JP2014132831A - Water purification method and liquid for water purification - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide liquid for water purification and a water purification method which can efficiently reduce nitrate, nitrite, and the like generated in the natural world, such as a marsh, a pond, a lake, a river, the sea, or wetlands, and a water environment of artificial equipment, such as a water bath for viewing or cultivation.SOLUTION: Liquid for water purification loads an external force to a culture solution containing raw water containing a microbe group including anaerobic microorganisms and particles composed of polyhydroxyalkanoate, and destroys at least a part of a biofilm formed on particle surfaces after cultivation starts before it completes.

Description

  The present invention relates to a water purification method and a water purification solution, and more specifically, swamps, ponds, lakes, rivers, seas, wetlands, water tanks, etc., using polyhydroxyalkanoate (hereinafter also referred to as “PHA”). The present invention relates to a water purification method and a water purification liquid.

  In the natural environment such as swamps, ponds, lakes, rivers, seas, and wetlands, and in the aquatic environment of artificial devices such as aquariums for amusement and aquaculture, various organisms survive. In these aquatic environments, for example, excreta such as vertebrates, organic substances containing nitrogen atoms such as proteins and nucleic acids contained in the carcasses thereof are decomposed by aerobic microorganisms or directly from invertebrates. Excretion produces ammonia, which is converted into nitrate and nitrite by microorganisms present in the water environment, and further converted into nitrogen and released into the atmosphere. In addition, it is known that organic matter containing phosphorus atoms contained in the excrement of living organisms and carcasses is converted by various microorganisms and produced as phosphates in the water environment or soil.

  By the way, conventionally, in wastewater treatment using activated sludge, methane fermentation, septic tanks, etc., methods using the action of microorganisms as described above have been used, and the contaminated environment has been repaired by focusing on the functions of various microorganisms. Attempts have been made. However, for example, in a closed aquarium for ornamental use or aquaculture, or in a natural environment where water currents are stagnant, excrement by aquatic organisms, food residues, etc. are converted to nitrogen to some extent as a result of the action of microorganisms. However, nitrates may accumulate in the water environment. As a result, in the natural environment, harmful water-blooms may be generated, and in the case of a water tank, moss may be generated and attached to the inner wall surface of the water tank.

  As a countermeasure, for example, in order to reduce the concentration of nitrate in the water tank and to suppress turbidity, a method using a specific microorganism (Non-Patent Document 1), a pellet of biodegradable polymer as a component serving as a nutrient source of the microorganism is used. A method to be used (Non-Patent Document 2) has been proposed. Non-Patent Document 2 proposes a method of using a reactor filled with biodegradable polymer pellets to circulate water in the water tank between the water tank and the reactor.

  However, the method described in Non-Patent Document 1 has a problem that a desired water purification effect cannot be obtained unless an appropriate microorganism is selected. In addition, it is practically very difficult to select a specific microorganism suitable for the natural environment or the water environment of an artificial device. Further, in the method described in Non-Patent Document 2, a biofilm is formed on the surface of the pellet and a certain water purification effect is obtained. However, for example, the nitrate treatment effect reaches a peak, and then hydrogen sulfide is generated. In some cases, the water environment may be degraded. In addition, in the reactor described in Non-Patent Document 2, pellets may be clogged around the reactor outlet where the water in the reactor is returned to the water tank, or some of the pellets may become hard to manage.

  In addition, for example, in an aquarium for breeding aquatic organisms for ornamental use or aquaculture, it is generally said that the nitrate concentration of water in the aquarium is preferably 50 mg / L or less, and further suppresses the mass generation of moss. The nitrate concentration is preferably 10 mg / L or less. However, the methods described in Non-Patent Documents 1 and 2 do not necessarily reduce the nitrate concentration sufficiently.

Coral Fish, A Publishing Company, April 10, 2011, No. 31, p111 Coral Fish, A Publishing Company, February 10, 2011, No. 30, p85

  In view of the above problems, the object of the present invention is, for example, the natural environment of swamps, ponds, lakes, rivers, seas, wetlands, etc., and the water environment of artificial devices such as aquariums for ornamental and aquaculture It is an object of the present invention to provide a water purification liquid and a water purification method capable of efficiently reducing nitrates, nitrites and the like generated in the process. Further, for example, as described in Non-Patent Document 2, when a reactor filled with particles such as pellets is used to purify the water environment in the water tank, the reactor is returned to the water tank when the liquid in the reactor is returned to the water tank. It is possible to provide a simple water purification method capable of suppressing clogging or solidification of particles or the like at or near the discharge port of water and a water tank reactor used in the method.

  As a result of intensive studies to solve the above problems, the inventors of the present invention, for example, with respect to a culture solution containing raw water collected from an aquarium in which aquatic organisms are bred and particles composed of polyhydroxyalkanoate, During the period from the start to the end, for example, an external force is applied by stirring or the like, and the culture solution obtained by destroying at least part of the biofilm formed on the surface of the PHA particles is put into the original water tank from which the raw water is collected When it is done (returned), it has been found that the nitrate concentration of water in the water tank is significantly reduced, and the present invention has been completed. The gist of the present invention is as follows.

(1) An external force is applied to the surface of the particles by applying an external force to the culture solution containing raw water containing a group of microorganisms containing anaerobic microorganisms and particles made of polyhydroxyalkanoate until the end of the culture. A water purification solution obtained by destroying at least a part of a formed biofilm.
(2) The water purification solution according to (1), wherein the external force is applied when the increase rate of the number of cells of the anaerobic microorganisms is slowed down.
(3) The water purification liquid according to (1) or (2), wherein the external force is loaded by stirring and / or vibration.
(4) Raw water containing microorganisms including anaerobic microorganisms collected from the natural environment or the water environment of artificial devices and particles made of polyhydroxyalkanoate are mixed before starting the culture and ending the culture In addition, when the increase rate of the number of anaerobic microorganisms slows down, at least a part of the culture solution obtained by applying an external force and destroying at least a part of the biofilm formed on the particle surface, A water purification method in which the raw water is collected and introduced into the water environment.
(5) The water purification method according to (4), wherein the external force is applied twice or more between the start of culture and the end of culture.
(6) The water purification method according to (4) or (5), wherein the polyhydroxyalkanoate is poly (3-hydroxybutyrate-co-3-hydroxyhexanoate).
(7) The water environment of the artificial device is a water environment including water in an aquarium where aquatic organisms are bred, and the nitrate concentration of water in the aquarium after the culture solution is introduced is 0 to 50 mg. / L is the water purification method according to any one of (4) to (6).
(8) An aquarium reactor for taking at least a part of a culture solution obtained by taking water in an aquarium where aquatic organisms are bred and culturing using the taken water. There,
A culture tank filled with the water taken up and particles made of polyhydroxyalkanoate;
An outlet for discharging the culture solution from the culture tank to the water tank;
Means for applying an external force for breaking at least a part of the biofilm formed on the particle surface;
Aquarium reactor equipped with.
(9) The reactor for a water tank according to (8), further comprising means for preventing the discharge port from being blocked in the vicinity of the discharge port.
(10) A water tank comprising the water tank reactor according to (8) or (9).

