CN113136343A - Culture method of photosynthetic microorganism and culture system and application thereof - Google Patents
Culture method of photosynthetic microorganism and culture system and application thereof Download PDFInfo
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Abstract
The invention relates to the field of microalgae culture, in particular to a culture method of photosynthetic microorganisms, a culture system and application thereof. The method comprises the following steps: under the first illumination and the first ventilation, the photosynthetic microorganisms are sent to a reactor A for mixotrophic culture to obtain a first culture solution; and (3) sending part or all of the first culture solution to a reactor B without dilution, and performing autotrophic culture under second illumination and second ventilation to obtain a second culture solution. The method provided by the invention can effectively improve the quality of the photosynthetic microorganisms obtained by mixotrophic culture.
Description
Technical Field
The invention relates to the field of microalgae culture, in particular to a culture method of photosynthetic microorganisms, a culture system and application thereof.
Background
Microalgae are a group of lower plants which grow in water in a wide variety and distribution, and comprise three culture methods of autotrophic culture, heterotrophic culture and mixotrophic culture. Wherein, the autotrophic culture is mainly to absorb CO through the high-efficiency photosynthesis of microalgae cells2Converting light energy into chemical energy of carbohydrate such as fat or starch, and releasing O2. The biological energy and chemicals produced by utilizing microalgae are expected to simultaneously replace fossil energy and reduce CO2Discharge, etc. Microalgae have received much attention in recent years because of their extremely high production efficiency. The heterotrophic culture is a growth mode utilizing an organic carbon source, the growth rate can be improved by more than ten times or even dozens of times, and the culture is mainly carried out in a fermentation tank. The mixotrophic culture can simultaneously utilize light energy and chemical energy in an organic carbon source for growth and CO2And an organic carbon source, and the growth rate is higher than that of the autotrophy and the heterotrophy. The light energy mixotrophic culture of the microalgae is also called mixed nutrient culture, can promote the growth of a plurality of microalgae and the synthesis of protein thereof, has become a new technology for culturing the microalgae because of the advantages of shortening the culture period and realizing the high-density culture of cells, and has important significance in production and economy.
The cost is the core problem of microalgae cultivation, organic carbon sources are required in the process of heterotrophic or mixotrophic cultivation of microalgae, and the organic carbon sources are a large part of the cost of the heterotrophic or mixotrophic cultivation of microalgae. In order to reduce the growth cost of microalgae, foreign scholars study the influence of glucose, acetic acid, lactic acid, glycerol, glycine and the like on the growth of the micro-vacant algae, the phaeodactylum tricornutum, the chlorella pyrenoidosa, the spirulina and the like and the accumulation of bioactive substances, and the study result shows that the soluble organic matter with proper concentration is beneficial to the growth of the microalgae and the accumulation of the active substances. Kirrolia et al (Renewable and susteable Energy Reviews 20: 642) compared the costs of microalgae in three different cultivation modes, namely open raceway ponds, photobioreactors and conventional fermenters in 2013, and the comparison showed that the costs of producing oil per unit mass were $ 7.64, $ 24.6 and $ 1.54, and the costs of producing microalgal biomass per unit mass were $ 1.54, $ 7.32 and $ 1.02, respectively. Although organic carbon sources are required to be added for culturing the microalgae in the fermentation tank, the production cost is not increased, which probably is because the microalgae grow in the fermentation tank at high efficiency, the production period of the microalgae is shortened, and other expenses such as labor cost, equipment depreciation cost, land occupation cost and the like for producing the microalgae with unit mass are reduced, thereby reducing the production cost of the microalgae.
Disclosure of Invention
And light conditions are provided during mixotrophic cultivation, but compared with the microalgae cultivated by autotrophy, the contents of protein and pigment are still obviously reduced, and the effect of improving the quality of the microalgae biomass is limited only by supplementing light during the mixotrophic cultivation. The microalgae needs to consume a large amount of O in the process of growing the microalgae2Although oxygen released by algae cell autotrophy can relieve oxygen supply pressure, a large amount of O needs to be supplemented2If gas exchange cannot be carried out in time, the growth of microalgae can be seriously influenced, and even cultivation failure can be caused.
