CN107937475B - Process for producing hydrogen by using scenedesmus obliquus and fungi in synergy - Google Patents

Process for producing hydrogen by using scenedesmus obliquus and fungi in synergy Download PDF

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CN107937475B
CN107937475B CN201810080741.3A CN201810080741A CN107937475B CN 107937475 B CN107937475 B CN 107937475B CN 201810080741 A CN201810080741 A CN 201810080741A CN 107937475 B CN107937475 B CN 107937475B
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黄永根
洪迪明
胡三强
白雪高
白健明
白永炜
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Jinan Xinjin hydrogen energy Co.,Ltd.
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Abstract

The invention belongs to the technical field of biology, and discloses a process for producing hydrogen by using scenedesmus obliquus and fungi in a synergistic manner, which comprises the following steps: step 1) preparing a compound bacterial liquid, step 2) preparing an algae liquid, step 3) preparing a hydrogen production culture solution, and step 4) synergistically producing hydrogen. The method has the advantages of simple and feasible process, high hydrogen production efficiency and wide application prospect.

Description

Process for producing hydrogen by using scenedesmus obliquus and fungi in synergy
Technical Field
The invention belongs to the technical field of biology, and particularly relates to a process for producing hydrogen by using scenedesmus obliquus and fungi in a synergistic manner.
Background
With the development of industrialization and the improvement of the living standard of human beings, the utilization of fossil energy is increased year by year, on one hand, the utilization causes serious environmental pollution, and on the other hand, the fossil energy is non-renewable. Therefore, it is urgent to find a clean and efficient renewable new energy source. Compared with other energy substances, hydrogen has a series of advantages of rich resources, small density, light weight, convenience in transportation, various utilization forms, wide application, high combustion value, cleanness, renewability, water generation after combustion and the like, so that the hydrogen is considered to be the most potential environment-friendly renewable energy source in the future society. With the further development of fuel cell technology, the direct conversion of hydrogen energy into electric energy can be completely realized, and the research on hydrogen production technology has great development potential. Thus, many countries around the world have invested a great deal of research expenditures for the development of hydrogen energy.
At present, the preparation of hydrogen mainly comprises the steps of preparing hydrogen by using fossil raw materials, preparing hydrogen by electrolyzing water, preparing hydrogen by organisms and the like. However, at present, hydrogen generation mainly depends on two ways, namely thermochemistry and photoelectrochemistry, which are not only expensive in production cost, large in energy requirement and serious in environmental pollution, but also emit a large amount of greenhouse gases, so that the method is non-environment-friendly, and the former needs to consume a large amount of valuable non-renewable resources such as coal, petroleum, natural gas and the like: the latter is high in consumption
The amount of electrical energy comes at a cost. In order to enable the hydrogen to be widely applied in the future, the key point is to establish a method for efficiently, simply, quickly and sustainably preparing the hydrogen. Therefore, making hydrogen a clean, renewable energy source in future society faces great challenges. Therefore, the preparation of hydrogen by using low-cost and renewable biomass resources as raw materials is receiving much attention.
Bio-hydrogen production is a new technology with such characteristics. The biological hydrogen-producing group mainly comprises: photosynthetic organisms (anaerobic photosynthetic bacteria, cyanobacteria, and green algae), non-photosynthetic organisms (strictly photosynthetic bacteria, facultative anaerobic bacteria, and aerobic bacteria), and archaeal flora. Because the hydrogen production mechanisms are different, the potential application value of the method is also evaluated differently. Currently, the research of biohydrogen production mainly focuses on photosynthetic bacteria and microalgae. Although the photosynthetic bacteria do not emit oxygen when hydrogen is emitted, the solar energy cannot be directly utilized to produce hydrogen; meanwhile, the application range of the organic matter for hydrogen production is limited due to factors such as the production area and the quantity of the organic matter, and the like, so that the hydrogen source is difficult to provide for human beings on a large scale. The hydrogen production of microalgae (blue algae and green algae) is realized by decomposing water into hydrogen and oxygen by using solar energy through a photosynthesis system and a special hydrogen producing enzyme, so that the hydrogen production method has a good application prospect.
