阅读说明：本技术 一种用于废纸脱墨的生物酶制剂、制备方法及脱墨工艺 (Biological enzyme preparation for deinking waste paper, preparation method and deinking process ) 是由 李少涛 吕金荣 孙尚先 于 2021-07-26 设计创作，主要内容包括：本发明具体涉及一种用于废纸脱墨的生物酶制剂、制备方法及脱墨工艺。生物酶技术在造纸领域具有广泛的应用,采用生物酶处理废纸脱墨相比化学处理能够有效减少生产工程中的废水排放。但固定化酶应用于废纸脱墨目前还在存在酶活力下降、回收不方便等缺陷。本发明提供了一种用于废纸脱墨的复合生物酶制剂,所述复合生物酶制剂包括复合酶及载体,所述载体为壳聚糖修饰的磁性纳米粒子。在上述载体的构建中,本发明优化了载体交联工艺,降低了对酶活力的破坏,提供了一种方便回收且更加稳定的脱墨处理剂,处理后纸张具有良好的白度及机械性能,具有较高的经济价值。(The invention particularly relates to a biological enzyme preparation for deinking waste paper, a preparation method and a deinking process. The biological enzyme technology has wide application in the field of papermaking, and compared with chemical treatment, the biological enzyme treatment of waste paper deinking can effectively reduce the waste water discharge in production engineering. However, the immobilized enzyme applied to waste paper deinking still has the defects of reduced enzyme activity, inconvenient recovery and the like. The invention provides a compound biological enzyme preparation for deinking waste paper, which comprises a compound enzyme and a carrier, wherein the carrier is chitosan-modified magnetic nanoparticles. In the construction of the carrier, the invention optimizes the carrier crosslinking process, reduces the damage to the enzyme activity, provides a deinking treatment agent which is convenient to recycle and more stable, and the treated paper has good whiteness and mechanical property and higher economic value.)

1. The composite biological enzyme preparation for deinking waste paper is characterized by comprising a composite enzyme and a carrier, wherein the carrier is chitosan-modified magnetic nanoparticles, and the composite enzyme is selected from one or a combination of several of lipase, cellulase, hemicellulase, pectinase and laccase.

2. The complex bio-enzyme preparation for deinking waste paper of claim 1, wherein said magnetic nanoparticles are metal oxides of iron element;

preferably, it is ZnFe₂O₄Nanoparticles, magnetic Fe₃O₄Nanoparticles, NiFe₂O₄Magnetic nanoparticles and nano-Fe₂O₃Nanoparticles and nano Fe₃S₄One of the nanoparticles; further, it is magnetic Fe₃O₄Nanoparticles of said magnetic Fe₃O₄The nanoparticles were treated with oleic acid.

3. The complex bio-enzyme preparation for deinking waste paper as claimed in claim 1, wherein said complex enzyme contains at least cellulase in a combination of cellulase and other enzymes, and examples of said combination include a combination of cellulase and laccase, a combination of lipase and cellulase or a combination of cellulase and pectinase.

4. The complex bioenzyme preparation for deinking waste paper according to claim 3, wherein the cellulase is selected from the group consisting of endoglucanase, exoglucanase or cellobiase, β -glucosidase;

or, the hemicellulase is selected from xylanase, beta-glucanase, galactanase, or xylanase;

or, the pectinase is selected from the group consisting of protopectinase, polygalacturonase, or lyase.

5. The method for preparing the composite biological enzyme preparation for deinking waste paper of any one of claims 1 to 4, wherein the chitosan is surface-modified by using polyethyleneimine, and the magnetic nanoparticles are added into the modified chitosan system to be combined to obtain the chitosan-modified magnetic nanoparticles.

