CN111411105A - Magnetic immobilized enzyme nano reactor and preparation method and application thereof - Google Patents
Magnetic immobilized enzyme nano reactor and preparation method and application thereof Download PDFInfo
- Publication number
- CN111411105A CN111411105A CN202010313183.8A CN202010313183A CN111411105A CN 111411105 A CN111411105 A CN 111411105A CN 202010313183 A CN202010313183 A CN 202010313183A CN 111411105 A CN111411105 A CN 111411105A
- Authority
- CN
- China
- Prior art keywords
- immobilized enzyme
- magnetic
- enzyme
- reaction
- magnetic immobilized
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Withdrawn
Links
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N11/00—Carrier-bound or immobilised enzymes; Carrier-bound or immobilised microbial cells; Preparation thereof
- C12N11/14—Enzymes or microbial cells immobilised on or in an inorganic carrier
Landscapes
- Chemical & Material Sciences (AREA)
- Genetics & Genomics (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Wood Science & Technology (AREA)
- Health & Medical Sciences (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Zoology (AREA)
- Biomedical Technology (AREA)
- Biotechnology (AREA)
- Inorganic Chemistry (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Microbiology (AREA)
- Enzymes And Modification Thereof (AREA)
- Immobilizing And Processing Of Enzymes And Microorganisms (AREA)
Abstract
The invention relates to a magnetic immobilized enzyme nano reactor and a preparation method and application thereof, wherein ferric chloride and ferrous chloride are mixed and react in an aqueous environment to synthesize ferroferric oxide magnetic nano particles; mixing the obtained reactant with polydopamine hydrochloride to synthesize ferroferric oxide magnetic nanoparticles with polydopamine attached to the surface; wrapping enzyme on the outer layer of the ferroferric oxide magnetic nano-particles with polydopamine attached on the surface by using a pH circulation method to synthesize a magnetic immobilized enzyme; and further reacting the magnetic immobilized enzyme with metal salt and an imidazole compound, and drying the obtained product to obtain the magnetic immobilized enzyme nano-reactor. The invention endows the enzyme catalyst with the characteristics of easy cleaning and repeated use and good organic and heat resistance.
Description
Technical Field
The invention belongs to the technical field of biological catalysis, relates to a biocatalyst enzyme, and particularly relates to a magnetic immobilized enzyme nano reactor, and a preparation method and application thereof.
Background
Magnetic Nanoparticles (MNPs) are a new type of Magnetic material developed after the 80's of the 20 th century, and mainly include a composite of elements such as iron, manganese, and cobalt, oxygen, and sulfur. The magnetic nanoparticles have small particle size, large specific surface area, strong adsorption capacity and flexible and easily-modified surface, so the magnetic nanoparticles are widely applied to the fields of catalytic reaction, biological separation, targeted drug loading, magnetic resonance imaging, analytical chemistry and the like. The immobilized enzyme usually needs to be recovered by a method such as centrifugal separation or filtration after use, and the catalyst is easily lost during the operation, which is complicated and time-consuming. The magnetic immobilized enzyme can be recovered by using an external magnetic field after the catalysis is finished, and is cleaned by using different organic solvents and salt solutions. However, when the magnetic immobilized enzyme is washed or used in a salt solution, the enzyme is easily desorbed from the surface of the magnetic particle, and the activity of the magnetic immobilized enzyme is lost.
Metal-organic frameworks (MOFs) are porous crystalline materials formed by Metal ions or Metal clusters and organic ligands through coordination bonds, and have flexible surface characteristics, large specific surface area and pore volume. Due to its unique properties, including tunable porosity, extremely high surface area and chemical/thermal stability, and high reusability, the metal-organic framework has great application prospects in enzyme immobilization. Depending on the synthesis method, metal organic framework-enzyme composites can be divided into five types: surface physical adsorption, surface covalent attachment, pore capture, biomimetic mineralization and coprecipitation. Surface adsorption and covalent attachment are easily achieved, and enzymes are immobilized on the surface of a metal organic framework through physical and chemical forces, but both methods have no protection effect on the immobilized enzymes, and the enzymes are easily inactivated when directly exposed to the environment. The immobilized enzyme by the pore capture method has good reusability and stability, however, most of the enzyme molecules have the size larger than the pore diameter of the metal-organic framework, and the enzyme which can enter the cavity of the metal-organic framework is few, so the method is not a universal method. The enzyme is wrapped by the coprecipitation method, polyvinylpyrrolidone is used as a capping agent, the cost is high, the preparation is difficult, the temperature range of the bioactivity of the enzyme is small, and the stability is poor.
