CN101381721B - Immobilized lipase and preparation method thereof - Google Patents
Immobilized lipase and preparation method thereof Download PDFInfo
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- CN101381721B CN101381721B CN2008102230765A CN200810223076A CN101381721B CN 101381721 B CN101381721 B CN 101381721B CN 2008102230765 A CN2008102230765 A CN 2008102230765A CN 200810223076 A CN200810223076 A CN 200810223076A CN 101381721 B CN101381721 B CN 101381721B
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Abstract
The invention discloses immobilized lipase and a method for preparing the same. The method for preparing the immobilized lipase comprises the following steps: evenly mixing lipase solution with polymer emulsion the weight of which is between 3 and 60 percent against that of the lipase solution, immersing a non-woven fabric by the obtained mixed liquid, and drying the mixed liquid to obtain the immobilized lipase, wherein the polymer emulsion is polyacrylate emulsion or the mixed emulsion containing polyacrylate and polyurethane as solutes. Experiments prove that the immobilized lipase has high catalysis performance and long service life, can still keep the catalysis efficiency above 90 percent after various continuous catalytic esterification reactions and transesterification reactions, and the required amount of the immobilized lipase for catalytically producing each ton of bio-diesel is less than or equal to 4 kilograms.
Description
Technical field
The present invention relates to a kind of immobilized lipase and preparation method thereof.
Background technology
The immobilization technology of biological enzyme is one of core content of enzyme engineering, it successfully overcome resolvase in industrial applications poor stability, poor heat resistance, can not reuse and be difficult for and the isolating shortcoming of product.Immobilized enzyme is all improving a lot aspect stability and the catalytic activity, and its reuse can reduce production cost again greatly.The immobilization technology of enzyme can improve the microenvironment of enzyme Journal of Molecular Catalysis, is convenient to the contact of enzyme-to-substrate and the diffusion of product.In addition, fixation support has been given certain physical behavior of immobilized enzyme and mechanical property, and it can be conveniently used in the various bio-reactor, is convenient to realize the serialization and the automatization of catalyzed reaction.The application of immobilized enzyme in organic catalytic system more shown its superiority especially.The process for fixation of the immobilization effect of enzyme and carrier character and enzyme has very big relation, and enzyme immobilization method commonly used at present mainly comprises: absorption method, covalent attachment method, entrapping method and crosslinking etc.Several diverse ways respectively have the relative merits of himself, and the enzyme of different sorts and character also often uses different process for fixation, usually use several process for fixation in case of necessity simultaneously.
Lipase is the wider a kind of enzyme of industrial application, it can be on oil-water interface catalytic hydrolysis, esterification, transesterificationization, lactone is synthetic, polypeptide is synthetic and the multiple reactions such as fractionation of chipal compounds, the lipase of different sources is also often different on catalytic activity and catalysis specificity.The lipase immobilization technology is a decision lipase key in application, and bibliographical information has had the various fixed method to be used for lipase immobilization.The fixation support that lipase is commonly used comprises natural porous material, each gellike, synthetic resins and composite modification material etc.On the market the commercial immobilized lipase of many kinds has been arranged now, wherein most widely used is the particulate state immobilized lipase, and it is carrier with synthetic resins, and the enzyme molecule is realized immobilization by covalently bound mode.This immobilized lipase enzyme activity is higher, and the enzyme carrying capacity is big, long service life, but complex process, and cost is higher, and the immobilization process enzyme is lived and is lost serious (generally losing more than 60%), and technology is also very high to enzyme self purity requirement.This type of immobilized enzyme price is very high, generally more than thousand yuan/kilogram, have in addition surpass ten thousand yuan, be unsuitable for the market large-scale application.
Summary of the invention
The purpose of this invention is to provide a kind of immobilized lipase and preparation method thereof.
The preparation method of immobilized lipase provided by the present invention is that 3%～60% polymer emulsion with lipase solution and described lipase solution weight mixes, and will obtain the mixed solution immersion non-woven fabrics, being fixed of drying lipase; Described polymer emulsion is polyacrylate dispersion or contains polyacrylic ester and the mixed emulsion of urethane, and polyacrylic ester and urethane ratio are not limit in the described mixed emulsion, and the scope of preferred mass ratio is 0.5:1～4:1.Described polymer emulsion and described lipase solution blended proportional range can be 5%-32%, 5%-8%, 5%-12%, 8%-24%, 24%-32% or 12%-32%.