One of the purposes of the invention is to improve the quality of mixotrophic photosynthetic microorganisms, and the other is to enhance the mixotrophic process by autotrophic culture. Therefore, the invention provides a culture method of photosynthetic microorganisms, a culture system and application thereof.
In order to achieve the above object, the present invention provides, in one aspect, a method for culturing a photosynthetic microorganism, the method being performed in a reactor a and a reactor B connected in series, the method comprising:
(1) under the first illumination and the first ventilation, the photosynthetic microorganisms are sent to a reactor A for mixotrophic culture to obtain a first culture solution;
(2) and (3) sending part or all of the first culture solution to a reactor B without dilution, and performing autotrophic culture under second illumination and second ventilation to obtain a second culture solution.
In a second aspect, the present invention provides a culture system for photosynthetic microorganisms, the system comprising: a reactor A for mixotrophic cultivation of photosynthetic microorganisms and a reactor B for autotrophic cultivation of photosynthetic microorganisms, wherein the reactor A comprises a first illumination device and a first ventilation device, the reactor B comprises a second illumination device and a second ventilation device,
and the culture solution outlet of the reactor A is communicated with the algae solution inlet of the reactor B, so that the photosynthetic microorganisms directly enter the reactor B for autotrophic culture after the mixotrophic culture in the reactor A.
In a third aspect, the present invention provides a method for producing biomass, comprising cultivating a photosynthetic microorganism using the above method, and extracting biomass from the resulting photosynthetic microorganism.
In a fourth aspect, the present invention provides a method for producing a bioenergy source, which comprises cultivating a photosynthetic microorganism using the above method.
The method of the invention significantly improves the quality of the mixotrophic photosynthetic microorganisms by a high concentration autotrophic process. In one embodiment of the present invention, the present invention provides a method that also effectively enhances the mixotrophic culture of photosynthetic microorganisms using autotrophic culture of photosynthetic microorganisms. In particular, the invention makes it possible to obtain the following advantages:
(1) although mixotrophic culture provides light conditions, mixotrophic culture also produces CO2However, mixotrophic alone has limited effect on improving biomass quality of photosynthetic microorganisms. The invention effectively implements the enhanced autotrophic culture of the high-concentration photosynthetic microorganisms from the mixotrophic process by providing specific illumination conditions, and obviously improves the biomass quality of the mixotrophic photosynthetic microorganisms.
(2) The use ratio of organic carbon source by mixotrophy is higher than that by heterotrophy. The invention strengthens the mixotrophic culture process, improves the production efficiency and reduces the culture cost by collecting and utilizing the exhaust gas in the autotrophic process and providing an air source for the mixotrophic culture.
Drawings
FIG. 1 is a chlorella growth curve of a fermenter according to an example of the present invention.
FIG. 2 is a curve of chlorella growth in a tubular reactor of examples 1-6 of the present invention.
FIG. 3 is a chlorophyll-a/dry weight change curve of chlorella in a fermenter according to examples 1 to 6 of the present invention with time.
FIG. 4 is a chlorophyll-a/dry weight change curve of Chlorella vulgaris over time in a tubular reactor according to examples 1-6 of the present invention.
Detailed Description
The endpoints of the ranges and any values disclosed herein are not limited to the precise range or value, and such ranges or values should be understood to encompass values close to those ranges or values. For ranges of values, between the endpoints of each of the ranges and the individual points, and between the individual points may be combined with each other to give one or more new ranges of values, and these ranges of values should be considered as specifically disclosed herein.
In one aspect, the present invention provides a method for culturing photosynthetic microorganisms, the method being performed in a reactor a and a reactor B connected in series, the method comprising:
(1) under the first illumination and the first ventilation, the photosynthetic microorganisms are sent to a reactor A for mixotrophic culture to obtain a first culture solution;
(2) and (3) sending part or all of the first culture solution to a reactor B without dilution, and performing autotrophic culture under second illumination and second ventilation to obtain a second culture solution.
