Methods of cultivating and producing composite algae as well as composite algae compositions

Abstract

To provide efficient methods of cultivating and producing algae, and to provide algae compositions.SOLUTION: An aspect of the present invention relates to a method of cultivating algae comprising a preparation step of preparing a culture solution containing a first algae and a second algae, and a combined culture step of culturing the first algae and the second algae in the culture solution. In the method of cultivating algae, when the first algae and the second algae are each individually cultivated under the same conditions that allow the first algae and the second algae to grow, the cell growth rate of the first algae after 6 hours from the start of the cultivation is 20% or more higher than the cell growth rate of the second algae after 6 hours from the start of the cultivation.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to methods for cultivating and producing algae, as well as composite algal compositions. [Background technology]

[0002] In recent years, biofuels have been attracting attention as an alternative to exhaustible resources such as petroleum. Examples of biofuels include bioethanol obtained by alcoholic fermentation of raw materials such as sugarcane and corn, and biodiesel obtained by processing vegetable oils such as rapeseed oil and soybean oil. However, since the raw materials for these biofuels are also used as food, there has been a problem in securing the raw materials.

[0003] Therefore, algae have been attracting attention as one of the biomass resources that do not compete with food. In addition, algae can be used not only as biofuel, but also contribute to the removal of microplastics in water (Patent Document 1) and carbon dioxide fixation (Patent Document 2). As an algae cultivation technique, for example, Patent Document 3 discloses an agitation system that can be used for large-scale algae production. [Prior art documents] [Patent documents]

[0004] [Patent Document 1] Patent No. 7099766 [Patent Document 2] Patent Publication No. 2022-061957 [Patent Document 3] Patent Publication No. 2022-511195 Summary of the Invention [Problem to be solved by the invention]

[0005] However, conventional methods for culturing algae have problems in that algae grow slowly and are vulnerable to environmental changes, etc. An object of the present invention is to provide an efficient method for culturing and producing algae, and an algae composition. [Means for solving the problem]

[0006] The present inventors have completed the present invention by combining specific algae.

[0007] That is, according to the first aspect of the present invention, A preparation step of preparing a culture solution containing a first algae and a second algae; a composite culture step of culturing the first algae and the second algae in the culture solution; Equipped with The present invention provides a method for culturing algae, in which, when the first algae and the second algae are cultured separately under the same conditions allowing their proliferation, the cell growth rate of the first algae after 6 hours from the start of culture is 20% or more higher than the cell growth rate of the second algae after 6 hours from the start of culture.

[0008] According to a second aspect of the present invention, A preparation step of preparing a culture solution containing a first algae and a second algae; a composite culture step of culturing the first algae and the second algae in the culture solution; Equipped with The present invention provides a method for culturing algae, in which, when the first algae and the second algae are cultured separately under the same conditions allowing their proliferation, the cell number of the first algae at the time when the cell number of the second algae reaches 150% of the cell number at the start of the culture is 200% or more of the cell number at the start of the culture.

[0009] According to a third aspect of the present invention, A preparation step of preparing a culture solution containing a first algae and a second algae; a composite culture step of culturing the first algae and the second algae in the culture solution; Equipped with The present invention provides a method for culturing algae, in which, when the first algae and the second algae are cultured individually under the same conditions allowing their proliferation, the number of cells of the first algae is greater than the number of cells of the second algae at the beginning of the culture, and after a predetermined time has elapsed, the number of cells of the first algae is less than the number of cells of the second algae.

[0010] According to a fourth aspect of the present invention, A preparation step of preparing a culture solution containing a first algae and a second algae; a composite culture step of culturing the first algae and the second algae in the culture solution; Equipped with The present invention provides a method for culturing algae, in which, when the first algae and the second algae are cultured individually under the same conditions allowing their proliferation, the number of cells in the stationary phase of the first algae is smaller than the number of cells in the stationary phase of the second algae.

[0011] In the first, second, third and fourth aspects, the same conditions may include the algae being in a logarithmic growth phase immediately before culturing.

[0012] In the first, second, third and fourth aspects, the identical conditions may include conditions in which the initial cell number, medium composition, culture temperature, aeration amount and light amount are substantially the same.

[0013] In the first, second, third and fourth aspects, the first algae may secrete a substance necessary for the growth of the second algae.

[0014] In the first, second, third and fourth aspects, the culture method may be included in a method for producing algae.

