CN100362015C - Method for separating and extracting protein and/or enzyme from ion liquid - Google Patents
Method for separating and extracting protein and/or enzyme from ion liquid Download PDFInfo
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- CN100362015C CN100362015C CNB2006100385160A CN200610038516A CN100362015C CN 100362015 C CN100362015 C CN 100362015C CN B2006100385160 A CNB2006100385160 A CN B2006100385160A CN 200610038516 A CN200610038516 A CN 200610038516A CN 100362015 C CN100362015 C CN 100362015C
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Abstract
The present invention discloses a method for extracting and separating proteins and/or enzymes from ionic liquids. The method comprises the steps of the pretreatment of raw materials, the addition of an extraction agent containing the ionic liquid, the centrifugation, the filtration, the purification and the concentration of the extraction liquid. The method which adopts the ionic liquid as the main component of the extraction agent has the advantages of high efficient and quick extraction and separation, safety and repeated use of the extraction agent, activity maintenance of the proteins and the enzymes obtained by extraction and separation, integration of both steps of extraction and separation into one and simple and convenient operation.
Description
Technical field
The present invention relates to the isolating method of a kind of protein and/or enzyme extraction, particularly relate to a kind of method with ionic liquid extract isolated protein and/or enzyme.
Background technology
Protein and enzyme are crucial living matters, and its range of application is very extensive, comprise every field such as medicine, food, chemical industry, makeup, daily necessities, agricultural.The extraction separation of protein and enzyme is the problem that people attach great importance to all the time, its extraction and separation method is a lot, as organic molecular solvent extracting and separating, membrane sepn, inverse micelle abstraction, supercritical fluid extraction separation, ion-exchange separation, precipitate and separate etc.Traditional method is to utilize most of protein all water soluble, rare salt, diluted acid or alkaline solution, the protein of minority and contaminated with lipid then is dissolved in the organic solvents such as ethanol, acetone, butanols, therefore, can adopt different solvents extraction separation and protein purification and enzyme.But these methods always exist some shortcomings more or less, see Table 1.
The deficiency of table 1 protein (comprising enzyme) traditional extraction process
| Project | The extraction with aqueous solution method | The organic solvent extraction method |
| The deficiency that exists | 1. protein (enzyme) can be degraded in leaching process and be lost; 2. temperature is wayward, easily causes protein (enzyme) inactivation; 3. can not extract fat-soluble protein; 4. be difficult to obtain high purity product; 5. operate cumbersome. | 1. can not extract water soluble protein; 2. this solvent usually is inflammable, explosive, toxic property; 3. easily cause protein (enzyme) inactivation; 4. be only applicable to cold condition; 5. operation is complicated, and range of application is narrow. |
(Room temperature ionic liquids RTILs), often is called ionic liquid for short to ionic liquid at room temperature, claims the room temperature melting salt again.Be meant the material of only being made up of ion that is in a liquid state under near temperature room temperature or the room temperature, it is as a kind of environment amenable solvent, is familiar with by people and accepts.Form ion liquid positively charged ion and be generally organic cation (as alkyl imidazole positively charged ion, alkyl pyridine positively charged ion, quaternary ammonium alkyl ion, Wan Ji quaternary phosphine ion etc.), negatively charged ion can be inorganic anion or organic anion (as [PF
6]-, [BF
4]-, [AlCl
4]-, [CF
3SO
3]-etc.).Since 1914 find first ionic liquid-nitro ethamine, particularly 20th century the mid-80 so far during this period of time, ionic liquid research in a lot of fields all presents very active situation.Ion liquid research can be divided into two stages substantially: the research before (1) 1992 year mainly concentrates on chloro aluminate class ionic liquid, this class ionic liquid is comparatively responsive to air, moisture, be easy to hydrolysis, need the anaerobic waterless operation, working conditions is comparatively harsh, thereby has limited its development and application; (2) research of being carried out in recent years mainly concentrates on the research and development to the insensitive non-chloro aluminate class ion liquid system of empty G﹠W.
