Method for preparing recoverable 3D microalgae gel and removing antibiotics in water by using recoverable 3D microalgae gel

Abstract

The invention discloses a method for preparing recoverable 3D microalgae gel and removing antibiotics in water, based on a 3D gel packaging technology, an isolation semipermeable membrane is constructed on the surface of a gel matrix, and a recoverable microalgae solid purification system is developed to cooperate with a mild environment provided by the gel matrix and the isolation effect of the semipermeable membrane. While the activity of the microalgae is protected and efficient biodegradation is realized, the leakage of the microalgae is prevented, and the potential microbial biohazard is restrained. Meanwhile, through the 3D printing technology, recoverable, low-cost, rapid and large-area industrial manufacturing can be achieved, the defects existing in traditional microalgae water treatment are overcome, and wide application prospects are achieved.

Description

Technical Field

[0002] The invention relates to the application of algae composite materials in the field of wastewater treatment, and in particular to a method for preparing a recyclable 3D microalgae gel and removing antibiotics from water. Background Art

[0003] The emergence of antibiotics has brought great convenience to the treatment of bacterial infections in humans and animals. However, the influx of a large amount of untreated antibiotics into natural waters will damage the ecosystem and aggravate the crisis of bacterial resistance to antibiotics, resulting in about 1.2 million deaths from antibiotic-resistant bacterial infections each year. Therefore, antibiotic resistance has been identified by the World Health Organization (WHO) as one of the most serious challenges facing human health. Although various methods for removing antibiotics, including advanced oxidation, adsorption, and ultrafiltration, have shown good efficiency, these methods are complex to prepare, costly, and may produce harmful byproducts. In contrast, bioremediation uses microorganisms to degrade antibiotics, providing an environmentally friendly technology for solving the problem of antibiotic pollution. In particular, photosynthetic algae is a low-carbon bioremediation candidate technology that can accumulate valuable biomass and produce oxygen while processing antibiotics, while achieving biological remediation and carbon sequestration of wastewater. For example, a method for using microalgae to degrade cefotaxime in water is disclosed in the public invention patent CN 114561293 A, but its degradation efficiency is low, and the activity of microalgae will be inhibited at high concentrations of antibiotics. At the same time, typical microalgae-mediated remediation technologies require the construction of large treatment pools or ponds, which require a large area and water volume. In addition, suspended microalgae in sewage are usually difficult to recycle, and the flow of these microorganisms (especially engineered species) into natural water bodies may affect the distribution of local microbial communities (such as causing algal blooms).

[0004] Cell-living materials provide an isolated environment by embedding cells in a matrix to protect them from the environment while allowing material exchange to ensure cell growth. Currently, most hydrogel matrices (such as sodium alginate) encapsulate cells in the form of gel particles. These systems have weak mechanical properties, limited diffusion of nutrients, and easy leaching of microorganisms, which limits their application. Summary of the invention

[0005] In order to solve the problems of difficult recycling of traditional microalgae treatment, poor biological activity of microalgae at high concentration, low degradation efficiency, etc., and to overcome the problems of weak mechanical properties of traditional packaging materials, easy leakage of microorganisms leading to algal bloom pollution, and difficulty in industrial manufacturing, the present invention provides a method for preparing a recyclable 3D microalgae gel network and removing antibiotics from water, which mainly includes the following steps:

[0006] 1. Preparation of microalgae bio-ink: dissolve polyethylene glycol diacrylate in deionized water, then add photoinitiator LAP, add sodium alginate after complete dissolution, and finally mix the concentrated algae solution with the system to obtain bio-ink;

[0007] 2. 3D printing of microalgae bio-ink: transfer the bio-ink into the printing barrel and load it into the 3D printer. Use the printer's software to build the model structure, and adjust the diameter of the monofilament by switching the needle. Perform light-curing 3D printing according to the set parameters. After the model is printed, it is further cured by UV light irradiation and calcium chloride cross-linking.

[0008] 3. Constructing an isolation coating: Immerse the printed and solidified microalgae gel into a tannic acid solution to construct an isolation coating. The coating can be applied multiple times to ensure uniform coating.

[0009] Furthermore, the addition ratio of the polyethylene glycol diacrylate in step 1 is 4wt% to 8wt%;

[0010] Furthermore, the addition ratio of the photoinitiator LAP in step 1 is 0.2wt% to 0.6wt%;

[0011] Furthermore, the sodium alginate added in step 1 is 5wt% to 7wt%;

[0012] Furthermore, the microalgae in step 1 is Chlorella, Chlamydomonas reinhardtii, Spirulina or diatom;

[0013] Furthermore, the microalgae cell density OD 680 0.5~2.0;

[0014] Furthermore, the amount of algae liquid added in step 1 is 50 to 300 mL.

