CN102728410B - Magnetic light-enzyme composite catalyst and preparation method thereof - Google Patents
Magnetic light-enzyme composite catalyst and preparation method thereof Download PDFInfo
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- CN102728410B CN102728410B CN201210193905.6A CN201210193905A CN102728410B CN 102728410 B CN102728410 B CN 102728410B CN 201210193905 A CN201210193905 A CN 201210193905A CN 102728410 B CN102728410 B CN 102728410B
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Abstract
The invention relates to a magnetic light-enzyme composite catalyst and a preparation method thereof. The magnetic light-enzyme composite catalyst is laccase immobilized on magnetic nano TiO2 processed by surface functionalization, particularly laccase immobilized on magnetic nano TiO2 particles modified by copper ion chelating. According to the invention, the magnetic light-enzyme composite catalyst provided by the invention realizes the transmission of electrons between laccase and TiO2 in the catalytic oxidation process, solves the problems of slow speed of receiving electrons by laccase from the substrate and slow combination between photoinduced electrons and O2; by using the synergism of enzyme catalysis and photocatalysis to raise catalytic oxidation efficiency, the oxidation of substrates with high oxidation-reduction potential and macromolecule substrates can be realized; the catalyst has superparamagnetism, can be conveniently and effectively controlled and separated under the effect of external magnetic field, the preparation process is simple, and the catalyst is suitable for large scale production.
Description
Technical field
The present invention relates to a kind of novel magnetic light enzyme composite catalyst, particularly, the present invention relates to a kind of by magnetic light-enzyme composite catalyst of optically catalytic TiO 2 and the enzymatic electron transmission link coupling of laccase and preparation method thereof.
Background technology
Laccase (Benzenediol: oxidoreducing enzyme, EC l.10.3.2) is a kind of polyphenol oxidase of cupric, and the ascorbic acid oxidase in plant, mammiferous ceruloplasmin belong to blue blue multicopper oxidase family together.Laccase by 4 electronics by O
2be catalysed and reduced into water, and along with the oxidation of some phenols substrates.The catalytic reaction of the laccase oxidation material such as phenols, aromatic amine has reaction condition gentleness, selective height, advantage that catalytic efficiency is high.Laccase is one of oxidoreducing enzyme with application prospect most, is mainly used in the degraded etc. of association with pulp bleaching in organic synthesis, the degraded of environmental contaminants and dye decolored, biological detection and electrochemical analysis, paper industry and wastewater treatment, fibrous raw material.
But, in the catalytic oxidation process of laccase, electronics is from substrate to the rate-limiting step that the transmission of laccase is this reaction, become Laccase Catalyzed and need one of the key problem solved (Oxidation of Phenols, Anilines, and Benzenethiols by Fungal Laccases:Correlation between Activity and RedoxPotentials as Well as Halide Inhibition, Feng Xu, Biochemistry, 1996,35:7608-7614).In addition, due to free laccase easy in inactivation, cannot to reclaim and product price is higher, cause the rising of its application cost.Magnetic carrier is utilized to carry out enzyme immobilizatio, not only greatly can improve the stability of enzyme, catalytic activity, selective and anti-inhibition, be conducive to immobilised enzymes be separated from reaction system and reclaim, be suitable for large-scale continuous operation, and motion mode and the direction of Magnetic control immobilised enzymes can be utilized, improve the catalytic efficiency of enzyme.
Utilize the research of semiconductor light-catalyst process each pollutant waste water to have a large amount of reports, its degraded object relates to multiple important organic compounds and the part inorganic compounds such as phenols, dyestuff, polycyclic aromatic hydrocarbon.Photochemical catalytic oxidation take N-type semiconductor as catalyst, wherein TiO
2active high, harmless, there is stable chemical property, ultraviolet shielded effect, fast light burn into indissoluble, and there is darker valence-band level, some chemical reactions absorbed heat can be made in and acceleration accomplished by the surface of light radiation, in addition nontoxic, cost is low, so be widely used as the catalyst of photocatalysis oxidation reaction.
The research of magnetic titanium dioxide nano particle and photocatalysis characteristic thereof has bibliographical information, and the doping of magnetic core improves TiO
2to the absorption efficiency (preparation of magnetic titanium dioxide nano particle and photocatalysis performance thereof, Li Heping etc., Journal of Molecular Catalysis, 2008,6 (22), 555-560) of visible ray.At enzyme and TiO
2on synergy process phenols wastewater, Meizler etc. adopt Surface coating TiO
2bead carry out the immobilization of horseradish peroxidase, its objective is and pass through TiO
2photocatalysis destructive enzyme oxidation formed polymer, reduce its inhibitory action to enzyme (A novel glass support for the immobilization and UV-activation of horseradishperoxidase for treatment of halogenated phenols, Alon Meizler., etc., ChemicalEngineering Journal, 2011,172:792-798).Existing research shows, O in water
2with TiO
2surface electronic (e
-) be combined into TiO
2light-catalysed rate-limiting step, and the effect adopting the method passing into excessive oxygen or loaded inert metal to accelerate electron transmission is limited (Photocatalytic oxidation of organicsin water using pure and silver-modified titanium dioxide particles, Vamathevan V., etc., Journal of Photochemistry and Photobiology A:Chemistry, 2002,148:233-245.).
The oxidation-reduction potential of laccase is at 0.5-0.8V, and wherein the oxidation-reduction potential of Trametes versicolor laccase is 0.8V; For the aldehydes matter of laccase oxidation, its oxidation-reduction potential mainly concentrates 0.5-0.8V, this explains substrate electron becomes reason (Kinetic and biochemicalproperties of high and low redox potential laccases from fungal and plant origin from rate-limiting step to laccase natural mode intermediate, Marco Frasconi, etc., Biochimica et Biophysica Acta, 2010,1804:899-908).And at TiO
2in light-catalysed process, O
2be good electron acceptor, but be adsorbed in TiO
2the O on surface
2with electronics (e
-) combination be decided by TiO
2the density on shallow top layer, the oxidation-reduction potential on this shallow top layer is 0.1-0.3V, and O
2change O into
2-oxidation-reduction potential be-0.28V, this is O
2with electronics (e
-) combination reason (TiO slowly
2photocatalysis:A Historical Overview and Future Prospects, KazuhitoHashimoto, ProspectsJapanese Journal of Applied Physics, 2005,44:8269-8285).The people such as Jia report and adopt the immobilized laccase of Bio-Glas and TiO
2photocatalysis synergy carries out the research that phenol catalysis can improve point catalytic effect, think that enzymatic and photocatalysis are carried out respectively, producing synergistic reason may be that the free radical metal enzymatic pathway acceleration enzymatic produced in photocatalytic process causes, although the method achieves Laccase Catalyzed and TiO
2light-catalysed synergy, but fail the rate-limiting step problem that solves in each self-catalysis, and immobilised enzymes and TiO
2not easily reclaim (Degradation of high concentration2,4-dichlorophenol by simultaneous photocatalytic – enzymatic process usingTiO2/UV and laccase, Jingbo Jia, etc., Journal of Hazardous Materials, 2012,205-206:150-155).