More traditional fluent meterial is compared, and ionic liquid has following impayable advantage: (1) generally can not become steam, and is not volatile, thereby in use can not cause very big pollution to environment; (2) have bigger equilibrium temperature scope (100～200 ℃, part scholar thinks that ion liquid liquid journey can reach 400 ℃), the electrochemical stability potential window of better chemical stability, broad, thermostability and fabulous oxidation-resistance; (3) physical and chemical performance such as ion liquid solvability, liquid state scope, depend on that the yin, yang ionic constitutes and pairing, can be as required, directed design ion liquid system, regulate its solvability to inorganics, water, organism and polymkeric substance, make many chemical reactions be able in homogeneous phase, finish, and reactor volume greatly reduces, and its acidity can transfer to hyper acidic.(4) ionic liquid can repeated multiple times use.(5) it also can be used as the enzyme catalyst in the novel material production process.Jamaal Wilkes finds also that recently ionic liquid can also be used to handle junked tire, reclaims polymkeric substance wherein.The nearest achievement in research of scientist also shows, can extract carbonic acid gas in the industrial gaseous waste effectively with ionic liquid.(6) density is big, does not dissolve each other with many solvents.When with another solvent extraction target compound, by action of gravity, just can realize separating of solvent and target compound, thereby guarantee the efficient use of solvent and catalyzer.Using at present some more fields mainly contains: the medicine in sensitive material in plating galvanic deposit in catalysis and organic synthesis field, compartment analysis field, the electrochemistry and electro-conductive material and electrochemical device, the field of novel and lubricant etc., the life science is synthetic with screening etc.
The application of ionic liquid in extracting and separating: separate the recovery product of purifying is the difficult problem of synthetic chemistry always.The water extracting and separating only is applicable to the wetting ability product, and distillation technique is not suitable for the product of volatility difference yet, with an organic solvent can cause crossed contamination again.Therefore, design safety, eco-friendly isolation technique seems more and more important.Ionic liquid is suitable as and separates the solvent of purifying owing to have unique physicochemical property very much.Especially on liquid-liquid extraction separated, the ionic liquid physical efficiency was dissolved some organic compound, mineral compound and organometallic compound, simultaneously with most organic solvent unmixings, was suitable as very much the new medium of liquid-liquid extraction.Present research majority is to extract the organic or inorganic thing with ionic liquid from the aqueous solution, and the different selected ionic liquids of extract are also different.
During with the ion liquid abstraction volatile organic matter, because of the ionic liquid steam forces down, Heat stability is good with the extraction phase heating, can be driven extract out of after extraction is finished, and ionic liquid is easy to recycle.Huddlestou etc. use and water-insoluble ionic liquid 1-methyl-3-butyl imidazole hexafluorophosphate ([BMIm] [PF
6]) derivative of benzene extraction such as toluene, aniline, phenylformic acid, chlorobenzene etc. from water.The result shows that the partition ratio of multiple organic bronsted lowry acids and bases bronsted lowry is opposite fully under the different pH, and this is directly related with the protonation state of solute in the solution.The size of partition ratio fluctuates up and down 1.0 with the variation of the pH of solution.By the pH of regulator solution, can control certain solute in two alternate distribution state, thereby improve the controllability of extraction process.Ionic liquid also can be applicable to industrial production as extraction agent, and as in the important method acetone-ethanol-butylic fermentation method of producing biofuel, the ethanol in the product is removed by distillation, and butanols often adopts the method for extracting and separating to be separated because boiling point is higher.Extraction agent commonly used comprises tributylphosphine, 1-octanol and ether, and their partition ratio is respectively 14.6,7.6 and 4.1.Though tributylphosphine has higher partition ratio, organism of fermentation there is big toxicity.Faddev etc. find to use [BMIm] [PF
6] and [OMIm] [PF
6] (1-methyl-3-octyl group imidazoles hexafluorophosphate) during as the extraction agent of butanols, partition ratio can reach 25.7 and 55.3 respectively, and extraction agent does not almost have toxicity to organism of fermentation.These studies show that the huge advantage of ionic liquid in separating organic matters, but utilize the report of ionic liquid extract isolated protein and/or enzyme still rare at present.
Summary of the invention
The objective of the invention is provides a kind of method with ionic liquid extract isolated protein and/or enzyme at above-mentioned technical problem.
