Method for rapidly forming algal biofilms

Abstract

To provide a method for rapidly forming algal biofilms.SOLUTION: The method of the present invention includes mixing activated sludge and filtered wastewater, heating, stirring, standing or centrifuging, collecting activated sludge extracellular extract, applying it to the surface of a carrier, evaporating the water, applying the algae cell mixture to the carrier surface carrying the activated sludge extracellular extract, repeating the application-evaporation operation of the activated sludge extracellular extract several times to obtain a carrier carrying the algae cell mixture, and immersing the carrier carrying the algae cell mixture in the filtered wastewater for biofilm culture. The method can shorten the film formation period to within 2 weeks, saving the biofilm initiation time of the algae biofilm wastewater treatment process. The algae biofilm formed in only one week can achieve the treated wastewater of the first grade A standard specified in GB 18918-2002, reducing the use of chemical reagents and special apparatus, reducing costs and technical thresholds, and being universal and having a very good promotion effect on different carrier materials, which is conducive to rapid popularization.SELECTED DRAWING: Figure 4

Description

[Technical field]

[0001] The present invention relates to the technical field of wastewater biological treatment, in particular to the rapid growth of algal biofilms. It relates to membrane methods. [Background technology]

[0002] Algal biofilms are biofilms dominated by algae, including eukaryotic microalgae and cyanobacteria. It is a film community that grows on surfaces with a certain amount of light and humidity. The wastewater treatment technology using aluminum has high nitrogen and phosphorus utilization efficiency, low carbon dependency, and low pollutant due to their resistance to toxic substances, their ability to fix carbon dioxide, and their ability to produce valuable biomass products. Widely attracting attention, under the background of "carbon neutrality", the low-carbonization and development of the sewage treatment industry The formation of algal biofilms involves the reversible attachment of microorganisms. The stages of adhesion and colonization, irreversible attachment and aggregation, biofilm development, and mature spreading are This process can take several weeks or even months. Excessively long biofilm initiation times can increase the initial costs of engineering applications and make algal biofilms more difficult to grow. Therefore, the fast growth of algal biofilms is not suitable for the rapid diffusion of algae in the algae growth process. There is an urgent need to develop a method. For other biofilm rapid biofilm formation methods, see CN112305029 discloses a method for rapid formation of electrode biofilms. The lysine and adhesive are mixed to form a composite formulation, which is then spin-coated onto the collector to form the electrode bilayer. CN114524508 rapidly forms a membrane in the biofilter. A reinforcing agent and method for promoting biological biofilms is disclosed. The solution comprises: High concentration of L-cysteine ​​and complementary amino acids (phenylalanine, proline, methionine) Adding one or more of these compounds promotes rapid film formation of the filler. No. 3,998,776 discloses a method for modifying fillers to promote rapid biofilm formation. In this method, the filler surface is treated with an acidic oxidizing agent, and then a hydrophilic dopant is applied. By introducing amine and oxidizing and self-polymerizing it on the surface of the filler layer, the hydrophilicity of the filler surface is improved. It changes the adhesiveness and adhesion of the material, and promotes rapid film formation. The present invention discloses a method for rapid biofilm formation for a biofilm reactor. The first step is to inoculate the wastewater with aerobic activated sludge, then add organic carbon and humus to stabilize the wastewater. Anaerobic activated sludge is inoculated into the reactor, and sugar alcohol and humus are added to promote membrane formation. do. There are few technical solutions for the rapid initiation of algal biofilms. CN10959365 7 discloses a rapid inoculation method for microalgae biofilms based on pH adjustment. The steps of the method include selecting a supplementary algae species, selecting an algae species to be inoculated and activating the supplementary algae species. The algae-algae copolymer suspension is mixed with the algae and pre-cultured, and the pH is adjusted to make it acidic and and obtaining an alkaline algae-algae copolymer suspension, mixing them, and adding the microorganisms to be inoculated. This involves inoculating a microalgal biofilm culture system to achieve rapid inoculation. The method has the following problems: (1) it is targeted at a dual pure algal culture system; Not applicable to the mixed algae-bacteria symbiosis in real wastewater treatment scenarios, (2)p It requires adjustment of H and measurement of zeta potential, and consumes a large amount of acid and alkali reagents, which makes the process tedious. (3) It is a costly process because it requires a certain hardware threshold. The final result is rapid sedimentation of the algae-algae mixture onto the carrier, resulting in the formation of a large number of algae such as microalgae. It is not guaranteed that the microorganisms in question will be able to stably attach to the growth surface. Methods have been developed for rapid initiation of films, but they involve the addition of organic agents or strong oxidizing agents. These are expensive, prone to cross-contamination, and difficult to understand microbial composition, growth conditions, and Because the application scenarios are different, these methods can be directly applied to the rapid formation of algal biofilms. There is a common problem that cannot be solved. Summary of the Invention