In the described method, the lipase activity scope of described lipase solution is 4000～20000U/g solution, and vigor is preferably 6000～15000U/g solution.
In the described method, described lipase solution adds stablizer earlier with before polymer emulsion mixes in described lipase solution; Described stablizer is PEG or gelatin; The mass percentage concentration of the adding of described stablizer is 1～5 ‰.
In the described method, described polyacrylic ester is so that two or more is that monomer carries out the multipolymer that polyreaction obtains in butyl acrylate, ethyl propenoate, methyl acrylate, Isooctyl acrylate monomer, vinyl acrylate, methyl methacrylate, vinyl acetate between to for plastic and the vinylformic acid; Described polyacrylic ester is preferably the multipolymer that butyl acrylate, ethyl propenoate and methyl methacrylate obtain than the ratio polyreaction that is 2～3:2～4:2～6 in massfraction.
In the described method, described urethane is the segmented copolymer of polyisocyanates and oligomeric polyols; Described oligomeric polyols is polyether glycol or polyester glycol, and the average molecular mass of described oligomeric polyols is 0.3～3kDa; Described polyisocyanates is aromatic isocyanate or aliphatic diisocyanate; Described aromatic isocyanate is preferably tolylene diisocyanate or '-diphenylmethane diisocyanate, and described aliphatic diisocyanate is preferably methylene diisocyanate six times.
The material of described non-woven fabrics is terylene, polypropylene fibre, polyamide fibre or viscose fiber.When described immobilized lipase is that described non-woven fabrics is preferably 30～80g/m when being used for the immobilized lipase of bio-reactor
2Filament polypropylene fibre non-woven fabric.
In the described method, described drying is oven dry below 80 ℃ or dries naturally.
In the described method, the quality percentage composition that contains of described immobilized lipase is below 10%.Reasonably water content helps the catalytic activity and the stability of being maintained fixed lipase.
In the described method, the solid content of described polymer emulsion is 20%～60%, and wherein preferred solid content is 35%～50%.Solid content determine by the control emulsion when synthetic that monomer is formed and content is realized.
In the described method, the band liquid quality of described non-woven fabrics dipping is 100%～900% of a described non-woven fabrics quality.
In the described method, described lipase liquid is the liquid that contains lipase that obtains behind the solution of lipase configuration or the microbial fermentation (as microbial fermentation solution, microbial fermentation concentrated liquor or microbial fermentation solution centrifugal after supernatant liquor).
Immobilized lipase provided by the present invention is the immobilized lipase of method for preparing.
Preparation method's technological operation of immobilized lipase of the present invention is simple, is convenient to industrialization continuity production, is fixation support with cheap non-woven fabrics, and production cost reduces greatly.Method of the present invention is less demanding to the character such as purity of lipase, can both carry out immobilization to fermented liquid, fermentation centrifugate, fermentation concentrated solution and high-purity enzyme powder.Polymer emulsion of the present invention is to the not influence of vigor of enzyme liquid; Enzyme liquid does not destroy the emulsification proterties of polymer emulsion self.The selected polymer emulsion of the present invention has film forming characteristic, can provide good microenvironment for the lipase Journal of Molecular Catalysis.Method of the present invention makes the lipase molecule be fixed on the non-woven fabrics fiber by complex methods such as absorption, crosslinked and embeddings, this combination is firm especially, being immobilized lipase is that catalytic activity or repeated use number of times all improve a lot, the lipase activity loss of immobilization process is less, in 25%, also different by the immobilized enzyme vigor that different vigor enzyme liquid make, vigor is generally between 10000～130000U/g.
Experimental results show that fixed lipase catalyzed performance height of the present invention, long service life, after it carried out repeatedly catalytic esterification and transesterification continuously, catalytic efficiency still can remain on more than 90%, immobilized enzyme amount≤4kg that its catalytic production biofuel per ton is required.
Description of drawings
Fig. 1 is the immobilized lipase of the present invention esterification yield of catalysis palmitinic acid and different monooctyl ester repeatedly.
