AU2020103345A4

AU2020103345A4 - Method for treating phosphorus-containing wastewater with microalgae

Info

Publication number: AU2020103345A4
Application number: AU2020103345A
Authority: AU
Inventors: Liang FU; Yutong Xie; Ge YAN; Zhenhao Zhao; Dandan ZHOU
Original assignee: Northeastern University China; Northeast Normal University
Current assignee: Northeast Normal University
Priority date: 2020-11-10
Filing date: 2020-11-10
Publication date: 2021-01-21
Anticipated expiration: 2028-11-10

Abstract

The present disclosure relates to a method for treating phosphorus-containing wastewater with microalgae, including: step (1): selecting microalgae in a logarithmic growth phase as a source of inoculum and controlling an initial cell density > 0.5x106 cells/mL; step (2): autotrophically cultivating the microalgae at 20-32°C with a light level of 2,000-20,000 Lux and a light-dark cycle of 10-24 h :14-0 h; step (3): directly adding phosphorus-containing wastewater with P043-- P < 50 mg/L at one time, or adding phosphorus-containing wastewater with P043-- P > 50 mg/L in batches at different times evenly distributed during a cultivation cycle to ensure a maximum phosphorus concentration < 100 mg/L after each addition. The present disclosure achieves direct use of phosphorus-containing wastewater with a relatively high phosphorus concentration in cultivation of microalgae. The present disclosure can effectively avoid osmotic or toxic effects due to a high concentration of substrate, and meet nutrient demands during a growth cycle of the microalgae without supplementing an organic carbon source. Moreover, in the present disclosure, the microalgae can be recovered and reused. 15 DRAWINGS 13 FIG.1 16

Description

DRAWINGS

13

FIG.1

METHOD FOR TREATING PHOSPHORUS-CONTAINING WASTEWATER WITH MICROALGAE TECHNICAL FIELD The present disclosure belongs to the field of biological treatment of phosphorus-containing wastewater, and specifically relates to a method for treating phosphorus-containing wastewater with microalgae. BACKGROUND Phosphorus is a main nutrient element for growth of organisms and an important indicator of water eutrophication. When a phosphorus content in water exceeds 20 mg/L, algae grow and reproduce rapidly and float on a water surface to form a blue-green or red-yellow water flower or film, that is, an algae bloom. This leads to decrease of dissolved oxygen in water and release of organic substances such as algal toxins, resulting in turbid and smelly water with seriously degraded water quality, and even potential threats to drinking water safety. Due to development of industrial and agricultural production and increase in population, phosphorus-containing pesticides and fertilizers have been used in a large quantity. This leads to increasingly serious phosphorus pollution in water. A considerable part of phosphorus pollution in surface water is caused due to industrial and agricultural production activities such as phosphate mining and processing, phosphate fertilizer production, production in pharmaceutical industry and livestock breeding. These production activities produce a large amount of phosphorus-containing wastewater where phosphorus is mainly present as phosphate (P0 4 3-) in a concentration of 3-300 mg/L. In China's "Integrated Wastewater Discharge Standard" (GB8978-1996), a primary standard level for phosphate (calculated in P) is < 0.5 mg/L, and a secondary standard level is < 1.0 mg/L. The environmental protection department sets that, wastewater must achieve the secondary standard level before entering urban sewage pipe network. Effective reduction of phosphorus content in wastewater to reduce environmental burden has attracted great attention from professionals in the industry. Methods for treating phosphorous-containing wastewater are divided into physicochemical methods and biological methods. The physicochemical methods are disadvantageous in the need to add chemicals, prepare special materials, and consume additional electrical energy and the like, which leads to a high cost in phosphorus removal and secondary pollution. At present, biological methods, compared with the physicochemical methods, can reduce the cost of treating phosphorus-containing wastewater. Biological methods mainly rely on phosphorus-accumulating bacteria with "phosphorus absorption under aerobic conditions and phosphorus release under anaerobic conditions" to achieve phosphorus removal through sludge discharge. But the sludge discharged still needs subsequent disposal. Microalgae use CO 2 to grow autotrophically through photosynthesis and absorb nutrients such as nitrogen and phosphorus. They are a highly adaptable single-celled microorganism and an excellent raw material for biodiesel. A concentration of phosphorus is 5.4 mg/L in normal cultivation of microalgae. When there is moderately excessive phosphorus in the environment (< 45 mg/L), microalgae cells absorb and convert the excessive phosphorus into polyphosphate (polyphosphorus, Poly-P) which is stored in the cells. Due to abundant phosphoric anhydride bonds which store energy, the Poly-P can provide additional energy for cell growth and lipid synthesis. Therefore, moderately excessive phosphorus positively promotes growth and metabolism of microalgae. However, a strategy with moderately excessive phosphorus will inevitably increase the cost of microalgae cultivation. Using phosphorus-containing wastewater as a source of excessive phosphorus can reduce the cost of microalgae cultivation and turn the phosphorus-containing wastewater to a useful resource. Therefore, in recent years, researchers generally believe that combination of microalgae cultivation and wastewater treatment is an effective way to reduce biodiesel production cost and turn wastewater into a useful resource. However, in practical applications, there are challenging problems as follows: 1. studies have found that a high concentration of phosphorus (> 150 mg/L) can lead to cytotoxicity, where combination of the high concentration of phosphorus with intracellular components can damage organelles, thereby inhibiting the growth of microalgae cells. In actualphosphorus-containing wastewater, the phosphorus concentration can be as high as 300 mg/L or more. 2. In a conventional method for cultivating microalgae, adding a high concentration of nutrients at an initial stage when a cell density is relatively low will show inhibition to a certain extent. As cell number increases with nutrients consumed, nutrient supply will be insufficient for cells at a high density in a later stage, which is also not conducive to cell growth and metabolism. Imbalance of nutrient supply and demand prevents progress to optimal microalgae growth and lipid accumulation. 3. An autotrophic growth rate of microalgae is relatively low, and can be easily affected by many factors such as light, temperature, pH, C02 concentration and gas-liquid mass transfer efficiency. For the above reasons, phosphorus-containing wastewater, especially wastewater with a relatively high phosphorus concentration, cannot be directly used for cultivation of microalgae. It is an object of the present invention to address the foregoing problems or at least to provide the public with a useful choice. All references, including any patents or patent applications cited in this specification are hereby incorporated by reference. No admission is made that any reference constitutes prior art. The discussion of the references states what their authors assert, and the applicants reserve the right to challenge the accuracy and pertinency of the cited documents. It will be clearly understood that, although a number of prior art publications are referred to herein, this reference does not constitute an admission that any of these documents form part of the common general knowledge in the art, in