6. The method for preparing the composite biological enzyme preparation for deinking waste paper according to claim 5, which comprises the following specific steps: adding polyethyleneimine into an acidic solution of chitosan, and heating under the condition of isolating oxygen to modify the surface of chitosan; inverse directionAdding magnetic Fe after finishing₃O₄Then incubating for a period of time to obtain the chitosan modified magnetic nanoparticles; adding the magnetic nanoparticles into a buffer solution of a complex enzyme for incubation to obtain the complex biological enzyme preparation;

preferably, the chitosan acid solution is prepared as follows: dissolving chitosan by adopting a 10% acetic acid solution, wherein the ratio of the chitosan to the acetic acid solution is 3-10 g: 150-400 mL;

preferably, the concentration of the polyethyleneimine is 8% -12%;

preferably, the chitosan surface modification mode is as follows: adding chitosan into an acetic acid solution for dissolving, adding a polyethyleneimine solution, and then placing the mixed solution in an inert gas atmosphere for heating; the heating temperature is 70-80 ℃, and the heating time is 1-2 hours;

preferably, the magnetic Fe₃O₄Adding the nano particles into the modified chitosan solution for fully mixing, wherein the mixing mode comprises one or the combination of vibration, stirring and ultrasound;

preferably, the magnetic nanoparticles are added into a buffer solution of the compound biological enzyme, and the shake incubation is carried out for 2-3 h at room temperature to obtain the compound biological enzyme preparation.

7. A composition comprising at least the complex bio-enzyme preparation of any of claims 1 to 4 for deinking waste paper, and a surfactant.

8. The composition of claim 7, wherein the surfactant is an anionic surfactant or a nonionic surfactant; further, the surfactant is one or a combination of more of potassium soap, sulfonated oil, linear alkyl benzene sodium sulfonate, fatty alcohol sodium sulfate, sulfo succinic acid zinc acetate sodium salt, fatty alcohol polyoxyethylene sodium sulfate, alkylphenol, fatty alcohol and propylene oxide polymer.

9. A method for deinking waste paper, comprising treating the pulp to be deinked with the complex biological enzyme preparation of any one of claims 1 to 4 or the composition of claim 7 or 8.

10. The method for deinking waste paper as claimed in claim 9, wherein said method for deinking waste paper comprises the steps of: crushing waste paper to be deinked, adding a surfactant solution for pretreatment, and pulping to obtain waste paper pulp; adding the composite biological enzyme preparation into waste paper pulp for deinking;

preferably, the weight ratio of the surfactant to the waste paper is 0.1-0.5: 100, respectively;

preferably, the concentration of the waste paper pulp is 30-50 g/L;

preferably, the addition amount of the complex enzyme preparation is 0.2-1.5% of the mass of the oven-dried waste paper pulp;

preferably, the treatment temperature of the complex enzyme preparation is 30-75 ℃, the treatment time is 25-120 min, the pH is 6.0-9.0, and the stirring speed is 100-150 rpm;

preferably, the deinking process further comprises a process of bleaching the deinked pulp, wherein the bleaching can be performed by a method which is conventional in the field, such as hydrogen peroxide bleaching and the like.

Technical Field

The invention belongs to the technical field of waste paper deinking, and particularly relates to a biological enzyme preparation for waste paper deinking, a preparation method thereof and a deinking process using the composite biological enzyme preparation.

Background

The information in this background section is only for enhancement of understanding of the general background of the invention and is not necessarily to be construed as an admission or any form of suggestion that this information forms the prior art that is already known to a person of ordinary skill in the art.

The pulping and papermaking industry is a large household using water in the prior art, is also an important pollution source causing water pollution in China, and has the problem that papermaking sewage is always subjected to scaling due to serious pollution. The recycling of waste paper is an effective countermeasure for solving the problems of shortage of raw materials, energy shortage and serious pollution in the paper industry at present, and in the production of waste paper as a raw material, the waste paper is first required to be removed of ink, resin and the like, and the process of the treatment is called waste paper deinking. The deinking process at present mainly comprises the following three steps: (1) defibering and separating fibers; (2) releasing the ink from the fibers; (3) the removed ink particles are removed from the slurry. Among them, the deinking process is particularly important for the recycled paper to meet the use requirement. Modern large deinking lines typically employ flotation or a process in which flotation is the primary wash. The two deinking processes are chemical deinking, consume certain chemicals, easily cause environmental pollution, simultaneously reduce the performance of secondary fibers, and easily cause alkaline blackening of paper pulp.