[0015] According to a fifth aspect of the present invention, a first algae and a second algae; An algae composition is provided in which, when the first algae and the second algae are cultured separately under the same conditions allowing them to grow, the cell growth rate of the first algae after 6 hours from the start of culture is 20% or more higher than the cell growth rate of the second algae after 6 hours from the start of culture.

[0016] According to a sixth aspect of the present invention, a first algae and a second algae; The present invention provides an algae composition in which, when the first algae and the second algae are cultured separately under the same conditions allowing them to grow, the cell number of the first algae at the time when the cell number of the second algae reaches 150% of the cell number at the start of culture is 200% or more of the cell number at the start of culture.

[0017] According to a seventh aspect of the present invention, a first algae and a second algae; An algae composition is provided in which, when the first algae and the second algae are cultured separately under the same conditions allowing their proliferation, the number of cells of the first algae is greater than the number of cells of the second algae at the beginning of the culture, and after a predetermined time has elapsed, the number of cells of the first algae is less than the number of cells of the second algae.

[0018] According to an eighth aspect of the present invention, a first algae and a second algae; An algal composition is provided in which, when the first algae and the second algae are cultured individually under the same conditions allowing their growth, the number of cells in the stationary phase of the first algae is smaller than the number of cells in the stationary phase of the second algae. Effect of the Invention

[0019] SUMMARY OF THE DISCLOSURE The present invention provides efficient methods for culturing and producing algae, as well as algal compositions. [Brief description of the drawings]

[0020] [Figure 1]FIG. 1 is a graph showing the cell density at each predetermined time when each algae was cultured using a synthetic medium. [Diagram 2] FIG. 1 is a graph showing the cell density at each time point when each algae was cultured using a low-nutrient medium. [Diagram 3] FIG. 1 is a graph showing the cell density at each specified time when various algae are cultured using dechlorinated tap water. [Figure 4] FIG. 13 is a diagram showing the cell density at each predetermined time when each algae is cultured under conditions in which the culture environment is changed. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0021] The present invention will be described in detail below. Hereinafter, when multiple upper limit values ​​and multiple lower limit values ​​are listed separately, it is assumed that all numerical ranges that can be set by freely combining these upper limit values ​​and lower limit values ​​are listed. In the following, unless otherwise specified, it is assumed that various measurements were performed at room temperature (for example, 25° C.).

[0022] Hereinafter, the algae according to this embodiment will be described, and then the method for culturing the algae according to this embodiment will be described.

[0023] (algae) The algae in this embodiment are not particularly limited, but are typically microalgae. In this specification, algae include algae that exhibit colonial properties due to adhesion between cells. Examples of types of algae in this embodiment include diatoms, green algae, xanthophyte algae, red algae, brown algae, dinoflagellates, raphidophytes, euglenids, glaucophytes, ophthalmophytes, pinguiophytes, chlorarachniophytes, haptophytes, and cryptophytes.

[0024] The size of the algae in this embodiment is not particularly limited. The size of the algae is typically 0.1 μm or more and 1000 μm or less. In this specification, the "size of the algae" refers to the maximum diameter part of the algae cell (for example, the long diameter part in the case of rod-shaped algae), and specifically refers to the maximum diameter average value of 100 cells randomly obtained.

[0025] The algae in this embodiment is preferably capable of producing a useful substance. Specifically, the algae in this embodiment is preferably capable of producing ethanol, a hydrocarbon, a lipid, or an adhesive substance (adhesive polysaccharide). Ethanol, a hydrocarbon, and a lipid can be used, for example, as biofuel. The adhesive substance can remove, for example, microplastics in water. Examples of algae capable of producing adhesive substances include diatoms belonging to Stramenopile, green algae, red algae, and zygophytes belonging to Archaeplastida, dinoflagellates belonging to Alveolata, euglenids belonging to Excavata, and chlorarachniophytes belonging to Rhizaria.

[0026] In this embodiment, a composite culture in which multiple types of algae are cultured, including a first algae and a second algae, is preferred. Regarding the first algae and the second algae, when the first algae and the second algae are cultured independently under the same conditions allowing the first algae and the second algae to grow, it is preferable that any one of the following, preferably two or more, more preferably three or more, and particularly preferably all of the following apply: (1) the cell growth rate of the first algae after 6 hours from the start of the culture is 20% or more higher than the cell growth rate of the second algae after 6 hours from the start of the culture; (2) the cell number of the first algae at the time when the cell number of the second algae is 150% of the cell number at the start of the culture is 200% or more of the cell number at the start of the culture; (3) the cell number of the first algae is greater than the cell number of the second algae at the beginning of the culture, and after a predetermined time has elapsed, the cell number of the first algae is less than the cell number of the second algae; and (4) the cell number of the first algae in the stationary phase is less than the cell number of the second algae in the stationary phase.