Australia or in any other country. It is acknowledged that the term 'comprise' may, under varying jurisdictions, be attributed with either an exclusive or an inclusive meaning. For the purpose of this specification, and unless otherwise noted, the term 'comprise' shall have an inclusive meaning - i.e. that it will be taken to mean an inclusion of not only the listed components it directly references, but also other non-specified components or elements. This rationale will also be used when the term 'comprised' or 'comprising' is used in relation to one or more steps in a method or process. Further aspects and advantages of the present invention will become apparent from the ensuing description which is given by way of example only.

DISCLOSURE OF INVENTION In order to solve the above problems, the present disclosure provides a method for treating phosphorus-containing wastewater with microalgae, which achieves direct use of phosphorus-containing wastewater, especially wastewater with a relatively high phosphorus concentration, in cultivation of microalgae. The method of the present disclosure can effectively avoid osmotic or toxic effects due to a high concentration of substrate, and fully meet nutrient demands during a growth cycle of the microalgae without supplementing an organic carbon source. Moreover, with the method of the present disclosure, the microalgae can be recovered and reused, turning wastewater into a useful resource. An objective of the present disclosure is achieved by the following technical solutions. A method for treating phosphorus-containing wastewater with microalgae includes the following steps: step 1: selecting microalgae in a logarithmic growth phase as a source of inoculum, and inoculating into an autotrophic cultivation system with an initial cell density controlled to be > 0.5x106 cells/mL; step 2: autotrophically cultivating the microalgae at 20-32°C with a light level of 2,000-20,000 Lux and a light-dark cycle of 10-24 h :14-0 h; step 3: directly adding phosphorus-containing wastewater with P0 43-- P < 50 mg/L to a microalgae cultivation system at one time, or addingphosphorus-containing wastewater with P0 4 3 --P > 50 mg/L in batches at different times evenly distributed during a microalgae cultivation cycle to ensure a maximum phosphorus concentration < 100 mg/L after each addition; step 4: monitoring change of phosphorus concentration and phosphorus removal, and discharging the phosphorus-containing wastewater when the phosphorus concentration is qualified; at the same time, determining microalgae biomass, lipid content and composition, and recovering the microalgae.