  According to the present invention, for example, nitrate, nitrite, etc. generated in the natural environment such as swamps, ponds, lakes, rivers, seas, wetlands, etc., and in the water environment of artificial devices such as aquariums for ornamental and aquaculture are efficiently used. It is possible to reduce well. In addition, when a reactor filled with particles such as pellets is used to purify the water environment in the aquarium, when returning the liquid in the reactor to the aquarium, particles are clogged at or near the reactor outlet. It is possible to provide a simple water purification method and an aquarium reactor used in the method.

It is the figure which showed an example of embodiment of the water tank provided with the reactor for water tanks of this invention. It is the figure which showed the other example of embodiment of the water tank provided with the reactor for water tanks of this invention. (A) It is a figure which shows a time-dependent change of the nitrate density | concentration of the water in the water tank in Example 1,3,5. (B) It is a figure which shows a time-dependent change of the nitrate density | concentration of the water in the water tank in Examples 2, 4 and Comparative Examples 1,2.

    Hereinafter, the present invention will be described in more detail.

  The liquid for water purification of the present invention is a culture solution containing raw water containing a group of microorganisms containing anaerobic microorganisms and particles composed of polyhydroxyalkanoate (PHA). An external force is applied to destroy at least a part of the biofilm formed on the particle surface.

  In the present invention, as described above, a mixture of predetermined raw water and particles made of PHA is used as a culture solution. After starting the culture and before ending the culture, an external force is applied to the culture solution. The biofilm formed on the surface of the PHA particles in the culture solution is destroyed. In the process of culturing, among the microbial groups contained in the raw water, first, aerobic microorganisms proliferate predominantly while utilizing PHA on the surface of the PHA particles to form a biofilm on the surface of the PHA particles. As this biofilm grows, an aerobic environment and an anaerobic environment are formed in the biofilm, so that various microbial groups that lived in the water environment from which the raw water was collected are included in the biofilm. . As the biofilm grows further, the inside of the biofilm becomes an anaerobic environment. Therefore, on the surface of the PHA particles, the layer in which anaerobic microorganisms live predominantly and the surface of the layer are aerobic microorganisms. It is thought that a layer that survives predominantly is formed. When the anaerobic microorganisms cover the entire surface of the PHA particles, it is difficult for PHA to decompose (utilize), that is, it is difficult for the various microorganisms contained in the biofilm to grow. Therefore, by destroying the biofilm formed on the surface of the PHA particles and peeling the biofilm from at least a part of the surface of the PHA particles so that the surface of the PHA particles is in contact with the culture solution, it is included in the biofilm. It is possible to increase the number of microorganism groups including aerobic and anaerobic microorganisms present on the culture medium side while maintaining a high growth rate of various microorganisms (mainly aerobic microorganisms) Conceivable.

According to a certain classification, the group of microorganisms contained in the biofilm can include protozoa, algae, fungi, Bacteria, cyanobacteria. In addition, these microorganism groups are classified into aerobic microorganisms (obligate aerobic microorganisms) that require oxygen to grow, and anaerobic microorganisms (obligate anaerobic microorganisms) that inhibit oxygen growth. It is said that there are facultative anaerobic microorganisms that grow with or without oxygen. In the present invention, facultative anaerobic microorganisms are included in anaerobic microorganisms.
And it is thought that the seed | species suitable for water environment grows among these various microorganisms, and contributes to a water purification effect | action.

  Among these, when attention is focused on the production of nitrate in the water environment, for example, the microorganisms present in various water environments include ammonia-oxidizing bacteria that are obligately aerobic bacteria that convert ammonia to nitrate. Nitrite-oxidizing bacteria, and vaginal bacteria that are anaerobic bacteria that convert the generated nitrate to nitrogen gas are considered to be included. And, it seems that the production of nitrate is controlled to some extent by microorganisms containing these bacteria, but it is not always easy to activate and stably function such microorganisms.

However, in the present invention, as described above, a culture solution obtained by culturing using predetermined raw water and PHA particles and deliberately destroying the biofilm in the course of culturing is used as a water purification solution. By simply returning the working solution to the original water environment, the concentration of nitrates in the water environment can be stably reduced and purified.
The detailed reason why such an effect can be obtained is not clear, but there is a balance of microbial species and amount suitable for each original water environment, and it originally exists for each water environment by culturing the collected raw water In other words, in order to increase the variety of microorganisms suitable for the water environment in a balanced manner, when they are put into the original water environment, they can be efficiently purified by the action of the microorganisms contained in the water purification solution that has been put in. it is conceivable that.

In this invention, it culture | cultivates using the raw | natural water containing the microorganism group containing an anaerobic microorganism as mentioned above. In the present invention, raw water means water collected from the natural environment such as swamps, ponds, lakes, rivers, seas, wetlands, and the like, and the aquatic environment of artificial devices such as ornamental and aquaculture tanks. Therefore, the raw water contains various microorganisms (microorganism group) suitable for the water environment according to the collected water environment. The microorganism group can include anaerobic microorganisms and aerobic microorganisms. Further, the natural world includes a place where an artificial facility is provided in part, such as an aquaculture farm or a fishing pond provided in the sea.
In the present invention, raw water may be simply referred to as “water”.

  In addition, from the viewpoint of purifying the water environment, the liquid for water purification of the present invention is collected from a place where purification of the water environment can be expected from the viewpoint of purging the water environment. Good. For example, in the case of a water environment existing in nature such as a swamp, a pond, a lake, a river, the sea, and a wetland, it may be collected from a place where the water flow tends to stagnate locally. In addition, in the case of an artificial device such as an aquarium or aquaculture device, the water environment circulates only in the artificial device, the inflow from the outside and the discharge to the outside, but the residence time in the device It is good to collect from the water environment in a long device. From the viewpoint of further enjoying the effects of the present invention, it is preferable to collect raw water from the water environment of the artificial device, and it is particularly preferable to collect raw water from the water environment of an aquarium for ornamental use or aquaculture. Examples of such aquarium include, but are not limited to, aquariums used in aquariums, exhibition aquariums for selling ornamental and edible aquatic organisms, domestic aquariums, etc. Absent. In addition, the aquatic organisms bred in such an aquarium are not particularly limited, and include various ornamental fish, edible fish such as scallops and goby, marine mammals, crustaceans, corals, and other aquatic animals and plants. It is applicable to the aquatic environment of aquariums that breed various aquatic organisms.