[0003] The method for high-speed formation of an algal biofilm of the present invention comprises the steps of: S1. Prepare activated sludge, algae mixture and wastewater, filter the wastewater, and collect the filtered wastewater. and S2: Activated sludge and filtered wastewater are mixed in a ratio of 1g:20-50mL and heated to 60-80℃. After heating, keep warm and stir for 10 to 60 minutes, then leave to stand or centrifuge. collecting the supernatant to obtain the activated sludge extracellular extract; S3, activated sludge extracellular extract at 100-200 mL / m 2 and applying the coating solution to the carrier surface in an amount of A step of naturally evaporating for 3 to 8 minutes to obtain a carrier carrying an activated sludge extracellular extract; S4: Algae mixture at 0.1-100g / m 2 The activated sludge extracellular extract was supported at a coating amount of The mixture of algae and fungi is then applied to the surface of the carrier. Then, step S3 is repeated several times to support the mixture of algae and fungi. obtaining a support; S5: The carrier carrying the algae-bacteria mixture is immersed in the filtered wastewater to culture the biofilm. After the biofilm was cultured for 3-4 days and a thin green mucus layer formed, the wastewater was replaced once a day. and continuing the biofilm culture. In one aspect of the present invention, in step S1, The activated sludge is obtained from the aerobic part of a biological tank in a municipal sewage treatment plant. Sludge is a common product in this field, is easily available, has low cost, The algae-bacteria mixture is scraped off the light-exposed surface of the biological tank wall or the secondary sedimentation tank wall of a municipal wastewater treatment plant. The microbial community is a mixture of algae and bacteria, and the mixture of algae and bacteria is a microbial community that is a mixture of algae and bacteria. It is a universal product and any municipal wastewater treatment plant, regardless of the process employed, will use bioreactors and and a secondary settling tank, wastewater treatment is a continuous process, therefore, The biomass from the walls of the bioreactor or secondary sedimentation tank at the site was relatively stable and did not change over time. , easy to obtain, low cost, The wastewater is urban sewage, which is one of the common wastewaters in this field; The filtration is performed using a 100 mesh nylon cloth to remove large particulate matter. In one aspect of the present invention, in step S2, the standing time is 15 to 60 minutes. The centrifugation speed is 5000 to 7500 rpm, and the centrifugation time is 5 to 10 min. in. In one aspect of the present invention, in step S3, the carrier is glass, silica gel filter, or the like. The material may be rubber, fabric or flexible plastic. In one aspect of the present invention, in step S4, the number of times is 2 to 5 times. In one aspect of the present invention, in step S5, the biofilm culture is grown under natural light or sunlight. It must be done under fluorescent light. In one aspect of the present invention, in step S5, the total time for biofilm culture is 2 weeks. is less than. Effect of the Invention

[0004] Compared with the prior art, the present invention has the following significant advantages: (1) The method of the present invention successfully shortens the film formation cycle of algal biofilms to within two weeks. This has significantly reduced the biofilm initiation time of the algae biofilm wastewater treatment process. It is possible. (2) The method of the present invention requires only one week to form an algal biofilm. The discharge standard for pollutants from urban wastewater treatment plants (G B 18918-2002) is sufficient to achieve the Grade 1A standard. (3) The method of the present invention uses fewer chemical reagents and specific Effectively reduce the use of special equipment, reduce the cost and technical threshold required for the method, and achieve rapid It is conducive to popularization, universally applicable, and has good promotion effect on different carrier materials. [Brief description of the drawings]

