CN112557526B - Quality-controllable isatis root extract and method for identifying isatis root containing raw material medicine in Chinese patent medicine - Google Patents

Abstract

The invention provides an isatis root extract with controllable quality and a method for identifying isatis root containing raw material medicine in Chinese patent medicine. The isatis root extract is detected by adopting a gas chromatography, and at least contains the following characteristic peaks: peak 1: retention time 9.46 ± 0.050min, peak 2: the retention time is 10.95 plus or minus 0.050 min; the gas chromatography conditions are as follows: a chromatographic column: DB-WAX capillary column; carrier gas: nitrogen with the flow rate of carrier gas being 1-3 mL/min; a detector: a FID detector for detecting the temperature of 220-240 ℃; sample inlet temperature: 220 to 240 ℃. The radix isatidis extract is further used as a reference substance, the radix isatidis medicinal material in the Chinese patent medicine (such as anemopyretic cold granules) can be accurately identified by adopting the gas chromatography condition, the identification method is safe and environment-friendly, has good specificity, good durability, good selectivity and high sensitivity, and provides a basis for establishing the quality standard of the Chinese patent medicine containing the radix isatidis medicinal material.

Description

Quality-controllable isatis root extract and method for identifying isatis root containing raw material medicine in Chinese patent medicine

Technical Field

The invention belongs to the field of quality control of traditional Chinese medicine extracts and Chinese patent medicines, and particularly relates to an isatis root extract with controllable quality and a method for identifying isatis root containing raw material medicines in a Chinese patent medicine.

Background

The Isatis root is a dried root of Isatis indigotica fort of cruciferae, is a long-history traditional Chinese medicine and has the effects of clearing away heat and toxic materials, cooling blood, relieving sore throat and the like. Since 1977, isatis roots were collected in the calendar edition "Chinese pharmacopoeia", and were used not only as decoction pieces in Chinese medicine formulas, but also as industrial raw materials for various Chinese patent medicines. The chemical composition of isatis root is extremely complex, and over the past decades, many scholars have been devoted to the study of the active ingredients of isatis root, and have isolated up to about 200 compounds from it, which are related to the structural types of alkaloids, organic acids, anthraquinones, flavones, lignans, triterpenes, sterols, sinaposide, nucleosides, fatty acids, amino acids, carbohydrates, etc. Chinese patent application No. CN200510012767.7 discloses an isatis root extract, which is prepared by the following preparation steps: taking radix isatidis decoction pieces, adding 5-15 times of 50-85% ethanol solution, placing in a suitable container, sealing, and cold soaking; heating and reflux extracting, filtering, and recovering ethanol under reduced pressure to obtain medicinal liquid; controlling the vacuum degree to be 20-90 kPa, and concentrating the liquid medicine under reduced pressure to obtain thick paste; drying the thick paste at 50-80 ℃ to obtain a product. However, the components in the extract are complex, the quality controllability is poor, and the application does not provide a method for identifying the Chinese patent medicine containing the isatis root by using the extract.

The wind-heat type common cold granule is a Chinese patent medicine containing isatis root which is commonly used in clinic, is prepared from 11 traditional Chinese medicines such as isatis root, weeping forsythia, reed rhizome, great burdock achene, chrysanthemum, weeping forsythia and the like, has the functions of clearing away heat and toxic materials, freeing lung and relieving sore throat, and is used for treating cold fever, nasal obstruction, headache, cough and excessive phlegm. The legal standard of the wind-heat type common cold granule is the ministerial issued drug standard, namely the first volume of the drug standard Chinese medicinal prescription preparation of Ministry of health (WS 3-0044-089). Only the prescription, preparation and properties of the granules are required from the issuance to the present, and the granules are checked according to the general rule of the granules without other quality control items. At present, the variety can only be tested according to the characters of the standard and the content under the general regulation of the granules in the four parts of the 2015 edition of Chinese pharmacopoeia. Therefore, in order to further accurately control the quality of the anemopyretic cold particles, more quality control items need to be established.

