Method for treating pig farm biogas slurry through phycomycete co-culture by adopting inclined plane flat plate type photobioreactor

Abstract

The invention discloses a method for treating pig farm biogas slurry through phycomycete co-culture by adopting an inclined plane flat plate type photobioreactor. The method comprises five steps of culturing a bacterial single strain, culturing a pure algae single colony, adjusting the component proportion of the pig farm biogas slurry, building the inclined plane flat plate type photobioreactor and treating the pig farm biogas slurry through phycomycete co-culture. According to the method, the symbiotic property of phycomycetes can be improved through preparation work such as selection of bacillus megaterium single strains and Desmodesmus sp pure single algae and adjustment of the component proportion of the pig farm biogas slurry, the treatment capacity of the pig farm biogas slurry is improved, and meanwhile the conversion efficiency of biomass is improved. According to the inclined plane flat plate type photobioreactor, flowing pig farm biogas slurry can be in full contact with phycomycetes on a carrier, continuous efficient innocent treatment can be conducted on the pig farm biogas slurry, and the inclined plane flat plate type photobioreactor is suitable for large-scale application and harvesting of phycomycetes biomass. The phycomycete carrier is low in price, easy to obtain, non-toxic and high in adhesion rate, phycomycetes in circulating liquid are not prone to loss, and solid-liquid separation is facilitated.

Description

Technical Field

[0001] The present invention relates to the treatment of biomass wastewater, and particularly to a method for co-culturing algae and bacteria to treat pig farm biogas slurry by using an inclined plate photobioreactor. Background Art

[0002] Pig farm biogas slurry is usually rich in nitrogen, phosphorus and organic carbon. If the pig farm biogas slurry is not properly treated and discharged into the environment, it will cause water pollution, eutrophication, algal blooms, reduction of DO, changes in aquatic ecology, serious odor problems, etc.

[0003] Existing relatively mature treatment methods for pig farm biogas slurry include: composting treatment, wastewater stabilization pond treatment, constructed wetland dilution and separation treatment, aquatic plant absorption treatment, microalgae treatment and other methods. Among them, microalgae treatment has attracted much attention due to its characteristics of environmental protection, high efficiency and low cost. In addition, microalgae have high contents of protein, antioxidants, vitamins and minerals, and can be used as feed additives for animal husbandry and aquaculture. However, since pig farm biogas slurry contains a large amount of alkaline substances, especially mainly NH 4+ , the pH of the biogas slurry can be close to 10. Different microalgae have different tolerances to NH 4+ under different environmental conditions. For most algae, when the pH is 9 to 10, the growth of algae will be severely inhibited, and the algae cannot grow continuously in pig farm biogas slurry, and cannot continuously convert and utilize the elements in the pig farm biogas slurry. Therefore, a co-culture form of microalgae and bacteria is usually adopted for sustainable large-scale harmless treatment of pig farm biogas slurry.

[0004] The algae-bacteria co-culture system can produce biomass energy while treating biogas slurry. However, in the co-culture system, microalgae cells and bacteria cells grow in suspension. If it is scaled up and applied in a reactor, there are problems such as easy loss of algae and bacteria and difficulty in solid-liquid separation. Summary of the Invention

[0005] Aiming at the problems of easy loss of algae and bacteria and difficulty in solid-liquid separation when using algae-bacteria co-culture to treat pig farm biogas slurry on a large scale, the present invention provides a method for co-culturing algae and bacteria to treat pig farm biogas slurry by using an inclined plate photobioreactor. This method adopts an algae-bacteria immobilized culture method to effectively separate algae-bacteria cells and the culture medium, improve the biomass, and reduce the biomass harvesting cost. The specific operation steps of the method of the present invention are as follows:

[0006] 1) Cultivate a single bacterial strain

[0007] Agar was added to the LB liquid culture medium, and after heating and sterilization, an LB solid medium was prepared. A dilution solution containing the Bacillus megaterium strain was spread on the LB solid medium and cultured in an incubator at 37 °C for 1 - 2 days. Single colonies with different morphologies were selected and transferred to a new LB solid medium, and isolation and purification were carried out by the streaking method to screen out the Bacillus megaterium single strain;