Disclosure of Invention
The invention aims to overcome the defects of the prior art and provide a magnetic immobilized enzyme nano reactor and a preparation method and application thereof.
The purpose of the invention can be realized by the following technical scheme:
a preparation method of a magnetic immobilized enzyme nano reactor comprises the following steps:
1) mixing ferric chloride and ferrous chloride with water, and then heating for reaction to obtain ferroferric oxide magnetic nanoparticles;
2) mixing the ferroferric oxide magnetic nanoparticles, polydopamine hydrochloride and a buffer solution, and then stirring for reaction to obtain the ferroferric oxide magnetic nanoparticles with polydopamine attached to the surfaces;
3) mixing ferroferric oxide magnetic nanoparticles with polydopamine attached on the surface, enzyme and water, then firstly adjusting the pH value to 8.0-10.0 and stirring, then adjusting the pH value to 7.0-8.0 and stirring to obtain magnetic immobilized enzyme;
4) mixing the magnetic immobilized enzyme, the metal salt and the imidazole compound with water, and then stirring for reaction to obtain the magnetic immobilized enzyme nano reactor.
Further, in the step 1), the mass concentration of the ferric chloride in the water is 0.01-100 g/L, and the mass ratio of the ferric chloride to the ferrous chloride is (0.1-10): 1.
Further, in the step 1), in the heating reaction process, the reaction temperature is 100-200 ℃, the reaction time is 1-2000min, and the reaction pressure is 2.5-3.5 MPa. Within the numerical range, the ferroferric oxide magnetic nanoparticles can be synthesized under different reaction temperatures and reaction times, and the difference is that the sizes of the particles are different.
Further, in the step 2), the buffer solution is a Tris-HCl buffer solution, the pH value of the Tris-HCl buffer solution is 7.0-10.0, the concentration of the Tris-HCl buffer solution is 0.01-100 g/L, the mass concentration of the polydopamine hydrochloride in the buffer solution is 0.01-15 g/L, and the mass ratio of the ferroferric oxide magnetic nanoparticles to the polydopamine hydrochloride is (0.1-10): 1.
Further, in the step 2), the reaction temperature is 4-40 ℃ and the reaction time is 1-2000min in the stirring reaction process. The stirring reaction aims to attach a layer of polydopamine hydrochloride on the surface of the ferroferric oxide magnetic nano particles, and the attached thicknesses are different as a result of different reaction time lengths.
Further, in the step 3), the mass concentration of the enzyme in water is 0.01-10 g/L, the mass ratio of the ferroferric oxide magnetic nanoparticles with polydopamine attached to the surface to the enzyme is (0.2-5):1, the enzyme comprises one or more of lipotransferase or lipohydrolase, the pH value is adjusted to 8.0-10.0 and then stirred for 0.1-5.0h, the pH value is adjusted to 7.0-8.0 and then stirred for 0.1-5.0h, and the pH value is adjusted by using sodium hydroxide as alkali and hydrochloric acid as acid.
Further, in the step 4), the mass concentration of the metal salt in water is 0.01-15 g/L, the mass ratio of the magnetic immobilized enzyme, the metal salt and the imidazole compound is (0.2-5):1 (0.5-5), the metal salt comprises one or more of zinc chloride, zinc nitrate or cobalt nitrate, the imidazole compound comprises one or more of dimethyl imidazole, benzimidazole or nitroimidazole, and the imidazole compound is used as an organic ligand and can be connected with a metal cation through a coordination bond, so that the pore size and the pore channel structure of the metal organic framework are influenced.
