CN115372535B - Thin layer chromatography detection method for two-day oil - Google Patents
Thin layer chromatography detection method for two-day oil Download PDFInfo
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- 238000004809 thin layer chromatography Methods 0.000 title claims abstract description 22
- 238000001514 detection method Methods 0.000 title claims abstract description 18
- 238000000034 method Methods 0.000 claims abstract description 19
- 239000003795 chemical substances by application Substances 0.000 claims abstract description 17
- 239000012488 sample solution Substances 0.000 claims abstract description 13
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims abstract description 12
- 239000013558 reference substance Substances 0.000 claims abstract description 12
- 238000010438 heat treatment Methods 0.000 claims abstract description 8
- HYGYSIDIKIGPJA-UHFFFAOYSA-N chloroform;ethyl acetate;methanol Chemical compound OC.ClC(Cl)Cl.CCOC(C)=O HYGYSIDIKIGPJA-UHFFFAOYSA-N 0.000 claims abstract description 7
- ROAYSRAUMPWBQX-UHFFFAOYSA-N ethanol;sulfuric acid Chemical compound CCO.OS(O)(=O)=O ROAYSRAUMPWBQX-UHFFFAOYSA-N 0.000 claims abstract description 7
- 238000005507 spraying Methods 0.000 claims abstract description 5
- 238000007605 air drying Methods 0.000 claims abstract description 4
- 238000007689 inspection Methods 0.000 claims abstract description 4
- 239000003921 oil Substances 0.000 claims description 48
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 claims description 21
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 21
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 18
- 239000000463 material Substances 0.000 claims description 15
- 238000002360 preparation method Methods 0.000 claims description 11
- 239000003208 petroleum Substances 0.000 claims description 9
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 claims description 8
- 239000000706 filtrate Substances 0.000 claims description 5
- 239000007788 liquid Substances 0.000 claims description 5
- 239000010495 camellia oil Substances 0.000 claims description 4
- 238000002156 mixing Methods 0.000 claims description 4
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 3
- 238000001914 filtration Methods 0.000 claims description 3
- 238000007670 refining Methods 0.000 claims description 3
- 238000002791 soaking Methods 0.000 claims description 3
- 238000001704 evaporation Methods 0.000 claims description 2
- 239000009286 sanguis draxonis Substances 0.000 claims description 2
- 239000009136 dragon's blood Substances 0.000 abstract description 37
- 239000003814 drug Substances 0.000 abstract description 17
- 238000003908 quality control method Methods 0.000 abstract description 9
- 238000004451 qualitative analysis Methods 0.000 abstract description 2
- 241001043298 Croton draco Species 0.000 abstract 2
- 235000019198 oils Nutrition 0.000 description 42
- 240000004824 Trimezia steyermarkii Species 0.000 description 33
- 238000012360 testing method Methods 0.000 description 31
- 239000000523 sample Substances 0.000 description 29
- 239000000243 solution Substances 0.000 description 28
- 239000013642 negative control Substances 0.000 description 20
- 229910052799 carbon Inorganic materials 0.000 description 12
- NOOLISFMXDJSKH-KXUCPTDWSA-N (-)-Menthol Chemical compound CC(C)[C@@H]1CC[C@@H](C)C[C@H]1O NOOLISFMXDJSKH-KXUCPTDWSA-N 0.000 description 11
- NOOLISFMXDJSKH-UHFFFAOYSA-N DL-menthol Natural products CC(C)C1CCC(C)CC1O NOOLISFMXDJSKH-UHFFFAOYSA-N 0.000 description 11
- 229940041616 menthol Drugs 0.000 description 10
- 239000000126 substance Substances 0.000 description 8
- 239000001525 mentha piperita l. herb oil Substances 0.000 description 6
- 235000019477 peppermint oil Nutrition 0.000 description 6
