CN113136341A - Heterotrophic-autotrophic photosynthetic microorganism culture method and system and biomass and biological energy production method - Google Patents

Heterotrophic-autotrophic photosynthetic microorganism culture method and system and biomass and biological energy production method Download PDF

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CN113136341A
CN113136341A CN202010060653.4A CN202010060653A CN113136341A CN 113136341 A CN113136341 A CN 113136341A CN 202010060653 A CN202010060653 A CN 202010060653A CN 113136341 A CN113136341 A CN 113136341A
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reactor
culture
heterotrophic
gas source
autotrophic
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朱俊英
荣峻峰
程琳
李煦
管炳伟
郄凤翔
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Sinopec Research Institute of Petroleum Processing
China Petroleum and Chemical Corp
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China Petroleum and Chemical Corp
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/12—Unicellular algae; Culture media therefor
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    • C12M21/00—Bioreactors or fermenters specially adapted for specific uses
    • C12M21/02—Photobioreactors
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    • C12M23/00—Constructional details, e.g. recesses, hinges
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    • C12M23/00—Constructional details, e.g. recesses, hinges
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    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/58—Reaction vessels connected in series or in parallel
    • C—CHEMISTRY; METALLURGY
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Abstract

The invention relates to the field of microalgae culture, in particular to a heterotrophic-autotrophic culture method and a system for photosynthetic microorganisms and a method for producing biomass and biological energy. The culture method comprises the following steps: under a first gas source, sending photosynthetic microorganisms into a reactor A for heterotrophic 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 the condition of illumination and a second gas source to obtain a second culture solution. The method provided by the invention can effectively improve the quality of the photosynthetic microorganisms obtained by heterotrophy.

Description

Heterotrophic-autotrophic photosynthetic microorganism culture method and system and biomass and biological energy production method
Technical Field
The invention relates to the field of microalgae culture, in particular to a heterotrophic-autotrophic culture method and a system for photosynthetic microorganisms and a method for producing biomass and biological energy.
Background
Microalgae are a wide variety of lower plants growing in water, and are cell factories driven by sunlight to absorb CO through efficient 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. In addition, part of microalgae can grow heterotrophically by using an organic carbon source like bacteria, the growth rate can be increased by tens of times or even tens of times, and the microalgae are mainly cultured in a fermentation tank. Besides the two nutrition modes, part of microalgae can also grow in a light energy mixotrophic way, namely, the microalgae can grow by utilizing the light energy and the chemical energy in the organic carbon source at the same time and utilize CO at the same time2And 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.
There are advantages and problems associated with each of autotrophy and heterotrophy. Heterotrophic culture, which utilizes chemical energy to accumulate biomass at a high rate, has the disadvantage of reducing the quality of the biomass, for example, the protein and pigment content of heterotrophic microalgae are significantly reduced compared to autotrophic culture of microalgae, and light supplementation is very difficult for large-volume fermenters. On the other hand, a large amount of O is consumed in the heterotrophic culture process2If the gas exchange cannot be carried out in time, the growth of the photosynthetic microorganisms can be seriously influenced, and even the cultivation fails.
Disclosure of Invention
The present invention is directed to solve the above problems of the prior art, and provides a method and a system for heterotrophic-autotrophic cultivation of photosynthetic microorganisms and a method for producing biomass and bioenergy, which are low in cost and suitable for large-scale cultivation of microalgae.
In order to achieve the above objects, the present invention provides, in one aspect, a method for heterotrophic-autotrophic cultivation of a photosynthetic microorganism, the method being carried out in a reactor a and a reactor B connected in series, the method comprising:
(1) under a first gas source, sending photosynthetic microorganisms into a reactor A for heterotrophic 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 the condition of illumination and a second gas source to obtain a second culture solution.
In a second aspect, the present invention provides a heterotrophic-autotrophic culture system for photosynthetic microorganisms, the system comprising: a reactor A and a reactor B, wherein the reactor A is set for heterotrophic culture of photosynthetic microorganisms; the reactor B is set to be used for autotrophic culture of photosynthetic microorganisms, wherein the reactor A comprises a first gas source device, the reactor B comprises a lighting device and a second gas source device,
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 heterotrophic 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 present invention significantly improves the quality of heterotrophic photosynthetic microorganisms through a high concentration autotrophic process. In one embodiment of the present invention, the present invention provides a method that also effectively enhances the heterotrophic culture of photosynthetic microorganisms using autotrophic culture of photosynthetic microorganisms. In particular, the invention makes it possible to obtain the following advantages:
(1) heterotrophic can employ larger volume fermenters than mixotrophic, but heterotrophic culture has a low utilization of organic carbon sources and the biomass quality of the photosynthetic microorganisms is low. The invention effectively implements the intensified autotrophic culture of the high-concentration photosynthetic microorganisms from the heterotrophic process by providing the specific illumination condition, and obviously improves the biomass quality of the heterotrophic photosynthetic microorganisms.