  After collecting raw water from such a water environment and performing a predetermined culture, the water purification liquid is diffused widely by introducing (returning) the water purification liquid of the present invention back to the original water environment. In addition, since the function of the microorganism group contained in the water purification solution is effectively exhibited, the concentration of nitrate and the like is reduced in the original water environment, and purification in the water environment is more suitably realized.

  The PHA in the particles composed of polyhydroxyalkanoate (PHA) used in the present invention is not particularly limited. For example, 3-hydroxyalkanoate represented by the following formula (1), 4 represented by the following formula (2), and the like. -A polymer having a repeating unit consisting of a hydroxyalkanoate.

(In the formulas (1) and (2), R is H or an alkyl group represented by C _n H _{2n + 1} , and n is an integer of 1 to 15.)

It may be a homopolymer of formula (1), formula (2) or from a copolymer comprising two or more combinations, ie, a di-copolymer, a tri-copolymer, a tetra-copolymer, etc., or their homopolymers, copolymers, etc. The blended material of 2 or more types chosen is mentioned. Among them, poly (3-hydroxybutyrate) (PHB), poly (3-hydroxybutyrate-co-3-hydroxyvalerate (PHBV), poly (3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBH) and poly (3-hydroxybutyrate-co-4-hydroxybutyrate) (P3HB4HB) are preferred, and since R is bulky, the degree of crystallinity is low, and there are many amorphous parts that are susceptible to anaerobic decomposition. In this respect, PHBH is more preferable.
In the present invention, as described above, PHA is assimilated by a group of microorganisms other than autotrophic contained in raw water.

The shape of the particles composed of PHA (hereinafter sometimes referred to as “PHA particles”) is not particularly limited, and examples thereof include powder, granules, pellets, and the like. It can be selected as appropriate depending on the manner of input to the environment.
The size of the particles is not particularly limited, and can be appropriately selected depending on the culture method, the manner in which the water purification solution is introduced into the water environment, and the like.
When particles made of PHA are used in the form of powder, it may be preferable to use a fine powder from the viewpoint of increasing the surface area. However, if the size of the powder is too small, it may be mixed with raw water. The average particle size is preferably 50 μm or more because the powder may rise and workability may deteriorate. Moreover, in the case of such a fine powder, it can be made into the emulsification form and suspension form which were disperse | distributed to water. Thereby, since a fine powder does not rise at the time of mixing with raw | natural water, the average particle diameter of particle | grains can be made smaller than 50 micrometers. Such an emulsified liquid of PHA particles is obtained by refining PHA particles with an end mill or the like using a mixed liquid of powder of 50 μm or more and water such as tap water (preferably not raw water). Can be prepared.
Moreover, when using granular and pellet-shaped particles, it is preferable to use porous particles from the viewpoint of securing the surface area. The size of the granular and pellet-like particles is preferably about 0.5 to 10 mm. Further, as the shape, various structures such as a cube, a rectangular parallelepiped, a cylinder, a sphere, and an ellipsoid can be used.

In the present invention, the raw water and PHA particles are mixed and cultured. The mixing ratio of the raw water and the PHA particles is not particularly limited as long as a biofilm is formed on the surface of the PHA particles of a certain level or more within a desired period when cultured, and can be appropriately determined. Further, depending on the method of applying an external force, the mixing ratio may be such that a shearing force is applied to the PHA particle surface so that at least a part of the biofilm formed on the PHA particle surface is destroyed.
In addition to raw water and PHA particles, inorganic salts and other components that can contribute to the growth of the microorganism group may be added as necessary.

The culture tank used when mixing the raw water and the PHA particles is not particularly limited, and a batch-type culture tank may be used, and raw water is collected (taken in water) continuously or intermittently from the water environment. Further, it may be a circulation type culture tank provided with means capable of being continuously or intermittently charged into the original water environment.
The batch type culture tank is not particularly limited, and examples thereof include a transparent or opaque culture tank that can be sealed, a simple cup and a plastic bottle with a lid, and the like. In addition, as the circulation culture tank, various culture tanks generally used for culturing microorganisms can be used. In the case of a circulation type culture tank, the culture tank includes a water intake port for taking water in the water environment from the water environment to the culture tank, and a drain for discharging the culture solution from the culture tank to the water environment. However, in order to prevent the PHA particles from being discharged from the culture tank, PHA particles such as a filter are present at or near the discharge port from the culture tank to the original water environment. In order to prevent or prevent the clogging or solidification of PHA particles at the discharge port and its vicinity, the discharge port such as a stirring blade should be blocked. It is preferable to provide a means for preventing.

  In the present invention, after starting the culture by mixing the raw water and the PHA particles and before ending the culture, an external force is applied to the culture solution in which the raw water and the PHA particles are mixed to At least part of the biofilm formed on the particle surface is destroyed. As a result, as described above, it is possible to effectively increase the surface of the PHA particles in the culture solution and the number of microorganism groups in the solution.

The external force is not particularly limited as long as at least a part of the biofilm formed on the surface of the PHA particles can be destroyed when loaded on the culture solution. Examples include, but are not limited to, means for stirring the culture medium to impart shearing force to the biofilm, means for vibrating the culture liquid to impart shearing force to the biofilm, and the like. These means can be appropriately selected according to the form of the culture tank.
As a culture tank, for example, when using a PET bottle, the PET bottle is filled with an appropriate amount of culture solution, shaken together with the PET bottle in a tightly sealed state, and the inside culture solution is stirred, so that the PHA particles At least a part of the biofilm formed on the surface of the PHA particles can be broken and peeled off by, for example, collision. Also, if it is larger than a PET bottle and it is difficult to shake the entire culture tank, a stirring blade, baffle plate, etc. are provided in the culture tank to stir the culture solution, or the water in the culture tank is pumped with a pump By causing the PHA particles to collide with each other, at least a part of the biofilm formed on the surface of the PHA particles can be broken and peeled off.
Furthermore, when using the above circulating culture tank, the raw water is taken from the water environment, and at the same time, the flow of the liquid generated when the culture liquid is drained from the culture tank to the original water environment is used. At least a part of the biofilm formed on the surface of the PHA particles can be broken and peeled off by convection of the PHA particles and causing the PHA particles to collide with each other. At the same time, the culture solution may be stirred by providing a stirring blade, a baffle plate, or the like in the culture tank.
In addition, in the case of using any culture tank, the culture solution can be vibrated using ultrasonic waves or the like, and at least a part of the biofilm formed on the PHA particle surface can be broken and peeled off by cavitation.