- 241000196324 Embryophyta Species 0.000 description 5
- 239000003153 chemical reaction reagent Substances 0.000 description 5
- 229940079593 drug Drugs 0.000 description 5
- DTGKSKDOIYIVQL-WEDXCCLWSA-N (+)-borneol Chemical compound C1C[C@@]2(C)[C@@H](O)C[C@@H]1C2(C)C DTGKSKDOIYIVQL-WEDXCCLWSA-N 0.000 description 4
- 238000000605 extraction Methods 0.000 description 4
- REPVLJRCJUVQFA-UHFFFAOYSA-N (-)-isopinocampheol Natural products C1C(O)C(C)C2C(C)(C)C1C2 REPVLJRCJUVQFA-UHFFFAOYSA-N 0.000 description 3
- 244000246386 Mentha pulegium Species 0.000 description 3
- 235000016257 Mentha pulegium Nutrition 0.000 description 3
- 235000004357 Mentha x piperita Nutrition 0.000 description 3
- YXFVVABEGXRONW-UHFFFAOYSA-N Toluene Chemical compound CC1=CC=CC=C1 YXFVVABEGXRONW-UHFFFAOYSA-N 0.000 description 3
- 229940116229 borneol Drugs 0.000 description 3
- CKDOCTFBFTVPSN-UHFFFAOYSA-N borneol Natural products C1CC2(C)C(C)CC1C2(C)C CKDOCTFBFTVPSN-UHFFFAOYSA-N 0.000 description 3
- DTGKSKDOIYIVQL-UHFFFAOYSA-N dl-isoborneol Natural products C1CC2(C)C(O)CC1C2(C)C DTGKSKDOIYIVQL-UHFFFAOYSA-N 0.000 description 3
- 239000000284 extract Substances 0.000 description 3
- 239000003205 fragrance Substances 0.000 description 3
- 235000001050 hortel pimenta Nutrition 0.000 description 3
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 2
- 235000003261 Artemisia vulgaris Nutrition 0.000 description 2
- 240000006891 Artemisia vulgaris Species 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- 240000001972 Gardenia jasminoides Species 0.000 description 2
- 235000006679 Mentha X verticillata Nutrition 0.000 description 2
- 235000002899 Mentha suaveolens Nutrition 0.000 description 2
- 235000001636 Mentha x rotundifolia Nutrition 0.000 description 2
- 208000002193 Pain Diseases 0.000 description 2
- 201000007100 Pharyngitis Diseases 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 2
- WORJEOGGNQDSOE-UHFFFAOYSA-N chloroform;methanol Chemical compound OC.ClC(Cl)Cl WORJEOGGNQDSOE-UHFFFAOYSA-N 0.000 description 2
- 230000000052 comparative effect Effects 0.000 description 2
- 239000013078 crystal Substances 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- DSSYKIVIOFKYAU-XCBNKYQSSA-N (R)-camphor Chemical compound C1C[C@@]2(C)C(=O)C[C@@H]1C2(C)C DSSYKIVIOFKYAU-XCBNKYQSSA-N 0.000 description 1
- 206010000087 Abdominal pain upper Diseases 0.000 description 1
- 241000208838 Asteraceae Species 0.000 description 1
- 240000000572 Blumea balsamifera Species 0.000 description 1
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 1
- 244000025254 Cannabis sativa Species 0.000 description 1
- 206010010071 Coma Diseases 0.000 description 1
- 206010051625 Conjunctival hyperaemia Diseases 0.000 description 1
- 206010010726 Conjunctival oedema Diseases 0.000 description 1
- 241000255925 Diptera Species 0.000 description 1
- 240000005636 Dryobalanops aromatica Species 0.000 description 1
- 206010019233 Headaches Diseases 0.000 description 1
- 206010019345 Heat stroke Diseases 0.000 description 1
- 241000218195 Lauraceae Species 0.000 description 1
- 206010068319 Oropharyngeal pain Diseases 0.000 description 1
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- 208000025865 Ulcer Diseases 0.000 description 1
- 239000002269 analeptic agent Substances 0.000 description 1
- 208000002399 aphthous stomatitis Diseases 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- RTEXIPZMMDUXMR-UHFFFAOYSA-N benzene;ethyl acetate Chemical compound CCOC(C)=O.C1=CC=CC=C1 RTEXIPZMMDUXMR-UHFFFAOYSA-N 0.000 description 1
- MDHYEMXUFSJLGV-UHFFFAOYSA-N beta-phenethyl acetate Natural products CC(=O)OCCC1=CC=CC=C1 MDHYEMXUFSJLGV-UHFFFAOYSA-N 0.000 description 1