Fig. 2 is the repeatedly esterification yield of catalysis oleic acid and methyl alcohol of immobilized lipase of the present invention
Fig. 3 is a repeatedly transesterification transformation efficiency of immobilized lipase of the present invention.
Embodiment
The described method of following embodiment if no special instructions, is ordinary method.
Percentage composition among the following embodiment if no special instructions, is the quality percentage composition.
The preparation of embodiment 1, immobilized lipase of the present invention and compliance test result thereof
1, the preparation of immobilized lipase of the present invention
With candiyeast Candida sp 99-125 enlarged culturing in the 1000L fermentor tank, substratum comprises dregs of beans 50g/L, soya-bean oil 40g/L, K
2HPO
41g/L, KH
2PO
41g/L, defoamer 3g/L under 28 ℃, stir radius 0.2m, stirring frequency 175r/min.Get fermented liquid, show that with olive oil hydrolysis method (industry standard QB/T1803-1993) detection the lipase hydrolysis vigor of fermented liquid is the 4000U/g fermented liquid.
Get the above-mentioned Candida sp 99-125 fermented liquid that obtains, through 2200g centrifuging and taking supernatant liquor, add PEG6000, making its mass percentage concentration is 2 ‰, fully after the dissolving, 5% the polyacrylate dispersion (solid content 35% that adds fermented liquid supernatant liquid quality, viscosity is 305cP, polyacrylic ester in the emulsion by weight fraction than being the butyl acrylate of 1:1:2, the multipolymer that ethyl propenoate and methyl methacrylate polymerization reaction obtain, polymkeric substance relative molecular mass distribution range is 10～50kDa (weight-average molecular weight 36kDa)) stir 10 minutes mixings, then with terylene non-woven fabric (45g/m
2) be carrier impregnation in liquid after 20 minutes, take out, to obtain liquid carrying rate (carrying the ratio of liquid and non-woven fabrics weight) be 300% non-woven fabrics by padding, place 50 ℃ dry to water content be 4%, i.e. being fixed lipase.Show that with the detection of olive oil hydrolysis method the vigor of this immobilized lipase is 10000U/g.Divided by the enzyme yield alive in the total activity calculating immobilization process of the adsorbed enzyme liquid of non-woven fabrics, above-mentioned preparation immobilized lipase enzyme yield alive is 76% with the total activity of measuring.
2, immobilized lipase of the present invention is used for the synthetic compliance test result of iso-octyl palmitate
The immobilized lipase of step 1 preparation is carried out repeatedly catalytic esterification continuously according to following reaction system:
0.200g the immobilized enzyme of step 1 preparation, the 1.000g palmitinic acid, 0.559g isooctyl alcohol (the acid alcohol mol ratio is 1:1.1), the 5mL normal hexane, 40 ℃, the 160r/min shaking table reacted 12 hours.
After each catalyzed reaction, used immobilized enzyme is taken out, after normal hexane is washed, be reentered into new above-mentioned same system and continue reaction.After each reaction, adopt acid base titration to determine esterification yield, record reaction esterification yield still remains on the reaction times more than 80%.
The result as shown in Figure 1, the result shows, the immobilized lipase endonuclease capable of step 1 preparation is the esterification of catalysis palmitinic acid and isooctyl alcohol well, carry out catalyzed reaction continuously 50 times, its catalytic esterification rate is still very high, the 36th esterification yield of continuous catalytic reaction is that 90.22%, the 47 secondary response esterification yield is 80.99%.Illustrate that immobilized lipase of the present invention not only has good catalytic efficiency, and have very long work-ing life.
The preparation of embodiment 2, immobilized lipase of the present invention and compliance test result thereof
1, the preparation of immobilized lipase of the present invention
Precipitate lipase with organic solvent precipitation method from the fermented liquid that the step 1 of embodiment 1 obtains, lipase makes the enzyme powder after drying.
After the enzyme powder added an amount of water dissolution, the lipase powder lysate that obtains detected the lipase hydrolysis vigor 5000U/g enzyme liquid that shows enzyme liquid with olive oil hydrolysis method (industry standard QB/T1803-1993).