The objective of the invention is to realize by following technical measures:
The isolating method of a kind of protein and/or enzyme extraction comprises the following steps:
A. raw materials pretreatment;
B. add and contain ion liquid extraction agent in right amount;
C. centrifugal, the filtration of extraction liquid, filtrate is purified, concentrates to get final product.
The isolating method of described protein and/or enzyme extraction, wherein the method for raw materials pretreatment is removal of impurities, regulates that pH value is that 3-10, adjusting moisture are 0%～90%, does not contain poisonous objectionable impurities and avoid solar radiation in the controlled temperature-5 ℃～85 ℃, environment.
The isolating method of described protein and/or enzyme extraction, wherein raw material is organism or its tissue, chemical preparations, biological products, biological chemistry goods, food, medicine and the intermediate thereof that contains protein and/or enzyme.
The isolating method of described protein and/or enzyme extraction, wherein containing ion liquid extraction agent is to prepare by following method: configuration proportion is: by weight percentage, at extraction agent intermediate ion liquid portion be: 15%～99%, the auxiliary portion is: 1%～85%, and ionic liquid and auxiliary share sum are 100%; Ionic liquid, auxiliary mixed get final product.
The isolating method of described protein and/or enzyme extraction, wherein ionic liquid adopts non-chloro aluminate class ionic liquid.
The isolating method of described protein and/or enzyme extraction, wherein ionic liquid is one or more in glyoxaline ion liquid, guanidine class ionic liquid, quaternary phosphine class ionic liquid, quaternary amines ionic liquid or the pyridines ionic liquid.
The isolating method of described protein and/or enzyme extraction, wherein ion liquid density is 1.1～1.6g/cm
3, be linear relationship between density and the temperature.
The isolating method of described protein and/or enzyme extraction, wherein auxiliary is one or more in neutral solvent, acid solvent, basic solvent or the acidity regulator.
The isolating method of described protein and/or enzyme extraction, wherein neutral solvent is water, methyl acetate, ethyl acetate, ethyl formate, methyl propionate, ethyl propionate, ethyl butyrate, methyl alcohol, ethanol, propyl alcohol, acetone, hexane in the auxiliary; Acid solvent is phosphoric acid, formic acid, acetate, propionic acid, butyric acid; Basic solvent is ethamine, propylamine, pyridine, picoline; Acidity regulator is yellow soda ash, sodium bicarbonate, phosphoric acid, SODIUM PHOSPHATE, MONOBASIC, Sodium phosphate dibasic, sodium hydrogen phosphate, monoammonium sulfate, hydrochloric acid, sodium hydroxide.
The isolating method of described protein and/or enzyme extraction, wherein the method for purifying adopts electrophoretic method or column chromatography.
Beneficial effect of the present invention:
The present invention has adopted novel green extracting and separating agent---ionic liquid, and this material has a series of outstanding superiority: 1. colourless, odorless, almost non-toxic, and non-volatile, almost there is not vapour pressure.2. good thermostability is arranged, the thermal stable temperature wide ranges.Nonflammable, viscosity is little, good fluidity.Generally about 400 ℃, equilibrium temperature is interval about 300 ℃ when being in a liquid state, and is that general molecular solvent is incomparable for initial decomposition temperature.3. the soluble material scope of ionic liquid is wide, many inorganic, organic, the organo-metallic of solubilized, synthetic or natural macromolecular material, biologically active substance, and solubleness is relatively large.4. ionic liquid has bigger polarity Modulatory character, high 1～2 order of magnitude of the viscosity of viscosity ratio common organic solvents or water, but still have good flowability, and can form two-phase or heterogeneous system, be fit to make to separate solvent or constitute reaction-separation coupling new system.5. ion liquid density is generally 1.1～1.6g/cm
3, be linear relationship between density and the temperature.6. ion liquid perveance is 10 under the normal temperature
-1Ω
-1M
-1About, ion is little usually, and viscosity is low, and density is big, then good conductivity.Wide (about the 4V) of its electrochemical stability potential window is that common solvent does not possess.7. ionic liquid is not only on solvent or catalyst performance as a kind of green and environment-friendly solvent or catalyzer and is had environmental safety, and self biodegradable be the environmental safety material.8. ionic liquid and some ion etc. also have stronger coordinative activity.
