CN101665478A - Isoaloeresin D and separation and extraction method for aloin - Google Patents
Isoaloeresin D and separation and extraction method for aloin Download PDFInfo
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- CN101665478A CN101665478A CN200910192487A CN200910192487A CN101665478A CN 101665478 A CN101665478 A CN 101665478A CN 200910192487 A CN200910192487 A CN 200910192487A CN 200910192487 A CN200910192487 A CN 200910192487A CN 101665478 A CN101665478 A CN 101665478A
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- barbaloin
- aloeresin
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- extracting method
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- OUGNWRCWQLUXHX-IAVFCEFGSA-N Isoaloeresin D Natural products COc1cc(C)c2C(=O)C=C(C[C@H](C)O)Oc2c1[C@@H]3O[C@H](CO)[C@@H](O)[C@H](O)[C@H]3OC(=O)C=Cc4ccc(O)cc4 OUGNWRCWQLUXHX-IAVFCEFGSA-N 0.000 title claims abstract description 38
- 238000000605 extraction Methods 0.000 title claims abstract description 28
- 238000000926 separation method Methods 0.000 title claims abstract description 20
- AFHJQYHRLPMKHU-XXWVOBANSA-N Aloin Natural products O=C1c2c(O)cc(CO)cc2[C@H]([C@H]2[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O2)c2c1c(O)ccc2 AFHJQYHRLPMKHU-XXWVOBANSA-N 0.000 title abstract description 15
- AFHJQYHRLPMKHU-UHFFFAOYSA-N isobarbaloin Natural products OC1C(O)C(O)C(CO)OC1C1C2=CC(CO)=CC(O)=C2C(=O)C2=C(O)C=CC=C21 AFHJQYHRLPMKHU-UHFFFAOYSA-N 0.000 title abstract description 15
- CPUHNROBVJNNPW-UHFFFAOYSA-N aloin A Natural products OC1C(O)C(O)C(CO)OC1OC1C2=CC(CO)=CC(O)=C2C(=O)C2=C(O)C=CC=C21 CPUHNROBVJNNPW-UHFFFAOYSA-N 0.000 title abstract description 11
- OUGNWRCWQLUXHX-SSJBJWSWSA-N [(2s,3r,4s,5s,6r)-4,5-dihydroxy-6-(hydroxymethyl)-2-[2-[(2s)-2-hydroxypropyl]-7-methoxy-5-methyl-4-oxochromen-8-yl]oxan-3-yl] (e)-3-(4-hydroxyphenyl)prop-2-enoate Chemical compound O([C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1C1=C2C(C(C=C(C[C@H](C)O)O2)=O)=C(C)C=C1OC)C(=O)\C=C\C1=CC=C(O)C=C1 OUGNWRCWQLUXHX-SSJBJWSWSA-N 0.000 title abstract description 9
- AFHJQYHRLPMKHU-WEZNYRQKSA-N aloin B Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1[C@H]1C2=CC(CO)=CC(O)=C2C(=O)C2=C(O)C=CC=C21 AFHJQYHRLPMKHU-WEZNYRQKSA-N 0.000 title abstract 4
- 241001116389 Aloe Species 0.000 claims abstract description 31
- 235000011399 aloe vera Nutrition 0.000 claims abstract description 31
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical group CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims abstract description 29
- 239000002904 solvent Substances 0.000 claims abstract description 26
- 238000000034 method Methods 0.000 claims abstract description 24
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical group CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims abstract description 19
- 239000002994 raw material Substances 0.000 claims abstract description 11
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical group CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 claims abstract description 9
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical group O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 8
- AFHJQYHRLPMKHU-OSYMLPPYSA-N aloin A Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1[C@@H]1C2=CC(CO)=CC(O)=C2C(=O)C2=C(O)C=CC=C21 AFHJQYHRLPMKHU-OSYMLPPYSA-N 0.000 claims description 47
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 30
- VNGDRRQGKMEADM-LLVKDONJSA-N Aloesaponol I Natural products CC(=O)c1c(O)cc2cc3C[C@@H](O)CC(=O)c3c(O)c2c1C VNGDRRQGKMEADM-LLVKDONJSA-N 0.000 claims description 29
- OUGNWRCWQLUXHX-UHFFFAOYSA-N [4,5-dihydroxy-6-(hydroxymethyl)-2-[2-(2-hydroxypropyl)-7-methoxy-5-methyl-4-oxochromen-8-yl]oxan-3-yl] 3-(4-hydroxyphenyl)prop-2-enoate Chemical compound COC1=CC(C)=C(C(C=C(CC(C)O)O2)=O)C2=C1C1OC(CO)C(O)C(O)C1OC(=O)C=CC1=CC=C(O)C=C1 OUGNWRCWQLUXHX-UHFFFAOYSA-N 0.000 claims description 29
- 230000002411 adverse Effects 0.000 claims description 17
- 230000005526 G1 to G0 transition Effects 0.000 claims description 15
