Algae-bacterium co-culture complex, culture method thereof, method for producing biomass by using algae-bacterium co-culture complex, biomass and related products

Abstract

The invention relates to the technical field of bioengineering, and discloses an algae-bacterium co-culture complex which is obtained by co-culturing microalgae and magnetotactic bacteria. The composition of the biological hydrocarbon on the surface of the microalgae can provide a relatively anaerobic growth environment for magnetotactic bacteria, and attachment and symbiosis of the microalgae and the magnetotactic bacteria are achieved. Meanwhile, magnetosome particles, glycopeptides and other substances synthesized by magnetotactic bacteria in cells can enter microalgae cells through endocytosis to promote hydrocarbon synthesis of the microalgae cells, so that hydrocarbon accumulation of the microalgae cells is increased; magnetotactic bacteria attached to the surface and magnetosomes in algae cells endow the microalgae with the characteristics of magnetic cells, the cells can be recovered through an external magnetic field, and the magnetic separation and recovery operation is simple, the separation speed is high, the energy consumption is low, and the recovery rate is high. The invention also discloses a culture method of the algae-bacterium co-culture complex, a method for producing biomass by using the algae-bacterium co-culture complex, the biomass and related products.

Description

Technical Field

[0001] The present application relates to the field of bioengineering technology, for example, to an algae-bacteria co-cultivation system and a cultivation method thereof, and a method for producing biomass using the same, biomass and related products. Background Art

[0002] With the increase in energy demand, the production and use of fossil fuels have caused serious environmental problems and carbon emission crises. Global warming caused by the rapid increase of greenhouse gases caused by industrialization has posed a major challenge to the sustainable development of the world, and has brought many problems including sea level rise, species extinction and biodiversity crisis, and food safety crisis. Photosynthesis is the process by which organisms use light energy as an energy source to absorb CO2 into the body and convert it into organic matter. It is an efficient means of CO2 fixation evolved in nature over billions of years, and it is also the driving force of the flow of matter and energy in the biosphere. In recent years, the use of photosynthetic autotrophic microalgae for CO2 biological fixation has attracted more and more attention. There are many advantages to using microalgae to fix carbon. High cell density can be obtained through photoautotrophic culture without competing for arable land. Secondly, many microalgae species usually have good tolerance to high concentrations of CO2 and can fix CO2 from different sources in an eco-friendly way. Bioactive substances such as lipids, proteins, vitamins and carotenoids produced by microalgae using CO2 can be used in food, feed, biofuels and pharmaceutical products, thereby reducing the cost of carbon fixation.

[0003] Microalgae can use light energy to convert CO2 and water into a variety of chemical substances, including sugars, proteins, lipids, vitamins, etc. through their own photosynthesis, which can be widely used in the production of food, medicines and food additives. Microalgae can also accumulate a large amount of oil and hydrocarbon substances, and is an excellent energy organism. Compared with other energy organisms, microalgae have the advantages of fast growth, high oil content, wide distribution range, and diverse energy forms. The composition of microalgae oil production does not contain sulfur elements, and no harmful and toxic gases are emitted when burned, which is beneficial to the ecological environment. Therefore, the use of microalgae to produce biofuels has important application prospects. The growth of microalgae can consume a large amount of carbon dioxide, and the production process from microalgae to oil can also achieve zero emissions, which has good environmental benefits and has great potential to alleviate the three major crises of food, energy and environment facing mankind. So far, at least the algae with the ability to produce hydrocarbons include Brown Staphylococcus aureus, Dunaliella, Chlorella, Inverted Cyclotria, Webwing Algae, and Cyclotria. The algae with the highest hydrocarbon content, the most research, and the most promising industrial algae are still Grape Algae.

[0004] The harvesting of microalgae has always been a bottleneck in the microalgae production process. Seeking an efficient, economical and suitable harvesting method for most microalgae is of great significance for the production of microalgae bioenergy.

[0005] It should be noted that the information disclosed in the above background technology section is only used to enhance the understanding of the background of the present application, and therefore may include information that does not constitute the prior art known to ordinary technicians in the field. Summary of the invention

[0006] In order to provide a basic understanding of some aspects of the disclosed embodiments, a brief summary is given below. The summary is not an extensive review, nor is it intended to identify key / critical components or delineate the scope of protection of these embodiments, but rather serves as a prelude to the detailed description that follows.

[0007] The disclosed embodiments provide an algae-bacteria co-culture complex and a culture method thereof, and a method for producing biomass using the same, biomass and related products. The algae-bacteria co-culture complex obtained by co-culturing microalgae and magnetotactic bacteria is endowed with the characteristics of magnetic cells, and the cell recovery is achieved through an external magnetic field, and the recovery is simple; and it can promote the synthesis of hydrocarbons, thereby increasing the accumulation of algae cell hydrocarbons.

[0008] In some embodiments, an algae-bacteria co-culture complex is obtained by co-culturing microalgae and magnetotactic bacteria.

[0009] In some embodiments, the method for culturing the algae-bacteria co-culture complex comprises: dispersing microalgae cells into a microalgae culture medium to obtain a microalgae cell suspension; inoculating a magnetotactic bacterial culture mixture into the microalgae cell suspension, co-culturing, and obtaining an algae-bacteria co-culture complex.

[0010] In some embodiments, the method of obtaining biomass using an algae-bacteria co-culture complex includes: adding an extractant to the algae-bacteria co-culture complex, and after extraction treatment, using magnetic force to separate the extracted algae-bacteria co-culture complex to obtain an extract; and processing the extract to obtain biomass.

[0011] In some embodiments, the biomass is obtained by extracting the aforementioned algae-bacteria co-culture complex; or, it is obtained by extracting the algae-bacteria co-culture complex obtained by culturing the aforementioned algae-bacteria co-culture complex; or, it is obtained by the aforementioned method of obtaining biomass using the algae-bacteria co-culture complex.

[0012] In some embodiments, the biodiesel comprises the aforementioned biomass.

[0013] In some embodiments, the aerospace kerosene comprises the aforementioned biomass.

[0014] In some embodiments, the feed includes the aforementioned algae-bacteria co-culture complex; or, includes the algae-bacteria co-culture complex obtained by culturing the aforementioned algae-bacteria co-culture complex by the aforementioned culturing method of the algae-bacteria co-culture complex; or, includes the aforementioned biomass.

[0015] The algae-bacteria co-cultivation complex and its cultivation method, and the method for producing biomass using the same, biomass and related products provided in the embodiments of the present disclosure can achieve the following technical effects:

[0016] The algae-bacteria co-culture complex of the disclosed embodiment is obtained by co-cultivating microalgae and magnetotactic bacteria in a preset ratio. The composition of biological hydrocarbons on the surface of the microalgae can provide a relatively anaerobic growth environment for magnetotactic bacteria, thereby achieving attachment and symbiosis between the two. At the same time, magnetosome particles, glycopeptides and other substances synthesized by magnetotactic bacteria in the cells can enter the microalgae cells through endocytosis. The entry of magnetotactic bacteria intracellular substances such as magnetosomes can cause stress reactions such as increased reactive oxygen species (ROS) in microalgae with the ability to produce biomass, promote the synthesis of its hydrocarbons (or oils), and thus increase the accumulation of algae cell biomass (hydrocarbons or oils). The magnetotactic bacteria attached to the surface and the magnetosomes in the algae cells give the microalgae the characteristics of magnetic cells, and the cells can be recovered by applying an external magnetic field. The magnetic separation and recovery operation is simple, the separation speed is fast, the energy consumption is low, and the recovery rate is high.