Another objective of the present disclosure is to provide a closed photoautotrophic cultivation apparatus, which can automatically control light, temperature, pH, and feed, and promote gas-liquid mass transfer through stirring and gas disperser. The apparatus includes a reactor body 15, a lighting device 8, a lighting control system 9, a stirring rotor 6, a stirring and temperature control device 7, a gas mixing device 3 and a gas disperser 5. The lighting device 8 is arranged on both sides of the reactor body 15 and in control connection with the lighting control system 9. A lower end of the reactor body 15 is provided with the stirring and temperature control device 7. An upper end of the stirring and temperature control device 7 is provided with the stirring rotor 6 which is located inside the reactor body 15. A pH probe 12 is provided inside the reactor body 15 and in control connection with a pH online monitoring and adjusting system 13 outside the reactor body 15. An inlet of the reactor body 15 is provided with a feeding port 10 which is in communication with an automatically feeding device 11. An inlet of the reactor body 15 is connected to a gas exhaust pipe 16 which connects to an exhaust gas absorption device 17. The gas disperser 5 is provided inside the reactor body 15 and in communication with the gas mixing device 3. A gas sterilization purifier 4 is provided on pipeline between the gas disperser 5 and the gas mixing device 3. A gas flow regulating valve 1 and a gas flow meter 2 are connected to the gas mixing device 3. The closed photoautotrophic cultivation apparatus can overcome many shortcomings of an open system, such as low cell density, easy contamination, water evaporation, insufficient C02 supply, and greater environmental impact. With the apparatus, cultivation conditions are stable and can be accurately controlled, aseptic operations can be carried out to effectively prevent contamination, a small area is needed, a high density culture can be enabled, and algae cells are easier to be harvested. Another objective of the present disclosure is to provide a method for treating phosphorus-containing wastewater with microalgae based on the above closed photoautotrophic cultivation apparatus, including steps as follows: step: selecting microalgae in a logarithmic growth phase as a source of inoculum, and inoculating through a sampling port into the reactor body of a closed photoautotroph system with an initial cell density > 0.5x106 cells/mL; step 2: lighting with a plant light in two directions with a light level of 2,000-20,000 Lux and a light-dark cycle of 10-24 h : 14-0 h, aerating with a mixed gas of C02 and air with a volume ratio of the two being 1-20% and an aeration rate per liter of culture medium of 0.05-1 L/min at 20-32°C, stirring a solution at 200-500 rpm, and controlling pH of liquid phase at 6.5-7.5 by the pH online monitoring and adjusting system; step 3: setting feeding volume and time of the automatically feeding device, adding phosphorus-containing wastewater to the reactor body through the feeding port, specifically, directly adding phosphorus-containing wastewater with P03 4 -- P< 50 mg/L to a microalgae cultivation system at one time at an initial stage of microalgae cultivation, or adding phosphorus-containing wastewater with P0 4 3 -P > 50 mg/L in batches at different times evenly distributed during a microalgae cultivation cycle to ensure a maximum phosphorus concentration < 100 mg/L; step 4: monitoring change of phosphorus concentration and phosphorus removal, and discharging the phosphorus-containing wastewater when the phosphorus concentration is qualified, determining microalgae biomass, lipid content and composition, and recovering the microalgae for preparation of biodiesel. Beneficial effects of the present disclosure are as follows: The present disclosure solves the problem that phosphorus-containing wastewater, especially wastewater with a relatively high phosphorus concentration, cannot be directly used for cultivation of microalgae. The present disclosure can effectively avoid osmotic or toxic effects due to a high concentration of substrate, and fully meet nutrient demands during a growth cycle of the microalgae without supplementing an organic carbon source. Moreover, with the present disclosure, the microalgae can be recovered and reused, turning wastewater into a useful resource. The microalgae in a logarithmic phase are inoculated to an apparatus for autotrophic cultivation. During an entire growth cycle of the microalgae, the phosphorus-containing wastewater is added for multiple times in batches with number of additions determined by the phosphorus concentration of the wastewater. Using phosphorus in batches to cultivate microalgae and treat