CN110794080A

CN110794080A - Method for detecting quality of medicine for treating colitis

Info

Publication number: CN110794080A
Application number: CN201911141768.XA
Authority: CN
Inventors: 刘金磊; 周建军; 曹珍荣; 石红艳; 赵颖
Original assignee: Liu Shengmei
Current assignee: Liu Shengmei
Priority date: 2019-11-20
Filing date: 2019-11-20
Publication date: 2020-02-14

Abstract

The invention provides a method for detecting the quality of a medicament for treating colitis, which adopts a microscope method to identify pseudo-ginseng and bletilla striata; identifying radix astragali and Notoginseng radix by thin layer chromatography; identifying Galla chinensis by thin layer chromatography; measuring the content of Galla chinensis by high performance liquid chromatography; measuring radix astragali content by high performance liquid chromatography; the detection method of the invention can effectively ensure the quality of the medicine in the production and use processes, thereby ensuring the curative effect of the medicine.

Description

Method for detecting quality of medicine for treating colitis

Technical Field

The invention relates to a method for detecting the quality of a medicine for treating colitis, belonging to the technical field of medicines.

Technical Field

The medicine for treating colitis has the functions of invigorating qi, promoting blood circulation, and relieving diarrhea, and can be used for treating diarrhea caused by qi deficiency and blood stasis, with symptoms of abdominal pain, diarrhea, tenesmus, mucus or purulent bloody stool, dark or black blood, listlessness of limbs, pale or purple tongue, and soft, thready or wiry and unsmooth pulse; chronic ulcerative colitis refers to the above-mentioned syndromes. Prepared by using 650 parts of 550-90 parts of astragalus root, 75-90 parts of pseudo-ginseng, 700 parts of 550-700 parts of Japanese ampelopsis root, 215 parts of 195-containing bletilla striata, 210 parts of frankincense and 210 parts of 195-containing gallnut as raw materials and 3000 parts of 800-containing auxiliary materials (for short, colitis medicine), and has good curative effect on various symptoms of chronic ulcerative colitis.

The colitis medicine prescription is proved by clinical application of traditional Chinese medicine for many years, and the clinical application results show that the medicine has definite curative effect on chronic ulcerative colitis, good long-term effect and no adverse reaction, and overcomes the defects of symptom control, poor long-term effect and the like of chemical medicines to a certain extent.

The colitis medicament of the invention, the prior art does not have a method for detecting the quality of the colitis medicament, and the product quality can not be effectively ensured in production and use, so that it is very important to provide a set of simple and reliable quality standards for ensuring the quality of the colitis medicament and the clinical medication effect.

Disclosure of Invention

The invention aims to overcome the defects of the prior art and provide a method for detecting the quality of a medicament for treating colitis, so that the quality of the product can be effectively ensured during production and use of the medicament, and the curative effect of the medicament is ensured.

The purpose of the invention is realized by the following technical scheme:

a quality detection method of a medicine for treating colitis comprises identifying Notoginseng radix and rhizoma Bletillae by microscopy; identifying radix astragali and Notoginseng radix by thin layer chromatography; identifying Galla chinensis by thin layer chromatography; and measuring the content of the gallnut by adopting a high performance liquid chromatography.

A method for detecting the quality of a medicine for treating colitis is characterized by comprising the following steps:

(1) identifying pseudo-ginseng and bletilla striata by using a microscope: taking a detected product, and observing the characteristics of the rhizoma bletillae powder under a microscope: the superficial vertical wall of the epidermal cell is wavy and curved, rarely, the calcium oxalate needle crystal bundles exist in large round-like mucus fine bubbles or are scattered everywhere, the needle crystal length is 18-88 mu m, the fibers are bundled, the diameter is 11-30 mu m, and the needle crystal bundles have herringbone or elliptical holes, and the diameters of the ladder conduit and the threaded conduit are 10-55 mu m; the characteristics of the pseudo-ginseng powder are as follows: the resin channel fragment contains yellow secretion, and the diameter of the ladder conduit and the threaded conduit is 10-55 μm, thus determining the preparation containing rhizoma bletilla and Notoginseng radix powder.

(2) Identifying radix astragali and Notoginseng radix by thin layer chromatography: weighing 5-10g of a sample, placing the sample in a conical flask with a plug, adding 50ml of trichloromethane, carrying out ultrasonic treatment for 30 minutes, filtering, adding 50ml of methanol into residues, placing the residue on a water bath, heating and refluxing for 1-3 hours, cooling, filtering, evaporating filtrate, dissolving the residues by adding 30ml of water, extracting the residues by using water saturated n-butyl alcohol for 1-5 times, 20-40ml each time, combining n-butyl alcohol solutions, washing the residues by using 10-20ml of 0.25mol/L sodium hydroxide solution, washing the residues by using 10-30ml of water, evaporating the n-butyl alcohol solution, dissolving the residues by adding 2ml of methanol to obtain a sample solution; taking proper amount of astragaloside IV reference substance, adding methanol to obtain 1mg solution per 1ml, and using as astragaloside IV reference substance solution; adding methanol into appropriate amount of ginsenoside Rb1 and ginsenoside Rg1 to obtain mixed solution containing 0.5mg of ginsenoside Rb1 and ginsenoside Rg1 per 1ml as Notoginseng radix reference solution; sucking 5-10 μ L of test solution, 5-10 μ L of astragaloside IV control solution and 5-10 μ L of Notoginseng radix control solution, respectively dropping on the same silica gel G thin layer plate, developing with lower layer solution of chloroform, methanol and water = 10-15: 5-9: 1-3 at 10 deg.C as developing agent, taking out, air drying, spraying with ethanol solution containing 1mol/L of sulfuric acid, and heating at 105 deg.C until the spots are clearly developed. Spots with the same color appear on the chromatogram of the test solution at the positions corresponding to the chromatograms of the reference solution; inspecting under 365nm ultraviolet lamp, and displaying the same fluorescent spot to obtain the preparation containing radix astragali and Notoginseng radix.

(3) Identifying gallnut by thin layer chromatography: weighing 1-5g of a sample, placing in a conical flask with a plug, adding 50ml of methanol, placing on a water bath, heating and refluxing for 10-50 minutes, taking down, placing at 0-5 ℃ for 3 hours, filtering, evaporating filtrate to dryness, adding 30ml of water into residue, heating to dissolve, cooling, filtering, evaporating filtrate to dryness, and adding 5ml of methanol into residue to dissolve to obtain a sample solution; adding methanol into gallic acid control to obtain 1mg solution per 1ml as control solution; sucking sample solution and reference solution 5 μ l each, respectively dropping on the same silica gel GF254 thin layer plate, developing with chloroform, ethyl formate and formic acid = 3-7: 1-3 as developing agent, taking out, air drying, inspecting under 254nm ultraviolet lamp, and displaying spots of the same color in the sample chromatogram at the position corresponding to the reference chromatogram to determine the preparation containing Galla chinensis.

(4) Measuring gallnut by high performance liquid chromatography: chromatographic conditions and system applicability tests, and octadecylsilane chemically bonded silica is used as a filler; taking a 5-15mmol/L phosphoric acid solution containing 2-5mol/L methanol as a mobile phase; the detection wavelength is 273 nm; the number of theoretical plates is not less than 5000 according to the peak of gallic acid; precisely weighing gallic acid reference substance, and preparing into 12-120 μ g/ml solution with 12mol/L methanol to obtain reference substance solution; weighing 1-5g of a sample, precisely weighing, placing the sample in a conical flask with a plug, precisely adding 50ml of 2-6mol/L hydrochloric acid, sealing the plug, weighing, heating at 90-120 ℃ for 2-5 hours, cooling, weighing again, supplementing the lost weight with 2-6mol/L hydrochloric acid, shaking up, filtering, precisely weighing 1ml of subsequent filtrate, placing the subsequent filtrate in a 50ml measuring flask, adding 12mol/L methanol to dilute to scale, and shaking up to obtain a sample solution; precisely sucking 5-20 μ l of reference solution and sample solution respectively, injecting into liquid chromatograph, and measuring; the content of Galla chinensis in the tested medicine is calculated by gallic acid according to external standard method.