Further, in the step 4), the reaction temperature is 4-40 ℃ and the reaction time is 10-3000min in the stirring reaction process. The stirring is magnetic stirring or mechanical stirring. After stirring and reacting, drying by adopting a freeze drying, vacuum drying or forced air drying mode. The stirring reaction aims to mineralize a layer of metal organic framework on the surface of the magnetic immobilized enzyme, and the mineralized thickness is different as a result of different reaction time lengths.
A magnetic immobilized enzyme nano reactor is prepared by the method.
An application of a magnetic immobilized enzyme nano reactor in biocatalytic reaction.
When the magnetic immobilized enzyme nano reactor is prepared, ferric chloride and ferrous chloride are mixed and react in an aqueous phase environment to synthesize ferroferric oxide magnetic nano particles; mixing the obtained reactant with polydopamine hydrochloride to synthesize ferroferric oxide magnetic nanoparticles with polydopamine attached to the surface; then, uniformly dispersing the lipase in an alkaline water phase, wherein the carboxyl of the lipase is deprotonated, and a hydrophobic group of the lipase is exposed; then adjusting the pH value of the system to be neutral, wherein the magnetic nanoparticles are positively charged, and the lipase is in an environment above the isoelectric point and negatively charged, and the lipase are combined due to electrostatic attraction and hydrophobic effect, and the magnetic nanoparticles are loaded in the lipase in the process of closing the structure of the hydrophobic part of the lipase, so that the magnetic immobilized enzyme is synthesized; and further reacting the magnetic immobilized enzyme with metal salt and an imidazole compound, and drying the obtained product to obtain the magnetic immobilized enzyme nano-reactor. The invention endows the enzyme catalyst with the characteristics of easy cleaning and repeated use and good organic and heat resistance.
The invention takes magnetic nano particles and a metal organic framework as carriers, and adopts a biomimetic mineralization mode to fix enzyme in the carriers to form a loaded catalytic system. The biomimetic mineralization technology can enlarge the biological activity temperature range of the enzyme, improve the stability of the enzyme, and has the characteristics of low cost, convenient preparation and high efficiency. Moreover, through biomimetic mineralization and synthesis of the metal organic framework/enzyme, the enzyme is surrounded by the framework of the metal organic framework, the shell of the metal organic framework can protect the enzyme from the influence of reaction environment, and meanwhile, the pores of the framework allow selective and rapid diffusion of small molecular substrates and products. Therefore, the metal organic framework layer is covered on the outer surface of the magnetic immobilized enzyme, so that the enzyme has the functions of easy cleaning and magnetic recovery, and the stability of the enzyme is improved.
In the invention, the magnetic nano-particles are used as the inner core, so that the catalyst can be more easily recycled, and the cost is reduced; the metal organic framework as a housing can protect the catalyst from the interference of the reaction environment, thereby improving the stability of the catalyst, including good thermal stability and chemical stability. The combined use of the two can open up a wider market in the field of biocatalysis.
Compared with the prior art, the invention has the following characteristics:
1) the magnetic nano particles are used as the inner core, and the separation and recovery of the immobilized enzyme can be realized by using an external magnetic field, so that the cost required by recovery, cleaning and reutilization is obviously reduced;
2) the hydrophobic polydopamine layer is wrapped on the surface of the magnetic nano-particle, so that the stability of the magnetic nano-particle is improved, and a binding site is provided for the hydrophobic adsorption of subsequent enzymes;
3) the enzyme is attached to the surface of the magnetic nano-particle with the outer layer of hydrophobic polydopamine by utilizing a pH circulation method, so that the active site of the enzyme is fully exposed, and the catalytic performance of the enzyme is improved;
4) the metal organic framework outer layer can protect the enzyme from being damaged by reaction environment, so that the stability of the enzyme, including thermal stability and organic solvent stability, is improved;
5) the invention has simple process flow and easy control of reaction conditions, prolongs the service life of the enzyme, saves the cost and can realize large-scale industrial production.