The document "thin-layer chromatography identification of wind-heat common cold granules, great dawn yoge, etc" records a method for qualitatively identifying the main medicinal materials of the medicine, namely isatis root, reed rhizome, great burdock achene and weeping forsythia, by adopting a thin-layer chromatography (TLC) method. The result shows that the radix isatidis in the wind-heat type common cold granules can be qualitatively identified by using the thin-layer chromatography and taking n-butyl alcohol, anhydrous ethanol, chloroform, acetic acid and water (5: 5: 2: 1: 1) as a developing agent. However, the end of the document also describes that the isatis root medicinal material contains glutamic acid, proline, monoaminobutyric acid, arginine and the like, wherein the polarity of arginine is stronger, and when the temperature is lower, the polarity of the thin layer chromatography developing agent can be changed. Therefore, the durability of the radix isatidis in the wind-heat type common cold granules is poor when the thin layer chromatography is used for qualitatively identifying the radix isatidis. In addition, the method described in this document does not use the specific index components of isatis root for identification, and therefore, the isatis root in the wind-heat type common cold granule cannot be effectively distinguished from other medicinal materials, which is not favorable for identification of isatis root in the wind-heat type common cold granule. Moreover, thin-layer chromatography often uses toxic and harmful organic reagents, which are harmful to the environment and personnel to some extent. Therefore, a method which is safe, environment-friendly, good in durability, good in specificity and high in sensitivity is urgently needed to be developed to further identify the raw material medicines in the wind-heat type common cold granules, and a basis is provided for establishing the quality standard of the wind-heat type common cold granules.

At present, no report is available for identifying the isatis root in a Chinese patent medicine (such as anemopyretic cold granules) containing the isatis root by using an isatis root extract with controllable quality as a reference through gas chromatography.

Disclosure of Invention

The invention aims to provide an isatis root extract with controllable quality, and the invention also aims to provide a method for identifying the isatis root containing the raw material medicine in the Chinese patent medicine, so that a basis is provided for establishing the quality standard of the Chinese patent medicine containing the isatis root as the raw material medicine.

The invention provides an isatis root extract, which is detected by adopting gas chromatography and at least comprises the following characteristic peaks: peak 1: retention time 9.46 ± 0.050min, peak 2: the retention time is 10.95 plus or minus 0.050 min;

the gas chromatography conditions are as follows:

a chromatographic column: DB-WAX capillary column;

carrier gas: nitrogen with the flow rate of carrier gas being 1-3 mL/min;

a detector: a FID detector for detecting the temperature of 220-240 ℃;

sample inlet temperature: 220 to 240 ℃.

Further, in the gas chromatography conditions, the chromatographic column is a DB-WAX capillary column, and the size of the chromatographic column is 30m multiplied by 0.32mm multiplied by 0.50 mu m; the flow rate of the carrier gas is 2 mL/min; the detection temperature is 230 ℃; the temperature of the sample inlet is 230 ℃;

And/or in the gas chromatography condition, the sample injection amount is 0.5-5.0 mu L, preferably 1 mu L; the gas chromatography condition adopts split sampling, and the split ratio is 1: 1.

Further, in the gas chromatography conditions, the column temperature is a programmed temperature: initial temperature 50 ℃, then 5 ℃ per minute to 140 ℃, held for 4 minutes; then raising the temperature to 200 ℃ at 20 ℃ per minute, and keeping the temperature for 1 minute; the temperature was then increased to 235 ℃ at 50 ℃ per minute and held for 3 minutes.

Further, the preparation method of the isatis root extract comprises the following steps: decocting radix Isatidis in water, filtering, collecting filtrate, adding alkali, distilling with steam, collecting distillate, extracting with organic solvent, standing, and collecting organic phase to obtain radix Isatidis extract.

Further, the decocting time is 1-4 hours; the mass volume ratio of the isatis root medicinal material to water is 1: (15-25) g/mL; the alkali is inorganic alkali, preferably sodium hydroxide;

and/or the organic solvent is petroleum ether; the volume ratio of the distillate to the petroleum ether is (40-60) to 1; the extraction mode is shaking extraction.

