CN102206209B - Method for extracting and separating soybean isoflavone monomer compounds from soybeans - Google Patents
Method for extracting and separating soybean isoflavone monomer compounds from soybeans Download PDFInfo
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- CN102206209B CN102206209B CN 201110089865 CN201110089865A CN102206209B CN 102206209 B CN102206209 B CN 102206209B CN 201110089865 CN201110089865 CN 201110089865 CN 201110089865 A CN201110089865 A CN 201110089865A CN 102206209 B CN102206209 B CN 102206209B
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Abstract
The invention discloses a method for extracting and separating soybean isoflavone monomer compounds from soybeans, comprising the following steps of: (1) preparation of a soybean isoflavone extractive and (2) column chromatography purification. The soybean isoflavone extractive obtained by using the method has high content of soybean isoflavone compounds and less impurities and is beneficial for follow-up further purification; in the purifying process, chloroform and other organic solvents harmful to the environment are not used so that the method is environment-friendly; and a simple, convenient and high-efficient way capable of realizing the purpose of the invention is established so that the condition of the chromatography method is optimized and the purity and the efficiency are both greatly improved.
Description
Technical field
The invention belongs to chemical field, specifically relating to a kind of method of separating multiple high-purity soybean isoflavone monomeric compound of extracting from soybean, mainly is the compounds such as separation and purification glycitin (Glycitin), daidzin (Daidzin), Genistoside (Genistin), glycitein (Glycitein), daidzein (Daidzein), genistein (Genistein).
Background technology
Soybean isoflavones is the secondary metabolite that soybean forms in process of growth, is the mixture of Polyphenols.The multiple nutrients of soybean, health-care effect are relevant with soybean isoflavones.Modern pharmacology studies show that soybean isoflavones has estrogen-like effects, can prevent menopausal syndrome; Have prevention, improve the osteoporosis effect, reduce bone-loss, increase simultaneously body to the absorption of calcium, increase bone density; Have prevention and the effect for the treatment of cancer, mammary cancer, colorectal carcinoma, lung cancer, prostate cancer and skin carcinoma and leukemia are had obvious therapeutic action, but the also generation of preventing ovarian cancer, colorectal carcinoma, cancer of the stomach and prostate cancer; Has the preventing cardiovascular disease effect, reducing cholesterol, adjusting blood fat, the generation of preventing cardiovascular disease; Has the effect of prevention degenerative brain disorder.
At present, the method for separating and purifying high-purity compound has been reported from soybean.Chinese patent (the separation method of major monomeric compoent of soybean isoflavone, application number 03135952.3) is not less than the mixture of 40% soybean isoflavones as raw material take content, take silica gel as sorbent material, take chloroform-methanol solution as eluent system, separate the major monomeric compoent that obtains genistein, daidzein, Genistoside, four kinds of soybean isoflavones of daidzin, the content of each monomer component all reaches more than 90%.Chinese patent (the method for separating genistein from isoflavone genin mixture, application number 201010114937.3) binary mixed solvent that forms take ionic liquid or by ionic liquid and other polar solvents adopts fractional-distillation extraction to separate from the isoflavone genin mixture that contains Daidezin, genistein and glycitein and obtains genistein as extraction agent.(the standby Standard multi-component soybean isoflavone glucoside of middle compacting such as full of leaves grain husk, grain science and technology is with economical, 04 phase in 2009) take the soybean isoflavones primary extract as raw material, the standby system of compacting and ODSC18 chromatographic column in the use, separate preparing daidzin, glycitin and 3 kinds of soybean isoflavone glucosides of Genistoside, purity is respectively 99.69%, 98.62% and 96.26%.Kang Shaohua etc. (silica gel column chromatography separates the research of isoflavone genin, and China brewages, 01 phase in 2009) utilize silica gel column chromatography to separate and obtain genistein, daidzein, 3 kinds of soybean isoflavones of glycitein, and purity is all above 90%.(the Isolation of 4 major soybean isoflavones with macroporous resins such as Liu Ye, Central-South pharmacy, 04 phase in 2007) utilizes behind the macroporous adsorbent resin ethanolic soln gradient elution with 20%, 25%, 35%, 45%, 70%, obtain daidzin, Genistoside, daidzein, 4 kinds of soybean isoflavones compositions of genistein, purity is respectively 49.31%, 60.38%, 19.33% and 8.14%.(the separation method of major monomeric compoent of soybean isoflavone such as Yao Kai, Sichuan University's journal (engineering science version), 03 phase in 2004) adopt 300~400 order silica gel, with chloroform-methanol eluent system wash-out, obtain Genistoside, daidzin, genistein and 4 kinds of major monomeric compoents of daidzein.(high-speed countercurrent chromatography separates daidzin and the Genistoside in the preparation soybean isoflavones for river and source etc., Food science, 01 phase in 2004) daidzin and the Genistoside in the employing high-speed countercurrent chromatography purification of soybean isoflavones, purity reaches respectively 98.2% and 99.2%.Xu De equality (isolation identification of soybean isoflavones and the research of antioxygenation, Agricultural University Of Nanjing's journal, 03 phase of calendar year 2001) is utilized silica gel column chromatography and chloroform, ethanol gradient elution, obtains daidzein and genistein.With Saphadex LH-20 post, methyl alcohol, water gradient elution, in conjunction with the repeated multiple times chromatography of TOYOpearl HW-40 chromatography column, obtain daidzin and Genistoside.