(5) Determining radix astragali by high performance liquid chromatography: chromatographic conditions and system applicability tests, and octadecylsilane chemically bonded silica is used as a filler; 13mol/L acetonitrile solution is used as a mobile phase; an evaporative light scattering detector; the number of theoretical plates is not less than 5000 calculated according to astragaloside IV peak; accurately weighing appropriate amount of astragaloside IV reference substance, and adding methanol to obtain 0.5mg/1ml reference substance solution; weighing 5-20g of sample, precisely weighing, placing in a conical flask with a plug, adding 100ml of chloroform, performing ultrasonic treatment for 30 minutes, filtering, adding 100ml of methanol into residue, heating to slightly boil for 1-3 hours, cooling, filtering, evaporating filtrate, adding 30ml of water into residue, shaking and extracting with water saturated n-butanol for 2 times, 30ml each time, mixing n-butanol solution, washing with ammonia test solution for 2 times, 30ml each time, discarding ammonia solution, evaporating n-butanol solution to dryness, adding 5ml of water into residue to dissolve, passing through a D101 type macroporous resin column with an inner diameter of 1.5cm and a length of 10cm, eluting with 50ml of water, discarding water solution, eluting with 5-20ml of chloroform, 5-20ml of ethyl acetate and 20-40ml of 7mol/L ethanol, discarding eluent, eluting with 40-100ml of methanol, collecting eluent, evaporating at 30-60 ℃, dissolving with methanol and fixing volume to a 5ml measuring bottle, obtaining a test solution; precisely sucking 5-20 μ l of reference solution and sample solution respectively, injecting into liquid chromatograph, and measuring; the content of Astragalus membranaceus in the tested medicine is calculated by Astragaloside IV according to an external standard method.

The invention has been completed, and the beneficial effects of the invention are that the method adopts various methods to carry out comprehensive quality control on the colitis medicine, can ensure the quality of the medicine, strictly controls the used medicinal materials, and ensures that the content of the effective components meets the requirements.

In order to further illustrate the quality controllability of the invention for colitis drugs, the beneficial effects of the invention are further illustrated by the quality standard research process of the invention. The quality criteria study is intended to further illustrate the role of the invention and not to limit the invention.

Preparation of determination sample

1 preparing colitis suppository

Taking 600g of astragalus, 80g of pseudo-ginseng, 200g of frankincense, 200g of gallnut, 600g of ampelopsis japonica and 200g of bletilla striata. Wherein the notoginseng, the frankincense, the gallnut and the common bletilla pseudobulb are crushed into fine powder; decocting the astragalus and the ampelopsis japonica in water twice, each time for 1.5 hours, mixing decoctions, filtering, concentrating under reduced pressure to obtain clear paste with the relative density of 1.15-1.20 (80 ℃), adding the fine powder, mixing uniformly, drying under reduced pressure, and crushing into fine powder. And heating and melting a proper amount of mixed fatty glyceride at 50-60 ℃, adding the fine powder, uniformly mixing, pouring into a mold, cooling, and preparing into 1000 granules.

2 preparing negative suppository without bletilla striata and notoginseng

Taking 600g of astragalus, 200g of frankincense, 200g of gallnut and 600g of ampelopsis japonica, and preparing the negative suppository without containing bletilla striata and pseudo-ginseng according to the method for preparing the colitis suppository.

3 preparing negative suppository without astragalus root and notoginseng

200g of frankincense, 200g of gallnut, 600g of ampelopsis japonica and 200g of bletilla striata are taken to prepare the negative suppository without containing astragalus and pseudo-ginseng according to the method for preparing the colitis suppository.

4 preparing negative suppository without Galla chinensis

Taking 600g of astragalus, 80g of pseudo-ginseng, 200g of frankincense, 600g of ampelopsis japonica and 200g of bletilla striata, and preparing the negative suppository without gallnut according to the method for preparing the colitis suppository.

Second, quality standard research process

1 differentiation of rhizoma Bletillae and Notoginseng radix

The samples were mounted on a microscope with chloral hydrate and observed under a microscope to detect microscopic features: the sag of the peripheral wall on the surface of the epidermal cell is wavy and is rare; the calcium oxalate needle crystal bundles exist in large round-like mucus fine bubbles or are scattered everywhere, and the needle crystal growth is in the range of 18-88 mu m; the fibers are bundled, have the diameter within the range of 11-30 mu m and are provided with herringbone or elliptical pores (bletilla striata). The resin tract fragments contained yellow secretions (notoginseng).

And taking rhizoma bletillae and pseudo-ginseng negative preparations for inspection by the same method, wherein the microscopic characteristics are not detected.

The result shows that the method has strong specificity and can effectively identify the bletilla striata and the pseudo-ginseng powder in the sample.

2 differentiation of Astragalus membranaceus and Panax notoginseng

Taking 4 granules of the product, cutting into pieces, placing in a conical flask with a plug, adding 50ml of trichloromethane, carrying out ultrasonic treatment (power 200W and frequency 40kHz) for 30 minutes, filtering, adding 50ml of methanol into residues, placing on a water bath, heating and refluxing for 2 hours, cooling, filtering, evaporating filtrate, dissolving residues by heating with 30ml of water, extracting with water-saturated n-butanol for 3 times, 30ml each time, combining n-butanol solutions, washing with 15ml of 0.25mol/L sodium hydroxide solution once, washing with 20ml of water again, evaporating n-butanol solutions to dryness, and dissolving residues by adding 2ml of methanol to obtain a sample solution.

And adding methanol to a proper amount of astragaloside IV reference substance to obtain a solution containing 1mg of astragaloside IV reference substance per 1 ml. Collecting ginsenoside Rb₁Reference substance, ginsenoside Rg₁Adding appropriate amount of reference substance, and adding methanol to obtain mixed solution containing 0.5mg of reference substance per 1ml, and making into Notoginseng radix reference substance solution.

Preparing the negative solution by taking the astragalus root and the pseudo-ginseng negative preparation and the preparation method of the test solution.

Performing thin layer chromatography test, sucking 10 μ L of sample solution, 5 μ L of astragaloside IV control solution and Notoginseng radix control solution, respectively dropping on the same silica gel G thin layer plate, spreading with chloroform-methanol-water (13: 7: 2) lower layer solution at 10 deg.C below as developing agent, taking out, air drying, spraying with 1mol/L sulfuric acid-containing ethanol solution, and heating at 105 deg.C until the spots are clearly developed.

The result shows that spots with the same color appear on the chromatogram of the test solution at the positions corresponding to the chromatograms of the reference solution; when the sample is inspected under an ultraviolet lamp (365nm), the same fluorescent spots are displayed, and the negative preparation is displayed without spots at the corresponding position. The method shows that the method has strong specificity for identifying the astragalus and the pseudo-ginseng and has no interference to other raw and auxiliary materials.

3 Chinese gall identification

Chromatographic conditions are as follows: with silica gel GF₂₅₄Is a thin layer plate fixed phase; the sample counting amount is 5 mu l; the method comprises the following steps of (1) mixing trichloromethane: ethyl formate: formic acid (5: 1) is used as developing agent.

Preparing a test solution: taking 2 samples, cutting into pieces, placing into a conical flask with a plug, adding 50ml of methanol, placing on a water bath, heating and refluxing for 30 minutes, taking down, placing at 0-5 ℃ for 3 hours, filtering, evaporating filtrate to dryness, adding 30ml of water into residues, heating to dissolve, cooling, filtering, evaporating filtrate to dryness, and adding 5ml of methanol into residues to dissolve to obtain the product.

Preparation of gallic acid reference solution: adding methanol into gallic acid control to obtain solution containing 1mg of gallic acid per 1 ml.

Preparation of negative solution: preparing a solution by taking the negative preparation without gallnut and a preparation method of a test solution.

Performing thin layer chromatography, sucking 5 μ l of the above solutions, respectively dropping on the same silica gel GF254 thin layer plate, developing, taking out, air drying, and inspecting under ultraviolet lamp (365 nm).

The result shows that spots of the same color appear in the chromatogram of the test solution at positions corresponding to those in the chromatogram of the control solution. The negative solution was spot-free at the corresponding position. The method is proved to have strong specificity for identifying the gallnut, and other raw and auxiliary materials have no interference.

4 content determination

The method for measuring the content of the gallic acid in the gallnut adopts high performance liquid chromatography, and a methodology research test is carried out on the content measurement of the gallic acid in the colitis suppository.

4.1 specificity test

Chromatographic conditions and system applicability tests, and octadecylsilane chemically bonded silica is used as a filler; taking a phosphoric acid solution containing 3.8mol/L methanol and 8.7mmol/L as a mobile phase; the detection wavelength is 273 nm; the number of theoretical plates is not less than 5000 according to gallic acid peak.