[0017] The cultivation method of the algae-bacteria co-cultivation complex of the disclosed embodiment is simple and easy, and the magnetism of the algae-bacteria co-cultivation complex makes it easy to recover and has a high recovery rate. In addition, in the process of obtaining biomass (biohydrocarbons, bio-oils, etc.) using the algae-bacteria co-cultivation complex, the magnetism of the extracted algae-bacteria co-cultivation complex also makes it easy to recover and has a high recovery rate.

[0018] The embodiments of the present disclosure include biomass-related products, such as bio-jet fuel, biodiesel, and feed, which reduce carbon emissions and improve environmental quality, and are green and environmentally friendly; after being added to feed, the nutritional value of the feed can be improved.

[0019] The above general description and the following description are exemplary and explanatory only and are not intended to limit the present application. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] One or more embodiments are exemplarily described by corresponding drawings, which do not limit the embodiments. Elements with the same reference numerals in the drawings are shown as similar elements, and the drawings do not constitute a scale limitation, and wherein:

[0021] Figure 1 It is a flowchart of a method for cultivating an algae-bacteria co-cultivation complex provided by an embodiment of the present disclosure;

[0022] Figure 2 It is a flowchart of a method for obtaining biomass by using an algae-bacteria co-cultivation complex provided by an embodiment of the present disclosure;

[0023] Figure 3is a flowchart of another method for obtaining biomass by using an algae-bacteria co-cultivation complex provided by an embodiment of the present disclosure;

[0024] Figure 4 is a graph showing the results of determination of biomass, total hydrocarbon production and total hydrocarbon yield of different culture systems of Example 3 of the embodiments of the present disclosure;

[0025] Figure 5 is a bar graph showing the determination of the recovery rate of algae magnetic separation at different algae-bacteria ratios in Example 3 of the present disclosure;

[0026] Figure 6 is a graph showing the results of determination of biohydrocarbon extraction rate and total hydrocarbon yield after re-cultivation under different monohydric alcohol extractants of Example 4 of the present disclosure;

[0027] Figure 7 is a graph showing the results of measuring the biomass hydrocarbon extraction rate and the total hydrocarbon yield after re-cultivation at different n-hexane to ethanol volume ratios of Example 4 of the present disclosure;

[0028] Figure 8 This is a graph showing the results of measuring the stability of hydrocarbons produced by the algae-bacteria co-cultivation system in Example 4 of the present disclosure. DETAILED DESCRIPTION

[0029] In order to be able to understand the features and technical contents of the embodiments of the present disclosure in more detail, the implementation of the embodiments of the present disclosure is described in detail below in conjunction with the accompanying drawings. The attached drawings are for reference only and are not used to limit the embodiments of the present disclosure. In the following technical description, for the convenience of explanation, a full understanding of the disclosed embodiments is provided through multiple details. However, one or more embodiments can still be implemented without these details. In other cases, to simplify the drawings, well-known structures and devices can be simplified for display.

[0030] The terms "first", "second", etc. in the specification and claims of the embodiments of the present disclosure and the above-mentioned drawings are used to distinguish similar objects, and are not necessarily used to describe a specific order or sequence. It should be understood that the terms used in this way can be interchanged where appropriate, so that the embodiments of the embodiments of the present disclosure described herein. In addition, the terms "including" and "having" and any variations thereof are intended to cover non-exclusive inclusions.

[0031] Unless otherwise stated, the term "plurality" means two or more.

[0032] In the embodiment of the present disclosure, the character " / " indicates that the preceding and following objects are in an "or" relationship. For example, A / B indicates: A or B.

[0033] The term "and / or" is a description of the association relationship between objects, indicating that three relationships can exist. For example, A and / or B means: A or B, or, A and B.

[0034] It should be noted that, in the absence of conflict, the embodiments and features in the embodiments of the present disclosure may be combined with each other.

[0035] The disclosed embodiment provides an algae-bacteria co-culture complex obtained by co-culturing microalgae and magnetotactic bacteria.

[0036] The algae-bacteria co-culture complex of the disclosed embodiment is obtained by co-culturing microalgae and magnetotactic bacteria in a preset ratio. The magnetotactic bacteria can synthesize magnetosome particles, glycopeptides and other substances composed of biofilm-coated, chain-arranged, nanoscale, single-domain magnetite (Fe3O4) in the cells, which have good biocompatibility and enter the microalgae cells through endocytosis. The entry of magnetotactic bacteria intracellular substances such as magnetosomes causes stress responses such as increased reactive oxygen species (ROS) in the microalgae cells, promoting the growth of the microalgae. The magnetotactic bacteria attached to the surface and the magnetosomes in the algae cells give the microalgae the characteristics of magnetic cells, and the cells are recovered / harvested by an external magnetic field. The recovery / harvesting of microalgae using magnetism is efficient, economical, and applicable to a variety of microalgae, which is of great significance to the production of microalgae, especially the production of microalgae bioenergy.

[0037] In the embodiments of the present disclosure, microalgae include microalgae capable of producing biomass, wherein biomass includes but is not limited to biohydrocarbons (hydrocarbon substances), biooils, sugars, proteins, lipids, vitamins, etc. The composition of biomass (biohydrocarbons / biooils, etc.) on the surface of microalgae can provide a relatively anaerobic growth environment for magnetotactic bacteria, achieving attachment and symbiosis of the two. In the algae-bacteria co-culture system, the integrity of the cell membrane of algae cells changes, and the intracellular and extracellular mass transfer capacity is enhanced, which is beneficial to the synthesis of metabolites of microalgae. Microalgae (e.g., Staphylococcus brownii) release extracellular metabolites such as polysaccharides and proteins during growth, which can be used by the surrounding magnetotactic bacteria to promote the growth of magnetotactic bacteria. Magnetotactic bacteria can synthesize magnetosome particles, glycopeptides and other substances composed of biofilm-coated, chain-arranged, nanoscale, single-domain magnetite (Fe3O4) in the cell, which have good biocompatibility and enter the microalgae cells through endocytosis. The entry of magnetotactic bacteria intracellular substances such as magnetosomes causes stress responses such as increased reactive oxygen species (ROS) in the microalgae cells, promoting the synthesis of hydrocarbons, thereby increasing the accumulation of hydrocarbons in the algae cells. The magnetotactic bacteria attached to the surface and the magnetosomes in the algae cells endow the microalgae with the characteristics of magnetic cells, and the cells can be recovered by applying an external magnetic field.

[0038] In the disclosed embodiment, after the algae-bacteria co-culture complex is co-cultured and biohydrocarbons are extracted through a suitable solvent system, the algae-bacteria co-culture complex can continue to be cultured in a culture medium to achieve the reuse of algae-bacteria cells and produce biohydrocarbons.

[0039] The disclosed embodiment constructs a stable and sustainable algae-bacteria co-culture complex for the production of biohydrocarbons by co-culturing microalgae and magnetotactic bacteria. The magnetotactic bacteria magnetosomes with good biocompatibility can promote the accumulation of microalgae biohydrocarbons and increase the biohydrocarbon yield. At the same time, the complex is endowed with magnetic properties, which can be separated by magnetic separation technology to achieve efficient recovery and reuse of cells. Moreover, magnetic separation has the advantages of simple operation, fast separation speed, and low energy consumption.

