CN106770882B - A kind of detection method of the Chinese medicine preparation containing ginseng or pseudo-ginseng - Google Patents
A kind of detection method of the Chinese medicine preparation containing ginseng or pseudo-ginseng Download PDFInfo
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Abstract
The present invention is a kind of detection method of the Chinese medicine preparation containing ginseng or pseudo-ginseng, and this method comprises the following steps：（1）Chinese medicine composition or Chinese medicine preparation containing ginseng or pseudo-ginseng are taken, prepares need testing solution；（2）Take and prepare negative sample solution without ginseng, the finished product of pseudo-ginseng；（3）Take ginsenoside Rb1's reference substance, ginsenoside Re's reference substance, ginsenoside Rg1's reference substance, ginsenoside Rf's control and notoginsenoside R reference substance, add methanol that the mixed solution that every 1ml methanol respectively contains 0.5 2mg above-mentioned reference substance is made, as reference substance mixed solution；（4）Detection；（5）Interpretation of result.The present invention can differentiate ginseng, the taste medicinal material of pseudo-ginseng two simultaneously, and index components ginsenoside Rb1, ginsenoside Re, notoginsenoside R, ginsenoside Rf and ginsenoside Rg1 are differentiated simultaneously.The detection method has the characteristics of easy, easy, specificity is strong, reproducible.
Description
Technical field
The present invention relates to field of medicine preparing technology, the inspection of specially a kind of Chinese medicine preparation containing ginseng or pseudo-ginseng
Survey method.
Background technology
Ginseng, pseudo-ginseng belong to tonic, often alone or share in Chinese medicine preparation, are mostly monarch-minister drug when sharing, the two
Contained chemical composition belongs to saponins together, and structure is similar, and thin-layer chromatography behavior is also close, to the Chinese medicine containing ginseng, pseudo-ginseng
It is difficult that preparation formulates index zone.
2015《Chinese Pharmacopoeia》In to the Chinese medicine preparation containing ginseng, pseudo-ginseng using TLC Identification identify
When, differentiated using ginsenoside Rb1, ginsenoside Rg1 and notoginsenoside R as reference substance, but in real work, by
It is very close in the Rf values of notoginsenoside R and ginsenoside Re, it is difficult to separate, pseudo-ginseng is substituted with ginseng so that being likely to occur
Or pseudo-ginseng substitutes the false positive results that ginseng feeds intake, the specificity differentiated in Chinese medicine preparation to ginseng, pseudo-ginseng is influenceed.
The content of the invention
The present invention is exactly to be directed to above technical problem, there is provided a kind of detection of the Chinese medicine preparation containing ginseng or pseudo-ginseng
Method.The detection method has the characteristics of easy, easy, specificity is strong, reproducible, is advantageous to improve and contains ginseng or pseudo-ginseng
Chinese medicine preparation quality standard, prevent from producing and occur substituting pseudo-ginseng with ginseng or pseudo-ginseng substitutes the Chinese medicine system that ginseng feeds intake
Agent, it is favorably improved the Clinical practice safety and stability of such Chinese medicine preparation.
The concrete technical scheme of the present invention is as follows：
A kind of detection method of the Chinese medicine preparation containing ginseng or pseudo-ginseng, comprises the following steps：
（1）Prepare need testing solution：Chinese medicine composition or Chinese medicine preparation containing ginseng or pseudo-ginseng are taken, coating is removed, grinds
Carefully, powder 2-5g is taken, adds chloroform 40ml, is heated to reflux 1 hour, discards chloroform liquid, the dregs of a decoction volatilize solvent, add water
0.5ml stirring moistenings, add water-saturated n-butanol 10ml, are ultrasonically treated 30 minutes, and Aspirate supernatant adds 3 times of amount ammonia solutions, shakes up,
Layering is placed, takes upper liquid to be evaporated, residue adds methanol 2-3ml to make dissolving, as need testing solution；
（2）Prepare negative sample solution：Prepared by the preparation technology of Chinese medicine composition or Chinese medicine preparation without ginseng, pseudo-ginseng
Finished product, it is finely ground, take powder 2-5g, add chloroform 40ml, be heated to reflux 1 hour, discard chloroform liquid, the dregs of a decoction volatilize molten
Agent, add water 0.5ml stirring moistenings, add water-saturated n-butanol 10ml, be ultrasonically treated 30 minutes, Aspirate supernatant adds 3 times of amount ammonia examinations
Liquid, shake up, place layering, take upper liquid to be evaporated, residue adds methanol 2-3ml to make dissolving, as negative sample solution；
（3）Prepare reference substance solution：Take ginsenoside Rb1's reference substance, ginsenoside Re's reference substance, ginsenoside Rg1
Reference substance, ginsenoside Rf's control and notoginsenoside R reference substance, add methanol that every 1ml methanol is made respectively containing the above-mentioned of 0.5-2mg
The mixed solution of reference substance, as reference substance mixed solution；
（4）Detection：Take step（3）Described mixed reference substance solution 1-3 μ l, step（1）Described need testing solution（Can
Prepare 3 batches, respectively 1 batch, 2 batches, 3 batches）0.3-3 μ l and step（2）Described negative sample solution 0.3-3 μ l, put respectively in
On same silica gel g thin-layer plate, with volume ratio 90：45：6.5 dichloromethane-absolute ethyl alcohol-water is solvent, and solvent is placed in
In chromatography cylinder, in 5 DEG C -20 DEG C of temperature, relative humidity 40%-80% environment in saturation 20 minutes, be then placed in lamellae, continue
After presaturation 30 minutes, deploy, take out, dry, spray with 10% ethanol solution of sulfuric acid, heating 2-3 minutes to spot at 105 DEG C shows
Color is clear, is inspected respectively under daylight and ultraviolet light 365nm；（The configuration of 10% ethanol solution of sulfuric acid:Refer to the dense sulphur with 10ml
Acid is added in 90ml absolute ethyl alcohol, is produced.The mass fraction of the concentrated sulfuric acid of use is in 95%-98%.
（5）Interpretation of result：Containing ginseng, the Chinese medicine preparation of pseudo-ginseng test sample chromatogram in, with reference substance ginsenoside
Rb1, reference substance ginsenoside Re, reference substance notoginsenoside R, reference substance ginsenoside Rf and reference substance ginsenoside Rg1
On chromatogram relevant position, the spot of same color is shown under daylight, the fluorescence spot of same color is shown under ultraviolet light；Containing ginseng,
In the test sample chromatogram of Chinese medicine preparation without pseudo-ginseng, with reference substance ginsenoside Rb1, reference substance ginsenoside Re, control
On the chromatogram relevant position of product ginsenoside Rf and reference substance ginsenoside Rg1, the spot of same color is shown under daylight, it is ultraviolet
Show the fluorescence spot of same color under light；In the test sample chromatogram of Chinese medicine preparation containing pseudo-ginseng, without ginseng, reference substance ginseng
Saponin(e Rb1, reference substance ginsenoside Re, reference substance notoginsenoside R and the chromatogram relevant position of reference substance ginsenoside Rg1
On, the spot of same color is shown under daylight, the fluorescence spot of same color is shown under ultraviolet light；Negative sample without ginseng, pseudo-ginseng
In product solution chromatogram, with reference substance chromatogram relevant position, not showing spot under daylight, do not show fluorescence spot under ultraviolet light.