There is no particular limitation on the timing for applying the external force. For example, after the culture starts, a biofilm is formed on the surface of the PHA particles, and at the stage where almost all of the used PHA particles are coated with the biofilm. It is preferable to apply external force while confirming the biofilm formation on the surface of the PHA particles. When the surface of the PHA particles is coated with a biofilm, the rate of increase in the number of microorganisms in the culture solution tends to be slowed down, so that the biofilm formation state can be an index of the external load period.
In addition to confirming the biofilm formation status, it is also possible to estimate the rate of increase of microbial groups in the culture solution. The increase rate of the microbial group in the culture solution can be estimated by, for example, grasping the change with time of the weight of the microbial group in the culture solution. As a method for this, for example, a sample is collected from the culture solution over time, and a microorganism group (bacteria etc.) is captured using filter paper, etc., and the weight is measured according to a conventional method to grasp the rate of increase. It is possible.
In addition to the method for obtaining the change in weight over time as described above, it is possible to grasp the rate of increase of microbial groups in the culture solution by estimating the number of microbial cells in the culture solution. . The number of cells can be measured by culturing on a medium such as agar and teaching the generated colonies. Moreover, the number of anaerobic microorganisms should just cultivate on anaerobic conditions using a commercially available anaerobic culture pack etc., and should count the number of colonies. In the present invention, since the aim is to reduce nitrate, it is preferable to apply an external force particularly when the rate of increase in the number of cells of anaerobic microorganisms has slowed.
In the present invention, the time when the rate of increase in the number of bacterial cells slows down means that the slope becomes gentle when the relationship between the number of culture days and the number of bacterial cells is plotted on a graph.

  However, the external force may be periodically applied regardless of the formation state of the biofilm.

In the present invention, it is preferable that the external force is loaded once every 1 to 7 days in consideration of the formation state of the biofilm. However, once the biofilm is formed on the surface of the PHA particles, it is experienced. Specifically, it is preferably at least once every 7 days, more preferably once every 3 days, and even more preferably once a day. Although depending on the culture conditions, it is preferable to apply the initial external force for biofilm destruction after the start of the culture after 3 days or more have elapsed since the start of the culture in which the biofilm is appropriately formed. However, depending on the conditions, the growth of the microbial group in the culture solution may be observed even on one day, so an external force may be applied on the first day after the start of the culture.
Further, the number of external force loads is not particularly limited, but from the viewpoint of more effectively purifying water by increasing the number of microorganism groups in the culture solution, the load is applied twice or more during the culture period. preferable.

  The temperature at the time of culture is not particularly limited as long as it is the temperature according to the water environment from which the raw water is collected. More preferred. Considering ease of temperature control, 80 ° C. or lower is preferable. Moreover, it is also possible to carry out at room temperature (room temperature) without performing temperature control.

  The culture period varies depending on various conditions such as the form, size, raw water condition, culture temperature, etc. of the PHA particles, but it cannot be said unconditionally. However, from the viewpoint of appropriately growing the microorganism group in the culture solution, it is 3 days or more. Is preferred. However, depending on the conditions, the growth of the microorganism group in the culture solution may be observed even on one day, so the culture period may be one day.

  Since the water purification solution of the present invention is a culture solution obtained by mixing raw water and PHA particles and performing a predetermined treatment as described above, the culture solution was coated with a biofilm. PHA particles and microbial populations outside the biofilm are included. When this water purification solution is introduced into the original water environment from which raw water is collected, the supernatant of the water purification solution (a solution containing almost no PHA particles) may be used. A liquid containing PHA particles coated with a turbid biofilm may be used. Since PHA is a polymer with high biodegradability, even if it is put into various water environments, it is easily decomposed and disappears in various environments, so it hardly deteriorates the original water environment. Conceivable.

  The water purification method of the present invention comprises mixing raw water containing a group of microorganisms containing anaerobic microorganisms collected from the natural environment or the water environment of an artificial device and particles made of polyhydroxyalkanoate, and starting culture. When the rate of increase in the number of anaerobic microorganisms slows down until the end of the culture, the culture solution obtained by destroying at least part of the biofilm formed on the particle surface by applying an external force At least a part is thrown into the water environment where the raw water is collected.

That is, the water purification method of the present invention is one that inputs (returns) the above-described water purification solution of the present invention (which is also a culture solution) to the original water environment from which the raw water was collected. Therefore, detailed description of common parts is omitted.
In the water purification method of the present invention, the culture solution to be introduced into the original water environment may be added according to the state of the original water environment. For example, a method of periodically spraying a suitable amount of water purification solution prepared in advance using raw water to each location, one or more circulating culture tanks installed in the vicinity of the water environment to be purified, and continuously For example, a method of collecting raw water and adding a culture solution (water purification solution) together with collection of raw water may be mentioned.
In this case, the amount of the culture solution (water purification solution) to be introduced into the original water environment may be determined as appropriate according to the environment. Since there is a possibility that the balance of microorganisms in the original water environment from which the raw water was collected may be lost, it is preferable to add it little by little while measuring the nitrate concentration and confirming the situation.

  Depending on the method of charging the culture solution, the culture solution (water purification solution) prepared in advance may decrease, so raw water may be collected again from the same water environment and added to the culture tank. PHA particles may also be added. And after adding these suitably, it culture | cultivates, providing external force as mentioned above, and it is good to prepare a culture solution (liquid for water purification).

  In particular, when the water purification method of the present invention is applied to the water environment of an aquarium where aquatic organisms are bred, the nitrate concentration of water in the aquarium can be 0 to 50 mg / L.

The water tank reactor of the present invention mainly comprises a culture tank used for preparing the above-described water purification liquid of the present invention and means for loading an external force. In addition, the water environment in which raw water is collected can be used particularly suitably in the case of a water environment in an aquarium where aquatic organisms are bred. Therefore, detailed description of common parts is omitted.
The aquarium reactor of the present invention may be connected to the aquarium so that raw water from the aquarium directly flows into the culture tank, or may be connected via a filter medium system. In the filter medium system, the microorganism group contained in the raw water may be partially captured and maintained in the filter medium part, and the species of the microorganism group supplied to the aquarium reactor may be adjusted. The material of the filter medium is not particularly limited, and a commercially available filter medium can be appropriately selected and used. For example, various fibers, porous stones and the like can be mentioned. In addition, since phosphate may accumulate in the filter medium, phosphorus atom-containing substances such as phosphate may be removed from the water environment in the water tank by periodically replacing the filter medium.