- 239000000284 extract Substances 0.000 claims description 13
- 239000000203 mixture Substances 0.000 claims description 11
- RSOWICXTDPTUSX-UHFFFAOYSA-N O.CC(C)=O.CCCCCC.CCOC(C)=O Chemical compound O.CC(C)=O.CCCCCC.CCOC(C)=O RSOWICXTDPTUSX-UHFFFAOYSA-N 0.000 claims description 6
- 238000001035 drying Methods 0.000 claims description 6
- 239000002798 polar solvent Substances 0.000 claims description 4
- 230000008569 process Effects 0.000 claims description 3
- NIQQIJXGUZVEBB-UHFFFAOYSA-N methanol;propan-2-one Chemical compound OC.CC(C)=O NIQQIJXGUZVEBB-UHFFFAOYSA-N 0.000 claims description 2
- 239000007858 starting material Substances 0.000 claims description 2
- 238000010262 high-speed countercurrent chromatography Methods 0.000 abstract description 7
- 239000000178 monomer Substances 0.000 abstract description 4
- 238000009776 industrial production Methods 0.000 abstract description 2
- 235000019439 ethyl acetate Nutrition 0.000 abstract 2
- 239000000047 product Substances 0.000 description 27
- 239000012071 phase Substances 0.000 description 24
- 239000000523 sample Substances 0.000 description 14
- 239000000843 powder Substances 0.000 description 10
- YDQWDHRMZQUTBA-UHFFFAOYSA-N Aloe emodin Chemical compound C1=CC=C2C(=O)C3=CC(CO)=CC(O)=C3C(=O)C2=C1O YDQWDHRMZQUTBA-UHFFFAOYSA-N 0.000 description 8
- 239000007788 liquid Substances 0.000 description 7
- 238000005303 weighing Methods 0.000 description 6
- 239000000470 constituent Substances 0.000 description 5
- 239000000287 crude extract Substances 0.000 description 5
- 239000007787 solid Substances 0.000 description 5
- 239000000243 solution Substances 0.000 description 5
- 239000000126 substance Substances 0.000 description 5
- 238000001644 13C nuclear magnetic resonance spectroscopy Methods 0.000 description 4
- 238000005160 1H NMR spectroscopy Methods 0.000 description 4
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 4
- OKKJLVBELUTLKV-MZCSYVLQSA-N Deuterated methanol Chemical compound [2H]OC([2H])([2H])[2H] OKKJLVBELUTLKV-MZCSYVLQSA-N 0.000 description 4
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 4
- 239000003814 drug Substances 0.000 description 4
- 230000000694 effects Effects 0.000 description 4
- 238000010828 elution Methods 0.000 description 4
- 238000004128 high performance liquid chromatography Methods 0.000 description 4
- 230000014759 maintenance of location Effects 0.000 description 4
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 239000000706 filtrate Substances 0.000 description 3
- 238000001819 mass spectrum Methods 0.000 description 3
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- 239000012488 sample solution Substances 0.000 description 3
- 238000005070 sampling Methods 0.000 description 3
- 238000007789 sealing Methods 0.000 description 3
- 238000011105 stabilization Methods 0.000 description 3
- 239000006228 supernatant Substances 0.000 description 3
- 238000001291 vacuum drying Methods 0.000 description 3
- 102100027324 2-hydroxyacyl-CoA lyase 1 Human genes 0.000 description 2
- 101001009252 Homo sapiens 2-hydroxyacyl-CoA lyase 1 Proteins 0.000 description 2
- 150000004056 anthraquinones Chemical class 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- SIHHLZPXQLFPMC-UHFFFAOYSA-N chloroform;methanol;hydrate Chemical compound O.OC.ClC(Cl)Cl SIHHLZPXQLFPMC-UHFFFAOYSA-N 0.000 description 2
- 238000004587 chromatography analysis Methods 0.000 description 2
- OTAFHZMPRISVEM-UHFFFAOYSA-N chromone Chemical compound C1=CC=C2C(=O)C=COC2=C1 OTAFHZMPRISVEM-UHFFFAOYSA-N 0.000 description 2
- 239000013078 crystal Substances 0.000 description 2
- 230000007812 deficiency Effects 0.000 description 2
- 235000008216 herbs Nutrition 0.000 description 2
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- 238000002156 mixing Methods 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
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- 230000001988 toxicity Effects 0.000 description 2
- 231100000419 toxicity Toxicity 0.000 description 2