[0015] Further, the printing parameters described in step 2 are: layer height: 0.25-0.4mm; number of layers: 6-12 layers; random printing filling, printing outline, and starting point; model: quadrilateral; speed: 10-20mm / s; broken wire lifting: 0.4-0.8mm; air pressure: 0.2-0.4MPa; early wire exit: 200-400ms; early wire closing: 0.1-0.4mm; light curing power: 0.2%-3%; filling mode: linear filling; filling method: linear filling; filling offset: 0.7mm; filling spacing: 2mm; rotation angle: 90°; number of rotations: 2 times;

[0016] Furthermore, the UV curing time in step 2 is 2 to 10 minutes, and the ion cross-linking time is 5 to 20 minutes; the calcium chloride concentration in step 2 is 2 wt% to 6 wt%.

[0017] Furthermore, the concentration of tannic acid in step 3 is 5 to 15 mg / mL;

[0018] Furthermore, the coating time in step three is 1 to 10 minutes.

[0019] The present invention also provides a method for removing antibiotics from wastewater using a recyclable 3D microalgae gel membrane, the specific steps of which are:

[0020] The printed microalgae gel film is placed in the antibiotic wastewater to be treated, and the antibiotic wastewater is purified after 1 to 5 days of degradation. After treatment, the microalgae gel film can be directly taken out for recycling or enter the next wastewater treatment cycle.

[0021] Further, the antibiotic wastewater includes one or more of tetracyclines, macrolides, and sulfonamides;

[0022] Furthermore, the antibiotic wastewater concentration is 0 to 500 mg / L;

[0023] Furthermore, the degradation conditions include a temperature of 22 to 25°C, a humidity of 30 to 70%, and a light intensity of 45 to 55 μmolphotons m -2 s -1 .

[0024] The technical advantages of the present invention are:

[0025] 1. The present invention realizes recyclable antibiotic wastewater treatment by constructing a foldable microalgae gel membrane;

[0026] 2. Compared with the widely used suspended microalgae, the solid microalgae purification system has a relatively high tolerance to antibiotics and a stronger degradation ability;

[0027] 3. The constructed semi-permeable membrane isolation layer can prevent the leakage of microalgae and avoid potential microbial biohazards;

[0028] 4. Microalgae gel is biodegradable and can be used as a biofertilizer without generating solid waste;

[0029] 5. Microalgae gel membrane can be mass-produced on a large scale using 3D printing technology and has the potential for industrial application. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 is a rheological curve diagram of the microalgae bio-ink of Example 1;

[0031] Figure 2 is the tensile stress-strain curve of the gel of Example 1;

[0032] Figure 3is a microalgae leaching diagram of the microalgae gel (denoted as MHN) in Example 1 before and after being coated with a tannic acid isolation layer;

[0033] Figure 4 is a comparison diagram of the effects of suspended microalgae (denoted as SM), MHN and MHN@TA in treating tetracycline at different concentrations in Example 2, and a diagram of the removal efficiency of tetracycline by MHN@TA at different times; C e represents the antibiotic concentration in the solution after degradation, and C0 represents the initial antibiotic concentration in the solution;

[0034] Figure 5 It is a graph of the degradation efficiency of antibiotics under different treatments;

[0035] Figure 6 This is a graph showing the efficiency of MHN@TA cyclic degradation of antibiotics in Example 3; Specific embodiments

[0036] Embodiment 1:

[0037] A method for preparing a recyclable 3D microalgae gel network, the specific implementation method is as follows:

[0038] 1. Preparation of microalgae bio-ink: 0.4 g of polyethylene glycol diacrylate was dissolved in 5 mL of deionized water, and then 10 mg of photoinitiator LAP was added. After the solution was completely dissolved, 0.3 g of sodium alginate was added. Finally, 200 mL of OD 680 The Chlorella algae liquid with a g / mL of 1.0 is centrifuged at 5000 g for 4 minutes, and then the precipitate is collected and mixed evenly with the system to obtain the bio-ink;

[0039] 2. 3D printing of microalgae bio-ink: The bio-ink prepared in step 1 is transferred into the printing cartridge and loaded into the 3D printer (Bio- SR, Regenovo, Hangzhou, China). The model structure was constructed using the printer software, and the diameter of the monofilament was adjusted by switching the needle. The light-cured 3D printing was performed according to the set parameters. After the model was printed, it was further cured by UV light irradiation and calcium chloride cross-linking. The specific printing parameters were: layer height: 0.3 mm; number of layers: 10 layers; random printing filling, printing contour, and starting point; model: quadrilateral; speed: 15 mm / s; broken wire elevation: 0.5 mm; air pressure: 0.3 MPa; early wire exit: 300 ms; early wire closing: 0.3 mm; light curing power: 1%; filling mode: linear filling; filling method: linear filling; filling bias: 0.7 mm; filling spacing: 2 mm; rotation angle: 90°; number of rotations: 2 times. The printed microalgae gel was cured under UV light for 3 min, and then further cross-linked in a calcium chloride solution with a concentration of 5 wt% for 10 min to obtain a molded microalgae gel, which was recorded as MHN.

