thin-layer chromatography is adopted to analyze the test solution, the reference solution and the negative sample solution to identify the components of the sculellaria barbata medicinal material in the rhododendron simsii planch, and the method comprises the following steps:
the test solution, the reference solution and the negative sample solution are developed according to the thin-layer chromatography, when spots with the same color appear in the chromatogram of the test solution at the positions corresponding to the chromatogram of the reference solution and the negative sample is not interfered, the sample is identified as containing the medicinal component of the barbed skullcap herb;
preferably, the volume ratio of ethyl acetate, butanone, formic acid and water in the developing solvent is 9: 4: 2: 2;
preferably, developing according to thin layer chromatography comprises the steps of:
taking the independent amount of each of the three solutions to be 4-8 mu l, respectively dropping the three solutions on the same silica gel G plate, and mixing the three solutions according to the volume ratio of 9: 4: 2: 2, developing with a mixed solution of ethyl acetate, methyl ethyl ketone, formic acid and water, taking out, drying and developing with a color-developing agent.
Preferably, the color developing agent is ferric trichloride ethanol solution;
preferably, the mass concentration of the ferric trichloride ethanol solution is 4-6%;
further, the mass concentration of the ferric trichloride ethanol solution is 5%.
Preferably, the development temperature of the thin layer chromatography is less than or equal to 10 ℃;
preferably, the development temperature of the thin layer chromatography is 16 ℃ to 26 ℃.
Preferably, the developed relative humidity of the thin layer chromatography is greater than or equal to 75%;
preferably, the developed relative humidity of the thin layer chromatography is less than or equal to 50%.
In a second aspect, the thin-layer chromatography method for identifying the barbed skullcap herb is applied to the quality evaluation of the Zilongjin tablets.
Compared with the prior art, the invention has the beneficial effects that:
the thin-layer chromatography method provided by the invention can accurately identify the medicinal material components of the sculellaria barbata in the zilongjin tablets, and the method not only has the advantages of high accuracy and small identification error, but also has the advantages of strong specificity, strong durability, good reproducibility, simple operation and the like; the thin-layer chromatography method can be applied to the evaluation of the quality of the traditional Chinese medicine, the control of the quality of the traditional Chinese medicine and the development of new traditional Chinese medicines.
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 is a graph showing the results of thin layer panels provided in example 1 of the present invention;
FIG. 2 is a graph showing the results of thin layer panels provided in example 2 of the present invention;
FIG. 3 is a graph showing the results of thin layer panels provided in example 3 of the present invention;
FIG. 4 is a graph showing the results of thin layer panels provided in example 4 of the present invention.
Detailed Description
The technical solutions of the present invention will be described clearly and completely with reference to the following embodiments, and it should be understood that the described embodiments are some, but not all, embodiments of the present invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
According to the first aspect of the invention, the thin-layer chromatography method for identifying the medicinal material components of the sculellaria barbata in the zilongjin tablets comprises the following steps:
s1: providing a Zilongjin tablet solution as a test solution, providing a scutellarin solution as a reference solution, and providing a preparation solution prepared from the rest medicines except the sculellaria barbata in the Zilongjin tablet as a negative sample solution;
s2: analyzing the test solution, the reference solution and the negative sample solution by adopting a thin-layer chromatography to identify whether the barbat skullcap medicinal ingredient is contained in the Zilongjin tablets;
zilongjin tablets
The Zilongjin tablet mainly comprises radix astragali, radix Angelicae sinensis, herba Solani Lyrati, herba Solani Nigri, Saviae Miltiorrhizae radix, herba Scutellariae Barbatae, herba Duchesneae Indicae and radix Curcumae; is film coated tablet. Zilongjin tablets are produced by Longshun banyan pharmaceutical factory, a patent number of ZL01144161.5, from Tianjin New pharmaceutical industry group, Inc.