- 206010003210 Arteriosclerosis Diseases 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 206010061218 Inflammation Diseases 0.000 description 1
- 241000234280 Liliaceae Species 0.000 description 1
- 238000005481 NMR spectroscopy Methods 0.000 description 1
- 239000004952 Polyamide Substances 0.000 description 1
- 238000012356 Product development Methods 0.000 description 1
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 238000000862 absorption spectrum Methods 0.000 description 1
- 239000003463 adsorbent Substances 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- PYKYMHQGRFAEBM-UHFFFAOYSA-N anthraquinone Natural products CCC(=O)c1c(O)c2C(=O)C3C(C=CC=C3O)C(=O)c2cc1CC(=O)OC PYKYMHQGRFAEBM-UHFFFAOYSA-N 0.000 description 1
- 230000000844 anti-bacterial effect Effects 0.000 description 1
- 208000011775 arteriosclerosis disease Diseases 0.000 description 1
- 238000000889 atomisation Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 150000001793 charged compounds Chemical class 0.000 description 1
- GCHZQTJYIFBIDC-UHFFFAOYSA-N chloroform methanol propan-2-one hydrate Chemical compound O.OC.CC(C)=O.ClC(Cl)Cl GCHZQTJYIFBIDC-UHFFFAOYSA-N 0.000 description 1
- 238000011208 chromatographic data Methods 0.000 description 1
- 238000004440 column chromatography Methods 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 229960000935 dehydrated alcohol Drugs 0.000 description 1
- GLYLMXARZJNUEY-UHFFFAOYSA-N dichloromethane;methanol;hydrate Chemical compound O.OC.ClCCl GLYLMXARZJNUEY-UHFFFAOYSA-N 0.000 description 1
- 238000007599 discharging Methods 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 230000009977 dual effect Effects 0.000 description 1
- HZQXXYJHLCSUGQ-UHFFFAOYSA-N ethyl acetate hexane methanol hydrate Chemical compound O.OC.CCCCCC.CCOC(C)=O HZQXXYJHLCSUGQ-UHFFFAOYSA-N 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 210000003746 feather Anatomy 0.000 description 1
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- 238000001294 liquid chromatography-tandem mass spectrometry Methods 0.000 description 1
- 238000000622 liquid--liquid extraction Methods 0.000 description 1
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- 235000013372 meat Nutrition 0.000 description 1
- 239000000401 methanolic extract Substances 0.000 description 1
- 125000000956 methoxy group Chemical group [H]C([H])([H])O* 0.000 description 1
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- 229920002647 polyamide Polymers 0.000 description 1
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- 229920001343 polytetrafluoroethylene Polymers 0.000 description 1
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- Medicines Containing Plant Substances (AREA)
- Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)
Abstract
The invention discloses an isoaloeresin D and a separation and extraction method for aloin, which takes aloe as a raw material to obtain a preliminary extraction; in high-speed counter-current chromatography, n-hexane, acetic ether, acetone and water form mobile phase at the upper part and fixed phase at the lower part so as to carry out separation and extraction, wherein the volume ratio of n-hexane to acetic ether to acetone to water is 0.5 to 0.1:4 to 6:0.5 to 2:4 to 6. The method simultaneously obtains high-purity isoaloeresin D and aloin, two monomer products, during one separation process for the first time. Used solvents have little pollution and high safety and the method has high extraction rate and simple operation and is suitable for industrial production and application. The method has wide application prospect in the extraction of active components of aloin and the bio-medical field.
Description
Technical field
The invention belongs to medicine and biological field, relate to a kind of method of extracting aloe effective constituent, be specifically related to the separating and extracting method that a kind of employing high speed adverse current chromatogram extracts Barbaloin A (Aloin) and different aloeresin D (Isoaloeresin D).