Further, the decoction time is 2 hours; the mass volume ratio of the isatis root medicinal material to water is 1:20 g/mL; the mass-volume ratio of the alkali to the filtrate is 0.05-0.40 g/mL, preferably 0.20 g/mL;

And/or the petroleum ether is 60-90 ℃ petroleum ether; the volume ratio of the distillate to the petroleum ether is 50: 1.

The invention also provides a method for identifying the isatis root contained in the raw material medicine in the Chinese patent medicine, and the identification method comprises the following steps:

a. preparing the Chinese patent medicine into a test solution;

b. preparing the radix Isatidis extract into reference solution;

c. detecting by adopting the gas chromatography condition;

d. if the gas chromatogram of the test solution at least contains the following characteristic peaks: the retention time of peak 1 is 9.46 plus or minus 0.050min, and the retention time of peak 2 is 10.95 plus or minus 0.050min, so that the Chinese patent medicine contains radix isatidis; if the following characteristic peaks are not contained or only contained, the method comprises the following steps: the retention time of peak 1 is 9.46 plus or minus 0.050min, and the retention time of peak 2 is 10.95 plus or minus 0.050min, so that the Chinese patent medicine does not contain isatis root.

Further, in step a, the preparation method of the test solution comprises: adding an alkaline aqueous solution into the Chinese patent medicine, distilling with steam, collecting distillate, adding an organic solvent into the distillate for extraction, standing after extraction, and taking an organic phase to obtain a test solution;

preferably, the Chinese patent medicine is wind-heat type common cold granules.

Further, the alkaline aqueous solution is an aqueous solution of an inorganic base, preferably an aqueous solution of sodium hydroxide; the mass volume ratio of the Chinese patent medicine to the alkaline aqueous solution is 0.5-2.0: 1 g/mL;

and/or the organic solvent is petroleum ether; the volume ratio of the distillate to the petroleum ether is (40-60) to 1; the extraction mode is shaking extraction.

Further, the concentration of the sodium hydroxide aqueous solution is 0.05-0.40 g/mL, preferably 0.20 g/mL; the mass volume ratio of the Chinese patent medicine to the alkaline aqueous solution is 1: 1 g/mL;

and/or the petroleum ether is 60-90 ℃ petroleum ether; the volume ratio of the distillate to the petroleum ether is 50: 1.

In the invention, the petroleum ether (60-90 ℃), namely the petroleum ether is 60-90 ℃, refers to the fraction of the petroleum ether with the initial boiling point not lower than 60 ℃ and the final boiling point not higher than 90 ℃.

In the present invention, the sodium hydroxide solution concentration refers to a mass-volume concentration. For example, a 20% sodium hydroxide solution refers to a solution having a sodium hydroxide concentration of 20% g/mL, i.e., 0.2 g/mL.

The isatis root extract provided by the invention is controllable in quality, and by taking the isatis root extract as a reference substance, the isatis root medicinal material in the wind-heat type common cold particle can be accurately identified by utilizing the gas chromatography condition of the isatis root extract.

The method of the invention can not only qualitatively identify the isatis root medicinal material in the wind-heat common cold granules, and take the compound 2, 4-pentadienitrile corresponding to the main peaks 1 and 2 as a reference substance, but also quantitatively determine the content of the isatis root medicinal material in the wind-heat common cold granules.

The method of the invention can be used for identifying the isatis root medicinal material in the wind-heat type common cold granules and can also be used for identifying the isatis root medicinal material in other Chinese patent medicines, thereby providing a basis for establishing the quality standard of the Chinese patent medicine containing the isatis root medicinal material.

Obviously, many modifications, substitutions, and variations are possible in light of the above teachings of the invention, without departing from the basic technical spirit of the invention, as defined by the following claims.

The present invention will be described in further detail with reference to the following examples. This should not be understood as limiting the scope of the above-described subject matter of the present invention to the following examples. All the technologies realized based on the above contents of the present invention belong to the scope of the present invention.

Drawings

FIG. 1 is a gas chromatogram of a control solution.

FIG. 2 is a gas chromatogram of a sample solution.