[0040] In the disclosed embodiments, microalgae include microalgae capable of producing biomass, for example, industrial algae species such as Botrytis cinerea, Dunaliella, Chlorella vulgaris, Cyclotella, Dictyopsis spp., Cyclotella and Chlorella spp.

[0041] Optionally, the microalgae include one or more of the following microalgae: Botryococcus brownii, Dunaliella, Chlorella, Cyclotella, Dictyoptera, Cyclotella and Nannochloropsis.

[0042] Optionally, the microalgae include brown staphylococcus algae. Brown staphylococcus algae is a freshwater or brackish water unicellular microalgae belonging to Chlorophyceae, Chlorophyceae, Chlorococcales, Brown staphylococcus family, Brown staphylococcus genus, containing products such as extracellular polysaccharides, fatty acids and hydrocarbons, and has a high hydrocarbon yield, which is higher than the hydrocarbon production of other microalgae and microorganisms, and the composition and structure of the hydrocarbons produced by Brown staphylococcus algae are extremely similar to petroleum, so Brown staphylococcus algae can be used to produce pollution-free, renewable petroleum substitutes, which has potential application value. Brown staphylococcus algae has a distinct cell wall, which is divided into two layers, the inner layer is composed of viscous polysaccharides and is fibrous, and the outer layer is formed by three thin layers to form a sheath. And Brown staphylococcus algae can convert 3% of solar energy into hydrocarbons, and the calorific value of Brown staphylococcus hydrocarbons can reach 30,000-42,000KJ / Kg. The hydrocarbons accumulated by Brown staphylococcus algae are stored on the outer wall of the cell.

[0043] In the embodiments of the present disclosure, the algae species of microalgae can be purchased, for example, from the Freshwater Algae Seed Bank of the Chinese Academy of Sciences (FACHB).

[0044] In the disclosed embodiment, the magnetotactic bacteria can be purchased, for example, from the China General Microorganism Collection Center.

[0045] Optionally, the magnetotactic bacteria include α-proteobacteria magnetotactic bacteria. The α-proteobacteria magnetotactic bacteria are easy to obtain, and the α-proteobacteria magnetotactic bacteria are all microaerobic or anaerobic bacteria, the optimal growth temperature is usually below 30°C, and they also have the characteristic of moving to low oxygen areas.

[0046] Optionally, the magnetotactic bacteria include magnetotactic spirochetes that are helical in shape.

[0047] Optionally, the magnetotactic bacteria include magnetotactic spirochetes of the class α-Proteobacteria, which is spiral in shape.

[0048] Optionally, the magnetotactic bacteria include Magnetospirillum. Magnetospirillum can be purchased. For example, the magnetotactic bacteria include Magnetospirillum gryphiswaldense, which can be purchased from the China General Microorganism Collection Center.

[0049] In some embodiments, the algae-bacteria co-culture complex is obtained by co-culturing microalgae and magnetotactic bacteria, including: co-culturing a microalgae cell suspension and a magnetotactic bacteria culture mixture according to a preset ratio. The preset ratio includes a volume ratio of the microalgae cell suspension to the magnetotactic bacteria culture solution of 100:1 to 1000:1. Optionally, the volume ratio of the microalgae cell suspension to the magnetotactic bacteria culture solution can be 100:1, 200:1, 300:1, 400:1, 500:1, 600:1, 700:1, 800:1, 900:1, 1000:1 or any ratio within the above volume ratio range.

[0050] In this embodiment, the microalgae cell suspension is a suspension obtained by dispersing microalgae cells in a microalgae culture medium. The dispersion ratio is not limited and is determined according to actual conditions. Optionally, in the microalgae cell suspension, the microalgae cell ratio is 2 to 5 g cell dry weight / L. Optionally, the microalgae cell ratio is 2, 3, 4, 5 or any value within 2 to 5, without limitation.

[0051] In this embodiment, the microalgae cells in the microalgae cell suspension are logarithmic growth phase microalgae cells collected when cultured to the logarithmic growth phase.

[0052] In this embodiment, the magnetotactic bacteria culture mixture is obtained by adding magnetotactic cells to a magnetotactic cell culture medium for culturing. Optionally, the magnetotactic bacteria culture mixture is obtained by adding magnetotactic cells to a magnetotactic cell culture medium for culturing to a logarithmic growth phase.

[0053] Optionally, the magnetotactic bacteria culture mixture is a magnetotactic bacteria culture mixture obtained by diluting the magnetotactic bacteria culture mixture stock solution when the magnetotactic bacteria are cultured to the logarithmic growth phase by a preset dilution multiple. In this embodiment, the preset dilution multiple is not limited and is determined based on actual conditions. Optionally, the preset dilution multiple is 2 to 6 times. Optionally, the preset dilution multiple is 2 to 5 times. Optionally, the preset dilution multiple is 3 to 4 times.

[0054] In some embodiments, the algae-bacteria co-culture complex is obtained by co-culturing microalgae and magnetotactic bacteria, and the co-culture conditions include one or more of the following three items: a culture temperature of 20 to 30° C.; a light intensity of 30 to 50 μmol / (m 2 ·s); the light-dark cycle is 14-20h(L) / 7-10h(D). Among them, 14-20h(L) / 7-10h(D) refers to a culture method in which the culture is alternating between culturing under light for 14-20h and culturing in darkness for 7-10h. That is, "L" represents light conditions and "D" represents dark conditions. It can be understood that the co-culture conditions are not limited to the above items and their ranges, and can be determined according to actual conditions during the actual co-culture process.

[0055] Optionally, the co-culture conditions include one or more of the following three items: a culture temperature of 23 to 28°C; a light intensity of 30 to 40 μmol / (m 2 ·s); light-dark cycle is 15-18h L / 7-9h D.

[0056] Optionally, the co-culture conditions include one or more of the following three items: a culture temperature of 25°C; a light intensity of 35 μmol / (m 2 ·s); the light-dark cycle was 16h(L) / 8h(D).

[0057] Combination Figure 1 As shown, the embodiment of the present disclosure provides a method for culturing an algae-bacteria co-cultivation complex, comprising the following steps:

[0058] S11, dispersing the microalgae cells into a microalgae culture medium to obtain a microalgae cell suspension;

[0059] S12, inoculating the magnetotactic bacteria culture mixture into the microalgae cell suspension, co-culturing, and obtaining an algae-bacteria co-culture complex.

[0060] In the cultivation method of the algae-bacteria co-culture complex of the disclosed embodiment, during the co-culture process of microalgae cells and magnetotactic bacteria, the magnetotactic bacteria and the microalgae attach and coexist, and at the same time, magnetosome particles and other substances synthesized by the magnetotactic bacteria in its cells enter the microalgae cells based on endocytosis, forming an algae-bacteria co-culture complex. In the algae-bacteria co-culture complex, the magnetotactic bacteria attached to the surface and the magnetosomes in the microalgae cells give the complex the characteristics of magnetic cells, and the recovery of the algae-bacteria co-culture complex is achieved by applying an external magnetic field.

[0061] In step S11 of the disclosed embodiment, the amount of dispersed microalgae cells in the microalgae cell suspension is not limited and is determined according to actual needs. In some embodiments, the cell dry weight concentration of the microalgae cell suspension in step S10 is 2 to 5 g / L. Optionally, the cell dry weight concentration is 2 g / L, 3 g / L, 4 g / L, 5 g / L or any value within 2 to 5 g / L, without limitation.

