CN108192889B - Method for treating wastewater by using bacterial cellulose immobilized microalgae - Google Patents
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Abstract
The invention relates to a method for treating wastewater by using bacterial cellulose immobilized microalgae, which comprises the following steps: after the microalgae is cultured at high density under proper illumination and temperature, the acetobacter xylinum culture medium is inoculated into the culture solution of the microalgae according to a certain proportion, cellulose is generated during the growth process of the acetobacter xylinum to capture the microalgae and is immobilized to form a cluster or a sphere, and the immobilized microalgae cluster can be used for treating various waste waters. The method introduces an immobilization technology of collecting microalgae by bacterial cellulose to treat wastewater, and immobilized microalgae agglomerates after wastewater treatment can be recycled, so that the requirement of wastewater industrial treatment application of microalgae is met; the invention is suitable for various wastewater treatments, has lower price and simple operation, is beneficial to wastewater treatment, and is a new way for economically and efficiently preparing microalgae wastewater treatment.
Description
Technical Field
The invention belongs to the technical field of environmental management, and relates to a method for treating wastewater by using bacterial cellulose immobilized microalgae.
Background
With the development of the social industrialization process, a large amount of industrial wastewater, aquaculture wastewater and domestic sewage are directly discharged into the natural environment without being effectively treated, and great threat is caused to the ecological environment. Compared with the traditional method, the method for treating the wastewater by using the microalgae can overcome the defects of secondary pollution, potential nutrient loss, incomplete utilization of resources and the like easily caused by the traditional wastewater treatment method, and can effectively remove nutrient substances such as nitrogen, phosphorus and the like causing water eutrophication at low cost, thereby having wide application prospect.
Compared with free algae, the method has the advantages that free cells are positioned in a limited space area by physical and chemical means to immobilize the microalgae, so that the microalgae has the characteristics of high cell density, high reaction speed, strong load capacity, stable and reliable operation, easiness in solid-liquid separation and the like, and the catabolic activity of the algae can be reduced to a certain degree, so that the biological activity of the algae is maintained and the algae can be repeatedly utilized.
The traditional immobilized microalgae technology mainly comprises an adsorption method and an embedding method. The adsorption is to attach the algae cells on the surface of the carrier, and the embedding is to embed or seal the algae cells in the carrier, and the obtained carrier mainly comprises organic and inorganic high molecular materials and the like. The synthesis cost of the carriers is high, the preparation procedure is complex, more steps such as centrifugation, flushing and the like are needed, after the carriers are used for a certain time, the growth of algae cells causes the shedding of the algae cells or the rupture of immobilized algae balls, and the adverse factors influence the application of the immobilized microalgae to a certain extent.
The invention adopts bacterial cellulose generated by acetobacter xylinum widely existing in nature to capture microalgae and is used as a carrier to realize immobilization, and researches on treating wastewater by using the bacterial cellulose immobilized microalgae are developed. The method is simple and efficient, has low cost, and greatly reduces the cost of microalgae for treating wastewater.
Disclosure of Invention
In view of the above, the present invention aims to provide a method for treating wastewater by immobilized microalgae using bacterial cellulose, which uses acetobacter xylinum to generate bacterial cellulose to trap microalgae into large clusters or spheres, thereby developing a simple, efficient and low-cost technology for treating wastewater by immobilized microalgae.
In order to achieve the above object, the present invention provides a method for treating wastewater by using microalgae immobilized by bacterial cellulose, comprising the following steps:
(1) high-density culture of microalgae;
(2) culturing acetobacter xylinum;
(3) adding a acetobacter xylinum culture medium into the microalgae culture solution according to a certain proportion, inoculating the cultured acetobacter xylinum, and generating bacterial cellulose by the acetobacter xylinum to capture and immobilize the microalgae;
(4) the immobilized microalgae cells are used for treating wastewater.
Further, the high-density culture of the microalgae in the step (1) specifically comprises: after the microalgae are subjected to amplification culture under the conditions of illumination intensity of 3000-5000 lux and temperature of 20-28 ℃, the microalgae are inoculated to a culture device for culture until the cell logarithmic phase reaches 10 cell density6～1010As described above. The culture method may be an autotrophic culture or a heterotrophic culture, and is intended to obtain a high-density algal solution. The microalgae is cultured to the logarithmic phase because the density of the microalgae is high, the activity is strong, the wastewater treatment effect after immobilization is good, the acetobacter xylinum is cultured to the logarithmic growth phase and then is inoculated into the algae liquid for capture, the activity of the generated cellulose is strong, and the rapid capture and immobilization of the microalgae are facilitated.