[0027] In this embodiment, "the same conditions capable of growing the first algae and the second algae" refers to any conventionally known conditions that can be set as long as the target algae can grow.

[0028] In this embodiment, "same conditions" preferably includes the algae being in the logarithmic growth phase immediately before culture. In this specification, "logarithmic growth phase" refers to the period during which cells are growing, excluding the lag phase, stationary phase, and death phase, and does not necessarily mean that the cells are growing logarithmically in the strict sense. Specifically, for example, when the following formula 1 is satisfied, it can be determined that the algae are in the logarithmic growth phase. In formula 1, N x is the elapsed time t x represents the total number of cells in t2>t1, and A is an arbitrary constant A>0.

[0029]

number

[0030] In addition, algae in the logarithmic growth phase can be obtained by culturing algae after freezing and thawing or after subculture for a certain period of time under conditions that allow the algae to grow. Specifically, for example, algae can be obtained by culturing algae for one day or more after freezing and thawing or after subculture. In this case, it is preferable that the algae are not saturated in the culture solution. Specifically, for example, when the algae are in the culture solution for 10 10 It is preferable that the concentration is less than cells / mL.

[0031] In this embodiment, "same conditions" preferably includes that the initial cell numbers are approximately the same. Furthermore, in this embodiment, "same conditions" more preferably includes that the initial cell numbers are approximately the same. Note that "approximately the same initial cell numbers" refers to the number of (seeded) cells at the start of culture having an error of ±10%.

[0032] In this embodiment, "same conditions" preferably includes that the medium composition is substantially the same. In addition, in this embodiment, "same conditions" more preferably includes that the medium composition is substantially the same. In addition, "substantially the same medium composition" means that the content of each medium component includes an error of ±10%. In this embodiment, the medium may be, for example, purified water, synthetic medium such as AF-6 medium, BN medium, C medium, HUT medium, MA medium, and VT medium, enriched medium such as ES medium, PES medium, PESI medium, ESS medium, and f / 2 medium, and modified medium thereof, or seawater, freshwater, brackish water, etc. in which algae live in nature.

[0033] In this embodiment, "same conditions" preferably includes that the culture temperature, aeration amount, or light amount is approximately the same. Furthermore, in this embodiment, "same conditions" more preferably includes that the culture temperature, aeration amount, and light amount are approximately the same, and even more preferably includes that the culture temperature, aeration amount, and light amount are approximately the same. Note that "approximately the same culture temperature, aeration amount, or light amount" refers to the value of each parameter during the entire culture period having an error of ±10%.

[0034] In this embodiment, the first algae preferably has a higher growth rate in the early stage of culture than the second algae. Specifically, the cell growth rate of the first algae after 6 hours from the start of culture is preferably 20% or more higher than the cell growth rate of the second algae after 6 hours from the start of culture. When such algae are used in a composite culture, the second algae can efficiently utilize substances secreted during growth of the first algae.

[0035] In this embodiment, the second algae preferably has a longer induction period (lag period) than the first algae. Specifically, it is preferable that the number of cells of the first algae is 200% or more of the number of cells at the start of culture at the time when the number of cells of the second algae reaches 150% of the number at the start of culture. When such algae are used for composite culture, the substances secreted during the growth of the first algae can be efficiently utilized during the growth of the second algae.

[0036] In this embodiment, the second algae preferably has a higher growth rate in the logarithmic growth phase than the first algae. Specifically, it is preferable that the number of cells of the first algae is greater than the number of cells of the second algae at the beginning of the culture, and that the number of cells of the first algae is smaller than the number of cells of the second algae after a predetermined time has elapsed. By using such algae in a composite culture, the algae can be cultured efficiently.

[0037] In this embodiment, the number of cells in the stationary phase of the first algae is preferably smaller than the number of cells in the stationary phase of the second algae. In this specification, the term "stationary phase" refers to a state in which cells increase and then cell growth almost stops. When such algae are used in a composite culture, the algae can be efficiently cultured.

