A method for photoenzymatic synergistic catalysis of Microcystis aeruginosa based on nanozymes
By employing a photocatalytic synergistic method combining nano-zero-valent silver/copper nanozymes and peroxides, the problem of low removal efficiency of Microcystis aeruginosa in traditional methods has been solved, achieving efficient and rapid water body restoration.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- FUZHOU UNIV
- Filing Date
- 2026-05-28
- Publication Date
- 2026-07-10
AI Technical Summary
Existing technologies are insufficient for efficiently removing Microcystis aeruginosa from water. Traditional methods are costly, time-consuming, and may cause secondary pollution. The preparation of integrated catalysts combining nanozymes and photocatalysts is difficult, and enzyme molecules are prone to inactivation.
By employing zero-valent silver/copper nanozymes and peroxides for synergistic catalysis, a suspension of Microcystis aeruginosa was mixed with nanozymes under light conditions. The photogenerated electrons released by the photocatalyst excited the active sites of the nanozymes, leading to a redox reaction that disrupted the integrity of the algal cell wall and membrane.
It significantly improves algae removal efficiency. The photo-enzyme synergistic catalysis method has an algae removal capacity that is 1.35 times that of enzyme catalysis and 11 times that of photocatalysis within 20 minutes, thus enhancing the removal effect of Microcystis aeruginosa.
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Abstract
Description
Technical Field
[0001] This invention relates to the field of environmental water treatment technology, specifically to a method for removing Microcystis aeruginosa based on photoenzyme synergistic catalysis using nanozymes. Background Technology
[0002] Currently, the rapid pace of industrialization and urbanization is leading to the discharge of industrial and domestic wastewater containing large amounts of nitrogen and phosphorus into natural water bodies, exacerbating eutrophication. The massive algal blooms caused by eutrophication have become a widespread concern in drinking water production. The high tolerance and toxin-producing capacity of algae make them a significant source of pollution in the aquatic environment. Traditional methods for controlling algal blooms include physical methods (such as mechanical removal and ultrasonic algae removal), chemical methods (addition of algaecides), and biological methods. However, these methods are often limited (e.g., high manpower and material costs, small effective range, secondary pollution, and geographical constraints). Furthermore, due to electrostatic repulsion and spatial effects, cyanobacterial cells are very stable in water and difficult to remove in traditional drinking water treatment processes. Researchers have employed various methods to enhance cyanobacterial cell removal, including chlorination, ozone oxidation, and ultraviolet radiation. However, these methods are often costly and time-consuming, leading to the release of intracellular organic matter and the production of disinfection byproducts. Therefore, finding a practical and gentle algae removal method is crucial for water body restoration.
[0003] In recent years, the construction of a photocatalytic-enzyme synergistic catalytic system by combining biological enzymes and photocatalysts has been considered a potential strategy to improve pollutant degradation capabilities and has attracted widespread attention from researchers. By combining the interdisciplinary advantages of photocatalysis and enzyme catalysis, the photocatalyst-enzyme hybrid system has efficiently carried out various light-driven biotransformations under environmentally friendly conditions, and is seen as a promising pathway to achieving green chemistry using solar energy. This system can generate synergistic catalytic effects between photocatalysis and biocatalysis, constructing a photo / enzyme heterojunction, and utilizing solar energy to provide energy and substrates for the enzyme catalytic process. The photocatalyst releases photogenerated electrons, which can be readily absorbed by the active sites of the enzyme and participate in redox reactions. The natural enzyme consumes electrons and inhibits the recombination of electron-hole pairs in the photocatalyst, further improving catalytic efficiency.