Summary of the invention
For laccase enzymatic and TiO in prior art
2the rate-limiting step problem existed respectively in photocatalysis, an object of the present invention is to provide a kind of magnetic light-enzyme composite catalyst, and described catalyst adopts magnetic Nano TiO
2as fixation support, form laccase and magnetic Nano TiO by paint enzyme immobilizatio
2complex, laccase plays the effect of electron transmission in phenol catalysis process, and it is directly from TiO
2surface accepts electronics (e
-) be water by reducing molecular oxygen, accelerate H
+with e
-separation, solve laccase and accept the slow and light induced electron of electron velocity and O from substrate
2in conjunction with problem slowly.
Magnetic light-enzyme composite catalyst of the present invention is for being immobilized in magnetic Nano TiO
2on laccase, described magnetic Nano TiO
2for through surface-functionalized magnetic Nano TiO
2.
Preferably, laccase of the present invention and magnetic Nano TiO
2mass ratio be 1:1-1:20(w/w), such as 1:1.1,1:1.6,1:2.7,1:3.0,1:4.3,1:5.1,1:5.7,1:6.2,1:6.9,1:7.4,1:8.0,1:8.4,1:8.8,1:9.2,1:9.9,1:11,1:12.8,1:14.3,1:15.7,1:17.2,1:18.8,1:19.2 etc.As laccase and magnetic Nano TiO
2mass ratio when being less than 1:1, the Electron Transfer of laccase is weak, and catalytic performance is bad; As laccase and magnetic Nano TiO
2mass ratio when being greater than 1:20, photocatalysis effect is not good.Especially as laccase and magnetic Nano TiO
2mass ratio be 1:3-1:15(w/w), preferred 1:5-1:10(w/w) time, the catalytic effect of preparation-obtained magnetic light-enzyme composite catalyst is better.
Magnetic Nano TiO of the present invention
2for the nano-TiO of magnetic loading
2, the carrier of the magnetic loading that those skilled in the art can obtain all can be used for the present invention, such as Fe
3o
4, γ-Fe
2o
3or spinels material such as NiFe
2o
4, MnFe
2o
4deng.Meanwhile, magnetic Nano TiO
2preparation method also have a lot, those skilled in the art have the ability to obtain magnetic Nano TiO completely
2preparation method, typical but non-limiting example has: in sol-gel process, high speed ball-milling method, reverse micelle method or Hydrolyze method any a kind or at least 2 in combination.
Magnetic Nano TiO of the present invention
2carrier be preferably Fe
3o
4, and, described magnetic Nano TiO
2middle Fe
3o
4content be 1-4wt%, such as 1.1wt%, 1.3wt%, 1.8wt%, 2.0wt%, 2.2wt%, 2.9wt%, 3.1wt%, 3.7wt%, 3.9wt% etc.As magnetic Nano TiO
2middle Fe
3o
4content be less than 1wt%, then magnetic effect is not obvious; Work as Fe
3o
4content when being greater than 4wt%, affect magnetic Nano TiO
2catalytic effect.
Magnetic Nano TiO of the present invention
2preparation method adopt method (the Comparison of magnetic-manometer titanium dioxide/ferriferous oxide(TiO of existing bibliographical information
2/ Fe
3o
4) composite photocatalyst prepared by acid-sol and homogeneousprecipitation methods.Jinyuan Chen, Journal of Materials Science, 2010,45:6018 – 6024) prepare magnetic Nano TiO
2nano particle.
Magnetic Nano TiO is better immobilized in order to laccase can be made
2on, the present invention is preferably by magnetic Nano TiO
2change copper ion huge legendary turtle into and close the magnetic Nano TiO modified
2particle.Of the present invention by magnetic Nano TiO
2change copper ion huge legendary turtle into and close the magnetic Nano TiO modified
2the method of particle has a variety of, and those skilled in the art can obtain by consulting pertinent literature, and the process that the present invention chooses is by magnetic Nano TiO
2first by imino group grafting agent, by magnetic Nano TiO
2change the magnetic Nano TiO with imino group functional group into
2nano particle, then by with copper ion solution hybrid reaction, obtain copper ion huge legendary turtle close magnetic Nano TiO
2nano particle.
Two of object of the present invention is to provide a kind of method preparing magnetic light-enzyme composite catalyst.Described method is by magnetic Nano TiO
2particle is modified through Silanization reaction, graft reaction and chelating copper ions and is obtained surface-functionalized magnetic Nano TiO
2, then by Laccase Immobilization at surface-functionalized magnetic Nano TiO
2on, obtain magnetic light-enzyme composite catalyst.Particularly, the preparation method of magnetic light-enzyme composite catalyst of the present invention comprises the steps:
(1) Silanization reaction solution is prepared;
(2) Silanization reaction is carried out;
(3) graft reaction is carried out;
(4) chelating copper ions is carried out;
(5) laccase curing reaction is carried out.
Step of the present invention (1) institute Silanization reaction solution comprises silane reagent, monohydric alcohol, magnetic Nano TiO
2nano particle and water.
Described silylating reagent is selected from the combination of any a kind or at least 2 kinds in 3-r-chloropropyl trimethoxyl silane, dimethyldichlorosilane, trichloromethyl silane, and described combination is 3-r-chloropropyl trimethoxyl silane/dimethyldichlorosilane, trichloromethyl silane/3-r-chloropropyl trimethoxyl silane, 3-r-chloropropyl trimethoxyl silane/dimethyldichlorosilane/trichloromethyl silane etc. such as; More preferably 3-r-chloropropyl trimethoxyl silane.