Above all of %, unless otherwise specified, it is represented as its weight/mass percentage composition.
The positive effect of the present invention is embodied in：
（One）, the detection method have that easy, easy, toxicity is low（Agents useful for same compares in the detection method development system
Relevant ginseng that Chinese Pharmacopoeia provides, agents useful for same toxicity is low in pseudo-ginseng detection method, mainly this detection method dichloromethane
Alkane substitutes chloroform, avoids the carcinogenicity of chloroform）, the characteristics of specificity is strong, reproducible.
（Two）, be advantageous to improve the quality standard of the Chinese medicine preparation containing ginseng or pseudo-ginseng, prevent from producing appearance with people
Ginseng substitutes pseudo-ginseng or pseudo-ginseng substitutes the Chinese medicine preparation that ginseng feeds intake, and is favorably improved the Clinical practice security of such Chinese medicine preparation
And stability.
Brief description of the drawings：
Fig. 1-1-1 are the thin-layer chromatogram that development system I deploys；
Fig. 1-1-2 are the thin-layer chromatogram that development system II deploys；
Fig. 1-1-3 are the thin-layer chromatogram that development system III deploys；
Fig. 1-1-4 are the thin-layer chromatogram that development system IV deploys,
Wherein, 1 is mixing reference substance 1 μ l；2 be the μ l of test sample point sample 0.5；3 be the μ l of test sample point sample 0.5；4 be test sample
The μ l of point sample 0.5；5 be the μ l of negative sample 0.5.
Fig. 1-2-1 is the thin-layer chromatogram that deploys after spot application mode point sample；
Fig. 1-2-2 is the thin-layer chromatogram that deploys after band application mode point sample；
Wherein, 1 is mixing reference substance 1 μ l；2 be the μ l of test sample point sample 0.2；3 be the μ l of test sample point sample 0.3；4 be test sample
The μ l of point sample 0.4；5 be the μ l of test sample point sample 0.5.
Fig. 1-3 need testing solutions thin layer differentiates the thin-layer chromatogram of point sample amount；
Wherein, 1 is mixing reference substance 1 μ l；2 be the μ l of test sample point sample 0.2；3 be the μ l of test sample point sample 0.3；4 be test sample
The μ l of point sample 0.4；5 be the μ l of test sample point sample 0.5.
Fig. 1-4-1 be using Tianjin Si Lida Science and Technology Ltd.s --- can clipboard expansion thin-layer chromatogram；
Fig. 1-4-2 are using Tianjin Si Lida Science and Technology Ltd.s --- the thin-layer chromatogram of efficient plate expansion；
Fig. 1-4-3 are to be made people rich source silica gel chemical reagent work using Qingdao --- the thin-layer chromatogram of glass plate expansion；
Fig. 1-4-4 are using subsidiary factory of Haiyang Chemical Plant, Qingdao --- the thin-layer chromatogram of glass plate expansion；
Wherein, 1：Mix the μ l of reference substance 1；2：The μ l of test sample point sample 0.3；3：The μ l of test sample point sample 0.3；4：Test sample point sample
0.3μl；5：The μ l of negative sample 0.3.
Fig. 1-5-1 are the thin-layer chromatogram deployed in 35 DEG C of environment of temperature；
Fig. 1-5-2 are the thin-layer chromatogram deployed in 30 DEG C of environment of temperature；
Fig. 1-5-3 are the thin-layer chromatogram deployed in 25 DEG C of environment of temperature；
Fig. 1-5-4 are the thin-layer chromatogram deployed in 20 DEG C of environment of temperature；
Fig. 1-5-5 are the thin-layer chromatogram deployed in 5 DEG C of environment of temperature；
Wherein, 1：Mix the μ l of reference substance 1；2：The μ l of test sample point sample 0.3；3：The μ l of test sample point sample 0.3；4：Test sample point sample
0.3μl；5：The μ l of negative sample point sample 0.3.
Fig. 1-6-1 are the thin-layer chromatogram deployed in the humidity environment of relative humidity 40%；
Fig. 1-6-2 are the thin-layer chromatogram deployed in the humidity environment of relative humidity 88%；
Wherein, 1：Mix the μ l of reference substance 1；2：The μ l of test sample point sample 0.3；3：The μ l of test sample point sample 0.3；4：For examination
The μ l of product point sample 0.3；5：The μ l of negative sample point sample 0.3.
The thin-layer chromatogram that Fig. 1-7-1 specificities are investigated；
Wherein, 1：Mix the μ l of reference substance 1；2：The μ l of test sample point sample 0.3；3：The μ l of test sample point sample 0.3；4：Test sample point sample
0.3μl；5：The μ l of negative sample point sample 0.3.
Ginseng, the thin layer of pseudo-ginseng need testing solution point sample amount differentiate chromatogram in Fig. 2-1 Kang Er heart-soothing capsules；
Wherein, 1：Mix the μ l of reference substance 1.5；2：The μ l of test sample point sample 0.2；3：The μ l of test sample point sample 0.3；4：Test sample point
The μ l of sample 0.4；5：The μ l of test sample point sample 0.5；6：The μ l of test sample point sample 0.6.
The thin-layer chromatogram that ginseng, pseudo-ginseng specificity are investigated in Fig. 2-2 Kang Er heart-soothing capsules,
Wherein, 1：Mix the μ l of reference substance 1.5；2：The μ l of 1 point sample of test sample 0.5；3：The μ l of 2 point sample of test sample 0.5；4：Test sample
The μ l of 3 point sample 0.5；5：The μ l of negative sample point sample 0.5
The thin layer of ginseng need testing solution point sample amount differentiates chromatogram in Fig. 3-1 Yixinshu tablets;
Wherein, 1：Mix the μ l of reference substance 2; 2：The μ l of test sample point sample 1; 3：The μ l of test sample point sample 2; 4：Test sample point sample 3
μl; 5：The μ l of test sample point sample 4; 6：The μ l of test sample point sample 5
The thin-layer chromatogram that ginseng specificity is investigated in Fig. 4 Yixinshu tablets；
Wherein, 1：Mix the μ l of reference substance 2；2：The μ l of 1 point sample of test sample 3；3：The μ l of 2 point sample of test sample 3；4：The point sample 3 of test sample 3
μl；5：The μ l of negative sample point sample 3.