Precisely weighing gallic acid reference substance, and preparing into solution containing 40 μ g per 1ml with 12mol/L methanol to obtain reference substance solution.

Taking 10 grains of a sample, precisely weighing, shearing, weighing 2g, precisely weighing, placing in a conical flask with a plug, precisely adding 50ml of 4mol/L hydrochloric acid, sealing the plug, weighing, heating at 100 ℃ for 3.5 hours, cooling, weighing again, supplementing the lost weight with 4mol/L hydrochloric acid, shaking up, filtering, precisely weighing 1ml of a subsequent filtrate, placing in a 50ml measuring flask, adding 12mol/L methanol to dilute to scale, and shaking up to obtain a sample solution.

Preparation of negative solution A Galla chinensis negative preparation is prepared by the same method.

The measurement method precisely pipetted 10. mu.l each of the above solutions, and the solution was injected into a liquid chromatograph for measurement. The content of Galla chinensis in the tested medicine is calculated by gallic acid according to external standard method.

TABLE 1 System suitability test results

Item	Number of theoretical plates	Repeatability (n = 7)	Degree of separation	Tailing factor
					Gallic acid reference substance	8300	0.10%	——	1.050
Test article	8500	——	2.445	1.050

The result shows that the repeatability of the chromatographic peak, the number of theoretical plates, the separation degree and the tailing factor of the gallic acid determined by the method all accord with the corresponding regulations in the high performance liquid chromatography, and the method has good applicability.

The specificity test result shows that the test solution has a corresponding chromatographic peak within the retention time of the chromatographic peak of the reference solution, and the negative preparation has no chromatographic peak within the corresponding retention time. The result shows that the negative property has no interference to the determination of gallic acid by the method, and the specificity is strong.

4.2 Linear test

A control solution containing gallic acid 0.1mg per 1ml was used as a control stock solution. Precisely sucking 1ml, 3 ml, 5ml and 7ml respectively, putting into a 10ml measuring flask, and adding 50% methanol to dilute to scale to obtain linear solutions with different concentrations.

Respectively and precisely sucking 10 μ l of the linear solution, injecting into a liquid chromatograph, and measuring to obtain the final product.

TABLE 2 Gallic acid Linear test results

Taking the concentration of the gallic acid reference substance as a horizontal coordinate and the peak area as a vertical coordinate, performing linear regression to obtain a regression equation: y =33038x-9506, r = 1.0000. The result shows that the method has a good linear relation between the sample injection amount of gallic acid in the range of 0.12-1.2 mu g and the peak area.

4.3 accuracy test

Precisely weighing 6 parts of the product about 1g (containing 52.9992mg/g of gallic acid), placing in a conical flask with a plug, precisely adding 50mg of gallic acid reference substance respectively, heating in water bath at 60 ℃ to melt, uniformly mixing, precisely adding 50ml of 4mol/L hydrochloric acid, sealing the plug, weighing, placing on the water bath for heating for 3.5 hours, cooling, weighing again, supplementing the lost weight with 4mol/L hydrochloric acid, shaking up, filtering, precisely weighing 1ml of subsequent filtrate, placing in a 50ml measuring flask, adding 50% methanol to dilute to scale, shaking up to obtain the accurate solution.

Precisely sucking 10 μ l of each of the reference solution and the accuracy solution, injecting into a liquid chromatograph, and measuring.

TABLE 3 accuracy test results

Injecting: the content of the gallic acid control product in 110831-201204 batch is 89.9%.

The result shows that the method has good accuracy in determination of gallic acid.

4.4 durability test

4.4.1 solution stability test

And (3) taking the test solution, standing at room temperature, precisely sucking 10 mu l of the test solution at different times, injecting the test solution into a liquid chromatograph, and recording the chromatogram. The results are as follows.

TABLE 4 test results of solution stability of test article

Time of day	0h	4h	5h	8h	9h	12h	RSD
								Peak area of gallic acid	1393145	1397855	1396387	1393334	1387635	1386668	0.33%

The result shows that the area of the peak of the gallic acid is not obviously changed when the test solution is placed at room temperature for 12 hours, which indicates that the test solution is still stable when the method is used for determination within 12 hours of being placed at room temperature, and the determination requirement of the method is met.

4.4.2 mobile phase ratio durability test

Durability tests were performed with varying ratios of mobile phases in chromatographic conditions. The results are as follows.

TABLE 5 mobile phase ratio for durability test results

The test result shows that the durability of the gallic acid for the change of the mobile phase proportion is good.

4.4.3 chromatographic column durability test

And (4) adopting chromatographic columns of different brands and batch numbers to measure the content of the gallic acid. The results are as follows.

TABLE 6 mobile phase ratio for durability test results

The results show that the durability of chromatographic columns for determining gallic acid in different brands is good.

4.5 precision test

4.5.1 repeatability test

6 parts of the test solution were prepared as described above, and the results were as follows.

TABLE 7 results of the repeatability tests

The result shows that the method has good repeatability in determination of the content of the gallic acid.

4.5.2 intermediate precision test

Samples were taken and subjected to intermediate precision tests, and the results are shown in each test item.

4.5.3 precision tests on different dates

Samples were taken for 3 consecutive days to determine gallic acid content. The results are as follows:

TABLE 8 precision test results on different dates

The result shows that the method has good precision in measuring different dates.

4.5.4 testing precision of different persons

Samples were taken and tested for gallic acid content on the same HPLC chromatograph by experimenter A, B. The results were obtained as follows:

TABLE 9 results of precision tests of different persons

The result shows that the method has good precision for measuring different people.

4.5.5 precision test of different instruments

The gallic acid content of the samples was measured by the same experimenter on A, B two HPLC chromatographs. The results were obtained as follows:

TABLE 10 results of precision tests of different instruments

The result shows that the method has good precision for measuring different instruments.

Radix astragali

The chromatographic condition takes octadecylsilane chemically bonded silica as a filler; 13mol/L acetonitrile solution is used as a mobile phase; an evaporative light scattering detector. The number of theoretical plates is not less than 5000 calculated according to astragaloside IV peak.

Preparation of reference substance solution A proper amount of astragaloside IV reference substance is precisely weighed, and methanol is added to prepare 0.5mg/1ml solution.

Preparing test solution by weighing 10 granules, precisely weighing, cutting, placing into conical flask with plug, adding chloroform 100ml, ultrasonic treating for 30 min, filtering, adding methanol 100ml into residue, placing in 80 deg.C water bath, maintaining slight boiling for 2 hr, cooling, filtering, and evaporating filtrate to dryness. Adding water 30ml into residue, shaking with water saturated n-butanol to extract for 2 times, 30ml each time, mixing n-butanol solution, washing with ammonia solution for 2 times, 30ml each time, discarding ammonia solution, evaporating n-butanol solution to dryness, dissolving residue with 5ml water, passing through D101 type macroporous resin column (inner diameter 1.5cm, length 10 cm), eluting with 50ml water, discarding water solution, eluting with chloroform 10ml, ethyl acetate 10ml, 7mol/L ethanol 30ml, discarding eluate, eluting with 50ml methanol, collecting eluate, evaporating to dryness at 40 deg.C, dissolving with methanol, and fixing volume to 5ml measuring flask.

The result shows that the test solution has a corresponding chromatographic peak within the retention time of the chromatographic peak of the reference solution, and the method can be used for detecting the astragaloside contained in the product.

Detailed Description

Example 1: the colitis suppository is determined according to the quality standard detection method of the invention

1) Identifying pseudo-ginseng and bletilla striata by using a microscope: taking a detected product, and observing the characteristics of the rhizoma bletillae powder under a microscope: the superficial vertical wall of the epidermal cell is wavy and curved, rarely, the calcium oxalate needle crystal bundles exist in large round-like mucus fine bubbles or are scattered everywhere, the needle crystal length is 18 mu m, the fibers are bundled, the diameter is 11 mu m, and the needle crystal bundles have herringbone or oval holes, and the diameters of the ladder conduit and the threaded conduit are 10 mu m; the characteristics of the pseudo-ginseng powder are as follows: the resin channel fragment contains yellow secretion, and the diameter of the ladder conduit and the threaded conduit is 10 μm, namely the preparation containing rhizoma bletillae and pseudo-ginseng powder is determined.