Background technology
Aloe (Aloe) is the perennial meat herbaceous plant of Liliaceae Aloe, is widely used in medicine, daily use chemicals and health care of food field.But, because the various complexity of aloe effective constituent, and much the character of chemical ingredientss is very close, therefore separates various compositions in the aloe, obtain hectogram and even the monomeric technology of feather weight is the difficult problem that aloe basis and high-quality product development are produced always.Anthraquinone analog compound is an aloetic main pharmacodynamics composition, and Barbaloin A wherein (Aloin) and different aloeresin D (Isoaloeresin D) more become the research focus in this field in recent years.Barbaloin A has another name called Barbaloin, aloin, Barbaloin in discharging body dirt, prevent arteriosclerosis, suppressing the tumor growth aspect has important effect; Different aloeresin D belongs to heteroside, because it has antibacterial and anti-inflammation functions, is fragrant local flavor prerequisite material simultaneously, has been subjected to suitable attention in field of medicaments, field of fine chemical.
At present for the extraction of aloe effective constituent, adopt silica gel, polyamide column chromatography method or macroporous adsorbent resins to carry out separation and Extraction both at home and abroad, but above-mentioned separation method complex operation more, the cost height is renderd a service lowly, is difficult to carry out industrial production.High speed adverse current chromatogram (High Speed Counter Current Chromatography, HSCCC) be a kind of liquid liquid distribution chromatography isolation technique without solid-state carrier, compare with other chromatographic technique, HSCCC is simple to operate, render a service highly, can eliminate sample irreversible adsorption that carrier causes fully and influences such as being infected with of sample, inactivation, sex change.Once set up good separation condition, it can realize efficient, the high purity separation of each component in the complex mixture and the preparation of a large amount of samples at short notice, so the research of separation condition and foundation are that HSCCC realizes successfully isolating key.
Because separating by it, the advantage of HSCCC on separation and Extraction, existing more at present research obtain aloetic effective constituent.Wang Chunyan etc. are moving phase down to be stationary phase mutually on solvent systems chloroform-methanol-water (9: 10.5: 8) mutually, and disposable separation obtains Barbaloin A and rhabarberone from the aloe crude drug; Chen Cunshe etc. draw by analysis, use chloroform-methanol-acetone-water (9: 8: 1: 8) be the best results of solvent systems separating and purifying aloe effective constituent anthraquinones rhabarberone and Barbaloin; Pan Xia etc. are raw material with the aloe full leaf, obtain the aloe chromone crude extract through a series of preprocessing means, adopting chloroform-methanol-water (4: 3: 2) mixing solutions and methylene chloride-methanol-water (5: 4: 2) mixing solutions is the separated from solvent system, goes out purity at the aloe chromone monomer more than 95% through the HSCCC separation and purification of two steps.But above research all exists certain deficiency: (1) disposable separating obtained sample is single, though can obtain Barbaloin A (Barbaloin) and rhabarberone simultaneously, in fact Barbaloin A is the precursor substance of rhabarberone, and effect is similar; (2) solvent toxicity that is adopted is bigger, as chloroform, methylene dichloride etc., the insecurity that pollutes and produce.
And for different aloeresin D, have not yet to see any report it is being obtained with once separating preparation process simultaneously with Barbaloin A.
Summary of the invention
The objective of the invention is at the deficiencies in the prior art, provide a kind of with high speed adverse current chromatogram once simultaneously high efficiency separation prepare the method for different aloeresin D of high purity (Isoaloeresin D) and Barbaloin A (Aloin).
The present invention is achieved through the following technical solutions above-mentioned purpose:
The separating and extracting method of a kind of different aloeresin D and Barbaloin A comprises the steps:
With the aloe is that starting material obtain primary extract, carries out separation and Extraction in high speed adverse current chromatogram.
Described primary extract is to be raw material with the aloe, adds the polar solvent supersound extraction, and the gained extracting solution reclaims the solvent drying under reduced pressure and gets.Aloe raw material can be fresh aloe or aloe products, and wherein preferred raw material is the aloe medicinal material.
Described polar solvent is acetone, methyl alcohol or acetone: methyl alcohol (8～6: 2～4).Preferred solvent is an acetone.The weight ratio of solvent and raw material is 15～25: 1.
The moving phase of described high speed adverse current chromatogram solvent systems and be fixed as normal hexane-ethyl acetate-acetone-water and constitute.Volume ratio wherein is a normal hexane: ethyl acetate: acetone: water=0.5～0.1: 4～6: 0.5～2: 4～6.Preferred volume ratio is a normal hexane: ethyl acetate: acetone: water=02: 5: 0.5～2: 5 can be 0.2: 5: 0.5: 5,0.2: 5: 1: 5,0.2: 5: 1.5: 5 or 0.2: 5: 2: 5.
The solvent systems of described high speed adverse current chromatogram adopts to be gone up as moving phase, following to stationary phase.