Described monohydric alcohol is the combination of any a kind or at least 2 kinds in the straight chain of C1-C5 or straight chained alkyl monohydric alcohol, the example of described monohydric alcohol has methyl alcohol, ethanol, normal propyl alcohol, n-butanol, n-amyl alcohol, isopropyl alcohol, the tert-butyl alcohol etc., the combination of monohydric alcohol of the present invention preferred any a kind or at least 2 kinds in methyl alcohol, ethanol, normal propyl alcohol further, described combination is methanol/ethanol, ethanol/normal propyl alcohol/methyl alcohol, normal propyl alcohol/ethanol etc. such as.Be 1 weight portion with the weight of silylating reagent, the addition of monohydric alcohol is 20-70 weight portion, and the parts by weight of such as monohydric alcohol are 22,27,31,39,45,52,57,63,67,69 etc., preferred 40-60 weight portion.
Described magnetic Nano TiO
2nano particle is the nano-TiO of magnetic-particle load
2, be preferably nano-TiO
2load is at Fe
3o
4on, and preferred vector Fe
3o
4account for magnetic Nano TiO
2the 1-4wt% of nanoparticle mass.Be 1 weight portion with the weight of silylating reagent, magnetic Nano TiO
2the addition of nano particle is 0.1-5 weight portion, such as magnetic Nano TiO
2the parts by weight of nano particle are 0.1,0.5,1.0,1.6,2.1,2.8,3.4,4.0,4.2,4.7,4.9 etc., preferred 0.5-2 weight portion.Magnetic Nano TiO of the present invention
2nano particle adopts method (the Comparison of magnetic-manometertitanium dioxide/ferriferous oxide(TiO of existing bibliographical information
2/ Fe
3o
4) composite photocatalyst prepared byacid-sol and homogeneous precipitation methods.Jinyuan Chen, Journal of MaterialsScience, 2010,45:6018 – 6024) prepare magnetic Nano TiO
2nano particle, its key step comprises: 1. synthesize the Fe that PEG modifies
3o
4magnetic fluid, 2. regulates magnetic fluid pH and adds Ti (SO
4)
2, 3. add urea and be hydrolyzed and neutralization reaction under condition of heating and stirring, synthesizing magnetic nano-TiO
2nano particle, 4. by magnetic Nano TiO
2nano particle is for subsequent use after Magneto separate, washing.
Preferably, magnetic Nano TiO of the present invention
2fe in nano particle
3o
4content is 1-4%(w/w), its content is regulated by solid content in regulation and control magnetic fluid, it is made to be scattered in deionized water through ultrasonic, form stable suspension, concentration is 2-10%(w/v), the mass/volume concentration of such as suspension is 2.1,2.4,2.9,3.5,4.1,4.8,5.3,6.0,7.5,8.7,9.4,9.8 etc., then puts into refrigerator and cooled and hides for subsequent use.
Described water is use for laboratory water, is experiment reagent well-known to those skilled in the art, in the preferred deionized water of water of the present invention, distilled water, redistilled water any 1.Be 1 weight portion with the weight of silylating reagent, the addition of water is 5-20 weight portion, such as whose parts by weight are 5.1,5.6,6.0,6.3,6.9,7.3,7.8,8.4,9.0,9.8,10.4,10.6,11.3,11.8,12.4,12.9,13.4,13.9,14.8,14.9 etc., preferred 10-15 weight portion.
Preferably, step (1) described Silanization reaction solution comprises following component by weight:
The parts by weight of each component of described Silanization reaction solution are all 1 weight portion for benchmark obtains with silylating reagent.
Further preferably, step (1) described Silanization reaction solution comprises following component by weight:
The parts by weight of each component of described Silanization reaction solution are all 1 weight portion for benchmark obtains with silylating reagent.
Preferably, the process for preparation of the described Silanization reaction solution of step (1) is: under room temperature, stirring condition, adds the silylating reagent of formula ratio, monohydric alcohol, magnetic Nano TiO in reaction vessel successively
2nano particle, water, adjust ph, obtained Silanization reaction solution; Described stir speed (S.S.) can select 200-300 rev/min, also can carry out the selection of other scopes according to actual conditions, such as 150 revs/min or 500 revs/min etc.
Preferably, step (1) described pH value is 3-5, such as 3.1,3.4,3.9,4.2,4.7,4.9 etc.PH value is less than 3 Silanization reaction solution facile hydrolysis, and unstable, condensation is not thorough yet; PH value is greater than 5, and monohydric alcohol is unstable, and Silanization reaction carries out not thorough.
Step of the present invention (2) described Silanization reaction is: Silanization reaction solution step (1) obtained, in protective atmosphere, carries out ultrasonic wave added reaction, then obtains magnetic Nano TiO through Magneto separate
2particle.
Preferably, the temperature of step (2) described Silanization reaction is 30-50 DEG C, such as 31-48 DEG C, 40-45 DEG C, 31 DEG C, 38 DEG C, 43 DEG C, 47 DEG C, 49 DEG C etc., temperature is too slow lower than 30 DEG C of reaction rates, temperature is too high higher than 50 DEG C of temperature, and reactant is easily polymerized; Reaction time is 20-60min, such as 25-57min, 29-42min, 21min, 28min, 33min, 37min, 47min, 59min etc., and the time is less than 20min, and reaction is not thorough, and the reaction time is after 60min, substantially reaches molecular balance.Consider the reaction temperature of the Silanization reaction of step (2), step of the present invention (2) is preferably carried out in heating water bath.Described Silanization reaction preferably carries out in the reactor that reflux is housed.
Preferably, the described ultrasonic power of step (2) is 60-120W, such as 61W, 67W, 75W, 82W, 88W, 96W, 102W, 107W, 113W, 117W etc., ultrasonic power is lower than 60W, reaction rate is too low, the required reaction time is oversize, and ultrasonic power is too high, the magnetic Nano TiO of the silylating reagent obtained
2structure bad.
Preferably; step (2) described protection gas is selected from the combination of any a kind or at least 2 kinds in nitrogen, helium, argon gas, neon, Krypton or xenon; described combination is nitrogen/helium, argon gas/nitrogen, helium/Krypton/argon gas, xenon/nitrogen/helium etc. such as; the combination of any a kind or at least 2 kinds in preferred nitrogen, helium or argon gas, further preferred nitrogen.