In order to improve the influence of the paper industry on the environment, the application of biotechnology in the paper industry is gradually increased, and the application comprises material preparation, pulping, fiber performance improvement, pitch obstacle control, bleaching, secondary fiber deinking, wastewater treatment, sludge biological composting and the like. Compared with the traditional chemical treatment mode, the enzymatic treatment does not need a large amount of chemical reagents, can obviously reduce the sewage generation amount, and has development significance because the fiber performance after the enzymatic treatment is superior to that of the chemical reagent treatment method. In the waste paper deinking process, the water absorption swelling capacity of waste paper fibers can be well improved, the strength of regenerated paper is improved, and the printing energy consumption is reduced by adopting biological enzyme deinking; commonly used biological enzymes include cellulases, lipases, laccases, and mixed enzymes. However, the cost of the enzyme treatment agent is higher than that of the chemical agent, and the release treatment using the free enzyme has the disadvantages of poor stability, difficulty in product separation and incapability of recycling. The immobilized enzyme can realize the stability improvement and the recycling of enzyme products, reduce the cost of an enzyme treatment process and overcome the technical defect of instability of free enzyme.

Currently, means for immobilizing enzymes include immobilizing enzymes on stationary phases of inorganic or organic materials by means of physical or chemical bonding, so as to achieve recycling of enzyme products. Immobilization carriers commonly used in the art include organic polymers such as chitosan and resin materials, and inorganic materials such as porous glass, silica gel, and activated carbon. The inorganic carrier material has good mechanical strength, wider sources and lower cost, but has the defects of difficult structure regulation and poor capability of bond synthase, leads the immobilized enzyme to be easy to desorb and inactivate, adopts organic polymer for modification and can complement so as to exert the excellent characteristics of the two,

disclosure of Invention

Against the above technical background, the present inventors have aimed to provide an immobilized enzyme product suitable for use in a wastepaper deinking process. The inventor conjectures that the immobilized enzyme needs to be added with a cross-linking agent to link the enzyme and the carrier in the preparation process, and the chemical combination mode can destroy a part of active sites of the enzyme, so that the enzyme activity is reduced. The immobilized enzyme product and the waste paper pulp are indispensable in the separation process and can generate a part of loss. In order to improve the enzyme catalysis efficiency of waste paper deinking, the invention provides a composite biological enzyme preparation, which takes chitosan modified magnetic nano particles as a carrier to load deinking composite enzyme.

The invention provides a compound biological enzyme preparation for deinking waste paper, which comprises a compound enzyme and a carrier, wherein the carrier is chitosan-modified magnetic nanoparticles, and the compound enzyme is selected from one or more of lipase, cellulase, hemicellulase, pectinase and laccase.

In the compound biological enzyme preparation of the first aspect, the magnetic nanoparticles are attached to the surface of chitosan, and the compound enzyme is embedded in a network structure formed by crosslinking chitosan through a group adsorption effect.

Aiming at the selection of the complex enzyme, the invention discovers that the cellulase has better affinity to the vector constructed by the invention, and the loss of enzyme activity is smaller by adopting the biological enzyme preparation constructed by compounding the cellulase and other enzymes.

In a second aspect of the present invention, a preparation method of the composite bio-enzyme preparation for deinking waste paper in the first aspect is provided, wherein the preparation method is characterized in that the surface of chitosan is modified by using polyethyleneimine, and magnetic nanoparticles are added into a modified chitosan system to be combined to obtain chitosan-modified magnetic nanoparticles.