Thin layer chromatography
Thin-layer chromatography, also called as plate chromatography, is one of chromatography, is an important experimental technique for rapid separation and qualitative analysis of a small amount of substances, and belongs to solid-liquid adsorption chromatography.
Developing agent
In the extraction and separation of chemical substances, a solvent used for separating two substances having different polarities is called a developing solvent, and it is a primary task to select a proper kind and ratio of developing solvent.
In a preferred embodiment, the developing agent for thin layer chromatography is a mixed solution of ethyl acetate, butanone, formic acid and water, and the volume ratio of the developing agent to the water is 8-9: 3-4: 2-3: 2-3.
The volume ratio is 8-9: 3-4: 2-3: the mixed solution of ethyl acetate, butanone, formic acid and water of 2-3 is used as developing solvent to separate the analyzed substances effectively.
Thin layer chromatography silica gel plate
The thin-layer chromatography silica gel plate is prepared by mixing high-purity thin-layer chromatography silica gel (powder) with a certain amount of binder and spraying, and the plate surface is pure white, flat, uniform and fine; the chemical property is stable, and the reaction with other acids and bases except strong base and hydrofluoric acid is avoided; the thin layer chromatography silica gel plate can be directly used for qualitative or quantitative analysis of various organic substances.
In a preferred embodiment, the silica gel plate of the thin layer chromatography is a silica gel G plate.
Further, the silica gel G plate comprises a smoke bench silica gel G plate or a MERCK silica gel G plate.
In a preferred embodiment, the thin layer chromatography is adopted to analyze the test solution, the reference solution and the negative sample solution to identify whether the barbat skullcap herb ingredient is contained in the rhododendron simsii planch tablet or not, and the method comprises the following steps:
developing the test solution, the reference solution and the negative sample solution according to thin-layer chromatography, and identifying as containing herba Scutellariae Barbatae medicinal material when spots of the same color appear at the corresponding positions of the test chromatogram and the reference chromatogram and the negative sample has no interference;
wherein the volume ratio of ethyl acetate-butanone-formic acid-water in the developing solvent is 9: 4: 2: 2;
in a preferred embodiment, developing according to thin layer chromatography comprises the steps of:
taking the independent amount of each of the three solutions to be 4-8 mu l, respectively dropping the three solutions on the same silica gel G plate, and mixing the three solutions according to the volume ratio of 9: 4: 2: 2 ethyl acetate-butanone-formic acid-water solution, taking out, drying and developing with developer.
Typical but non-limiting independent amounts of test solution are for example 4. mu.l, 5. mu.l, 6. mu.l, 7. mu.l, 8. mu.l;
preferably, the color developing agent is ferric chloride ethanol solution;
further, the mass concentration of the ferric trichloride ethanol solution is 4-6%;
further preferably, the mass concentration of the ferric trichloride ethanol solution is 5%;
the ferric trichloride ethanol solution can realize effective color development on target analysis substances.
When spots with the same color appear on the chromatogram of the test sample at the positions corresponding to the chromatogram of the reference sample, and the negative sample has no interference, the sample is identified as containing the scutellariae barbatae medicinal material.
Typical but not limiting mass concentrations of ferric chloride ethanol solution are 4%, 5%, 6%.
Further, the development temperature of the thin layer chromatography is less than or equal to 10 ℃ and 16-26 ℃;
further, the development relative humidity of the thin layer chromatography is greater than or equal to 75%, less than or equal to 50%.
Typical, but not limiting, development temperatures for thin layer chromatography are, for example, 1 ℃, 2 ℃, 3 ℃, 4 ℃, 5 ℃, 6 ℃, 7 ℃, 8 ℃, 9 ℃, 10 ℃.
Typical, but not limiting, development temperatures of thin layer chromatography are, for example, 16 ℃, 17 ℃, 18 ℃, 19 ℃, 20 ℃, 21 ℃, 22 ℃, 23 ℃, 24 ℃, 25 ℃, 26 ℃.