Step (3) described graft reaction is: join in graft reaction reagent by the product that step (2) obtains, and in protective atmosphere, stirs and carries out graft reaction, then obtain the magnetic Nano TiO with imino group functional group through Magneto separate
2nano particle.Preferably, described graft reaction reagent is iminodiacetic acid sodium solution; Described graft reaction carries out in heating water bath; Described graft reaction carries out in the reactor that reflux is housed; The rotating speed of described stirring is 100-300 rev/min, such as 110-290 rev/min, 100-287 rev/min, 101 revs/min, 105 revs/min, 110 revs/min, 143 revs/min, 167 revs/min, 187 revs/min, 190 revs/min, 196 revs/min etc.Described protection gas is selected from the combination of any a kind or at least 2 kinds in nitrogen, helium, argon gas, neon, Krypton or xenon; described combination is nitrogen/helium, argon gas/nitrogen, helium/Krypton/argon gas, xenon/nitrogen/helium etc. such as; the combination of any a kind or at least 2 kinds in preferred nitrogen, helium or argon gas, further preferred nitrogen.
Preferably, the pH scope 8-10 of the graft reaction described in step (3), such as 8.1,8.4,8.8,9.2,9.5,9.8,9.9 etc.; Reaction temperature 40-80 DEG C, such as 41 DEG C, 47 DEG C, 53 DEG C, 44 DEG C, 59 DEG C, 63 DEG C, 66 DEG C, 72 DEG C, 77 DEG C, 79 DEG C etc.; Reaction time 30-90min, such as 31min, 35min, 38min, 42min, 49min, 57min, 69min, 73min, 78min, 84min, 88min, 89min etc.
Preferably, the preferred 0.5-2%(w/v of concentration of the iminodiacetic acid sodium solution described in step (3)), such as 0.51%, 0.56%, 0.63%, 0.79%, 0.85%, 0.93%, 1.32%, 1.54%, 1.85%, 1.92%, 1.96% etc.; The magnetic Nano TiO that described step (2) obtains
2the concentration of nano particle is 1-4%(w/v), such as 1.1%, 1.6%, 2.1%, 2.9%, 3.5%, 4.0%, 1.23%, 1.38%, 1.72%, 2.34%, 3.96%, 3.41% etc.
Step (4) described chelating copper ions is: join in copper ion solution by the product that step (3) obtains, oscillating reactions under room temperature condition, obtains copper ion huge legendary turtle and close the magnetic Nano TiO modified after Magneto separate, washing
2nano particle.
Copper ion solution of the present invention be in order to by copper ion by magnetic Nano TiO
2nano particle carries out chelating modification, obtains copper ion huge legendary turtle and closes the magnetic Nano TiO modified
2nano particle.Described copper ion solution is the solution containing bivalent cupric ion of solubility, the combination of such as, any a kind or at least 2 kinds in copper chloride solution, copper-bath, copper nitrate solution or Schweinfurt green solution, described combination is copper chloride solution/copper-bath, copper nitrate solution/copper chloride solution, copper chloride solution/copper-bath/Schweinfurt green solution etc. such as.Preferably, step (4) described copper ion solution preferably sulfuric acid copper solution.
Preferably, the concentration of step (4) described copper ion solution is preferably 2-5%(w/v), such as 2.1%, 2.6%, 3.0%, 3.6%, 4.2%, 4.8%, 4.9% etc., such as concentration is 2.3%(w/v) copper-bath and/or concentration be 4.25%(w/v) copper chloride solution.
The described laccase curing reaction of step (5) is: copper ion huge legendary turtle step (4) prepared closes the magnetic Nano TiO modified
2particle mixes with laccase solution, and at ambient temperature, vibration, carries out Laccase Immobilization reaction, then after Magneto separate, washing, obtains magnetic light-enzyme composite catalyst.
Preferably, step (5) described laccase solution is the free laccase be dissolved in buffer solution, and the typical but non-limiting example of described buffer solution has citrate buffer solution, tartaric acid buffer, acetate buffer solution, phosphate buffer etc.The buffer solution that buffer solution of the present invention is well known to the skilled person, its process for preparation is simple to operation, in the present invention, select which kind of buffer solution laccase, the professional knowledge that those skilled in the art can grasp according to oneself and actual experiment condition are selected.
Preferably, step (5) described laccase is selected from the combination of in fungal laccase a kind or at least 2 kinds, typical but non-limiting example has Trametes laccase, Agaricus laccase, Podospora laccase, Rhizoctonia laccase, Polyporus laccase, Pholiota laccase, Coriolus laccase, Neurosporus crassa laccase or Plebia radiata laccase etc., described combination is Agaricus laccase/Podospora laccase such as, Rhizoctonia laccase/Plebia radiata laccase/Pholiota laccase, Coriolus laccase/Neurosporuscrassa laccase/Podospora laccase etc., described laccase molecular weight ranges is preferably 50-100kDa, such as 51kDa, 55kDa, 58kDa, 62kDa, 69kDa, 71kDa, 76kDa, 85kDa, 90kDa, 94kDa, 98kDa etc.
Preferably, the concentration of step (5) described laccase solution is 0.1-0.8g/L, such as 0.11g/L, 0.17g/L, 0.23g/L, 0.31g/L, 0.37g/L, 0.42g/L, 0.48g/L, 0.57g/L, 0.63g/L, 0.68g/L, 0.73g/L, 0.78g/L, 0.79g/L etc.
Preferably, step (5) described laccase and copper ion huge legendary turtle close the magnetic Nano TiO modified
2the mass ratio of particle is (0.1-0.4): 1, such as 0.11:1,0.13:1,0.18:1,0.25:1,0.34:1,0.41:1,0.42:1,0.31:1,0.37:1,0.39:1 etc.
Preferably, the pH value of the described laccase curing reaction of step (5) is 3-6, such as 3.1,3.4,3.7,4.2,4.7,5.1,5.6,5.9 etc.
Preferably, the time of the described laccase curing reaction of step (5) is 60-120min, such as 61min, 63min, 67min, 75min, 84min, 91min, 99min, 103min, 108min, 112min, 115min, 119min etc.
As optimal technical scheme, the preparation method of magnetic light-enzyme composite catalyst of the present invention is with magnetic Nano TiO
2for Laccase Immobilization carrier, be coupling agent with silylating reagent, add a certain amount of monohydric alcohol, in acid condition, by after ultrasonic, Magneto separate, appropriate iminodiacetic acid sodium solution is added, under alkali condition, after stirring, Magneto separate, washing, add finite concentration CuSO
4solution, obtains the magnetic Nano TiO of chelating metal ion after stirring, Magneto separate
2nano particle.The magnetic Nano TiO of appropriate chelating metal ion is added in finite concentration laccase solution
2nano particle, after vibration, Magneto separate and washing, finally obtains novel magnetic light enzyme composite catalyst.