[0062] In step S11, the microalgae cells are logarithmic growth phase microalgae cells collected when they are in the logarithmic growth phase. That is, the logarithmic growth phase microalgae cells are microalgae cells collected when the microalgae are cultured and cultured to the logarithmic growth phase. The logarithmic growth phase generally refers to the growth stage in which the number of microalgae cells multiplies exponentially.

[0063] The microalgae culture medium in step S11 is not limited, and a conventional culture medium adapted to the type of microalgae is used. For example, when the microalgae includes Staphylococcus brownii, the microalgae culture medium uses Chu13×2 culture medium (Chu13 culture medium components are doubled). Optionally, the Chu13 culture medium also includes a mineral mixture; wherein the concentration of the mineral mixture is 2 to 10 mL / L. Optionally, the concentration of the mineral mixture is 4 to 8 mL / L. Optionally, the concentration of the mineral mixture is 5 to 6 mL / L.

[0064] In some embodiments, Figure 2 As shown, before step S11, step S10 is also included, adding microalgae to the microalgae culture medium, culturing under preset microalgae culture conditions, culturing for a first preset time, centrifuging, and obtaining microalgae cells. In this embodiment, the preset microalgae culture conditions are not limited, and the appropriate culture conditions can be determined according to the type of microalgae. The first preset time is also determined according to the type of microalgae. For example, when the microalgae is Staphylococcus brownii, the first preset time is 20 to 25 days. Within this time range, Staphylococcus brownii is in a logarithmic growth phase.

[0065] Optionally, in step S10, the preset microalgae culture conditions include one or more of the following: culture temperature of 20-30°C; rotation speed of 80-150 rpm; light intensity of 30-50 μmol / (m 2 ·s); the light-dark cycle is 14-20h(L) / 7-10h(D).

[0066] In this embodiment, the centrifugation operation in step S10 can be carried out under any suitable centrifugation conditions according to the type of microalgae. Optionally, the centrifugation conditions include: centrifugation at 6000-8000 g for 8-15 min. Optionally, in step S10, the centrifugation conditions include: centrifugation at 7000 g for 10 min.

[0067] In the disclosed embodiment, the algae species of the microalgae can be purchased, for example, from the freshwater algae species bank of the Institute of Hydrobiology, Chinese Academy of Sciences. The liquid algae species are purified to obtain sterile microalgae; and then the sterile microalgae are cultured in step S10 to obtain microalgae cells.

[0068] In step S12 of the disclosed embodiment, the magnetotactic bacteria culture mixture is obtained by adding magnetotactic bacteria to a magnetotactic bacteria culture medium for cultivation. In some embodiments, the method for obtaining the magnetotactic bacteria culture mixture comprises: adding magnetotactic bacteria to a magnetotactic bacteria culture medium, culturing for a second preset time under the culture conditions of 25 to 30° C. and 80 to 150 rpm, and obtaining the magnetotactic bacteria culture mixture. The second preset time is not limited and is determined according to the specific type of magnetotactic bacteria.

[0069] Optionally, the second preset time is the culture time required for the magnetotactic bacteria to be cultured to the logarithmic growth phase. In this embodiment, the magnetotactic bacteria culture mixture is a logarithmic phase culture mixture of magnetotactic bacteria in the logarithmic growth phase. For example, the OD of the magnetotactic bacteria culture mixture can be measured by 565 The OD value was tested to determine whether the magnetotactic bacteria entered the logarithmic growth phase. 565 A value in the range of 1.5 to 2.5 can determine that the magnetotactic bacteria are in the logarithmic growth phase.

[0070] Optionally, in step S12, the magnetotactic bacteria culture mixture is a magnetotactic bacteria culture mixture obtained by culturing the magnetotactic bacteria to the logarithmic growth phase and diluting the magnetotactic bacteria culture mixture by a preset dilution multiple. That is, the amount of bacteria in the magnetotactic bacteria culture mixture used for inoculation can be adjusted by dilution, the ratio of algae and fungi cells can be adjusted, and the co-cultivation effect can be improved. In this embodiment, the preset dilution multiple is not limited and can be determined based on actual conditions. Optionally, the preset dilution multiple is 2 to 6 times. Optionally, the preset dilution multiple is 2 to 5 times. Optionally, the preset dilution multiple is 3 to 4 times. Among them, the magnetotactic bacteria can be purchased, for example, they can be purchased from the China General Microorganism Collection Management Center.

[0071] Optionally, in step S12, the magnetotactic bacteria include magnetotactic bacteria of the class α-Proteobacteria.

[0072] Optionally, in step S12, the magnetotactic bacteria include magnetotactic spirochetes in a spiral shape.

[0073] Optionally, in step S12, the magnetotactic bacteria include magnetotactic spirochetes of the class α-Proteobacteria that are spiral in shape.

[0074] Optionally, the magnetotactic bacteria include Magnetospirillum. For example, the magnetotactic bacteria include Magnetospirillum gryphiswaldense, which can be purchased from China General Microorganism Collection Center.

[0075] In step S12 of this embodiment, the mixed culture solution of magnetotactic bacteria is inoculated into the microalgae cell suspension, and the volume ratio of the microalgae cell suspension to the culture solution of magnetotactic bacteria is controlled to be 100: 1 to 1000: 1. The appropriate ratio can ensure that the ratio of the growth or induction factors required by each is appropriate, ensuring the photosynthesis of the microalgae, and the appropriate ratio of magnetotactic bacteria can ensure the subsequent recovery and the enhancement of hydrocarbon and oil synthesis.

[0076] Optionally, the culture solution of magnetotactic bacteria is inoculated into the microalgae cell suspension, and the volume ratio of the microalgae cell suspension to the culture solution of magnetotactic bacteria is controlled to be 200:1 to 800:1.

[0077] Optionally, the culture solution of magnetotactic bacteria is inoculated into the microalgae cell suspension, and the volume ratio of the microalgae cell suspension to the culture solution of magnetotactic bacteria is controlled to be 400:1 to 600:1.

[0078] In some embodiments, in step S12, the magnetotactic bacteria culture mixture is inoculated into the microalgae cell suspension, co-cultured, and an algae-bacteria co-culture complex is obtained; including: inoculating the magnetotactic bacteria culture mixture into the microalgae cell suspension, co-culturing, and obtaining a co-culture system having an algae-bacteria co-culture complex; using magnetic force to separate the algae-bacteria co-culture complex in the co-culture system to obtain the algae-bacteria co-culture complex. In this embodiment, the algae-bacteria co-culture complex is separated from the culture solution of the culture system to facilitate the subsequent extraction of biomass.

[0079] Optionally, the algae-bacteria co-culture complex in the co-culture system is separated by magnetic force, including: placing a container containing the co-culture system on a strong magnetic magnet, the algae-bacteria co-culture complex is attracted by the magnet to achieve magnetic separation, and removing the supernatant to obtain the algae-bacteria co-culture complex.

[0080] The algae-bacteria co-culture complex of the disclosed embodiments may exist in any form and may be present in the co-culture system in a mixed form with the culture solution, or may be a separate algae-bacteria co-culture complex separated. It may be determined according to actual conditions.