2) Identifying radix astragali and Notoginseng radix by thin layer chromatography: weighing 5g of a sample, placing the sample in a conical flask with a plug, adding 50ml of trichloromethane, carrying out ultrasonic treatment for 30 minutes, filtering, adding 50ml of methanol into residues, placing the residues on a water bath, heating and refluxing for 1 hour, cooling, filtering, evaporating the filtrate to dryness, adding 30ml of water into the residues, heating and dissolving the residues, extracting with 20ml of water-saturated n-butyl alcohol, washing with 10ml of 0.25mol of sodium hydroxide solution, washing with 10ml of water, evaporating the n-butyl alcohol solution to dryness, and adding 2ml of methanol into the residues to dissolve the residues to obtain a sample solution; taking proper amount of astragaloside IV reference substance, adding methanol to obtain 1mg solution per 1ml, and using as astragaloside IV reference substance solution; adding methanol into appropriate amount of ginsenoside Rb1 and ginsenoside Rg1 to obtain mixed solution containing 0.5mg of ginsenoside Rb1 and ginsenoside Rg1 per 1ml as Notoginseng radix reference solution; sucking 5 mul of the test solution, 5 mul of each of the astragaloside IV reference solution and the pseudo-ginseng reference solution, respectively dropping on the same silica gel G thin layer plate, adding chloroform: methanol: the lower layer solution which is placed at the temperature of below 10 ℃ with the water = 10: 5: 1 is used as a developing agent, the developing agent is developed, taken out, dried, sprayed with an ethanol solution containing 1mol/L of sulfuric acid, and heated at the temperature of 105 ℃ until the spots are clearly developed. Spots with the same color appear on the chromatogram of the test solution at the positions corresponding to the chromatograms of the reference solution; inspecting under 365nm ultraviolet lamp, and displaying the same fluorescent spot to obtain the preparation containing radix astragali and Notoginseng radix.

3) Identifying gallnut by thin layer chromatography: weighing 1g of sample, placing into a conical flask with a plug, adding 50ml of methanol, placing on a water bath, heating and refluxing for 10 minutes, taking down, placing at 0 ℃ for 3 hours, filtering, evaporating filtrate to dryness, adding 30ml of water into residue, heating to dissolve, cooling, filtering, evaporating filtrate to dryness, and adding 5ml of methanol into residue to dissolve to obtain a sample solution; adding methanol into gallic acid control to obtain 1mg solution per 1ml as control solution; sucking 5 mul of each of the test solution and the reference solution, respectively dropping on the same silica gel GF254 thin layer plate, adding chloroform: ethyl formate: developing with formic acid = 3: 1 as developing agent, taking out, air drying, and inspecting under 254nm ultraviolet lamp, wherein spots with the same color appear on the chromatogram of the test solution at the position corresponding to the chromatogram of the control solution, thus determining the preparation containing Galla chinensis.

4) Gallnut is determined by high performance liquid chromatography: chromatographic conditions and system applicability tests, and octadecylsilane chemically bonded silica is used as a filler; taking a 5mmol/L phosphoric acid solution containing 2mol/L methanol as a mobile phase; the detection wavelength is 273 nm; the number of theoretical plates is not less than 5000 according to gallic acid peak.

Precisely weighing gallic acid reference substance, and preparing into 12 μ g/ml solution with 12mol/L methanol to obtain reference substance solution;

weighing 1g of a sample, precisely weighing, placing in a conical flask with a plug, precisely adding 50ml of 2mol/L hydrochloric acid, sealing the plug, weighing, heating at 90 ℃ for 5 hours, cooling, weighing again, supplementing the lost weight with 2mol/L hydrochloric acid, shaking up, filtering, precisely weighing 1ml of a subsequent filtrate, placing in a 50ml measuring flask, adding 12mol/L methanol for diluting to a scale, and shaking up to obtain a sample solution.

Precisely sucking 5 μ l of each of the reference solution and the sample solution, injecting into a liquid chromatograph, and measuring; the content of Galla chinensis in the tested medicine is calculated by gallic acid according to external standard method.

5) Determining radix astragali by high performance liquid chromatography: chromatographic conditions and system applicability tests, and octadecylsilane chemically bonded silica is used as a filler; 13mol/L acetonitrile solution is used as a mobile phase; an evaporative light scattering detector; the number of theoretical plates is not less than 5000 calculated according to astragaloside IV peak.

Accurately weighing appropriate amount of astragaloside IV reference substance, and adding methanol to obtain 0.5mg/1ml reference substance solution.

Taking 5g of a sample, precisely weighing, placing in a conical flask with a plug, adding 100ml of trichloromethane, performing ultrasonic treatment for 30 minutes, filtering, adding 100ml of methanol into residue, heating to slightly boil for 1 hour, cooling, filtering, evaporating filtrate to dryness, adding 30ml of water into residue, shaking and extracting with water-saturated n-butyl alcohol for 2 times, 30ml each time, combining n-butyl alcohol solutions, washing with ammonia test solution for 2 times, 30ml each time, discarding ammonia solution, evaporating n-butyl alcohol solution to dryness, adding 5ml of water into residue to dissolve, passing through a D101 type macroporous resin column with the inner diameter of 1.5cm and the length of 10cm, eluting with 50ml of water, discarding water solution, eluting with 5ml of trichloromethane, 5ml of ethyl acetate and 20ml of 7mol/L ethanol, discarding eluent, then eluting with 40ml of methanol, collecting eluent, evaporating to dryness at 30 ℃, dissolving with methanol, and fixing the volume to a 5ml volumetric flask to obtain a sample solution.

Precisely sucking 5 μ l and 10 μ l of reference solution and 5 μ l of test solution respectively, injecting into liquid chromatograph, and measuring; the content of Astragalus membranaceus in the tested medicine is calculated by Astragaloside IV according to an external standard method.

6) Test results

The test results are shown in Table 11:

TABLE 11 colitis suppository test results

The results show that the methods meet the regulations, and the method can effectively control the quality of the colitis suppository.

Example 2: the enteric granule for treating colitis is determined according to the quality standard detection method of the invention.

1) Identifying pseudo-ginseng and bletilla striata by using a microscope: taking a detected product, and observing the characteristics of the rhizoma bletillae powder under a microscope: the superficial vertical wall of the epidermal cell is wavy and curved, rarely, the calcium oxalate needle crystal bundles exist in large round-like mucus fine bubbles or are scattered everywhere, the needle crystals have the length of 70 mu m, the fibers are bundled, the diameter is 30 mu m, and the needle crystals have herringbone or oval holes, and the diameters of the ladder conduit and the threaded conduit are 50 mu m; the characteristics of the pseudo-ginseng powder are as follows: the resin channel fragment contains yellow secretion, and the diameter of the ladder conduit and the threaded conduit is 60 μm, namely the preparation containing rhizoma bletillae and radix notoginseng powder is determined.

2) Identifying radix astragali and Notoginseng radix by thin layer chromatography: weighing 10g of a sample, placing the sample in a conical flask with a plug, adding 50ml of trichloromethane, carrying out ultrasonic treatment for 30 minutes, filtering, adding 50ml of methanol into residues, placing the residues on a water bath, heating and refluxing for 3 hours, cooling, filtering, evaporating the filtrate to dryness, adding 30ml of water into the residues, heating and dissolving the residues, extracting with water-saturated n-butanol for 5 times, 40ml each time, combining n-butanol solutions, washing with 20ml of 0.25mol sodium hydroxide solution once, washing with 30ml of water again, evaporating the n-butanol solution to dryness, and adding 2ml of methanol into the residues to dissolve the residues to obtain a sample solution; taking proper amount of astragaloside IV reference substance, adding methanol to obtain 1mg solution per 1ml, and using as astragaloside IV reference substance solution; adding methanol into appropriate amount of ginsenoside Rb1 and ginsenoside Rg1 to obtain mixed solution containing 0.5mg of ginsenoside Rb1 and ginsenoside Rg1 per 1ml as Notoginseng radix reference solution; sucking 10 mul of the test solution, 10 mul of each of the astragaloside IV reference solution and the pseudo-ginseng reference solution, respectively dropping the solutions on the same silica gel G thin layer plate, adding chloroform: methanol: the lower layer solution which is placed at the temperature of below 10 ℃ with the water = 15: 9: 3 is used as a developing agent, the developing agent is developed, taken out, dried, sprayed with an ethanol solution containing 1mol/L of sulfuric acid, and heated at the temperature of 105 ℃ until the spots are clearly developed. Spots with the same color appear on the chromatogram of the test solution at the positions corresponding to the chromatograms of the reference solution; inspecting under 365nm ultraviolet lamp, and displaying the same fluorescent spot to obtain the preparation containing radix astragali and Notoginseng radix.