(2) The invention strengthens the heterotrophic 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 heterotrophic culture.
Drawings
FIG. 1 is a graph showing the growth of Chlorella in the fermentor of examples 1-7 of the present invention.
FIG. 2 is a curve of chlorella growth in a tubular reactor of examples 1-7 of the present invention.
FIG. 3 is a chlorophyll-a/dry weight change curve of Chlorella vulgaris over time in a tubular reactor according to examples 1-7 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 heterotrophic-autotrophic cultivation of photosynthetic microorganisms, the method being carried out in a reactor A and a reactor B connected in series, the method comprising:
(1) under a first gas source, sending photosynthetic microorganisms into a reactor A for heterotrophic 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 the condition of illumination and a second gas source to obtain a second culture solution.
In a second aspect, the present invention provides a heterotrophic-autotrophic culture system for photosynthetic microorganisms, the system comprising: a reactor A and a reactor B, wherein the reactor A is set for heterotrophic culture of photosynthetic microorganisms; the reactor B is set to be used for autotrophic culture of photosynthetic microorganisms, wherein the reactor A comprises a first gas source device, the reactor B comprises a lighting device and a second gas source device,
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 heterotrophic 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 one embodiment of the present invention, the gas discharged from the reactor B is used as part or all of the first gas source, and when used as part of the gas source, the gas discharged from the reactor B is mixed with other conventionally supplied gas to be used as the first gas source.
In the present invention, it is understood that when the autotrophic culture is performed in the reactor B, the heterotrophic culture is performed in the reactor A, so that the gas discharged from the reactor B can be used as the source of aeration gas for the reactor A.
In order to better promote the series heterotrophic-autotrophic secondary culture, the gas discharged from the reactor B is preferably collected and compressed to serve as the gas source of the first gas source.
In the system of the present invention, for this purpose, preferably, the exhaust port of the reactor B is sequentially connected to the gas collecting device and the pressure boosting device, and then is connected to the first gas source 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 gas source device.
The reactor B can be an open type or a closed type, is preferably a tubular type, plate type or column type photobioreactor structure or an open type raceway pond structure, and is more preferably a closed type photobioreactor structure, so that the discharged gas can be collected conveniently. While the reactor a is a closed reactor, preferably a fermenter configuration.
According to the invention, the first gas source can be the gas discharged from the reactor B, 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 the air, and the heterotrophic culture is more facilitated. The aeration of the first source of gas is preferably 0.1-10L/(L.min), preferably 0.2-5L/(L.min), to enable better heterotrophic growth of photosynthetic microorganisms in the first culture unit of the invention.
In order to be suitable for high-concentration autotrophic cultures, the gas used by the second gas source is preferably a carbon dioxide-rich gas, said gas used by the second gas source having a carbon dioxide content of 0.03-5%, for example 0.1-2% by volume. Wherein, the ventilation quantity of the second air source is preferably 0.1-10L/(L.min), and more preferably 0.2-5L/(L.min).
According to the present invention, the ventilation means used for the first gas source and the second gas source can be any ventilation means conventionally used in the art as long as the ventilation means can be used for ventilating the first gas source and the second gas source of the present invention.
According to the invention, the heterotrophic culture can be carried out with 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 the heterotrophic culture. In this case, a part of the culture solution may be extracted from the culture solution after the heterotrophic culture in the reactor A each time and sent to the reactor B, while the remaining culture solution is continuously heterotrophic cultured in the reactor A (by additionally supplementing the 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 obtained after the heterotrophic 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 heterotrophic culture can be directly compressed air during the first heterotrophic culture, and the compressed air after the sterile treatment can be used.
Wherein, the culture solution after the heterotrophic culture which is sent from the reactor A is sent to the reactor B without dilution for the autotrophic culture, so that the photosynthetic microorganism products with higher quality can be obtained.
In the system of the present invention, in order to allow the heterotrophic culture liquid fraction of the reactor a to be fed into 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 fed into the reactor B for autotrophic culture after the heterotrophic 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.
In order to be more beneficial to the autotrophic culture, a two-stage illumination mode is adopted, namely preferably, the 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 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 be more beneficial to the growth of the photosynthetic microorganisms in the invention, the illumination wavelength is preferably 380-780nm, more preferably 490-460nm and/or 620-760nm, and under the illumination wavelength, the photosynthetic microorganism cells can better utilize the light energy and reduce the energy consumption for cultivating the photosynthetic microorganisms.