Method with ionic liquid extract isolated protein and/or enzyme provided by the invention; it is low and protect the shortcoming of poor activity that dangerous, the extraction agent that had both overcome traditional extraction recycles and reuses rate, also avoided the severe condition and the hidden danger of supercritical extraction method.Extraction separation of the present invention is efficient, quick, and extraction agent safety can repeatedly be used repeatedly, and it is active that the protein of extraction separation (enzyme) can keep, and has extraction is united two into one advantages such as convenience simple to operation with separating.
Description of drawings
Fig. 1 is a process flow sheet of the present invention.
Embodiment
Come by the following examples the present invention is made a more detailed description.Described extraction and separation process flow process and extraction agent preparation method are only for reference, but do not mean that limitation of the present invention.
Extraction agent in following examples (ionic liquid+auxiliary) can use more than 10 times repeatedly.Adopting stirrer, mulser or ultrasonic wave that each component is mixed during the preparation extraction agent gets final product.The fragmentation of unreceipted actual conditions and step in the enforcement, separate, method such as filtration, purifying, ultrasonic oscillation, lipidated protein mensuration is the technology that those of ordinary skills know.
Embodiment 1
Extract plant protein.With the 1000g soybean meal is raw material, carry out raw materials pretreatment, comprise fragmentation, sieve, removal of impurities, with distilled water to regulate the raw material water content be 1.0%～3.0% (w/w), controlled temperature 15 ℃～45 ℃, transfer pH 6.5～9.5 with sodium bicarbonate, note not containing poisonous objectionable impurities in the environment and avoiding solar radiation, in order to avoid protein active is impacted.Add extraction agent (by weight percentage, extraction agent is by the six alkyl guanidine fluoroborates that account for extraction agent gross weight 88.5%, 4% ethyl butyrate, 5% ethanol and 2.5% acetic acid are formed, mixing standby before the use) 5000g (divides and extracts for 3 times, the consumption of extraction agent is 40% of a total amount for the first time, second, be 30% three times), under the vibrator vibration, extract 3～5h, then on the whizzer in the centrifugation of carrying out under the 5000r/min about 10min, filter, and filtrate regulated moisture to 5 ± 0.5% once more, pH 3.5～5.5, and then it is centrifugal, filter, filtrate is crude product, carry out purifying (method of purifying is method known to a person of ordinary skill in the art, and is as follows) with electrophoretic method again, the purifying material concentrates with vacuum freeze-drying method and makes the soy-protein product.After measured, this method extraction rate reached is more than 85.6%, and the purity of proteinaceous product (Kjeldahl determination with GB/T5009.5-1996 GB regulation is measured) reaches more than 97.8%.
Embodiment 2
Extract plant protein.With the 1000g wheat is raw material, carry out pre-treatment, comprising pulverizing, sieving, regulate the raw material water content with distilled water is that 60 ± 5% (w/w), controlled temperature are 40 ± 5 ℃, transfer pH 6.5～9.5 with sodium hydrogen phosphate, notes not containing poisonous objectionable impurities in the environment and avoiding solar radiation.Add extraction agent (by weight percentage, extraction agent is by the six alkyl guanidine fluorophosphate ionic liquids that account for extraction agent gross weight 88.5%, 10% methyl propionate, 1% hexane and 0.5% butyric acid are formed, mixing standby before the use) 4000g (divides and extracts for 3 times, the consumption of extraction agent is 40% of a total amount for the first time, second, be 30% three times), under the vibrator vibration, extract 3～5h, then on the whizzer under 5000r/min centrifugal 10min, filter, and to filtrate adjusting moisture to 5 ± 1%, pH 4.5 ± 1, centrifuging then, filtrate is crude product, carry out purifying (method of purifying is method known to a person of ordinary skill in the art, and is as follows) with column chromatography again, the purifying material concentrates with vacuum freeze-drying method and makes the soy-protein product.After measured, this method extraction rate reached is more than 87%, and the purity of proteinaceous product (Kjeldahl determination with GB/T5009.5-1996 GB regulation is measured) reaches more than 98.1%.