In a second aspect, the present invention provides a culture system for photosynthetic microorganisms, the system comprising: a reactor A for mixotrophic cultivation of photosynthetic microorganisms and a reactor B for autotrophic cultivation of photosynthetic microorganisms, wherein the reactor A comprises a first illumination device and a first ventilation device, the reactor B comprises a second illumination device and a second ventilation device,
and the culture solution outlet of the reactor A is communicated with the algae solution inlet of the reactor B, so that the photosynthetic microorganisms directly enter the reactor B for autotrophic culture after the mixotrophic culture in the reactor A.
The above methods and systems of the present invention will be described concurrently below, but it should be understood that the methods and systems of the present invention can be used in conjunction with each other, or independently as the subject of the present invention.
In the invention, the gas discharged from the reactor B is used as a part or all of the gas source of the first aeration, and when the gas discharged from the reactor B is used as a part of the gas source, the gas discharged from the reactor B is mixed with other gases for conventional gas supply and then is used as the gas source of the first aeration.
In the present invention, it is understood that, when the autotrophic culture is carried out in the reactor B, the mixotrophic culture is also carried out in the reactor A, so that the gas discharged from the reactor B can be used as a source of aeration gas for the reactor A.
In order to better promote the series mixotrophic-autotrophic auxiliary culture, it is preferable that the gas discharged from the reactor B is collected and compressed to be used as a gas source for the first aeration.
In the system of the present invention, for this purpose, preferably, the exhaust port of the reactor B is sequentially connected to a gas collecting device and a pressure boosting device, and then is connected to the first ventilation device of the reactor a, so as to collect and compress the gas discharged from the reactor B and use the gas as the gas source of the first ventilation device.
The reactor B may be open or closed, preferably of tubular, plate or column photobioreactor construction or open raceway pond construction. In order to effectively utilize the gas discharged from the reactor B, the reactor B is of a closed type photobioreactor structure, and the discharged gas is convenient to collect. While the reactor a is a closed reactor, preferably a fermenter configuration.
According to the invention, any existing compatible gas can be used for the first aeration. Preferably, the gas discharged from the reactor B is used, and the gas is doped with oxygen generated in the autotrophic culture process, so that the content of the oxygen is higher than that of air, and the mixotrophic culture is more facilitated. The aeration rate of the first aeration is preferably 0.1-10L/(L.min), preferably 0.2-5L/(L.min), to enable better mixotrophic growth of the photosynthetic microorganisms in the reactor A of the present invention.
In order to be suitable for the high-concentration autotrophic culture, the gas source used for the second aeration is preferably a carbon dioxide-rich gas, such as carbon dioxide-rich air. The second source of aerated gas has a carbon dioxide content of 0.03 to 5% by volume, for example 0.1 to 2%. In the case of autotrophic culture, the aeration rate of the second aeration is preferably 0.1 to 10L/(L.min), more preferably 0.2 to 5L/(L.min).
According to the present invention, the aeration device used for the first aeration and the second aeration may be an aeration device structure conventionally used in the art as long as it can be used for performing the first aeration and the second aeration of the present invention.
According to the invention, the mixotrophic culture can be carried out under stirring, wherein the stirring speed is preferably 200-500 r/min. Therefore, a stirring structure may be added to the reactor a of the system of the present invention.
According to the invention, the photosynthetic microorganisms of the reactor B originate from the culture broth of the reactor A after mixotrophic culture. In this case, a part of the culture solution may be withdrawn from the culture solution after the mixotrophic culture in the reactor A each time and sent to the reactor B, and the remaining culture solution may be continuously mixotrophic cultured in the reactor A (by additionally supplying a nutrient solution), whereby the gas supply from the reactor B to the reactor A in the method of the present invention means during the culture in both the reactor B and the reactor A. Preferably, wherein the part of the culture broth from reactor A that is fed to reactor B is 70-90% by volume of the total culture broth of reactor A.