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 present invention, in order to enable the microorganisms to better utilize light energy, the light source arrangement of the present invention is preferably such that the distance between the light sources in the light direction is 2-300mm, preferably 60-200mm, or the light sources can be enclosed and directly inserted into the culture solution.
According to the present invention, the temperature of the heterotrophic culture is preferably 20 to 35 ℃. The cultivation time may vary within wide limits, for example from 3 to 10 days. The time of heterotrophic cultivation is understood here to be the period of time from each start of cultivation to the re-inoculation of a new photosynthetic microorganism species or the 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 invention, the reactor A of the invention is closed, so that sterile heterotrophic culture can be carried out, which requires the addition of an organic carbon source. 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 heterotrophic culture system of the photosynthetic microorganism may also be supplemented with other agents conventionally employed 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 certain sterile environment for heterotrophic culture, the gas used for aeration may be sterile, and antibiotics may be added to the heterotrophic 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 invention, the culture medium adopted by the culture system of the photosynthetic microorganism preferably consists of: 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, adding glucose-induced chlorella strain and 50mg/L kanamycin, introducing 1L/(L.min) sterile air at 28 deg.C and 250r/min, supplementing 15g/L glucose every day, supplementing other nutrient salts according to consumption, and heterotrophic culturing 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 3000lux, the illumination intensity is adjusted to 15000lux after 24h, and the illumination wavelength is 380-. Meanwhile, 2.4L of BG11 medium, 15g/L of glucose and 50mg/L of kanamycin which are sterilized are continuously added into the closed fermentation tank, the nutrition is supplemented every day, and the heterotrophic culture is continuously carried out in the closed fermentation tank by taking the rest part of algae liquid as algae seeds.
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 in the fermentor and tubular photobioreactor was measured at the indicated times with the simultaneous heterotrophic and autotrophic culture as shown in table 1, and the chlorophyll a/dry weight (weight ratio) of chlorella in the tubular photobioreactor was measured as shown in table 2.
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, sterilizing at 120 deg.C for 30min, cooling, inoculating, adding glucose-induced chlorella strain and 10mg/L chloramphenicol, introducing 0.8L/(L.min) sterile air at 28 deg.C and 250r/min, supplementing 15g/L glucose every day, supplementing other nutrient salts according to consumption, and culturing 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 2000lux, the illumination intensity is adjusted to 12000lux after 24h, and the illumination wavelength is 380-. Meanwhile, 2.4L of BG11 culture medium, 15g/L of glucose and 10mg/L of chloramphenicol and azithromycin which are sterilized are continuously added into the closed fermentation tank, the nutrition is supplemented every day, and the rest algae liquid is taken as algae seeds to be continuously cultured in a heterotrophic way 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 in the fermentor and tubular photobioreactor was measured at the indicated times with the simultaneous heterotrophic and autotrophic culture as shown in table 1, and the chlorophyll a/dry weight (weight ratio) of chlorella in the tubular photobioreactor was measured as shown in table 2.
Example 3
This example is intended to illustrate the method for culturing a photosynthetic microorganism of the present invention.
The method as described in example 1, except that, when the heterotrophic culture and the autotrophic culture are performed simultaneously, the autotrophic culture is performed by irradiating light at an irradiation intensity of 1000lux for 24 hours and then irradiating light at 20000lux for 24 hours; the dry weight of chlorella in the fermentor and tubular photobioreactor was measured at the indicated times with the simultaneous heterotrophic and autotrophic culture as shown in table 1, and the chlorophyll a/dry weight (weight ratio) of chlorella in the tubular photobioreactor was measured as shown in table 2.
Example 4
This example is intended to illustrate the method for culturing a photosynthetic microorganism of the present invention.
The method as described in example 1, except that, when the heterotrophic culture and the autotrophic culture are performed simultaneously, the autotrophic culture is performed by irradiating light at a light intensity of 5000lux for 24 hours and then irradiating light at 8000 lux; the dry weight of chlorella in the fermentor and tubular photobioreactor was measured at the indicated times with the simultaneous heterotrophic and autotrophic culture as shown in table 1, and the chlorophyll a/dry weight (weight ratio) of chlorella in the tubular photobioreactor was measured as shown in table 2.
Example 5
This example is intended to illustrate the method for culturing a photosynthetic microorganism of the present invention.