Further wherein the culture device is selected from one of a shake flask, a gas-through bottle, a photobioreactor, a fermenter, or an open culture tank.
Further, the microalgae include, but are not limited to, one of the genera Chlorella (chlorella sp.), Cylindrocarpus (cy1 indetheca sp.), Diatom (diatom), Nitzschia sp, Schizochytrium sp, Scenedesmus sp, Chlorococcus sp, Nannochloropsis sp, Chlamydomonas sp, Tetraselmis sp, Eudorina sp, or a mixture of such species of microalgae, among others.
Further, the culture medium of the microalgae includes, but is not limited to, one of BG-11 medium, SE medium, BBM medium or TAP medium, and any other modified microalgae culture medium and heterotrophic culture medium thereof.
Further, wherein the culture conditions of acetobacter xylinum in step (2) are as follows: culturing at 25-28 deg.C and 130r/min for 5 days.
Further, the formula of the acetobacter xylinum culture medium is that 30g/l of fructo-oligosaccharide, 20g/l of yeast extract (powder), 0.4g/l of anhydrous magnesium sulfate, 3.3g/l of ammonium sulfate, 1g/l of potassium dihydrogen phosphate, 1.75g/l of citric acid monohydrate, 2.4g/l of sodium citrate dihydrate and the initial pH value is 4.6.
Further, the step (3) specifically comprises: adding the acetobacter xylinum culture medium into the microalgae culture solution obtained in the step (1) according to the proportion of 10-50% (v/v), inoculating the acetobacter xylinum cultured in the step (2), controlling the culture condition until the acetobacter xylinum generates bacterial cellulose to trap the microalgae to form a dough or a ball, and harvesting the microalgae by using a simple separation method.
Further, the step (3) specifically comprises the steps of adding a acetobacter xylinum culture medium into the microalgae culture solution according to the proportion of 10% -40% (v/v), inoculating 1% -5% (v/v) acetobacter xylinum, oscillating or stirring for 4-48 hours at the temperature of 24-30 ℃ and at the speed of 0-80 r/min, enabling the acetobacter xylinum to generate bacterial cellulose to capture microalgae and form a cluster or sphere, and obtaining the immobilized microalgae material with the bacterial cellulose as a carrier by a fishing or filtering method.
Further, wherein the wastewater in step (5) includes, but is not limited to, one of municipal wastewater, industrial wastewater (such as brewery wastewater and paper mill wastewater), aquaculture wastewater (such as pig manure wastewater), or biogas fermentation wastewater.
Further, the step (5) specifically comprises: the immobilized microalgae agglomerates are put into various wastewater which can be adapted to microalgae (different microalgae have different characteristics and can treat different wastewater), the immobilized microalgae agglomerates can keep the photosynthetic activity under certain temperature and illumination, and the immobilized microalgae agglomerates have good effects of removing nitrogen, phosphorus and COD and high adsorbability.
The invention has the following beneficial effects:
the method has low cost, and the immobilized microalgae can be used for treating the wastewater repeatedly, and is safe and nontoxic. As long as the acetobacter capable of producing the bacterial cellulose can be used for trapping and immobilizing the microalgae for treating wastewater, the applicant finds that the acetobacter can grow quickly and has high bacterial cellulose yield and can obtain good effect, the degree of immobilizing the microalgae by the bacterial cellulose can be controlled to keep the metabolic activity of the microalgae, the immobilized aggregate has high tensile strength and is not easy to break, the algae is not easy to fall off, the operation is stable and reliable, and the acetobacter can be used for treating wastewater.
The method introduces an immobilization technology of collecting microalgae by bacterial cellulose to treat wastewater, and immobilized algae cells after the wastewater is treated can be recycled, so that the requirement of industrial wastewater treatment of microalgae is met; the invention is suitable for various wastewater treatments, has lower price and simple operation, is beneficial to wastewater treatment, and is a new way for economically and efficiently preparing microalgae wastewater treatment.