Drawings
FIG. 1 is a TEM image of a magnetic immobilized enzyme nanoreactor prepared in example 1;
FIG. 2 is a FTIR chart of the magnetic immobilized enzyme nanoreactor prepared in example 1;
FIG. 3 is an XRD pattern of the magnetic immobilized enzyme nanoreactor prepared in example 1;
FIG. 4 is a TGA diagram of a magnetic immobilized enzyme nanoreactor made in example 1;
FIG. 5 is a graph showing the magnetic properties of the magnetic immobilized enzyme nanoreactor prepared in example 1.
In fig. 1 to 5, a represents a ferroferric oxide magnetic nanoparticle, b represents a ferroferric oxide magnetic nanoparticle with polydopamine attached to the surface, c represents a magnetic immobilized enzyme, and d represents a magnetic immobilized enzyme nanoreactor.
Detailed Description
The invention is described in detail below with reference to the figures and specific embodiments. The present embodiment is implemented on the premise of the technical solution of the present invention, and a detailed implementation manner and a specific operation process are given, but the scope of the present invention is not limited to the following embodiments.
Example 1:
0.35g of ferric chloride and 1.20g of ferrous chloride are taken to be mixed with 1L water and fully dissolved, the mixed solution is placed in a high-pressure reaction kettle, sealing is carried out, the reaction temperature is 198 ℃, the reaction pressure is 3MPa, the heat preservation time is 1990min, after the reaction is completed, the obtained liquid reactant is cooled, then ultrapure water and ethanol are respectively used for washing three times, vacuum drying is carried out, 0.24g of the obtained dried substance and 0.24g of polydopamine hydrochloride are taken to be mixed with 0.02L Tris-HCl buffer solution (0.02 g/L, pH 7.5), ultrasonic dispersion is uniform, the mixed solution is vigorously stirred on a mechanical stirring device for 2000min, the obtained liquid reactant is respectively used for washing three times with ultrapure water and ethanol, vacuum drying is carried out, 0.2g of the obtained dried substance and 0.4g of non-immobilized Tris-transesterified lipase are taken to be uniformly mixed in 0.1L water, the pH value is firstly adjusted to 9.8 by sodium hydroxide, then is mechanically adjusted to 0.1h, the pH value is adjusted to 0.7 min, the obtained dried by using a mechanical stirring device, the obtained liquid reactant is further stirred with 0.6.6 h of sodium hydroxide, the obtained liquid reactant is further stirred, the obtained by using a mechanical stirring, the obtained by using a freeze-2 g of sodium hydroxide, the obtained three times of the obtained sodium hydroxide, the obtained solid phase, the solid phase is stirred, the solid phase is further.
The obtained magnetic immobilized enzyme nano reactor particles are subjected to morphology observation and performance characterization, and the morphology observation result is shown in figure 1. As can be seen from a TEM image, the synthesized ferroferric oxide magnetic nanoparticles have a relatively uniform spherical shape, the diameter of the nanoparticles is about 400nm, the poly-dopamine hydrochloride layer attached to the surface of the nanoparticles is about 27nm, and the shapes of the particles are still complete and smooth without changing the shapes of the particles by enzyme immobilization and metal organic framework mineralization. The chemical group and crystallinity results of the particles are shown in fig. 2 and fig. 3, and it can be seen that characteristic peaks in the figures prove that the particles contain various components including ferroferric oxide, polydopamine hydrochloride, enzyme and metal organic framework. The thermal stability results of the particles are shown in fig. 4, and it can be seen that the loss rate of the particles at high temperature is significantly reduced after the metal organic framework layer is mineralized. The magnetic properties of the particles are shown in fig. 5, which shows that after the reaction is completed, the magnetic immobilized enzyme nanoreactor particles can be easily separated and recovered from the reaction medium by an external magnet.
The magnetic immobilized enzyme nanoreactor is used for catalyzing transesterification of annatto and glycerol, 2g of annatto, 30g of glycerol and 0.5g of magnetic immobilized enzyme nanoreactor particles are mixed with a 4L solvent (a mixed solution of 2-methyl-2-butanol and tetrahydrofuran in a volume ratio of 1:4) and are fully dissolved, the mixed solution is placed in a shaking incubator, the temperature is set to be 55 ℃, the rotating speed is 180rpm, and the time is 40 h.