The existing research shows that immobilized enzyme has certain enzyme activity reduction compared with free enzyme, which is probably because certain loss occurs in the immobilization process of the enzyme, the process that the enzyme is connected with a carrier through a cross-linking reaction tends to occupy a part of enzyme activity sites, and the carrier also causes a part of steric hindrance on the contact of the enzyme and a substrate. The invention designs a method for improving the immobilization of enzyme, reduces the use of cross-linking agent, and promotes the catalytic efficiency of enzyme by matching with active metal groups. Therefore, in the preparation method provided by the invention, the adsorption effect of the enzyme and the carrier is realized by increasing the amino coordination group without adopting traditional cross-linking agents such as glutaraldehyde. The composite biological enzyme preparation obtained by the preparation method has good stability, the enzyme activity is not obviously reduced after repeated use, and the composite biological enzyme preparation can be conveniently recycled through an external magnetic field by depending on the action of the magnetic nanoparticles in the carrier.

In a third aspect of the invention, a composition is provided, which comprises at least the complex bio-enzyme preparation for deinking waste paper of the first aspect, and further comprises a surfactant.

In the deinking process in the prior art, the waste paper is generally pretreated by adopting a surfactant, and the swelling effect of the surfactant on fibers and surface ink is realized, so that the catalytic efficiency of enzyme treatment is improved. In the research aiming at the deinking process, the anionic or nonionic surfactant and the complex enzyme biological agent in the first aspect are combined for application, so that the paper treatment effect is better.

In a fourth aspect of the invention, there is provided a method for deinking waste paper, the method comprising treating the pulp to be deinked with the complex biological enzyme preparation of the first aspect or the composition of the third aspect.

Drawings

The accompanying drawings, which are incorporated in and constitute a part of this specification, are included to provide a further understanding of the invention, and are incorporated in and constitute a part of this specification, illustrate exemplary embodiments of the invention and together with the description serve to explain the invention and not to limit the invention.

FIG. 1 is a scanning electron micrograph of the complex biological enzyme described in example 1.

Detailed Description

It is to be understood that the following detailed description is exemplary and is intended to provide further explanation of the invention as claimed. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

It is noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of exemplary embodiments according to the invention. As used herein, the singular forms "a", "an" and "the" are intended to include the plural forms as well, and it should be understood that when the terms "comprises" and/or "comprising" are used in this specification, they specify the presence of stated features, steps, operations, devices, components, and/or combinations thereof, unless the context clearly indicates otherwise.

As introduced in the background art, immobilized enzymes often suffer from the defects of reduced enzyme activity, incomplete separation and the like. The invention aims to provide a stripping agent which is convenient and rapid to recover and has more stable enzyme activity.

The invention provides a compound biological enzyme preparation for deinking waste paper, which comprises a compound enzyme and a carrier, wherein the carrier is chitosan-modified magnetic nanoparticles, and the compound enzyme is selected from one or more of lipase, cellulase, hemicellulase, pectinase and laccase.

In the technical solution of the first aspect, the magnetic nanoparticles are metal oxides of iron element, and further are ZnFe₂O₄Nanoparticles, magnetic Fe₃O₄Nanoparticles、NiFe₂O₄Magnetic nanoparticles and nano-Fe₂O₃Nanoparticles and nano Fe₃S₄One kind of nano particle.

In one embodiment of the present invention, the magnetic nanoparticles are magnetic Fe₃O₄In a specific embodiment, the magnetic Fe₃O₄The nanoparticles are not limited to a specific preparation method, and are magnetic Fe prepared by a coprecipitation method, hydrothermal synthesis method, or the like known in the art₃O₄The nano particles can meet the catalytic deinking effect of the complex enzyme preparation. The composite biological enzyme preparation adopts magnetic Fe₃O₄The nano particles are combined with the chitosan, so that the complex enzyme preparation can be conveniently recycled through an external magnetic field.

In a further preferred embodiment, the magnetic Fe₃O₄The surface of the nano particle is treated by oleic acid: magnetic Fe₃O₄The nano particles are placed in an oleic acid/absolute ethyl alcohol solution for treatment, and after the treatment is finished, the nano particles are alternately cleaned by water and absolute ethyl alcohol.