phosphorus-containing wastewater can significantly reduce toxicity and inhibition due to the high concentration of phosphorus and increase the biomass by 0.9-2.3 times. As the number of phosphorus additions increases, the lipid content of the microalgae also gradually increases (from 23.5% to 40.5%) where cells are more inclined to synthesize saturated fatty acids (from 30% to more than 50%), and stability of the lipid increases. Moreover, A phosphorus removal rate reaches %-100%. In order to further improve growth rate of microalgae and treatment efficiency of phosphorus-containing wastewater, a closed photoautotrophic cultivation apparatus is designed and manufactured. The apparatus can automatically control light, temperature, pH, and feed, and promote gas-liquid mass transfer through stirring and gas disperser. In the method for treating phosphorus-containing wastewater with microalgae based on the above closed photoautotroph culture apparatus, the microalgae have a shortened growth cycle which is 2/3-1/4 of the original cycle, a maximum biomass of 5.5 g/L which is 1-2 times greater, and a growth rate increased by 2-3 times, and a phosphorus removal rate is increased by 1-3 times. Reference herein is made to various aspects and embodiments of the present invention. For clarity and to aid prolixity every possible combination, iteration or permutation of features, aspects and embodiments are not described explicitly. Thus, it should be appreciated that the disclosure herein includes any combination, iteration, multiple or permutation unless explicitly and specifically excluded. BRIEF DESCRIPTION OF DRAWINGS Further aspects and advantages of the present invention will become apparent from the following description which is given by way of example only and with reference to the accompanying drawings in which: FIG. 1 is a schematic structural diagram of a closed photoautotrophic cultivation system provided by an embodiment of the present disclosure; where: 1 is gas flow regulating valve, 2 is gas flow meter, 3 is gas mixing device, 4 is gas sterilization purifier, 5 is gas disperser, 6 is stirring rotor, 7 is stirring and temperature control device, 8 is lighting device, 9 is lighting control system, 10 is feeding port, 11 is automatically feeding device, 12 is pH probe, 13 is pH online monitoring and adjusting system, 14 is sampling port, 15 is reactor body, 16 is gas exhaust pipe, and 17 is exhaust gas absorption device. BEST MODES FOR CARRYING OUT THE INVENTION The present disclosure will be described in detail below with reference to specific embodiments. 1. Method for treating phosphorus-containing wastewater with microalgae Comparative Embodiment 1 Chlorella regularis was used in an experiment, and autotrophically cultivated in a 250 mL Erlenmeyer flask cultivation system at 28°C with a rotation speed of 160 rpm and a light level of 2,000 Lux. A culture medium was BG11 medium added with phosphorus-containing wastewater, in which nitrogen was 50 mg/L and phosphorus was 5.4 mg/L. In a cultivation cycle of 30-40 d, microalgae had maximum biomass of 1.8 g/L and a maximum growth rate of 0.23 g/(L-d), and a phosphorus removal rate reached 100% with a removal speed of 0.38 mg/( L-d). Algae cells had a lipid content of 23.5% in which unsaturated fatty acids took up more than 70%. Comparative Embodiment 2: Chlorella regularis was used in an experiment, and autotrophically cultivated in a 250 mL Erlenmeyer flask culture system at 28°C with a rotation speed of 160 rpm and a light level of 2,000 Lux. A culture medium was BG11 medium added with phosphorus-containing wastewater, in which nitrogen was 50 mg/L and phosphorus was 45 mg/L. In a cultivation cycle of 30-40 d, compared with Comparative Embodiment 1, microalgae had maximum biomass of 3.5 g/L which was increased by 93%, a maximum growth rate increased by nearly 2 times, and a phosphorus removal rate reached 100% with a removal speed increased by 4 times. Algae cells had a lipid content of 35% which was increased by 49% compared with control group 1, in which unsaturated fatty acids took up more than 70%. Comparative Embodiment 3: Chlorella regularis was used in an experiment, and autotrophically cultivated in a 250 mL Erlenmeyer flask cultivation system at 28°C with a rotation speed of 160 rpm and a light level of 2,000 Lux. A culture medium was BG11 medium added with phosphorus-containing wastewater, in which nitrogen was 50 mg/L and phosphorus was 200 mg/L. In a cultivation cycle of 30-40 d, compared with Comparative Embodiment 1, microalgae had maximum biomass (1.1 g/L) decreased by 40%, a maximum growth rate decreased by 37.5%, a lipid content decreased by 16%, and a phosphorus removal rate was 65% which was close to that in experimental group 1. Algae