Example 2:
a magnetic immobilized enzyme nano-reactor is prepared by the following steps:
1) mixing ferric chloride and ferrous chloride with water, and then heating for reaction to obtain ferroferric oxide magnetic nanoparticles;
2) mixing the ferroferric oxide magnetic nanoparticles, polydopamine hydrochloride and a buffer solution, and then stirring for reaction to obtain the ferroferric oxide magnetic nanoparticles with polydopamine attached to the surfaces;
3) mixing ferroferric oxide magnetic nanoparticles with polydopamine attached on the surface, enzyme and water, then adjusting the pH value to 8.0 and stirring, and then adjusting the pH value to 8.0 and stirring to obtain a magnetic immobilized enzyme;
4) mixing the magnetic immobilized enzyme, the metal salt and the imidazole compound with water, and then stirring for reaction to obtain the magnetic immobilized enzyme nano reactor.
In the step 1), the mass concentration of the ferric chloride in the water is 0.01 g/L, the mass ratio of the ferric chloride to the ferrous chloride is 10:1, and in the heating reaction process, the reaction temperature is 100 ℃ and the reaction time is 2000 min.
In the step 2), the buffer solution is a Tris-HCl buffer solution, the pH value of the Tris-HCl buffer solution is 7.0, the concentration of the Tris-HCl buffer solution is 100 g/L, the mass concentration of the polydopamine hydrochloride in the buffer solution is 0.01 g/L, the mass ratio of the ferroferric oxide magnetic nanoparticles to the polydopamine hydrochloride is 10:1, and in the stirring reaction process, the reaction temperature is 4 ℃ and the reaction time is 2000 min.
In the step 3), the mass concentration of the enzyme in water is 0.01 g/L, the mass ratio of the ferroferric oxide magnetic nanoparticles with polydopamine attached to the surface to the enzyme is 5:1, the enzyme comprises lipotransferase and lipohydrolase, the pH value is adjusted to 8.0, stirring is carried out for 5.0 hours, and the pH value is adjusted to 7.0, stirring is carried out for 5.0 hours.
In the step 4), the mass concentration of the metal salt in water is 0.01 g/L, the mass ratio of the magnetic immobilized enzyme to the metal salt to the imidazole compound is 5:1:0.5, the metal salt is zinc chloride, and the imidazole compound is dimethyl imidazole, and in the stirring reaction process, the reaction temperature is 40 ℃ and the reaction time is 10 min.
Example 3:
a magnetic immobilized enzyme nano-reactor is prepared by the following steps:
1) mixing ferric chloride and ferrous chloride with water, and then heating for reaction to obtain ferroferric oxide magnetic nanoparticles;
2) mixing the ferroferric oxide magnetic nanoparticles, polydopamine hydrochloride and a buffer solution, and then stirring for reaction to obtain the ferroferric oxide magnetic nanoparticles with polydopamine attached to the surfaces;
3) mixing ferroferric oxide magnetic nanoparticles with polydopamine attached on the surface, enzyme and water, then adjusting the pH value to 10.0 and stirring, then adjusting the pH value to 7.0 and stirring to obtain a magnetic immobilized enzyme;
4) mixing the magnetic immobilized enzyme, the metal salt and the imidazole compound with water, and then stirring for reaction to obtain the magnetic immobilized enzyme nano reactor.
In the step 1), the mass concentration of ferric chloride in water is 100 g/L, the mass ratio of ferric chloride to ferrous chloride is 0.1:1, and in the heating reaction process, the reaction temperature is 200 ℃ and the reaction time is 1 min.
In the step 2), the buffer solution is a Tris-HCl buffer solution, the pH value of the Tris-HCl buffer solution is 10.0, the concentration of the Tris-HCl buffer solution is 0.01 g/L, the mass concentration of the polydopamine hydrochloride in the buffer solution is 15 g/L, the mass ratio of the ferroferric oxide magnetic nanoparticles to the polydopamine hydrochloride is 0.1:1, and in the stirring reaction process, the reaction temperature is 40 ℃ and the reaction time is 1 min.