[0004] However, the preparation of such integrated catalysts still faces a series of challenges. This is because chemical and enzyme catalysts typically operate in different media at different temperatures and can deactivate each other. For example, when a photocatalyst is in direct contact with an enzyme molecule, although adjacent active sites can enhance mass transfer and accelerate the reaction process, photoexcitation-generated free radicals can lead to enzyme deactivation. Furthermore, the presence of natural enzymes on the surface of the photocatalyst affects the excitation of electron-hole pairs. More importantly, due to the structural limitations of the photocatalyst itself, low enzyme loading and cycling stability are major drawbacks of photoenzyme catalytic materials. Using nanozymes to replace natural enzymes holds promise for overcoming these problems. Unfortunately, there is currently no research on nanozyme-based photoenzyme synergistic catalytic removal of Microcystis aeruginosa. Summary of the Invention
[0005] To address the aforementioned technical problems, the present invention aims to provide a method for removing Microcystis aeruginosa based on nanozymes and photoenzyme synergistic catalysis, thereby solving the problem of low algae removal efficiency in existing single systems.
[0006] The technical solution of this invention to solve the above-mentioned technical problems is as follows: A method for the photoenzymatic synergistic catalytic removal of Microcystis aeruginosa based on nanozymes is provided, comprising the following steps: Nanoenzymes were added to a suspension of Microcystis aeruginosa, followed by the addition of peroxide, and algae were removed under light conditions. The nanozyme was prepared by the following method: (1) Dissolve silver nitrate and copper nitrate trihydrate in water to prepare solution A; (2) Dissolve sodium borohydride in water to prepare solution B; (3) Add solution B to solution A, wash and vacuum dry to obtain nZVS / nZVC, i.e. nanozyme.
[0007] Based on the above technical solution, the present invention can be further improved as follows: Furthermore, the volume-to-mass ratio of Microcystis aeruginosa suspension to nanozyme is 1L:100-700mg.
[0008] Furthermore, the volume-to-mass ratio of Microcystis aeruginosa suspension to nanozyme was 1L:500mg.
[0009] Furthermore, the density of Microcystis aeruginosa in the Microcystis aeruginosa suspension was OD. 680 =0.1-0.5.
[0010] Furthermore, the density of Microcystis aeruginosa in the Microcystis aeruginosa suspension was OD. 680 =0.2.
[0011] Furthermore, the peroxide is potassium persulfate.
[0012] Furthermore, the concentration of peroxide in the Microcystis aeruginosa suspension was 0.1-2 mmol / L.
[0013] Furthermore, the peroxide concentration in the Microcystis aeruginosa suspension was 1 mmol / L.
[0014] Furthermore, the lighting conditions are LED light sources, tungsten lamps, xenon lamps, or fixed wavelength light sources. Furthermore, the LED light source power is 63W.
[0015] Furthermore, algae removal was carried out under magnetic stirring conditions.
[0016] Furthermore, in step (1), the molar volume ratio of silver nitrate, copper nitrate trihydrate and water is 0.2-0.8 mmol: 0.2-0.8 mmol: 30 mL.
[0017] Furthermore, in step (1), ultrapure water is used.
[0018] Furthermore, in step (2), the mass-to-volume ratio of sodium borohydride and water is 100 mg: 10 mL.
[0019] Furthermore, in step (2), ultrapure water is used.
[0020] Furthermore, in step (3), under stirring and nitrogen atmosphere conditions, solution B is added dropwise to solution A.
[0021] Furthermore, in step (3), the volume ratio of solution B to solution A is 1:3.
[0022] Furthermore, in step (3), water and ethanol are used for centrifugal cleaning in sequence.
[0023] Furthermore, in step (3), the product is vacuum dried at 80°C for 12 hours.
[0024] The present invention also provides the application of the above method in algae removal.
[0025] The present invention has the following beneficial effects: 1. Single nanozyme systems are highly dependent on exogenous oxidants and have low activation efficiency, leading to high oxidant dosage, high costs, and a high risk of secondary pollution. Single photocatalytic systems have rapid photogenerated carrier recombination and low visible light utilization, resulting in limited algae removal efficiency. In the absence of light, the algae removal effect is difficult to guarantee, making it unsuitable for complex natural aquatic environments. Therefore, this invention provides a photocatalytic synergistic catalytic algae removal method based on zero-valent silver / copper nanozymes. This system can generate a synergistic catalytic effect between photocatalysis and nanozyme catalysis, utilizing light energy to provide energy and substrate for the nanozyme catalytic process. The photocatalyst releases photogenerated electrons, which can be easily absorbed by the active sites of the nanozyme, participating in redox reactions and further improving catalytic efficiency. The algae removal effect is significantly better than that of photocatalytic systems and nanozyme catalytic systems.