The blue algae and the green algae are different in enzyme for catalyzing hydrogen production, the blue algae is nitrogen fixation enzyme for catalyzing hydrogen production, and energy ATP is consumed in the hydrogen production process, so that the overall hydrogen production efficiency is low. The reversible hydrogen-producing enzyme for catalyzing hydrogen production in green algae takes solar energy as energy and water as raw materials, and the theoretical quantum efficiency of catalyzing hydrogen production is high, so that the research on hydrogen production by green algae is most likely to become the subject of hydrogen production research in the future. Because the hydrogen production mechanisms of green algae are different, more high-hydrogen-yield algae species are obtained, and the hydrogen discharge characteristics of the algae species are researched. The hydrogenase of green algae is extremely sensitive to oxygen and is easily inhibited by oxygen to lose activity, and the oxygen is a specific product of photosynthesis, so that the hydrogen production efficiency of the green algae is low, and the development of the hydrogen production of the green algae is limited to a great extent, so that the content of oxygen in cells of the green algae needs to be reduced when the hydrogen production of the green algae is improved. At present, the method for reducing the oxygen content mainly comprises the steps of removing sulfur elements in a culture medium, so that the oxygen release activity of a photosynthetic system is inhibited, and the oxygen content generated by water photolysis is reduced. Therefore, to improve the hydrogen production efficiency of algae, the amount of oxygen in cells is reduced while ensuring the normal supply of electrons.
Disclosure of Invention
The invention aims to overcome the defects of low hydrogen production efficiency of algae and the like in the prior art and provides a process for producing hydrogen by using scenedesmus obliquus and fungi in a synergistic manner.
The invention is realized by the following technical scheme:
the process for producing hydrogen by using scenedesmus obliquus and fungi in a synergistic manner comprises the following steps: step 1) preparing a compound bacterial liquid, step 2) preparing an algae liquid, step 3) preparing a hydrogen production culture solution, and step 4) synergistically producing hydrogen.
Specifically, the process comprises the following steps:
step 1) preparing a compound bacterial liquid, namely respectively culturing enterococcus faecium and rhodopseudomonas palustris according to a conventional method to obtain 1 × 108Mixing the CFU/ml seed solution according to the volume ratio of 1:2 to obtain a mixed seed solution, transferring the mixed seed solution into a fermentation culture medium according to the inoculation amount of 10%, and culturing for 12 hours at 30 ℃ to obtain a compound bacterial solution;
step 2) preparing an algae solution: inoculating Scenedesmus obliquus in logarithmic growth phase into TAP culture solution according to the inoculation amount of 5-7% for culturing for 2-3 days to obtain algae solution;
step 3) preparing hydrogen production culture solution: adding sodium glycerophosphate and linoleic acid glyceride into the TAP culture solution to obtain a hydrogen production culture solution;
step 4), producing hydrogen cooperatively: transferring the hydrogen-producing culture solution into a reactor, and introducing N2Maintaining an anaerobic environment; then inoculating the algae liquid into the hydrogen production culture solution according to the volume ratio of 8-10%, performing dark culture for 12h, then inoculating the compound bacteria liquid into the hydrogen production culture solution according to the volume ratio of 0.5-1%, and performing synergistic culture for 72-96 h.
Preferably, the first and second electrodes are formed of a metal,
the formula of the fermentation medium comprises the following components in percentage by mass: molasses 3%, corn steep liquor 2.5%, soybean meal 2%, K2HPO40.1％、KH2PO40.1％、CaCO30.01％、FeSO40.005％、MnSO40.005% and the balance water, pH 7.0.
Preferably, the first and second electrodes are formed of a metal,
in the step 2), the culture conditions are as follows: the illumination intensity is 100 mu mol.m-2·s-1The ratio of light to dark is 12h to 12h, the water temperature is 25 ℃, and the pH value is 8.
Preferably, the first and second electrodes are formed of a metal,
the concentration of the sodium glycerophosphate is 0.5 mg/L.
Preferably, the first and second electrodes are formed of a metal,
the concentration of the linoleic acid glyceride is 1 mg/L.
Preferably, the first and second electrodes are formed of a metal,
in the step 4), the co-culture conditions are as follows: the illumination intensity is 100 mu mol.m-2·s-1The light-dark ratio is 12h:12 h.
The starting point and the beneficial effects of the invention mainly comprise but are not limited to the following aspects:
microalgae usually can release substances such as carbohydrates, amino acids, lipids and the like to the environment for bacteria to utilize during growth, meanwhile, the bacteria can also provide growth promoting factors such as inorganic nitrogen, phosphorus, carbohydrate substances, vitamins and the like for algae, and the algae and the bacteria form a commensal relationship; it is particularly difficult to select a proper strain for compatibility with algae.
Enterococcus faecium belongs to anaerobic bacteria and can be fermented to produce formate, acetate, organic acid and the like, and only hydrogen and carbon dioxide are produced in the fermentation process; the rhodopseudomonas palustris takes the substances as substrates, so that the hydrogen production effect is good;
the mixed bacteria liquid of the enterococcus faecium and the rhodopseudomonas palustris has good synergistic performance, can grow rapidly under the anaerobic illumination condition and can also grow under the aerobic condition, and the oxygen generated by the algae is effectively utilized, so that the oxygen content is reduced to the minimum, and the better hydrogen production capability is achieved; the carbon dioxide generated by the compound bacterial liquid can meet the requirement of the rapid growth of algae cells, and further the biomass of algae is improved.