Further preferably, the preparation method of magnetic light-enzyme composite catalyst of the present invention comprises the following steps:
A, magnetic Nano TiO
2nano particle surface-functionalized
(1) under room temperature, stirring condition, silylating reagent, monohydric alcohol, magnetic Nano TiO in reaction system, is added successively
2nano particle, water, wherein, based on 1 weight portion silylating reagent, add monohydric alcohol 40-60 weight portion, magnetic Nano TiO
2nano particle 0.5-2 weight portion, water 10-15 weight portion, adjust ph;
(2) mixed solution step (1) obtained pass into nitrogen protection, heating water bath and install reflux additional condition under carry out the Silanization reaction of ultrasonic wave added, obtain silanization magnetic Nano TiO through Magneto separate
2nano particle;
(3) product that step (2) obtains is joined in iminodiacetic acid sodium solution; stir, pass into nitrogen protection, heating water bath and install reflux additional condition under carry out graft reaction, through Magneto separate, washing after obtain the magnetic Nano TiO with imino group functional group
2nano particle;
(4), in the copper-bath joined by the product that step (3) obtains, oscillating reactions under room temperature condition, obtains the magnetic Nano TiO that copper ion huge legendary turtle is closed after Magneto separate, washing
2nano particle.
B, surface-functionalized magnetic Nano TiO
2the Laccase Immobilization of nano particle
(5) the magnetic Nano TiO of surface copper ion chelating will prepared
2nano particle mixes with the free laccase be dissolved in buffer solution, and at ambient temperature, vibration, carries out Laccase Immobilization reaction, after Magneto separate, washing, obtain magnetic light-enzyme composite catalyst.
Particularly, of the present invention by magnetic Nano TiO
2the method that nano grain surface functionalization and Laccase Immobilization prepare magnetic light-enzyme composite catalyst can comprise the following steps:
(1 ') adopts method (the Comparison of magnetic-manometertitanium dioxide/ferriferous oxide(TiO of existing bibliographical information
2/ Fe
3o
4) composite photocatalyst prepared byacid-sol and homogeneous precipitation methods.Jinyuan Chen, Journal of MaterialsScience, 2010,45:6018 – 6024) prepare magnetic Nano TiO
2nano particle, wherein Fe
3o
4content is 1-4%(w/w), make it be scattered in deionized water through ultrasonic, form stable suspension, concentration is 2-10%(w/v), then put into refrigerator and cooled and hide for subsequent use;
(1) under room temperature (about 25 DEG C), the 200-400 rev/min condition stirred, silylating reagent, monohydric alcohol, magnetic Nano TiO is added successively in reaction system
2nano particle, water, wherein, based on 1 weight portion silylating reagent, preferably, add monohydric alcohol 8-12 weight portion, magnetic Nano TiO
2nano particle 0.1-0.4 weight portion, water 2-3 weight portion, adjust ph, to 3-5, fully stirs 10-30min;
(2) mixed solution step (2) obtained pass into nitrogen protection, heating water bath, install reflux, 60-120W additional ultrasound condition under carry out Silanization reaction, reaction temperature 30-50 DEG C, reaction time 20-60min, gained silanization magnetic Nano TiO
2nano particle is after Magneto separate, without the need to washing, for subsequent use;
(3) product that step (3) obtains is joined 0.5-2%(w/v) in iminodiacetic acid sodium solution, wherein, silanization magnetic Nano TiO
2the concentration of nano particle is 1-4%(w/v); regulate pH to 8-10; 100-300 rev/min of stirring, pass into nitrogen protection, heating water bath and install reflux additional condition under carry out graft reaction; reaction temperature 40-80 DEG C; reaction time 30-90min, obtains the magnetic Nano TiO with imino group functional group after Magneto separate, washing
2nano particle;
(4) product that step (4) obtains is joined 2-5%(w/v) copper-bath in, wherein, silanization magnetic Nano TiO
2the concentration of nano particle is 2-6%(w/v), under room temperature condition, 100-200 rev/min of oscillating reactions, reaction time 45-90min, obtains the magnetic Nano TiO that copper ion huge legendary turtle is closed after Magneto separate, washing
2nano particle;
(5) the magnetic Nano TiO of surface copper ion chelating will prepared
2nano particle mixes with the free laccase be dissolved in buffer solution, and wherein, the concentration of laccase is 0.1-0.8g/L, the magnetic Nano TiO of free laccase and surface copper ion chelating
2the mixed proportion of nano particle is 0.1-0.4g laccase/1g carrier, pH3-6, and at ambient temperature, Laccase Immobilization reaction is carried out in 100-200 rev/min of vibration, reaction time 60-120min, after Magneto separate, washing, obtains magnetic light-enzyme composite catalyst.
Described monohydric alcohol is selected from a kind of in methyl alcohol, ethanol, normal propyl alcohol or both mixtures.
Described laccase is selected from one or more the mixture in fungal laccase, and laccase molecular weight ranges is 50-100kDa.
Three of object of the present invention is to provide a kind of purposes of magnetic light-enzyme composite catalyst, and described magnetic light-enzyme composite catalyst is used for the degraded of association with pulp bleaching in organic synthesis, the degraded of environmental contaminants and dye decolored, biological detection and electrochemical analysis, paper industry and wastewater treatment, fibrous raw material; Be preferred for organic compound and the part inorganic compound of the contaminated environment such as degrading phenol, dyestuff, polycyclic aromatic hydrocarbon.
Magnetic light-enzyme composite catalyst of the present invention is with magnetic Nano TiO
2for fixation support, realize laccase and magnetic Nano TiO by the means of metal ion huge legendary turtle and affine modification
2the establishment of complex, described catalyst to achieve in catalytic oxidation process electronics at laccase and TiO
2between transmission, solve laccase and accept the slow and light induced electron of electron velocity and O from substrate
2in conjunction with problem slowly, enzymatic and light-catalysed synergy is utilized to improve catalytic oxidation efficiency, the oxidation to the higher substrate of oxidation-reduction potential and macromolecule substrate can be realized, this catalyst has superparamagnetism, control easily and effectively can be carried out and be separated under the effect of externally-applied magnetic field, preparation technology is simple, is easy to amplify.Especially as laccase in described catalyst and magnetic Nano TiO
2mass ratio be 1:5-10, wherein for the magnetic Nano TiO of catalyst preparing
2fe
3o
4content is 1-4%(w/w) time, better effects if.