Get the above-mentioned enzyme powder lysate that obtains, add PEG6000, making its mass percentage concentration is 4 ‰, and fully after the dissolving, (viscosity is 200cP, and solid content is 46% to add 8% polyacrylate dispersion of enzyme liquid quality; Polyacrylic ester is the multipolymer that butyl acrylate, ethyl propenoate, Isooctyl acrylate monomer and vinylformic acid obtain for the 3:2:1:1 polyreaction according to mass ratio, polymkeric substance relative molecular mass distribution range is that 20～50kDa (weight-average molecular weight 40kDa) stirs 10 minutes mixings, then with polyamide fibre non-woven fabrics (42g/m
2) for carrier impregnation contains in the polymer emulsion of enzyme liquid after 15 minutes in this, take out, to obtain liquid carrying rate (carrying the ratio of liquid and non-woven fabrics weight) be 360% non-woven fabrics by padding, place 70 ℃ dry to water content be 5%, i.e. being fixed lipase.Show that with the detection of olive oil hydrolysis method the vigor of this immobilized lipase is 16000U/g.Divided by the enzyme yield alive in the total activity calculating immobilization process of the adsorbed enzyme liquid of non-woven fabrics, above-mentioned preparation immobilized lipase enzyme yield alive is 79% with the total activity of measuring.
2, immobilized lipase of the present invention is used for the synthetic compliance test result of Witconol 2301
The immobilized lipase of step 1 preparation is carried out repeatedly catalytic esterification continuously according to following reaction system:
0.200g the immobilized enzyme of step 1 preparation, 1.680g oleic acid, 0.191g methyl alcohol (dividing 3 addings), the 5mL normal hexane, 40 ℃, the 180r/min shaking table reacted 12 hours.
After each catalyzed reaction, used immobilized enzyme is taken out, after the normal hexane washing, be reentered into new above-mentioned same system and continue reaction.After each reaction, adopt acid base titration to determine esterification yield, record reaction esterification yield still remains on the reaction times more than 90%.
The result as shown in Figure 2, the result shows, the immobilized lipase endonuclease capable of step 1 preparation is the esterification of catalysis oleic acid and methyl alcohol well, carries out catalyzed reaction continuously 25 times, its catalytic esterification rate remains on more than 90%.Illustrate that immobilized lipase of the present invention not only has good catalytic efficiency, and have long work-ing life.
The preparation of embodiment 3, immobilized lipase of the present invention and compliance test result thereof
1, the preparation of immobilized lipase of the present invention
Fermented liquid is from 5 tons of fermentor tanks, and bacterial strain and substratum stir radius 0.35m, stirring frequency 136r/min with the step 1 of embodiment 1.
Get new following jar fermented liquid, this fermented liquid lipase hydrolysis vigor is the 8000U/g fermented liquid, remove by filter impurity such as thalline, add gelatin, making its mass percentage concentration is 3 ‰, fully dissolving, (this emulsion is that solute is that mass ratio is the polyacrylic ester of 2:1 and the mixed emulsion of urethane to add 12% polymer emulsion of fermented liquid quality, viscosity is 280cP, total solid content 36%, wherein polyacrylic ester is a butyl acrylate, the multipolymer that ethyl propenoate and methyl methacrylate obtain for the ratio polyreaction of 3:4:6 by ratio of weight and the number of copies, polymkeric substance relative molecular mass distribution range are 20～50kDa (weight-average molecular weight 30kDa); Urethane is the multipolymer that tolylene diisocyanate and polyether glycol (the relative molecular mass distribution range is 1.8～2.6kDa (weight-average molecular weight 2.2kDa)) polymerization obtain, the molecular weight distribution scope is 20～60kDa (weight-average molecular weight 45kDa)), stir mixed in 15 minutes after, respectively with 33g/m
2Long filament spunbond polypropylene non-woven fabrics, 35g/m
2, 40g/m
2And 45g/m
2Short-fibre hot-rolled nonwoven polypropylene fabric immerse after 15 minutes, take out, be 500% by padding the control liquid carrying rate, place 70 ℃ dry to water content be 7%, promptly obtain with 33g/m respectively
2Long filament spunbond polypropylene non-woven fabrics, 35g/m
2, 40g/m
2Or 45g/m
2Short-fibre hot-rolled nonwoven polypropylene fabric be the immobilized lipase of carrier.Show with the detection of olive oil hydrolysis method, with 33g/m
2Long filament spunbond polypropylene non-woven fabrics, 35g/m
2, 40g/m
2Or 45g/m
2Short-fibre hot-rolled nonwoven polypropylene fabric be that the vigor of the immobilized lipase of carrier is respectively 32000U/g, 31000U/g, 31000U/g, 32500U/g, prepare this with 33g/m
2Long filament spunbond polypropylene non-woven fabrics, 35g/m
2, 40g/m
2Or 45g/m
2Short-fibre hot-rolled nonwoven polypropylene fabric be that the immobilized lipase enzyme of the carrier yield of living is respectively 80%, 78%, 79%, 81%%.