The consumption of described stationary phase, moving phase and the weight ratio of raw material are 94～120: 230～250: 0.3～0.6.
In the sepn process of described high speed adverse current chromatogram, flow rate of mobile phase is 0.8 ~ 1.2ml/min.
The column diameter that adopts when high speed adverse current chromatogram is 2.16mm, column volume 120mm, flow rate of mobile phase 1.0ml/min, engine speed 860rpm, during sample size 500mg condition, the collection time period of different aloeresin D is 100 ~ 140min, and the collection time period of Barbaloin A is 170 ~ 250min.
The present invention is by big quantity research and cut-and-try work, determined just to carry treatment technology, selected normal hexane-ethyl acetate-acetone-water as solvent systems (what adopt usually is normal hexane-ethyl acetate-methanol-water system), optimized its configuration proportion, will go up as moving phase, make different aloeresin D and Barbaloin A obtain ideal separation and Extraction effect.
Compared with prior art, the present invention has following beneficial effect:
1. the present invention can obtain the different aloeresin D of high purity (Isoaloeresin D) and two monomer products of Barbaloin A (Aloin) simultaneously in a sepn process;
2. the present invention need not to use the big solvents of toxicity such as chloroform, methylene dichloride, pollutes little, safe;
3. the present invention adopts as moving phase, following to stationary phase (normal hexane: ethyl acetate: methyl alcohol: aqueous systems), when reclaiming normal hexane, ethyl acetate, just obtain product subsequently, compare with phase (water) is as the method for moving phase down, the present invention is removing solvent, is obtaining aspect the solid phase prod tangible advantage is arranged;
4. extraction yield height of the present invention, different aloeresin D reaches more than 21%; Barbaloin A reaches more than 42%;
5. stationary phase retention rate height of the present invention reaches more than 78%;
6. the present invention is raw materials used clear and definite, and promptly aloe or aloe medicinal material therefrom can obtain a large amount of different aloeresin D (Isoaloeresin D) and Barbaloin A (Aloin) monomer;
7. the present invention is simple to operate, and separation condition is fixed, and is suitable for suitability for industrialized production.
Extract the effect of different aloeresin D (Isoaloeresin D) and Barbaloin A (Aloin) in order to verify present method, the present invention also adopts HPLC that extract is carried out purity check, carry out structural analysis with UV, IR, 1H NMR, 13C NMR and MS, calculate its extraction yield with " product quality/sample introduction quality * 100% ".
Description of drawings
The high speed adverse current chromatogram figure of Fig. 1 aloe acetone crude extract
The spectrogram of different aloeresin D among Fig. 2 HPCL
The spectrogram of Barbaloin A among Fig. 3 HPCL
The chemical structure of the different aloeresin D of Fig. 4
The chemical structure of Fig. 5 Barbaloin A
Embodiment
Below further specify technical scheme of the present invention by specific embodiment.
Take by weighing 25g aloe medicinal powder in beaker, divide three times with 500ml acetone, ultrasonic 20min extracts, and each back of extracting is centrifugal, collect supernatant liquid, No. three times extracting solution mixes, and reclaims solvent with 60 ℃ of Rotary Evaporators and makes extract be paste clearly, then in 60 ℃ of drying under reduced pressure of vacuum drying oven, obtain pale brown look glossiness solid, weigh, obtain acetone extract 16.26g, extraction yield is 65.06%.Sealing is stored in below 10 ℃, and is standby.
The high speed adverse current chromatogram separation and Extraction adopts Mk5QuilkPrep500 high-speed counter-current chromatograph (Britain AECS company), is furnished with polyfluortetraethylene pipe (PTFE) separator column (pipe diameter 2.16mm, separated volume 120ml), SeriesII constant flow pump (Scientific System company), SPD-10Avp UV, visible light detector (island, Suzhou Tianjin instrument company), N2000 chromatographic data workstation (Zhejiang University's intelligence reaches Information Technology Co. Ltd).