Compared with prior art, the present invention has following beneficial effect:
(1) magnetic light-enzyme composite catalyst provided by the invention can to realize in catalytic oxidation process electronics at laccase and TiO
2between transmission, relieve the electron transmission rate-limiting step in both catalytic reactions, utilize enzymatic and light-catalysed synergy to improve catalytic oxidation efficiency;
(2) magnetic light-enzyme composite catalyst provided by the invention can realize the oxidation to the higher substrate of oxidation-reduction potential and macromolecule substrate;
(3) magnetic light-enzyme composite catalyst provided by the invention has superparamagnetism, can carry out control easily and effectively and be separated under the effect of externally-applied magnetic field;
(4) preparation technology of magnetic light-enzyme composite catalyst provided by the invention is simple, is easy to amplify.
Detailed description of the invention
For ease of understanding the present invention, it is as follows that the present invention enumerates embodiment.Those skilled in the art should understand, described embodiment is only help to understand the present invention, should not be considered as concrete restriction of the present invention.
Embodiment 1
The magnetic Nano TiO that (1 ') will prepare
2nanoparticulate dispersed forms suspension in deionized water, magnetic Nano TiO
2fe
3o
4content is 4%(w/w), magnetic Nano TiO
2concentration be 2%(w/v), then put into refrigerator and cooled hide for subsequent use.
(1) under room temperature (25 DEG C), 200 revs/min of conditions stirred, silylating reagent 2g, monohydric alcohol 120g, magnetic Nano TiO is added successively in reaction system
2nano particle 4g, water 30g, adjust ph to 5, after fully stirring 30min, obtains Silanization reaction solution;
(2) Silanization reaction solution is reacted 20min under passing into nitrogen protection, heating water bath 50 DEG C, installing reflux, 120W ultrasound condition additional, products therefrom, after Magneto separate, obtains silanization magnetic Nano TiO
2nano particle;
(3) by silanization magnetic Nano TiO
2nano particle joins 2%(w/v) iminodiacetic acid sodium solution in; granule density is 4%(w/v); regulate pH to 10; 300 revs/min of stirrings, pass into nitrogen protection, heating water bath 80 DEG C and install reflux additional condition under reaction time 30min, through Magneto separate, washing after obtain the magnetic Nano TiO with imino group functional group
2nano particle;
(4) nano particle that step (3) obtains is joined 5%(w/v) copper-bath in, granule density is 6%(w/v), under room temperature condition, 200 revs/min of oscillating reactions 45min, through Magneto separate, washing after obtain copper ion huge legendary turtle close magnetic Nano TiO
2nano particle;
(5) the magnetic Nano TiO that the copper ion huge legendary turtle obtained with step (4) is closed
2nano particle mixes as carrier and the 0.1g/L be dissolved in the pH3.0 tartaric acid buffer laccase that dissociates, and the concentration 1g/L of nano particle, at ambient temperature, 100 revs/min of oscillating reactions 60min, after Magneto separate, washing, obtain magnetic light-enzyme composite catalyst.
Catalytic activity is tested:
Be that substrate carries out catalysis activity mensuration with phenol, the vigor of composite catalyst comparatively equivalent dissociates laccase and magnetic Nano TiO
2catalysis activity sum improve 50%.
Embodiment 2
The magnetic Nano TiO that (1 ') will prepare
2nanoparticulate dispersed forms suspension in deionized water, magnetic Nano TiO
2fe
3o
4content is 1%(w/w), its concentration is 10%(w/v), then put into refrigerator and cooled and hide for subsequent use;
(1) under room temperature (25 DEG C), 200 revs/min of conditions stirred, silylating reagent 2g, monohydric alcohol 80g, magnetic Nano TiO is added successively in reaction system
2nano particle 1g, water 20g, adjust ph to 3, after fully stirring 10min, obtains Silanization reaction solution;
(2) Silanization reaction solution is reacted 60min under passing into nitrogen protection, heating water bath 30 DEG C, installing reflux, 60W ultrasound condition additional, gained silanization magnetic Nano TiO
2nano particle, after Magneto separate, obtains silanization magnetic Nano TiO
2nano particle;
(3) by silanization magnetic Nano TiO
2nano particle joins 0.5%(w/v) in iminodiacetic acid sodium solution; granule density is 1%(w/v); regulate pH to 8; 100 revs/min of stirrings, pass into nitrogen protection, heating water bath 40 DEG C and install reflux additional condition under reaction time 90min, through Magneto separate, washing after obtain the magnetic Nano TiO with imino group functional group
2nano particle;
(4) nano particle that step (3) obtains is joined 2%(w/v) copper-bath in, granule density is 2%(w/v), under room temperature condition, 100 revs/min of oscillating reactions 90min, through Magneto separate, washing after obtain copper ion huge legendary turtle close magnetic Nano TiO
2nano particle;
(5) the magnetic Nano TiO that the copper ion huge legendary turtle obtained with step (4) is closed
2nano particle mixes as carrier and the 0.8g/L be dissolved in the pH6.0 phosphate buffer laccase that dissociates, and granule density 2g/L, at ambient temperature, 200 revs/min of oscillating reactions 120min, after Magneto separate, washing, obtain magnetic light-enzyme composite catalyst.
Catalytic activity is tested:
Be that substrate carries out catalysis activity mensuration with phenol, the vigor of composite catalyst comparatively equivalent dissociates laccase and magnetic Nano TiO
2catalysis activity sum improve 40%.