FIG. 3 is a gas chromatogram of a isatis root-lacking negative sample solution.

FIG. 4 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles produced by company A.

FIG. 5 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles produced by company B.

FIG. 6 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles produced by company C.

FIG. 7 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles produced by company D.

FIG. 8 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles produced by company E.

FIG. 9 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles produced by F corporation.

FIG. 10 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles produced by G corporation.

FIG. 11 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles produced by the company H.

FIG. 12 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles produced by company I.

FIG. 13 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles manufactured by J.

FIG. 14 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles manufactured by K corporation.

FIG. 15 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles manufactured by L company.

FIG. 16 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles produced by M corporation.

FIG. 17 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles produced by N corporation.

FIG. 18 is a gas chromatogram of a test solution obtained by using anemopyretic cold particles produced by O corporation.

FIG. 19 is a total ion flow chart of the reference radix Isatidis solution by GC-MS.

FIG. 20 is an ion mass spectrum of a compound corresponding to main peak 1.

FIG. 21 is a structural diagram of a compound corresponding to main peak 1.

FIG. 22 is an ion mass spectrum of a compound corresponding to main peak 2.

FIG. 23 is a structural diagram of a compound corresponding to main peak 2.

FIG. 24 is a gas chromatogram of the sample solution corresponding to the sodium hydroxide solution of Table 2 at a concentration of 20%.

Detailed Description

The raw materials and equipment used in the invention are known products and are obtained by purchasing commercial products.

The instrument comprises the following steps: agilent 7890B gas chromatograph (Agilent technologies, ltd); XPE26 electronic balance (mettler-toledo switzerland); AE-240 electronic balance (sydow sedolis, germany); BUCHI Distillation Unit K-350 steam still.

The radix Isatidis control drug (121177) -201608) was provided by China food and drug assay institute.

Example 1 gas chromatography identification method of Isatis indigotica fort in anemopyretic cold granules

The method is tested according to gas chromatography (0521 of the general rules of the four departments in the Chinese pharmacopoeia 2020 edition), and comprises the following specific steps:

preparation of reference drug solution: taking 5g of radix isatidis reference medicinal material, adding 100mL of water, decocting for 2 hours, filtering to obtain 50mL of filtrate, placing the filtrate in a distillation tube, adding 10g of sodium hydroxide, performing steam distillation, collecting 250mL of distillate, placing the distillate in a volumetric flask (the volumetric flask is filled with 5mL of petroleum ether (60-90 ℃) in advance), shaking for extraction, shaking uniformly, standing, and taking the upper petroleum ether solution as the reference medicinal material solution.

Preparation of a test solution: taking 50g of commercial anemopyretic cold particles, placing the particles in a distillation tube, adding 50mL of 20% sodium hydroxide solution, carrying out steam distillation, collecting 250mL of distillate, placing the distillate in a volumetric flask (5 mL of petroleum ether (60-90 ℃) is filled in the volumetric flask in advance), shaking for extraction, standing after shaking uniformly, and taking the upper layer petroleum ether solution as a test solution.

The determination method comprises precisely sucking 1 μ L of reference medicinal material solution and sample solution respectively, injecting into gas chromatograph, and determining. The chromatographic conditions were as follows:

DB-WAX (polyethylene glycol) capillary column (30 m.times.0.32 mm.times.0.50 μm) was used; FID detector, temperature 230 ℃; the injection port temperature is 230 ℃; split-flow sample injection is carried out, and the split-flow ratio is 1: 1; the carrier gas is nitrogen, and the flow rate of the carrier gas is as follows: 2 mL/min; column temperature is temperature programmed (see table 1): the initial temperature was 50 ℃ at 5 ℃ to 140 ℃ per minute for 4 minutes, 20 ℃ to 200 ℃ per minute for 1 minute, and 50 ℃ to 235 ℃ per minute for 3 minutes. The number of theoretical plates is calculated according to the main peak 1 of radix Isatidis and should not be lower than 2 × 10⁵。

TABLE 1 column temperature programmed temperature settings

Rate of temperature rise (. degree. C./minute)	Column oven temperature (. degree. C.)	Retention time (minutes)
			-	50	0
5	140	4
			20	200	1
50	235	3

The results are shown in FIGS. 1-2. In the embodiment, the chromatogram of the sample solution extracted by the steam distillation method has less impurities, good peak shape and small interference. And 2 main chromatographic peaks (main peak 1: 9.46min, main peak 2: 10.95min) with the same retention time as that in the chromatogram of the reference medicinal material solution appear in the chromatogram of the test solution, and the radix isatidis medicinal material is judged to be contained in the test product wind-heat common cold granules.