In a preferred embodiment of the first aspect, the complex enzyme at least contains cellulase, and is in the form of a combination of cellulase and other enzymes, and examples of the combination include a combination of cellulase and laccase, a combination of lipase and cellulase, or a combination of cellulase and pectinase.

Further, the cellulase is selected from endoglucanase, exoglucanase or cellobiase, and beta-glucosidase.

Further, the hemicellulase is selected from xylanase, beta-glucanase, galactanase or xylanase.

Further, the pectinase is selected from protopectinase, polygalacturonase or lyase.

In a second aspect of the present invention, a preparation method of the composite bio-enzyme preparation for deinking waste paper in the first aspect is provided, wherein the preparation method is characterized in that the surface of chitosan is modified by using polyethyleneimine, and magnetic nanoparticles are added into a modified chitosan system to be combined to obtain chitosan-modified magnetic nanoparticles.

Preferably, the preparation method comprises the following specific steps: adding polyethyleneimine into an acidic solution of chitosan, and heating under the condition of isolating oxygen to modify the surface of chitosan; adding magnetic Fe after the reaction is finished₃O₄Then incubating for a period of time to obtain the chitosan modified magnetic nanoparticles; and adding the magnetic nanoparticles into a buffer solution of a complex enzyme for incubation to obtain the complex biological enzyme preparation.

Further, the chitosan acid solution is prepared as follows: dissolving chitosan by adopting a 10% acetic acid solution, wherein the ratio of the chitosan to the acetic acid solution is 3-10 g: 150-400 mL.

When the acid solution of chitosan is prepared, chitosan solid powder can be added into acetic acid solution for dissolving for a few times under the condition of continuous stirring; or spraying chitosan directly on the surface of acetic acid solution, and standing to swell completely.

In a preferred embodiment, the chitosan is prepared in an acidic solution as follows: 5g of chitosan was added to 200mL of 10% acetic acid solution and left to stand for 0.8h to sufficiently swell.

Further, the concentration of the polyethyleneimine is 8% -12%.

Further, the chitosan surface modification mode is as follows: adding chitosan into an acetic acid solution for dissolving, adding a polyethyleneimine solution, and then placing the mixed solution in an inert gas atmosphere for heating; the heating temperature is 70-80 ℃, and the heating time is 1-2 hours.

Further, the magnetic Fe₃O₄Adding the nano particles into the modified chitosan solution for fully mixing, wherein the mixing mode comprises one or the combination of vibration, stirring and ultrasound; in the embodiment with better effect, the chitosan modified magnetic nanoparticles are obtained by low-speed stirring after a period of ultrasonic treatment.

Further, adding the magnetic nanoparticles into a buffer solution of the composite biological enzyme, and performing shake incubation for 2-3 h at room temperature to obtain the composite biological enzyme preparation.

In a third aspect of the invention, a composition is provided, which comprises at least the complex bio-enzyme preparation for deinking waste paper of the first aspect, and further comprises a surfactant.

Preferably, the surfactant is an anionic surfactant or a nonionic surfactant; further, the surfactant is one or a combination of more of potassium soap, sulfonated oil, linear alkyl benzene sodium sulfonate, fatty alcohol sodium sulfate, sulfo succinic acid zinc-sodium acetate, fatty alcohol polyoxyethylene sodium sulfate, alkylphenol (octylphenol, nonylphenol), fatty alcohol and propylene oxide polymer.

In a fourth aspect of the invention, there is provided a method for deinking waste paper, the method comprising treating the pulp to be deinked with the complex biological enzyme preparation of the first aspect or the composition of the third aspect.

Preferably, the wastepaper deinking method comprises the steps of: crushing waste paper to be deinked, adding a surfactant solution for pretreatment, and pulping to obtain waste paper pulp; adding the composite biological enzyme preparation of the first aspect into waste paper pulp for deinking.