3) Identifying gallnut by thin layer chromatography: weighing 5g of sample, placing in a conical flask with a plug, adding 50ml of methanol, placing on a water bath, heating and refluxing for 50 minutes, taking down, placing at 5 ℃ for 3 hours, filtering, evaporating filtrate to dryness, adding 30ml of water into residue, heating to dissolve, cooling, filtering, evaporating filtrate to dryness, and adding 5ml of methanol into residue to dissolve to obtain a sample solution; adding methanol into gallic acid control to obtain 1mg solution per 1ml as control solution; sucking 5 mul of each of the test solution and the reference solution, respectively dropping on the same silica gel GF254 thin layer plate, adding chloroform: ethyl formate: formic acid = 7: 3 as developing agent, developing, taking out, air drying, placing under 254nm ultraviolet lamp for inspection, and in the chromatogram of the test solution, spots with the same color appear on the corresponding positions of the chromatogram of the reference solution, thus determining the preparation containing Galla chinensis.

4) Gallnut is determined by high performance liquid chromatography: chromatographic conditions and system applicability tests, and octadecylsilane chemically bonded silica is used as a filler; taking a 15mmol/L phosphoric acid solution containing 5mol/L methanol as a mobile phase; the detection wavelength is 273 nm; the number of theoretical plates is not less than 5000 according to the peak of gallic acid; precisely weighing gallic acid reference substance, and preparing into 120 μ g/ml solution with 12mol/L methanol to obtain reference substance solution; weighing 5g of a sample, precisely weighing, placing the sample in a conical flask with a plug, precisely adding 50ml of 6mol/L hydrochloric acid, sealing the plug, weighing, heating at 120 ℃ for 2 hours, cooling, weighing again, supplementing the lost weight with 6mol/L hydrochloric acid, shaking up, filtering, precisely weighing 1ml of a subsequent filtrate, placing the subsequent filtrate in a 50ml measuring flask, adding 12mol/L methanol for diluting to a scale, and shaking up to obtain a sample solution; precisely sucking 20 μ l of each of the reference solution and the sample solution, injecting into a liquid chromatograph, and measuring; the content of Galla chinensis in the tested medicine is calculated by gallic acid according to external standard method.

5) Determining radix astragali by high performance liquid chromatography: chromatographic conditions and system applicability tests, and octadecylsilane chemically bonded silica is used as a filler; 13mol/L acetonitrile solution is used as a mobile phase; an evaporative light scattering detector; the number of theoretical plates is not less than 5000 calculated according to astragaloside IV peak; accurately weighing appropriate amount of astragaloside IV reference substance, and adding methanol to obtain 0.5mg/1ml reference substance solution; taking 20g of a sample, precisely weighing, placing in a conical flask with a plug, adding 100ml of trichloromethane, performing ultrasonic treatment for 30 minutes, filtering, adding 100ml of methanol into residue, heating to slightly boil for 3 hours, cooling, filtering, evaporating filtrate to dryness, adding 30ml of water into residue, shaking and extracting with water-saturated n-butyl alcohol for 2 times, 30ml each time, combining n-butyl alcohol solutions, washing with an ammonia test solution for 2 times, 30ml each time, discarding the ammonia solution, evaporating the n-butyl alcohol solution to dryness, adding 5ml of water into the residue to dissolve, passing through a D101 type macroporous resin column with the inner diameter of 1.5cm and the length of 10cm, eluting with 50ml of water, discarding a water solution, eluting with 20ml of trichloromethane, 20ml of ethyl acetate and 40ml of 7mol/L ethanol, discarding the eluent, then eluting with 100ml of methanol, collecting the eluent, evaporating to dryness at 60 ℃, dissolving with methanol, and fixing the volume to a 5ml volumetric flask to obtain a sample; precisely sucking 10 μ l and 20 μ l of reference solution and 20 μ l of test solution respectively, injecting into liquid chromatograph, and measuring; the content of Astragalus membranaceus in the tested medicine is calculated by Astragaloside IV according to an external standard method.

6) Test results

The test results are shown in Table 12:

TABLE 12 test results of colitis enteric granules

The results all meet the requirements of the quality standard detection method, and show that the method can effectively control the quality of the colitis enteric-coated granules.

Example 3: the enteric capsule for treating colitis is determined according to the quality standard detection method of the invention.

The results all meet the requirements of the quality standard detection method, and show that the method can effectively control the quality of the colitis enteric-coated capsules.

1) Identifying pseudo-ginseng and bletilla striata by using a microscope: taking a detected product, and observing the characteristics of the rhizoma bletillae powder under a microscope: the superficial vertical wall of the epidermal cell is wavy and curved, rarely, the calcium oxalate needle crystal bundles exist in large round-like mucus fine bubbles or are scattered everywhere, the needle crystals are 90 mu m long, the fibers are bundled, the diameter is 20 mu m, and the needle crystals have herringbone or oval holes, and the diameters of the ladder conduit and the threaded conduit are 43 mu m; the characteristics of the pseudo-ginseng powder are as follows: the resin channel fragment contains yellow secretion, and the diameter of the ladder conduit and the threaded conduit is 35 μm, namely the preparation containing rhizoma bletillae and radix notoginseng powder is determined.

2) Identifying radix astragali and Notoginseng radix by thin layer chromatography: weighing 6g of a sample, placing the sample in a conical flask with a plug, adding 50ml of trichloromethane, carrying out ultrasonic treatment for 30 minutes, filtering, adding 50ml of methanol into residues, placing the residues on a water bath, heating and refluxing for 2 hours, cooling, filtering, evaporating the filtrate to dryness, adding 30ml of water into the residues, heating and dissolving the residues, extracting with water-saturated n-butanol for 3 times, 30ml each time, combining n-butanol solutions, washing with 15ml of 0.25mol sodium hydroxide solution once, washing with 20ml of water again, evaporating the n-butanol solution to dryness, and adding 2ml of methanol into the residues to dissolve the residues to obtain a sample solution; taking proper amount of astragaloside IV reference substance, adding methanol to obtain 1mg solution per 1ml, and using as astragaloside IV reference substance solution; adding methanol into appropriate amount of ginsenoside Rb1 and ginsenoside Rg1 to obtain mixed solution containing 0.5mg of ginsenoside Rb1 and ginsenoside Rg1 per 1ml as Notoginseng radix reference solution; sucking 7 mul of the test solution, 7 mul of each of the astragaloside IV reference solution and the pseudo-ginseng reference solution, respectively dropping the solution on the same silica gel G thin layer plate, adding chloroform: methanol: the lower layer solution which is placed at the temperature of below 10 ℃ with the water = 12: 7: 2 is used as a developing agent, the developing agent is developed, taken out, dried, sprayed with an ethanol solution containing 1mol/L of sulfuric acid, and heated at the temperature of 105 ℃ until the spots are clearly developed. Spots with the same color appear on the chromatogram of the test solution at the positions corresponding to the chromatograms of the reference solution; inspecting under 365nm ultraviolet lamp, and displaying the same fluorescent spot to obtain the preparation containing radix astragali and Notoginseng radix.

3) Identifying gallnut by thin layer chromatography: weighing 2g of a sample, placing the sample in a conical flask with a plug, adding 50ml of methanol, placing the sample on a water bath, heating and refluxing for 30 minutes, taking down the sample, placing the sample at 2 ℃ for 3 hours, filtering, evaporating filtrate to dryness, adding 30ml of water into residue, heating to dissolve the residue, cooling, filtering, evaporating filtrate to dryness, and adding 5ml of methanol into the residue to dissolve the residue to obtain a sample solution; adding methanol into gallic acid control to obtain 1mg solution per 1ml as control solution; sucking 5 mul of each of the test solution and the reference solution, respectively dropping on the same silica gel GF254 thin layer plate, adding chloroform: ethyl formate: formic acid = 5: 2 as developing agent, developing, taking out, air drying, placing under 254nm ultraviolet lamp for inspection, and in the chromatogram of the test solution, spots with the same color appear on the corresponding positions of the chromatogram of the reference solution, thus determining the preparation containing Galla chinensis.

4) Gallnut is determined by high performance liquid chromatography: chromatographic conditions and system applicability tests, and octadecylsilane chemically bonded silica is used as a filler; taking a 10mmol/L phosphoric acid solution containing 3mol/L methanol as a mobile phase; the detection wavelength is 273 nm; the number of theoretical plates is not less than 5000 according to gallic acid peak.

