CN104865341A - Thin-layer chromatography detection method of traditional Chinese medicine composition for increasing animal immunity - Google Patents
Thin-layer chromatography detection method of traditional Chinese medicine composition for increasing animal immunity Download PDFInfo
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Abstract
The invention discloses a thin-layer chromatography detection method of a traditional Chinese medicine composition. The thin-layer chromatography detection method comprises the following steps: a, preparing a test solution; b, preparing a reference solution; c, respectively applying the test solution, the reference solution of astragaloside, the reference solution of oleanolic acid and the reference solution of pinoresinol diglucoside to the same silica gel plate, developing by taking the lower solution of chloroform-ethyl acetate-methanol-water as a developing agent, taking out, airing, and heating by taking 2% vanillic aldehyde sulfuric acid solution as a colour developing agent till the colour of spots is developed clearly; and d, viewing the silica gel plate after the colour is developed. The thin-layer chromatography detection method of the traditional Chinese medicine composition disclosed by the invention can be used for simultaneously detecting astragalus membranaceus, glossy privet fruits and the bark of eucommia in the traditional Chinese medicine composition; the detection result is accurate and reliable; and simultaneously, the thin-layer chromatography detection method disclosed by the invention has the advantages of being few in process, simple and convenient to operate, short in detection time, low in cost and the like and is particularly applied to industrial application.
Description
Technical field
The present invention relates to a kind of thin-layered chromatography detection method improving the Chinese medicine composition of animal resistibility.
Background technology
Thin-layer chromatography (method) is a kind of conventional Chinese medical herb method, has that equipment is simple, easy and simple to handle, velocity of separation is fast, sensitivity and resolution comparatively advantages of higher.
The Radix Astragali, have another name called yellow over sixty years of age, be the dry root of legume astragalus mongolicus, having the effects such as invigorating qi for strengthening superficies, sharp water detumescence, pus draining and toxin expelling, myogenic, is a kind of Chinese crude drug.The complex chemical composition of the Radix Astragali, mainly contains saponins, flavonoids and glycoside thereof, polysaccharide, amino acids etc.The fruit of glossy privet is the dry mature fruit of Oleaceae Ligustrum plant glossy privet, sweet, the micro-bitterness of taste, have merit that is nourishing liver and kidney, black of improving eyesight, can be used for the symptoms such as the deficiency of liver-yin and kidney-yin, dizzy tinnitus, poliosis, order are secretly failed to understand, interior heat is quenched one's thirst, osteopyrexia and fever, is a kind of Chinese crude drug.The complex chemical composition of the fruit of glossy privet, mainly contains three mushroom classes, flavonoids, phenylethanol etc.The bark of eucommia is the bark of Angiospermae Eucommiaceae bark of eucommia platymiscium, is a kind of Chinese crude drug.
In prior art, the Radix Astragali, the fruit of glossy privet, the medicinal material such as the bark of eucommia and composition thereof adopts thin-layer chromatography to differentiate usually, but, existing discrimination method can only to the Radix Astragali, the fruit of glossy privet, a kind of medicinal material in the bark of eucommia carries out Qualitive test, as: Liu Xiaoqing etc. (Liu Xiao Qing Li army is scape Lian Lixiao gorgeous Men Qiming Kang Ting state recklessly. and in Milkvetch Root, calycosin glucoside thin-layer chromatography is differentiated fast. Liaoning University of TCM's journal, in July, 2013, 15th volume, 7th phase .) prepare need testing solution by specific pre-treatment, and select specific methenyl choloride-methanol-water (7.5:2.5:0.2) to be developping agent, thin layer clear spot could be obtained, the good testing result of degree of separation, Zhou Yang etc. (raise Liu Li Xu De week raw. the indentification by TLC of special glossy privet hardship in fruit of glossy privet medicine materical crude slice. Chinese medicine company, on October 5th, 2013,22nd volume, 19th phase .) prepare need testing solution by specific pre-treatment, and select specific acetic acid ethyl ester-acetone-formic acid-water (6:3:1:1) to be developping agent, thin layer clear spot, the good testing result of degree of separation could be obtained, Liu Ling etc. (Liu Ling Bao Jia section. Eucommia ulmoides indentification by TLC is studied. China National folk medicine, 2013 .) prepare need testing solution by specific pre-treatment, and select specific methenyl choloride-n-butanol-water-acetic acid solution (1:7:1:1) to be developping agent, the small adjustment that the shaker test of its developping agent demonstrates each group component in developping agent thin layer clear spot, the good testing result of degree of separation could be obtained, even also can bring very large impact to testing result.