In separating funnel, prepare two-phase system, with normal hexane-ethyl acetate-acetone-water (0.2: 5: 1.5: 5) put in the separating funnel, leave standstill behind the shake well, following to stationary phase, last as moving phase, ultrasonic degas; Take by weighing an amount of above-mentioned aloe crude extract powder, with mutually ultrasonic assist in dissolving under the 6ml, be made into the sample solution that concentration is 85.5mg/ml, filter, get filtrate and use sampling valve sample introduction 5ml, adopt the type of elution of unicoil, tail joint, at first make pipeline be full of stationary phase, when instrument during, moving phase is pumped into pipeline with the 1.0ml/min flow velocity with the operation of the velocity-stabilization of 860rpm.When moving phase obviously being arranged when the pipeline tail end flows out, illustrate to reach biphase equilibrium that calculate the stationary phase retention rate, the stationary phase retention rate of this experiment is 79.1%.30 seconds after cut receptors begin to connect automatically flow point behind the sample introduction, and speed is 5min/tube, and effluent liquid detects at the 254nm place with UV-detector, writes down color atlas simultaneously, collect flow point according to color atlas.
The flow point of gained corresponding to peak I (different aloeresin D) and peak II (Barbaloin A) (seeing accompanying drawing 1) merged, reclaim solvent, get component I and component I I.Carry out purity check with HPLC, Waters1525 high performance liquid chromatograph, Waters717 type automatic sampler, 2996 type diode-array detectors, Empower chromatographic working station (U.S. waters company).Moving phase is methyl alcohol: water:, gradient elution, elution program is: 0～30min:40%A-80%A, 60%B-20%B; 30～40min, 80%A-95%A, 20%B-5%B; A is the methyl alcohol phase, and B is water (containing acetic acid 0.34%) flow velocity: 1.0mL/min; Column temperature is 25 ℃, and DAD scanning wavelength scope is 190～370nm, and the ultraviolet detection wavelength is 254nm;
Carry out structural analysis with UV, IR, 1H NMR, 13C NMR and MS.TENSOR37 Red external spectrum instrument (German Bruker company) KBr compressing tablet, Bruker AvanceIII 400MHz nuclear magnetic resonance analyser (German Bruker company), LCMS-IT-TOF high-resolution mass spectrometer (day island proper Tianjin company).Mass spectrum condition: ion source ESI, atomization air pressure 35psi, dry gas flow 11mL/min, 350 ℃ of drying temperatures, scanning of the mass spectrum mass range 50～1000.Full scan one-level mass spectrum and selection ion full scan second order ms dual mode are measured simultaneously.
Analytical results:
Component I is an one matter, and purity is 98.62%; Component I I is the component that binary becomes, total purity 99.54% (area normalization method) (seeing accompanying drawing 2,3).
Component I, component I I reclaim solvent respectively, obtain product I and product II.Product I is the faint yellow solid powder, heavy 108.6mg, and extraction yield (product quality/sample introduction quality) is 21.2%; Product II is the yellow crystal powder, heavy 219.2mg, and extraction yield is 42.7%.
Product I its structured data after measured is: UV λ max (nm) MeOH:299.1,299.9; IR (KBr) cm-1:3446,1649,1602,1514,1383; TOF-MS (m/z): 555.1878[M-H]-, 1H NMR and 13C NMR (CD3OD) data see Table 1.Above data are consistent with the different aloeresin D of bibliographical information, are defined as different aloeresin D, its structure such as accompanying drawing 4.
Product II is two mixture of ingredients of A, B, and total purity 99.54% is analyzed through HPLC-DAD, and composition B is consistent with the retention time and the UV spectrum absorption at Barbaloin A B reference substance peak under the identical chromatographic conditions, tentatively is defined as Barbaloin A B; The ultra-violet absorption spectrum of composition A is consistent with composition B, tentatively is defined as Barbaloin A isomer-Barbaloin A A.
Through further measuring UV λ max (nm) MeOH:223.3 of A, two compositions of B, 270.3,297.5,357.0, IR (KBr) cm-1:3442,1619,1458,1386,1261.Spectroscopic data conforms to Barbaloin A B reference substance spectroscopic data; Analyze through LC-MS/MS, the m/z of composition A, B and Barbaloin A B reference substance is 417, and molecular ion peak, fragmention are identical, and comprehensively above analytical results determines that product II is the mixture of Barbaloin A A, B, its structure such as accompanying drawing 5.
Take by weighing 30g aloe raw medicinal herbs powder in beaker, use methyl alcohol: acetone (7: 3) mixed solution 500ml, divide three ultrasonic 30min to extract, centrifugal, collect supernatant liquid and merge, reclaiming solvent with 60 ℃ of Rotary Evaporators makes extract be paste clearly, in 60 ℃ of drying under reduced pressure of vacuum drying oven, obtain pale brown look glossiness solid then, weigh, obtain methanol extract 19.38g, extraction yield is 64.60%.Sealing is stored in below 10 ℃ standby.