Embodiment 3
Extract animal protein.With 500g pig blood meal is raw material, removes impurity, ultrasonication again, sodium bicarbonate adjusting pH 7.5～9.0 earlier, regulates 10 ℃～15 ℃ of moisture 0.1%～2.0% (w/w), temperature, notes not containing poisonous objectionable impurities in the environment and avoiding solar radiation.Add extraction agent (by weight percentage, extraction agent is by the ionic liquid that the 1-methyl-3-octyl group imidazoles hexafluorophosphate is formed of the 1-methyl-3-butyl imidazole a tetrafluoro borate that accounts for extraction agent gross weight 25% and 55%, 3% SODIUM PHOSPHATE, MONOBASIC, 7% monoammonium sulfate, 1% ethyl formate, 9% acetone is formed, mix standby before the use), the consumption of extraction agent is that 10 times of raw materials quality (divide and extract for 3 times, the consumption of extraction agent is 40% of a total amount for the first time, second, be 30% three times), under sonic oscillation, extract 20min, then on the whizzer in the centrifugation of carrying out under the 6000r/min about 10min, filter, and to filtrate adjusting moisture to 10 ± 2%, pH 4.5～6.0,5% silica powder helps precipitation, and then centrifuging, filtrate is crude product, carry out purifying with electrophoretic method again, the purifying material concentrates with vacuum freeze-drying method and makes the soy-protein product.After measured, this method extraction rate reached is more than 90.2%, and the purity of proteinaceous product (Kjeldahl determination with GB/T5009.5-1996 GB regulation is measured) reaches more than 98.3%.
Embodiment 4
Extract animal protein.With 500g pig blood meal is raw material, removes impurity, ultrasonication again, sodium hydrogen phosphate and sodium phosphate earlier and regulates 10 ℃～15 ℃ of pH 8.5～9.0, moisture 0.1%～2.0% (w/w), temperature, notes not containing poisonous objectionable impurities in the environment and avoiding solar radiation.Add extraction agent (by weight percentage, the ionic liquid that extraction agent is made up of the N-butyl-pyridinium a tetrafluoro borate of the 1-octyl group-3-Methylimidazole hexafluorophosphate that accounts for extraction agent gross weight 65% and 15%, 5% sodium hydrogen phosphate, 5% monoammonium sulfate, 3% methyl acetate, 7% acetone is formed, mix standby before the use), the consumption of extraction agent is that 5 times of raw materials quality (divide and extract for 3 times, the consumption of extraction agent is 40% of a total amount for the first time, second, be 30% three times), under sonic oscillation, extract 80min, then on the whizzer in the centrifugation of carrying out under the 4000r/min about 10min, filter, and filtrate regulated moisture to 8 ± 3%, acidity pH 4.5～5.5,3% silica powder helps precipitation, and then centrifuging, filtrate is crude product, carry out purifying with column chromatography again, the purifying material makes proteinaceous product with vacuum freeze-drying method.After measured, this method extraction rate reached is more than 91.2%, and the purity of proteinaceous product (Kjeldahl determination with GB/T5009.5-1996 GB regulation is measured) reaches more than 98.5%.
Embodiment 5
Extract enzyme.Cellulase solution with 100g vigor 〉=100U/g is a raw material, the coarse filtration removal of impurities, is that 10% phosphoric acid is regulated pH 3.5～4.5, moisture 60 ± 5% (w/w), 25 ℃～30 ℃ of temperature, noted not containing poisonous objectionable impurities and avoiding solar radiation in the environment with concentration.Add extraction agent (by weight percentage, extraction agent is by accounting for the 1-methyl-3-butyl imidazole hexafluoro borate ion liquid of extraction agent gross weight by 50%, 7% SODIUM PHOSPHATE, MONOBASIC, 8% monoammonium sulfate, 5% formic acid and 30% methyl alcohol, mix standby before the use) 500g, under the ultrasonator vibration, extract, extraction time 60min, then in the centrifugation of under 5000r/min, carrying out 10min on the whizzer, filter, filtrate is regulated moisture to 20 ± 5% once more, acidity pH 4.5～5.0, centrifuging then, filtrate is crude product, carries out purifying with column chromatography again, and the purifying material concentrates with vacuum freeze-drying method and makes the cellulase product.After measured, this method extraction rate reached is more than 84.4%, and the purity of cellulase product (measurement result of measuring cellulase activity with filter paper sugar filter paper method is represented) reaches more than 95.0%.