  Below, embodiment of the reactor for water tanks and the water tank provided with the same of the present invention is described based on a drawing.

  FIG. 1 is a view showing an example of an embodiment of a water tank provided with a water tank reactor of the present invention. In the example shown in FIG. 1, the main configuration includes a water tank 10, a water tank reactor 20, and a filter medium system 30. Although not shown, the aquarium may be provided with various devices for raising aquatic organisms such as a ventilation pump and a heater for adjusting the water temperature in the aquarium.

  In this example, the raw water 11 in the water tank 10 is overflowed from the drain port 15 of the drain pipe 12, and the raw water 11 is filtered from the water receiving port 31 provided at the connecting portion between the filtration container 32 and the drain pipe 12 of the filter medium system 30. Water is passed through the material system 30. The raw water 11 reaches the space 35 through the filter medium 34 disposed in the filtration space 33 of the filtration container 32. In this example, the filtration container 32 is divided into a filtration space 33 and a space 35 by a separation plate 37 having a plurality of small holes 38. The filtered raw water 11 ′ in the space 35 is sent from the drain port 36 to the aquarium reactor 20 by the pump 40.

  In the case where the raw water 11 is not filtered, the filter medium 34 may not be provided, or the entire filter medium system 30 (reference numerals 30 to 38) may not be used.

  The culture tank 21 of the aquarium reactor 20 is provided with a water intake pipe 22 extending from the upper part to the lower part of the substantially central part, and the filtered raw water 11 ′ is fed into the culture tank 21 from the water intake 23 at the lower part of the water intake pipe 22. To be liquidated. The culture tank 21 has a cylindrical shape, and an inclined surface 21a is provided at the lower part thereof so that the culture solution 24 and the PHA particles 25 in the culture tank can flow without staying. Then, the filtered raw water 11 ′ flowing from the water intake port 23 arranged at the bottom of the substantially central portion of the culture tank 21 forms an upward flow by the inclined surface 21 a (see arrow 27 in FIG. 1), and the PHA particles are generated by the upward flow. 25 rises particularly at the side wall surface portion of the culture tank 21. The inclined surface 21a may be configured by two or more planes, may be configured by a situation, and can be appropriately determined according to the shape of the culture tank 21 and the like. On the other hand, when the raised PHA particles hit the upper wall surface 21 b of the upper part of the culture tank 21, the PHA particles descend near the intake pipe 22, and a part of the culture solution 24 is drainage provided on the upper side wall of the culture tank 21 to protrude from the side wall. It is discharged from the discharge port 29 a of the part 29 and is put into the water tank 10. Thus, in this example, when the PHA particles in the culture tank 21 described above rise and fall, the PHA particles collide with each other, so that the biofilm formed on the surface of the PHA particles is destroyed. . Although not shown, as a configuration for destroying other biofilms, an ultrasonic transducer may be arranged in the culture tank 21 or ultrasonic waves may be irradiated from the outside of the culture tank 21. .

  Moreover, when adjusting the temperature of a culture solution, you may provide a heater in the culture tank 21, and may arrange | position a jacket etc. outside the culture tank 21. FIG.

  In this example, as described above, the drainage part 29 that protrudes from the side wall and communicates with the inside of the culture tank 21 is provided on the upper side wall of the culture tank 21. In addition, a discharge port 29 a communicating with the water tank 10 and a filter 28 are provided on the upper wall surface of the discharge unit 29. Furthermore, a stirring blade 26 is arranged on the inner side wall portion of the discharge unit 29, and when the stirring blade 26 rotates, a flow toward the inside of the culture tank 21, a flow toward the filter 28, and the like are generated. It is possible to suppress or prevent clogging or hardening of PHA particles in the vicinity (for example, the filter 28).

  The configuration of the filter 28 is not limited as long as the culture solution 24 (water purification solution) can pass through and a small hole or the like that can suppress or prevent the passage of the PHA particles 25 is provided.

  The culture solution 24 (which is also a water purification solution) discharged from the discharge port 29 a of the aquarium reactor 20 is input from the water injection port 14 into the water tank 10 through the water injection pipe 13 of the water tank 10. In this example, the water injection pipe 13 is arranged inside the drain pipe 12 and is configured such that the lower part of the water injection pipe 13 protrudes from an opening provided in the side wall of the water injection pipe 12. It may be adopted.

  With the configuration described above, the pump 40 can be operated to take the raw water in the water tank 10 and culture it in the water tank reactor 20, and a part of the culture solution 24 can be put into the water tank 10. While the pump 40 is being operated, the biofilm is continuously broken in the culture tank 21 of the aquarium reactor 20 due to the formation of the biofilm on the surface of the PHA particles and the flow and collision of the PHA particles. That is, it occurs once or twice or more, and the group of microorganisms in the culture tank 21 is propagated. Even when the pump 40 is stopped, in order to destroy at least a part of the biofilm and to propagate the microorganism group in the culture tank 21, other stirring blades and ultrasonic vibrators (not shown) are used as the culture tank. 21 may be arranged. By adopting such a configuration, the intermittent culture solution 24 can be introduced into the water tank 10.

  When the aquarium reactor 20 is not used, between the drain port 36 of the filter medium system 30 and the intake pipe 22 of the aquarium reactor 20, and between the drain port 29a of the aquarium reactor 20 and the intake pipe 13. In addition, it is preferable that the drain port 36 and the water injection pipe 13 are directly communicated with each other by using the three-way cocks 41 arranged respectively. In the meantime, the culture solution (water purification solution) prepared using the above-described PET bottle may be put into the water tank 10.

  FIG. 2 is a schematic view showing another example of the embodiment of the water tank provided with the water tank reactor of the present invention. As shown in FIG. 2, the present embodiment is different in that six water tanks 10 a to 10 f are used instead of the water tank 10 in FIG. 1. Since the configuration of the filter medium system 30, the aquarium reactor 20, and the connecting portion thereof is substantially the same as the configuration shown in FIG. 1, the detailed description thereof will be omitted, and the same components are denoted by the same reference numerals. It was attached. The configurations of the water tanks 10a to 10f, the drain pipes 12a to 12f, and the water injection pipes 13a to 13f are the same as the configurations of the water tank 10, the drain pipe 12, and the water injection pipe 13 shown in FIG. Description is omitted. In addition, although the example which connected the six water tanks is shown in this example, it can change suitably.