The beneficial effects of the present invention are demonstrated by the following experimental examples.

Experimental example 1 chromatographic column screening experiment

1. Experimental methods

Referring to the test method of example 1, the DB-WAX (polyethylene glycol) capillary column (30 m.times.0.32 mm.times.0.50 μm) was replaced with a DB-WAX (polyethylene glycol) capillary column (30 m.times.0.53 mm.times.1.0 μm) or an HP-5 capillary column (30 m.times.0.32 mm. times.0.25 μm), respectively, and the resulting chromatograms were analyzed with the remaining test conditions unchanged.

2. Results of the experiment

As a result, it was found that when a DB-WAX (polyethylene glycol) capillary column (30 m. times.0.32 mm. times.0.50 μm) was used, the theoretical plate number was high, the degree of separation was the best, and the retention time was moderate. Therefore, a DB-WAX (polyethylene glycol) capillary column (30m multiplied by 0.32mm multiplied by 0.50 mu m) is selected.

Experimental example 2 screening of method for preparing test solution

2.1 screening of the extraction solvent by shaking

A test solution was prepared by referring to the method of example 1, and only petroleum ether (60 to 90 ℃ C.) was replaced with ethyl acetate. However, it was found that the system could not be separated and the solvent was miscible when it was allowed to stand after shaking.

Therefore, petroleum ether (60-90 ℃) is selected as a shaking extraction solvent.

2.2 screening of extraction methods

Referring to the test method of example 1, only the preparation methods of the test solution and the reference solution were modified to the following volatile oil extraction methods:

the preparation method of the test solution comprises the following steps: taking 50g of anemopyretic cold particles, placing the anemopyretic cold particles in a volatile oil extractor, connecting the volatile oil extractor, adding 350mL of 20% sodium hydroxide solution, adding 5mL of petroleum ether (60-90 ℃) into a branch pipe of the volatile oil extractor, keeping slight boiling for 3 hours according to a volatile oil determination method (2204 in the four departments of the 2015 version of Chinese pharmacopoeia), and taking petroleum ether liquid to a constant volume of 5mL to serve as a test solution.

The preparation method of the reference medicinal material solution comprises the following steps: taking 5g of isatis root as a reference medicinal material, adding 100mL of water, decocting for 2 hours, filtering to obtain 50mL of filtrate, placing the filtrate in a volatile oil extractor, adding 10g of sodium hydroxide, adding 5mL of petroleum ether (60-90 ℃) into a branch pipe of the volatile oil extractor, keeping slight boiling for 3 hours according to a volatile oil determination method (2204 in 2015 edition of the four-part general rules), taking petroleum ether liquid, and fixing the volume to 5mL to be used as a reference medicinal material solution.

The result shows that the chromatogram of the sample solution obtained by the volatile oil extraction method has many impurities and unclear peak shape. Thus, the steam distillation method of example 1 was selected to prepare a test solution.

2.3 examination of extraction Effect by different concentrations of alkali

Referring to the test method of example 1, only the preparation method of the test article solution was modified: taking 50g of a reference preparation (anemopyretic cold particles provided by taiji group, sichuan mianyang pharmaceutical limited, batch number 200403), respectively adding 50mL of 5%, 10%, 20%, 30% and 40% sodium hydroxide solution, performing steam distillation, collecting 250mL of distillate, placing in a volumetric flask (petroleum ether (60-90 ℃) is pre-filled in the volumetric flask for 5mL), shaking for extraction, shaking uniformly, standing, and taking an upper layer petroleum ether solution as a test solution.