The thin layer of pseudo-ginseng need testing solution point sample amount differentiates chromatogram in Fig. 4-1 sheep leaves of pulse plants notoginseng capsules；
Wherein, 1：Mix the μ l of reference substance 1；2：The μ l of test sample point sample 0.2；3：The μ l of test sample point sample 0.3；4：Test sample point sample
0.4μl；5：The μ l of test sample point sample 0.5；6：The μ l of test sample point sample 0.6.
Fig. 4-2 is the thin-layer chromatogram that pseudo-ginseng specificity is investigated in sheep leaves of pulse plants notoginseng capsule.
Wherein, 1：Mix the μ l of reference substance 1；2：The μ l of 1 point sample of test sample 0.3；3：The μ l of 2 point sample of test sample 0.3；4：Test sample 3
The μ l of point sample 0.3；5：The μ l of negative sample point sample 0.3.
Embodiment
In order to make the purpose , technical scheme and advantage of the present invention be clearer, with reference to embodiment pair
The present invention is described in further detail, but the scope that this should not be interpreted as to the above-mentioned theme of the present invention is only limitted to following implementations
Example.
1 instrument and reagent：
KQ5200DB type numerical control ultrasonic cleaners（Kunshan Ultrasonic Instruments Co., Ltd.）；The electric heating constant temperature water of DK-98- II
Bath（Tianjin Qin Site Instrument Ltd.）, silica gel g thin-layer plate（Tianjin Si Lida Science and Technology Ltd.s, can clipboard, batch
Number：160122）, silica gel 60A（Tianjin Si Lida Science and Technology Ltd.s, the efficient plate of type, lot number can be cut：16025）, silica G is thin
Laminate（Qingdao wave silica-gel desiccant Co., Ltd, glass plate, lot number：20150502）, silica gel g thin-layer plate（Qingdao Haiyang
Subsidiary factory of factory, glass plate, lot number：20160611）, thin layer chromatogram scanner（CAMAG, made in Switzerland）, it is automatic
Point sample instrument（CAMAG, made in Switzerland）, ginsenoside Rf（National Institute for Food and Drugs Control, for containing measurement
It is fixed to use, lot number：111719-201104）, ginsenoside Rg1（National Institute for Food and Drugs Control, for assay, criticize
Number：110703-201529）, ginsenoside Re（National Institute for Food and Drugs Control, for assay, lot number：110754-
201525）, ginsenoside Rb1（National Institute for Food and Drugs Control, for assay, lot number：110704-201424）、
Panax Notoginseng saponin R1（National Institute for Food and Drugs Control, for assay, lot number：110745-201318）；It is methanol, anhydrous
Ethanol, ethyl acetate, chloroform, dichloromethane, n-butanol are that analysis is pure.
Anti- silly pellet, the formula composition of its Chinese medicine composition are：Ginseng 181.1g, Ligusticum wallichii 181.1g, pseudo-ginseng 90.5g, leech
90.5g, borneol 5.434g, Moschus 1.811g.
Anti- red specific preparation method of being crazy about is as follows：Above Six-element bulk drug is weighed in proportion, and leech is ground into coarse powder, adds 10
Times quality water seethes with excitement decoction slightly, decocts 2 times, 1 hour every time, merges decocting liquid, filtration, is concentrated into relative density as 1.0, adds
Ethanol to concentration is 60wt%, stands 24 hours, filtration, and slag of getting it filled is standby；Ginseng, Ligusticum wallichii, the taste medicine of pseudo-ginseng three add up 6 times of quality
60wt% alcohol reflux extracts 3 times, 1.5 hours every time, merges extract solution, filtration, reclaims ethanol, is concentrated into right amount, will be above-mentioned
The dregs of a decoction are miscible with concentrate, spray drying, dried cream powder are made；Borneol and muscone grind altogether, add dried cream powder and appropriate amount of auxiliary materials,
Mix, particle is made, suppress 1000, film coating, produce.
Experimental example 1：Ginseng, the indentification by TLC of pseudo-ginseng in a kind of i.e. anti-silly pellet of Chinese medicine composition
1. the preparation of need testing solution：Anti- silly pellet 20 is taken, removes coating, it is finely ground, powder 5g is taken, adds chloroform
40ml, it is heated to reflux 1 hour, discards chloroform liquid, the dregs of a decoction volatilizes solvent, add water 0.5ml stirring moistenings, add the positive fourth of water saturation
Alcohol 10ml, it is ultrasonically treated 30 minutes, Aspirate supernatant adds 3 times of amount ammonia solutions, shakes up, places layering, take upper liquid to be evaporated, residue
Methanol 1.5ml is added to make dissolving, as need testing solution.
2. prepare negative sample solution：The finished product without ginseng, pseudo-ginseng is prepared by the anti-red preparation technology that is crazy about, it is finely ground, take
Powder 5g, add chloroform 40ml, be heated to reflux 1 hour, discard chloroform liquid, the dregs of a decoction volatilize solvent, add water 0.5ml to stir
Moistening, adding water-saturated n-butanol 10ml, be ultrasonically treated 30 minutes, Aspirate supernatant adds 3 times of amount ammonia solutions, shakes up, places layering,
Upper liquid is taken to be evaporated, residue adds methanol 1.5ml to make dissolving, as negative sample solution.
3. the preparation of reference substance solution：Take ginsenoside Rb1Reference substance, ginsenoside Re's reference substance, ginsenoside Rf couple
According to product, ginsenoside Rg1Reference substance and Panax Notoginseng saponin R1Reference substance adds methanol that every 1ml respectively mixed solutions containing 2mg are made, as
Mixed reference substance solution.
4. thin layer method selects
The selection of 4.1 development systems
Development system I：Chloroform-acetate-methanol-water（Volume ratio is 15:40:22:10,10 DEG C arranged below
Lower floor's solution）
Development system II：Chloroform-methanol-water（Volume ratio is 13:7:2,10 DEG C of lower floor's solution arranged below）
Development system III：N-butanol-ethyl acetate-water（Volume ratio is 4:1:5,10 DEG C of upper solutions arranged below）
Development system IV：Dichloromethane：Absolute ethyl alcohol：Water（Volume ratio is 90：45：6.5）
It is each to draw above-mentioned μ l of reference substance solution 1, need testing solution a, need testing solution b, need testing solution c and negative sample
0.5 μ l, put respectively on same silica gel g thin-layer plate, be respectively solvent with development system I, II, III and IV, deploy, take out,
Dry, spray with 10% ethanol solution of sulfuric acid, it is clear to be heated to spot development at 105 DEG C, puts daylight and ultraviolet lamp respectively
Inspected under (365nm).Test result is shown in the thin-layer chromatogram that Fig. 1-1-1 deploy to Fig. 1-1-4 differences development system, is specially：
The thin-layer chromatogram that Fig. 1-1-1 development systems I deploy；The thin-layer chromatogram that Fig. 1-1-2 development systems II deploy；Fig. 1-1-3 exhibitions
The thin-layer chromatogram that open system III is deployed；The thin-layer chromatogram that Fig. 1-1-4 development systems IV deploy, wherein, 1 is mixing reference substance
1μl；2 be the μ l of test sample point sample 0.5；3 be the μ l of test sample point sample 0.5；4 be the μ l of test sample point sample 0.5；5 be the μ of negative sample 0.5
l。
Conclusion：Through analyzing knowable to the thin-layer chromatogram of four kinds of different development systems, the thin-layer chromatography of development system I, II, III
Figure can only show four spots, illustrate that this three kinds of systems can not separate five kinds of reference substances, and development system IV can be right by five kinds
Separated according to product, and development system IV compares development system I, II, III, and collocation method is easier, system agents useful for same toxicity compared with
Development system that is low, therefore selecting development system IV to differentiate as this product thin layer.