  In the present embodiment, the raw waters 11a to 11f in the water tanks 10a to 10f are all passed through one filter medium system 30 and the water tank reactor 20. Then, the culture solution cultured in the water tank reactor 20 is introduced (returned) to each of the water tanks 10a to 10f as a water purification liquid. In addition, each arrow in FIG. 2 shows the flow direction. This embodiment is suitable when breeding aquatic organisms of the same species in separate aquariums.

  EXAMPLES Hereinafter, the present invention will be described more specifically with reference to examples. However, the present invention is not limited to these examples.

(Method of measuring nitrate concentration)
The water in the water tank was periodically sampled and measured using a “NO3 tester” manufactured by Red Sea.

(Production Example 1) Production of PHBH with 3HH rate of 12 mol% <Production of plasmid vector for gene insertion>
A. DNA for insertion A DNA comprising a base sequence containing a Caviae phaC promoter and a ribosome binding site was prepared as follows. A. PCR was carried out using the Caviae genomic DNA as a template and the primers shown in SEQ ID NO: 1 and SEQ ID NO: 2. PCR was (1) 98 ° C. for 2 minutes, (2) 98 ° C. for 15 seconds, 60 ° C. for 30 seconds, and 68 ° C. for 20 seconds for 25 cycles, and the polymerase used was KOD-plus- (manufactured by TOYOBO). The DNA fragment obtained by PCR was terminally phosphorylated and digested with EcoRI. This DNA fragment was designated as PAc-5P + Eco.
Next, the DNA insertion site is set immediately before the start codon of the bktB gene of the chromosomal DNA of the KNK-005AS strain described in JP 2008-029218 A [0038], and upstream of the start codon of the gene by the following procedure. A DNA consisting of the nucleotide sequence was prepared.
Using the genomic DNA of the KNK-005 strain described in JP-A-2008-029218 [0036] as a template DNA source, PCR was performed using the primers shown in SEQ ID NO: 3 and SEQ ID NO: 4 to initiate the bktB gene. DNA consisting of a base sequence upstream of the codon was obtained. PCR was (1) 98 ° C. for 2 minutes, (2) 98 ° C. for 15 seconds, 64 ° C. for 30 seconds, and 68 ° C. for 30 seconds for 25 cycles, and the polymerase used was KOD-plus-. The DNA fragment obtained by PCR was simultaneously digested with the restriction enzymes BamHI and EcoRI. This DNA fragment was designated as PbktB-Bam + Eco.
PAc-5P + Eco and PbktB-Bam + Eco were ligated, and PCR was performed using the primers shown in SEQ ID NO: 3 and SEQ ID NO: 2 with the DNA produced in the ligation solution as the template DNA. PCR was (1) 98 ° C. for 2 minutes, (2) 98 ° C. for 15 seconds, 60 ° C. for 30 seconds, and 68 ° C. for 50 seconds for 25 cycles, and the polymerase used was KOD-plus-. The DNA fragment obtained by PCR was terminally phosphorylated and digested with BamHI. This DNA fragment was designated as bPac-5P + Bam.
Next, a DNA having a base sequence downstream from the initiation codon of the gene was prepared. PCR was performed using the genomic DNA of the KNK-005 strain as a template DNA source and the primers shown in SEQ ID NO: 5 and SEQ ID NO: 6 to obtain a DNA comprising a base sequence downstream from the start codon of the bktB gene. . PCR was (1) 98 ° C. for 2 minutes, (2) 98 ° C. for 15 seconds, 64 ° C. for 30 seconds, and 68 ° C. for 30 seconds for 25 cycles, and the polymerase used was KOD-plus-. The DNA fragment obtained by PCR was terminally phosphorylated and digested with ClaI. This DNA fragment was designated ORF-5P + Cla.
bPac-5P + Bam and ORF-5P + Cla were ligated, and PCR was performed using the primers shown in SEQ ID NO: 3 and SEQ ID NO: 6 with the DNA generated in the ligation solution as the template DNA. For PCR, (1) 98 ° C. for 2 minutes, (2) 98 ° C. for 15 seconds, 60 ° C. for 30 seconds, and 68 ° C. for 1 minute 30 seconds were repeated 25 cycles, and the polymerase used was KOD-plus-. The DNA fragment obtained by PCR was co-digested with BamHI and ClaI. This DNA fragment was subcloned into a site digested with the same restriction enzyme of the vector pBluescript II KS (-) (manufactured by TOYOBO). The obtained vector was designated as bAO / pBlu. The base sequence was determined using a DNA sequencer 3130xl Genetic Analyzer manufactured by APPLIED BIOSSYSTEMS, and it was confirmed that it was the same as the base sequence of the template DNA.
Subsequently, the pSACKm described in JP2008-029218A [0037] was treated with the restriction enzyme NotI to cut out a DNA fragment of about 5.7 kb containing the kanamycin resistance gene and the sacB gene. This was inserted into a site cleaved with the same enzyme of bAO / pBlu to prepare a plasmid bAO / pBlu / SacB-Km for gene disruption / insertion.

<Preparation of gene insertion strain Pac-bktB / AS strain>
Next, using KNK-005AS as a parent strain, a strain was prepared using bAO / pBlu / SacB-Km in which a DNA comprising a phaC promoter and a base sequence containing a ribosome binding site was inserted immediately before the start codon of the bktB gene. E. coli strain S17-1 (ATCC 47005) was transformed with the gene insertion plasmid bAO / pBlu / SacB-Km. The obtained transformant was mixed and cultured on KNK-005AS strain and Nutrient Agar medium (manufactured by Difco) to perform conjugation transfer. Simmons agar medium containing 250 mg / L kanamycin (sodium citrate 2 g / L, sodium chloride 5 g / L, magnesium sulfate heptahydrate 0.2 g / L, ammonium dihydrogen phosphate 1 g / L, dihydrogen phosphate 2 A strain that had grown on potassium 1 g / L, agar 15 g / L, pH 6.8) was selected to obtain a strain in which the plasmid was integrated on the chromosome of the KNK-005AS strain. This strain is cultured for 2 generations in Nutrient Broth medium (manufactured by Difco), then diluted and applied to Nutrient Agar medium containing 15% sucrose, and the grown strain is selected to perform the second recombination. The resulting stock was acquired. Furthermore, a desired gene insertion strain was isolated by PCR analysis.
This gene insertion strain was named Pac-bktB / AS strain, its nucleotide sequence was determined using a DNA sequencer 3130xl Genetic Analyzer, and DNA comprising a phaC promoter and a ribosome binding site immediately before the start codon of the bktB gene Was confirmed to be an inserted strain.