Precisely weighing gallic acid reference substance, and preparing into 50 μ g/ml solution with 12mol/L methanol to obtain reference substance solution.

Weighing 3g of a sample, precisely weighing, placing in a conical flask with a plug, precisely adding 50ml of 4mol/L hydrochloric acid, sealing the plug, weighing, heating at 100 ℃ for 3 hours, cooling, weighing again, supplementing the lost weight with 4mol/L hydrochloric acid, shaking up, filtering, precisely weighing 1ml of a subsequent filtrate, placing in a 50ml measuring flask, adding 12mol/L methanol for diluting to a scale, and shaking up to obtain a sample solution.

Precisely sucking 10 μ l of each of the reference solution and the sample solution, injecting into a liquid chromatograph, and measuring; the content of Galla chinensis in the tested medicine is calculated by gallic acid according to external standard method.

Taking 10g of a sample, precisely weighing, placing in a conical flask with a plug, adding 100ml of trichloromethane, performing ultrasonic treatment for 30 minutes, filtering, adding 100ml of methanol into residue, heating to slightly boil for 2 hours, cooling, filtering, evaporating filtrate to dryness, adding 30ml of water into residue, shaking and extracting with water-saturated n-butyl alcohol for 2 times, 30ml each time, combining n-butyl alcohol solutions, washing with ammonia test solution for 2 times, 30ml each time, discarding ammonia solution, evaporating n-butyl alcohol solution to dryness, adding 5ml of water into residue to dissolve, passing through a D101 type macroporous resin column with the inner diameter of 1.5cm and the length of 10cm, eluting with 50ml of water, discarding water solution, eluting with 10ml of trichloromethane, 10ml of ethyl acetate and 30ml of 7mol/L ethanol, discarding eluent, then eluting with 80ml of methanol, collecting eluent, evaporating to dryness at 40 ℃, dissolving with methanol, and fixing the volume to a 5ml volumetric flask, thus obtaining a sample solution.

Precisely sucking 5 μ l and 20 μ l of reference solution and 5 μ l of test solution respectively, injecting into liquid chromatograph, and measuring; the content of Astragalus membranaceus in the tested medicine is calculated by Astragaloside IV according to an external standard method.

6) Test results

The test results are shown in Table 13:

TABLE 13 colitis enteric Capsule test results

The results all meet the requirements of the quality standard detection method, and show that the method can effectively control the quality of the colitis enteric-coated capsule.

Example 4: the enteric-coated tablets for treating colitis are determined according to the quality standard detection method of the invention.

1) Identifying pseudo-ginseng and bletilla striata by using a microscope: taking a detected product, and observing the characteristics of the rhizoma bletillae powder under a microscope: the superficial vertical wall of the epidermal cell is wavy and curved, and calcium oxalate needle crystal bundles are rarely present in large round mucus fine bubbles or scattered everywhere, the needle crystals have the length of 48 mu m, the fibers are bundled, the diameter is 26 mu m, and the needle crystals have herringbone or oval holes, and the diameters of the ladder conduit and the threaded conduit are 41 mu m; the characteristics of the pseudo-ginseng powder are as follows: the resin channel fragment contains yellow secretion, and the diameter of the ladder conduit and the threaded conduit is 39 μm, namely the preparation containing rhizoma bletillae and radix notoginseng powder is determined.

2) Identifying radix astragali and Notoginseng radix by thin layer chromatography: weighing 8g of a sample, placing the sample in a conical flask with a plug, adding 50ml of trichloromethane, carrying out ultrasonic treatment for 30 minutes, filtering, adding 50ml of methanol into residues, placing the residues on a water bath, heating and refluxing for 3 hours, cooling, filtering, evaporating the filtrate to dryness, adding 30ml of water into the residues, heating and dissolving, extracting with water-saturated n-butanol for 4 times, 40ml each time, combining n-butanol solutions, washing with 20ml of 0.25mol sodium hydroxide solution once, washing with 20ml of water again, evaporating the n-butanol solution to dryness, and adding 2ml of methanol into the residues to dissolve to obtain a sample solution; taking proper amount of astragaloside IV reference substance, adding methanol to obtain 1mg solution per 1ml, and using as astragaloside IV reference substance solution; adding methanol into appropriate amount of ginsenoside Rb1 and ginsenoside Rg1 to obtain mixed solution containing 0.5mg of ginsenoside Rb1 and ginsenoside Rg1 per 1ml as Notoginseng radix reference solution; sucking 10 mul of the test solution, 8 mul of each of the astragaloside IV reference solution and the pseudo-ginseng reference solution, respectively dropping on the same silica gel G thin layer plate, adding chloroform: methanol: the lower layer solution which is placed at the temperature of below 10 ℃ with the water = 13: 8: 2 is used as a developing agent, the developing agent is developed, taken out, dried, sprayed with an ethanol solution containing 1mol/L of sulfuric acid, and heated at the temperature of 105 ℃ until the spots are clearly developed. Spots with the same color appear on the chromatogram of the test solution at the positions corresponding to the chromatograms of the reference solution; inspecting under 365nm ultraviolet lamp, and displaying the same fluorescent spot to obtain the preparation containing radix astragali and Notoginseng radix.

3) Identifying gallnut by thin layer chromatography: weighing 4g of a sample, placing the sample in a conical flask with a plug, adding 50ml of methanol, placing the sample on a water bath, heating and refluxing for 40 minutes, taking down the sample, placing the sample at 4 ℃ for 3 hours, filtering, evaporating filtrate to dryness, adding 30ml of water into residue, heating to dissolve the residue, cooling, filtering, evaporating filtrate to dryness, and adding 5ml of methanol into the residue to dissolve the residue to obtain a sample solution; adding methanol into gallic acid control to obtain 1mg solution per 1ml as control solution; sucking 5 mul of each of the test solution and the reference solution, respectively dropping on the same silica gel GF254 thin layer plate, adding chloroform: ethyl formate: formic acid = 6: 3 as developing agent, developing, taking out, air drying, placing under 254nm ultraviolet lamp for inspection, and in the chromatogram of the test solution, spots with the same color appear on the corresponding positions of the chromatogram of the reference solution, thus determining the preparation containing the gallnut.

4) Gallnut is determined by high performance liquid chromatography: chromatographic conditions and system applicability tests, and octadecylsilane chemically bonded silica is used as a filler; taking a 12mmol/L phosphoric acid solution containing 4mol/L methanol as a mobile phase; the detection wavelength is 273 nm; the number of theoretical plates is not less than 5000 according to gallic acid peak.

Precisely weighing gallic acid reference substance, and preparing into 40 μ g/ml solution with 12mol/L methanol to obtain reference substance solution.

Weighing 4g of a sample, precisely weighing, placing in a conical flask with a plug, precisely adding 50ml of 5mol/L hydrochloric acid, sealing the plug, weighing, heating at 110 ℃ for 4 hours, cooling, weighing again, supplementing the lost weight with 5mol/L hydrochloric acid, shaking up, filtering, precisely weighing 1ml of a subsequent filtrate, placing in a 50ml measuring flask, adding 12mol/L methanol for diluting to a scale, and shaking up to obtain a sample solution.

Precisely sucking 15 μ l of each of the reference solution and the sample solution, injecting into a liquid chromatograph, and measuring; the content of Galla chinensis in the tested medicine is calculated by gallic acid according to external standard method.

Taking 15g of a sample, precisely weighing, placing in a conical flask with a plug, adding 100ml of trichloromethane, performing ultrasonic treatment for 30 minutes, filtering, adding 100ml of methanol into residue, heating to slightly boil for 3 hours, cooling, filtering, evaporating filtrate to dryness, adding 30ml of water into residue, shaking and extracting with water-saturated n-butyl alcohol for 2 times, 30ml each time, combining n-butyl alcohol solutions, washing with ammonia test solution for 2 times, 30ml each time, discarding ammonia solution, evaporating n-butyl alcohol solution to dryness, adding 5ml of water into residue to dissolve, passing through a D101 type macroporous resin column with the inner diameter of 1.5cm and the length of 10cm, eluting with 50ml of water, discarding water solution, eluting with 20ml of trichloromethane, 20ml of ethyl acetate and 30ml of 7mol/L ethanol, discarding eluent, then eluting with 90ml of methanol, collecting eluent, evaporating at 50 ℃ to dryness, dissolving with methanol, and fixing the volume to a 5ml volumetric flask, thus obtaining a sample solution.