[0005] [Figure 1] 1A-1D are cross-sectional views of an algal biofilm of Example 1 of the present invention at different times. [Diagram 2] FIG. 11 is a comparison diagram of the film formation effect between an experimental group and a control group in Example 4 of the present invention. [Diagram 3] Photographs showing changes during the biofilm culture process in a reactor in Example 5 of the present invention. [Figure 4] FIG. 1 shows the promoting effect of different treatment methods examined in the test examples of the present invention on the rapid formation of algal biofilms on four material surfaces (cotton cloth, glass, silica gel, and flexible plastic). [Diagram 5]FIG. 1 shows the wastewater treatment effectiveness of membranes on which algal biofilms were formed after two weeks on four material surfaces (cotton cloth, glass, silica gel, and flexible plastic) using different treatment methods examined in the test examples of the present invention. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0006] The technical solutions of the present invention are further described below in conjunction with the accompanying drawings. Example 1 In this example, glass was used as a carrier, and the algae biofilm profile rapid film formation process Observe. S1: Activated sludge freshly discharged from the aerobic section of a municipal wastewater treatment plant was collected and analyzed. The algae mixture was scraped off the wall of the secondary sedimentation tank with a scraper, and at the same time, a water sample after the grid was taken. The wastewater is collected and filtered through a 100 mesh nylon cloth, then collected in a container. After filtering, the wastewater is obtained. S2. Measure the sludge concentration (MLSS) of activated sludge by weighing method and calculate the concentration per 1g (dry weight) of activated sludge. Add 25mL of filtered wastewater to each container, place in a magnetic stirrer, and heat to 60℃ for 30min. The mixture was centrifuged at 5000 rpm for 5 minutes, and the supernatant was collected and used for analysis. Obtaining an extracellular extract of the sludge; S3, Place the glass in a culture dish and infuse the activated sludge extracellular extract at 125 mL / m 2 Glass with a coating amount of The slides containing the activated sludge extracellular extract were then applied to the slides and allowed to evaporate for 3 min. Gain, S4, 75g / m of algal cell mixture 2 The glass surface containing activated sludge extracellular extract was coated with The application-evaporation step of step S3 is repeated three times to make the algae mixture into a smooth mixture. The results showed that the algae-bacteria mixture was strongly attached to the slide surface. In the first iteration of step S3 and in the last iteration of step S3, the evaporation time is The incubation time was changed to 5 min to obtain a slide containing the algae-bacteria mixture. S5. Place the culture dish containing the slide containing the algae-fungal mixture in an artificial climate incubator. , light intensity 260 μmol / s m 2 The light:dark cycle was 12h:12h, and the soiling after filtration was The specimens were slowly immersed in water (filtered wastewater was used as the culture medium) to culture the biofilm. After 3 days, The culture medium was changed for the first time, the algal biofilm profile was recorded, and then cultured once a day. The liquid was replaced and the algal biofilm profile was recorded on the 6th and 14th days. The film profile is shown in FIG. FIG. 1 is a cross-sectional view of an algal biofilm in Example 1 of the present invention at different times. As shown, when biofilms were cultured using the method of the present invention, the cells were already visible to the naked eye on the third day. A thin layer of visible algal biofilm was present, and after 2 weeks, a dense layer of algae more than 2 mm thick appeared. The formation of biofilms has been reported in previous experiments and in the literature. Compared with the time required for deposition (one to several months), the deposition time of this method is significantly shortened.