Above-mentioned the whole bag of tricks or take multistep to separate operates cumbersome; Or only can obtain a kind of or a few soybean isoflavones composition; Or the composition purity that obtains is not high; Or to use the chloroform noxious solvent in the sepn process.
Summary of the invention
The object of the invention is to overcome the deficiencies in the prior art, provide a kind of easy and simple to handle, fractional dose is large, comprehensive cost is low, the method of the multiple high-purity soybean isoflavone compound of quick preparation with short production cycle, a step separation and purification can obtain the multiple soybean isoflavones monomeric compounds such as highly purified glycitin, daidzin, Genistoside, glycitein, daidzein, genistein.
The solution of the present invention is as follows:
Extract the method for separating the soybean isoflavones monomeric compound from soybean, step is:
(1) preparation of soybean isoflavone: skimmed soy beans dregs of beans, soak with ethanol or methyl alcohol, ethanol or methyl alcohol diacolation with 5-20 times of volume extract, percolate spraying drying after being recycled to without alcohol obtains the extract meal, the extract meal with acetone refluxing extraction, condensing crystal, is obtained soybean isoflavone;
(2) column chromatography purification: with the soybean isoflavone dissolve with methanol, upper Superose 12 posts carry out the chromatographic separation purifying, be that elutriant carries out wash-out with methanol-water or alcohol-water, eluent flow rate is 1-10 times of column volume per hour, effluent liquid is detected at 254 nm by UV-detector, collect the target components cut, cut concentrating under reduced pressure with obtaining obtains the multiple soybean isoflavones monomeric compounds such as highly purified glycitin, daidzin, Genistoside, glycitein, daidzein, genistein.
Foregoing method, preferred scheme be, uses the double-deck diacolation post with the chuck heat tracing when step (1) diacolation extracts, and medium temperature is 30-80 ℃, the percolate flow velocity be 1-5 times of column volume/hour.More preferably, medium temperature is 40-60 ℃ (preferred 50 ℃), and the percolate flow velocity is 2-4 times of column volume/hour (preferred 3 times of column volumes/hour).
Foregoing method, preferred scheme be, step (1) is during with ethanol or methyl alcohol soak degreasing soybean meal, the mass concentration of ethanol or methyl alcohol is the preferred 50%-80% of 40%-85%(, more preferably 60%), soak time is 6-12 hour (preferred 7-10 hour, more preferably 8 hours).
Foregoing method, preferred scheme be, the mass concentration of used ethanol or methyl alcohol was the preferred 50%-80% of 40%-85%(when step (1) diacolation extracted, more preferably 60%).
Foregoing method, preferred scheme be, used acetone quality concentration is the preferred 85%-95% of 80%-100%(during step (1) refluxing extraction extract meal, more preferably 90%).
Foregoing method, preferred scheme is that step (2) elutriant is the methanol-water elutriant of 20%-80%.
Foregoing method, preferred scheme is that type of elution is degree such as grade, substep or gradient.
Foregoing method, preferred scheme is that elutriant is 60% methanol-water isocratic elution/50% methanol-water isocratic elution/40% methanol-water and 60% methanol-water gradient elution/30% methanol-water and 70% methanol-water gradient elution.
Foregoing method, preferred scheme be, when carrying out wash-out with elutriant, eluent flow rate is 3-8 times of column volume per hour (preferably per hour 5 times of column volumes).