[0038] In this embodiment, the first algae preferably secretes a substance necessary for the growth of the second algae. Examples of substances necessary for the growth of algae include vitamins such as thiamine, minerals such as Fe, Mn, Co, and plant hormones such as indole acetic acid, gibberellin, and abscisic acid. By subjecting such algae to a co-cultivation, the second algae can be efficiently cultured. In addition, it is preferable that the first algae according to this embodiment is highly autotrophic and the second algae is highly heterotrophic.

[0039] In this embodiment, in addition to the first algae and the second algae, a further algae (a third algae) may be included. In one aspect of this embodiment, it is preferable that the third algae has similar properties to the first algae described above. Specifically, when the second algae and the third algae are cultured independently under the same conditions allowing them to grow, it is preferable that any one of the following, preferably two or more, more preferably three or more, even more preferably four or more, and particularly preferably all of the following apply: (1) the cell growth rate of the third algae after 6 hours from the start of the culture is 20% or more higher than the cell growth rate of the second algae after 6 hours from the start of the culture; (2) the cell number of the third algae at the time when the cell number of the second algae is 150% of the cell number at the start of the culture is 200% or more of the cell number at the start of the culture; (3) the cell number of the third algae is greater than the cell number of the second algae at the beginning of the culture, and after a predetermined time has elapsed, the cell number of the third algae is less than the cell number of the second algae; and (4) the stationary phase cell number of the third algae is less than the stationary phase cell number of the second algae; and (5) the third algae secretes a substance necessary for the growth of the second algae. When such algae are used in a composite culture, when the second algae grows, substances secreted by the first algae and the third algae during their growth can be efficiently utilized, and the algae can be efficiently cultured.

[0040] In another aspect of this embodiment, the third algae preferably has similar properties to those of the second algae when the above-mentioned second algae is the first algae. Specifically, when the second algae and the third algae are cultured independently under the same conditions allowing their proliferation, it is preferable that any one of the following, preferably two or more, more preferably three or more, even more preferably four or more, and particularly preferably all of the following apply: (1) the cell growth rate of the second algae after 6 hours from the start of the culture is 20% or more higher than the cell growth rate of the third algae after 6 hours from the start of the culture; (2) the cell number of the second algae at the time when the cell number of the third algae is 150% of the cell number at the start of the culture is 200% or more of the cell number at the start of the culture; (3) the cell number of the second algae is greater than the cell number of the third algae at the beginning of the culture, and after a predetermined time has elapsed, the cell number of the second algae is less than the cell number of the third algae; and (4) the stationary phase cell number of the second algae is less than the stationary phase cell number of the third algae; and (5) the second algae secretes a substance necessary for the proliferation of the third algae. When such algae are used in a composite culture, when the third algae grows, the substances secreted by the first algae and the second algae during their growth can be efficiently utilized, and the algae can be efficiently cultured.

[0041] In this embodiment, the number of types of algae is not particularly limited as long as it is 2 or more. When the number of types of algae in this embodiment is n (n≧3), any two types of algae among them correspond to any one, preferably two or more, more preferably three or more, even more preferably four or more, and particularly preferably all, of the above (1) to (5).

[0042] In one aspect of this embodiment, the photosynthetic bacteria can be the first algae or the second algae. Examples of photosynthetic bacteria include cyanobacteria (blue-green algae), red bacteria, and green bacteria. When growing, the photosynthetic bacteria can secrete substances necessary for the growth of algae, such as vitamins, minerals, and plant hormones. By co-culturing such photosynthetic bacteria and algae, the algae can be efficiently cultured.

[0043] Specific examples of the combination of the first algae and the second algae according to this embodiment include Chlorella or Chlamydomonas as the first algae and Microcystis or Anabaena as the second algae. Specific examples of the combination of the first algae and the second algae according to this embodiment include Rhodobacteria as the first algae and Skeletonema or Chaetoceros as the second algae.

[0044] (Method of culturing algae) The algae cultivation method in this embodiment preferably includes a preparation step of preparing a culture solution containing a first algae and a second algae, and a composite cultivation step of culturing the first algae and the second algae in the culture solution.