[0007] Example 2 In this example, glass was used as a carrier, and the algae biofilm profile rapid film formation process Observe. S1: Activated sludge freshly discharged from the aerobic section of a municipal wastewater treatment plant was collected and The algae mixture was scraped off the wall of the secondary sedimentation tank with a scraper, and a water sample after the grid was simultaneously taken. The wastewater is collected and filtered through a 100 mesh nylon cloth before being collected in a container. After filtering, the wastewater is obtained. S2. The sludge concentration (MLSS) of activated sludge was measured by the weighing method. Add 20 mL of filtered wastewater to each flask, place in a magnetic stirrer, heat to 60°C, and maintain for 10 min. The mixture was centrifuged at 6000 rpm for 6 minutes, and the supernatant was collected and analyzed for activity. Obtaining a sludge extracellular extract; S3, Place the glass in a culture dish and infuse the activated sludge extracellular extract at 100 mL / m 2 Glass with a coating amount of The slides containing the activated sludge extracellular extracts were then applied to the surfaces of the slides and allowed to evaporate for 5 min. Gain, S4, 0.1g / m of algal cell mixture 2 The activated sludge extracellular extract was applied to a glass substrate at a coating amount of The application-evaporation step of step S3 is repeated five times to obtain an algae-bacteria mixture. The algae-fungus mixture was observed to be firmly attached to the slide surface, and a slide containing the algae-fungus mixture was obtained. S5. Place the culture dish containing the slide containing the algae-fungal mixture in an artificial climate incubator. , light intensity 260 μmol / s m 2 The light:dark cycle was 12h:12h, and the soiling after filtration was The specimens were slowly immersed in water (filtered wastewater was used as the culture medium) to culture the biofilm for 3 days. The culture medium was changed for the first time after that, and then changed once a day for a total of 13 days.

[0008] Example 3 In this example, glass was used as a carrier, and the algae biofilm profile rapid film formation process Observe. S1: Activated sludge freshly discharged from the aerobic section of a municipal wastewater treatment plant was collected and The algae mixture was scraped off the wall of the secondary sedimentation tank with a scraper, and the water sample after the grid was simultaneously collected. The wastewater is collected and filtered through a 100 mesh nylon cloth before being collected in a container. After filtering, the wastewater is obtained. S2. Measure the sludge concentration (MLSS) of activated sludge by weighing method and calculate the concentration per 1g (dry weight) of activated sludge. Add 50L of filtered wastewater to the tank, place in a magnetic stirrer, heat to 80℃ and maintain for 60min. The mixture was then centrifuged at 7500 rpm for 10 minutes, and the supernatant was collected and used for analysis. Obtaining an extracellular extract of the sludge; S3, Place the glass in a culture dish and infuse the activated sludge extracellular extract at 200 mL / m 2 Glass with a coating amount of The slides containing the activated sludge extracellular extract were then applied to the slides and allowed to evaporate for 8 min. Gain, S4: 100g / m of algae mixture 2 The activated sludge extracellular extract was applied to a glass substrate at a coating amount of The application-evaporation step of step S3 is repeated twice to obtain an algae-bacteria mixture. It was observed that the algae-microbe mixture was firmly attached to the slide surface. The first iteration of S3 and the last iteration of step S3 had an evaporation time of 5 min. to obtain a slide containing a mixture of algae and fungi. S5. Place the culture dish containing the slide containing the algae-fungal mixture in an artificial climate incubator. , light intensity 260 μmol / s m 2 The light:dark cycle was 12h:12h, and the soiling after filtration was The specimens were slowly immersed in water (filtered wastewater was used as the culture medium) to culture the biofilm for 4 days. The culture medium was changed for the first time after 1 hour, and thereafter the culture medium was changed once a day for a total of 14 days of culture.

[0009] Example 4 In this embodiment, the step of applying the activated sludge extract is performed by treating the activated sludge with the extract and treating the activated sludge with the extract. The surface film formation rates of untreated cotton fabrics were compared. S1, the sample collection step is the same as step S1 in Example 1; S2: Add 50 mL of filtered wastewater per 1 g of activated sludge (dry weight) and place in a magnetic stirrer. Heat to 60°C and maintain for 60 min, then centrifuge at 4000 g for 10 min. Centrifuge and collect the supernatant to obtain the activated sludge extracellular extract. S3, Material surface treatment Experimental group: The activated sludge extracellular extract was evenly applied to the surface of cotton fabric. The experimental process was the same as in Example 1. It is the same as step S3, Control group: The sewage water after filtration was evenly applied to the surface of the cotton fabric. The experimental process was the same as in Example 1. Same as S3, but replacing the activated sludge extracellular extract with sewage treatment plant post-filtration effluent. Steps S4 and S5 correspond to steps S4 and S5 in Example 1, respectively. The experimental procedure was the same as in 5, but the ratio of the amount of algae mixture applied to the surface area of ​​the cotton fabric was 0.5 g / m 2 was reduced to. The loading conditions on the carrier surface of each experimental group and control group were recorded, as shown in Figure 2. On the second day, the film formation conditions of the experimental group and the control group were compared again. As can be seen from FIG. 2, the membrane formation effect of the activated sludge extracellular extract was The experimental group treated with activated sludge extracellular extract showed more microbial activity than the control group not treated with activated sludge extracellular extract. Many of the initial algal-bacterial symbionts were immobilized on the support surface, and after 3 days a dense algal biofilm was formed. is formed, which indicates that the step of applying the activated sludge extracellular extract proposed in the present method is It is the key to promote fast biofilm formation.