It should be understood that if the culture solution after the mixotrophic culture in the reactor A is used as the source of the photosynthetic microorganisms in the reactor B, the aeration gas source used in the mixotrophic culture may be compressed air directly or compressed air after aseptic treatment may be used in the first mixotrophic culture.
Wherein the culture solution after the mixotrophic culture sent from the reactor A is sent to the reactor B without dilution for autotrophic culture, so that a photosynthetic microorganism product with higher quality can be obtained.
In the system of the present invention, in order to allow the mixotrophic culture liquid fraction of the reactor a to be sent to the reactor B for autotrophic culture, the culture liquid outlet of the reactor a is communicated with the algal liquid inlet of the reactor B, so that the photosynthetic microorganisms are directly sent to the reactor B for autotrophic culture after mixotrophic culture in the reactor a. The reactor A and the reactor B are communicated through a culture solution conveying pipeline, a conveying pump can be arranged on the pipeline, and the position of the reactor A is higher than that of the reactor B and has enough height difference so as to be beneficial to conveying the culture solution from the reactor A to the reactor B.
According to the present invention, the first illumination and the second illumination should adopt illumination intensity suitable for the mixotrophic culture and the autotrophic culture, respectively, and in order to be more suitable for the culture method of the present invention, to obtain higher quality photosynthetic microorganisms, preferably, the illumination intensity of the first illumination is 2000-.
In order to be more beneficial to the autotrophic culture, a two-stage illumination mode is adopted, namely preferably, the second illumination comprises first-stage illumination and second-stage illumination, the light intensity of the first-stage illumination is below 5000lux, preferably 2000 + 5000lux, and more preferably 3000 + 4000 lux;
the light intensity of the second stage illumination is greater than 5000lux, preferably 6000-20000lux, and more preferably 10000-15000 lux.
According to the present invention, the duration of the first stage illumination is preferably 10 hours or more, preferably 10 to 36 hours, more preferably 12 to 30 hours, especially 20 to 24 hours.
According to the present invention, the duration of the second-stage light irradiation is not particularly limited, and the second-stage light irradiation is performed for the remaining culture time after the first-stage light irradiation. It is understood that the reactor B is used for a new round of inoculation of photosynthetic microorganisms for the first stage of illumination and then for the second stage of illumination.
In the invention, the first illumination adopts an artificial light source; the second illumination adopts an artificial light source.
According to the invention, the illumination wavelength can be varied in a wide range, and can be partial wavelength light or full wavelength light, in order to facilitate the growth of the photosynthetic microorganisms in the invention, preferably, the wavelengths of the first illumination and the second illumination are both 380-780nm, more preferably 490-460nm and/or 620-760nm, under the illumination wavelength, the photosynthetic microorganism cells can better utilize the light energy, and the energy consumption for cultivating the photosynthetic microorganisms is reduced.
The light source used for the above illumination may be an LED light source, in particular a blue and red LED light source. In order to isolate water vapor, the light source can be sealed by adopting a transparent material.
According to the invention, in order to enable the microorganisms to better utilize the light energy, it is preferred for the inventive light source arrangement that the spacing of the light sources in the direction of the light rays is 2-300mm, preferably 60-200 mm; the light source can also be sealed and directly inserted into the culture solution.
According to the present invention, it is preferable that the temperature of the mixotrophic culture is 20 to 35 ℃. The cultivation time may vary within wide limits, for example from 3 to 10 days. The time of mixotrophic culture is herein understood to be the period of time from each start of culture to re-inoculation of a new photosynthetic microorganism species or addition of a new nutrient solution.
According to the present invention, the temperature of the autotrophic culture is preferably 20-35 ℃. The incubation time may vary within wide limits, for example from 3 to 20 days. The time of autotrophic cultivation is understood here to be the period of time from each start of cultivation to the re-inoculation of a new species of photosynthetic microorganisms.