[0026] 2. Algae removal based on photoenzyme synergistic catalysis: Nanozymes simultaneously serve as both photocatalytic active centers and peroxidase-like active sites. Under the combined action of light and peroxidase catalysis, the efficiency of reactive oxygen species generation can be significantly improved. Compared to single photocatalysis or single nanozyme systems, this invention accelerates the cycling process of the reactive sites, thereby rapidly disrupting the cell wall and cell membrane integrity of *Microcystis aeruginosa*, leading to leakage of intracellular electrolytes and chlorophyll. Within the same treatment time, the algae removal capacity of photoenzyme synergistic catalysis is significantly superior to both photocatalysis and enzyme catalysis. At 20 min, the algae removal capacity of photoenzyme synergistic catalysis is 1.35 times that of enzyme catalysis and 11 times that of photocatalysis. Attached Figure Description
[0027] Figure 1 SEM and EDS images of nZVS, nZVC, and nZVS / nZVC; Figure 2 Crystal structure analysis diagrams for nZVS, nZVC, and nZVS / nZVC; Figure 3 A three-dimensional graph showing the relationship between time, power, and enzyme activity; Figure 4 The fitting results for nZVS / nZVC with H2O2 as the reaction substrate; Figure 5 The fitting results for nZVS / nZVC with TMB as the reaction substrate; Figure 6 Figure 1 shows the effect of light on the removal of Microcystis aeruginosa by nZVS / nZVC. Figure 7 Figure 1 shows the effect of different silver-copper ratios (nZVS / nZVC) on the removal of Microcystis aeruginosa. Figure 8 This is a comparative analysis diagram of photocatalysis, enzyme catalysis, and photoenzyme synergistic catalysis. Detailed Implementation
[0028] The principles and features of the present invention are described below with reference to the accompanying drawings. The examples given are for illustrative purposes only and are not intended to limit the scope of the invention. Unless otherwise specified in the examples, conventional conditions or conditions recommended by the manufacturer should be followed. Reagents or instruments whose manufacturers are not specified are all commercially available products.
[0029] nZVS / nZVC represents nano-zero valence silver / copper.
[0030] Example 1: A method for photoenzymatic synergistic catalytic removal of Microcystis aeruginosa based on nanozymes includes the following steps: (1) Nanozyme preparation (1.1) Add silver nitrate and copper nitrate trihydrate to ultrapure water and sonicate for 30 min to achieve full dissolution of the silver source and copper source to obtain solution A; wherein the molar volume ratio of silver nitrate, copper nitrate trihydrate and ultrapure water is 0.2 mmol: 0.8 mmol: 30 mL; (1.2) Dissolve sodium borohydride in ultrapure water to prepare solution B; wherein the mass-volume ratio of sodium borohydride to ultrapure water is 100 mg: 10 mL; (1.3) Under magnetic stirring and nitrogen protection, solution B was added dropwise to solution A at a volume ratio of 1:3. The solution was washed by centrifugation with water and ethanol in sequence, and then dried under vacuum at 80°C for 12 h to obtain nZVS / nZVC (silver-copper ratio of 2:8), i.e., nanozyme. (2) According to the volume-to-mass ratio of 1L:500mg, in a suspension of Microcystis aeruginosa (the density of Microcystis aeruginosa is OD) 680 =0.2) Add the nanozyme obtained in step (1.3), then add peroxide (potassium persulfate, the peroxide concentration in the Microcystis aeruginosa suspension is 1 mmol / L), place it in a reactor with an LED light source (power of 63W) on top and a magnetic stirrer on the bottom, and remove algae under light conditions.