[0008] 2) Cultivate pure algal single colonies

[0009] Agar for the medium and penicillin were added to the BG11 liquid culture medium, and after heating to 45 °C and stirring, it was allowed to stand and solidify into a BG11 solid medium. The culture solution containing the Desmodesmus sp. microalgae was diluted with sterile water, and the diluted solution was spread on the solid medium. After culturing for 6 days, microalgal single colonies were picked for plate streaking separation. After 4 generations of separation, pure algal single colonies of Desmodesmus sp. were obtained. The solid medium with single colonies was placed in a 4 °C refrigerator for storage and standby;

[0010] 3) Adjust the component ratio of pig farm biogas slurry

[0011] The pig farm biogas slurry was filtered, and samples were taken to detect the C, N, and P elements in the filtered pig farm biogas slurry. Soluble nitrate and / or soluble phosphate were added to adjust C / N and N / P, and then it was transferred to a storage tank for standby;

[0012] 4) Build an inclined - plane plate - type photobioreactor

[0013] The reactor is a cuboid made of plexiglass, including a reaction tank and a top cover. The top cover can be opened, and a light source is provided inside the top cover. On the corresponding short vertical surfaces of the reaction tank, there are several horizontal liquid inlets and outlets respectively. A filter plate parallel to the short vertical surface is provided inside the reaction tank. The distance from the filter plate to the liquid outlet surface is 1 / 5 - 1 / 8 of the distance from the filter plate to the liquid inlet surface; the space between the filter plate and the liquid inlet surface is evenly divided into several culture tanks along the liquid flow direction, and the culture tanks are filled with algal - bacteria carriers; the bottom surface of the reaction tank is raised by 3° - 7° from the filter plate to the liquid inlet surface, and the bottom surface of the reaction tank is raised by 12° - 18° from the liquid outlet surface to the filter plate; the liquid inlet is connected to a peristaltic pump, the peristaltic pump is connected to the outlet of the storage tank, and the liquid outlet is connected to the inlet of the storage tank;

[0014] 5) Co - culture of algae and bacteria for treating pig farm biogas slurry

[0015] Desmodesmus sp. microalgae and Bacillus megaterium bacteria cultured to the logarithmic growth phase were added to the algal - bacteria carriers according to the quantity ratio. The algal - bacteria carriers were filled in the culture tanks, the light source cover plate was covered, the peristaltic pump was started, the light source was turned on, and circulation was carried out at room temperature for 4 - 6 days. During this period, samples were regularly taken to detect the biogas slurry indexes. After the indexes were stable, the circulation was stopped and the microalgae were recovered.

[0016] Specifically, in step 3), the C / N is adjusted to 100:(15-80), and the N / P is (15-40):1; in step 4), the algal-bacterial carrier is pine sawdust or cotton cloth; in step 5), the quantity ratio of Desmodesmus sp microalgae to Bacillus megaterium bacteria is (3-15):1.

[0017] The present invention can improve the algal-bacterial symbiosis by selecting single strain of Bacillus megaterium, pure algae of Desmodesmus sp, and adjusting the component ratio of pig farm biogas slurry, etc., improve the treatment capacity of pig farm biogas slurry and at the same time improve the conversion efficiency of biomass. The inclined-plane flat-plate photobioreactor of the present invention can make the flowing pig farm biogas slurry fully contact with the algal-bacteria co-cultured on the carrier, can continuously and efficiently harmlessly treat the pig farm biogas slurry, is suitable for large-scale application, and harvests algal-bacterial biomass. The algal-bacterial carrier of the present invention is cheap and easy to obtain, non-toxic and has a high attachment rate, and the algal-bacteria are not easy to lose in the circulating liquid, which is convenient for solid-liquid separation. Specific embodiments

[0018] The present invention is described below in conjunction with examples. The examples given are only used to explain the present invention and are not intended to limit the scope of the present invention.

[0019] The algal species and their sources used in the present invention are as follows: Chlorella vulgaris (FACHB-1072), Chlorella pyrenoidosa (FACHB-10), and Desmodesmus sp. (FACHB-2919) were purchased from the Freshwater Algae Culture Collection of the Institute of Hydrobiology, Chinese Academy of Sciences, and then separated and purified by the Marine Microalgae Biology Laboratory for standby; Desmodesmus sp. G41-M is a mutant algal strain with high biomass and high lipid yield isolated by the Marine Microalgae Biotechnology Laboratory from the inland river water sample in Xinjiang. The biogas slurry was taken from a pig farm in Yantai, Shandong Province.