China document (Zhai less admire Lee Cao Guowen become monarch. the TLC distinguish of compound gold granules. Chinese veterinarian's medical magazine, 2010, 6th phase .) disclose the thin-layer identification method of compound gold granules, the method needs the Radix Astragali, the fruit of glossy privets etc. adopt different pre-treatments to prepare need testing solution respectively, select different developping agents respectively, carry out thin-layer chromatography detection respectively, the method can not to the Radix Astragali in compound granular, the components such as the fruit of glossy privet carry out pre-treatment simultaneously and thin-layer chromatography detects, there is operation many, complicated operation, detection time is long, the many defects of high in cost of production.
Therefore, few, the easy and simple to handle and new method that simultaneously can detect the Radix Astragali, the fruit of glossy privet, the bark of eucommia of a kind of operation is needed to invent.
Summary of the invention
The object of the present invention is to provide a kind of thin-layered chromatography detection method of Chinese medicine composition.
The thin-layered chromatography detection method of a kind of Chinese medicine composition provided by the invention, comprises the following steps:
A, prepare need testing solution: get Chinese medicine composition, add methyl alcohol A, ultrasonic process, filter, get supernatant; Supernatant miillpore filter filters, evaporate to dryness, and residue, with after 10%v/v ammonia solvent, with water-saturated n-butanol extraction, is extracted liquid; By extract evaporate to dryness, residue methyl alcohol B dissolves, and obtains need testing solution;
B, preparation reference substance solution: get Astragaloside IV, oleanolic acid, Pinoresinol diglucoside and dissolve with methyl alcohol respectively, obtain the reference substance solution of Astragaloside IV, the reference substance solution of oleanolic acid, the reference substance solution of Pinoresinol diglucoside;
C, by the reference substance solution of the reference substance solution of need testing solution, Astragaloside IV, oleanolic acid, the reference substance solution of Pinoresinol diglucoside, point sample is in same silica gel plate respectively, with lower floor's solution of chlorofonn-ethylacetate-methanol-water for developping agent, launch, take out, dry, with 2% vanillin-sulfuric acid solution for developer, 105 DEG C to be heated to spot development clear;
D, inspect the silica gel plate after colour developing.
Further, in step a, described Chinese medicine composition is made up of following raw material: the Radix Astragali, the fruit of glossy privet, the bark of eucommia.
Further, in step a, described Chinese medicine composition is made up of the raw material of following parts by weight: the Radix Astragali 18 parts, the fruit of glossy privet 9 parts, the bark of eucommia 9 parts.
Further, in step a, described Chinese medicine composition is prepared from according to following steps:
1., get the Radix Astragali, the fruit of glossy privet, the bark of eucommia, be crushed to 80 orders, dry, obtain radix astragali coarse powder, fruit of glossy privet meal, bark of eucommia meal respectively;
2., under-10 DEG C ~-20 DEG C conditions, radix astragali coarse powder step 1. obtained is crushed to 100 order ~ 800 object granularities further, obtains Radix Astragali Ultramicro-powder;
3., by fruit of glossy privet meal mix with bark of eucommia meal, with soak by water twice, filter, concentrated, obtain concentrate;
4. the Radix Astragali Ultramicro-powder that 2. concentrate, by step 3. obtained and step obtain mixes, and granulates, and dries, and crosses 80 mesh sieves, to obtain final product.
Further, in step a, the w/v of Chinese medicine composition and methyl alcohol A is 1:15g/ml; The volume ratio of methyl alcohol A, 10%v/v ammoniacal liquor, water-saturated n-butanol, methyl alcohol B is 6:2:3:1.
Further, in step a, the time of ultrasonic process is 60min; The aperture of miillpore filter is 0.45 μm.
Further, in step b, the concentration of the reference substance solution of the reference substance solution of Astragaloside IV, the reference substance solution of oleanolic acid, Pinoresinol diglucoside is respectively 1mg/ml.
Further, in step c, described silica gel plate is silica gel g thin-layer plate.
Further, in step c, in chlorofonn-ethylacetate-methanol-water, the volume ratio of chloroform, ethyl acetate, methyl alcohol, water is 20:40:22:1.
Further, in steps d, the silica gel plate after colour developing, on the position corresponding to reference substance chromatogram, the spot of test sample Chromatogram display same color.