[0010] Example 5 In this embodiment, a transfer device is used to realize automation of the coating-evaporation steps, and at the same time, the method It was used in an actual algal biofilm reactor to verify its effectiveness in wastewater treatment. S1, sample collection is the same as step S1 in Example 1; S2: Add 20 mL of filtered wastewater per 1 g (dry weight) of activated sludge and place in a magnetic stirrer. Heat to 80°C and maintain for 60 min, then centrifuge at 6000 g for 10 min. Centrifuge and collect the supernatant to obtain the activated sludge extracellular extract. S3. The silica gel film is placed on the double rotating shaft of the transmission device, like a vertical conveyor belt. A reactor is formed, a pool is formed at the bottom end, and the activated sludge extracellular extract is poured into the pool. When the Rikagel film is rotated to the bottom, it is just touching the activated sludge extracellular extract. The rotation speed of the silica gel film was controlled so that one turn period was 5 minutes. This allows the application and evaporation of activated sludge extracellular extract to be automated, and the activated sludge extracellular extract Obtain a silica gel film, S4: After rotating the silica gel film once, the algae mixture and the coating surface area ratio of 10 0g / m 2 The algae mixture was then applied to the silica gel film containing the activated sludge extracellular extract. The sludge extracellular extract was applied to the surface. The rotation speed was adjusted to 1 turn per 7 minutes, and the application-evaporation step was performed. The slide was automatically cycled five times and it was observed that the algae-fungus mixture was firmly attached to the slide surface. As a result, a silica gel film containing the algae-bacteria mixture was obtained, S5, as shown in Figure 3, the reactor with the silica gel film containing the algae-bacteria mixture was exposed to sunlight. The pool was filled with filtered wastewater and the pool was then cooled for the first three days. The water in the tank was not exchanged, and after that, the water retention time was set at 24 hours every day, and then the wastewater was exchanged. Film culture was performed, and the results were compared before and after application of the algae-bacteria mixture, and after application of the algae-bacteria mixture. The state of biofilm on the carrier surface on the 7th day was recorded. FIG. 3 is a photograph showing the change in the reactor biofilm culture process in Example 5 of the present invention; As can be seen from FIG. 3, after the algae-bacteria mixture was applied to the carrier surface, a large amount of initial algae was inoculated. The bacterial symbionts attach and provide the basis for the subsequent rapid expansion of the algal biofilm. Based on the enhanced community attachment, after 7 days, the algal biofilm grew over the entire surface of the carrier, and the biofilm 32.6g / m 2 reached. The chemical acidity of the water was then measured to evaluate the effectiveness of the formed algal biofilm in treating the wastewater. Calculates the values ​​of COD, total nitrogen (TN) and total phosphorus (TP) and measures the chemical oxygen demand. COD, total nitrogen (TN) and total phosphorus (TP) are the pollutants discharged from municipal wastewater treatment plants. Detected according to the method specified in Table 7 of the GB 18918-2002 standard. The quality of the wastewater was measured and found to be COD 21.4±5.9mg / L and TN 5.2±2. 3mg / L, and TP 0.23±0.19mg / L. These values ​​were compared with those of the urban wastewater treatment plant. Within the Class A standard specified in the Pollutant Discharge Standard (GB 18918-2002) This means that the fast-starting algal biofilm of the present method has a very high wastewater treatment effect. Furthermore, this example demonstrates that the method used in the present invention is based on a rational device design. It can be seen that this can be automated and has a wide range of potential.