[0040] 3. Construction of isolation coating: The printed and solidified microalgae gel was immersed in a tannic acid solution with a concentration of 10 mg / mL for 2 minutes to construct an isolation coating. The coating process was repeated twice to ensure uniform coating, which was recorded as MHN@TA.

[0041] Embodiment 2:

[0042] The printed microalgae gel film with a size of 40 cm × 50 cm × 0.5 cm was placed in 1000 mL of wastewater containing tetracycline at 23 °C, 50% humidity, and a light intensity of 50 μmol photons m -2 s -1 , the antibiotic wastewater was purified after 3 days of degradation.

[0043] Example 3: The microalgae gel after purification in Example 2 is taken out and placed in new antibiotic wastewater to enter the next cycle of degradation.

[0044] Comparative Example 1: Preparation method of PEGDA single network hydrogel

[0045] 0.4 g of polyethylene glycol diacrylate was dissolved in 5 mL of deionized water, and then 10 mg of photoinitiator LAP was added. The solution was poured into a mold and cured under ultraviolet light for 3 minutes to obtain a formed hydrogel, which was recorded as PEGDA.

[0046] Comparative Example 2: Preparation method of SA single network hydrogel

[0047] Preparation of sodium alginate single network hydrogel: 0.3 g of sodium alginate was dissolved in 5 mL of deionized water, the solution was poured into a mold, and cross-linked with a 5 wt % calcium chloride solution for 10 min to obtain a formed hydrogel, which was recorded as SA.

[0048] Comparative Example 3: Preparation method of 3D double network hydrogel network

[0049] The preparation method is the same as in Example 1, except that no microalgae is added, denoted as DNH and DNH@TA.

[0050] Comparative Example 4: Suspended microalgae treatment method

[0051] The exponentially growing Chlorella algae solution was collected and centrifuged at 5000 g for 4 min. The precipitate was then collected and the microalgae cells were resuspended in tetracycline wastewater and maintained at 23 °C, 50% humidity, and a light intensity of 50 μmol photons m - 2 s -1 After 3 days of degradation, the antibiotic wastewater was purified. It is recorded as SM.

[0052] Detection Example

[0053] Tetracycline detection: Take 1mL of the sample to be tested, filter it with a 0.22μm filter membrane, and then use a high performance liquid chromatograph to detect the residual antibiotic concentration. The detection conditions are: using a COSMOSIL C18 column (4.6×150mm, 5μm); the mobile phase is ultrapure water and acetonitrile-trifluoroacetic acid (0.1%, v / v); the mobile phase gradient is 85% to 20% (v / v); column temperature: 25℃; flow rate: 2mL / min; injection volume: 20μL; detection wavelength: 350nm.

[0054] Figure 1 It can be seen that the bio-ink has shear-thinning properties, and the addition of microalgae does not affect its rheological properties, indicating that the bio-ink has good printability;

[0055] Figure 2 It can be seen that the tensile strength of the double network hydrogel (denoted as DNH) is 134.3 kPa, which is higher than that of the single network hydrogel (polyethylene glycol diacrylate denoted as PEGDA, sodium alginate denoted as SA). At the same time, due to the hydrogen bond interaction between PEGDA and TA, the mechanical properties of the DNH@TA hydrogel coated with tannic acid (denoted as TA) are further improved, indicating that the microalgae gel film has good tensile properties and can be folded and bent;

[0056] Figure 3 It can be seen that after 7 days of immersion, the microalgae leakage rate of MHN without TA coating was 12.9%, while the leakage rate of MHN@TA was only 1.2%, showing excellent sealing performance;

[0057] Depend on Figure 4 a It can be seen that compared with SM, MHN and MHN@TA have higher degradation efficiency, especially in the environment of high antibiotic concentration; Figure 4 b It can be seen that after 72 hours of treatment, the degradation efficiency of MHN@TA for tetracycline at a concentration of 100 mg / L is as high as 99.3%, indicating that MHN@TA has excellent degradation performance and its degradation efficiency is not affected before and after coating.