4.2 point sample modes select
Notoginsenoside R and the spot of ginsenoside Re are closer to the distance in the thin-layer chromatogram deployed due to development system IV,
Separating degree is poor, therefore investigates spot application and band application, to make this two kinds of compositions preferably separate.Specific test result is shown in
Fig. 1-2-1 and Fig. 1-2-2, the thin-layer chromatogram deployed after the sample loading mode point sample of difference, it is specially：Fig. 1-2-1 is spot application
The thin-layer chromatogram of expansion；Fig. 1-2-2 is the thin-layer chromatogram of band application expansion, wherein, 1 is mixing reference substance 1 μ l；2 are
The μ l of test sample point sample 0.3；3 be the μ l of test sample point sample 0.3；4 be the μ l of test sample point sample 0.3；5 be the μ of negative sample 0.3.
Conclusion：It is visible to compare the thin-layer chromatogram deployed after the sample loading mode point sample of difference, the thin layer deployed after band application
Chromatogram spot is apparent, and separating degree is preferable, therefore selects band application.
4.3 point sample amounts are investigated
According to selected thin-layer identification method, take reference substance solution 1 μ l, μ l of need testing solution 0.2,0.3 μ l, 0.4 μ l,
0.5 μ l, put on same silica gel g thin-layer plate, deploy respectively, take out, dry, spray with 10% ethanol solution of sulfuric acid, 105 DEG C of heating
It is clear to spot development, put daylight and ultraviolet lamp（365nm）Under inspect.Test result is shown in Fig. 1-3 need testing solution point sample amounts
Thin layer differentiate chromatogram, wherein, 1 is mixing reference substance 1 μ l；2 be the μ l of test sample point sample 0.2；3 be the μ l of test sample point sample 0.3；
4 be the μ l of test sample point sample 0.4；5 be the μ l of test sample point sample 0.5.
Conclusion：The thin-layer chromatogram of analysis need testing solution difference point sample amount can obtain, during 0.3 μ l of point sample principal spot colour developing compared with
Clearly, and separating degree is preferable, therefore the point sample amount of determination this product need testing solution is 0.3 μ l.
5. durability is investigated
The investigation of 5.1 different lamellaes
The silica gel g thin-layer plate of four different manufacturers is chosen, takes the μ l of mixed reference substance solution 1 respectively, and need testing solution a,
Need testing solution b, need testing solution c and each 0.3 μ l of negative sample solution, put on same silica gel g thin-layer plate, according to having selected
The expansion of thin-layer chromatography condition, take out, dry, spray with 10% ethanol solution of sulfuric acid, 105 DEG C to be heated to spot development clear, puts day
Light and ultraviolet lamp（365nm）Under inspect.The effect of different lamellae expansion is tested, Fig. 1-4-1 are that Tianjin Si Lida science and technology has
Limit company --- can clipboard；Fig. 1-4-2 are Tianjin Si Lida Science and Technology Ltd.s --- efficient plate；Fig. 1-4-3 are that Qingdao is abundant
Min Yuan silica gel chemical reagent work --- glass plate；Fig. 1-4-4 are subsidiary factory of Haiyang Chemical Plant, Qingdao --- glass plate.Wherein, 1：Mixing pair
According to the μ l 2 of product 1：The μ l of test sample point sample 0.3；3：The μ l of test sample point sample 0.3；4：The μ l of test sample point sample 0.3；5：The μ of negative sample 0.3
l。
Conclusion：The thin-layer chromatogram of com-parison and analysis difference lamellae expansion, the results showed that in selected thin-layer chromatography bar
Under part, different lamellaes can reach discriminating purpose.
5.2 temperature are investigated
Under conditions of relative humidity is fixed as 60%, changes the temperature of thin-layer developing environment respectively, take a little excellent thin layer
Plate, put in 5 DEG C and 35 DEG C of environment and deploy, take out, dry, spray with 10% ethanol solution of sulfuric acid, 105 DEG C to be heated to spot development clear
It is clear, put daylight and ultraviolet lamp（365nm）Under inspect.The thin-layer chromatogram deployed tested in different relative temperature environment, Fig. 1-
5-1 is the thin-layer chromatogram deployed in 35 DEG C of environment of temperature；Fig. 1-5-2 are the thin-layer chromatogram deployed in 30 DEG C of environment of temperature；
Fig. 1-5-3 are the thin-layer chromatogram deployed in 25 DEG C of environment of temperature；Fig. 1-5-4 are the thin-layer chromatography deployed in 20 DEG C of environment of temperature
Figure；Fig. 1-5-5 are the thin-layer chromatogram deployed in 5 DEG C of environment of temperature.Wherein, 1：Mix the μ l of reference substance 1；2：Test sample point sample
0.3μl；3：The μ l of test sample point sample 0.3；4：The μ l of test sample point sample 0.3；5：The μ l of negative sample point sample 0.3.
Conclusion：Each principal spot of thin-layer chromatogram deployed in 5 DEG C of environment is clear, and separating degree is preferable；In 35 DEG C of environment
The thin-layer chromatogram spot of middle expansion obscures, and separating degree is poor, shows that high temperature has a great influence to this product indentification by TLC.Separately
Investigate the feasibility of the discrimination method in 30 DEG C, 25 DEG C and 20 DEG C environment of room temperature, it is seen that 30 DEG C, the thin layer deployed in 25 DEG C of environment
Chromatogram spot is still unintelligible, and separating degree is poor, can reach discriminating purpose at 20 DEG C, therefore the thin-layer identification method is 5 DEG C~20
DEG C good tolerance.