[0011] Test Example 1 The method of Example 1 of the present invention and the pH adjustment method of Patent CN109593657 (inoculation z (Adjust the pH of the inoculation solution until the eta potential is 0) Acid method (L-cysteine ​​2.0 g / L and phenylalanine, proline, and methionine Add 0.4g / L of each amino acid), oxidation + dopamine method of patent CN113998776 (a method of treating the filler surface with an acidic oxidizing agent and then introducing hydrophilic dopamine), and A blank control set without any treatment was used for the study. The oxidation + dopamine method was performed using cotton and silicone. Since this test method cannot be applied to gel materials, the experimental results using these two materials are No results. According to the results in Figure 4, this method is more effective in promoting rapid algal biofilm formation than other existing methods. significantly higher than Test Example 2 This study mainly examined the effect of different treatment methods on the growth of algal biofilms on four material surfaces (cotton, Wastewater treatment efficiency of membranes formed on glass, silica gel and flexible plastic for two weeks. The results of the test are shown in Figure 5. The algae biofilm formed by the method of Example 1 of the invention showed very good sewage treatment efficiency in 2 weeks. The results were achieved, and the concentrations of COD, TN, and TP in the wastewater were significantly lower than those of other methods. This shows that the algae biofilm rapidly formed by this method is also very effective in wastewater treatment. It is clear that this is beneficial.

Claims

1. Step S1: Prepare activated sludge, an algae-cell mixture, and wastewater, filter the wastewater, and then filter the sludge. collecting water; Step S2: Mix activated sludge and filtered wastewater in a ratio of 1 g:20-50 mL, and add 60-80 mL of activated sludge to the filtered wastewater. After heating to 0°C, the mixture is stirred while keeping the temperature constant for 10 to 60 minutes. The mixture is then left to stand or centrifuged. separating and collecting the supernatant to obtain an activated sludge extracellular extract; Step S3: 100 to 200 mL / m of activated sludge extracellular extract ２ On the carrier surface with a coating amount of The extract is applied to the carrier and allowed to evaporate for 3 to 8 minutes to obtain the carrier carrying the extracellular extract of activated sludge. Top and Step S4: Add the algae mixture to a volume of 0.1 to 100 g / m ２ Activated sludge extracellular extraction at application amount of The mixture of algae and fungi is applied to the surface of the carrier carrying the substance, and then step S3 is repeated several times to form the mixture of algae and fungi. Obtaining a support carrying the Step S5: The carrier carrying the algae-bacteria mixture is immersed in the filtered wastewater to form a biofilter. The cells were cultured in a 3-4 day biofilm culture, and after a thin green mucus layer formed, the cells were stained once a day. and continuing the biofilm culture by replacing the water. A method for rapid biofilm formation.

2. In step S1, the activated sludge is obtained from the aerobic section of a biological tank of a municipal wastewater treatment plant. The activated sludge is a mixture of algae and bacteria, and the mixture is applied to the walls of a biological tank or a secondary sedimentation tank of a municipal wastewater treatment plant. The microbial community is a mixture of algae and bacteria scraped off from the light-exposed surface, and the wastewater is from an urban area. The sewage is filtered using a 100 mesh nylon cloth. The method according to claim 1 .

3. In step S2, the standing time is 15 to 60 minutes, and the centrifugation speed is The centrifugation time is 5 to 10 minutes. The method of claim 1,

4. In step S3, the carrier is glass, a silica gel film, a fabric or a flexible plastic.

2. The method of claim 1, wherein the substrate is a plastic.

5. 2. The method according to claim 1, wherein in step S4, the number of times is 2 to 5. method.

6. In step S5, the biofilm culture needs to be performed under natural or artificial light. The light intensity of the artificial light is 260 μmol / s m ２ 4. The method according to claim 3, 2. The method according to claim 1.

7. In step S5, the total time of the biofilm culture is less than two weeks. The method according to claim 1, wherein

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