2, immobilized lipase of the present invention is used for the compliance test result of transesterification
With step 1 preparation with 33g/m
2Long filament spunbond polypropylene non-woven fabrics, 35g/m
2, 40g/m
2Or 45g/m
2Short-fibre hot-rolled nonwoven polypropylene fabric be that the immobilized lipase of carrier carries out repeatedly catalysis transesterification respectively continuously, this transesterificationization is shaken bottle reaction system: 2g oil (good fortune is the one-level soybean oil near the house), the 4ml sherwood oil, 0.15g water, 279 μ l methyl alcohol (dividing 3 addings), the immobilized enzyme of 0.6g step 1 preparation, 40 ℃, the 170r/min shaking bath reacted 12 hours.After each reaction, immobilized enzyme is reentered into new above-mentioned system and continues reaction after sherwood oil cleans.All use the content of gas chromatography determination fatty acid methyl ester after each reaction, calculate and change esterification yield.
The result as shown in Figure 3, successive reaction 12 times, step 1 preparation with 33g/m
2Long filament spunbond polypropylene non-woven fabrics, 35g/m
2, 40g/m
2Or 45g/m
2Short-fibre hot-rolled nonwoven polypropylene fabric be that the immobilized lipase of carrier all still keep to change esterification yield all more than 90%, each immobilized enzyme catalysis activity remains unchanged substantially.Immobilized enzyme shows good tolerability to methyl alcohol, glycerine etc.Among Fig. 3,1 is 33g/m
2Long filament spunbond polypropylene non-woven fabrics be the result of continuous 12 the catalysis transesterifications of immobilized lipase of carrier, 2-4 are respectively 35g/m
2, 40g/m
2And 45g/m
2Short-fibre hot-rolled nonwoven polypropylene fabric be the result of continuous 12 the catalysis transesterifications of immobilized lipase of carrier.With immobilized lipase airing after normal hexane cleans of 12 batches of above-mentioned reactions, placed 2 days, continue to repeat above-mentioned reaction, continue 8 batches of reactions, change esterification yield and still remain on 90%.
The result show the preparation of this step 1 with 33g/m
2Long filament spunbond polypropylene non-woven fabrics, 35g/m
2, 40g/m
2Or 45g/m
2Short-fibre hot-rolled nonwoven polypropylene fabric be that the immobilized lipase of carrier all can continuous 20 batches of above-mentioned transesterifications of catalysis, and change esterification yield all more than 90%.