[0031] Example 2: A method for photoenzymatic synergistic catalysis of Microcystis aeruginosa based on nanozymes includes the following steps: In step (1.1), the molar volume ratio of silver nitrate, copper nitrate trihydrate and water is 0.5 mmol: 0.5 mmol: 30 mL; nZVS / nZVC (silver-copper ratio is 5:5) is obtained, and the rest is the same as in Example 1.
[0032] Example 3: A method for photoenzymatic synergistic catalysis of Microcystis aeruginosa based on nanozymes includes the following steps: In step (1.1), the molar volume ratio of silver nitrate, copper nitrate trihydrate and water is 0.8 mmol: 0.2 mmol: 30 mL; nZVS / nZVC (silver-copper ratio of 8:2) is obtained, and the rest is the same as in Example 1.
[0033] Comparative Example 1: A method for photoenzymatic synergistic catalysis of Microcystis aeruginosa based on nanozymes includes the following steps: In step (1.1), copper nitrate trihydrate is not used to prepare nZVS (nano zero-valent silver), and the rest is the same as in Example 1.
[0034] Comparative Example 2: A method for photoenzymatic synergistic catalysis of Microcystis aeruginosa based on nanozymes includes the following steps: In step (1.1), silver nitrate is not included, and nZVC (nano zero-valent copper) is prepared. The rest is the same as in Example 1.
[0035] Comparative Example 3: A method for photocatalytic removal of Microcystis aeruginosa includes the following steps: In step (2), there is no peroxide, and the rest is the same as in Example 1.
[0036] Comparative Example 4: A method for enzyme-catalyzed removal of Microcystis aeruginosa includes the following steps: In step (2), the LED power is 0 W, and the rest is the same as in Example 1.
[0037] Test case I. Surface Morphology Analysis of nZVS / nZVC 1. The nZVS / nZVC prepared in Example 1, the nZVS prepared in Comparative Example 1, and the nZVC prepared in Comparative Example 2 were subjected to SEM and EDS detection. The results are shown in [Figure 1]. Figure 1 ( Figure 1 In the middle (a)-(c), nZVS is used; (d)-(f) is used; nZVC is used; and (g)-(i) is used; nZVS / nZVC is used.
[0038] 2. Crystal structure analysis was performed on the nZVS / nZVC prepared in Examples 1-3, the nZVS prepared in Comparative Example 1, and the nZVC prepared in Comparative Example 2. The results are shown in [the table below]. Figure 2 .
[0039] Depend on Figure 1 It can be seen that the distributions of Ag, Cu, and O elements are highly overlapping, indicating that nZVS and nZVC are uniformly co-dispersed in the composite material.
[0040] Depend on Figure 2 It can be seen that the characteristic peak of nZVS is related to zero-valent silver (Ag) 0The crystal planes in the standard card (JCPDS No. 04-0783) correspond consistently; the characteristic peaks of nZVC also match those of zero-valent copper (Cu). 0 The standard crystal planes of (JCPDS No. 80-0836) and copper oxide (CuO, JCPDS No. 84-1548) indicate that oxidation exists on the nZVC surface. Ag can also be observed simultaneously in the XRD pattern of the nZVS / nZVC composite material. 0 The characteristic peaks of Ag and Ag are observed, and the Ag content increases with the increase of the nZVS doping ratio. 0 The gradual increase in the intensity of the characteristic peaks and the gradual decrease in the intensity of Cu-related characteristic peaks indicate that different doping ratios successfully controlled the phase composition ratio of the composite material.
[0041] II. Detection of the enhancing effect of light irradiation on the activity of nZVS / nZVC (nanozymes, peroxidase-like enzymes) The effect of light irradiation on enhancing the activity of the nZVS / nZVC nanozyme prepared in Example 1 was detected. The specific detection method was as follows: Step 1: Using 3,3',5,5'-tetramethylbenzidine (TMB) as the enzyme labeling reagent, Berritan-Robisen buffer (pH=4.0), H2O2, TMB, and the nZVS / nZVC nanozyme prepared in Example 1 were added sequentially. The reaction was then carried out at room temperature, and samples were taken at 60s intervals. The absorbance was measured at 652 nm using a spectrophotometer. To better observe the enhancing effect of light on the activity of nZVS / nZVC nanozyme, the light power gradient was set to 0 W (Dark), 63 W, 96 W, and 126 W.