In the step 3), the mass concentration of the enzyme in water is 10 g/L, the mass ratio of the ferroferric oxide magnetic nanoparticles with polydopamine attached to the surface to the enzyme is 0.2:1, the enzyme is a lipase, the pH value is adjusted to 10.0, then the mixture is stirred for 0.1h, and the pH value is adjusted to 8.0, then the mixture is stirred for 0.1 h.
In the step 4), the mass concentration of the metal salt in water is 15 g/L, the mass ratio of the magnetic immobilized enzyme to the metal salt to the imidazole compound is 0.2:1:5, the metal salt comprises zinc nitrate and cobalt nitrate, and the imidazole compound comprises benzimidazole and nitroimidazole, and in the stirring reaction process, the reaction temperature is 4 ℃ and the reaction time is 3000 min.
Example 4:
a magnetic immobilized enzyme nano-reactor is prepared by the following steps:
1) mixing ferric chloride and ferrous chloride with water, and then heating for reaction to obtain ferroferric oxide magnetic nanoparticles;
2) mixing the ferroferric oxide magnetic nanoparticles, polydopamine hydrochloride and a buffer solution, and then stirring for reaction to obtain the ferroferric oxide magnetic nanoparticles with polydopamine attached to the surfaces;
3) mixing ferroferric oxide magnetic nanoparticles with polydopamine attached to the surface, enzyme and water, then adjusting the pH value to 9.0 and stirring, then adjusting the pH value to 7.5 and stirring to obtain a magnetic immobilized enzyme;
4) mixing the magnetic immobilized enzyme, the metal salt and the imidazole compound with water, and then stirring for reaction to obtain the magnetic immobilized enzyme nano reactor.
In the step 1), the mass concentration of ferric chloride in water is 50 g/L, the mass ratio of ferric chloride to ferrous chloride is 5:1, and in the heating reaction process, the reaction temperature is 150 ℃ and the reaction time is 1200 min.
In the step 2), the buffer solution is a Tris-HCl buffer solution, the pH value of the Tris-HCl buffer solution is 8.0, the concentration of the Tris-HCl buffer solution is 50 g/L, the mass concentration of the polydopamine hydrochloride in the buffer solution is 4 g/L, the mass ratio of the ferroferric oxide magnetic nanoparticles to the polydopamine hydrochloride is 6:1, and in the stirring reaction process, the reaction temperature is 24 ℃ and the reaction time is 1300 min.
In the step 3), the mass concentration of the enzyme in water is 4 g/L, the mass ratio of the ferroferric oxide magnetic nanoparticles with polydopamine attached to the surface to the enzyme is 3:1, the enzyme is fat transferase, the pH value is adjusted to 9.0 and then stirred for 2.4 hours, and the pH value is adjusted to 7.5 and then stirred for 2.4 hours.
In the step 4), the mass concentration of the metal salt in water is 7 g/L, the mass ratio of the magnetic immobilized enzyme to the metal salt to the imidazole compound is 2:1:2, the metal salt is zinc nitrate, and the imidazole compound is nitroimidazole, and in the stirring reaction process, the reaction temperature is 23 ℃ and the reaction time is 2100 min.
In conclusion, the method for constructing the magnetic immobilized enzyme nano reactor has the advantages that the required equipment is simple, the separation or recycling of the catalyst can be realized by using the external magnetic field, the method is convenient and quick, and the cost is reduced; the enzyme is wrapped on the surface of the modified magnetic nano-particles by utilizing a pH circulation method, so that the active sites of the enzyme are fully exposed, and the catalytic performance of the enzyme is improved; the metal organic framework as a shell can protect the enzyme from being damaged by the reaction environment, so that the thermal stability, the stability of an organic solvent and the ionic stability are improved; the invention has simple process flow and easy control of reaction conditions, prolongs the service life of the enzyme and saves the cost, so the technology is suitable for large-scale industrial production.