[0081] In step S12 of the disclosed embodiment, in some embodiments, the co-culture conditions include one or more of the following: a culture temperature of 20 to 30° C., a light intensity of 30 to 50 μmol / (m 2·s); the light-dark cycle is 14-20h(L) / 7-10h(D); and the culture time is 5-10 days.

[0082] Optionally, the co-culture conditions include one or more of the following: a culture temperature of 23-28°C, a light intensity of 30-40 μmol / (m 2 ·s); the light-dark cycle is 15-18h(L) / 7-9h(D); and the culture time is 6-9 days.

[0083] Optionally, the co-culture conditions include one or more of the following: a culture temperature of 25°C, a light intensity of 35 μmol / (m 2 ·s); the light-dark cycle was 16h(L) / 8h(D); and the culture time was 8 days.

[0084] Combination Figure 3 As shown, the embodiment of the present disclosure also provides a method for obtaining biomass using an algae-bacteria co-cultivation complex, comprising the following steps:

[0085] S21, adding an extractant to the algae-bacteria co-culture complex, and after extraction, separating the extracted algae-bacteria co-culture complex by magnetic force to obtain an extract. The algae-bacteria co-culture complex is a simple algae-bacteria co-culture complex separated from the co-culture system.

[0086] S22, processing the extract to obtain biomass.

[0087] In the method for obtaining biomass in the embodiment of the present disclosure, the biomass in the algae-bacteria co-culture complex is extracted into an extracting solution by using an extractant, and the algae-bacteria co-culture complex is magnetically fixed by using magnetic force to separate from the extracting solution, and the separation operation is simple, so that the extracting solution can be processed to obtain biomass. Among them, biomass includes biohydrocarbons, biooils, sugars, proteins, lipids, vitamins, etc.

[0088] In the disclosed embodiment, the extractant in step S21 is determined according to the type of microalgae, without limitation. In some embodiments, the extractant includes a n-hexane-monohydric alcohol system, wherein the volume ratio of n-hexane to monohydric alcohol is 100:5 to 100:25. The n-hexane-monohydric alcohol system extractant of this embodiment can ensure the effective extraction of biomass.

[0089] Optionally, the volume ratio of n-hexane to monohydric alcohol is 100:10 to 100:20.

[0090] Optionally, the volume ratio of n-hexane to monohydric alcohol is 100:13 to 100:18.

[0091] Optionally, the volume ratio of n-hexane to monohydric alcohol is 100:15.

[0092] Optionally, the monohydric alcohol includes a monohydric alcohol having a carbon number of 1 to 3. For example, the monohydric alcohol includes methanol, ethanol, isopropanol, n-propanol, and the like.

[0093] In some embodiments, in step S21, the extract is processed to obtain biomass, including: vacuum evaporating the extract to remove the extractant, collecting crude hydrocarbon extracts; and purifying the crude hydrocarbon extracts to obtain biomass.

[0094] The purification process includes: purifying the crude hydrocarbon extract on a silica gel column, collecting all samples before the yellow band comes out, removing the elution solvent by vacuum evaporation, and blowing the remaining residue with an inert gas.

[0095] In some embodiments, Figure 3 As shown, the method for obtaining biomass using algae-bacteria co-culture complexes also includes: S23, adding the extracted algae-bacteria co-culture complexes separated by magnetic force in step S21 to a culture medium, resuspending to obtain a suspension, and re-culturing to obtain a re-cultured co-culture system; separating the re-cultured co-culture system by magnetic force to obtain the re-cultured algae-bacteria co-culture complex; S24, treating the re-cultured algae-bacteria co-culture complexes by the operations of steps S21 and S22 to obtain biomass and the extracted algae-bacteria co-culture complexes. The extracted algae-bacteria co-culture complexes obtained in step S24 can be re-cultured by step S23, and the culture is repeated in this way to achieve the cyclic acquisition of biomass and the reuse of the algae-bacteria co-culture complexes.

[0096] The embodiments of the present disclosure also provide a biomass obtained by extracting the algae-bacteria co-culture complex of any of the aforementioned embodiments; or, obtained by extracting the algae-bacteria co-culture complex obtained by culturing the algae-bacteria co-culture complex of any of the aforementioned embodiments; or, obtained by the method of obtaining biomass using the algae-bacteria co-culture complex of any of the aforementioned embodiments.

[0097] The biomass in the embodiments of the present disclosure includes, but is not limited to, biohydrocarbons (hydrocarbon substances), biooils, sugars, proteins, lipids, vitamins, etc., which can be extracted from the algae-bacteria co-culture complex according to actual needs.

[0098] The disclosed embodiment also provides a biodiesel, including the aforementioned biomass. The biodiesel of this embodiment is made from raw materials including the aforementioned biomass, is a "green energy", has good environmental performance, good engine starting performance, good fuel performance, and is renewable.

[0099] The disclosed embodiments also provide a bio-jet fuel, comprising the aforementioned biomass.

[0100] The bio-jet fuel of this embodiment is made from raw materials including the aforementioned biomass, which reduces carbon emissions and improves environmental quality. In this embodiment, the biomass includes bio-hydrocarbons and / or bio-oils.

[0101] The disclosed embodiments also provide a feed, including the algae-bacteria co-culture complex of any of the aforementioned embodiments; or, the algae-bacteria co-culture complex obtained by culturing the algae-bacteria co-culture complex of any of the aforementioned embodiments; or, the aforementioned biomass.

[0102] In the embodiments of the present disclosure, the algae-bacteria co-culture complex can be directly added to the feed, and the biomass content is high, making the feed more nutritious. The biomass extracted from the algae-bacteria co-culture complex can also be added to the feed as an additive to improve the nutritional value of the feed. The feed of the embodiments of the present disclosure includes animal feed.

[0103] Specific examples are provided below to more specifically illustrate the algae-bacteria co-cultivation system and its cultivation method and the method for producing biomass using the same according to the embodiments of the present disclosure.

[0104] Example 1 Obtaining Brown Staphylococcus aureus Cells

[0105] 100 μL of the liquid algae of Brown Staphylococcus aureus was evenly spread on a Chu13 solid plate containing 20-50 mg / L carbenicillin sodium for purification. After 10 days of culture, a single colony of Chlorella vulgaris was picked and streaked on a new Chu13 medium agar plate. This was repeated 3 times and examined under a microscope to obtain sterile Brown Staphylococcus aureus. The liquid algae of Brown Staphylococcus aureus was purchased from the Freshwater Algae Seed Bank of the Chinese Academy of Sciences (FACHB).

[0106] Chu13×2 medium (Chu13 medium with double components) was used, and the culture temperature, light intensity, light-dark cycle and rotation speed were 25℃, 35μmol / (m 2 ·s), 16h L / 8h D and 100rpm. After culturing the brown Staphylococcus algae for 3 weeks (the microalgae were in the logarithmic growth phase), the culture solution was centrifuged at 7000×g for 10 min to collect the brown Staphylococcus algae cells to obtain the brown Staphylococcus algae cells.

[0107] In this Example 1, the components and dosage of Chu13 culture medium are shown in Table 1 below.