The high speed adverse current chromatogram separation and Extraction is prepared two-phase system in separating funnel, with normal hexane-ethyl acetate-acetone-water (0.2: 5: 1.0: 5) put in the separating funnel, leave standstill behind the shake well, following to stationary phase, last as moving phase, ultrasonic degas; Take by weighing an amount of above-mentioned aloe crude extract powder, with mutually ultrasonic assist in dissolving under the 6ml, be made into the sample solution that concentration is 80.1mg/ml, filter, get filtrate and use sampling valve sample introduction 5ml, adopt the type of elution of unicoil, tail joint, at first make pipeline be full of stationary phase, when instrument during, moving phase is pumped into pipeline with the 1.0ml/min flow velocity with the operation of the velocity-stabilization of 860rpm.When moving phase obviously being arranged when the pipeline tail end flows out, sample introduction, the after cut receptor began to connect automatically flow point in 30 seconds, and speed is 5min/tube, and effluent liquid detects at the 254nm place with UV-detector, writes down color atlas simultaneously, collects flow point according to color atlas.
Analytical results:
Component I, component I I reclaim solvent respectively, obtain product I and product II with the dehydrated alcohol recrystallization.Product I is the heavy 83.2mg of faint yellow solid powder, and extraction yield (product quality/sample introduction quality) is 20.8%; Product II is the yellow crystal powder, heavy 172.8mg, and extraction yield is 43.2%.
Carry out purity check with HPLC, product I (different aloeresin D) purity is 99.04%; Product II (Barbaloin A A, B) purity 98.66%
Take by weighing 100g aloe raw medicinal herbs powder in beaker, use methyl alcohol: acetone (1: 2) mixed solution 1500ml, divide three ultrasonic 30min to extract, centrifugal, collect supernatant liquid and merge, reclaiming solvent with 60 ℃ of Rotary Evaporators makes extract be paste clearly, in 60 ℃ of drying under reduced pressure of vacuum drying oven, obtain pale brown look glossiness solid then, weigh, obtain methanol acetone extract 67.18g, extraction yield is 67.18%.Sealing is stored in below 10 ℃ standby.
The high speed adverse current chromatogram separation and Extraction is prepared two-phase system in separating funnel, with normal hexane-ethyl acetate-acetone-water (0.15: 5: 0.5: 4.5) put in the separating funnel, leave standstill behind the shake well, following to stationary phase, last as moving phase, ultrasonic degas; Take by weighing an amount of above-mentioned aloe crude extract powder, with mutually ultrasonic assist in dissolving under the 7ml, be made into the sample solution that concentration is 85.03mg/ml, filter, get filtrate and use sampling valve sample introduction 5ml, adopt the type of elution of unicoil, tail joint, at first make pipeline be full of stationary phase, when instrument during, moving phase is pumped into pipeline with the 1.0ml/min flow velocity with the operation of the velocity-stabilization of 860rpm.When moving phase obviously being arranged when the pipeline tail end flows out, sample introduction, the after cut receptor began to connect automatically flow point in 30 seconds, and speed is 5min/tube, and effluent liquid detects at the 254nm place with UV-detector, writes down color atlas simultaneously, collects flow point according to color atlas.
Interpretation of result:
Component I, component I I reclaim solvent respectively, obtain product I and product II.The heavy 90.68mg of product I, extraction yield (product quality/sample introduction quality) is 21.33%; The heavy 178.9mg of product II, extraction yield is 42.17%.
Carry out purity check with HPLC, product I (different aloeresin D) purity is 98.15%; Product II (Barbaloin A A, B) purity 98.02%.
The 1H NMR of table 1. product I and 13C NMR (CD3OD) data
| The position | ??1H(□，ppm) | ??13C(□，ppm) |
| ??2 | ??167.4 | |
| ??3 | ??6.11 | ??112.5 |
| ??4 | ??182.3 | |
| ??4a | ??116.8 | |
| ??5 | ??144.6 | |
| ??6 | ??6.79 | ??112.6 |
| ??7 | ??162.0 | |
| ??8 | ??111.8 | |
| ??1a | ??159.6 | |
| ??9 | ??2.76 | ??44.6 |
| ??10 | ??4.50m | ??66.0 |
| ??11 | ??1.35d(J＝6.2) | ??23.7 |
| ??12 | ??2.72 | ??23.6 |
| ??7-OMe | ??3.88 | ??57.1 |
| ??1’ | ??5.19d(J＝10.1) | ??72.3 |
| ??2’ | ??5.73t(J＝9.6) | ??73.8 |
| ??3’ | ??77.9 | |
| ??4’ | ??72.8 | |
| ??5’ | ??83.0 | |
| ??6’ | ??63.1 | |
| ??1” | ??168.0 | |
| ??2” | ??6.04d(J＝15.9) | ??114.7 |
| ??3” | ??7.36d(J＝16.8) | ??146.6 |
| ??4” | ??127.0 | |
| ??5”，9” | ??7.33d(J＝8.9) | ??131.1 |
| ??6”，8” | ??6.75d(J＝8.6) | ??117.2 |
| ??7” | ??161.3 |
Claims (9)
1. the separating and extracting method of different aloeresin D and Barbaloin A is characterized in that comprising the steps:
With the aloe is that starting material obtain primary extract, carries out separation and Extraction in high speed adverse current chromatogram.