- 235000013361 beverage Nutrition 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 210000004556 brain Anatomy 0.000 description 1
- 239000001110 calcium chloride Substances 0.000 description 1
- 229910001628 calcium chloride Inorganic materials 0.000 description 1
- 230000005465 channeling Effects 0.000 description 1
- 239000003086 colorant Substances 0.000 description 1
- 235000009508 confectionery Nutrition 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 208000002173 dizziness Diseases 0.000 description 1
- 150000002148 esters Chemical class 0.000 description 1
- 229930003935 flavonoid Natural products 0.000 description 1
- 150000002215 flavonoids Chemical class 0.000 description 1
- 235000017173 flavonoids Nutrition 0.000 description 1
- 239000000796 flavoring agent Substances 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 235000013355 food flavoring agent Nutrition 0.000 description 1
- 231100000869 headache Toxicity 0.000 description 1
- 238000003384 imaging method Methods 0.000 description 1
- 208000014674 injury Diseases 0.000 description 1
- 238000011835 investigation Methods 0.000 description 1
- 230000007803 itching Effects 0.000 description 1
- 230000007774 longterm Effects 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 210000004400 mucous membrane Anatomy 0.000 description 1
- 201000009240 nasopharyngitis Diseases 0.000 description 1
- 239000002674 ointment Substances 0.000 description 1
- 239000002304 perfume Substances 0.000 description 1
- 230000008092 positive effect Effects 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 238000001953 recrystallisation Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 235000013599 spices Nutrition 0.000 description 1
- 238000001256 steam distillation Methods 0.000 description 1
- 230000008961 swelling Effects 0.000 description 1
- -1 terpenoid organic compound Chemical class 0.000 description 1
- 239000012085 test solution Substances 0.000 description 1
- 229940034610 toothpaste Drugs 0.000 description 1
- 239000000606 toothpaste Substances 0.000 description 1
- 230000008736 traumatic injury Effects 0.000 description 1
- 231100000397 ulcer Toxicity 0.000 description 1
- MWOOGOJBHIARFG-UHFFFAOYSA-N vanillin Chemical compound COC1=CC(C=O)=CC=C1O MWOOGOJBHIARFG-UHFFFAOYSA-N 0.000 description 1
- FGQOOHJZONJGDT-UHFFFAOYSA-N vanillin Natural products COC1=CC(O)=CC(C=O)=C1 FGQOOHJZONJGDT-UHFFFAOYSA-N 0.000 description 1
- 235000012141 vanillin Nutrition 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/90—Plate chromatography, e.g. thin layer or paper chromatography
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/90—Plate chromatography, e.g. thin layer or paper chromatography
- G01N30/94—Development
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
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- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Medicines Containing Plant Substances (AREA)
Abstract
The invention relates to the technical field of medicine quality control, and particularly discloses a thin-layer chromatography detection method of two-day oil. The method of the invention comprises the following steps: respectively dispensing the two-day oil sample solution and the reference substance solution on the same silica gel G plate, developing with chloroform-ethyl acetate-methanol with volume ratio of 5-10:1-5:1 as developing agent, taking out, air drying, spraying 5-15% sulfuric acid ethanol solution as color developing agent, heating until spots develop clearly, and placing under ultraviolet light 365nm for inspection. The method can perform qualitative analysis on the content of the dragon's blood in the second-day oil, thereby performing quality control on the dragon's blood in the second-day oil and further effectively standardizing market competition.