<Cloning of ccr gene and expression unit construction>
The ccr gene encoding an enzyme that converts crotonyl-CoA to butyryl-CoA, a precursor of 3HH monomer, was cloned from the chromosomal DNA of Streptomyces cinnamonensis Okami strain (DSM 1042). PCR was performed using the DNAs of SEQ ID NO: 7 and SEQ ID NO: 8 as primers. The conditions were (1) 98 ° C. for 2 minutes, (2) 94 ° C. for 10 seconds, 55 ° C. for 20 seconds and 68 ° C. for 90 seconds for 25 cycles, and the polymerase used was KOD-plus-. The fragment amplified by PCR was purified and then cleaved with restriction enzymes BamHI and AflII. Subcloning the EE32d13 fragment (J. Bacteriol., 179, 4821 (1997)) into the EcoRI site of the pUC19 vector, cutting this plasmid with BglII and AflII, and replacing the ccr gene with the BamHI and AflII fragments. The ccr expression unit was constructed by

<Cloning of phaC gene and preparation of expression unit>
A phaC expression unit containing SEQ ID NO: 11 was prepared as a SpeI fragment. PCR was performed using HG :: PRe-N149S / D171G-T / pBlu described in JP-A-2007-228894 [0031] as a template and primers shown in SEQ ID NO: 9 and SEQ ID NO: 10. The conditions were (1) 98 ° C. for 2 minutes, (2) 94 ° C. for 10 seconds, 55 ° C. for 30 seconds and 68 ° C. for 2 minutes for 25 cycles, and the polymerase used was KOD-plus-. The fragment amplified by PCR was purified and then cleaved with the restriction enzyme SpeI to prepare an expression unit.
The expression vector pCUP2-631 was constructed as follows.
C. As a plasmid vector for constructing an expression vector in necator, pCUP2 described in International Publication WO / 2007/049716 [0041] was used. First, the ccr gene expression unit constructed in Example 7 was excised by EcoRI treatment, and this fragment was ligated with pCUP2 cleaved with MunI. Next, the phaC expression unit prepared in Example 8 was prepared as a SpeI fragment and inserted into the SpeI site of pCUP2 containing the ccr gene expression unit to construct a pCUP2-631 vector.

<Production of transformant>
The introduction of the pCUP2-631 vector into various cells was carried out by electrical introduction as follows. The gene introduction apparatus used was a gene pulser manufactured by Biorad, and the cuvette used was a gap 0.2 cm manufactured by Biorad. 400 μl of competent cells and 20 μl of expression vector were injected into a cuvette and set in a pulse device, and an electric pulse was applied under the conditions of electrostatic capacity 25 μF, voltage 1.5 kV, and resistance value 800Ω. After the pulse, the bacterial solution in the cuvette was cultured with shaking on Nutrient Broth medium (manufactured by DIFCO) at 30 ° C. for 3 hours, and then on a selection plate (NutrientAgar medium (manufactured by DIFCO), kanamycin 100 mg / L) at 30 ° C. The cultured transformants were cultured for a day to obtain transformed transformants.

<Culture of transformant>
The transformant prepared above was cultured. The composition of the pre-medium is 1% (w / v) Meat-extract, 1% (w / v) Bacto-Trypton, 0.2% (w / v) Yeast-extract, 0.9% (w / v) Na _The pH was adjusted to 6.7 with ₂ HPO ₄ · 12H ₂ O and 0.15% (w / v) KH ₂ PO ₄ .
The composition of the polyester production medium is 1.1% (w / v) Na ₂ HPO ₄ · 12H ₂ O, 0.19% (w / v) KH ₂ PO ₄ , 0.6% (w / v) (NH ₄ ) ₂ SO ₄ , 0.1% (w / v) MgSO ₄ .7H ₂ O, 0.5% (v / v) trace metal salt solution (1.6% (w / v) FeCl in 0.1N hydrochloric acid) ₃ · 6H ₂ O, 1% (w / v) CaCl ₂ · 2H ₂ O, 0.02% (w / v) CoCl ₂ · 6H ₂ O, 0.016% (w / v) CuSO ₄ · 5H ₂ O, 0.012% (w / v) NiCl ₂ .6H ₂ O, and 0.01% (w / v) CrCl ₃ .6H ₂ O were dissolved. This was carried out by fed-batch culture in which PKOO (Palm kernel olein, palm kernel oil olein fraction) was fed as a carbon source.
Glycerol stock of each transformant was inoculated into the pre-culture medium and cultured for 20 hours, and 10% (v / V) Inoculated. The operating conditions were a culture temperature of 28 ° C., a stirring speed of 420 rpm, an aeration rate of 0.6 vvm, and a pH controlled between 6.6 and 6.8. For control, 14% aqueous ammonia was used. Incubation was carried out for up to 65 hours. After incubation, the cells were collected by centrifugation, washed with methanol, and lyophilized to obtain PHBH.
The 3HH composition analysis of the produced polyester was measured by gas chromatography as follows. To about 20 mg of dry PHBH, 2 ml of sulfuric acid-methanol mixture (15:85) and 2 ml of chloroform were added and sealed, and heated at 100 ° C. for 140 minutes to obtain methyl ester of PHBH decomposition product. After cooling, 1.5 g of sodium bicarbonate was added little by little to neutralize it, and the mixture was allowed to stand until the generation of carbon dioxide gas stopped. After adding 4 ml of diisopropyl ether and mixing well, the mixture was centrifuged and the monomer unit composition of the PHBH degradation product in the supernatant was analyzed by capillary gas chromatography. The gas chromatograph used was Shimadzu Corporation GC-17A, and the capillary column used was GL Science's NEUTRA BOND-1 (column length 25 m, column inner diameter 0.25 mm, liquid film thickness 0.4 μm). He was used as the carrier gas, the column inlet pressure was set to 100 kPa, and 1 μl of the sample was injected. As temperature conditions, the temperature was raised from an initial temperature of 100 to 200 ° C. at a rate of 8 ° C./min, and further from 200 to 290 ° C. at a rate of 30 ° C./min. As a result of analysis under the above conditions, it was PHBH having a 3HH ratio of 12%. The weight average molecular weight was about 660,000, and Tg was 0 ° C.