Drawings
FIG. 1A is a light mirror image of a bacterial cellulose-trapped immobilized scenedesmus obliquus pellet;
FIG. 1B is a scanning electron microscope image of a bacterial cellulose-trapped immobilized scenedesmus obliquus pellet;
FIG. 2A is a diagram showing the effect of bacteria cellulose in capturing immobilized Scenedesmus obliquus cells to remove high-concentration culture wastewater;
FIG. 2B is a second graph showing the effect of bacteria cellulose in capturing immobilized Scenedesmus obliquus cells to remove high-concentration culture wastewater;
FIG. 2C is a third diagram showing the effect of bacteria cellulose in capturing immobilized Scenedesmus obliquus cells to remove high-concentration culture wastewater;
FIG. 2D is a fourth diagram showing the effect of bacteria cellulose in capturing immobilized Scenedesmus obliquus cells to remove high-concentration culture wastewater.
Detailed Description
Other features and advantages of the present invention will be further illustrated by the following examples, which are intended to be illustrative only and not limiting.
Example 1: chlorella immobilization treatment wastewater
The SE medium for chlorella culture was as follows: NaNO30.25g/L；K2HPO4-3H2O 0.075g/L；MgSO4-7H2O 0.075g/L；CaCl2-2H2O 0.025g/L；KH2PO40.175g/L；NaCl 0.025g/L；FeCl3-6H2O0.005 g/L; 1mL of EDTA-Fe; a51 mL; 40mL of soil extract.
Preparing a Soil extracting solution (Soil extract): putting a proper amount of garden soil (without being fertilized) into an oven, drying at 60 ℃ overnight, weighing 250g of garden soil, pouring into a 3000mL triangular flask, adding 1000mL of distilled water, sealing the opening of the flask by using a vent plug, heating in water bath for 3h by boiling water, cooling, precipitating overnight, centrifuging at 1000rpm/min for 5min under the aseptic condition, taking supernatant, adding sterilized distilled water into the supernatant to reach the volume of 1000mL, and storing the soil extract at 4 ℃ for later use.
EDTA-Fe solution: weighing 4.1ml of concentrated hydrochloric acid and diluting the concentrated hydrochloric acid into 500ml of distilled water; 0.9306g of 1N EDTA-Na2 are weighed and dissolved in 50ml of distilled water; 10ml of 1N HCl containing 0.901g of FeCl3 & 6H2O and 10ml of 0.1N EDTA-Na2 were weighed out and mixed, and diluted to 1000 ml.
A5(Trace mental solution) solution: h3BO32.8g/L dH2O；MnCl·4H2O 1.86g/L，ZnSO4·7H2O 0.22g/L；Na2MoO4·2H2O 0.39g/L；CuSO4·5H2O 0.08g/L；Co(NO3)2·6H2O is 0.05 g/L. The heterotrophic culture medium is prepared by the same method, and organic carbon sources (such as glucose and fructose) are added to the medium until the concentration of the initial reducing sugar is 1-15 g/L.
At 1 to 5 × 106Inoculating chlorella into SE culture medium at cell density of one/ml, culturing under aeration at 20-30 deg.C and 30-54 μmol/m2Culturing for 10 days under the s-illumination condition, wherein the cell density of the algae is 2-6 multiplied by 107And obtaining the chlorella culture solution per milliliter. For chlorella with heterotrophic culture characteristics, the ratio is 1-5 multiplied by 106Inoculating the chlorella into a heterotrophic culture medium, and culturing for 6 days at 25-30 ℃ in a shaking table at 80-120 r/min until the cell density of the chlorella reaches 2-9 multiplied by 109And obtaining the chlorella culture solution per milliliter. The obtained Chlorella culture solution was used for the following experimentsIn the examples.
Taking 500ml of chlorella culture solution cultured in an autotrophic or heterotrophic mode, adding 120ml of sterilized acetobacter xylinum culture medium, inoculating 6% (v/v) of acetobacter xylinum, starting to generate lumps after 12 hours, and taking out immobilized microalgae lumps captured by cellulose when the capture rate in 48 hours reaches more than 98%. And transferring the formed immobilized microalgae mass into high-concentration (1000-1500 mg/l COD) municipal wastewater and 1500ml pig manure wastewater, and carrying out intermittent aeration treatment without stirring. The result shows that the bacterial cellulose capture immobilized chlorella has the effects of efficiently removing ammonia, phosphorus and COD (after 6 days, the ammonia nitrogen removal rate reaches over 90 percent, the total nitrogen removal rate reaches over 65 percent, the total phosphorus removal rate reaches over 65 percent, and the COD removal rate reaches over 60 percent), and also has efficient adsorbability.