5.3 humidity are investigated
Under conditions of temperature is fixed as 5 DEG C, changes the relative humidity of thin-layer developing environment respectively, take a little excellent thin layer
Plate, put in the environment that relative humidity is 40% and 88% and deploy respectively, take out, dry, spray with 10% ethanol solution of sulfuric acid, 105 DEG C add
Heat is clear to spot development, puts daylight and ultraviolet lamp（365nm）Under inspect.Determine deploy in different relative humidity conditions it is thin
Layer chromatography figure, Fig. 1-6-1 are the thin-layer chromatogram deployed in the humidity environment of relative humidity 40%；Fig. 1-6-2 are relative humidity
The thin-layer chromatogram deployed in 88% humidity environment.Wherein, 1：Mix the μ l of reference substance 1；2：The μ l of test sample point sample 0.3；3：For examination
The μ l of product point sample 0.3；4：The μ l of test sample point sample 0.3；5：The μ l of negative sample point sample 0.3.
Conclusion：Thin-layer developing principal spot in the environment of relative humidity 40% and 88% is clear, and separating degree is good, therefore the method is in phase
To good tolerance in the environment of humidity 40%~88%.
6. specificity is investigated
Take μ l of mixed reference substance solution 1 described in step 3, the need testing solution a described in step 1, need testing solution b, confession
Each 0.3 μ l of the test sample solution c and μ l of negative sample solution 0.3 described in step 2, put respectively on same silica gel g thin-layer plate, with body
Product ratio 90：45：6.5 dichloromethane-absolute ethyl alcohol-water be solvent, solvent is placed in chromatography cylinder, in 5 DEG C of temperature, relative
Saturation 20 minutes in the environment of humidity 40%, lamellae is then placed in, after continuing presaturation 30 minutes, deploys, take out, dry, spray
It is clear to spot development in 105 DEG C of heating 3 minutes with 10% ethanol solution of sulfuric acid, respectively in daylight and ultraviolet light（365nm）Under
Inspect, observation feminine gender has noiseless.It is specifically shown in figure Fig. 1-7-1, the thin-layer chromatogram that specificity is investigated, wherein, 1：Mixing control
The μ l of product 1；2：The μ l of test sample point sample 0.3；3：The μ l of test sample point sample 0.3；4：The μ l of test sample point sample 0.3；5：Negative sample point sample 0.3
μl。
Conclusion：Without ginseng, pseudo-ginseng test sample on the corresponding position of saponins reference substance spot it is noiseless, show this
The specificity of thin-layer identification method is good.
In summary experimental result, while differentiate that ginseng in the Chinese medicinal composition preparation, the detection method of pseudo-ginseng are as follows：Take
The need testing solution a described in μ l of mixed reference substance solution 1, step 1, need testing solution b, need testing solution c described in step 3 is each
The 0.3 μ l and μ l of negative sample solution 0.3 described in step 2, put respectively on same silica gel g thin-layer plate, with volume ratio 90：45：
6.5 dichloromethane-absolute ethyl alcohol-water is solvent, and solvent is placed in chromatography cylinder, in 5 DEG C -20 DEG C of temperature, relative humidity
Saturation 20 minutes in 40%-80% environment, lamellae is then placed in, after continuing presaturation 30 minutes, deploys, take out, dry, spray
It is clear to spot development in 105 DEG C of heating 2-3 minutes with 10% ethanol solution of sulfuric acid, respectively in daylight and ultraviolet light（365nm）
Under inspect.In the test sample chromatogram of the Chinese medicine composition, with reference substance ginsenoside Rb1, reference substance ginsenoside Re, right
According on the chromatogram relevant position of product notoginsenoside R, reference substance ginsenoside Rf and reference substance ginsenoside Rg1, show under daylight
The spot of same color, the fluorescence spot of same color is shown under ultraviolet light, illustrate the Chinese medicine composition simultaneously containing ginseng, three
Seven；Without ginseng, pseudo-ginseng negative sample solution chromatogram in, with reference substance chromatogram relevant position, under daylight not show spot,
Do not show fluorescence spot under ultraviolet light, illustrate that the detection method specificity is strong, it is negative noiseless.
Experimental example 2：Commercially available Kang Er heart-soothing capsules prepare ginseng, pseudo-ginseng in Kang Er heart-soothing capsule medicinal extract according to Chinese Pharmacopoeia prescription
Indentification by TLC
1. prepare need testing solution：Kang Er heart-soothing capsule medicinal extract is taken, it is finely ground, powder 5g is taken, adds chloroform 40ml, is heated
Backflow 1 hour, discards chloroform liquid, and the dregs of a decoction volatilize solvent, adds water 0.5ml stirring moistenings, adds water-saturated n-butanol 10ml, surpass
Sonication 30 minutes, Aspirate supernatant add 3 times of amount ammonia solutions, shake up, place layering, take upper liquid to be evaporated, residue adds methanol 2ml
Make dissolving, as need testing solution.
2. prepare negative sample solution：The finished product without ginseng, pseudo-ginseng is prepared by the preparation technology of Kang Er heart-soothing capsules, is ground
Carefully, powder 5g is taken, adds chloroform 40ml, is heated to reflux 1 hour, discards chloroform liquid, the dregs of a decoction volatilize solvent, add water
0.5ml stirring moistenings, add water-saturated n-butanol 10ml, are ultrasonically treated 30 minutes, and Aspirate supernatant adds 3 times of amount ammonia solutions, shakes up,
Layering is placed, takes upper liquid to be evaporated, residue adds methanol 2ml to make dissolving, as negative sample solution.
3. prepare reference substance solution：Take ginsenoside Rb1's reference substance, ginsenoside Re's reference substance, ginsenoside Rg1
Reference substance, ginsenoside Rf's control and notoginsenoside R reference substance, add methanol that every 1ml methanol is made respectively above-mentioned right containing 0.5mg
According to the mixed solution of product, as reference substance mixed solution.
4. point sample amount is investigated
Take the μ l of mixed reference substance solution 1 in step 3, μ l of need testing solution 0.2 in step 2,0.3 μ l, 0.4 μ l, 0.5 μ l,
0.6 μ l, put respectively on same silica gel g thin-layer plate, with volume ratio 90：45：6.5 dichloromethane -10 DEG C of absolute ethyl alcohol-water is
Solvent, solvent are placed in chromatography cylinder, in 5 DEG C of temperature, relative humidity 40% environment in saturation 20 minutes, be then placed in thin
Laminate, after continuing presaturation 30 minutes, deploy, take out, dry, spray with 10% ethanol solution of sulfuric acid, 3 minutes are heated at 105 DEG C extremely
Spot development is clear, respectively in daylight and ultraviolet light（365nm）Under inspect.See that Fig. 2-1 need testing solutions thin layer differentiates point sample amount
Investigation, wherein, 1：Mix the μ l of reference substance 1.5；2：The μ l of test sample point sample 0.2；3：The μ l of test sample point sample 0.3；4：Test sample point
The μ l of sample 0.4；5：The μ l of test sample point sample 0.5；6：The μ l of test sample point sample 0.6.