cells showed signs of poisoning such as enlargement, separation of cytoplasm from cell wall, and cell wall damage. Embodiment 1: Chlorella regularis was used in an experiment, and autotrophically cultivated in a 250 mL Erlenmeyer flask culture system at 28°C with a rotation speed of 160 rpm and a light level of 2,000 Lux. A culture medium was BG11 medium added with phosphorus-containing wastewater, in which nitrogen was 50 mg/L. During a cultivation cycle of 30-40 d, phosphorus-containing wastewater were added in 2 batches (at an initial and an intermediate time point), with phosphorus concentration being 100 mg/L after each addition. Compared with Comparative Embodiment 1, microalgae had maximum biomass of 2.1 g/L which was increased by 14%, and a maximum growth rate increased by 15%. A phosphorus removal rate was 84% with a removal speed increased by 29% compared with Comparative Embodiment 3. This indicated that, phosphorus at a high concentration can be added in batches to reduce its biological toxicity. Algae cells had a lipid content of 36.5% which was increased by 4% compared with Comparative Embodiment 2, in which unsaturated fatty acids took up more than %. Embodiment 2: Chlorella regularis was used in an experiment, and autotrophically cultivated in a 250 mL Erlenmeyer flask cultivation system at 28°C with a rotation speed of 160 rpm and a light level of 2,000 Lux. A culture medium was BG11 medium added with phosphorus-containing wastewater, in which nitrogen was 50 mg/L. During a cultivation cycle of 30-40 d, phosphorus-containing wastewater were added in 4 batches (at 0, 1/4, 1/2, 3/4 cycle), with phosphorus concentration being mg/L after each addition. Microalgae had maximum biomass of 3.6 g/L which was increased by 3% compared with Comparative Embodiment 2, and a phosphorus removal rate was 80-90% which was close to that in Embodiment 1. Algae cells had a lipid content of 3 8 .5% which was increased by 10% compared with Comparative Embodiment 2, in which unsaturated fatty acids took up more than 50%. Embodiment 3: (experimental group 5, Embodiment 4, addition for 8 times) Chlorella regularis was used in an experiment, and autotrophically cultivated in a 250 mL Erlenmeyer flask culture system at 28°C with a rotation speed of 160 rpm and a light level of 2,000 Lux. A culture medium was BG11 medium added with phosphorus-containing wastewater, in which nitrogen was 50 mg/L. During a cultivation cycle of 30-40 d, phosphorus-containing wastewater were added in 8 batches (at 0, 1/8, 1/4, 3/8, 1/2, 5/8, 3/4, 7/8 cycle), with phosphorus concentration being 25 mg/L after each addition. Microalgae had maximum biomass of 3.6 g/L which was increased by 3% compared with Comparative Embodiment 2, and a phosphorus removal rate was 80-90% which was close to that in Embodiment 1. Algae cells had a lipid content of 40.5% which was increased by 16% compared with Comparative Embodiment 2, in which unsaturated fatty acids was significantly reduced in percentage while saturated fatty acids took up more than 50%. This indicated that, addition of phosphorus in batches in cultivation facilitated improvement in content, stability and quality of biological lipid. Comparison of experimental results of the above embodiments led to the following conclusions: Moderately excessive phosphorus (45 mg/L) significantly promoted growth of microalgae by 93% and increased lipid accumulation by 49%. However, a significantly excessive phosphorus concentration (200 mg/L) caused cytotoxicity to microalgae instead, inhibiting growth by 40% and reducing lipid content by 16 %. Phosphorus supplementation in batches significantly reduced the toxicity due to a high concentration of phosphorus, and further increased the microalgae biomass (by 3-14%) and lipid content (by 4-16%) by adjusting number of batches. With increase in the number of batches, the microalgae were more inclined to synthesize saturated fatty acids with a proportion gradually increasing to more than 50%, thereby improving the stability and quality of biological lipid. With the above optimal conditions, the phosphorus removal rate reached 8 0 -9 0 % with a removal speed of 4-6 mg/(L-d), and 160-180 mg/L of phosphorus was removed within 30-40 d. 2. Method for treating phosphorus-containing wastewater with microalgae based on a closed photoautotrophic cultivation system Comparative Embodiment 4: Chlorella regularis was used in an experiment and inoculated into a closed photoautotrophic cultivation system. A culture medium used was BG11 medium added with phosphorous-containing wastewater at pH 6.5-7.5 with a nitrogen concentration of 50 mg/L and an initial algae density of 0.5x106-1x107 cells/mL. Bidirectional lighting was used with an average light level of 2,000-20,000