Typical but non-limiting developed relative humidities of thin layer chromatography are, for example, 75%, 80%, 85%, 90%.
Typical but non-limiting developed relative humidities of thin layer chromatography are, for example, 10%, 20%, 30%, 40%, 50%.
Under the development conditions of the temperature and the relative humidity, the thin-layer chromatography experiment can effectively identify the sculellaria barbata.
According to the second aspect of the invention, the thin-layer chromatography method for identifying the barbed skullcap herb is applied to the quality evaluation of the zilongjin tablets.
The method provided by the invention can be used for simply and accurately determining whether the barbat skullcap medicinal ingredient is contained in the Zilongjin tablets, and has the advantages of strong specificity, high durability and small error.
The invention is further illustrated by the following examples. The materials in the examples are prepared according to known methods or are directly commercially available, unless otherwise specified.
The Zilongjin tablets in the examples were produced by Longshun banyan pharmaceutical factory, New pharmaceutical group, Inc. in Tianjin.
Example 1
Preparation of a test solution: taking 15 tablets of the product of Zilongjin tablets, removing coatings, grinding, adding 50ml of methanol, heating and refluxing for 40 minutes, cooling, filtering, evaporating filtrate to dryness, adding 5ml of water into residues to dissolve, and processing by a method D101Eluting with 10% ethanol 70ml, discarding eluate, eluting with 30% ethanol 50ml, collecting eluate, evaporating to dry, and dissolving residue with methanol 2ml to obtain the final product.
Preparation of control solutions: adding methanol into scutellarin control to obtain solution containing 1 mg/1 ml.
Preparation of negative sample solution: according to the proportion of the formula of the Zilongjin tablet, taking other medicines except the barbed skullcap herb, preparing the preparation according to the preparation process of the Zilongjin tablet, and then operating according to the preparation method of the test solution to obtain the Zilongjin tablet.
And (2) operating at the temperature T of 27 ℃ and the relative humidity RH of 75%, sucking 4-8 mu l of each of the two solutions, respectively dropping the two solutions on the same high-efficiency silica gel G thin-layer plate, developing by taking ethyl acetate-butanone-formic acid-water (9: 4: 2: 2) as a developing agent, taking out, drying in the air, and spraying a 5% ferric trichloride ethanol solution. In the chromatogram of the test sample, spots of the same color appear at the corresponding positions of the chromatogram of the reference sample, and the negative sample has no interference, i.e. the sample is identified as containing the medicinal component of the herba Scutellariae Barbatae, the result of the thin layer plate is shown in figure 1, from left to right: a negative sample; sample (batch: BE 35503); sample (batch: BE 35509); sample (batch number: CI 35548); scutellarin control (Zhongzhong hospital 842-200102).
Example 2
The operating conditions for this example were T21 ℃ and RH 37%, the results for the lamella plates are shown in fig. 2, from left to right: a negative sample; sample (batch: BE 35503); sample (batch: BE 35509); sample (batch number: CI 35548); scutellarin control (Zhongzhong hospital 842-200102).
Example 3
The operating conditions for this example were T4 ℃ and RH 42%, the results for the lamella plates are shown in fig. 3, from left to right, as in example 1: a negative sample; sample (batch: BE 35503); sample (batch: BE 35509); sample (batch number: CI 35548); scutellarin control (Zhongzhong hospital 842-200102).
Example 4
The operating conditions for this example were T24 ℃ and RH 75%, the results for the lamella plates are shown in fig. 4, from left to right, as in example 1: a negative sample; sample (batch: BE 35503); sample (batch: BE 35509); sample (batch number: CI 35548); scutellarin control (Zhongzhong hospital 842-200102).