The method of example 1, except that the light wavelength for the autotrophic culture is full wavelength when the heterotrophic culture and the autotrophic culture are performed simultaneously; the dry weight of chlorella in the fermentor and tubular photobioreactor was measured at the indicated times with the simultaneous heterotrophic and autotrophic culture as shown in table 1, and the chlorophyll a/dry weight (weight ratio) of chlorella in the tubular photobioreactor was measured as shown in table 2.
Example 6
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 heterotrophic culture and the autotrophic culture are simultaneously performed, the light intensity in the autotrophic culture is not divided into two stages, but the culture is continuously performed under 20000lux light; the dry weight of chlorella in the fermentor and tubular photobioreactor was measured at the indicated times with the simultaneous heterotrophic and autotrophic culture as shown in table 1, and the chlorophyll a/dry weight (weight ratio) of chlorella in the tubular photobioreactor was measured as shown in table 2.
Example 7
According to the method described in example 1, except that the heterotrophic culture and the autotrophic culture are performed simultaneously, the fermentation tank is filled with air instead of the gas discharged from the tubular photobioreactor; the dry weight of chlorella in the fermentor and tubular photobioreactor was measured at the indicated times with the simultaneous heterotrophic and autotrophic culture as shown in table 1, and the chlorophyll a/dry weight (weight ratio) of chlorella in the tubular photobioreactor was measured as shown in table 2.
TABLE 1
Figure BDA0002374343150000131
TABLE 2
Figure BDA0002374343150000132
Figure BDA0002374343150000141
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 7 according to Table 1 with time is plotted as a chlorella growth curve shown in FIG. 2; chlorophyll a/dry weight values over time for chlorella in the tubular reactor of table 2 were plotted as shown in fig. 3.
As can be seen from the results shown in the table and the figure, the method of the invention can implement the autotrophic culture of the high-concentration photosynthetic microorganisms, thereby significantly improving the quality of the heterotrophic photosynthetic microorganisms; in addition, the present invention can also utilize the autotrophic culture of photosynthetic microorganisms to enhance the heterotrophic 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 (12)

1. A method for heterotrophic-autotrophic cultivation of photosynthetic microorganisms, characterized in that it is carried out in a reactor A and a reactor B connected in series, the method comprising:
(1) under a first gas source, sending photosynthetic microorganisms into a reactor A for heterotrophic 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 the condition of illumination and a second gas source to obtain a second culture solution.
2. The method according to claim 1, wherein the photosynthetic microorganism is a microalgae, preferably a green algae, more preferably a chlorella.
3. The method of any of claims 1-2, wherein the first gas source has a ventilation of 0.1-10L/(L-min), preferably 0.2-5L/(L-min);
preferably, the gas discharged from the reactor B is used as part or all of the first gas source.
4. A method according to any one of claims 1 to 3, wherein the second source has a ventilation of from 0.1 to 10L/(L-min), preferably from 0.2 to 5L/(L-min);
the second gas source adopts carbon dioxide-containing gas, and the carbon dioxide-containing gas is air or carbon dioxide-rich air.
5. The method according to any one of claims 1 to 4, wherein the illumination comprises first-stage illumination and 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.
6. The method according to any one of claims 1 to 5, wherein the temperature of the heterotrophic culture is between 20 ℃ and 35 ℃; the temperature of the autotrophic culture is 20-35 ℃.
7. A heterotrophic-autotrophic culture system for photosynthetic microorganisms, the system comprising: a reactor A and a reactor B, wherein the reactor A is set for heterotrophic culture of photosynthetic microorganisms; the reactor B is set to be used for autotrophic culture of photosynthetic microorganisms, wherein the reactor A comprises a first gas source device, the reactor B comprises a lighting device and a second gas source device,
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 heterotrophic culture in the reactor A.
8. The system of claim 7, wherein the gas outlet of reactor B is in communication with the first gas source device of reactor A, such that the gas discharged from reactor B serves as a gas source for some or all of the first gas source device.
9. The system of claim 7 or 8, wherein the exhaust port of the reactor B is sequentially connected with a gas collecting device and a pressure boosting device, and then is connected with the first gas source device of the reactor A, so that the gas discharged by the reactor B is collected and compressed to be used as the gas source of the first gas source device.
10. The system of any one of claims 7-9, wherein the reactor a is a fermentor structure; the reactor B is in a tubular, plate or column type photobioreactor structure.
11. A method for producing biomass, comprising cultivating a photosynthetic microorganism according to the method of any one of claims 1 to 6, and extracting biomass from the resulting photosynthetic microorganism.
12. A method of producing a bioenergy source, comprising culturing a photosynthetic microorganism using the method of any one of claims 1 to 6.
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