The embodiments described above are described to facilitate an understanding and use of the invention by those skilled in the art. It will be readily apparent to those skilled in the art that various modifications to these embodiments may be made, and the generic principles described herein may be applied to other embodiments without the use of the inventive faculty. Therefore, the present invention is not limited to the above embodiments, and those skilled in the art should make improvements and modifications within the scope of the present invention based on the disclosure of the present invention.
Claims (10)
1. A preparation method of a magnetic immobilized enzyme nano reactor is characterized by comprising the following steps:
1) mixing ferric chloride and ferrous chloride with water, and then heating for reaction to obtain ferroferric oxide magnetic nanoparticles;
2) mixing the ferroferric oxide magnetic nanoparticles, polydopamine hydrochloride and a buffer solution, and then stirring for reaction to obtain the ferroferric oxide magnetic nanoparticles with polydopamine attached to the surfaces;
3) mixing ferroferric oxide magnetic nanoparticles with polydopamine attached on the surface, enzyme and water, then firstly adjusting the pH value to 8.0-10.0 and stirring, then adjusting the pH value to 7.0-8.0 and stirring to obtain magnetic immobilized enzyme;
4) mixing the magnetic immobilized enzyme, the metal salt and the imidazole compound with water, and then stirring for reaction to obtain the magnetic immobilized enzyme nano reactor.
2. The preparation method of the magnetic immobilized enzyme nanoreactor according to claim 1, wherein in step 1), the mass concentration of the ferric chloride in water is 0.01-100 g/L, and the mass ratio of the ferric chloride to the ferrous chloride is (0.1-10): 1.
3. The method for preparing a magnetic immobilized enzyme nanoreactor as claimed in claim 1, wherein in the step 1), the reaction temperature is 100-200 ℃ and the reaction time is 1-2000min during the heating reaction.
4. The preparation method of the magnetic immobilized enzyme nanoreactor according to claim 1, wherein in the step 2), the buffer solution is Tris-HCl buffer solution, the pH value of the Tris-HCl buffer solution is 7.0-10.0, the concentration of the Tris-HCl buffer solution is 0.01-100 g/L, the mass concentration of the polydopamine hydrochloride in the buffer solution is 0.01-15 g/L, and the mass ratio of the ferroferric oxide magnetic nanoparticles to the polydopamine hydrochloride is (0.1-10): 1.
5. The preparation method of the magnetic immobilized enzyme nanoreactor according to claim 1, wherein in the step 2), the reaction temperature is 4-40 ℃ and the reaction time is 1-2000min during the stirring reaction.
6. The preparation method of the magnetic immobilized enzyme nanoreactor according to claim 1, wherein in the step 3), the mass concentration of the enzyme in water is 0.01-10 g/L, the mass ratio of the ferroferric oxide magnetic nanoparticles with polydopamine attached to the surface to the enzyme is (0.2-5):1, the enzyme comprises one or more of lipotransferase or lipohydrolase, the pH value is adjusted to 8.0-10.0, then the stirring is carried out for 0.1-5.0h, and the pH value is adjusted to 7.0-8.0, then the stirring is carried out for 0.1-5.0 h.
7. The preparation method of the magnetic immobilized enzyme nanoreactor according to claim 1, wherein in the step 4), the mass concentration of the metal salt in water is 0.01-15 g/L, the mass ratio of the magnetic immobilized enzyme, the metal salt and the imidazole compound is (0.2-5):1 (0.5-5), the metal salt comprises one or more of zinc chloride, zinc nitrate or cobalt nitrate, and the imidazole compound comprises one or more of dimethyl imidazole, benzimidazole or nitroimidazole.
8. The preparation method of the magnetic immobilized enzyme nanoreactor according to claim 1, wherein in the step 4), the reaction temperature is 4-40 ℃ and the reaction time is 10-3000min during the stirring reaction.
9. A magnetic immobilized enzyme nanoreactor, characterized in that it is prepared by the method of any one of claims 1 to 8.