[0108] Table 1

[0109] Element Concentration (g / L) Trace Elements Concentration (g / L) <![CDATA[KNO3]]> 0.2 <![CDATA[H3BO3]]> 0.572 <![CDATA[K2HPO4﹒3H2O]]> 0.052 <![CDATA[ZnSO4﹒7H2O]]> 0.044 <![CDATA[CaCl2]]> 0.0407 <![CDATA[CuSO4﹒5H2O]]> 0.016 <![CDATA[Na2-EDTA]]> 0.01 <![CDATA[Na2MoO4﹒2H2O]]> 0.012 Trace element solution 5mL <![CDATA[CoSO4﹒7H2O]]> 0.018

[0110] (Note: The total volume of the culture medium is 1000 mL, and 1.5% agar is added to the solid culture medium)

[0111] Example 2 Obtaining a mixed culture solution of magnetotactic bacteria

[0112] The magnetospirilla was added to the magnetotactic bacteria culture medium and cultured at 30°C and 100 rpm to obtain the logarithmic phase magnetospirilla culture solution. The magnetospirilla was purchased from the Magnetospirilla of Griffiswald Town of the China General Microorganism Collection Center. The culture medium was sterilized at 121°C for 20 minutes.

[0113] The culture medium formula used in this Example 2 is shown in Table 2 below.

[0114] Table 2

[0115]

[0116]

[0117] (Note: Add distilled water to 1L, and adjust the pH to 6.5-7 with 0.1mol / L NaOH)

[0118] Example 3

[0119] A method for culturing an algae-bacteria co-cultivation complex comprises the following steps:

[0120] S31, dispersing the brown Staphylococcus algae cells obtained in Example 1 into Chu13 medium (see Table 1 for specific formula) added with 5 mL / L of mineral mixture to obtain a brown Staphylococcus algae cell suspension, and adjusting the cell dry weight concentration to 2-5 g / L. For example, adjusting the cell dry weight concentration to 3 g / L.

[0121] S32, adding the culture solution of Magnetospirillum obtained in Example 2 and cultured to the logarithmic phase to the culture solution of Magnetospirillum to dilute and adjust the OD 565 to 0.5-1 (or, the dilution multiple is controlled to be 2-5) to obtain a culture solution of Magnetospirillum to be inoculated; the culture solution of Magnetospirillum to be inoculated is inoculated into a suspension of Staphylococcus brownii cells at a volume ratio of 100:1, 200:1, 400:1, 600:1 and 800:1 respectively, and co-culture is performed to obtain a co-culture system having an algae-bacteria co-culture complex; wherein the co-culture conditions include: a culture temperature of 25°C; a light intensity of 35 μmol / (m 2 ·s); the light-dark cycle was 16h(L) / 8h(D); and the culture time was 6 days.

[0122] S33, placing the co-culture system on a strong magnetic magnet, the algae-bacteria co-culture complex is attracted by the magnet, magnetic separation of cells is achieved, and the supernatant is removed to obtain the algae-bacteria co-culture complex.

[0123] In this embodiment 3, in step S32, the inoculation container may be a conical shake flask, which is convenient for subsequent co-culture and rotation culture.

[0124] In this embodiment 3, according to the different algae-bacteria ratios (i.e., inoculation volume ratios) in the co-cultivation system, the co-cultivation systems obtained after co-cultivation in step S32 are correspondingly recorded as co-cultivation system I (100:1), co-cultivation system II (200:1), co-cultivation system III (400:1), co-cultivation system IV (600:1), and co-cultivation system V (800:1). Correspondingly, the algae-bacteria co-cultivation complexes obtained after magnetic separation in step S33 of each co-cultivation system are respectively recorded as complex I, complex II, complex III, complex IV, and complex V.

[0125] In this embodiment 3, pure algae culture of a single microalgae is performed. The pure algae culture is to culture the Staphylococcus braunii cell suspension in step S31 under the co-culture conditions in step S32 to obtain a pure algae culture system.

[0126] In this Example 3, a pure bacterial culture of Magnetospirillum was also carried out. The pure bacterial culture was a pure bacterial culture system obtained by culturing in Example 2.

[0127] In this Example 3, the biomass and total algae hydrocarbons of the co-cultivation systems I to V, as well as the pure algae culture system and the pure bacteria culture system were measured (the measurement method is described in the corresponding content below). The measurement results are as follows: Figure 4 As shown, and the corresponding data as shown in Table 3.

[0128] And, based on the magnetic separation method of step S33, the magnetic separation recovery rate of algae was measured (the measurement method is described in the corresponding content below), specifically, an equal amount of co-culture solution (co-culture system, pure algae culture system, pure bacteria culture system of step S32) was taken into a beaker, and it was placed on a magnet for 5 seconds to allow the algae-bacteria co-culture complex (algae-bacteria cells) to quickly aggregate and stick to the bottom, and the supernatant was removed by magnetic separation, and the algae-bacteria co-culture complex was collected. Then, the steps were the same as the biomass measurement method, and the magnetic separation recovery biomass obtained by magnetic separation was measured, and the magnetic separation recovery rate of algae was calculated. The measurement results are as follows: Figure 5 As shown in Table 3, where magnetic separation recovery rate = (magnetic separation recovery biomass / system biomass) × 100%.

[0129] Table 3

[0130]

[0131]

[0132] As shown in Table 3, in the co-culture system III with a ratio of 400:1 of the inoculation volume of the brown Staphylococcus algae to the magnetotactic bacteria, the biomass (4.12 g / L), total hydrocarbon content (72.1%), and total hydrocarbon yield (3 g / L) of the brown Staphylococcus algae were not affected by co-culture, and the hydrocarbon content and hydrocarbon yield of the brown Staphylococcus algae were increased by more than 100%. In the co-culture system III with a ratio of 400:1 of the brown Staphylococcus algae to the magnetotactic bacteria, the recovery rate of the algae by magnetic separation of microalgae reached 98.5%.

[0133] Example 4

[0134] The method for obtaining biomass by using an algae-bacteria co-culture complex comprises the following steps:

[0135] S41. Add an n-hexane-monohydric alcohol system extractant to the algae-bacteria co-culture complex. After the extraction treatment, separate the extracted algae-bacteria co-culture complex by magnetic force to obtain an extract; wherein the algae-bacteria co-culture complexes are respectively complex I to complex V in Example 3, and correspondingly, extracts I to V are obtained respectively.

[0136] S42, treating the extract to obtain biogenic hydrocarbons (recorded as primary biogenic hydrocarbons).

[0137] S43, adding the extracted algae-bacteria co-culture complex obtained by magnetic separation in step S41 to Chu13 medium added with 5 mL / L of mineral mixture, resuspending, obtaining a suspension with a cell dry weight of 3 g / L, and re-culturing under the same culture conditions as the co-culture conditions in Example 3 to obtain a re-cultivated co-culture system; placing the re-cultivated co-culture system on a strong magnetic magnet, the re-cultivated algae-bacteria co-culture complex is attracted by the magnet, achieving magnetic separation, removing the supernatant, and obtaining a re-cultivated algae-bacteria co-culture complex;

[0138] S44, performing the operations of step S41 and step S42 on the re-cultured algae-bacteria co-culture complex obtained in step S43 to obtain biohydrocarbons (referred to as secondary biohydrocarbons) and the extracted algae-bacteria co-culture complex.

[0139] In this embodiment 4, the extracted algae-bacteria co-culture complex obtained in step S44 can be circulated to be re-cultured in step S43, and this process can be repeated to achieve the cyclic acquisition of biohydrocarbons and the reuse of the algae-bacteria co-culture complex.