Description
Technical Field
The invention belongs to the technical field of medicine quality control, and particularly relates to a thin-layer chromatography detection method of two-day oil.
Background
The second day oil, including menthol, peppermint oil and borneol, is a brownish red clear liquid with peppermint fragrance, is a wind-dispelling and exciting medicine, and can be used for treating common cold, dizziness, heatstroke and stomachache, etc. Wherein, the mint isThe oil is a light grass green liquid or a light yellow clear liquid, a large amount of colorless crystals are separated out at a lower temperature, the oil is strong in mint fragrance and cool and slightly bitter, is spicy and cool, has strong channeling performance, is gradually deepened in color after long-term storage, is gradually sticky in quality, can be mixed with ethanol, chloroform or diethyl ether at will, and is often used for expelling mosquitoes and relieving physical fatigue. Menthol, also known as menthol, is a terpenoid organic compound having the formula C 10 H 20 O, menthol is extracted from leaves and stems of peppermint, is colorless needle-like or prismatic crystal or white crystalline powder, has special fragrance of peppermint, is cool after initial burning, generally has two isomers (D type and L type), natural menthol is mainly L-isomer (L-menthol), menthol can be used as a flavoring agent of toothpaste, perfume, beverage, candy and the like, is used as a stimulating agent in medicine, acts on skin or mucous membrane, has cool and itching relieving effect, can be taken orally as a wind-dispelling medicine, is used for headache, nose, throat inflammation and the like, and ester thereof can also be used for spices and medicines. Borneol, also called flake brain, plum ice, etc., is prepared from stems and leaves of blumea balsamifera of Compositae or branches and leaves of Cinnamomum camphora of Lauraceae by steam distillation and recrystallization, and has chemical composition of 2-alcohol and chemical formula of C 10 H 18 O, it can be used for treating coma, conjunctival congestion, swelling and pain, sore throat, aphtha, sore pain, and unhealed ulcer. In addition, the secondary oil also comprises black oil, and the different colors of the secondary oil indicate that the addition amount of the black oil is obviously different. The black oil is refined by dragon's blood, gardenia carbon and mugwort leaf carbon in tea oil, and the consumption of the black oil in the second-day oil is large and the black oil contains the dragon's blood which is a rare Chinese medicinal material, so that the quality control of the dragon's blood in the second-day oil is necessary, thereby effectively standardizing market competition.
Patent CN02129687.1 provides a quality control method of a resina Draconis total flavonoid preparation, which comprises the following steps: taking the sample solution and the reference substance solution, spotting on a silica gel G thin layer plate, developing with 13-16:1 chloroform-methanol as developing agent, and inspecting under ultraviolet lamp. Patent CN200810049281.4 discloses a quality control method of a shaolin rheumatism traumatic injury ointment, wherein the quality control method of dragon's blood specifically comprises the following steps: taking the sample solution and the reference substance solution, spotting on a silica gel G thin layer plate, developing with 95:5 chloroform-methanol as developing agent, and inspecting under ultraviolet lamp.
Currently, there is no method for quality control of resina Draconis in the world oil.
Disclosure of Invention
In order to solve the above problems in the prior art, the present application provides a thin layer chromatography detection method for two-day oil.
In order to achieve the above object, the present invention provides the following technical solutions:
in one aspect, the invention provides a thin layer chromatography detection method of the two-day oil, which uses chloroform-ethyl acetate-methanol as a developing agent and uses a silica gel G plate as a thin layer chromatography plate to carry out the thin layer chromatography detection of the two-day oil.
Specifically, the developing agent is chloroform-ethyl acetate-methanol solution with the volume ratio of 5-10:1-5:1.
Further specifically, the volume ratio of the developing agent is 6-10:1-3:1, preferably 8:2:1.
Specifically, the method uses sulfuric acid ethanol solution as a color developing agent.