Lux and a light-dark cycle of 10-24 h : 14-0 h. A temperature was controlled at 20-32°C. A solution was stirred at 200-500 rpm. Feeding volume and time were set on an automatically feeding device. Phosphorus-containing wastewater was added to a reactor body through a feeding port. During a cultivation cycle of 10-20 d, phosphorus-containing wastewater were added in 8 batches (at 0, 1/8, 1/4, 3/8, 1/2, 5/8, 3/4, 7/8 cycle), with phosphorus concentration being 25 mg/L after each addition. The closed photoautotrophic cultivation system was run under the above conditions. Microalgae had maximum biomass of 3.9 g/L with a maximum growth rate of 0.65 g/(L-d). A phosphorus removal rate was 80-90% with a removal speed of 8-18 mg/(L-d), and lipid content and composition were similar to those in Embodiment 3. Comparative Embodiment 5: Chlorella regularis was used in an experiment and inoculated into a closed photoautotrophic cultivation system. A culture medium used was BG11 medium added with phosphorous-containing wastewater at pH 6.5-7.5 with a nitrogen concentration of 50 mg/L. An initial algae density was 0.5x106-1x107 cells/mL. Bidirectional lighting was used with an average light level of 2,000-20,000 Lux and a light-dark cycle of 10-24 h : 14-0 h. A temperature was controlled at 20-32°C. Aeration was carried out with a mixed gas of C02 and air with a volume ratio of the two being 1-20% and an aeration rate per liter of culture medium of 0.05-1 L/min. Feeding volume and time were set on an automatically feeding device. Phosphorus-containing wastewater was added to a reactor body through a feeding port. During a cultivation cycle of 10-20 d, phosphorus-containing wastewater were added in 8 batches (at 0, 1/8, 1/4, 3/8, 1/2, 5/8, 3/4, 7/8 cycle), with phosphorus concentration being 25 mg/L after each addition. The closed photoautotrophic cultivation system was run under the above conditions. Microalgae had maximum biomass of 5.2 g/L with a maximum growth rate of 0.87 g/(L-d). A phosphorus removal rate was 85-95% with a removal speed of 8.5-19 mg/(L-d), and lipid content and composition were similar to those in Embodiment 3. Embodiment 4: Chlorella regularis was used in an experiment and inoculated into a closed photosynthetic autotrophic cultivation system. An initial algae density was 0.5x10 6-1x107 cells/mL. A culture medium used was BG11 medium added with phosphorous-containing wastewater at pH 7 with a nitrogen concentration of 50 mg/L. Bidirectional lighting was used with an average light level of 2,000-20,000 Lux and a light-dark cycle of 10-24 h : 14-0 h. A temperature was controlled at -32°C. Aeration was carried out with a mixed gas of C02 and air with a volume ratio of the two being 3%, and an aeration rate per liter of culture medium was 0.05-1 L/min. A solution was stirred at 200-500 rpm. Feeding volume and time were set on an automatically feeding device. Phosphorus-containing wastewater was added to a reactor body through a feeding port. During a cultivation cycle of 10-20 d, phosphorus-containing wastewater were added in 8 batches (at 0, 1/8, 1/4, 3/8, 1/2, 5/8, 3/4, 7/8 cycle), with phosphorus concentration being 25 mg/L after each addition. The closed photoautotrophic cultivation system was run under the above conditions. Microalgae had maximum biomass of 5.5 g/L with a maximum growth rate of 0.92 g/(L-d). A phosphorus removal rate was 85-100% with a removal speed of 8.5-20 mg/(L-d), and lipid content and composition were similar to those in Embodiment 3. Compared with Comparative Embodiment 4, the biomass and the growth rate were increased by 40%, and the phosphorus removal rate was increased by 6-10%. Compared with Comparative Embodiment 5, the biomass and the growth rate were increased by 5%. Comparison of the above experimental results of embodiments with the closed photoautotrophic cultivation system showed that, growth and metabolism of the microalgae were significantly promoted. A growth cycle was shortened to 2/3-1/4 of the original cycle. The maximum biomass was increased by 1-2 times. The growth rate was increased by 2-3 times, and the phosphorus removal rate was increased by 1-3 times. Embodiment 5: Chlorella regularis was used in an experiment and inoculated into a closed photoautotrophic cultivation system. An initial algae density was 0.5x106-1x107 cells/mL. A culture medium used was BG11 medium added with phosphorous-containing wastewater at pH 6.5 with a nitrogen concentration of 50 mg/L. Bidirectional lighting was used with an average light level of 2,000-20,000 Lux and a light-dark cycle of 10-24 h : 14-0 h. A temperature was controlled at -32°C. Aeration was carried out with a mixed gas of C02 and air with a volume ratio of the two being 20%, and an aeration rate per liter of culture medium was 0.05-1 L/min. A solution was stirred at 200-500 rpm. Feeding volume and time were set on an automatically feeding device. Phosphorus-containing wastewater was added to a reactor body through a feeding port. During a cultivation cycle of 10-20 d, phosphorus-containing wastewater were added in 8 batches (at 0, 1/8, 1/4, 3/8, 1/2, 5/8, 3/4, 7/8 cycle), with phosphorus concentration being 25 mg/L after each addition. The closed photoautotrophic cultivation system was run under the above conditions. Microalgae had maximum biomass of 2.8 g/L with a maximum growth rate of 0.47 g/(L-d). A phosphorus removal rate was 4 0 -5 0 % with a removal speed of 8-18 mg/(L-d), and lipid content and composition were similar to those in Embodiment 3. Embodiment 6: Chlorella regularis was used in an experiment and inoculated into a closed photoautotrophic cultivation system. An initial algae density was 0.5x106-1x107 cells/mL. A culture medium used was BG11 medium added with phosphorous-containing wastewater at pH 7.5 with a nitrogen concentration of 50 mg/L. Bidirectional lighting was used with an average light level of