The thin-layered chromatography detection method of Chinese medicine composition of the present invention, can detect the Radix Astragali in Chinese medicine composition, the fruit of glossy privet, the bark of eucommia simultaneously, testing result accurately, reliably, simultaneously, the inventive method has that operation is few, easy and simple to handle, detection time is short, low cost and other advantages, is applicable to very much the application in industry.
Obviously, according to foregoing of the present invention, according to ordinary technical knowledge and the customary means of this area, not departing under the present invention's above-mentioned basic fundamental thought prerequisite, the amendment of other various ways, replacement or change can also be made.
The embodiment of form by the following examples, is described in further detail foregoing of the present invention again.But this should be interpreted as that the scope of the above-mentioned theme of the present invention is only limitted to following example.All technology realized based on foregoing of the present invention all belong to scope of the present invention.
Accompanying drawing explanation
Fig. 1 is the thin-layer chromatography testing result of Chinese medicine composition of the present invention, wherein, and " pine " representative " Pinoresinol diglucoside ", " first " representative " Astragaloside IV ", " together " representative " oleanolic acid ", " sample " representative " test sample ".
Embodiment
The raw material used in the specific embodiment of the invention, equipment are known product, obtain by buying commercially available prod.
The thin-layered chromatography detection method of embodiment 1, Chinese medicine composition of the present invention
1, Chinese medicine composition is prepared
Count by weight: the Radix Astragali 18 parts, the fruit of glossy privet 9 parts, the bark of eucommia 9 parts; Use Universalpulverizer to be crushed to 80 orders respectively three kinds of medicinal materials, dry, make the moisture of each meal medicinal material control 6% ~ 8%; Under temperature is-10 DEG C ~-20 DEG C conditions, radix astragali coarse powder is crushed to the Ultramicro-powder that granularity is 100 ~ 800 order granularities; By the fruit of glossy privet and the mixing of bark of eucommia meal, with soak by water twice, filter, mixing filtrate concentrates, and finally by concentrate and the mixing of Radix Astragali Ultramicro-powder, granulation post-drying, crosses 80 mesh sieves, obtain Chinese medicine composition.
2, detection method
Above-mentioned traditional Chinese medicine particle ground, precision takes powder 2.0g, is placed in triangular flask, adds methyl alcohol 30ml, ultrasonic process 60min, filters, gets supernatant; Supernatant filters with 0.45 μm of miillpore filter (PTFE), is placed in evaporating dish evaporate to dryness; Residue 10ml 10% (V/V) ammonia solvent, places 10min, frequently jolting; Solution is moved in separating funnel, extract 3 times with 15ml water-saturated n-butanol; Combining extraction liquid, uses evaporating dish evaporate to dryness, and residue adds 5ml methyl alcohol and dissolves, and obtains need testing solution;
Get Astragaloside IV, oleanolic acid, Pinoresinol diglucoside, add methyl alcohol, make the solution of every 1ml containing 1mg respectively, product solution in contrast.
Test according to thin-layered chromatography (Pharmacopoeia of People's Republic of China 2010 editions annex VI B), draw above-mentioned need testing solution, each 3 μ l of reference substance solution, put respectively on same silica gel g thin-layer plate, with lower floor's solution of chlorofonn-ethylacetate-methanol-water (20:40:22:1) for developping agent, launch, take out, dry, spray, with 2% vanillin-sulfuric acid solution, is heated to spot development at 105 DEG C clear; Its testing result is shown in Fig. 1, on the position corresponding to reference substance chromatogram, and the spot of test sample Chromatogram display same color.
The thin-layered chromatography detection method of Chinese medicine composition of the present invention, can detect the Radix Astragali in Chinese medicine composition, the fruit of glossy privet, the bark of eucommia simultaneously, testing result accurately, reliably, simultaneously, the inventive method has that operation is few, easy and simple to handle, detection time is short, low cost and other advantages, is applicable to very much the application in industry.
Claims (10)
1. a thin-layered chromatography detection method for Chinese medicine composition, is characterized in that: described discrimination method comprises the following steps:
A, prepare need testing solution: get Chinese medicine composition, add methyl alcohol A, ultrasonic process, filter, get supernatant; Supernatant miillpore filter filters, evaporate to dryness, and residue, with after 10%v/v ammonia solvent, with water-saturated n-butanol extraction, is extracted liquid; By extract evaporate to dryness, residue methyl alcohol B dissolves, and obtains need testing solution;
B, preparation reference substance solution: get Astragaloside IV, oleanolic acid, Pinoresinol diglucoside and dissolve with methyl alcohol respectively, obtain the reference substance solution of Astragaloside IV, the reference substance solution of oleanolic acid, the reference substance solution of Pinoresinol diglucoside;
C, by the reference substance solution of the reference substance solution of need testing solution, Astragaloside IV, oleanolic acid, the reference substance solution of Pinoresinol diglucoside, point sample is in same silica gel plate respectively, with lower floor's solution of chlorofonn-ethylacetate-methanol-water for developping agent, launch, take out, dry, with 2% vanillin-sulfuric acid solution for developer, 105 DEG C to be heated to spot development clear, obtains the silica gel plate after developing the color;
D, inspect the silica gel plate after colour developing.