The present invention extracts the method for separating the soybean isoflavones monomeric compound from soybean, extract obtained soybean isoflavones kind compound content is high, and impurity is very low, and this advantage can be known from soybean isoflavone HPLC figure (Fig. 1) and find out.In addition, also have following advantage:
(1) resulting soybean isoflavone soybean isoflavones kind compound content is high, and impurity is low, is conducive to follow-up being further purified.
(2) through separation and purification can obtain the multiple high-purity soybean isoflavone monomeric compounds such as glycitin, daidzin, Genistoside, glycitein, daidzein, genistein simultaneously once the step.Do not need as prior art sample to be carried out separating for several times, only need a separating step can obtain multiple high-purity compound, method is simple to operate, and efficient is high, and process cycle is short, saves reagent, has reduced production cost.
(3) do not use the organic solvents such as chloroform large to environmental hazard, environmental protection in the purge process.
(4) set up easy, efficient, as to realize a goal of the invention approach, optimized the condition (eluent components and type of elution) of chromatography method, purity and efficient are all greatly improved.
Description of drawings
Fig. 1 is the high-efficient liquid phase chromatogram of soybean isoflavone.
Fig. 2 is embodiment 1 thin layer chromatography figure.
Fig. 3 is embodiment 2 thin layer chromatography figure.
Fig. 4 is embodiment 3 thin layer chromatography figure.
Fig. 5 is embodiment 4 thin layer chromatography figure.
Embodiment
Describe technical scheme of the present invention in detail below in conjunction with embodiment and accompanying drawing, but protection domain is not by this restriction.Equipment used or raw material all can obtain from market among the embodiment.Used Superose 12 is available from General Electric (China) Medical Group, and agents useful for same is all available from Tianjin reagent four factories, and institute's water is deionized water.
Embodiment:Skimmed soy beans dregs of beans 1000 gram is with 60% alcohol immersion 8 hours, with 60% ethanol percolate extraction of 10 times of volumes.Use the double-deck diacolation post with the chuck heat tracing when diacolation extracts, medium temperature is 50 ℃, the percolate flow velocity be 3 times of column volumes/hour.Percolate spraying drying after being recycled to without alcohol obtains the extract meal, and the extract meal with 90% acetone refluxing extraction, condensing crystal, is obtained soybean isoflavone.
With the soybean isoflavone dissolve with methanol, upper Superose 12 chromatographies, be that elutriant carries out wash-out with methanol-water, eluent flow rate is 5 times of column volumes per hour, effluent liquid is detected at 254 nm by UV-detector, collect the target components cut, the cut concentrating under reduced pressure with obtaining namely obtains the monomeric compound that will separate.
The contriver adopts different types of elution by using the methyl alcohol of different concns, has optimized the purification condition of realizing the object of the invention, relevant experimental result such as following table one:
Table one column chromatography condition
? | The |
Embodiment | |
1 | 60% methanol-water isocratic elution |
Embodiment 2 | 50% methanol-water isocratic elution |
Embodiment 3 | 40% methanol-water and 60% methanol-water gradient elution |
Embodiment 4 | 30% methanol-water and 70% methanol-water gradient elution |
Fig. 2 is thin layer chromatography figure when selecting embodiment 1 system, and as seen from Figure 2, glycitin and daidzin divide too busy to get away, and other component separating is good, can obtain four kinds of compositions such as Genistoside, glycitein, daidzein, genistein.Fig. 3 is the thin layer chromatography figure when with embodiment 2 system, and as seen from Figure 3, glycitin and separating of daidzin make moderate progress, but unsatisfactory, and other component separating is good, and in addition, the required disengaging time of genistein is oversize, does not elute in 500 minutes.Fig. 4 is the thin layer chromatography figure when selecting embodiment 3 system.Thin layer chromatography figure when Fig. 5 is embodiment 4 system, by Fig. 4,5 as seen, each component separating is good, and disengaging time also can be accepted.Collect each peak component according to color atlas, behind the recovery solvent, can obtain corresponding high-purity compound.According to
1H-NMR and
13The test of C-NMR is identified gained peak component, has obtained glycitin, daidzin, Genistoside, glycitein, daidzein, 6 soybean isoflavones compositions of genistein.
Through HPLC area normalization method analytical test, the purity of resulting each component of embodiment 1-4 is very high, all more than 98%.