2,000-20,000 Lux and a light-dark cycle of 10-24 h : 14-0 h. A temperature was controlled at -32C. Aeration was carried out with a mixed gas of C02 and air with a volume ratio of the two being 1%, and an aeration rate per liter of culture medium was 0.05-1 L/min. A solution was stirred at 200-500 rpm. Feeding volume and time were set on an automatically feeding device. Phosphorus-containing wastewater was added to a reactor body through a feeding port. During a cultivation cycle of 10-20 d, phosphorus-containing wastewater were added in 8 batches (at 0, 1/8, 1/4, 3/8, 1/2, 5/8, 3/4, 7/8 cycle), with phosphorus concentration being 25 mg/L after each addition. The closed photoautotrophic cultivation system was run under the above conditions. Microalgae had maximum biomass of 4.5 g/L with a maximum growth rate of 0.75 g/(L-d). A phosphorus removal rate was 80 - 9 0 % with a removal speed of 8-18 mg/(L-d), and lipid content and composition were similar to those in Embodiment 3. The above embodiments showed that, a method of removing phosphorus from wastewater with photobiological cells was feasible, and application of a closed photoautotrophc cultivation system to the method can further improve growth rate of the microalgae and phosphorus removal. Supplementation in multiple batches can significantly reduce cytotoxicity due to a high concentration of phosphorus, and promote the growth of the microalgae (by 3-14%) and lipid accumulation (by 4-16%). The phosphorus concentration in treated wastewater was < 300 mg /L, and the phosphorus removal rate can reach 90%-100%. The microalgae used in all the embodiments of the present disclosure had not undergone domestication. According to existing literature reports, phosphorus removal efficiency can be further improved by means such as domestication of algae species, increase of inoculum concentration, and optimization of cultivation conditions. The above contents are only preferred embodiments of the present disclosure. It should be noted that, these embodiments are only used to illustrate the present disclosure but not intended to limit the scope of the present disclosure. Moreover, after reading the contents of the present disclosure, those skilled in the art can make various changes or modifications to the present disclosure, and these equivalent forms also fall within the scope defined by the appended claims of this application. It should be understood that there exist implementations of other variations and modifications of the invention and its various aspects, as may be readily apparent to those of ordinary skill in the art, and that the invention is not limited by the specific embodiments described herein. Features and embodiments described above may be combined with and without each other. It is therefore contemplated to cover any and all modifications, variations, combinations or equivalents that fall within the scope of the basic underlying principals disclosed and claimed herein. Although specific advantages have been enumerated above, various embodiments may include some, none, or all of the enumerated advantages.