The results of comparing the characteristic peak areas of isatis roots in a gas chromatogram obtained by detecting the test solution in the presence of alkali with different concentrations are shown in table 2. The gas chromatogram obtained at a sodium hydroxide solution concentration of 20% is shown in FIG. 24.

TABLE 2 Effect of different concentrations of alkali on the identification of Isatis root

As is clear from Table 2, in the preparation of the test solutions, when the concentration of the sodium hydroxide solution was 20%, the peak area of the characteristic peak of Isatis root was the largest in the chromatograms obtained. So 20% sodium hydroxide solution is selected when preparing the test solution.

Experimental example 3 specificity test

Control solutions and test solutions were prepared in the same manner as in example 1.

Then preparing a isatis root-lacking negative sample solution: taking the other medicinal ingredients except the isatis root in the prescription of the wind-heat type common cold granule to prepare a negative sample lacking the isatis root. Then, referring to the method for preparing the test solution of example 1, only anemopyretic cold granules were replaced with the above-mentioned negative sample lacking isatis root to prepare a negative sample solution lacking isatis root.

Precisely sucking 1 μ L of each of the reference medicinal material solution, the test sample solution and the radix Isatidis-lacking negative sample solution, and respectively injecting into a gas chromatograph for determination. The chromatographic conditions were the same as in example 1.

The results are shown in FIGS. 1 to 3. 2 main chromatographic peaks (main peak 1: 9.46min, main peak 2: 10.95min) with the same retention time as that in the chromatogram of the reference medicinal material solution appear in the chromatogram of the test solution. And if the isatis root negative sample solution is absent, no chromatographic peak is present at the corresponding retention time of the chromatogram of the control medicinal material solution. The defect of the isatis root negative sample does not interfere the identification of the isatis root, and the test method has good specificity.

Experimental example 4 examination of durability

Each test sample solution was prepared by referring to the method of example 1 using commercially available anemopyretic cold particles produced by different manufacturers (A to O) as samples, and then a chromatogram of each test sample solution was obtained according to the test method of example 1.

The results are shown in FIGS. 4 to 18, and it can be seen that the test method of the present invention is excellent in durability.

Experimental example Structure confirmation of Compound corresponding to 52 Main chromatographic peaks

A control solution of Isatis root was prepared according to the method of example 1, followed by confirmation of composition by gas chromatography-mass spectrometry (gas chromatography conditions were the same as in example 1). The obtained Total Ion Current (TIC) spectrum is shown in FIG. 19, and the ion mass spectrum is shown in FIGS. 20 and 22.

After analysis, the compounds corresponding to the main peaks 1 and 2 of the isatis root reference medicinal material solution are determined to be 2, 4-pentadienitrile. The molecular weight and molecular formula of the compounds corresponding to the two main peaks are the same, cis-trans structures exist on the spatial conformation of the molecules, and the main peaks 1 and 2 are deduced to be cis-trans isomers. The NIST MS search structure of the compound corresponding to the main peak 1 is shown in FIG. 21, and the NIST MS search structure of the compound corresponding to the main peak 1 is shown in FIG. 23.

Therefore, by using the method of the invention, the compound 2, 4-pentadienitrile corresponding to the main peaks 1 and 2 as a reference substance, the content of the isatis root medicinal material in the wind-heat-sensing common cold particles can be quantitatively measured.

In conclusion, the invention provides an isatis root extract and a method for identifying the isatis root containing the raw material medicine in Chinese patent medicine. The radix isatidis extract is controllable in quality, the radix isatidis extract is used as a reference substance, the radix isatidis medicinal material in the wind-heat type common cold particles can be accurately identified by utilizing the gas chromatography condition of the method, the method is safe and environment-friendly, good in specificity, good in durability, good in selectivity and high in sensitivity, and a basis is provided for establishing the quality standard of a Chinese patent medicine containing the radix isatidis medicinal material.