More preferably, the photosynthetic microorganism is a microalgae, preferably a green alga, more preferably a chlorella.
According to the present invention, the reactor A of the present invention is hermetically sealed, so that a sterile mixotrophic culture, which requires the supplementation of an organic carbon source, can be performed. Preferably, the organic carbon source is a saccharide and/or an acetate. The saccharide may be one or more of glucose, fructose, sucrose, maltose, and the like. The solvent may be, for example, sodium acetate. More preferably, the organic carbon source is glucose.
Wherein the amount of the organic carbon source added may vary within a wide range, and preferably, the amount of the organic carbon source added in the culture system is 5 to 15 g/L.
According to the invention, the mixotrophic culture system of the photosynthetic microorganisms may also be supplemented with other agents conventionally used in the art, such as phosphate (e.g. K)2HPO4、Na2HPO4Etc.). They may be used in amounts conventional in the art, and the present invention is not particularly limited thereto.
According to the present invention, in order to maintain a constant sterile environment for mixotrophic culture, the gas used for aeration may be sterile, and antibiotics may be added to the mixotrophic culture system to prevent bacterial growth. Such antibiotics may be those conventionally employed in the art for controlling the aseptic culture of microorganisms, and may be, for example, one or more of kanamycin, chloramphenicol, streptomycin, gentamicin, vancomycin, azithromycin, and the like. The amount may vary within wide limits, for example from 10 to 65 mg/L.
According to the present invention, the culture system of the photosynthetic microorganism can be appropriately adjusted according to the algal species, and the culture medium used preferably has the following composition: k2HPO4·3H2O：20-50mg/L，NaNO3：1200-2000mg/L，Na2CO3：10-30mg/L，MgSO4·7H2O：50-90mg/L，CaCl2·2H2O: 30-50mg/L, citric acid: 1-10mg/L, ferric ammonium citrate: 1-10mg/L, sodium EDTA: 0.5-2mg/L, trace element A5: 0.5-2 ml/L.
The composition of the trace element A5 is preferably as follows: h3BO3：2500-3000mg/L，MnCl2·4H2O：1500-2000mg/L，ZnSO4·7H2O：200-250mg/L，CuSO4·5H2O：50-90mg/L，NaMoO4·5H2O：350-420mg/L，Co(NO3)2·6H2O：20-65mg/L。
In a third aspect, the present invention provides a method for producing biomass, comprising cultivating a photosynthetic microorganism using the above method, and extracting biomass from the resulting photosynthetic microorganism.
The biomass may be any of a variety of biomass conventional in the art, for example, may be one of a lipid, a protein, a carbohydrate, a nucleic acid, a pigment, a vitamin, a growth factor, or any combination thereof.
In a fourth aspect, the present invention provides a method for producing a bioenergy source, which comprises cultivating a photosynthetic microorganism using the above method.
The method is suitable for the culture of photosynthetic microorganisms, and can obtain high-quality photosynthetic microorganism fermentation liquor with higher yield under lower energy consumption.
The present invention will be described in detail below by way of examples.
In the following examples:
and (3) measuring the dry weight of the chlorella: taking an appropriate amount of algae liquid, 6000r/min, centrifuging for 5min, removing supernatant, freeze-drying algae mud for 72h, and weighing. Chlorella species are from the institute for aquatic organisms, academy of sciences, China. And an alga seed preparation stage, namely adding about 600mL of BG11 culture medium and 5g/L of glucose into a triangular flask, then sterilizing at 120 ℃ for 30min, cooling, adding a proper amount of alga seeds and 50mg/L of kanamycin, and then introducing sterile air to culture for about 3d under the conditions that the light intensity is 6000lux and the temperature is 28 ℃ to obtain induced alga seeds.
BG11 medium composition: k2HPO4·3H2O：40mg/L，NaNO3：1500mg/L，Na2CO3：20mg/L，MgSO4·7H2O：75mg/L，CaCl2·2H2O: 36mg/L, citric acid: 6mg/L, ferric ammonium citrate: 6mg/L, sodium EDTA: 1mg/L, trace element A5: 1 ml/L.