Conclusion：The thin-layer chromatogram of analysis need testing solution difference point sample amount can obtain, during 0.5 μ l of point sample principal spot colour developing compared with
Clearly, and separating degree is preferable, therefore the point sample amount of determination this product need testing solution is 0.5 μ l.
5. specificity is investigated
Take μ l of mixed reference substance solution 1.5 described in step 3, the need testing solution a described in step 1, need testing solution b,
Each 0.5 μ l of the need testing solution c and μ l of negative sample solution 0.5 described in step 2, put respectively on same silica gel g thin-layer plate, with
Volume ratio 90：45：6.5 dichloromethane-absolute ethyl alcohol-water is solvent, and solvent is placed in chromatography cylinder, in 5 DEG C of temperature, phase
To saturation in the environment of humidity 40% 20 minutes, lamellae is then placed in, after continuing presaturation 30 minutes, is deployed, is taken out, dry,
Spray is clear to spot development in 105 DEG C of heating 3 minutes with 10% ethanol solution of sulfuric acid, respectively in daylight and ultraviolet light（365nm）
Under inspect, observation feminine gender have it is noiseless.See the thin-layer chromatogram that Fig. 2-2 specificities are investigated, wherein, 1：Mix the μ l of reference substance 1.5；
2：The μ l of 1 point sample of test sample 0.5；3：The μ l of 2 point sample of test sample 0.5；4：The μ l of 3 point sample of test sample 0.5；5：The μ l of negative sample point sample 0.5.
Conclusion：Without ginseng, pseudo-ginseng test sample on the corresponding position of saponins reference substance chromatogram spot noiseless, table
The specificity of the bright thin-layer identification method is good.
In summary experimental result, while differentiate that ginseng in Kang Er heart-soothing capsules, the detection method of pseudo-ginseng are as follows：Take step 3
Need testing solution a, need testing solution b, need testing solution c each 0.5 described in described μ l of mixed reference substance solution 1.5, step 1
The μ l and μ l of negative sample solution 0.5 described in step 2, put respectively on same silica gel g thin-layer plate, with volume ratio 90：45：6.5
Dichloromethane-absolute ethyl alcohol-water be solvent, solvent is placed in chromatography cylinder, in 5 DEG C -20 DEG C of temperature, relative humidity 40%-
Saturation 20 minutes in 80% environment, are then placed in lamellae, after continuing presaturation 30 minutes, deploy, take out, dry, spray with
10% ethanol solution of sulfuric acid, it is clear to spot development in 105 DEG C of heating 2-3 minutes, respectively in daylight and ultraviolet light（365nm）Under
Inspect.In the test sample chromatogram of Kang Er heart-soothing capsules, with reference substance ginsenoside Rb1, reference substance ginsenoside Re, reference substance
Show identical on the chromatogram relevant position of notoginsenoside R, reference substance ginsenoside Rf and reference substance ginsenoside Rg1, under daylight
The spot of color, the fluorescence spot of same color is shown under ultraviolet light, illustrate to contain ginseng, pseudo-ginseng in Kang Er heart-soothing capsules simultaneously；No
Containing ginseng, pseudo-ginseng negative sample solution chromatogram in, with reference substance chromatogram relevant position, under daylight not show spot, it is ultraviolet
Do not show fluorescence spot under light, illustrate that the detection method specificity is strong, it is negative noiseless.
Experimental example 3：Commercially available Yixinshu tablet or the thin layer color that ginseng in Yixinshu tablet medicinal extract is prepared according to Chinese Pharmacopoeia prescription
Spectrum differentiates
1. prepare need testing solution：Yixinshu tablet medicinal extract is taken, it is finely ground, powder 2g is taken, adds chloroform 40ml, heats back
Stream 1 hour, discards chloroform liquid, and the dregs of a decoction volatilize solvent, adds water 0.5ml stirring moistenings, adds water-saturated n-butanol 10ml, ultrasound
Processing 30 minutes, Aspirate supernatant add 3 times of amount ammonia solutions, shake up, place layering, take upper liquid to be evaporated, residue adds methanol 2ml to make
Dissolving, as need testing solution.
2. prepare negative sample solution：The finished product without ginseng is prepared by the preparation technology of Yixinshu tablet, it is finely ground, take powder
Last 2g, add chloroform 40ml, be heated to reflux 1 hour, discard chloroform liquid, the dregs of a decoction volatilize solvent, add water 0.5ml stirrings wet
Profit, add water-saturated n-butanol 10ml, be ultrasonically treated 30 minutes, Aspirate supernatant adds 3 times of amount ammonia solutions, shakes up, places layering, take
Upper liquid is evaporated, and residue adds methanol 2ml to make dissolving, as negative sample solution.
3. prepare reference substance solution：Take ginsenoside Rb1's reference substance, ginsenoside Re's reference substance, ginsenoside Rg1
Reference substance, ginsenoside Rf's control and notoginsenoside R reference substance, add methanol that every 1ml methanol is made respectively above-mentioned right containing 0.5mg
According to the mixed solution of product, as reference substance mixed solution.
4. point sample amount is investigated
Take the μ l of mixed reference substance solution 2 in step 3, μ l of need testing solution 1,2 μ l, 3 μ l, 4 μ l, 5 μ l in step 2, respectively
Point is on same silica gel g thin-layer plate, with volume ratio 90：45：6.5 dichloromethane-absolute ethyl alcohol-water is solvent, solvent
Be placed in chromatography cylinder, in 5 DEG C of temperature, relative humidity 40% environment in saturation 20 minutes, be then placed in lamellae, continue pre- full
After 30 minutes, deploy, take out, dry, spray with 10% ethanol solution of sulfuric acid, 105 DEG C heating 3 minutes it is clear to spot development,
Respectively in daylight and ultraviolet light（365nm）Under inspect.See that Fig. 3-1 need testing solutions thin layer differentiates the investigation of point sample amount, wherein,
1：Mix the μ l of reference substance 2；2：The μ l of test sample point sample 1；3：The μ l of test sample point sample 2；4：The μ l of test sample point sample 3；5：Test sample point sample 4
μl；6：The μ l of test sample point sample 5.
Conclusion：The thin-layer chromatogram of analysis need testing solution difference point sample amount can obtain, and principal spot colour developing is more clear during 3 μ l of point sample
It is clear, and separating degree is preferable, therefore the point sample amount of determination this product need testing solution is 3 μ l.