[0045] In the preparation step in this embodiment, it is preferable to prepare a culture solution containing a first algae and a second algae. The first algae and the second algae before the preparation step may be in a state of being frozen or dried (by a drying method that allows the algae to survive, such as hot air drying, freeze drying, vacuum drying, etc.) independently or mixed together, or may be in a state of being cultured in separate culture tanks, or may be in a state of being cultured mixed together. In addition, when the first algae and the second algae are stored or cultured independently, the second algae may be added to a culture solution containing the first algae, or conversely, the first algae may be added to a culture solution containing the second algae, or the first algae and the second algae may be added to a separate culture tank approximately at the same time. The first algae and the second algae just before the composite culture step described later may be in a dormant state, induction phase, or stationary phase, etc., but are preferably in a logarithmic growth phase. By using such algae, the algae can be rapidly grown after the start of culture.

[0046] The medium in this embodiment is not particularly limited as long as it can grow algae. The medium may be, for example, purified water, synthetic medium such as AF-6 medium, BN medium, C medium, HUT medium, MA medium, and VT medium, enriched medium such as ES medium, PES medium, PESI medium, ESS medium, and f / 2 medium, and modified medium thereof, or seawater, fresh water, brackish water, etc. in which algae live in nature. In addition, as long as algae can grow, industrial water, industrial wastewater, sewage treatment water, domestic wastewater, agricultural wastewater, etc. may be used, and specific examples include wastewater from a garbage disposal plant and wastewater from a power plant.

[0047] In the composite culture step of the present embodiment, it is preferable that the first algae and the second algae are cultured in a culture solution. By culturing multiple types of algae having the above-mentioned relationship together (also called composite culture or co-culture), the algae can be efficiently cultured.

[0048] In the composite culture step of this embodiment, a culture tank is typically used. The culture tank according to this embodiment may be an open system (for example, an outdoor rice field, a reservoir, a lake, a marine aquaculture farm, etc.) or a closed system. Artificial algae culture is often performed in a closed system, but the culture method according to this embodiment is highly robust against changes in the culture environment, so the culture tank may be an open system and can be appropriately selected depending on the application.

[0049] The culture conditions in the composite culture step in this embodiment are not particularly limited. Specifically, as long as the algae can be cultured, the culture temperature, the amount of ventilation, and the amount of light can be set arbitrarily. Note that "algae can be cultured" refers to the algae being viable, and includes the case where the number of cells temporarily decreases as long as the algae are not completely killed.

[0050] The culture method according to the present embodiment is highly robust against changes in the culture environment. The term "change in the culture environment" refers to an environmental change that may reduce the number of algae cells in the system due to, for example, abnormal temperature, abnormal ventilation, abnormal light, contamination, etc. Specific examples of environmental changes include bad weather and the introduction of water with deteriorated water quality, such as sewage treatment water or industrial wastewater, into the culture tank. In the case of such an environmental change, the number of algae cells in the system may temporarily decrease in the present embodiment, but they are less likely to die than in the case of a single algae. Thereafter, the first algae rapidly grows, promoting the growth of the second algae, and the number of algae cells in the system quickly recovers. That is, the culture method according to the present embodiment is more robust against such changes in the culture environment than conventional single algae culture.

[0051] The composite culture process according to this embodiment may be batch culture or continuous culture (also called serial culture). The culture method according to this embodiment is highly robust against changes in the culture environment, and therefore can be applied to long-term culture such as continuous culture. During continuous culture, it is preferable to continuously or intermittently replace (recover and add) a part of the algae and a part of the culture solution in the culture tank with newly prepared algae and a new culture medium.

[0052] The period of the composite culture step according to the present embodiment is not particularly limited. The period of the composite culture step according to the present embodiment is, for example, 3 days or more, 1 week or more, 1 month or more, 1 year or more, etc. In the case of the above-mentioned continuous culture, it is possible to culture algae semi-permanently.

[0053] (How to use algae) The algae cultivation method according to this embodiment can be included in an algae production method.

[0054] According to the method of the present embodiment, algae can be efficiently cultured, and therefore can be used for nitrogen fixation, nitrification, carbon fixation, and the like in the atmosphere and water. In addition, in one aspect of the present embodiment, even if an algae has a low nitrification ability when cultured alone, it can exhibit a high nitrification effect by co-culturing it with a specific algae (e.g., Scutellaria baicalensis). In this case, it is preferable that the specific algae is the first algae and the algae with a low nitrification ability is the second algae. Therefore, the method of the present embodiment can be used in water treatment facilities, aquaculture facilities, and the like. In addition, the produced algae can be recovered and carbonized together with wood materials and the like, and used as carbonized pellets or biochar. Furthermore, the produced algae, or algae that has been processed after production, such as carbonized or dried, can be used as fertilizer or feed.