Composition of trace element a 5: h3BO3：2860mg/L，MnCl2·4H2O：1810mg/L，ZnSO4·7H2O：222mg/L，CuSO4·5H2O：79mg/L，NaMoO4·5H2O：390mg/L，Co(NO3)2·6H2O：50mg/L。
A culture system: the culture system comprises a reactor A and a reactor B which are sequentially connected in series, wherein the reactor A is a closed fermentation tank with the volume of 5L, and the reactor B is a closed tubular photobioreactor with the volume of 10L; wherein, the algae liquid outlet of the bottom of the closed fermentation tank is connected with an algae liquid conveying pipe, the algae liquid conveying pipe is connected to a conveying pump, and the outlet of the conveying pump is connected to the algae liquid inlet at the upper part of the closed tubular photobioreactor through another algae liquid conveying pipe; the top exhaust port of the closed tubular photobioreactor is connected to a collecting tank, the gas exhaust port of the collecting tank is connected to an air compressor, and the exhaust port of the air compressor is connected to the air inlet of the ventilation device in the closed fermentation tank.
Example 1
This example is intended to illustrate the method for culturing a photosynthetic microorganism of the present invention.
Adding 3L BG11 culture medium and 15g/L glucose into a closed fermentation tank of the culture system, sterilizing at 120 deg.C for 30min, cooling, inoculating Chlorella strain induced by glucose and kanamycin at 50mg/L, introducing 1L/(L.min) sterile air at 28 deg.C, 250r/min, 10000lux of light intensity, 380-: and (4) 12 h. Supplementing 15g/L glucose every day, supplementing other nutrient salts according to consumption, and performing mixotrophic culture until the growth of algae cells is slow.
Directly transferring 80 vol% algae solution from a closed fermentation tank to a closed tubular photobioreactor without dilution for autotrophic culture at 28 deg.C, and introducing 1L/(L.min) gas containing CO2The volume fraction is 0.2%, the illumination intensity is 15000lux, the illumination wavelength is 380-780nm, and the light-dark period is 12 h: and (4) 12 h. Simultaneously adding sterilized 2.4L BG11 culture medium, 15g/L glucose, and 50mg/L kanamycin into closed fermentation tank, supplementing nutrition every day, and sealing with the rest algae solution as algae seedPerforming mixotrophic culture in a closed fermentation tank.
Wherein, gas discharged from an exhaust port of the tubular photobioreactor is collected and compressed to be sent to a bottom gas inlet of the closed fermentation tank, the closed fermentation tank is ventilated, and the ventilation flow is kept to be 1L/(L.min); the gas discharged from the exhaust port of the closed fermentation tank is directly discharged, and the closed tubular photobioreactor adopts pure compressed air for ventilation, so that the ventilation flow is kept at 1L/(L.min).
The dry weight of chlorella and the chlorophyll a/dry weight (weight ratio) in the fermentor and tubular photobioreactor were measured at the designated time under the simultaneous performance of the mixotrophic culture and the autotrophic culture as shown in tables 1 and 2, respectively.
Example 2
This example is intended to illustrate the method for culturing a photosynthetic microorganism of the present invention.
Adding 3L BG11 culture medium and 15g/L glucose into a closed fermentation tank of the culture system, then sterilizing at 120 ℃ for 30min, cooling for later use, adding chlorella strain induced by glucose and 10mg/L chloramphenicol during inoculation, introducing 0.8L/(L.min) sterile air, controlling the temperature at 28 ℃, the rotation speed at 250r/min, the light intensity at 10000lux, the illumination wavelength at 380-780nm, and the light-dark period at 12 h: and (4) 12 h. Supplementing 15g/L glucose every day, supplementing other nutrient salts according to consumption, and performing mixotrophic culture until the growth of algae cells is slow.