Embodiment 6
Extract Escherichia coli protein.With intestinal bacteria (E.coli) MC1061 fermented liquid (OD after the 100g fermentation
260Be 0.5～0.6) be raw material, pre-treatment (comprises removal of impurities, regulate pH 7～8, moisture 80 ± 5%, 15 ℃～20 ℃ of temperature, note not containing poisonous objectionable impurities in the environment and avoiding solar radiation) after, add extraction agent (by weight percentage, extraction agent is by accounting for the 1-ethyl-3-Methylimidazole trifluoroacetate ionic liquid of extraction agent gross weight by 65%, 10% yellow soda ash, 15% ethyl acetate, 10% ethanol, mix standby before the use) 300g, under the ultrasonator vibration, extract, extraction time 50min, then in the centrifugation of under 5000r/min, carrying out 20min on the whizzer, filter, filtrate is regulated moisture to 65 ± 5% once more, acidity pH 7.5～8.0, centrifugal then, filter, filtrate is crude product, carry out purifying with column chromatography again, the purifying material concentrates with vacuum freeze-drying method and makes the Escherichia coli protein product.After measured, this method extraction rate reached is more than 91.2%, and the purity of Escherichia coli protein product (Kjeldahl determination with GB/T5009.5-1996 GB regulation is measured) reaches more than 92.4%.
Embodiment 7
Extract terramycin.With the terramycin fermented liquid after the 100g fermentation is raw material, pre-treatment (comprises removal of impurities, regulate pH2.5～4, moisture 70 ± 5%, temperature-5 ℃～5 ℃, note not containing poisonous objectionable impurities in the environment and avoiding solar radiation) after, add extraction agent (by weight percentage, extraction agent is by accounting for the 1-ethyl-3-Methylimidazole ethyl sulfonate ionic liquid of extraction agent gross weight by 60%, 5% yellow soda ash, 15% ethyl acetate, 20% ethanol, mix standby before the use) 300g, under the ultrasonator vibration, extract, extraction time 50min, then in the centrifugation of under 6000r/min, carrying out 15min on the whizzer, filter, filtrate is regulated moisture to 35 ± 5% once more, acidity pH 5.5～6.0, centrifugal then, filter, filtrate is crude product, carry out purifying with column chromatography again, the purifying material concentrates with vacuum freeze-drying method and makes the terramycin product.After measured, this method extraction rate reached is more than 92.2%, and the purity of terramycin product (representing with National Standard Method GB/T14931.01-1996 high performance liquid chromatography (HLPC) method measurement result) reaches more than 93.6%.
Embodiment 8
Extract yeast protein.A-acetate lactic acid decarboxylase (aldc) fermented liquid (OD with the short and small bud bacillus of saccharomyces cerevisiae after the 100g fermentation
260Be 0.6～0.7) be raw material, pre-treatment (comprises removal of impurities, regulate pH 5.5～6.5, moisture 70 ± 5%, temperature-5 ℃～5 ℃, control environment and do not contain poisonous objectionable impurities and lucifuge) after, add extraction agent (by weight percentage, extraction agent is by accounting for the Tetrylammonium sulfonate ion liquid of extraction agent gross weight by 70%, 8% sodium bicarbonate, 15% methyl acetate, 7% picoline, mix standby before the use) 300g, under the ultrasonator vibration, extract, extraction time 60min, then in the centrifugation of under 6000r/min, carrying out 15min on the whizzer, filter, filtrate is regulated moisture to 35 ± 5% once more, acidity pH 5.5～6.0, and is centrifugal then, filter, filtrate is crude product, carry out purifying with column chromatography again, the purifying material concentrates with vacuum freeze-drying method and makes the yeast proteinaceous product.After measured, this method extraction rate reached is more than 92.2%, and the purity of yeast proteinaceous product (Kjeldahl determination with GB/T5009.5-1996 GB regulation is measured) reaches more than 93.6%.
Claims (9)
1. protein or the isolating method of enzyme extraction is characterized in that comprising the following steps:
A. raw materials pretreatment;
B. add and contain ion liquid extraction agent in right amount;
C. centrifugal, the filtration of extraction liquid, filtrate is purified, concentrates to get final product;
Wherein: the method for raw materials pretreatment is removal of impurities, regulate that pH value is that 3-10, adjusting moisture are 0%～90%, do not contain poisonous objectionable impurities and avoid solar radiation in the controlled temperature-5 ℃～85 ℃, environment.