Other technical advantages may become readily apparent to one of ordinary skill in the art after review of the following figures and description. It should be understood at the outset that, although exemplary embodiments are illustrated in the figures and described above, the principles of the present disclosure may be implemented using any number of techniques, whether currently known or not. Unless otherwise specifically noted, articles depicted in the drawings are not necessarily drawn to scale. Modifications, additions, or omissions may be made to the systems, apparatuses, and methods described herein without departing from the scope of the disclosure. For example, the components of the systems and apparatuses may be integrated or separated. Moreover, the operations of the systems and apparatuses disclosed herein may be performed by more, fewer, or other components and the methods described may include more, fewer, or other steps. Additionally, steps may be performed in any suitable order. As used in this document, "each" refers to each member of a set or each member of a subset of a set.

Claims

What is claimed is: 1. A method for treating phosphorus-containing wastewater with microalgae, comprising: step (1): selecting microalgae in a logarithmic growth phase as a source of inoculum, and inoculating into an autotrophic cultivation system with an initial cell density controlled to be > 0.5x106 cells/mL; step (2): autotrophically cultivating the microalgae at 20-32°C with a light level of 2,000-20,000 Lux and a light-dark cycle of 10-24 h :14-0 h; step (3): directly adding phosphorus-containing wastewater with P0 43-- P < 50 mg/L to a microalgae cultivation system at one time, or addingphosphorus-containing wastewater with P0 43 --P > 50 mg/L in batches at different times evenly distributed during a microalgae cultivation cycle to ensure a maximum phosphorus concentration < 100 mg/L after each addition; step (4): monitoring change of phosphorus concentration and phosphorus removal, discharging the phosphorus-containing wastewater when the phosphorus concentration is qualified, and at the same time, recovering the microalgae.
2. The method for treating phosphorus-containing wastewater with microalgae according to claim 1, wherein in step (3), the phosphorus-containing wastewater is added in 8 batches during a microalgae cultivation cycle.
3. A closed photoautotrophic cultivation apparatus for use in the method for treating phosphorus-containing wastewater with microalgae according to claim 1, wherein the apparatus comprises a reactor body (15), a lighting device (8), a lighting control system (9), a stirring rotor (6), a stirring and temperature control device (7), a gas mixing device (3) and a gas disperser (5), wherein the lighting device (8) is arranged on both sides of the reactor body (15) and in control connection with the lighting control system (9), a lower end of the reactor body (15) is provided with the stirring and temperature control device (7), an upper end of the stirring and temperature control device (7) is provided with the stirring rotor (6) which is located inside the reactor body (15), a pH probe (12) is provided inside the reactor body (15) and in control connection with a pH online monitoring and adjusting system (13) outside the reactor body (15), an inlet of the reactor body (15) is provided with a feeding port (10) which is in communication with an automatically feeding device (11), an inlet of the reactor body (15) is connected to a gas exhaust pipe (16) which is connected to an exhaust gas absorption device (17), the gas disperser (5) is provided inside the reactor body (15) and in communication with the gas mixing device (3), a gas sterilization purifier (4) is provided on pipeline between the gas disperser (5) and the gas mixing device (3), a gas flow regulating valve (1) and a gas flow meter (2) are connected to the gas mixing device (3).
4. A method for treating phosphorus-containing wastewater with microalgae by the closed photoautotrophic cultivation apparatus according to claim 3, comprising: step (1): selecting microalgae in a logarithmic growth phase as a source of inoculum, and inoculating through a sampling port into the reactor body of a closed photoautotrophic cultivation system with an initial cell density > 0.5x106 cells/mL; step (2): lighting by a plant light in two directions with a light level of 2,000-20,000 Lux and a light-dark cycle of 10-24 h :14-0 h, aerating with a mixed gas of C02 and air with a volume ratio of the two being 1-20% and an aeration rate per liter of culture medium of 0.05-1 L/min at 20-32°C, stirring a solution at 200-500 rpm, and controlling pH of liquid phase at 6.5-7.5 by the pH online monitoring and adjusting system; step (3): setting feeding volume and time of the automatically feeding device, adding phosphorus-containing wastewater to the reactor body through the feeding port, specifically, directly adding phosphorus-containing wastewater with P03 4 -- P< 50 mg/L to a microalgae

cultivation system at one time at an initial stage of microalgae cultivation, or adding phosphorus-containing wastewater with P0 43-- P > 50 mg/L in batches at different times evenly distributed during a microalgae cultivation cycle to ensure a maximum phosphorus concentration < 100 mg/L.
5. The method for treating phosphorus-containing wastewater with microalgae according to claim 4, wherein step (2) is implemented by lighting by a plant light in two directions with a light level of 2,000-20,000 Lux and a light-dark cycle of 10-24 h :14-0 h, aerating with a mixed gas of C02 and air with a volume ratio of the two being 3% and an aeration rate per liter of culture medium of 0.05-1 L/min at 20-32°C, stirring a solution at 200-500 rpm, and controlling pH of liquid phase at 7 by the pH online monitoring and adjusting system.

FIG. 1 DRAWINGS