Embodiment 3
The magnetic Nano TiO that (1 ') will prepare
2nanoparticulate dispersed forms suspension in deionized water, magnetic Nano TiO
2fe
3o
4content is 2%(w/w), its concentration is 5%(w/v), then put into refrigerator and cooled and hide for subsequent use;
(1) under room temperature (25 DEG C), 300 revs/min of conditions stirred, silylating reagent 2g, monohydric alcohol 100g, magnetic Nano TiO is added successively in reaction system
2nano particle 2.5g, water 25g, adjust ph to 4, after fully stirring 20min, obtains Silanization reaction solution;
(2) Silanization reaction solution is reacted 40min under passing into nitrogen protection, heating water bath 40 DEG C, installing reflux, 90W ultrasound condition additional, gained silanization magnetic Nano TiO
2nano particle, after Magneto separate, obtains silanization magnetic Nano TiO
2nano particle;
(3) by silanization magnetic Nano TiO
2nano particle joins 1%(w/v) in iminodiacetic acid sodium solution; granule density is 2%(w/v); regulate pH to 9; 200 revs/min of stirrings, pass into nitrogen protection, heating water bath 60 DEG C and install reflux additional condition under reaction time 60min, through Magneto separate, washing after obtain the magnetic Nano TiO with imino group functional group
2nano particle;
(4) nano particle that step (3) obtains is joined 3%(w/v) copper-bath in, granule density is 4%(w/v), under room temperature condition, 150 revs/min of oscillating reactions 60min, through Magneto separate, washing after obtain copper ion huge legendary turtle close magnetic Nano TiO
2nano particle;
(5) the magnetic Nano TiO that the copper ion huge legendary turtle obtained with step (4) is closed
2nano particle mixes as carrier and the 0.3g/L be dissolved in the pH4.5 citrate buffer solution laccase that dissociates, and granule density 1g/L, at ambient temperature, 150 revs/min of oscillating reactions 90min, after Magneto separate, washing, obtain magnetic light-enzyme composite catalyst.
Catalytic activity is tested:
Be that substrate carries out catalysis activity mensuration with phenol, the vigor of composite catalyst comparatively equivalent dissociates laccase and magnetic Nano TiO
2catalysis activity sum improve 70%.
Embodiment 4
The magnetic Nano TiO that (1 ') will prepare
2nanoparticulate dispersed forms suspension in deionized water, magnetic Nano TiO
2fe
3o
4content is 3%(w/w), its concentration is 4%(w/v), then put into refrigerator and cooled and hide for subsequent use;
(1) under room temperature (25 DEG C), 200 revs/min of conditions stirred, silylating reagent 2g, monohydric alcohol 90g, magnetic Nano TiO is added successively in reaction system
2nano particle 3g, water 30g, adjust ph to 4.5, after fully stirring 30min, obtains Silanization reaction solution;
(2) Silanization reaction solution is reacted 50min under passing into nitrogen protection, heating water bath 30 DEG C, installing reflux, 60W ultrasound condition additional, gained silanization magnetic Nano TiO
2nano particle, after Magneto separate, obtains silanization magnetic Nano TiO
2nano particle;
(3) by silanization magnetic Nano TiO
2nano particle joins 1.5%(w/v) in iminodiacetic acid sodium solution; granule density is 3%(w/v); regulate pH to 8.5; 150 revs/min of stirrings, pass into nitrogen protection, heating water bath 50 DEG C and install reflux additional condition under reaction time 80min, through Magneto separate, washing after obtain the magnetic Nano TiO with imino group functional group
2nano particle;
(4) nano particle that step (3) obtains is joined 4%(w/v) copper-bath in, granule density is 5%(w/v), under room temperature condition, 120 revs/min of oscillating reactions 80min, through Magneto separate, washing after obtain copper ion huge legendary turtle close magnetic Nano TiO
2nano particle;
(5) the magnetic Nano TiO that the copper ion huge legendary turtle obtained with step (4) is closed
2nano particle mixes as carrier and the 0.6g/L be dissolved in the pH5 acetate buffer solution laccase that dissociates, and granule density 2g/L, at ambient temperature, 120 revs/min of oscillating reactions 100min, after Magneto separate, washing, obtain magnetic light-enzyme composite catalyst.
Catalytic activity is tested:
Be that substrate carries out catalysis activity mensuration with phenol, the vigor of composite catalyst comparatively equivalent dissociates laccase and magnetic Nano TiO
2catalysis activity sum improve 60%.
Embodiment 5
The magnetic Nano TiO that (1 ') will prepare
2nanoparticulate dispersed forms suspension in deionized water, magnetic Nano TiO
2fe
3o
4content is 1.4%(w/w), its concentration is 4.2%(w/v), then put into refrigerator and cooled and hide for subsequent use;
(1) under room temperature (25 DEG C), 200 revs/min of conditions stirred, silylating reagent 2g, monohydric alcohol 40g, magnetic Nano TiO is added successively in reaction system
2nano particle 0.2g, water 10g, adjust ph to 3.7, after fully stirring 30min, obtains Silanization reaction solution;
(2) Silanization reaction solution is reacted 45min under passing into nitrogen protection, heating water bath 35 DEG C, installing reflux, 100W ultrasound condition additional, gained silanization magnetic Nano TiO
2nano particle, after Magneto separate, obtains silanization magnetic Nano TiO
2nano particle;
(3) by silanization magnetic Nano TiO
2nano particle joins 0.7%(w/v) in iminodiacetic acid sodium solution; granule density is 3.7%(w/v); regulate pH to 9.4; 180 revs/min of stirrings, pass into helium/argon shield, heating water bath 62 DEG C and install reflux additional condition under reaction time 45min, through Magneto separate, washing after obtain the magnetic Nano TiO with imino group functional group
2nano particle;
(4) nano particle that step (3) obtains is joined 2.4%(w/v) copper-bath in, granule density is 4.1%(w/v), under room temperature condition, 125 revs/min of oscillating reactions 50min, obtain the magnetic Nano TiO that copper ion huge legendary turtle is closed after Magneto separate, washing
2nano particle;
(5) the magnetic Nano TiO that the copper ion huge legendary turtle obtained with step (4) is closed
2nano particle mixes as carrier and the 0.2g/L be dissolved in the pH5 acetate buffer solution laccase that dissociates, and granule density 1g/L, at ambient temperature, 120 revs/min of oscillating reactions 75min, after Magneto separate, washing, obtain magnetic light-enzyme composite catalyst.
Catalytic activity is tested:
Be that substrate carries out catalysis activity mensuration with phenol, the vigor of composite catalyst comparatively equivalent dissociates laccase and magnetic Nano TiO
2catalysis activity sum improve 55%.