Further, the weight ratio of the surfactant to the waste paper is 0.1-0.5: 100.

further, the concentration of the waste paper pulp is 30-50 g/L.

Further, the addition amount of the complex enzyme preparation is 0.2-1.5% of the mass of the oven-dried waste paper pulp.

Further, the treatment temperature of the complex enzyme preparation is 30-75 ℃, the treatment time is 25-120 min, the pH is 6.0-9.0, and the stirring speed is 100-150 rpm.

Further, the deinking process also comprises a process of bleaching deinking pulp, and the bleaching can be carried out by adopting a conventional method in the field, such as hydrogen peroxide bleaching and the like.

In order to make the technical solution of the present invention more clearly understood by those skilled in the art, the technical solution of the present invention will be described in detail below with reference to specific examples and comparative examples.

Example 1

In this embodiment, a composite bio-enzyme for deinking waste paper is provided, where the composite bio-enzyme includes a carrier and a composite enzyme, and the carrier is chitosan-modified magnetic Fe₃O₄The composite enzyme is laccase and lipase, and the preparation method of the composite biological enzyme comprises the following steps:

1) preparation of magnetic Fe by coprecipitation method₃O₄Nanoparticle:

2.1g FeCl₃·4H₂O and 5.5g FeCl₃·6H₂O dissolved in 100mL deionized water in N₂Heating water bath to 80 ℃ in the atmosphere, slowly dropwise adding concentrated ammonia water into the mixed solution until the solution is alkalescent, violently stirring for 1h at the rotating speed of 500rpm, separating by an additional magnet after the reaction is finished, washing with deionized water until the pH of the solution is neutral, and finally performing vacuum drying to obtain magnetic Fe₃O₄Nanoparticles.

2) Chitosan modified magnetic nanoparticles:

adding 6g of chitosan into 250mL of 10% acetic acid solution for 1 hour to fully swell the chitosan, adding 20mL of 15% polyethyleneimine solution, and placing the reaction system in N₂Heating to 80 ℃ in the atmosphere for reaction for 2h to modify the surface of the chitosan, cooling the temperature of the reaction system to room temperature after the reaction is finished, and carrying out magnetic Fe preparation in the step 1)₃O₄Adding the modified solution and placing the reaction system in an ultrasonic environment for treatment for 10 min; stirring the mixed solution after ultrasonic treatment for 3 hours at the rotating speed of 90r/min, dropwise adding a NaOH solution with the mass fraction of 3% in the stirring process, adjusting the pH of the mixed solution to 7.3, magnetically separating a product after the reaction is finished, and alternately cleaning the product with water and absolute ethyl alcohol to obtain the chitosan modified magnetic nanoparticles.

3) Mixing laccase and lipase in a mass ratio of 1: 1 is dissolved in PBS buffer solution with the concentration of 0.02mol/L and the pH value is 7.5 to prepare 0.1mg/mL enzyme buffer solution, 10mg of the chitosan modified magnetic nano particles prepared in the step 2) is weighed and added into the phosphate buffer solution, the mixed system is placed at 150rpm to be oscillated and incubated at room temperature for 3h, and then magnetic separation is carried out to obtain the compound biological enzyme preparation.

Example 2

In this embodiment, a composite bio-enzyme for deinking waste paper is provided, where the composite bio-enzyme includes a carrier and a composite enzyme, and the carrier is chitosan-modified magnetic Fe₃O₄The composite enzyme is composed of pectinase and lipase, and the preparation method of the composite biological enzyme comprises the following steps:

1) preparation of magnetic Fe by microwave hydrothermal method₃O₄Nanoparticle:

with FeSO₄·H₂O、Fe(NO₃)₃·H₂Preparation of magnetic Fe from O₃O₄Nano particles, modifying the surface of the product by ethanol of oleic acid, washing, centrifuging and drying the product by distilled water and absolute ethanol to obtain the magnetic Fe₃O₄Nanoparticles.