The hydrogen yield of the invention is obviously superior to that of the method of singly adopting algae fermentation, the concentration of algae cells is obviously improved, the oxygen concentration is kept at a very low level, and the invention is beneficial to maintaining the activity of catalase and improving the hydrogen yield; the addition of a proper amount of chemical stimulant in the culture solution can improve the hydrogen production of algae cells and also has an obvious promotion effect on the biomass of the algae.
Drawings
FIG. 1: influence of the composite bacterial liquid on hydrogen production of algae.
Detailed Description
Those skilled in the art can modify the process parameters appropriately to achieve the desired results with reference to the disclosure herein. It is expressly intended that all such similar substitutes and modifications which would be obvious to one skilled in the art are deemed to be included in the invention. While the products and methods of this invention have been described in terms of preferred embodiments, it will be apparent to those of skill in the art that variations and modifications, or appropriate alterations and combinations, of the products and methods described herein may be made and utilized without departing from the spirit, scope, and spirit of the invention. For a further understanding of the present invention, reference will now be made in detail to the following examples.
Example 1
The process for producing hydrogen by using scenedesmus obliquus and fungi in a synergistic manner comprises the following steps:
preparing composite bacterial liquid, namely culturing enterococcus faecium and rhodopseudomonas palustris respectively according to a conventional method to obtain 1 × 108Mixing the CFU/ml seed solution according to the volume ratio of 1:2 to obtain a mixed seed solution, transferring the mixed seed solution into a fermentation culture medium according to the inoculation amount of 10%, and culturing for 12 hours at 30 ℃ to obtain a compound bacterial solution; the formula of the fermentation medium comprises the following components in percentage by mass: molasses 3%, corn steep liquor 2.5%, soybean meal 2%, K2HPO40.1％、KH2PO40.1％、CaCO30.01％、FeSO40.005％、MnSO40.005 percent, the balance being water, and the pH value being 7.0;
preparing an algae solution: inoculating Scenedesmus obliquus in logarithmic growth phase into TAP culture solution according to 5% inoculation amount, culturing, and controlling illumination intensity to be 100 μmol · m-2·s-1Culturing for 3 days at a light-dark ratio of 12h to 12h, a water temperature of 25 ℃ and a pH of 8 to obtainto a concentration of 8 × 106cell/mL of algae solution;
preparing hydrogen production culture solution: adding sodium glycerophosphate and linoleic acid glyceride into the TAP culture solution, and controlling the concentration of the sodium glycerophosphate to be 0.5mg/L and the concentration of the linoleic acid glyceride to be 1mg/L to obtain a hydrogen production culture solution;
and (3) synergistic hydrogen production: transferring the hydrogen-producing culture solution into a reactor, and introducing N2Maintaining an anaerobic environment; then inoculating the algae liquid into the hydrogen production culture solution according to the volume ratio of 8%, carrying out dark culture for 12h, then inoculating the composite bacterial liquid into the hydrogen production culture solution according to the volume ratio of 0.5%, and carrying out co-culture for 96h, wherein the co-culture conditions are as follows: the illumination intensity is 100 mu mol.m-2·s-1The light-dark ratio is 12h:12 h.
Example 2
The process for producing hydrogen by using scenedesmus obliquus and fungi in a synergistic manner comprises the following steps:
preparing composite bacterial liquid, namely culturing enterococcus faecium and rhodopseudomonas palustris respectively according to a conventional method to obtain 1 × 108Mixing the CFU/ml seed solution according to the volume ratio of 1:2 to obtain a mixed seed solution, transferring the mixed seed solution into a fermentation culture medium according to the inoculation amount of 10%, and culturing for 12 hours at 30 ℃ to obtain a compound bacterial solution; the formula of the fermentation medium comprises the following components in percentage by mass: molasses 3%, corn steep liquor 2.5%, soybean meal 2%, K2HPO40.1％、KH2PO40.1％、CaCO30.01％、FeSO40.005％、MnSO40.005 percent, the balance being water, and the pH value being 7.0;
preparing an algae solution: inoculating Scenedesmus obliquus in logarithmic growth phase into TAP culture solution according to inoculation amount of 7% for culturing, and controlling illumination intensity to be 100 μmol · m-2·s-1culturing for 2 days at a light-to-dark ratio of 12h to 12h, a water temperature of 25 deg.C and a pH of 8 to obtain a concentration of 3 × 106cell/mL of algae solution;
preparing hydrogen production culture solution: adding sodium glycerophosphate and linoleic acid glyceride into the TAP culture solution, and controlling the concentration of the sodium glycerophosphate to be 0.5mg/L and the concentration of the linoleic acid glyceride to be 1mg/L to obtain a hydrogen production culture solution;
and (3) synergistic hydrogen production: transferring hydrogen production culture solutionInto the reactor, N is introduced2Maintaining an anaerobic environment; then inoculating the algae liquid into a hydrogen production culture solution according to the volume ratio of 8-10%, carrying out dark culture for 12h, then inoculating the composite bacterial liquid into the hydrogen production culture solution according to the volume ratio of 1%, and carrying out co-culture for 72h, wherein the co-culture conditions are as follows: the illumination intensity is 100 mu mol.m-2·s-1The light-dark ratio is 12h:12 h.