The preparation of embodiment 4, immobilized lipase of the present invention and compliance test result thereof
1, the preparation of immobilized lipase of the present invention
Fermented liquid is from 5 tons of fermentor tanks, and bacterial strain and fermentation condition are with the step 1 of embodiment 3.Fermented liquid is through the centrifugal supernatant liquor that gets of 2000g, and recording the supernatant liquor lipase activity is the 6500U/g supernatant liquor.Get centrifuged supernatant and add PEG6000 and gelatin, making PEG6000 quality percentage composition is 2 ‰, and gelatin quality percentage composition is 1 ‰.(this emulsion is to contain the mixed emulsion that mass ratio is 3:1 polyacrylic ester and urethane, and viscosity is 360cP, always solid content 42% to add 24% polymer emulsion of fermentation centrifugate quality; Wherein polyacrylic ester is the multipolymer that butyl acrylate, ethyl propenoate, vinyl acrylate and methyl methacrylate obtain for the 2:2:1:3 polyreaction according to mass ratio, and polymkeric substance relative molecular mass distribution range is 40～60kDa (weight-average molecular weight 49kDa); Urethane is the multipolymer that '-diphenylmethane diisocyanate and polyester glycol (the relative molecular mass distribution range is 0.8～1.6kDa (weight-average molecular weight 1.2kDa)) polymerization obtain, the molecular weight distribution scope is 30～50kDa (weight-average molecular weight 37kDa)), stir mixed in 20 minutes after, immerse grammes per square metre 38g/m
2The polypropylene filament yarn non-woven fabrics, take out after 10 minutes, to obtain liquid carrying rate (carrying the ratio of liquid and non-woven fabrics weight) be 900% non-woven fabrics by padding, natural airing, the mensuration water content is 9%, i.e. being fixed lipase.Show that with the detection of olive oil hydrolysis method the vigor of this immobilized lipase is 42000U/g, preparing this immobilized lipase enzyme yield alive is 75%.
2, immobilized lipase of the present invention is used for the compliance test result of fish oil ethyl esterization
Measurement result shows, step 1 prepared immobilized enzyme can continuous 26 batches of catalysis fish oil the ethyl ester reaction, and transformation efficiency is more than 92%.This enzyme can be used for the modification of fish oil and the production preparation of ethyl ester DHA.
The preparation of embodiment 5, immobilized lipase of the present invention and compliance test result thereof
1, the preparation of immobilized lipase of the present invention
Fermented liquid is from 5 tons of fermentor tanks, and bacterial strain and fermentation condition are with the step 1 of embodiment 3.Fermented liquid through 2000r/min centrifugal supernatant liquor, supernatant liquor through the rolling ultra-filtration membrane concentrate concentrated solution.
Get the centrifugal concentrated solution of fermentation, vigor 15000U/g, add gelatin and PEG8000, make the mass percentage concentration of gelatin and PEG8000 be 2 ‰, fully after the dissolving, (this emulsion is that to contain mass ratio be the polyacrylic ester of 1:1 and the mixed emulsion of urethane to add 32% polymer emulsion of the centrifugal concentrated solution quality of fermentation, total solid content 44.6%, viscosity is 420cP, wherein polyacrylic ester is a butyl acrylate, ethyl propenoate, methyl acrylate, the multipolymer that vinyl acetate between to for plastic obtains than polyreaction according to the quality of 3:1:4:10, polymkeric substance relative molecular mass distribution range is 40～52kDa (weight-average molecular weight 45kDa); Urethane is the multipolymer that six methylene diisocyanates and polyester glycol (the relative molecular mass distribution range is 1.2～2.1kDa (weight-average molecular weight 1.8kDa)) polymerization obtain, the molecular weight distribution scope is 60～110kDa (weight-average molecular weight 80kDa)), stir mixed in 20 minutes after, immerse grammes per square metre 60g/m
2The viscose fiber non-woven fabrics, take out after 10 minutes, to obtain liquid carrying rate (carrying the ratio of liquid and non-woven fabrics weight) be 600% non-woven fabrics by padding, 36 ℃ of warm braws dry to water content be 6%, i.e. being fixed lipase.Show that with the detection of olive oil hydrolysis method the vigor of this immobilized lipase is 68000U/g, preparing this immobilized lipase enzyme yield alive is 76%.
2, immobilized lipase of the present invention is used for the compliance test result of production of biodiesel
The immobilized lipase of step 1 preparation is carried out repeatedly catalysis biological diesel production continuously, production of biodiesel reaction system: at cylindrical reactor (diameter 15cm, high 0.6m) add the immobilized enzyme of step 1 preparation in, 330g carries out some batch of material liquid successive reactions; Every batch reaction adds the raw material of storage tank: 40kg purifies sewer oil, 2kg water, 5.6L methyl alcohol (dividing 6 addings), reaction raw materials is after mixing, through the pump cycling stream through above-mentioned cylindrical reactor, flow velocity is 350g/min, and the circulation retention time is 8hr, in cylindrical reactor and the storage tank with 40 ℃ of heating in water bath.After finishing a batch reaction feed liquid is emitted, add virgin material in the storage tank and proceed reaction same as described above, each reaction back is calculated and is surveyed transformation efficiency with gas Chromatographic Determination methyl esters content, when reaction conversion ratio at 80% stopped reaction when following.Calculate the biofuel amount that successive reaction obtains altogether according to transformation efficiency.Above-mentioned successive reaction can be carried out 3-4 batch.