2) Chitosan modified magnetic nanoparticles:

adding 5g of chitosan into 200mL of 10% acetic acid solution for 0.8h to fully swell the chitosan, adding 15mL of 10% polyethyleneimine solution, and placing the reaction system in N₂Heating to 75 ℃ in the atmosphere to react for 1.5h to modify the surface of the chitosan, and after the reaction is finished, cooling the temperature of the reaction system to room temperature to obtain the magnetic Fe prepared in the step 1)₃O₄Adding the modified solution and placing the reaction system in an ultrasonic environment for treatment for 15 min; stirring the mixed solution after ultrasonic treatment for 4 hours at the rotating speed of 50r/min, dropwise adding a NaOH solution with the mass fraction of 2%, adjusting the pH of the mixed solution to 7.0, magnetically separating a product after the reaction is finished, and alternately cleaning the product by using water and absolute ethyl alcohol to obtain the chitosan modified magnetic nanoparticles.

3) Mixing cellulase and lipase in a mass ratio of 0.5: dissolving the chitosan 1 in PBS buffer solution with the pH value of 7.0 and the concentration of 0.02mol/L to prepare 0.05mg/mL enzyme buffer solution, weighing 8mg of the chitosan modified magnetic nanoparticles prepared in the step 2) and adding the chitosan modified magnetic nanoparticles into the phosphate buffer solution, placing the mixed system at 100rpm, oscillating, incubating at room temperature for 2.5h, and then carrying out magnetic separation to obtain the compound biological enzyme preparation.

Example 3

In this embodiment, a composite bio-enzyme for deinking waste paper is provided, where the composite bio-enzyme includes a carrier and a composite enzyme, and the carrier is chitosan-modified magnetic Fe₃O₄The composite enzyme is cellulase and lipase, and the preparation method of the composite biological enzyme comprises the following steps:

1) preparation of magnetic Fe by ammonia coprecipitation method₃O₄Nanoparticle:

configuration of 0.5mol/LFeCl₃And FeSO₄Solution according to Fe³⁺:Fe²⁺1.75: 1, adding analytically pure ammonia water, continuously stirring and slowly dropwise adding the ammonia water until the pH value is 8.5, separating the obtained black sol-like substance by a magnetic field, washing, and adding the black sol-like substance into an oleic acid/absolute ethyl alcohol solution for treatment. Then washing, centrifuging and drying the product by adopting distilled water and absolute ethyl alcohol to obtain the magnetic Fe₃O₄Nanoparticles.

2) Chitosan modified magnetic nanoparticles:

adding 8g of chitosan into 300mL of 10% acetic acid solution for 1.5h to fully swell the chitosan, adding 25mL of 20% polyethyleneimine solution, and placing the reaction system in N₂Heating to 85 ℃ in the atmosphere for reaction for 2.5h to modify the surface of the chitosan, and after the reaction is finished, cooling the temperature of the reaction system to room temperature to obtain the magnetic Fe prepared in the step 1)₃O₄Adding the modified solution, stirring the mixed solution for 8 hours at the rotating speed of 80r/min, dropwise adding a NaOH solution with the mass fraction of 3%, adjusting the pH of the mixed solution to 7.3, magnetically separating a product after the reaction is finished, and alternately cleaning the product with water and absolute ethyl alcohol to obtain the chitosan modified magnetic nanoparticles.

3) Mixing cellulase and pectinase in a mass ratio of 0.5: dissolving 1 in PBS buffer solution with the concentration of 0.02mol/L and the pH value of 7.0 to prepare 0.2mg/mL enzyme buffer solution, weighing 15mg of the chitosan modified magnetic nanoparticles prepared in the step 2) and adding the chitosan modified magnetic nanoparticles into the phosphate buffer solution, placing the mixed system at 100rpm, oscillating, incubating at room temperature for 3.5h, and then carrying out magnetic separation to obtain the compound biological enzyme preparation.