5. specificity is investigated
Take μ l of mixed reference substance solution 2 described in step 3, the need testing solution a described in step 1, need testing solution b, confession
Each 3 μ l of the test sample solution c and μ l of negative sample solution 3 described in step 2, put respectively on same silica gel g thin-layer plate, with volume ratio
90：45：6.5 dichloromethane-absolute ethyl alcohol-water is solvent, and solvent is placed in chromatography cylinder, in 5 DEG C of temperature, relative humidity
Saturation 20 minutes in 40% environment, are then placed in lamellae, after continuing presaturation 30 minutes, deploy, take out, dry, spray with
10% ethanol solution of sulfuric acid, 105 DEG C heating 3 minutes it is clear to spot development, respectively in daylight and ultraviolet light（365nm）Lower inspection
Depending on observation feminine gender has noiseless.See the thin-layer chromatogram that Fig. 4 specificities are investigated, wherein, 1：Mix the μ l of reference substance 2；2：Test sample
The μ l of 1 point sample 3；3：The μ l of 2 point sample of test sample 3；4：The μ l of 3 point sample of test sample 3； 5：The μ l of negative sample point sample 3.
Conclusion：Test sample without ginseng is noiseless on the corresponding position of saponins reference substance chromatogram spot, shows this
The specificity of thin-layer identification method is good.
In summary experimental result, differentiate that the detection method of ginseng in Yixinshu tablet is as follows：Take the mixing pair described in step 3
According to described in the need testing solution a described in μ l of product solution 2, step 1, need testing solution b, each 3 μ l of need testing solution c and step 2
The μ l of negative sample solution 3, put respectively on same silica gel g thin-layer plate, with volume ratio 90：45：6.5 dichloromethane-anhydrous second
Alcohol-water is solvent, and solvent is placed in chromatography cylinder, in 5 DEG C -20 DEG C of temperature, relative humidity 40%-80% environment in saturation 20
Minute, lamellae is then placed in, after continuing presaturation 30 minutes, is deployed, is taken out, dry, spray with 10% ethanol solution of sulfuric acid,
105 DEG C of heating 2-3 minutes are clear to spot development, respectively in daylight and ultraviolet light（365nm）Under inspect.Yixinshu tablet supplies examination
In product chromatogram, with reference substance ginsenoside Rb1, reference substance ginsenoside Re, reference substance ginsenoside Rf, reference substance ginseng
On saponin(e Rg1 chromatogram relevant position, the spot of same color is shown under daylight, the fluorescence spot of same color is shown under ultraviolet light,
On the chromatogram relevant position with reference substance notoginsenoside R, the spot without same color under daylight, without identical face under ultraviolet light
The fluorescence spot of color, illustrate to contain ginseng in Yixinshu tablet, without pseudo-ginseng, and in the absence of the phenomenon for substituting ginseng with pseudo-ginseng；
In negative sample solution chromatogram without ginseng, with reference substance chromatogram relevant position, spot not being shown under daylight, under ultraviolet light
Do not show fluorescence spot, illustrate that the detection method specificity is strong, it is negative noiseless.
Experimental example 4：Commercially available sheep leaves of pulse plants notoginseng capsule prepares pseudo-ginseng in sheep leaves of pulse plants notoginseng capsule medicinal extract according to Chinese Pharmacopoeia prescription
Indentification by TLC
1. prepare need testing solution：Sheep leaves of pulse plants notoginseng capsule medicinal extract is taken, it is finely ground, powder 2g is taken, adds chloroform 40ml, adds
Heat backflow 1 hour, discards chloroform liquid, and the dregs of a decoction volatilize solvent, adds water 0.5ml stirring moistenings, adds water-saturated n-butanol 10ml,
It is ultrasonically treated 30 minutes, Aspirate supernatant adds 3 times of amount ammonia solutions, shakes up, places layering, take upper liquid to be evaporated, residue adds methanol
2ml makes dissolving, as need testing solution.
2. prepare negative sample solution：The finished product without pseudo-ginseng is prepared by the preparation technology of sheep leaves of pulse plants notoginseng capsule, it is finely ground,
Powder 2g is taken, adds chloroform 40ml, is heated to reflux 1 hour, discards chloroform liquid, the dregs of a decoction volatilize solvent, add water 0.5ml to stir
Moistening is mixed, adds water-saturated n-butanol 10ml, is ultrasonically treated 30 minutes, Aspirate supernatant adds 3 times of amount ammonia solutions, shakes up, places and divide
Layer, takes upper liquid to be evaporated, residue adds methanol 2ml to make dissolving, as negative sample solution.
3. prepare reference substance solution：Take ginsenoside Rb1's reference substance, ginsenoside Re's reference substance, ginsenoside Rg1
Reference substance, ginsenoside Rf's control and notoginsenoside R reference substance, add methanol that the respectively above-mentioned control containing 1mg of every 1ml methanol is made
The mixed solution of product, as reference substance mixed solution.
4. point sample amount is investigated
Take the μ l of mixed reference substance solution 1 in step 3, μ l of need testing solution 0.2 in step 2,0.3 μ l, 0.4 μ l, 0.5 μ l,
0.6 μ l, put respectively on same silica gel g thin-layer plate, with volume ratio 90：45：6.5 dichloromethane-absolute ethyl alcohol-water is expansion
Agent, solvent are placed in chromatography cylinder, in 5 DEG C of temperature, relative humidity 40% environment in saturation 20 minutes, be then placed in lamellae,
After continuing presaturation 30 minutes, deploy, take out, dry, spray with 10% ethanol solution of sulfuric acid, 3 minutes are heated at 105 DEG C to spot
Colour developing is clear, respectively in daylight and ultraviolet light（365nm）Under inspect.See that Fig. 4-1 need testing solutions thin layer differentiates examining for point sample amount
Examine, wherein, 1：Mix the μ l of reference substance 1；2：The μ l of test sample point sample 0.2；3：The μ l of test sample point sample 0.3；4：The μ of test sample point sample 0.4
l；5：The μ l of test sample point sample 0.5；6：The μ l of test sample point sample 0.6.
Conclusion：The thin-layer chromatogram of analysis need testing solution difference point sample amount can obtain, during 0.3 μ l of point sample principal spot colour developing compared with
Clearly, and separating degree is preferable, therefore the point sample amount of determination this product need testing solution is 0.3 μ l.
5. specificity is investigated
Take μ l of mixed reference substance solution 1 described in step 3, the need testing solution a described in step 1, need testing solution b, confession
Each 0.3 μ l of the test sample solution c and μ l of negative sample solution 0.3 described in step 2, put respectively on same silica gel g thin-layer plate, with body
Product ratio 90：45：6.5 dichloromethane -10 DEG C of absolute ethyl alcohol-water is solvent, and solvent is placed in chromatography cylinder, in 5 DEG C of temperature,
Saturation 20 minutes in the environment of relative humidity 40%, lamellae is then placed in, after continuing presaturation 30 minutes, deploys, take out, dry in the air
It is dry, spray with 10% ethanol solution of sulfuric acid, it is clear to spot development in 105 DEG C of heating 3 minutes, respectively in daylight and ultraviolet light
（365nm）Under inspect, observation feminine gender have it is noiseless.See Fig. 4-2, the thin-layer chromatogram that specificity is investigated, wherein, 1：Mixing control
The μ l of product 1；2：The μ l of 1 point sample of test sample 0.3；3：The μ l of 2 point sample of test sample 0.3；4：The μ l of 3 point sample of test sample 0.3；5：Negative sample point sample
0.3μl。
Conclusion：Without ginseng, pseudo-ginseng test sample on the corresponding position of saponins reference substance chromatogram spot noiseless, table
The specificity of the bright thin-layer identification method is good.