Directly transferring 80 vol% algae solution from a closed fermentation tank to a closed tubular photobioreactor without dilution for autotrophic culture at 28 deg.C, and introducing 1L/(L.min) gas containing CO2The volume fraction is 0.2%, the illumination intensity is 10000lux, and the illumination wavelength is 380-780 nm. Meanwhile, the sterilized 3L BG11 culture medium, 15g/L glucose and 10mg/L chloramphenicol were continuously added into the closed fermentation tank, and the nutrition was supplemented every day, and the remaining part of the algae solution was used as the algae seed to continue the mixotrophic culture in the closed fermentation tank.
Wherein, the gas discharged from the exhaust port of the tubular photobioreactor is collected and compressed to be sent to the bottom gas inlet of the closed fermentation tank, the closed fermentation tank is ventilated, and the ventilation flow is kept to be 0.8L/(L.min); the gas discharged from the exhaust port of the closed fermentation tank is directly discharged, and the closed tubular photobioreactor adopts pure compressed air for ventilation, so that the ventilation flow is kept at 0.8L/(L.min).
The dry weight of chlorella and the chlorophyll a/dry weight (weight ratio) in the fermentor and tubular photobioreactor were measured at the designated time under the simultaneous performance of the mixotrophic culture and the autotrophic culture as shown in tables 1 and 2, respectively.
Example 3
This example is intended to illustrate the method for culturing a photosynthetic microorganism of the present invention.
According to the method described in example 1, except that, in the case where the mixotrophic culture and the autotrophic culture are simultaneously performed, the mixotrophic culture is performed with illumination at 5000 lux; in the autotrophic culture, the autotrophic culture is performed under 6000lux for illumination; the dry weight of chlorella and the chlorophyll a/dry weight (weight ratio) in the fermentor and tubular photobioreactor were measured at the designated time under the simultaneous performance of the mixotrophic culture and the autotrophic culture as shown in tables 1 and 2, respectively.
Example 4
This example is intended to illustrate the method for culturing a photosynthetic microorganism of the present invention.
According to the method described in example 1, except that when the mixotrophic culture and the autotrophic culture are performed simultaneously, in the mixotrophic culture, the mixotrophic culture is performed under illumination of 30000 lux; in the autotrophic culture, the autotrophic culture is performed under 50000lux for illumination; the dry weight of chlorella and the chlorophyll a/dry weight (weight ratio) in the fermentor and tubular photobioreactor were measured at the designated time under the simultaneous performance of the mixotrophic culture and the autotrophic culture as shown in tables 1 and 2, respectively.
Example 5
This example is intended to illustrate the method for culturing a photosynthetic microorganism of the present invention.
The method according to example 1, except that, when the mixotrophic culture and the autotrophic culture are performed simultaneously, the illumination wavelength for the mixotrophic and autotrophic cultures is the full wavelength; the dry weight of chlorella and the chlorophyll a/dry weight (weight ratio) in the fermentor and tubular photobioreactor were measured at the designated time under the simultaneous performance of the mixotrophic culture and the autotrophic culture as shown in tables 1 and 2, respectively.
Example 6
According to the method described in example 1, except that the mixotrophic culture and the autotrophic culture are performed simultaneously, air is introduced into the fermenter instead of the gas discharged from the tubular photobioreactor; the dry weight of chlorella and the chlorophyll a/dry weight (weight ratio) in the fermentor and tubular photobioreactor were measured at the designated time under the simultaneous performance of the mixotrophic culture and the autotrophic culture as shown in tables 1 and 2, respectively.
TABLE 1
TABLE 2
The dry weight change of the chlorella in the fermenter according to Table 1 with time is plotted as a chlorella growth curve shown in FIG. 1, and the dry weight change of the chlorella in the tubular reactor according to examples 1 to 6 according to Table 1 with time is plotted as a chlorella growth curve shown in FIG. 2; the chlorophyll a/dry weight values of chlorella in the fermentor in table 2 over time were plotted as shown in fig. 3, and the chlorophyll a/dry weight values of chlorella in the tubular reactor in table 2 over time were plotted as shown in fig. 4.