Example 2: scenedesmus obliquus immobilized wastewater treatment method
The SE medium for culturing Scenedesmus obliquus is as follows: NaNO30.25g/L；K2HPO4-3H2O 0.075g/L；MgSO4-7H2O 0.075g/L；CaCl2-2H2O 0.025g/L；KH2PO40.175g/L；NaCl 0.025g/L；FeCl3-6H2O0.005 g/L; 1mL of EDTA-Fe; a51 mL; 40mL of soil extract.
Preparing a Soil extracting solution (Soil extract): putting appropriate amount of garden soil (not fertilized) into an oven, drying at 60 deg.C overnight, weighing 250g, pouring into 3000mL triangular flask, adding 1000mL distilled water, sealing the bottle mouth with a gas-permeable plug, heating in water bath with boiling water for 3 hr, cooling, precipitating overnight, and sterilizing
Centrifuging at 1000rpm/min for 5min at 800-.
EDTA-Fe solution: weighing 4.1ml of concentrated hydrochloric acid and diluting the concentrated hydrochloric acid into 500ml of distilled water; 0.9306g of 1N EDTA-Na2 are weighed and dissolved in 50ml of distilled water; 10ml of 1N HCl containing 0.901g of FeCl3 & 6H2O and 10ml of 0.1N EDTA-Na2 were weighed out and mixed, and diluted to 1000 ml.
A5(Trace mental solution) solution: h3BO32.8g/L dH2O；MnCl·4H2O 1.86g/L，ZnSO4·7H2O 0.22g/L；Na2MoO4·2H2O 0.39g/L；CuSO4·5H2O 0.08g/L；Co(NO3)2·6H2O 0.05g/L。
At 15X 106Inoculating Scenedesmus obliquus into SE culture medium at cell density of algae/ml, culturing in air, and culturing at 20-30 deg.C and 30-54 μmol/m2S light irradiation, after 10 days of culture, the cell density of algae reaches 2-6 × 107And obtaining the scenedesmus obliquus culture solution per milliliter. The obtained scenedesmus obliquus culture solution was used in the following examples.
Taking 500ml of scenedesmus obliquus culture solution, adding 110ml of sterilized acetobacter xylinum culture medium into the culture solution, inoculating 3% (v/v) of cultured acetobacter xylinum, collecting and agglomerating after 4 hours, wherein the collection efficiency reaches 95% after 8 hours, and then taking out the immobilized microalgae agglomerate (shown in fig. 1A and 1B) collected by cellulose, wherein fig. 1A shows the state of the immobilized microalgae agglomerate observed under a light mirror, and fig. 1B shows the microscopic state of the immobilized microalgae agglomerate observed under a scanning electron microscope; as can be seen from FIGS. 1A and 1B, Scenedesmus obliquus is gathered and wrapped, and is wrapped by cellulose, so that the Scenedesmus obliquus is relatively firm and not easy to break. The immobilized microalgae is 2-6 × 108The block mass is transferred into 2000ml of culture wastewater or biogas fermentation wastewater with the initial COD value of 1000mg/L, stirring is not needed, aeration is carried out once per hour, and the aeration rate is 200 ml/min; the results of fig. 2A-2D show that the bacteria cellulose capture immobilized scenedesmus obliquus has high ammonia, phosphorus and COD removal effects (after 6 days, ammonia nitrogen removal rate reaches more than 95%, total nitrogen removal rate reaches more than 85%, total phosphorus removal rate reaches more than 70%, and COD concentration removal rate reaches more than 75%), and also has high adsorption.
Example 3: mixed algae immobilized wastewater treatment
Preparing SE culture medium with 1-5 × 106 Inoculating 5% of each of Chlamydomonas reinhardtii, Chlorella and Scenedesmus obliquus to each of the cells/ml, culturing in an aerated medium at 20-30 deg.C and 30-54 μmol/m2S illuminationCulturing for 10 days under the condition to obtain a mixed algae solution, wherein the cell density of algae is 2-6 multiplied by 107One/ml. The resulting mixed algal culture liquid was used in the following examples. And taking 500ml of mixed algae culture solution, adding 100ml of sterilized acetobacter xylinum culture medium into the mixed algae culture solution, inoculating 5% (v/v) of cultured acetobacter xylinum, collecting the mixture into a mass after 4 hours, and collecting the mixture with the collection efficiency reaching 96% after 12 hours to obtain the immobilized microalgae mass collected by cellulose. The immobilized microalgae is 2-6 × 109The block is added into beer mill wastewater and paper mill wastewater of 1200mg/LCOD, aeration is carried out once every hour, and the result of aeration rate of 200ml/min shows that the bacterial cellulose capturing immobilized mixed algae block has the effects of high-efficiency ammonia removal, phosphorus removal and COD removal (after 6 days, the ammonia nitrogen removal rate reaches more than 95%, the total nitrogen removal rate reaches more than 70%, the total phosphorus removal rate reaches more than 75%, and the COD concentration removal rate reaches more than 65%).