Further specifically, the concentration of the sulfuric acid ethanol solution is 5-15%, preferably 10%.
Specifically, the method comprises the following steps: respectively dispensing the two-day oil sample solution and the reference substance solution on the same silica gel G plate, developing with chloroform-ethyl acetate-methanol with volume ratio of 5-10:1-5:1 as developing agent, taking out, air drying, spraying 5-15% sulfuric acid ethanol solution as color developing agent, heating until spots develop clearly, and placing under ultraviolet light 365nm for inspection.
Specifically, the preparation method of the sample solution comprises the following steps: adding sodium hydroxide solution into the second-day oil according to the volume ratio of the second-day oil to the 2% sodium hydroxide solution of 1:5, shaking and extracting, adjusting the pH of the extracting solution to 3-5 by using dilute hydrochloric acid, shaking and extracting by using chloroform with the same volume as the 2% sodium hydroxide solution, evaporating the extracting solution to dryness, and dissolving residues in ethyl acetate solution with the volume of the second-day oil to be used as a sample solution.
Specifically, the preparation method of the reference substance solution comprises the following steps: adding tea oil into sanguis Draxonis reference medicinal materials at a feed-liquid ratio of 1:10, mixing, standing, soaking for 36-60 hr, heating to 180-220deg.C, refining for 3-5 hr, filtering to obtain filtrate, and mixing with the filtrate: petroleum ether is added to dissolve according to the volume ratio of petroleum ether being 1:19, and the petroleum ether is used as a reference substance solution.
In some embodiments, the thin layer chromatography detection method of the invention can be used for detecting and identifying the content of the dragon's blood in the second day oil, thereby controlling the quality of the dragon's blood in the second day oil.
In some embodiments, for quality detection of the second day oil, the quality detection and identification of menthol and peppermint oil in the second day oil are also required, and the method mainly comprises the following steps: (1) 0.5mL of the two-day oil was diluted with ethyl acetate to 5mL to prepare a sample solution. (2) And adding ethyl acetate into menthol and peppermint oil reference substances to prepare solutions containing 0.01mg of menthol and peppermint oil reference substances per 1mL, respectively, to obtain reference substance solutions. (3) According to thin layer chromatography (rule 0502 of Chinese pharmacopoeia 2020 edition), sucking 1-2 μl of each of the above three solutions, respectively spotting on the same silica gel G thin layer plate, spreading with benzene-ethyl acetate (19:1) as developing agent, taking out, air drying, spraying 10% vanillin sulfuric acid solution, and heating until the spots develop clearly.
Compared with the prior art, the invention has the following positive and beneficial effects:
the method can perform qualitative analysis on the dragon's blood in the second-day oil, thereby performing quality control on the dragon's blood in the second-day oil, and further effectively standardizing market competition.
Drawings
FIG. 1 is a graph showing the results of two-day oil thin layer chromatography (T: 24.2 ℃ C., RH: 52.2%) in example 1, wherein 1 is a dragon's blood negative control, 2 is a dragon's blood control drug, 3 is a sample SC80008,4 is a sample SC80017,5 is a sample SC80019,6 is a sample SC80020.
FIG. 2 is a graph showing the results of two-day oil thin layer chromatography using a Merck thin layer plate in Germany (T: 24.6 ℃, RH: 69.2%), wherein 1. Dragon's blood negative control, 2. Dragon's blood control, 3. Test substance SC80008,4. Test substance SC80017,5. Test substance SC80019,6. Test substance SC80020.
Fig. 3 is a graph of the results of two-day oil thin layer chromatography using a Qingdao ocean thin layer plate (T: 24.6 ℃, RH: 69.2%), wherein 1. Dragon blood negative control, 2. Dragon blood control medicinal material, 3. Test article SC80008,4. Test article SC80017,5. Test article SC80019,6. Test article SC80020.