2. the separating and extracting method of different aloeresin D as claimed in claim 1 and Barbaloin A is characterized in that: described high speed adverse current chromatogram solvent systems is normal hexane-ethyl acetate-acetone-water.
3. the separating and extracting method of different aloeresin D as claimed in claim 2 and Barbaloin A is characterized in that: the solvent quality ratio of described high speed adverse current chromatogram solvent systems is a normal hexane: ethyl acetate: acetone: water=0.5～0.1: 4～6: 0.5～2: 4～6.
4. as the separating and extracting method of claim 2 or 3 described different aloeresin D and Barbaloin A, it is characterized in that: the solvent systems of described high speed adverse current chromatogram adopts to be gone up as moving phase, following to stationary phase.
5. as the separating and extracting method of arbitrary described different aloeresin D of claim 1～4 and Barbaloin A, it is characterized in that: the consumption of described stationary phase, moving phase and the weight ratio of raw material are 94～120: 230～250: 0.3～0.6.
6. the separating and extracting method of different aloeresin D as claimed in claim 1 and Barbaloin A is characterized in that: in the sepn process of described high speed adverse current chromatogram, flow rate of mobile phase is 0.8～1.2ml/min.
7. the separating and extracting method of different aloeresin D as claimed in claim 1 and Barbaloin A is characterized in that: described primary extract is to be raw material with the aloe, adds the polar solvent supersound extraction, and the gained extracting solution reclaims the solvent drying under reduced pressure and gets.
8. the separating and extracting method of different aloeresin D as claimed in claim 7 and Barbaloin A is characterized in that: described polar solvent is a kind of in acetone, methyl alcohol or the acetone-carbinol mixture.
9. as the separating and extracting method of claim 7 or 8 described different aloeresin D and Barbaloin A, it is characterized in that: the weight ratio of solvent and raw material is 15～25: 1.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102101893A (en) * | 2010-12-20 | 2011-06-22 | 大兴安岭林格贝有机食品有限责任公司 | Method for enriching and purifying aloe polysaccharides in aloe |
| CN104215614A (en) * | 2014-07-16 | 2014-12-17 | 中山大学 | Method for detecting aloin A, aloin B and aloe-emodin in series through high performance liquid chromatography-diode array/fluorescent detector |
| CN105211787A (en) * | 2015-11-13 | 2016-01-06 | 谢镜国 | A kind of extraction process of longan seed polyphenol |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1965852A (en) * | 2005-06-07 | 2007-05-23 | 中国医学科学院药物研究所 | Aloeresin D and effective component, preparation process, pharmaceutical composition and use thereof |
| CN100453548C (en) * | 2006-10-11 | 2009-01-21 | 上海华震科技有限公司 | Method for separating barbaloin in high purity from product of aloe |
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2009
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102101893A (en) * | 2010-12-20 | 2011-06-22 | 大兴安岭林格贝有机食品有限责任公司 | Method for enriching and purifying aloe polysaccharides in aloe |
| CN102101893B (en) * | 2010-12-20 | 2012-08-22 | 大兴安岭林格贝有机食品有限责任公司 | Method for enriching and purifying aloe polysaccharides in aloe |
| CN104215614A (en) * | 2014-07-16 | 2014-12-17 | 中山大学 | Method for detecting aloin A, aloin B and aloe-emodin in series through high performance liquid chromatography-diode array/fluorescent detector |
| CN104215614B (en) * | 2014-07-16 | 2016-11-23 | 中山大学 | A kind of high performance liquid chromatography-diode array/fluorescence detector Series detectors barbaloin A, B and the method for aloe-emodin |
| CN105211787A (en) * | 2015-11-13 | 2016-01-06 | 谢镜国 | A kind of extraction process of longan seed polyphenol |
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