10. Use of the magnetic immobilized enzyme nanoreactor of claim 9 in biocatalytic reactions.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010313183.8A CN111411105A (en) | 2020-04-20 | 2020-04-20 | Magnetic immobilized enzyme nano reactor and preparation method and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010313183.8A CN111411105A (en) | 2020-04-20 | 2020-04-20 | Magnetic immobilized enzyme nano reactor and preparation method and application thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN111411105A true CN111411105A (en) | 2020-07-14 |
Family
ID=71490066
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202010313183.8A Withdrawn CN111411105A (en) | 2020-04-20 | 2020-04-20 | Magnetic immobilized enzyme nano reactor and preparation method and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN111411105A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112029758A (en) * | 2020-08-12 | 2020-12-04 | 华南理工大学 | Multi-enzyme immobilization material and preparation method and application thereof |
-
2020
- 2020-04-20 CN CN202010313183.8A patent/CN111411105A/en not_active Withdrawn
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112029758A (en) * | 2020-08-12 | 2020-12-04 | 华南理工大学 | Multi-enzyme immobilization material and preparation method and application thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Liu et al. | Advances on methods and easy separated support materials for enzymes immobilization | |
| Liu et al. | Smart chemistry of enzyme immobilization using various support matrices–a review | |
| Du et al. | Enzyme nanocapsules armored by metal-organic frameworks: A novel approach for preparing nanobiocatalyst | |
| Sheng et al. | Enhanced activity and stability of papain by covalent immobilization on porous magnetic nanoparticles | |
| CN107267494A (en) | The@Fe of enzyme@ZIF 83O4Magnetic Nano enzyme reactor and preparation method thereof | |
| CN111359589B (en) | Chitosan/bacterial cellulose composite aerogel adsorbent and preparation method and application thereof | |
| CN110229807B (en) | Preparation method of organophosphorus immobilized enzyme catalyst, organophosphorus immobilized enzyme catalyst and application thereof | |
| Hou et al. | Formulation of robust organic–inorganic hybrid magnetic microcapsules through hard-template mediated method for efficient enzyme immobilization | |
| CN102206624A (en) | Magnetic composite microsphere immobilized laccase and preparation method thereof | |
| CN106807380B (en) | A kind of copper-based ternary composite metal oxide hollow nano-material, preparation method and application | |
| Wu et al. | Immobilization of carbonic anhydrase for facilitated CO2 capture and separation | |
| Zhang et al. | Preparation of a magnetically recoverable biocatalyst support on monodisperse Fe 3 O 4 nanoparticles | |
| CN106348349B (en) | A kind of high-ratio surface hollow-core construction cobaltosic oxide and its synthetic method and application | |
| CN111411105A (en) | Magnetic immobilized enzyme nano reactor and preparation method and application thereof | |
| Ren et al. | Multiscale immobilized lipase for rapid separation and continuous catalysis | |
| Zhao et al. | Ecofriendly construction of enzyme reactor based on three-dimensional porous cryogel composites | |
| CN114870807A (en) | Ionized magnetic microsphere material | |
| Tian et al. | Synthesis of L-aspartic acid-based bimetallic hybrid nanoflowers to immobilize snailase for the production of rare ginsenoside compound K | |
| Xiao et al. | Immobilization of Candida rugosa lipase (CRL) on a hierarchical magnetic zeolitic imidazole framework-8 for efficient biocatalysis | |
| CN107774227A (en) | A kind of preparation method of ferro manganese composite oxides/ferroso-ferric oxide core-shell nano material | |
| CN105969758B (en) | Immobilised enzymes, magnetic carbon material and preparation method thereof | |
| CN109439647B (en) | Magnetic immobilized enzyme carrier with core-shell structure and preparation method and application thereof | |
| CN116726817A (en) | Polymer-metal organic frame open nanocapsule, preparation method and application thereof | |
| CN112916027A (en) | Method for preparing iron phosphide/carbon composite material by using yeast biomass | |
| CN114805860A (en) | Preparation method of hollow magnetic polymer microspheres |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| WW01 | Invention patent application withdrawn after publication | ||
| WW01 | Invention patent application withdrawn after publication |
Application publication date: 20200714 |