[0055] When the algae according to the present embodiment produces a useful substance, the method according to the present embodiment can efficiently produce the useful substance. In particular, when the second algae produces a useful substance, the useful substance can be produced more efficiently than when a single algae is cultured. When the algae according to the present embodiment can produce one or more substances selected from the group consisting of ethanol, hydrocarbons, and lipids, these can be recovered and used as biofuel. Furthermore, when the algae according to the present embodiment can produce an adhesive substance, microplastics in the culture solution can be removed. Therefore, the method according to the present embodiment can be used in water treatment facilities, aquaculture facilities, and the like. Furthermore, when the adhesive substance is a polysaccharide, it can also be used as a raw material for bioplastics.

[0056] (Algae composition) The algae composition according to this embodiment is a composition containing multiple types of algae. Specifically, it is a composition containing at least a first algae and a second algae in any ratio. By obtaining such a composition, the above-mentioned preparation process becomes easier. The cell number ratio of the first algae to the second algae is preferably 1:0.01 to 100, more preferably 1:0.1 to 10, and even more preferably 1:0.5 to 2. A composition with such a cell number ratio allows efficient cultivation of algae.

[0057] In the algal composition according to this embodiment, the liquid containing the algae is arbitrary. The liquid according to this embodiment may be a conventionally known culture medium or a frozen storage solution. The algal composition according to this embodiment may be in a liquid or frozen state, so long as the algae are viable. When the algal composition according to this embodiment is in a liquid state, it can be used immediately without the need for thawing. On the other hand, when the algal composition according to this embodiment is in a frozen state, it has excellent storage stability.

[0058] The algae composition according to the present embodiment may further include any additives. Examples of the additives according to the present embodiment include vitamins such as thiamine, and minerals such as Fe, Mn, and Co.

[0059] The present invention is not limited to the above-described embodiments, and various modifications such as design changes may be made based on the knowledge of those skilled in the art, and such modified embodiments are also included in the scope of the present invention.

[0060] In one aspect of this embodiment, the type of algae and the cell number ratio may be selected according to the use of the algae, and multiple types of algae may be provided to a user cultivating algae either individually or as an algae composition. Such an aspect is also included in the scope of the present invention. EXAMPLES

[0061] 1. Cultivation of Complex Algae Green algae (Chlamydomonas) and blue-green algae (Microcystis) were pre-cultured in 500 mL cell culture flasks. BN medium (composition shown in Table 1) was used as the medium, and algal cells were seeded in 200 mL of the medium so that the algal cells would be 7000 cells / mL at the start of the culture. These were placed in an artificial climate chamber (LPH-241, manufactured by Nippon Medical and Chemical Equipment Co., Ltd.) set at 20°C and cultured statically for 14 days to prepare pre-culture flasks. The cell densities of Chlamydomonas and Microcystis at the end of the pre-culture were 1.2 x 10 6 cells / mL and 1.7 x 10 6 cells / mL.

[0062] [Table 1]

[0063] Next, 250 mL of BN medium was prepared in each of four 1 L cell culture flasks. The preculture solution was added so as to obtain the respective algae concentrations shown in Table 2, and these were placed in an artificial climate chamber and cultured under the following conditions. <Culture conditions> Temperature: 20℃ Light output: LED, 5000Lx Airflow rate: 0.8L / min

[0064] At each specified time interval, a small amount of the culture solution was sampled and the absorbance at a wavelength of 680 nm was measured using an ultraviolet-visible spectrophotometer (Shimadzu Corporation, BioSpec-Mini). From these results, the cell density was calculated from a cell density-absorbance calibration curve prepared in advance for each algae (for multiple algae, a standard solution with an equal number of cells was used to prepare the curve). The cell density at each specified time interval is shown in Table 2 and Figure 1, and the percentage of the cell number and the cell increase rate relative to the start of the culture are shown in Table 3.