[0020] Add 1.6% agar by weight of the medium and penicillin with a concentration of 25 U / mL to the BG11 liquid medium, heat to 45°C and stir until it solidifies, and let it stand to form the BG11 solid medium. Dilute the microalgae culture solution of Desmodesmus sp. with sterile water, take the diluted solution and spread it on the solid medium. After culturing for 6 days, pick the microalgae single colonies for streak plate separation. After separating 4 generations, obtain pure algae single colonies. Place the solid medium with single colonies in a 4°C refrigerator for storage and standby. The formula of the BG11 liquid medium is shown in Table 1.

[0021] Table 1. Formula of BG11 liquid medium

[0022]

[0023] The biogas slurry from the pig farm was serially diluted with sterile water at a ratio of 1:10 to obtain a biogas slurry dilution of 10 -4 ~10 -6 . The biogas slurry dilution was added to LB liquid culture medium with 1.5% agar by weight, sterilized by heating to make LB solid medium. The biogas slurry dilution was spread on the LB solid medium and cultured in an incubator at 37°C for 2 days. Different morphological single colonies were selected and transferred to a new LB solid medium, and separated and purified by streaking until the single strain of Bacillus megaterium was screened out. The formula of the LB liquid culture medium is shown in Table 2.

[0024] Table 2. Formula of LB liquid culture medium

[0025]

[0026]

[0027] The biogas slurry from the pig farm was filtered, and samples were taken to detect the C, N, and P elements in the filtered biogas slurry from the pig farm. Soluble nitrate and / or soluble phosphate were added to adjust the C / N to be between 100:(15 - 80), and the N / P to be between (15 - 40):1. Then it was transferred to a storage tank for standby.

[0028] An inclined - plane flat - type photobioreactor was built. The reactor is a cuboid made of plexiglass, including a reaction tank and a top cover. The top cover can be opened, and a light source is provided on the inner side of the top cover. A number of horizontal liquid inlets and outlets are respectively provided on the corresponding short vertical surfaces of the reaction tank. A filter plate parallel to the short vertical surface is arranged in the reaction tank. The distance from the filter plate to the liquid outlet surface is between 1 / 5 and 1 / 8 of the distance from the filter plate to the liquid inlet surface; the space between the filter plate and the liquid inlet surface is evenly divided into a number of culture tanks along the liquid flow direction, and the culture tanks are filled with algal - bacterial carriers; the bottom surface of the reaction tank is raised by 3° - 7° from the filter plate to the liquid inlet surface, and the bottom surface of the reaction tank is raised by 12° - 18° from the liquid outlet surface to the filter plate; the liquid inlet is connected to a peristaltic pump, the peristaltic pump is connected to the outlet of the storage tank, and the liquid outlet is connected to the inlet of the storage tank.

[0029] The Desmodesmus sp microalgae and Bacillus megaterium bacteria cultured to the logarithmic growth phase were added to the algal - bacterial carriers according to the quantity ratio, the algal - bacterial carriers were filled in the culture tanks, the light source cover plate was covered, the peristaltic pump was started, the light source was turned on, and it was circulated at room temperature for 4 - 6 days. During this period, samples were regularly taken to detect the biogas slurry indexes. After the indexes were stable, the circulation was stopped and the microalgae were recovered.

[0030] 1. When the immobilized algae-bacteria system with pine sawdust as the carrier is applied in a vertical flat-plate photobioreactor, its dry cell weight reaches 2.05 g / L, and the removal rates of COD, TP, and NH 4+ -N in biogas slurry are 91%, 89%, and 74% respectively. Compared with its application in a conical flask, the removal rates of TP and NH4+-N are increased by 61% and 73% respectively.