[0042] Step 2: The steady-state kinetic parameters of nZVS / nZVC were calculated. H₂O₂ solutions of different concentrations were prepared, and constant amounts of nanozyme, TMB, and Beritan-Robisen buffer (pH = 4.0) were added to each solution. After reacting for 10 min, the absorbance was measured at 652 nm, and Km and Vmax were calculated according to the Michaelis-Menten equation to evaluate the steady-state kinetic behavior of the photoenzyme catalyst and the substrate H₂O₂ during the reaction. Similarly, TMB solutions of different concentrations were prepared to evaluate the steady-state kinetic behavior of the nanozyme and the substrate TMB during the reaction.
[0043] See results Figure 3-5 ( Figure 4 and Figure 5 (b) are both double reciprocal curves corresponding to (a).
[0044] Depend on Figure 3 It can be seen that the three-dimensional relationship between time, power, and enzyme activity directly reflects the peroxidase-like catalytic activity of nZVS / nZVC nanozymes under different LED light irradiation powers. Under any LED power, OD...652 The values all increased with increasing reaction time, indicating that the oxidation reaction of TMB is a time-dependent process, which is consistent with enzyme-catalyzed reaction kinetics. At the same reaction time, the higher the LED power, the higher the corresponding OD. 652 The higher the value, the better. This indicates that the light power has a positive regulatory effect on the peroxidase-like activity of nanozymes: higher power provides more photons, which can excite the SPR effect of nZVS / nZVC to generate more photogenerated carriers. These carriers participate in the oxidation reaction of TMB, ultimately improving the catalytic efficiency.
[0045] Depend on Figure 4 and Figure 5 It can be seen that, when nZVS / nZVC uses H2O2 and TMB as reaction substrates, under both dark and light conditions, the increase in Vo gradually decreases as the TMB concentration increases from 0.05 mM to 2.00 mM, which is consistent with the Michaelis-Menten behavior. Subsequently, double reciprocal curves were plotted to fit the Michaelis-Menten equation, and the fitting results further confirmed the peroxidase-like characteristics of nZVS / nZVC.
[0046] Depend on Figure 4 As shown in (b), the slope of the fitting curve under dark conditions is [not specified in the original text]. K m / V max =0.62555, intercept 1 / V max =0.21811, calculated as follows K m =2.87, V max =4.58 × 10 -8 M·s -1 The slope of the fitted curve under light conditions. K m / V max =0.35638, intercept 1 / V max =0.21142, calculated as follows K m =1.69, V max =4.73 × 10 -8 M·s -1 .
[0047] Depend on Figure 5 As shown in (b), the slope of the fitting curve under dark conditions is [not specified in the original text]. Km / V max =0.60503, intercept 1 / V max =0.26144, calculated as follows K m =2.62, V max =4.33 × 10 -8 M·s -1 The slope of the fitted curve under light conditions. K m / V max =0.33078, intercept 1 / V max =0.22646, calculated as follows K m =1.46, V max =4.41 × 10 -8 M·s -1 .
[0048] in, K m This represents the Michaelis constant in steady-state dynamics, and its magnitude is related to the affinity of the nanozyme for the substrate; the smaller the value, the greater the affinity. V max This indicates the maximum reaction rate.
[0049] In summary, under illumination, nZVS / nZVC exhibited enhanced maximum reaction rate and substrate affinity.
[0050] III. Detection of the effect of photoenzyme synergistic catalysis in removing Microcystis aeruginosa 1. Effect of light on the removal of Microcystis aeruginosa by nZVS / nZVC The effect of illumination on the removal of *Microcystis aeruginosa* by nZVS / nZVC in Example 1 was investigated. Specifically, the LED light source power gradient was set to 0 W, 63 W, 96 W, and 126 W. The chlorophyll a concentration of *Microcystis aeruginosa* was measured using a phytoplankton fluorometer, where C0 is the initial chlorophyll a concentration and C is the instantaneous concentration at a certain time t. The results are shown below. Figure 6 .