[0140] In this Example 4, taking the complex III obtained by co-cultivation with an inoculation volume ratio of 400:1 of algae-bacteria ratio as an example, the extraction rate of biohydrocarbons of the algae-bacteria co-cultivation complex III (the volume ratio of n-hexane to monohydric alcohol is 100:15) was tested for step S41 (see the corresponding content below for the test method) and the total hydrocarbon production of the co-cultivation system after re-cultivation in step S43. The test results are as follows: Figure 6 As shown, and the data shown in Table 4.

[0141] Table 4

[0142]

[0143] Combination Figure 6 As can be seen from Table 4, the effects of different monohydric alcohols on the recovery rate of biohydrocarbons and the cell reuse performance of the algae-bacteria co-culture complex (algae-bacteria cells) show that ethanol has the highest recovery rate of biohydrocarbons for algae-bacteria cells, reaching 70.3%; after hydrocarbon extraction, the cells are reused, and the hydrocarbon production after cultivation reaches 2.96 g / L, with no significant decrease.

[0144] In this Example 4, taking the complex III obtained by co-cultivation with an inoculation volume ratio of 400:1 as an example, different n-hexane-ethanol system extractants with different volume ratios of n-hexane to ethanol were used to test the biohydrocarbon extraction rate of the algae-bacteria co-cultivation complex III (see the corresponding content below for the test method) and the total hydrocarbon production of the co-cultivation system after re-cultivation in step S43. The test results are as follows: Figure 7 As shown and the data shown in Table 5.

[0145] Table 5

[0146]

[0147] Combination Figure 7 From Table 5, it can be seen that the effects of different extractant solvent compositions on the hydrocarbon extraction rate and cell reuse performance show that under the 100:15 condition, the hydrocarbon recovery rate was 70.5%, and the hydrocarbon production of the algae-bacteria co-culture complex (algae-bacteria cells) after repeated culture after hydrocarbon extraction reached 2.96 g / L; under the 100:20 condition, the hydrocarbon recovery rate was increased to 75.9%, but the hydrocarbon production of the algae-bacteria cells after repeated culture after hydrocarbon extraction was significantly reduced to 2.01 g / L.

[0148] Meanwhile, in Example 4, taking the complex III obtained by co-cultivation with an inoculation volume ratio of 400:1 of algae-bacteria ratio as an example, the stability of hydrocarbons produced by repeated use of the algae-bacteria co-cultivation system was determined using an n-hexane-ethanol system extractant with a volume ratio of n-hexane to ethanol of 100:15. The results of the determination are as follows: Figure 8As shown. Specifically, the co-cultivation system of Example 3 is the first cycle, and the re-cultivated co-cultivation system obtained in step S43 is the second cycle. In this way, the extracted algae-bacteria co-cultivation complex obtained in step S44 is cycled to re-cultivate in step S43 and step S44, and the extraction operation in step S44 is performed for the third to tenth cycles in sequence. It can be seen that the stability of the hydrocarbons produced by the repeated use of the algae-bacteria co-cultivation system shows that the algae-bacteria co-cultivation complex is reused 10 times and extracted by a 100:15 n-hexane-ethanol system, and its hydrocarbon recovery rate is maintained at more than 70%. After the hydrocarbon extraction, the hydrocarbon production of cells after repeated culture is stable at 2.95-3.02 g / L, and no significant fluctuations are observed, indicating that the system has good reusability.

[0149] In the embodiments of the present disclosure, when the method of obtaining biomass using an algae-bacteria co-cultivation complex in Example 4 is used to extract biohydrocarbons from complex I, complex II, complex IV and complex V, the test results consistent with those of complex III recorded in Example 4 can be achieved, which will not be repeated here.

[0150] In the disclosed embodiment, the steps for determining the biomass of Staphylococcus aureus in the culture system are as follows: 100 mL of culture solution (the co-culture system, pure algae culture system, pure bacteria culture system in step S32 of Example 3) is taken and centrifuged at 7000×g for 10 min to collect cells, the cells are washed twice with deionized water and then repeatedly ground and extracted with 95% methanol, and then the absorbance of the methanol solution at 652 nm and 665 nm is measured, the chlorophyll content is calculated based on the absorbance value, and then the biomass is calculated based on the standard curve between chlorophyll and microalgae dry weight. The steps for determining the total hydrocarbons of microalgae are as follows: take an equal amount of culture solution (the co-culture system, pure algae culture system, and pure bacteria culture system in step S32 of Example 3) and centrifuge to collect algae cells or algae-bacteria cells, wash them twice and then freeze-dry them, freeze-dry algae powder or algae-bacteria solid (i.e., the dry weight of cells used for extraction), add n-hexane for ultrasonication for 15 minutes, centrifuge at 5000×g for 15 minutes to collect the supernatant n-hexane extract, repeat the extraction process 3 times, combine the n-hexane extracts, vacuum evaporate to remove the n-hexane, collect the crude hydrocarbon extract, purify it through a silica gel column, collect all samples before the yellow strip comes out, vacuum evaporate to remove the n-hexane, blow the remaining residue with nitrogen, weigh to obtain the hydrocarbon mass, calculate the hydrocarbon yield and cell hydrocarbon content, wherein, cell hydrocarbon content=(mass of hydrocarbons obtained by extraction / cell dry weight used for extraction)×100%, hydrocarbon yield=cell dry weight concentration of culture solution×cell hydrocarbon content.

[0151] In the disclosed embodiment, the steps for determining the biohydrocarbon extraction rate of the algae-bacteria co-culture complex (algae-bacteria cells) are as follows: take 100 mL of co-culture solution (the co-culture system of step S32 of embodiment 3), remove the supernatant by magnetic separation with a magnet, collect the algae-bacteria co-culture complex (algae-bacteria cells), wash twice and then freeze-dry, freeze-dry algae powder or algae-bacteria solid, add 3 times the volume of wet cell extractant, ultrasonicate for 15 minutes, magnetically separate algae-bacteria cells and collect the supernatant extract, vacuum evaporate to remove the extractant, collect crude hydrocarbon extract, purify by silica gel column, collect all samples before the yellow stripe comes out, vacuum evaporate to remove the elution solvent, blow nitrogen to the remaining residue, weigh to obtain the extracted hydrocarbon mass of different extractants, and obtain the extracted hydrocarbon amount. Take an equal amount of culture solution (the co-culture system of step S32 of embodiment 3) and determine the total hydrocarbon content according to the above-mentioned total hydrocarbon determination step (this step can be omitted, and the total hydrocarbon content obtained in embodiment 3 can be directly used), and calculate the extraction rate. Extraction rate = (extracted hydrocarbon amount / total hydrocarbon content) × 100%. Furthermore, the extracted wet algae-bacteria cells (i.e., the extracted algae-bacteria co-culture complex obtained in step S41) are added to the original culture volume of Chu13 culture medium with freshly added mineral mixture and cultured for 7 days. The total hydrocarbon production in the co-culture system after re-culture is determined by the aforementioned total hydrocarbon determination step to determine its reusability.

[0152] Example 5

[0153] A bio-jet fuel is prepared from raw materials including the bio-hydrocarbons extracted in Example 4. The bio-jet fuel of this example uses bio-hydrocarbons as raw materials, is green and environmentally friendly, reduces carbon emissions and improves environmental quality.