FIG. 4 is a graph showing the results of two-day oil thin layer chromatography under the conditions of T25.1 ℃ and RH 33.6%, wherein 1 is a dragon's blood negative control, 2 is a dragon's blood control medicine, 3 is a test SC80008,4 is a test SC80017,5 is a test SC80019, and 6 is a test SC80020.
FIG. 5 is a graph showing the results of two-day oil thin layer chromatography under the conditions of T24.4 ℃ and RH 87.7%, wherein 1 is a dragon's blood negative control, 2 is a dragon's blood control medicine, 3 is a test sample SC80008,4 is a test sample SC80017,5 is a test sample SC80019,6 is a test sample SC80020.
FIG. 6 is a graph showing the results of two-day oil thin layer chromatography under the conditions of T8.9 ℃ and RH 70.7%, wherein 1 is a dragon's blood negative control, 2 is a dragon's blood control medicine, 3 is a test sample SC80008,4 is a test sample SC80017,5 is a test sample SC80019,6 is a test sample SC80020.
FIG. 7 is a graph showing the results of two-day oil thin layer chromatography under the conditions of T37.1 ℃ and RH 44.6%, wherein 1 is a dragon's blood negative control, 2 is a dragon's blood control medicine, 3 is a test SC80008,4 is a test SC80017,5 is a test SC80019,6 is a test SC80020.
FIG. 8 is a graph showing the results of two-day oil thin layer chromatography under the conditions of T21.6deg.C and RH 64.7%, wherein 1 is a dragon's blood negative control, 2 is a dragon's blood control drug, 3 is a sample SC80008,4 is a sample SC80017,5 is a sample SC80019,6 is a sample SC80020.
FIG. 9 shows the results of partial sample detection (T: 21.6deg.C, RH: 64.7%) in Experimental example 1, wherein 1. Star group SC80021,2. Star group SC80022,3. Star group SC80023,4. Star group SC80024,5. Star group SC80025,6. Dragon's blood negative control, 7. Dragon's blood control, 8. Star group SC80026,9. Star group RC80014, 10. Star group SC80007, 11. Star group PC80009, 12. Star group PC80014.
FIG. 10 shows the results of partial sample detection (T: 21.5 ℃ C., RH: 64.7%) in Experimental example 1, wherein 1.Star group PC80016, 2.Star group RC80001, 3.Star group RC80007, 4.other company X2002102,5, other company X2109016, dragon's blood negative control, 7.dragon's blood control, 8.other company X2010110,9, other company X210401, 10.other company Y201201, 11.other company Y200701, 12.other company Y200301.
FIG. 11 shows the results of partial sample detection (T: 21.6deg.C, RH: 65.3%) in Experimental example 1, wherein 1.Star group SC80004, 2.Star group QC80014, 3.Star group QC80021, 4.other company X210301, 5.other company X210802,6, dragon's blood negative control, 7.Dragon's blood control, 8.other company X211001,9, other company X21080110, other company X210902, 11.other company X2009108, 12.other company X1911109.
FIG. 12 shows the results of partial sample detection (T: 21.6deg.C, RH: 65.3%) in Experimental example 1, wherein 1, star group QC80007,2, star group SC80012,3, star group SC80013,4, other company X210101,5, other company X2005105,6, dragon's blood negative control, 7, dragon's blood control, 8, other company X210701,9, other company X21020110, other company X1906105, 11, other company X2012112, 12, other company X210402.
Detailed Description
The present invention will be described in further detail with reference to the following examples, which are not intended to limit the present invention, but are merely illustrative of the present invention. The experimental methods used in the following examples are not specifically described, but the experimental methods in which specific conditions are not specified in the examples are generally carried out under conventional conditions, and the materials, reagents, etc. used in the following examples are commercially available unless otherwise specified.