[0051] Depend on Figure 6 It can be seen that the algae removal efficiency is relatively low under dark conditions; however, the algae removal effect is significantly improved after the introduction of LED light, and the rate of decrease of C / C0 gradually accelerates with the increase of LED power.
[0052] 2. Effects of different silver-copper ratios (nZVS / nZVC) on the removal of Microcystis aeruginosa The effects of nanozymes in Examples 1-3 and Comparative Examples 1-2 on the removal of Microcystis aeruginosa were detected. The specific detection method was as follows: C / C0 detection was performed. Results are shown below. Figure 7 .
[0053] Depend on Figure 7 It can be seen that the algae removal efficiency of the single material is significantly weaker than that of the composite material. The removal efficiency of *Microcystis aeruginosa* by the nZVS / nZVC composite material decreases rapidly with reaction time, and the algae removal rate gradually increases with the increase of the proportion of nZVC in the composite material. This result indicates that the combination of nZVS and nZVC effectively enhances the algae removal performance.
[0054] III. Comparative Analysis of Photocatalysis, Enzyme Catalysis, and Photoenzyme Co-catalysis The algae removal effects of Example 1 and Comparative Examples 3-4 were tested, and the results are shown below. Figure 8 .
[0055] Depend on Figure 8 It can be seen that, at the same reaction time, the algae removal capacity of photocatalysis and enzyme catalysis is significantly better than that of photocatalysis and enzyme catalysis. At 20 min, the algae removal capacity of photocatalysis and enzyme catalysis is 1.35 times that of enzyme catalysis and 11 times that of photocatalysis.
[0056] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.
Claims
1. A method for the photoenzymatic synergistic catalytic removal of Microcystis aeruginosa based on nanozymes, characterized in that, Includes the following steps: Nanoenzymes were added to a suspension of Microcystis aeruginosa, followed by the addition of peroxide, and algae were removed under light conditions. The nanozyme was prepared by the following method: (1) Dissolve silver nitrate and copper nitrate trihydrate in water to prepare solution A; (2) Dissolve sodium borohydride in water to prepare solution B; (3) Add solution B to solution A, wash and vacuum dry to obtain nZVS / nZVC, i.e. nanozyme.
2. The method for removing Microcystis aeruginosa based on nanozymes according to claim 1, characterized in that, The volume-to-mass ratio of Microcystis aeruginosa suspension to nanozyme is 1L:100-700mg.
3. The method for removing Microcystis aeruginosa based on nanozymes according to claim 1, characterized in that, The peroxide is potassium persulfate.
4. The method for removing Microcystis aeruginosa based on nanozymes according to claim 1, characterized in that, The lighting conditions are LED light source, tungsten lamp, xenon lamp or fixed wavelength light source.
5. The method for photoenzymatic synergistic catalysis of Microcystis aeruginosa based on nanozymes according to claim 1, characterized in that, Algae removal is performed under magnetic stirring conditions.
6. The method for removing Microcystis aeruginosa based on nanozymes according to claim 1, characterized in that, In step (1), the molar volume ratio of silver nitrate, copper nitrate trihydrate and water is 0.2-0.8 mmol: 0.2-0.8 mmol: 30 mL.
7. The method for removing Microcystis aeruginosa based on nanozymes according to claim 1, characterized in that, In step (2), the mass-to-volume ratio of sodium borohydride and water is 100 mg: 10 mL.
8. The method for removing Microcystis aeruginosa based on nanozymes according to claim 1, characterized in that, In step (3), under stirring and nitrogen atmosphere conditions, solution B is added dropwise to solution A.
9. The method for removing Microcystis aeruginosa based on nanozymes according to claim 1, characterized in that, In step (3), the volume ratio of solution B to solution A is 1:
3.
10. The application of the method for removing Microcystis aeruginosa based on nanozymes according to any one of claims 1-9 in algae removal.