[0154] Example 6

[0155] A biodiesel is prepared from a raw material including the biohydrocarbon extracted in Example 4. The biodiesel of this example uses biohydrocarbon as a raw material, is green and environmentally friendly, and has the characteristics of good engine starting performance, good fuel performance, and renewability.

[0156] Example 7

[0157] A feed includes the algae-bacteria co-culture complex obtained by culture and separated by magnetic force in Example 3. The algae-bacteria co-culture complex obtained in Example 3 has a high content of biohydrocarbons, so the feed including it is more nutritious. The feed of this Example 7 includes animal feed.

[0158] Example 8

[0159] A feed comprising the biohydrocarbons extracted in Example 4. In the feed of this embodiment, the biohydrocarbons are added to the feed as an additive to improve the nutritional value of the feed. The feed of this embodiment 8 comprises animal feed.

[0160] The present application is not limited to the structures that have been described above and shown in the drawings, and various modifications and changes may be made without departing from the scope thereof. The scope of the present application is limited only by the appended claims.

[0161] In the present application, each embodiment may focus on the differences from other embodiments, and the same or similar parts between the embodiments may refer to each other.

Claims

1. An algae-bacteria co-culture complex, characterized in that: The algae-bacteria co-culture complex is obtained by co-culturing microalgae and magnetotactic bacteria.

2. The algae-bacteria co-cultivation system according to claim 1, characterized in that: Microalgae include microalgae that have the ability to produce biomass.

3. The algae-bacteria co-cultivation system according to claim 2, characterized in that: Microalgae capable of producing biomass include one or more of the following microalgae: Botrytis cinerea, Dunaliella, Chlorella, Cyclotella, Dictyopsis truncatula, Cyclotella and Nannochloropsis truncatula.

4. The algae-bacteria co-cultivation system according to claim 1, characterized in that: Magnetotactic bacteria include magnetotactic bacteria of the class α-Proteobacteria.

5. The algae-bacteria co-cultivation system according to claim 4, characterized in that: Magnetotactic bacteria of the class α-Proteobacteria include Magnetospirillum.

6. A method for cultivating an algae-bacteria co-cultivation complex, characterized in that: include: dispersing the microalgae cells into a microalgae culture medium to obtain a microalgae cell suspension; The magnetotactic bacteria culture mixture is inoculated into the microalgae cell suspension, and co-cultured to obtain an algae-bacteria co-culture complex.

7. The culture method according to claim 6, characterized in that: The microalgae cells are microalgae cells in the logarithmic growth phase that are collected when the cells are cultured to the logarithmic growth phase.

8. The culture method according to claim 6, characterized in that: The cell dry weight concentration of the microalgae cell suspension is 2 to 5 g cell dry weight / L.

9. The culture method according to claim 8, characterized in that: The cell dry weight concentration of the microalgae cell suspension is 3-4 g / L.

10. The culture method according to claim 6, characterized in that: The magnetotactic bacteria culture mixture is a magnetotactic bacteria culture stock solution obtained when the magnetotactic bacteria are cultured to a logarithmic growth phase.

11. The culture method according to claim 6, characterized in that: The magnetotactic bacteria culture mixture is prepared by diluting a magnetotactic bacteria culture mixture stock solution when the magnetotactic bacteria are cultured to a logarithmic growth phase by a preset dilution multiple.

12. The culture method according to claim 11, characterized in that: The preset dilution multiple is 2 to 6 times.

13. The culture method according to claim 6, characterized in that: The mixed culture solution of magnetotactic bacteria is inoculated into the microalgae cell suspension, and the volume ratio of the microalgae cell suspension to the culture solution of magnetotactic bacteria is controlled to be 100:1 to 1000:

1.

14. The culture method according to claim 13, characterized in that: The culture solution of magnetotactic bacteria is inoculated into the microalgae cell suspension, and the volume ratio of the microalgae cell suspension to the culture solution of magnetotactic bacteria is controlled to be 200:1 to 800:

1.

15. The culture method according to claim 13, characterized in that: The culture solution of magnetotactic bacteria is inoculated into the microalgae cell suspension, and the volume ratio of the microalgae cell suspension to the culture solution of magnetotactic bacteria is controlled to be 400:1 to 600:

1.

16. The culture method according to any one of claims 6 to 15, characterized in that: The co-culture conditions include one or more of the following: culture temperature is 20-30°C, light intensity is 30-50 μmol / (m 2 ·s); the light-dark cycle is 14-20h(L) / 7-10h(D); and the culture time is 5-10 days.

17. The culture method according to any one of claims 6 to 15, characterized in that: The mixed culture solution of magnetotactic bacteria is inoculated into the microalgae cell suspension, and co-cultured to obtain an algae-bacteria co-culture complex; comprising: The magnetotactic bacteria culture mixture is inoculated into a microalgae cell suspension for co-cultivation to obtain a co-cultivation system having an algae-bacteria co-cultivation complex; the algae-bacteria co-cultivation complex in the co-cultivation system is separated by magnetic force to obtain an algae-bacteria co-cultivation complex.

18. A method for obtaining biomass using an algae-bacteria co-culture complex, characterized in that: include: adding an extractant to the algae-bacteria co-culture complex, and after extraction treatment, separating the extracted algae-bacteria co-culture complex by magnetic force to obtain an extract; The extract is treated to obtain biomass.

19. The method for obtaining biomass according to claim 18, characterized in that: The extractant comprises a n-hexane-monohydric alcohol system, wherein the volume ratio of n-hexane to monohydric alcohol is 100:5 to 100:

25.

20. The method for obtaining biomass according to claim 19, characterized in that: The volume ratio of n-hexane to monohydric alcohol is 100:15 to 100:

20.

21. The method for obtaining biomass according to any one of claims 18 to 20, characterized in that: The monohydric alcohol includes monohydric alcohols having 1 to 3 carbon atoms.

22. The method for obtaining biomass according to any one of claims 18 to 20, characterized in that: The extract is processed to obtain biomass; including: The extract is subjected to vacuum evaporation treatment to remove the extractant, and crude hydrocarbon extract is collected; the crude hydrocarbon extract is then purified to obtain biomass.

23. The method for obtaining biomass according to any one of claims 18 to 20, characterized in that: Also includes: adding the extracted algae-bacteria co-culture complex separated by magnetic force into a culture medium, resuspending to obtain a suspension, and re-culturing to obtain a re-cultured co-culture system; The re-cultivated co-cultivation system is separated by magnetic force to obtain the re-cultivated algae-bacteria co-cultivation complex; The re-cultured algae-bacteria co-culture complex is processed to obtain biomass and extracted algae-bacteria co-culture complex.

24. A biomass, characterized in that: Extracted from the algae-bacteria co-culture complex according to any one of claims 1 to 5; or, Extracted from the algae-bacteria co-cultivation complex obtained by culturing the algae-bacteria co-cultivation complex according to any one of claims 6 to 17; or, The method is obtained by using the method for obtaining biomass using an algae-bacteria co-cultivation complex as described in any one of claims 18 to 23.

25. A biodiesel, characterized in that: Comprising the biomass of claim 24.

26. A bio-jet fuel, characterized in that: Comprising the biomass of claim 24.

27. A feed, characterized in that: comprising the algae-bacteria co-cultivation complex according to any one of claims 1 to 5; or, An algae-bacteria co-cultivation complex obtained by culturing the algae-bacteria co-cultivation complex according to any one of claims 6 to 17; or, comprising the biomass according to claim 24.

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