As can be seen from the results shown in the table and the figure, the method of the invention can implement autotrophic culture of high-concentration photosynthetic microorganisms, thereby significantly improving the quality of mixotrophic photosynthetic microorganisms; in addition, the present invention can also utilize autotrophic culture of photosynthetic microorganisms to strengthen the mixotrophic process of photosynthetic microorganisms.
The preferred embodiments of the present invention have been described above in detail, but the present invention is not limited thereto. Within the scope of the technical idea of the invention, many simple modifications can be made to the technical solution of the invention, including combinations of various technical features in any other suitable way, and these simple modifications and combinations should also be regarded as the disclosure of the invention, and all fall within the scope of the invention.
Claims (13)
1. A method for cultivating photosynthetic microorganisms, the method being carried out in a reactor A and a reactor B connected in series, the method comprising:
(1) under the first illumination and the first ventilation, the photosynthetic microorganisms are sent to a reactor A for mixotrophic culture to obtain a first culture solution;
(2) and (3) sending part or all of the first culture solution to a reactor B without dilution, and performing autotrophic culture under second illumination and second ventilation to obtain a second culture solution.
2. The method according to claim 1, wherein the second illumination comprises a first-stage illumination and a second-stage illumination, and the light intensity of the first-stage illumination is below 5000lux, preferably 2000-5000lux, and more preferably 3000-4000 lux;
the light intensity of the second stage illumination is greater than 5000lux, preferably 6000-20000lux, and more preferably 10000-15000 lux.
3. The method according to claim 1 or 2, wherein the illumination intensity of the second illumination is 5000-.
4. The method according to any one of claims 1-3, wherein the intensity of the first illumination is 5000-.
5. The method according to any one of claims 1 to 4, wherein the first aeration has an aeration volume of 0.1 to 10L/(L-min), preferably 0.2 to 5L/(L-min);
preferably, the ventilation volume of the second ventilation is 0.1-10L/(L.min), preferably 0.2-5L/(L.min); more preferably, the second vent gas is carbon dioxide-containing gas, and the carbon dioxide-containing gas is air or carbon dioxide-rich air.
6. The process according to any one of claims 1 to 5, wherein the gas discharged from the reactor B is used as a source of part or all of the first aeration.
7. The method according to any one of claims 1 to 6, wherein the temperature of the mixotrophic culture is 20-35 ℃; the temperature of the autotrophic culture is 20-35 ℃.
8. Method according to any one of claims 1-7, wherein the photosynthetic microorganism is a microalgae, preferably a green algae, more preferably a chlorella.
9. A system for cultivating photosynthetic microorganisms, the system comprising: a reactor A for mixotrophic cultivation of photosynthetic microorganisms and a reactor B for autotrophic cultivation of photosynthetic microorganisms, wherein the reactor A comprises a first illumination device and a first ventilation device, the reactor B comprises a second illumination device and a second ventilation device,
and the culture solution outlet of the reactor A is communicated with the algae solution inlet of the reactor B, so that the photosynthetic microorganisms directly enter the reactor B for autotrophic culture after the mixotrophic culture in the reactor A.
10. The system of claim 9, wherein the exhaust port of the reactor B is connected with a gas collecting device and a pressure increasing device in sequence, and then is connected with the first ventilating device of the reactor a, so that the gas discharged from the reactor B is collected and compressed to be used as the gas source of the first ventilating device.
11. The system of claim 9 or 10, wherein the reactor a is a fermenter structure; the reactor B is in a tubular, plate or column type photobioreactor structure.
12. A method for producing biomass comprising cultivating a photosynthetic microorganism according to the method of any one of claims 1 to 8 and extracting biomass from the resulting photosynthetic microorganism.
13. A method of producing a bioenergy source comprising culturing a photosynthetic microorganism according to the method of any one of claims 1 to 8.
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