Claims (1)
1. from soybean, extract the method for separating the soybean isoflavones monomeric compound, it is characterized in that step is:
(1) preparation of soybean isoflavone: skimmed soy beans dregs of beans 1000g, with 60% alcohol immersion 8 hours, 60% ethanol percolate extraction with 10 times of volumes, percolate spraying drying after being recycled to without alcohol obtains the extract meal, the extract meal with 90% acetone refluxing extraction, condensing crystal, is obtained soybean isoflavone; Use the double-deck diacolation post with the chuck heat tracing when diacolation extracts, medium temperature is 50 ℃, the percolate flow velocity be 3 times of column volumes/hour;
(2) column chromatography purification: with the soybean isoflavone dissolve with methanol, upper Superose12 post carries out the chromatographic separation purifying, be that elutriant carries out wash-out with methanol-water, eluent flow rate is 5 times of column volumes per hour, effluent liquid is detected at 254nm by UV-detector, collect the target components cut, with the cut concentrating under reduced pressure that obtains, obtain glycitin, daidzin, Genistoside, glycitein, daidzein, 6 soybean isoflavones compositions of genistein; Type of elution is 30% methanol-water and 70% methanol-water gradient elution.
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CN103788156B (en) * | 2014-02-24 | 2015-07-01 | 聊城大学 | Method for separating and purifying soy isoflavone monomer compounds |
CN104546998B (en) * | 2014-10-20 | 2017-09-29 | 长春大学 | The production method of high genistin content isoflavones |
CN105614071A (en) * | 2015-12-28 | 2016-06-01 | 成都华罗生物科技有限公司 | Soybean isoflavone extract and extraction method thereof |
CN105533210A (en) * | 2016-01-21 | 2016-05-04 | 成都华罗生物科技有限公司 | Medicine composition for improving piglet production performance and preparation method and application of medicine composition |
CN107102069A (en) * | 2017-03-14 | 2017-08-29 | 上海优久生物科技有限公司 | A kind of method of 6 kinds of isoflavones in quick measure feed addictive |
CN108503675A (en) * | 2018-07-11 | 2018-09-07 | 黑龙江八农垦大学 | The method for extraction and purification and application thereof of Soybean Leaves glycitin |
CN110156856B (en) * | 2019-06-17 | 2020-09-29 | 山东师范大学 | Method for preparing glycitein by using primary scum for treating soybean milk water |
CN110540557B (en) * | 2019-06-17 | 2020-12-08 | 山东禹王生态食业有限公司 | Method for preparing glycitein by taking soybean navel powder as raw material |
CN110128386B (en) * | 2019-06-17 | 2021-05-11 | 山东师范大学 | Preparation method of pure soyabean extract |
Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN1528758A (en) * | 2003-09-30 | 2004-09-15 | 四川大学 | Method for separating major monomeric compoent of soybean isoflavone |
CN101781278A (en) * | 2010-02-26 | 2010-07-21 | 浙江大学 | Method for separating genistein from isoflavone genin mixture |
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JPS62126186A (en) * | 1985-11-28 | 1987-06-08 | Tsumura Juntendo Inc | Production of isoflavone derivative |
JP2000095792A (en) * | 1998-09-21 | 2000-04-04 | Showa Sangyo Co Ltd | Acquisition of isoflavone composition comprising genistin |
-
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CN1528758A (en) * | 2003-09-30 | 2004-09-15 | 四川大学 | Method for separating major monomeric compoent of soybean isoflavone |
CN101781278A (en) * | 2010-02-26 | 2010-07-21 | 浙江大学 | Method for separating genistein from isoflavone genin mixture |
Non-Patent Citations (4)
Title |
---|
G. Fiechter,等.Characterization of isoflavone composition in soy-based nutritional supplements via ultra performance liquid chromatography.《Analytica chimica acta》.2010,第672卷(第1-2期),72-78. * |
北京医学院,等.3.提取方法.《中草药成分化学》.人民卫生出版社,1980,(第1版),6-7. * |
琼脂糖凝胶分离纯化大黄等中药有效成分的研究;黄艳艳;《聊城大学硕士学位论文》;20080827;正文第25-38页 * |
黄艳艳.琼脂糖凝胶分离纯化大黄等中药有效成分的研究.《聊城大学硕士学位论文》.2008,正文25-38. |
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