Embodiment 6
The magnetic Nano TiO that (1 ') will prepare
2nanoparticulate dispersed forms suspension in deionized water, magnetic Nano TiO
2fe
3o
4content is 3.8%(w/w), its concentration is 8.8%(w/v), then put into refrigerator and cooled and hide for subsequent use;
(1) under room temperature (25 DEG C), 200 revs/min of conditions stirred, silylating reagent 2g, monohydric alcohol 140g, magnetic Nano TiO is added successively in reaction system
2nano particle 10g, water 40g, adjust ph to 4.7, after fully stirring 55min, obtains Silanization reaction solution;
(2) Silanization reaction solution is reacted 25min under passing into nitrogen protection, heating water bath 45 DEG C, installing reflux, 115W ultrasound condition additional, gained silanization magnetic Nano TiO
2nano particle, after Magneto separate, obtains silanization magnetic Nano TiO
2nano particle;
(3) by silanization magnetic Nano TiO
2nano particle joins 1.6%(w/v) in iminodiacetic acid sodium solution; granule density is 1.2%(w/v); regulate pH to 8.5; 280 revs/min of stirrings, pass into helium/argon shield, heating water bath 77 DEG C and install reflux additional condition under reaction time 85min, through Magneto separate, washing after obtain the magnetic Nano TiO with imino group functional group
2nano particle;
(4) nano particle that step (3) obtains is joined 4.7%(w/v) copper-bath in, granule density is 3.7%(w/v), under room temperature condition, 150 revs/min of oscillating reactions 85min, obtain the magnetic Nano TiO that copper ion huge legendary turtle is closed after Magneto separate, washing
2nano particle;
(5) the magnetic Nano TiO that the copper ion huge legendary turtle obtained with step (4) is closed
2nano particle mixes as carrier and the 0.6g/L be dissolved in the pH5 tartaric acid buffer laccase that dissociates, and granule density 4.2g/L, at ambient temperature, 180 revs/min of oscillating reactions 105min, after Magneto separate, washing, obtain magnetic light-enzyme composite catalyst.
Catalytic activity is tested:
Be that substrate carries out catalysis activity mensuration with phenol, the vigor of composite catalyst comparatively equivalent dissociates laccase and magnetic Nano TiO
2catalysis activity sum improve 75%.
Applicant states, the present invention illustrates detailed process equipment and process flow process of the present invention by above-described embodiment, but the present invention is not limited to above-mentioned detailed process equipment and process flow process, namely do not mean that the present invention must rely on above-mentioned detailed process equipment and process flow process and could implement.Person of ordinary skill in the field should understand, any improvement in the present invention, to equivalence replacement and the interpolation of auxiliary element, the concrete way choice etc. of each raw material of product of the present invention, all drops within protection scope of the present invention and open scope.
Claims (3)
1. a preparation method for magnetic light-enzyme composite catalyst, is characterized in that, described magnetic light-enzyme composite catalyst is for being immobilized in magnetic Nano TiO
2on laccase, described magnetic Nano TiO
2for closing the magnetic Nano TiO modified through copper ion huge legendary turtle
2, described laccase and copper ion huge legendary turtle close the magnetic Nano TiO modified
2mass ratio be 1:1-1:20 (w/w), described copper ion huge legendary turtle closes the magnetic Nano TiO modified
2fe
3o
4content is 1-4wt%, and described laccase is selected from the combination of in fungal laccase a kind or at least 2 kinds, and described laccase molecular weight ranges is 50-100kDa;
Described method comprises the steps:
(1) under room temperature, stirring condition, silylating reagent, monohydric alcohol, magnetic Nano TiO in reaction system, is added successively
2particle, water, wherein, based on 1 weight portion silylating reagent, add monohydric alcohol 40-60 weight portion, magnetic Nano TiO
2particle 0.5-2 weight portion, water 10-15 weight portion, adjust ph 3-5;
(2) mixed solution step (1) obtained pass into nitrogen protection, heating water bath and install reflux additional condition under carry out the Silanization reaction of ultrasonic wave added, obtain silanization magnetic Nano TiO through Magneto separate
2particle;
(3) product that step (2) obtains is joined in iminodiacetic acid sodium solution; stir, pass into nitrogen protection, heating water bath and install reflux additional condition under carry out graft reaction, through Magneto separate, washing after obtain the magnetic Nano TiO with imino group functional group
2particle;
(4), in the copper-bath joined by the product that step (3) obtains, oscillating reactions under room temperature condition, obtains the magnetic Nano TiO that copper ion huge legendary turtle is closed after Magneto separate, washing
2particle;
(5) copper ion huge legendary turtle step (4) prepared closes the magnetic Nano TiO modified
2particle mixes with the free laccase be dissolved in buffer solution, and at ambient temperature, vibration, carries out Laccase Immobilization reaction, after Magneto separate, washing, obtain described magnetic light-enzyme composite catalyst.
2. the method for claim 1, is characterized in that, step (5) described laccase and copper ion huge legendary turtle close the magnetic Nano TiO modified
2the mass ratio of particle is 1:3-1:15 (w/w).
3. method as claimed in claim 2, it is characterized in that, step (5) described laccase and copper ion huge legendary turtle close the magnetic Nano TiO modified
2the mass ratio of particle is 1:5-1:10 (w/w).
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| CN111167519B (en) * | 2020-01-13 | 2022-08-05 | 河北工业大学 | Photo-enzyme integrated nano catalyst and application thereof |
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| CN112592917B (en) * | 2020-12-31 | 2023-08-11 | 华南理工大学 | Magnetic lipase-supported photocatalyst, and preparation method and application thereof |
| CN114807112B (en) * | 2022-05-05 | 2024-02-27 | 江苏大学 | Method for immobilizing laccase by magnetic graphite phase carbon nitride and application thereof |
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| JP2006096653A (en) * | 2004-08-31 | 2006-04-13 | Toyota Central Res & Dev Lab Inc | Carbon gel composite material |
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| CN102284284A (en) * | 2011-06-03 | 2011-12-21 | 南昌航空大学 | Method for preparing molecularly imprinted TiO2/WO3 composite photocatalyst with visible light response through direct method |
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| US5443975A (en) * | 1990-06-13 | 1995-08-22 | Ente Per Le Nuove Technologie, L'energia E L'ambiente (Enea) | Method of binding enzymes to sintered expanded clays |
| JP2006096653A (en) * | 2004-08-31 | 2006-04-13 | Toyota Central Res & Dev Lab Inc | Carbon gel composite material |
| CN101856628A (en) * | 2010-04-30 | 2010-10-13 | 北京科技大学 | Conducting polymer modified magnetic photocatalyst and preparation method thereof |
| CN102284284A (en) * | 2011-06-03 | 2011-12-21 | 南昌航空大学 | Method for preparing molecularly imprinted TiO2/WO3 composite photocatalyst with visible light response through direct method |
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