Experimental materials
1. Experimental reagent: silica gel G precast slab (Tsingtao ocean chemical plant, lot number: 20210406) (Tsingtao ocean chemical plant, lot number: 20210202), silica gel G precast slab (Merck, germany, lot number: HX 14868326), petroleum ether (60-90 ℃) (analytical grade, guangzhou chemical reagent plant, lot number: 20200803 20), toluene (analytical grade, guangzhou chemical reagent plant, lot number: 20190101 08), acetone (analytical grade, guangzhou chemical reagent plant, lot number: 20210301 05).
2. Experimental instrument: ME5002T/02 type electronic balance (Metrele-Tolyx (Shanghai)), BP-221D type electronic balance (Sidolis, germany) CAMAG reprostar3 digital imaging system (Kma, switzerland).
3. Control medicinal materials: the resina Draconis control medicinal materials (batch numbers: 120906-201611), fructus Gardeniae control medicinal materials (batch numbers: 120906-201611), folium Artemisiae Argyi control medicinal materials (batch numbers: 121345-201804), mentholum control (batch numbers: 110728-201707), peppermint oil control extract (batch numbers: 111551-201704), and Borneolum Syntheticum control medicinal materials (batch numbers: 110743-2016706) are all purchased from Chinese food and drug verification institute.
4. Test article:
a total of 44 batches of two day oil samples were collected, of which 23 were available from Guangzhou white cloud Star group (pharmaceutical industry), X18 from other companies and Y3 from other companies. The batch list is shown in Table 1, where ∈ numbers 1-4 are the recipe study use batches.
TABLE 1 two day oil batch list and corresponding manufacturer
5. Resina Draconis negative control: according to the standard prescription proportion of the product, the medicinal materials except the dragon's blood, namely gardenia, mugwort leaf, peppermint oil, menthol and borneol are taken, and a dragon's blood negative control sample is prepared according to the standard process preparation method.
Example 1A two-day oil thin layer chromatography detection method
The extraction solvent and the extraction method of the sample solution preparation method are inspected, the sample solution preparation method is preferred, the developing agent, the inspection method, the sample application amount and other chromatographic conditions of the developing condition are inspected, and the finally determined thin layer identification method is as follows:
preparation of test solution: 2mL of the product is taken, 10mL of 2% sodium hydroxide solution is used for shaking extraction, the pH of the extract is adjusted to 3-5 by dilute hydrochloric acid, 10mL of chloroform is used for shaking extraction, the extract is evaporated to dryness, and 2mL of ethyl acetate is added into the residue to dissolve the residue to be used as a sample solution.
Preparation of control medicinal material solution: and (3) adding 0.4g of dragon's blood reference medicine, adding 4g of tea oil, soaking in a crucible for 48 hours, heating to about 200 ℃ for refining for 4 hours, filtering, taking 200 mu L of filtrate, adding 10mL of petroleum ether for dissolving, and preparing the reference medicine solution by the same method.
Preparation of negative control solution: a negative control solution was prepared by the same method as that of the sample.
Thin layer plate: silica gel G thin layer board.
Spotting: the above solution solutions were each 10. Mu.L.
Developing agent: chloroform-ethyl acetate-methanol (8:2:1).
The unfolding mode is as follows: and (5) expanding in an uplink mode.
And (5) checking: spraying 10% sulfuric acid ethanol solution, slightly heating, and inspecting under ultraviolet lamp (365 nm).
Results: four batches of test samples were taken and tested according to the conditions described above. In the chromatogram of the test sample, the same blue spots appear at the positions corresponding to the chromatogram of the control drug. The results were reproducible and the negative control was not significantly disturbed, as shown in FIG. 1.
EXAMPLE 2 methodology investigation
1. Comparison of different lamina plates: the prefabricated silica gel G thin layer plates of two brands of Merck and Qingdao ocean in Germany are respectively tested according to a planned method. The results are shown in figures 2-3, spots with the same color appear on the positions corresponding to the chromatogram of the control medicinal material in the chromatogram of the test sample, and no obvious interference is caused by the negative control.