Precisely sucking 10 μ l and 20 μ l of reference solution and 10 μ l of test solution respectively, injecting into liquid chromatograph, and measuring; the content of Astragalus membranaceus in the tested medicine is calculated by Astragaloside IV according to an external standard method.

6) Test results

The test results are shown in Table 14:

table 14 colitis enteric tablet test results

The results all meet the requirements of the quality standard detection method, and the method can effectively control the quality of the colitis enteric-coated tablet.

Example 5: the enema for treating colitis is determined according to the quality standard detection method of the invention.

1) Identifying pseudo-ginseng and bletilla striata by using a microscope: taking a detected product, and observing the characteristics of the rhizoma bletillae powder under a microscope: the superficial vertical wall of the epidermal cell is wavy and curved, rarely, the calcium oxalate needle crystal bundles exist in large round-like mucus fine bubbles or are scattered everywhere, the needle crystals are 63 mu m long, the fibers are bundled, the diameter is 24 mu m, and the needle crystals have herringbone or oval holes, and the diameters of the ladder conduit and the threaded conduit are 26 mu m; the characteristics of the pseudo-ginseng powder are as follows: the resin channel fragment contains yellow secretion, and the diameter of the ladder conduit and the threaded conduit is 31 μm, namely the preparation containing rhizoma bletillae and radix notoginseng powder is determined.

2) Identifying radix astragali and Notoginseng radix by thin layer chromatography: weighing 6g of a sample, placing the sample in a conical flask with a plug, adding 50ml of trichloromethane, carrying out ultrasonic treatment for 30 minutes, filtering, adding 50ml of methanol into residues, placing the residues on a water bath, heating and refluxing for 2 hours, cooling, filtering, evaporating the filtrate to dryness, adding 30ml of water into the residues, heating and dissolving the residues, extracting with water saturated n-butyl alcohol for 2 times, 30ml each time, combining n-butyl alcohol solutions, washing with 12ml of 0.25mol sodium hydroxide solution once, washing with 18ml of water once again, evaporating the n-butyl alcohol solution to dryness, and adding 2ml of methanol into the residues to dissolve the residues to obtain a sample solution; taking proper amount of astragaloside IV reference substance, adding methanol to obtain 1mg solution per 1ml, and using as astragaloside IV reference substance solution; adding methanol into appropriate amount of ginsenoside Rb1 and ginsenoside Rg1 to obtain mixed solution containing 0.5mg of ginsenoside Rb1 and ginsenoside Rg1 per 1ml as Notoginseng radix reference solution; sucking 10 mul of the test solution, 10 mul of each of the astragaloside IV reference solution and the pseudo-ginseng reference solution, respectively dropping the solutions on the same silica gel G thin layer plate, adding chloroform: methanol: the lower layer solution which is placed at the temperature of below 10 ℃ with the water = 11: 6: 1 is used as a developing agent, the developing agent is developed, taken out, dried, sprayed with an ethanol solution containing 1mol/L of sulfuric acid, and heated at the temperature of 105 ℃ until the spots are clearly developed. Spots with the same color appear on the chromatogram of the test solution at the positions corresponding to the chromatograms of the reference solution; inspecting under 365nm ultraviolet lamp, and displaying the same fluorescent spot to obtain the preparation containing radix astragali and Notoginseng radix.

3) Identifying gallnut by thin layer chromatography: weighing 2g of a sample, placing the sample in a conical flask with a plug, adding 50ml of methanol, placing the sample on a water bath, heating and refluxing for 20 minutes, taking down the sample, placing the sample at 1 ℃ for 3 hours, filtering, evaporating filtrate to dryness, adding 30ml of water into residue, heating to dissolve the residue, cooling, filtering, evaporating filtrate to dryness, and adding 5ml of methanol into the residue to dissolve the residue to obtain a sample solution; adding methanol into gallic acid control to obtain 1mg solution per 1ml as control solution; sucking 5 mul of each of the test solution and the reference solution, respectively dropping on the same silica gel GF254 thin layer plate, adding chloroform: ethyl formate: developing formic acid = 4: 1 as developing agent, taking out, air drying, and inspecting under 254nm ultraviolet lamp, wherein spots with the same color appear on the chromatogram of the test solution at the position corresponding to the chromatogram of the reference solution, thus determining the preparation containing Galla chinensis.

4) Gallnut is determined by high performance liquid chromatography: chromatographic conditions and system applicability tests, and octadecylsilane chemically bonded silica is used as a filler; taking a 6mmol/L phosphoric acid solution containing 3mol/L methanol as a mobile phase; the detection wavelength is 273 nm; the number of theoretical plates is not less than 5000 according to gallic acid peak.

Precisely weighing gallic acid reference substance, and preparing into 60 μ g/ml solution with 12mol/L methanol to obtain reference substance solution.

Weighing 2g of a sample, precisely weighing, placing in a conical flask with a plug, precisely adding 50ml of 3mol/L hydrochloric acid, sealing the plug, weighing, heating at 95 ℃ for 3 hours, cooling, weighing again, supplementing the lost weight with 3mol/L hydrochloric acid, shaking up, filtering, precisely weighing 1ml of a subsequent filtrate, placing in a 50ml measuring flask, adding 12mol/L methanol for diluting to a scale, and shaking up to obtain a sample solution.

Taking 8g of a sample, precisely weighing, placing in a conical flask with a plug, adding 100ml of trichloromethane, performing ultrasonic treatment for 30 minutes, filtering, adding 100ml of methanol into residue, heating to slightly boil for 2 hours, cooling, filtering, evaporating filtrate to dryness, adding 30ml of water into residue, shaking and extracting with water-saturated n-butyl alcohol for 2 times, 30ml each time, combining n-butyl alcohol solutions, washing with an ammonia test solution for 2 times, 30ml each time, discarding the ammonia solution, evaporating the n-butyl alcohol solution to dryness, adding 5ml of water into the residue to dissolve, passing through a D101 type macroporous resin column with the inner diameter of 1.5cm and the length of 10cm, eluting with 50ml of water, discarding a water solution, eluting with 8ml of trichloromethane, 8ml of ethyl acetate and 12ml of 7mol/L ethanol, discarding the eluent, subsequently eluting with 50ml of methanol, collecting the eluent, evaporating to dryness at 35 ℃, dissolving with methanol, and fixing the volume to a 5ml volumetric flask, thus obtaining a.

Precisely sucking 5 μ l and 20 μ l of reference solution and 20 μ l of test solution respectively, injecting into liquid chromatograph, and measuring; the content of Astragalus membranaceus in the tested medicine is calculated by Astragaloside IV according to an external standard method.

6) Test results

The test results are shown in Table 15:

TABLE 15 colitis enema test results

The results all meet the requirements of the quality standard detection method, and the method can effectively control the quality of the colitis enema.

Example 6: the enteric-coated pills for treating the colitis are determined according to the quality standard detection method.

1) Identifying pseudo-ginseng and bletilla striata by using a microscope: taking a detected product, and observing the characteristics of the rhizoma bletillae powder under a microscope: the superficial vertical wall of the epidermal cell is wavy and curved, rarely, the calcium oxalate needle crystal bundles exist in large round-like mucus fine bubbles or are scattered everywhere, the needle crystal length is 77 mu m, the fibers are bundled, the diameter is 28 mu m, and the needle crystal bundles have herringbone or oval holes, and the diameters of the ladder conduit and the threaded conduit are 52 mu m; the characteristics of the pseudo-ginseng powder are as follows: the resin channel fragment contains yellow secretion, and the diameter of the ladder conduit and the threaded conduit is 50 μm, namely the preparation containing rhizoma bletillae and radix notoginseng powder is determined.