2. thin-layered chromatography detection method according to claim 1, is characterized in that: in step a, and described Chinese medicine composition is made up of following raw material: the Radix Astragali, the fruit of glossy privet, the bark of eucommia.
3. thin-layered chromatography detection method according to claim 2, is characterized in that: in step a, and described Chinese medicine composition is made up of the raw material of following parts by weight: the Radix Astragali 18 parts, the fruit of glossy privet 9 parts, the bark of eucommia 9 parts.
4. the thin-layered chromatography detection method according to Claims 2 or 3, is characterized in that: in step a, and described Chinese medicine composition is prepared from according to following steps:
1., get the Radix Astragali, the fruit of glossy privet, the bark of eucommia, be crushed to 80 orders, dry, obtain radix astragali coarse powder, fruit of glossy privet meal, bark of eucommia meal respectively;
2., under-10 DEG C ~-20 DEG C conditions, radix astragali coarse powder step 1. obtained is crushed to 100 order ~ 800 object granularities further, obtains Radix Astragali Ultramicro-powder;
3., by fruit of glossy privet meal mix with bark of eucommia meal, with soak by water twice, filter, concentrated, obtain concentrate;
4. the Radix Astragali Ultramicro-powder that 2. concentrate, by step 3. obtained and step obtain mixes, and granulates, and dries, and crosses 80 mesh sieves, to obtain final product.
5. thin-layered chromatography detection method according to claim 1, is characterized in that: in step a, and the w/v of Chinese medicine composition and methyl alcohol A is 1:15g/ml; The volume ratio of methyl alcohol A, 10%v/v ammoniacal liquor, water-saturated n-butanol, methyl alcohol B is 6:2:3:1.
6. thin-layered chromatography detection method according to claim 1, is characterized in that: in step a, and the time of ultrasonic process is 60min; The aperture of miillpore filter is 0.45 μm.
7. thin-layered chromatography detection method according to claim 1, is characterized in that: in step b, and the concentration of the reference substance solution of the reference substance solution of Astragaloside IV, the reference substance solution of oleanolic acid, Pinoresinol diglucoside is respectively 1mg/ml.
8. thin-layered chromatography detection method according to claim 1, is characterized in that: in step c, and described silica gel plate is silica gel g thin-layer plate.
9. thin-layered chromatography detection method according to claim 1, is characterized in that: in step c, and in chlorofonn-ethylacetate-methanol-water, the volume ratio of chloroform, ethyl acetate, methyl alcohol, water is 20:40:22:1.
10. thin-layered chromatography detection method according to claim 1, is characterized in that: in steps d, the silica gel plate after colour developing, on the position corresponding to reference substance chromatogram, and the spot of test sample Chromatogram display same color.
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| CN106596829A (en) * | 2015-10-14 | 2017-04-26 | 镇江先创生物科技有限公司 | Thin-layer chromatography detection method for high-toxic pesticide carbofuran residues |
| CN108226369A (en) * | 2016-12-15 | 2018-06-29 | 江苏维赛科技生物发展有限公司 | A kind of remaining thin-layered chromatography detection method of high poison antistaling agent thiabendazole |
| CN109752480A (en) * | 2017-11-05 | 2019-05-14 | 江苏维赛科技生物发展有限公司 | A kind of remaining thin-layered chromatography detection method of high poison antistaling agent o-phenyl phenol |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN106596829A (en) * | 2015-10-14 | 2017-04-26 | 镇江先创生物科技有限公司 | Thin-layer chromatography detection method for high-toxic pesticide carbofuran residues |
| CN108226369A (en) * | 2016-12-15 | 2018-06-29 | 江苏维赛科技生物发展有限公司 | A kind of remaining thin-layered chromatography detection method of high poison antistaling agent thiabendazole |
| CN109752480A (en) * | 2017-11-05 | 2019-05-14 | 江苏维赛科技生物发展有限公司 | A kind of remaining thin-layered chromatography detection method of high poison antistaling agent o-phenyl phenol |
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