2. Comparison of different humidity: the relative humidity of the incubator was adjusted with calcium chloride and distilled water, and the spotted thin-layer plates were taken and developed in incubators with low relative humidity (33.6%) and high relative humidity (87.7%), respectively. The results are shown in figures 4-5, and the test sample chromatogram shows spots with the same color at the positions corresponding to the positions of the control chromatogram in the test sample chromatogram under the condition of the relative humidity of 33.6-87.7%, and the negative control has no obvious interference.
3. Comparison of different temperatures: the spotted thin-layer plates were taken and developed under low temperature (in a refrigerator, 8.9 ℃) and high temperature (in an incubator, 37.1 ℃), respectively. The results are shown in figures 6-7, and the test sample chromatogram shows spots with the same color at the positions corresponding to the positions of the control medicine chromatograms when tested at the temperature of 8.9-37.1 ℃, and the negative control has no obvious interference.
Comparative example 1.
This comparative example differs from example 1 in that: the developing agent used is petroleum ether: ethyl acetate (2:1), otherwise the conditions were the same as in example 1.
As a result, as shown in FIG. 8, no spots were found at the positions corresponding to the chromatograms, and no detection of the blood was possible.
Experimental example 1 sample measurement
The 44 batches of samples were tested according to the method of example 1, and as a result, it was found that the two-day oil samples of the respective batches of other companies X and Y were not qualified, and the spots at the corresponding positions of the spots of the reference medicinal material were not clear enough. The rest batches meet the regulations, the spots are clear, the separation effect of the method is good, the results are shown in figures 9-12, and the summary of the results is shown in table 2.
TABLE 2 thin layer identification results for batches of two days oil
The foregoing is a description of embodiments of the invention, which are specific and detailed, but are not to be construed as limiting the scope of the invention. It should be noted that it will be apparent to those skilled in the art that several variations and modifications can be made without departing from the spirit of the invention, which are all within the scope of the invention.
Claims (2)
1. A thin layer chromatography detection method of two-day oil is characterized in that: the method uses chloroform-ethyl acetate-methanol as developing agent, uses silica gel G plate as thin layer chromatography plate, carries out thin layer chromatography detection of the two days oil,
the method comprises the following steps: respectively dispensing two-day oil sample solution and reference substance solution on the same silica gel G plate, developing with chloroform-ethyl acetate-methanol with volume ratio of 8:2:1 as developing agent, taking out, air drying, spraying 5-15% sulfuric acid ethanol solution as color developing agent, heating until spots develop clearly, and placing under ultraviolet light 365nm for inspection;
the preparation method of the sample solution comprises the following steps: adding sodium hydroxide solution into the second-day oil according to the volume ratio of the second-day oil to the 2% sodium hydroxide solution of 1:5, shaking and extracting, adjusting the pH of the extracting solution to 3-5 by using dilute hydrochloric acid, shaking and extracting by using chloroform with the same volume as the 2% sodium hydroxide solution, evaporating the extracting solution to dryness, and dissolving residues in ethyl acetate solution with the volume of the second-day oil to be used as a sample solution;
the preparation method of the reference substance solution comprises the following steps: adding tea oil into sanguis Draxonis reference medicinal materials at a feed-liquid ratio of 1:10, mixing, standing, soaking for 36-60 hr, heating to 180-220deg.C, refining for 3-5 hr, filtering to obtain filtrate, and mixing with the filtrate: petroleum ether is added to dissolve according to the volume ratio of petroleum ether being 1:19, and the petroleum ether is used as a reference substance solution.
2. The method according to claim 1, characterized in that: the concentration of the sulfuric acid ethanol solution is 10%.
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| CN101530545A (en) * | 2009-02-07 | 2009-09-16 | 包头中药有限责任公司 | Quality standard for musk analgesic spray Chinese medicinal preparation and inspection method thereof |
| CN109781922A (en) * | 2019-03-18 | 2019-05-21 | 青海省药品检验检测院 | A kind of TLC Identification identifying dragon's blood and Resina Draconis |
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