[0065] [Table 2]

[0066] [Table 3]

[0067] In Comparative Example 1 (when Chlamydomonas was cultured alone) and Comparative Example 2 (when Microcystis was cultured alone), as shown in Table 3, the cell increase rate of Chlamydomonas after 6 hours from the start of culture was 20% or more higher than the cell increase rate of Microcystis after 6 hours from the start of culture (condition (1)). Also, as shown in FIG. 1 and Table 3, the cell number of Chlamydomonas was 200% or more from the start of culture at the time when the cell number of Microcystis was 150% from the start of culture (between 12 and 24 hours) (condition (2)). Furthermore, as shown in FIG. 1, the cell number of Chlamydomonas was higher than the cell number of Microcystis at the beginning of culture, and the cell number of Chlamydomonas was lower than the cell number of Microcystis after a certain time (about 55 hours) (condition (3)). In addition, as shown in Table 2, the cell number in the stationary phase (when cell growth has almost stopped, approximately 48 hours after the start of growth) in Chlamydomonas was found to be smaller than the cell number in the stationary phase (when cell growth has almost stopped, approximately 120 hours after the start of growth) in Microcystis (condition (4)).

[0068] In addition, for Examples 1 and 2, it was found that the final cell increase rate was higher by composite culturing of multiple algae than when each algae was cultured alone (Comparative Examples 1 and 2), as shown in Table 3. In addition, for Example 2, it was found that the final cell increase rate was higher by composite culturing of multiple types of algae than when each algae was cultured alone (Comparative Examples 1 and 2), as shown in Table 2.

[0069] 2. Cultivation using low-nutrient medium Pre-culture was carried out under the same conditions as in 1. A low-nutrient medium containing 110 mg / L (NH2)2CO in BN medium and dechlorinated tap water were prepared in a 1 L cell culture flask, and the pre-culture solution was added to the desired algae concentration. These were then cultured under the same conditions as in 1.

[0070] At each time point, a small amount of the culture medium was taken and the absorbance at a wavelength of 680 nm was measured using an ultraviolet-visible spectrophotometer. The cell density was calculated from the results. The cell densities at each time point are shown in Table 4, Figure 2 (low nutrition) and Figure 3 (dechlorinated tap water), and the percentage of cell count and cell growth rate compared to the start of culture are shown in Table 5.

[0071] [Table 4]

[0072] [Table 5]

[0073] It was found that Comparative Examples 3 and 4, as well as Comparative Examples 5 and 6, satisfied the above conditions (1) to (4), as shown in Tables 4 and 5, Figures 2 and 3. It was also found that even when a low-nutrient medium and dechlorinated tap water were used, the algae could be cultured (produced) more efficiently by co-culturing multiple types of algae compared to the case where each algae was cultured alone.

[0074] 3. Impact of environmental changes Pre-culture was carried out under the same conditions as in 1. As in 1, pre-culture solution was added to achieve the specified algae concentration. From the start of culture until 6 hours had elapsed, the LED was turned off and culture was carried out without ventilation, but otherwise the culture conditions were the same as in 1.

[0075] At each time point, a small amount of the culture medium was sampled and the absorbance at a wavelength of 680 nm was measured using a UV-Visible spectrophotometer. The cell density was calculated from the results. The cell density at each time point is shown in Table 6 and FIG. 4.

[0076] [Table 6]

[0077] As shown in Table 6 and Figure 4, by culturing multiple types of algae in combination, the algal cell count can be quickly recovered even when the culture environment changes, and the algae can be cultivated (produced) efficiently, compared to when each algae is cultured alone.

Claims

1. A preparation step of preparing a culture solution containing a first algae and a second algae; a composite culture step of culturing the first algae and the second algae in the culture solution; Equipped with the cell number ratio of the first algae to the second algae is 1:0.1 to 100; A method for culturing algae, wherein the first algae is a green algae and the second algae is a photosynthetic bacterium.

2. A method for culturing algae as described in claim 1, wherein the total cell number of the first algae and the second algae is greater than the total cell number when the first algae and the second algae are cultured alone under the same conditions capable of culturing the first algae and the second algae.

3. A method for producing a culture solution comprising: a preparation step of preparing a culture solution containing a first algae and a second algae; a composite culture step of culturing the first algae and the second algae in the culture solution; Equipped with the cell number ratio of the first algae to the second algae is 1:0.1 to 100; A method for producing algae, wherein the first algae is a green algae and the second algae is a photosynthetic bacterium.

4. A method for producing algae as described in claim 3, wherein the total cell number of the first algae and the second algae is greater than the total cell number when the first algae and the second algae are cultured alone under the same conditions capable of culturing the first algae and the second algae.

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