Analysis of
By analyzing the thin-layer chromatography results of examples 1-4, it can be seen that the thin-layer chromatography method for identifying the barbat skullcap component in the zilongjin tablets of the present application has strong durability and the influence of the operating temperature and the relative humidity on the thin-layer chromatography method is small. The thin-layer chromatography method provided by the invention can accurately identify the medicinal material components of the sculellaria barbata in the zilongjin tablets, and the method not only has the advantages of high accuracy and small identification error, but also has the advantages of strong specificity, strong durability, good reproducibility, simple operation and the like; the thin-layer chromatography method can be applied to the quality evaluation of the Zilongjin tablets.
Finally, it should be noted that: the above embodiments are only used to illustrate the technical solution of the present invention, and not to limit the same; while the invention has been described in detail and with reference to the foregoing embodiments, it will be understood by those skilled in the art that: the technical solutions described in the foregoing embodiments may still be modified, or some or all of the technical features may be equivalently replaced; and the modifications or the substitutions do not make the essence of the corresponding technical solutions depart from the scope of the technical solutions of the embodiments of the present invention.
Claims (10)
1. A thin-layer chromatography method for identifying the medicinal material components of sculellaria barbata in a zilongjin tablet is characterized by comprising the following steps:
providing a Zilongjin tablet solution as a test solution, providing a scutellarin solution as a reference solution, and providing a preparation solution prepared from the rest medicinal ingredients except the sculellaria barbata in the Zilongjin tablet as a negative sample solution;
analyzing the test solution, the reference solution and the negative sample solution by adopting a thin-layer chromatography to identify whether the barbat skullcap medicinal ingredient exists in the Zilongjin tablets;
the developing agent of the thin-layer chromatography is a mixed solution of ethyl acetate, butanone, formic acid and water, and the volume ratio is 8-9: 3-4: 2-3: 2-3.
2. The thin layer chromatography method of claim 1, wherein the silica gel plate of the thin layer chromatography is a silica gel G plate.
3. The thin layer chromatography method of claim 2, wherein the silica gel G plate comprises a bench silica gel G plate or a MERCK silica gel G plate.
4. The thin layer chromatography method of claim 1, wherein the thin layer chromatography is used for analyzing the test solution, the reference solution and the negative sample solution to identify the sculellaria barbata medicinal material component in the zilongjin tablet, and the method comprises the following steps:
the test solution, the reference solution and the negative sample solution are developed according to the thin-layer chromatography, when spots with the same color appear in the chromatogram of the test solution at the positions corresponding to the chromatogram of the reference solution and the negative sample is not interfered, the sample is identified as containing the medicinal component of the barbed skullcap herb;
preferably, the volume ratio of ethyl acetate, butanone, formic acid and water in the developing solvent is 9: 4: 2: 2;
preferably, developing according to thin layer chromatography comprises the steps of:
taking the independent amount of each of the three solutions to be 4-8 mu l, respectively dropping the three solutions on the same silica gel G plate, and mixing the three solutions according to the volume ratio of 9: 4: 2: 2, developing with a mixed solution of ethyl acetate, methyl ethyl ketone, formic acid and water, taking out, drying and developing with a color-developing agent.
5. The thin layer chromatography method of claim 4, wherein the color-developing agent is an ethanol solution of ferric chloride;
preferably, the mass concentration of the ferric trichloride ethanol solution is 4-6%;
preferably, the mass concentration of the ferric trichloride ethanol solution is 5%.
6. The thin layer chromatography method of claim 1, wherein the development temperature of the thin layer chromatography is less than or equal to 10 ℃.
7. The thin layer chromatography method of claim 1, wherein the thin layer chromatography is performed at a development temperature of 16 ℃ to 26 ℃.
8. The thin layer chromatography method of claim 1, wherein the developed relative humidity of the thin layer chromatography is greater than or equal to 75%.
9. The thin layer chromatography method of claim 1, wherein the developed relative humidity of the thin layer chromatography is less than or equal to 50%.
10. Use of the thin layer chromatography method according to any one of claims 1 to 9 in the quality evaluation of Zilongjin tablets.
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