[0031] 2. When the immobilized algae-bacteria system with waste cotton cloth as the carrier is applied in an inclined flat-plate photobioreactor, its dry cell weight reaches 3.45 g / L, and the removal rates of COD, TP, and NH 4+ -N in biogas slurry are 100%, 98%, and 93% respectively. Compared with its application in a conical flask, the removal rates of COD, TP, and NH4+-N are increased by 32%, 52%, and 42% respectively.

[0032] 3. Using pine sawdust and waste cotton cloth as the carriers for immobilizing algae and bacteria and applying them in an inclined flat-plate photobioreactor can achieve large-scale treatment of biogas slurry while harvesting algae-bacteria biomass. The concentrations of COD, TP, and NH 4+ -N in the biogas slurry treated by the reactor all meet the requirements of the "Pollutant Discharge Standard for Livestock and Poultry Breeding Industries GB 18596-2001".

[0033] The above are only the preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principle of the present invention shall be included within the protection scope of the present invention.

Claims

1. A method for co-culturing algae and bacteria to treat biogas slurry from pig farms using an inclined plane flat-plate photobioreactor, characterized in that, It includes the following steps: 1) Cultivate single bacterial strains Add agar to the LB liquid culture medium, sterilize it by heating to make LB solid medium, spread the dilution solution containing Bacillus megaterium strains on the LB solid medium, place it in an incubator at 37°C for 1 - 2 days, select single colonies with different morphologies to a new LB solid medium, and separate and purify them by the streaking method to screen out Bacillus megaterium single strains; 2) Cultivate single pure algal colonies Add medium agar and penicillin to the BG11 liquid culture medium, stir it after heating to 45°C, and let it stand and solidify into BG11 solid medium. Dilute the culture solution containing Desmodesmus sp. microalgae with sterile water, take the dilution solution and spread it on the solid medium. After culturing for 6 days, pick single algal colonies of microalgae for plate streaking separation. After separating for 4 generations, obtain Desmodesmus sp. pure algal single colonies, and place the solid medium with single colonies in a 4°C refrigerator for storage for later use; 3) Adjust the component ratio of pig farm biogas slurry Filter the pig farm biogas slurry, take samples to detect the C, N, and P elements in the filtered pig farm biogas slurry, add soluble nitrate and / or soluble phosphate to adjust C / N and N / P, and transfer it to a storage tank for later use; 4) Build an inclined - plane flat - type photobioreactor The reactor is a cuboid made of plexiglass, including a reaction tank and a top cover. The top cover can be opened, and a light source is provided on the inner side of the top cover. A number of horizontal liquid inlets and outlets are respectively provided on the corresponding short vertical surfaces of the reaction tank. A filter plate parallel to the short vertical surface is provided in the reaction tank. The distance from the filter plate to the liquid outlet surface is 1 / 5 - 1 / 8 of the distance from the filter plate to the liquid inlet surface; the space between the filter plate and the liquid inlet surface is evenly divided into several culture tanks along the liquid flow direction, and the culture tanks are filled with algal - bacterial carriers; the bottom surface of the reaction tank is raised by 3° - 7° from the filter plate to the liquid inlet surface, and the bottom surface of the reaction tank is raised by 12° - 18° from the liquid outlet surface to the filter plate; the liquid inlet is connected to a peristaltic pump, the peristaltic pump is connected to the outlet of the storage tank, and the liquid outlet is connected to the inlet of the storage tank; 5) Co - culture algae and bacteria to treat pig farm biogas slurry Add Desmodesmus sp. microalgae and Bacillus megaterium bacteria in the logarithmic growth phase to the algal - bacterial carriers according to the quantity ratio, fill the algal - bacterial carriers in the culture tanks, cover the light source cover plate, turn on the peristaltic pump, turn on the light source, and circulate at room temperature for 4 - 6 days. During this period, regularly take samples to detect the biogas slurry indexes. Stop circulating after the indexes are stable, and recover the microalgae.

2. The method according to claim 1, wherein In step 3), adjust C / N to 100:(15 - 80), and N / P to (15 - 40):

1.

3. The method according to claim 2, wherein In step 4), the algal - bacterial carrier is pine sawdust or cotton cloth.

4. The method according to claim 3, characterized in that, In step 5), the quantity ratio of Desmodesmus sp. microalgae and Bacillus megaterium bacteria is (3 - 15):1.

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