In summary experimental result, differentiate that the detection method of pseudo-ginseng in sheep leaves of pulse plants notoginseng capsule is as follows：Take mixed described in step 3
Need testing solution a, need testing solution b, each 0.3 μ l of need testing solution c and step 2 described in conjunction reference substance solution 1 μ l, step 1
The described μ l of negative sample solution 0.3, put respectively on same silica gel g thin-layer plate, with volume ratio 90：45：6.5 dichloromethane
Alkane-absolute ethyl alcohol-water is solvent, and solvent is placed in chromatography cylinder, in 5 DEG C -20 DEG C of temperature, relative humidity 40%-80% ring
Saturation 20 minutes in border, lamellae is then placed in, after continuing presaturation 30 minutes, deploys, take out, dry, spray with 10% sulfuric acid second
Alcoholic solution, it is clear to spot development in 105 DEG C of heating 2-3 minutes, respectively in daylight and ultraviolet light（365nm）Under inspect.The sheep leaves of pulse plants
In the test sample chromatogram of notoginseng capsule, with reference substance ginsenoside Rb1, reference substance ginsenoside Re, reference substance notoginsenoside
R1, reference substance ginsenoside Rg1 chromatogram relevant position on, the spot of same color is shown under daylight, identical face is shown under ultraviolet light
The fluorescence spot of color, on the chromatogram relevant position with reference substance ginsenoside Rf, the spot without same color, ultraviolet under daylight
Fluorescence spot without same color under light, illustrate to contain pseudo-ginseng in sheep leaves of pulse plants notoginseng capsule, without ginseng, and be not present and use ginseng
Substitute the phenomenon of pseudo-ginseng；In negative sample solution chromatogram without pseudo-ginseng, with reference substance chromatogram relevant position, under daylight not
Show spot, do not show fluorescence spot under ultraviolet light, illustrate that the detection method specificity is strong, it is negative noiseless.
The foregoing description of the disclosed embodiments, professional and technical personnel in the field are enable to realize or using the present invention.
A variety of modifications to these embodiments will be apparent for those skilled in the art, as defined herein
General Principle can be realized in other embodiments without departing from the spirit or scope of the present invention.Therefore, it is of the invention
The embodiments shown herein is not intended to be limited to, and is to fit to and principles disclosed herein and features of novelty phase one
The most wide scope caused.
Claims (1)
1. a kind of detection method of the Chinese medicine preparation containing ginseng or pseudo-ginseng, it is characterised in that comprise the following steps：
（1）Prepare need testing solution：Chinese medicine composition or Chinese medicine preparation containing ginseng or pseudo-ginseng are taken, removes coating, it is finely ground, take
Powder 2-5g, add chloroform 40ml, be heated to reflux 1 hour, discard chloroform liquid, the dregs of a decoction volatilize solvent, add water 0.5ml to stir
Moistening is mixed, adds water-saturated n-butanol 10ml, is ultrasonically treated 30 minutes, Aspirate supernatant adds 3 times of amount ammonia solutions, shakes up, places and divide
Layer, takes upper liquid to be evaporated, residue adds methanol 2-3ml to make dissolving, as need testing solution；
（2）Prepare negative sample solution：By the preparation technology of Chinese medicine composition or Chinese medicine preparation prepare without ginseng, pseudo-ginseng into
Product, it is finely ground, powder 2-5g is taken, adds chloroform 40ml, is heated to reflux 1 hour, discards chloroform liquid, the dregs of a decoction volatilize solvent,
Adding water 0.5ml stirring moistenings, add water-saturated n-butanol 10ml, be ultrasonically treated 30 minutes, Aspirate supernatant adds 3 times of amount ammonia solutions,
Shake up, place layering, take upper liquid to be evaporated, residue adds methanol 2-3ml to make dissolving, as negative sample solution；
（3）Prepare reference substance solution：Take ginsenoside Rb1's reference substance, ginsenoside Re's reference substance, ginsenoside Rg1's control
Product, ginsenoside Rf's control and notoginsenoside R reference substance, add methanol that the respectively above-mentioned control containing 0.5-2mg of every 1ml methanol is made
The mixed solution of product, as reference substance mixed solution；
（4）Detection：Take step（3）Described mixed reference substance solution 1-3 μ l, step（1）Described need testing solution 0.3-3 μ l
And step（2）Described negative sample solution 0.3-3 μ l, put respectively on same silica gel g thin-layer plate, with volume ratio 90：45：
6.5 dichloromethane-absolute ethyl alcohol-water is solvent, and solvent is placed in chromatography cylinder, in 5 DEG C -20 DEG C of temperature, relative humidity
Saturation 20 minutes in 40%-80% environment, lamellae is then placed in, after continuing presaturation 30 minutes, deploys, take out, dry, spray
It is clear to spot development in 105 DEG C of heating 2-3 minutes with 10% ethanol solution of sulfuric acid, respectively under daylight and ultraviolet light 365nm
Inspect；
（5）Interpretation of result：Containing ginseng, the Chinese medicine preparation of pseudo-ginseng test sample chromatogram in, with reference substance ginsenoside Rb1,
Reference substance ginsenoside Re, reference substance notoginsenoside R, the chromatogram of reference substance ginsenoside Rf and reference substance ginsenoside Rg1
On relevant position, the spot of same color is shown under daylight, the fluorescence spot of same color is shown under ultraviolet light；Containing ginseng, be free of
In the test sample chromatogram of the Chinese medicine preparation of pseudo-ginseng, with reference substance ginsenoside Rb1, reference substance ginsenoside Re, reference substance people
On the chromatogram relevant position for joining saponin(e Rf and reference substance ginsenoside Rg1, show the spot of same color under daylight, under ultraviolet light
The fluorescence spot of aobvious same color；In the test sample chromatogram of Chinese medicine preparation containing pseudo-ginseng, without ginseng, reference substance ginsenoside
On Rb1, reference substance ginsenoside Re, reference substance notoginsenoside R and the chromatogram relevant position of reference substance ginsenoside Rg1, day
Show the spot of same color under light, the fluorescence spot of same color is shown under ultraviolet light；Negative sample solution without ginseng, pseudo-ginseng
In chromatogram, with reference substance chromatogram relevant position, not showing spot under daylight, do not show fluorescence spot under ultraviolet light.
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| CN110917313A (en) * | 2019-11-29 | 2020-03-27 | 长春中医药大学 | Ten-ingredient xianglu capsules, preparation method thereof and ginsenoside content analysis method |
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