2. protein according to claim 1 or the isolating method of enzyme extraction is characterized in that raw material is organism or its tissue, chemical preparations, biological products, biological chemistry goods, food, medicine and the intermediate thereof that contains protein or enzyme.
3. according to right 1 described protein or the isolating method of enzyme extraction, it is characterized in that containing ion liquid extraction agent is to prepare by following method: configuration proportion is: by weight percentage, at extraction agent intermediate ion liquid portion be: 15%～99%, the auxiliary portion is: 1%～85%, and ionic liquid and auxiliary share sum are 100%; Ionic liquid, auxiliary mixed get final product.
4. according to right 1 or 3 described protein or the isolating method of enzyme extraction, it is characterized in that ionic liquid is non-chloro aluminate class ionic liquid.
5. according to right 4 described protein or the isolating method of enzyme extraction, it is characterized in that ionic liquid is one or more in glyoxaline ion liquid, guanidine class ionic liquid, quaternary phosphine class ionic liquid, quaternary amines ionic liquid or the pyridines ionic liquid.
6. protein according to claim 5 or the isolating method of enzyme extraction is characterized in that ion liquid density is 1.1～1.6g/cm
3, be linear relationship between density and the temperature.
7. protein according to claim 3 or the isolating method of enzyme extraction is characterized in that auxiliary is one or more in neutral solvent, acid solvent, basic solvent or the acidity regulator.
8. protein according to claim 7 or the isolating method of enzyme extraction is characterized in that neutral solvent is water, methyl acetate, ethyl acetate, ethyl formate, methyl propionate, ethyl propionate, ethyl butyrate, methyl alcohol, ethanol, propyl alcohol, acetone or hexane in the auxiliary; Acid solvent is phosphoric acid, formic acid, acetate, propionic acid or butyric acid; Basic solvent is ethamine, propylamine, pyridine or picoline; Acidity regulator is yellow soda ash, sodium bicarbonate, phosphoric acid, SODIUM PHOSPHATE, MONOBASIC, Sodium phosphate dibasic, sodium hydrogen phosphate, monoammonium sulfate, hydrochloric acid or sodium hydroxide.
9. protein according to claim 1 or the isolating method of enzyme extraction is characterized in that the method for purifying adopts electrophoretic method or column chromatography.
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| CN102796163A (en) * | 2012-09-06 | 2012-11-28 | 南京财经大学 | Method for extracting and separating proteins from cake by using ionic liquid and enzyme process |
| CN106323705A (en) * | 2015-07-03 | 2017-01-11 | 中国科学院大连化学物理研究所 | Rapid pretreatment method of proteome sample based on ionic liquid |
| CN105146047A (en) * | 2015-10-19 | 2015-12-16 | 内蒙古清谷新禾有机食品集团有限责任公司 | Method for extracting buckwheat protein from buckwheat |
| CN105348369B (en) * | 2015-11-27 | 2019-01-25 | 中国科学院过程工程研究所 | A method of using pupa albumen in ionic liquid extract separation silkworm chrysalis |
| CN105777863B (en) * | 2016-03-24 | 2019-06-18 | 陈铭 | A method of the isolated polypeptide from glyoxaline ion liquid |
| CN108529732B (en) * | 2018-03-19 | 2021-05-25 | 河南师范大学 | Method for purifying iron-containing protein wastewater based on ionic liquid |
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|---|---|---|---|---|
| WO2005103070A1 (en) * | 2004-04-21 | 2005-11-03 | The University Of York | Affinity chromatography using ionic liquids |
| CN1587240A (en) * | 2004-07-16 | 2005-03-02 | 中国科学院过程工程研究所 | New method for preparing and separating integrately antibiotic medicine in ionic liquid double water phase |
| CN1660901A (en) * | 2004-12-24 | 2005-08-31 | 中国科学院长春应用化学研究所 | Method for preparing solution of keratin |
Non-Patent Citations (1)
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| 室温离子液体在化学合成和萃取分离中的应用. 刘晓庚等.化学与生物工程，第4期. 2004 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US8143381B2 (en) | 2006-11-17 | 2012-03-27 | Merck Patent Gmbh | Use of ionic liquids for membrane protein extraction |
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| CN1807446A (en) | 2006-07-26 |
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