Claims (8)

1. A method for identifying the radix isatidis containing the raw material medicine in Chinese patent medicine is characterized by comprising the following steps: the method comprises the following steps:

a. preparing the Chinese patent medicine into a test solution;

in the step a, the preparation method of the test solution comprises the following steps: adding an alkaline aqueous solution into the Chinese patent medicine, distilling with steam, collecting distillate, adding an organic solvent into the distillate for extraction, standing after extraction, and taking an organic phase to obtain a test solution; the Chinese patent medicine is anemopyretic cold granules, the organic solvent is petroleum ether, and the alkaline aqueous solution is a sodium hydroxide aqueous solution;

b. Preparing radix Isatidis extract into reference solution; the isatis root extract is detected by gas chromatography, and at least contains the following characteristic peaks: peak 1: retention time 9.46 ± 0.050min, peak 2: the retention time is 10.95 plus or minus 0.050 min;

in the step b, the preparation method of the isatis root extract comprises the following steps: decocting radix Isatidis in water, filtering, collecting filtrate, adding alkali, distilling with steam, collecting distillate, extracting with organic solvent, standing, and collecting organic phase to obtain radix Isatidis extract; the organic solvent is petroleum ether, and the alkali is sodium hydroxide;

c. detecting by adopting gas chromatography;

the gas chromatography conditions are as follows:

and (3) chromatographic column: DB-WAX capillary column, 30 mm × 0.32mm × 0.50 μm;

carrier gas: nitrogen and carrier gas flow rate is 2 mL/min;

a detector: a FID detector, the detection temperature is 230 ℃;

sample inlet temperature: 230 ℃;

in the gas chromatography condition, the sample injection amount is 0.5-5.0 mu L; the gas chromatography condition adopts split sample injection, and the split ratio is 1: 1; column temperature is programmed temperature rise: initial temperature 50 ℃, then 5 ℃ per minute to 140 ℃, held for 4 minutes; then raising the temperature to 200 ℃ at 20 ℃ per minute, and keeping the temperature for 1 minute; then raising the temperature to 235 ℃ at 50 ℃ per minute, and keeping the temperature for 3 minutes;

d. If the gas chromatogram of the test solution at least contains the following characteristic peaks: the retention time of the peak 1 is 9.46 +/-0.050 min, and the retention time of the peak 2 is 10.95 +/-0.050 min, so that the Chinese patent medicine contains isatis root medicinal materials; if the following characteristic peaks are not contained or only contained: the retention time of the peak 1 is 9.46 +/-0.050 min, and the retention time of the peak 2 is 10.95 +/-0.050 min, so that the Chinese patent medicine does not contain isatis root medicinal materials.

2. The method of claim 1, wherein: the mass volume ratio of the Chinese patent medicine to the alkaline aqueous solution is 0.5-2.0: 1 g/mL;

and/or the volume ratio of the distillate to the petroleum ether is (40-60): 1; the extraction mode is shaking extraction.

3. The method of claim 1, wherein: the concentration of the sodium hydroxide aqueous solution is 0.05-0.40 g/mL; the mass volume ratio of the Chinese patent medicine to the alkaline aqueous solution is 1: 1 g/mL;

and/or the petroleum ether is 60-90 ℃ petroleum ether; the volume ratio of the distillate to the petroleum ether is 50: 1.

4. The method of claim 3, wherein: the concentration of the sodium hydroxide aqueous solution is 0.20 g/mL.

5. The method of claim 1, wherein: the sample size was 1 μ L.

6. The method of claim 1, wherein: in the step (b), the decoction time is 1-4 hours; the mass volume ratio of the isatis root medicinal materials to the water is 1: (15-25) g/mL; and/or the volume ratio of the distillate to the petroleum ether is (40-60): 1; the extraction mode is shaking extraction.

7. The method of claim 6, wherein: in the step (b), the decoction time is 2 hours; the mass volume ratio of the isatis root medicinal material to water is 1:20 g/mL; the mass volume ratio of the alkali to the filtrate is 0.05-0.40 g/mL;

and/or the petroleum ether is 60-90 ℃ petroleum ether; the volume ratio of the distillate to the petroleum ether is 50: 1.

8. The method of claim 7, wherein: the mass-to-volume ratio of the alkali to the filtrate was 0.20 g/mL.

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