Example 1
About 500 g of PHBH powder (average particle size 150 μm) prepared in Production Example 1 and about 400 mL of raw water collected from a water tank (water volume: 400 L) rearing corals were placed in a 500 ml PET bottle. This time was set as the start of culture. The PET bottle was capped, and the bottle was shaken strongly for about 5 seconds and stirred. Then, it was allowed to stand at room temperature with the lid closed. Thereafter, the mixture was shaken vigorously for about 5 seconds once a day.
Before returning the culture solution to the original water tank, the nitrate concentration of water in the water tank was measured by the above method, and the amount of the culture liquid (water purification solution) to be introduced into the water tank was set from the measured value. When the nitrate concentration is 50 mg / L or more, the culture solution is 10 mL with respect to 100 L of water in the aquarium, 4 mL with respect to 100 L when 25 mg / L or more and less than 50 mg / L, and 1 mL with respect to 100 L when less than 25 mg / L. .
The first nitrate concentration measurement was performed 72 hours after the start of the culture, which was defined as the first day, and the culture solution was returned immediately after the measurement. Thereafter, the culture solution was added on the 2nd day, 3rd day, 4th day, 7th day, 10th day, 11th day, 12th day, 13th day, and 20th day. The results of measuring the nitrate concentration over time are shown in FIG.
The water tank (1500 mm × 500 mm × 600 mm) has a water volume of 400 L, circulates water by overflow, and is a natural system that does not use a filter medium.

(Example 2)
In the same manner as in Example 1 except that a 200 L water tank (1100 mm × 400 mm × 450 mm) rearing the sago was used as the water environment, the culture and the culture solution (water purification solution) for the water tank were used. The nitrate concentration was measured. The results of measuring the nitrate concentration over time are shown in FIG.

(Example 3)
In the same manner as in Example 1, except that the water tank (1200 mm × 450 mm × 450 mm) with a water volume of breeding goby was used as the water environment, and the water in the water tank was circulated using the filter medium system, The culture solution (water purification solution) was added to the culture and the water tank, and the nitrate concentration was measured. The results of measuring the nitrate concentration over time are shown in FIG.

(Example 4)
Aquariums that are breeding ornamental fish and connected to each other through one filter media system (the size of each tank is 690 mm × 400 mm × 400 mm and the total water volume is 700 L) are used as the water environment. Except for this, in the same manner as in Example 1, the culture and the culture solution (water purification solution) were added to the water tank, and the nitrate concentration in each water tank was measured. The time-lapse measurement result of the nitrate concentration (average value of the measured values in each water tank) is shown in FIG. The configuration of the six connected water tanks and the filter medium system are the same as those in FIG.

(Example 5)
An aquarium that keeps ornamental fish and that is connected to four units through one filter media system (the size of each tank is 690 mm × 400 mm × 400 mm, and the total amount of water is 700 L) is used as the water environment. Except for this, in the same manner as in Example 1, the culture and the culture solution (water purification solution) were added to the water tank, and the nitrate concentration in each water tank was measured. FIG. 3A shows the results of measuring the nitrate concentration (average value of the measured values in each water tank) over time. The configuration of the four connected water tanks and the filter medium system are the same as those in FIG.

From the results of Examples 1 to 5 shown in FIGS. 3A and 3B, it can be seen that the nitrate concentration can be controlled at a low level.
It should be noted that immediately after the culture solution (water purification solution) was put into the water tank, the concentration of nitrate in the water tank sometimes increased (for example, in FIG. 3 (a), on the 20th day of Example 5). And the 21st day.), And decreased after a certain period. The cause of this is not clear, but it is presumed that nitrate increased temporarily in the process of shifting the balance of various microorganisms present in the water environment to a more desirable balance.

(Comparative Example 1)
The nitrate concentration was measured in the same manner as in Example 3 except that the culture solution was not added to the water tank. The results of measuring the nitrate concentration over time are shown in FIG. From the fifth day onward, it was always a high value exceeding 250 mg / L. In the figure, it is omitted after the fifth day.

(Comparative Example 2)
Except that the culture solution was never stirred, the culture solution and the culture solution (water purification solution) were charged into the water tank in the same manner as in Example 3, and the nitrate concentration was measured. The results of measuring the nitrate concentration over time are shown in FIG. From the 14th day onward, the value was always higher than 250 mg / L. In the figure, it is omitted after the 14th day.

10, 10a to 10f Water tank 11, 11a to 11f Raw water 11 'Raw filtered water 12, 12a to 12f Drain pipe 13, 13a to 13f Water pipe 14 Water inlet 15 Water outlet 20 Tank reactor 21 Culture tank 21a Inclined surface 21b Upper wall surface 22 Water intake tube 23 Water intake 24 Culture solution (water purification solution)
25 PHA particle 26 Stirring blade 27 Arrow 28 Filter 29 Drainage part 29a Drainage port 30 Filtering material system 31 Receiving port 32 Filtration container 33 Filtration space 34 Filtering material 35 Space 36 Drainage port 37 Separation plate 38 Small hole 40 Pump 41 Three-way cock

Claims (10)

  For the culture solution containing raw water containing a group of microorganisms including anaerobic microorganisms and particles made of polyhydroxyalkanoate, an external force was applied between the start and end of the culture, and the particles were formed on the surface of the particles. A water purification solution that destroys at least part of a biofilm.   The water purification solution according to claim 1, wherein the external force is loaded when the rate of increase in the number of cells of the anaerobic microorganisms slows down.   The water purification liquid according to claim 1 or 2, wherein the external force is loaded by stirring and / or vibration.   Anaerobic between the start of culture after mixing raw water containing microorganisms including anaerobic microorganisms collected from the natural environment or the water environment of artificial devices and particles of polyhydroxyalkanoate until the end of the culture When the increase rate of the number of bacterial cells of the sex microorganism has slowed, at least a part of the culture solution obtained by destroying at least a part of the biofilm formed on the particle surface by applying an external force is used as the raw water. A water purification method to be introduced into the collected water environment.   The water purification method according to claim 4, wherein the external force is applied twice or more between the start of culture and the end of culture.   The water purification method according to claim 4 or 5, wherein the polyhydroxyalkanoate is poly (3-hydroxybutyrate-co-3-hydroxyhexanoate).   The water environment of the artificial device is a water environment including water in an aquarium where aquatic organisms are bred, and the nitrate concentration of water in the aquarium after the culture solution is introduced is 0 to 50 mg / L. The water purification method according to any one of claims 4 to 6. A water tank reactor for taking water in an aquarium where aquatic organisms are bred and injecting at least part of a culture solution obtained by culturing using the taken water into the water tank,
A culture tank filled with a culture solution containing the water taken up and particles comprising polyhydroxyalkanoate;
An outlet for discharging the culture solution from the culture tank to the water tank;
Means for applying an external force for breaking at least a part of the biofilm formed on the surface of the particles comprising the polyhydroxyalkanoate;
Aquarium reactor equipped with.   The aquarium reactor according to claim 8, further comprising means for preventing the discharge port from being clogged in the vicinity of the discharge port. A water tank comprising the water tank reactor according to claim 8.

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