In summary, the above description is only a preferred embodiment of the present invention, and not intended to limit the scope of the present invention, so that all equivalent changes made in the description and drawings of the present invention should be included in the scope of the present invention.
Claims (6)
1. A method for treating wastewater by using bacterial cellulose immobilized microalgae is characterized by comprising the following steps:
(1) high-density culture of microalgae: after the microalgae are subjected to amplification culture under the conditions of illumination intensity of 3000-5000 lux and temperature of 20-28 ℃, the microalgae are inoculated to a culture device for culture until the cell logarithmic phase reaches 10 and the cell density reaches 106-1010So far;
(2) culturing acetobacter xylinum: culturing at 25-28 deg.C and 130r/min for 5 days;
(3) adding a acetobacter xylinum culture medium into a microalgae culture solution according to the proportion of 10-40% (v/v), inoculating 1-5% (v/v) acetobacter xylinum, oscillating or stirring for 4-48 hours at the temperature of 24-30 ℃ and at the speed of 0-80 r/min, generating bacterial cellulose by the acetobacter xylinum to capture microalgae and form a cluster or sphere, and obtaining an immobilized microalgae material taking the bacterial cellulose as a carrier by a salvage or filtration method;
(4) the immobilized microalgae cells are used for treating wastewater.
2. The method of claim 1, wherein the culture device is selected from one of a shake flask, an aeration bottle, a photobioreactor, a fermentor or an open culture pond.
3. The method of claim 2, wherein the microalgae comprises Chlorella (Chlorella)chlorella sp.) Cylindrocarpon (D) of Cylindrocarponcy1indrotheca sp.) Diatoms (A), (B), (C), (D), (C), (D), (C), (diatom) Diamond shaped algae (D.sp.)Nitzschia sp.) Schizochytrium limacinum (Schizochytrium limacinum (Fr.) pers.), (schizochytrium sp.) Scenedesmus (Scenedesmus)Scenedesmus sp.) Chlorococcum micranthum, (b) ANannochloris sp.) Chlamydomonas (A), (B), (CChlamydomonas sp.) Flat algae (D, E)Tetraselmis sp.) Genus Haematococcus (A. borealis)Eudorina sp.) One or a mixture of microalgae of these species.
4. The method of claim 1, wherein the culture medium of the microalgae comprises one of a BG-11 medium, a SE medium, a BBM medium, or a TAP medium.
5. The method of claim 1, wherein the culture medium for acetobacter xylinum is formulated with 30g/l fructo-oligosaccharide, 20g/l yeast extract, 0.4g/l anhydrous magnesium sulfate, 3.3g/l ammonium sulfate, 1g/l potassium dihydrogen phosphate, 1.75g/l citric acid monohydrate, 2.4g/l sodium citrate dihydrate, and an initial pH of 4.6.
6. The method of claim 1, wherein the wastewater in step (4) comprises one of municipal wastewater, industrial wastewater, aquaculture wastewater, or biogas fermentation wastewater.
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| CN109231488B (en) * | 2018-10-19 | 2021-06-18 | 浙江海洋大学 | Method for synchronously treating nitrogen and phosphorus inorganic nutrients and organic pollutants in water body |
| CN111018127A (en) * | 2020-02-12 | 2020-04-17 | 青岛卓越海洋集团有限公司 | Method for treating aquaculture wastewater by algae |
| CN115215506A (en) * | 2022-06-07 | 2022-10-21 | 海南省智慧环境投资控股有限公司 | Planting and breeding coupling method for purifying breeding sewage by using mangrove forest |
| CN117089544B (en) * | 2023-09-06 | 2024-05-14 | 山东悦翔生物有限公司 | Microalgae culture method based on modified cellulose carrier |
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