The result shows, enzyme amount≤4kg that process for producing biofuel per ton is required.This immobilized enzyme is that Production by Enzymes biofuel (particularly being raw material with the sewer oil) has been opened up wide road, has crucial meaning.
Claims (9)
1. the preparation method of an immobilized lipase is that lipase solution and the polymer emulsion that is the 3%-60% of described lipase solution weight are mixed, and will obtain the mixed solution immersion non-woven fabrics, being fixed of drying lipase; Described polymer emulsion is that polyacrylate dispersion or solute are the mixed emulsion of polyacrylic ester and urethane;
Described lipase solution adds stablizer earlier with before polymer emulsion mixes in described lipase solution; Described stablizer is PEG and/or gelatin;
Described polyacrylic ester is so that two or more is that monomer carries out the multipolymer that polyreaction obtains in butyl acrylate, ethyl propenoate, methyl acrylate, Isooctyl acrylate monomer, vinyl acrylate, methyl methacrylate, vinyl acetate between to for plastic and the vinylformic acid;
Described urethane is the segmented copolymer of polyisocyanates and oligomeric polyols; Described oligomeric polyols is polyether glycol or polyester glycol, and the relative molecular mass of described oligomeric polyols is 0.3～3kDa; Described polyisocyanates is aromatic isocyanate or aliphatic diisocyanate; Described aromatic isocyanate is tolylene diisocyanate or '-diphenylmethane diisocyanate, and described aliphatic diisocyanate is six methylene diisocyanates;
The material of described non-woven fabrics is terylene, polypropylene fibre, polyamide fibre or viscose fiber;
The mass ratio of polyacrylic ester and urethane is 0.5: 1～4: 1 in the described mixed emulsion.
2. method according to claim 1 is characterized in that: lipase content is a 4000-20000U/g solution in the described lipase solution.
3. method according to claim 1 is characterized in that: described polyacrylic ester is that butyl acrylate, ethyl propenoate and methyl methacrylate are 2～3: 2～4 in the massfraction ratio: the multipolymer that 2～6 ratio polyreaction obtains; The mass percentage concentration of the adding of described stablizer is 1～5 ‰; Described drying is oven dry below 80 ℃ or dries naturally.
4. according to any described method among the claim 1-3, it is characterized in that: the quality percentage composition that contains of described immobilized lipase is below 10%.
5. according to any described method among the claim 1-3, it is characterized in that: the solid content of described polymer emulsion is 20%～60%.
6. method according to claim 5 is characterized in that: the solid content of described polymer emulsion is 35%～50%.
7. according to any described method among the claim 1-3, it is characterized in that: the band liquid quality of described non-woven fabrics dipping is the 100%-900% of described non-woven fabrics quality.
8. method according to claim 1 is characterized in that: described lipase liquid is the liquid that contains lipase that obtains behind the solution of lipase configuration or the microbial fermentation.
9. the immobilized lipase of any described method preparation among the claim 1-8.
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| CN101724621A (en) * | 2009-12-17 | 2010-06-09 | 北京凯泰新世纪生物技术有限公司 | Microcarrier immobilized lipase and preparation method thereof |
| CN102863629A (en) * | 2012-09-28 | 2013-01-09 | 南开大学 | Preparation method of lipase microemulsion with high heat stability |
| CN103275814B (en) * | 2013-06-07 | 2015-02-18 | 华东理工大学 | Method for preparing biodiesel without by-product of glycerin by utilization of high acid value waste oil |
| CN107779444A (en) * | 2016-08-31 | 2018-03-09 | 北京超纳生物质化工技术研究院 | For preparing the catalyst of low-carbon chain triglyceride and the method using the catalyst preparation low-carbon chain triglyceride |
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| JP2886902B2 (en) * | 1989-08-08 | 1999-04-26 | 日本バイリーン株式会社 | Enzyme-containing material and immobilized enzyme using the same |
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