2) Identifying radix astragali and Notoginseng radix by thin layer chromatography: weighing 9g of a sample, placing the sample in a conical flask with a plug, adding 50ml of trichloromethane, carrying out ultrasonic treatment for 30 minutes, filtering, adding 50ml of methanol into residues, placing the residues on a water bath, heating and refluxing for 3 hours, cooling, filtering, evaporating the filtrate to dryness, adding 30ml of water into the residues, heating and dissolving the residues, extracting with water-saturated n-butanol for 4 times, 40ml each time, combining the n-butanol solutions, washing with 20ml of 0.25mol sodium hydroxide solution once, washing with 30ml of water again, evaporating the n-butanol solution to dryness, and adding 2ml of methanol into the residues to dissolve the residues to obtain a sample solution; taking proper amount of astragaloside IV reference substance, adding methanol to obtain 1mg solution per 1ml, and using as astragaloside IV reference substance solution; adding methanol into appropriate amount of ginsenoside Rb1 and ginsenoside Rg1 to obtain mixed solution containing 0.5mg of ginsenoside Rb1 and ginsenoside Rg1 per 1ml as Notoginseng radix reference solution; sucking 10 mul of the test solution, 6 mul of each of the astragaloside IV reference solution and the pseudo-ginseng reference solution, respectively dropping on the same silica gel G thin layer plate, adding chloroform: methanol: the lower layer solution which is placed at the temperature of below 10 ℃ with the water = 14: 8: 2 is used as a developing agent, the developing agent is developed, taken out, dried, sprayed with an ethanol solution containing 1mol/L of sulfuric acid, and heated at the temperature of 105 ℃ until the spots are clearly developed. Spots with the same color appear on the chromatogram of the test solution at the positions corresponding to the chromatograms of the reference solution; inspecting under 365nm ultraviolet lamp, and displaying the same fluorescent spot to obtain the preparation containing radix astragali and Notoginseng radix.

4) Gallnut is determined by high performance liquid chromatography: chromatographic conditions and system applicability tests, and octadecylsilane chemically bonded silica is used as a filler; taking a 13mmol/L phosphoric acid solution containing 4mol/L methanol as a mobile phase; the detection wavelength is 273 nm; the number of theoretical plates is not less than 5000 according to gallic acid peak.

Precisely weighing gallic acid reference substance, and preparing into 90 μ g/ml solution with 12mol/L methanol to obtain reference substance solution.

Weighing 4g of a sample, precisely weighing, placing in a conical flask with a plug, precisely adding 50ml of 5mol/L hydrochloric acid, sealing the plug, weighing, heating at 115 ℃ for 4 hours, cooling, weighing again, supplementing the lost weight with 5mol/L hydrochloric acid, shaking up, filtering, precisely weighing 1ml of a subsequent filtrate, placing in a 50ml measuring flask, adding 12mol/L methanol for diluting to a scale, and shaking up to obtain a sample solution.

Taking 15g of a sample, precisely weighing, placing in a conical flask with a plug, adding 100ml of trichloromethane, performing ultrasonic treatment for 30 minutes, filtering, adding 100ml of methanol into residue, heating to slightly boil for 2 hours, cooling, filtering, evaporating filtrate to dryness, adding 30ml of water into residue, shaking and extracting with water-saturated n-butyl alcohol for 2 times, 30ml each time, combining n-butyl alcohol solutions, washing with an ammonia test solution for 2 times, 30ml each time, discarding the ammonia solution, evaporating the n-butyl alcohol solution to dryness, adding 5ml of water into the residue to dissolve, passing through a D101 type macroporous resin column with the inner diameter of 1.5cm and the length of 10cm, eluting with 50ml of water, discarding a water solution, eluting with 15ml of trichloromethane, 15ml of ethyl acetate and 30ml of 7mol/L ethanol, discarding the eluent, eluting with 90ml of methanol, collecting the eluent, evaporating to dryness at 50 ℃, dissolving with methanol, and fixing the volume to a 5ml volumetric flask, thus obtaining a sample.

6) Test results

The test results are shown in Table 15:

table 15 colitis enteric pill test results

The results all meet the requirements of the quality standard detection method, and show that the method can effectively control the quality of the colitis enteric-coated pills.

Claims

1. A method for detecting the quality of a medicine for treating colitis is characterized by comprising the following steps:

(1) identifying pseudo-ginseng and bletilla striata by using a microscope: taking a detected product, and observing the characteristics of the rhizoma bletillae powder under a microscope: the superficial vertical wall of the epidermal cell is wavy and curved, rarely, the calcium oxalate needle crystal bundles exist in large round-like mucus fine bubbles or are scattered everywhere, the needle crystal length is 18-88 mu m, the fibers are bundled, the diameter is 11-30 mu m, and the needle crystal bundles have herringbone or elliptical holes, and the diameters of the ladder conduit and the threaded conduit are 10-55 mu m; the characteristics of the pseudo-ginseng powder are as follows: the resin channel fragment contains yellow secretion, and the diameter of the ladder conduit and the threaded conduit is 10-55 μm, thus determining the preparation containing rhizoma bletilla and Notoginseng radix powder;

(2) identifying radix astragali and Notoginseng radix by thin layer chromatography: weighing 5-10g of a sample, placing the sample in a conical flask with a plug, adding 50ml of trichloromethane, carrying out ultrasonic treatment for 30 minutes, filtering, adding 50ml of methanol into residues, placing the residue on a water bath, heating and refluxing for 1-3 hours, cooling, filtering, evaporating filtrate, dissolving the residues by adding 30ml of water, extracting the residues by using water saturated n-butyl alcohol for 1-5 times, 20-40ml each time, combining n-butyl alcohol solutions, washing the residues by using 10-20ml of 0.25mol/L sodium hydroxide solution, washing the residues by using 10-30ml of water, evaporating the n-butyl alcohol solution, dissolving the residues by adding 2ml of methanol to obtain a sample solution; taking proper amount of astragaloside IV reference substance, adding methanol to obtain 1mg solution per 1ml, and using as astragaloside IV reference substance solution; adding methanol into appropriate amount of ginsenoside Rb1 and ginsenoside Rg1 to obtain mixed solution containing 0.5mg of ginsenoside Rb1 and ginsenoside Rg1 per 1ml as Notoginseng radix reference solution; sucking 5-10 μ L of test solution, 5-10 μ L of astragaloside IV control solution and 5-10 μ L of Notoginseng radix control solution, respectively dropping on the same silica gel G thin layer plate, developing with lower layer solution of chloroform, methanol and water = 10-15: 5-9: 1-3 at 10 deg.C as developing agent, taking out, air drying, spraying with ethanol solution containing 1mol/L of sulfuric acid, and heating at 105 deg.C until the spots are clearly developed;

spots with the same color appear on the chromatogram of the test solution at the positions corresponding to the chromatograms of the reference solution; inspecting under 365nm ultraviolet lamp to show the same fluorescent spot, and determining to be preparation containing radix astragali and Notoginseng radix;

(3) identifying gallnut by thin layer chromatography: weighing 1-5g of a sample, placing in a conical flask with a plug, adding 50ml of methanol, placing on a water bath, heating and refluxing for 10-50 minutes, taking down, placing at 0-5 ℃ for 3 hours, filtering, evaporating filtrate to dryness, adding 30ml of water into residue, heating to dissolve, cooling, filtering, evaporating filtrate to dryness, and adding 5ml of methanol into residue to dissolve to obtain a sample solution; adding methanol into gallic acid control to obtain 1mg solution per 1ml as control solution; sucking sample solution and reference solution 5 μ l each, respectively dropping on the same silica gel GF254 thin layer plate, developing with chloroform, ethyl formate and formic acid = 3-7: 1-3 as developing agent, taking out, air drying, inspecting under 254nm ultraviolet lamp, and displaying spots of the same color in the sample chromatogram at the position corresponding to the reference chromatogram to determine the preparation containing Galla chinensis;

(4) measuring gallnut by high performance liquid chromatography: chromatographic conditions and system applicability tests, and octadecylsilane chemically bonded silica is used as a filler; taking a 5-15mmol/L phosphoric acid solution containing 2-5mol/L methanol as a mobile phase; the detection wavelength is 273 nm; the number of theoretical plates is not less than 5000 according to the peak of gallic acid; precisely weighing gallic acid reference substance, and preparing into 12-120 μ g/ml solution with 12mol/L methanol to obtain reference substance solution; weighing 1-5g of a sample, precisely weighing, placing the sample in a conical flask with a plug, precisely adding 50ml of 2-6mol/L hydrochloric acid, sealing the plug, weighing, heating at 90-120 ℃ for 2-5 hours, cooling, weighing again, supplementing the lost weight with 2-6mol/L hydrochloric acid, shaking up, filtering, precisely weighing 1ml of subsequent filtrate, placing the subsequent filtrate in a 50ml measuring flask, adding 12mol/L methanol to dilute to scale, and shaking up to obtain a sample solution; precisely sucking 5-20 μ l of reference solution and sample solution respectively, injecting into liquid chromatograph, and measuring; the content of Galla chinensis in the tested medicine is calculated by gallic acid according to external standard method;