[0058] Figure 5Water(dark) and Water(light) indicate that tetracycline wastewater without any treatment is naturally degraded under light and dark conditions; DNH@TA indicates that tetracycline wastewater is treated with hydrogel without adding microalgae (the preparation method of DNH@TA is the same as that of MHN@TA, the difference is that DNH@TA does not add microalgae); it can be seen that under light or dark conditions, the natural degradation efficiency of tetracycline in water is negligible, and the removal efficiency of DNH@TA hydrogel without microalgae for tetracycline is about 7.7%, indicating that the gel has little adsorption effect on tetracycline, while the removal efficiency of MHN@TA for tetracycline is as high as 96.3%, indicating that microalgae biodegradation is the main driving force for the removal of antibiotics.

[0059] The removal rate was calculated according to the formula: Removal rate (%) = (1-(Ce / C0)) x 100%, where Ce represents the concentration of antibiotics in the solution after degradation and C0 represents the initial concentration of antibiotics in the solution;

[0060] Figure 6 It can be seen that the microalgae gel film can be conveniently subjected to multiple antibiotic degradation treatments without a significant reduction in degradation efficiency. After five cycles of degradation, the degradation efficiency is still greater than 90%.

Claims

1. A method for preparing a recyclable 3D microalgae gel, comprising the following steps:

1. Preparation of microalgae bio-ink: dissolve polyethylene glycol diacrylate in deionized water, then add photoinitiator LAP, add sodium alginate after complete dissolution, and finally mix the concentrated microalgae solution with the system to obtain bio-ink; 2. 3D printing of microalgae bio-ink: transfer the bio-ink into the printing barrel and load it into the 3D printer. Use the printer's software to build the model structure, and adjust the diameter of the monofilament by switching the needle. Perform light-curing 3D printing according to the set parameters. After the model is printed, it is further cured by UV light irradiation and calcium chloride cross-linking.

3. Constructing an isolation coating: Immerse the printed and solidified microalgae gel into a tannic acid solution to construct an isolation coating.

2. The method for preparing a recyclable 3D microalgae gel according to claim 1, characterized in that: The addition ratio of the polyethylene glycol diacrylate in step 1 is 4wt% to 8wt%.

3. The method for preparing any one of the recyclable 3D microalgae gels according to any one of claims 1-2, characterized in that: The sodium alginate addition ratio in step 1 is 5wt% to 7wt%.

4. The method for preparing any one of the recyclable 3D microalgae gels according to any one of claims 1 to 3, characterized in that: The microalgae in step 1 is Chlorella, Chlamydomonas reinhardtii, Spirulina or diatom.

5. The method for preparing any one of the recyclable 3D microalgae gels according to any one of claims 1 to 4, characterized in that: The cell density of the microalgae in step 1 is OD 680 It is 0.5~2.0, and the amount of algae liquid added is 50~300mL.

6. The method for preparing any one of the recyclable 3D microalgae gels according to any one of claims 1 to 5, characterized in that: The printing parameters described in step 2 are: layer height: 0.25-0.4mm; Number of layers: 6-12; Printing filling, printing outline, random starting point; Model: quadrilateral; Speed: 10-20mm / s; Broken wire lifting: 0.4-0.8mm; Air pressure: 0.2-0.4MPa; Early wire drawing: 200-400ms; Early wire closing: 0.1-0.4mm; Light curing power: 0.2%-3%; Filling mode: linear filling; Filling method: linear filling; Filling offset: 0.7mm; Filling spacing: 2mm; Rotation angle: 90°; Number of rotations: 2 times.

7. The method for preparing any one of the recyclable 3D microalgae gels according to any one of claims 1 to 6, characterized in that: The UV curing time in step 2 is 2 to 10 minutes, and the ion cross-linking time is 5 to 20 minutes; the calcium chloride concentration in step 2 is 2 wt% to 6 wt%.

8. The method for preparing any one of the recyclable 3D microalgae gels according to any one of claims 1 to 7, characterized in that: The tannic acid concentration in step 3 is 5 to 15 mg / mL; the coating time is 1 to 10 min.

9. The method for removing antibiotics from wastewater using any one of the recyclable 3D microalgae gels according to claims 1-8, characterized in that: The recyclable 3D microalgae gel prepared according to any one of the methods described in claims 1-8 is placed in the antibiotic wastewater to be treated, and the antibiotic wastewater is purified after degradation for 1 to 5 days. After treatment, the microalgae gel film can be directly taken out for recycling or enter the next wastewater treatment cycle.

10. The method for removing antibiotics from wastewater using a recyclable 3D microalgae gel membrane according to claim 9, characterized in that: Antibiotic wastewater includes one or more of tetracyclines, macrolides and sulfonamides; the concentration of antibiotic wastewater is 0-500 mg / L; the degradation conditions include temperature of 22-25°C, humidity of 30-70%, and light of 45-55 μmolphotons m 2 s -1 .

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