Example 4

In this embodiment, there is also provided a waste paper deinking method, including crushing office waste paper to be treated into paper pieces, soaking in water, dehydrating to a water content of about 30%, and adding an aqueous solution of sodium fatty alcohol sulfate to the dehydrated office waste paper, wherein a weight ratio of the sodium fatty alcohol sulfate to the waste paper is 0.5: 100, soaking for 2 hours.

After soaking, adding the waste paper and the solution into a beater in batches to be beaten into paper pulp, adding the medium-complex biological enzyme preparation prepared in any one of the embodiments 1-3 with the absolute dry mass of 1% to be uniformly mixed, continuously adding water until the concentration of the slurry is reduced to 8g/L, treating at 37 ℃ for 1.5h, and stirring at 100 rpm.

Comparative example 1

In the research process of the invention, in order to enhance the coordination capacity of chitosan and metal particles, other preparation methods for providing a coordinating amino group are also researched. The preparation method comprises the following steps of grafting the chitosan with ethylenediamine, and is different from the embodiment 1 in that: 8g of chitosan was added to 300mL of a 10% acetic acid solution for 1.5 hours to sufficiently swell the chitosan, and 25mL of a 20% ethylenediamine methanol solution was added thereto to react in a water bath at 67 ℃.

In other embodiments, the chitosan is modified by triethylene tetramine.

Comparative example 2

In this embodiment, a composite bio-enzyme preparation coated with different bio-enzymes is provided, which is different from embodiment 1 in that in step 3) of this embodiment, the composite enzyme is laccase or cellulase; the laccase and the cellulase are mixed in a mass ratio of 0.5: 1 mixing and dissolving in a buffer solution, and carrying out mixed incubation with the chitosan modified nano particles.

Comparative example 3

In this embodiment, a composite bio-enzyme preparation coated with different bio-enzymes is provided, which is different from embodiment 1 in that in step 3) of this embodiment, the composite enzyme is pectinase and cellulase; the mass ratio of the pectinase to the cellulase is 0.5: 1 mixing and dissolving in a buffer solution, and carrying out mixed incubation with the chitosan modified nano particles.

Comparative example 4

In this example, a waste paper deinking method is provided, which is different from example 4 in that a cationic surfactant is used to pretreat crushed waste paper.

The enzyme activity of the complex enzyme preparation prepared in the above examples 1 to 3 and comparative examples 1 to 3 was measured, and the enzyme activity was evaluated by a Wootton (Worthington) method

The enzyme activity was calculated as follows:

wherein:

E₅₁₀: an increase in absorbance per minute at 510nm during the enzyme-catalyzed reaction;

v: the total volume of the reaction solution was 3.0 mL;

Δ A: specific absorption coefficient of the product, 6.58cm²(μmol·dye)^-1；

v: the volume of enzyme solution was added, and the value was 0.1 mL.

The results of the enzyme activity test described in the examples and comparative examples, and the results of the enzyme activity after deinking treatment are shown in the following table 1:

TABLE 1

As can be seen from the data in Table 1, the composite biological enzyme prepared in examples 1-3 still has higher enzyme activity after being used for many times, while the composite enzyme preparation in comparative examples 1-3 has lower enzyme activity, which may indicate that the cellulase has better affinity effect on the vector provided by the invention and can realize higher stability.

In the invention, the performance of the paper pulp treated by the composite biological enzyme preparation in the examples and the comparative examples is tested, the paper pulp after deinking in the examples is made into pulp sheets, and the pulp sheets are air-dried, dried and quantified by 60g/m²The sheet of paper of (1). Wherein, the paper whiteness test method is specified in GB/T7974-:

TABLE 2

As can be seen from the above data, the waste paper pulp treated by the composite biological enzyme preparation described in examples 1-3 has better whiteness, which proves that the deinking effect is better. In particular, the pulp whiteness after the treatment with the cationic surfactant in comparative example 4 has a significant deficiency compared to the other groups, which may indicate that the cationic surfactant may reduce the catalytic efficiency of the complex bio-enzyme preparation.

The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.