Comparative example 1
The process for producing hydrogen by utilizing Scenedesmus obliquus comprises the following steps:
preparing an algae solution: inoculating Scenedesmus obliquus in logarithmic growth phase into TAP culture solution according to 5% inoculation amount, culturing, and controlling illumination intensity to be 100 μmol · m-2·s-1culturing for 3 days at a light-to-dark ratio of 12h to 12h, a water temperature of 25 deg.C and a pH of 8 to obtain a concentration of 8 × 106cell/mL of algae solution;
preparing hydrogen production culture solution: adding sodium glycerophosphate and linoleic acid glyceride into the TAP culture solution, and controlling the concentration of the sodium glycerophosphate to be 0.5mg/L and the concentration of the linoleic acid glyceride to be 1mg/L to obtain a hydrogen production culture solution;
hydrogen production: transferring the hydrogen-producing culture solution into a reactor, and introducing N2Maintaining an anaerobic environment; then inoculating the algae liquid into a hydrogen production culture solution according to the volume ratio of 8%, carrying out dark culture for 12h, and then continuing hydrogen production culture for 96h, wherein the hydrogen production culture conditions are as follows: the illumination intensity is 100 mu mol.m-2·s-1The light-dark ratio is 12h:12 h.
Comparative example 2
The process for producing hydrogen by utilizing Scenedesmus obliquus comprises the following steps:
preparing an algae solution: inoculating Scenedesmus obliquus in logarithmic growth phase into TAP culture solution according to 5% inoculation amount, culturing, and controlling illumination intensity to be 100 μmol · m-2·s-1culturing for 3 days at a light-to-dark ratio of 12h to 12h, a water temperature of 25 deg.C and a pH of 8 to obtain a concentration of 8 × 106cell/mL of algae solution;
hydrogen production: transferring TAP culture solution into a reactor, and introducing N2Maintaining an anaerobic environment; then inoculating the algae solution into hydrogen production culture solution according to the volume ratio of 8%, dark culturing for 12h, and continuing to produceHydrogen culture is carried out for 96h, and the hydrogen production culture conditions are as follows: the illumination intensity is 100 mu mol.m-2·s-1The light-dark ratio is 12h:12 h.
Example 3
Testing the hydrogen production performance:
setting the liquid volume of the reactor to be 8L, setting the space above the liquid to be 2L, multiplying the gas composition by the volume of the gas to obtain the gas quantity generated in the reaction process, and measuring the gas phase product in the reactor by using a gas chromatography;
the hydrogen production amount and the hydrogen content of example 1 and comparative examples 1 to 2 were measured; the concentration of algal cells in the culture broth was also determined by cytometry. See table 1 specifically:
TABLE 1
Group of Hydrogen (L) Oxygen amount (L) algal cell concentration (× 10)6cell/mL）
Example 1 0.226 0.018 13.7
Comparative example 1 0.061 0.039 8.9
Comparative example 2 0.039 0.031 6.4
And (4) conclusion: the hydrogen yield of the embodiment 1 of the invention is obviously better than that of the comparative example 1 and the comparative example 2, the concentration of the algae cells is also better than that of the comparative example 1 and the comparative example 2, the oxygen concentration of the embodiment 1 is kept at a low level, the activity of producing catalase is kept, the hydrogen yield is improved, and the table 1 shows that the hydrogen yield of the algae cells can be improved by the compound bacteria liquid and the chemical stimulant, and the obvious promotion effect on the biomass of algae is also provided.
Example 4
Influence of the composite bacterial liquid on hydrogen production of algae:
example 2 was used as the experimental group, and the detailed operation was performed according to example 3; a control group was also set, wherein control group 1: the procedure of example 2 was repeated except that enterococcus faecium was used; control group 2: the procedure of example 2 was followed except that Rhodopseudomonas palustris was used. As shown in fig. 1, the hydrogen production of the experimental group is significantly higher than that of the control group 1 and the control group 2 using a single strain, and since enterococcus faecium belongs to anaerobic bacteria, organic substances such as formate, acetate and organic acid can be produced by fermentation, and only hydrogen and carbon dioxide are produced during the fermentation process; the rhodopseudomonas palustris can take the substances as substrates, and the hydrogen production effect is good; the mixed bacteria liquid can grow rapidly under the anaerobic illumination condition and can also grow under the aerobic condition, so that oxygen generated by algae is effectively utilized, the oxygen content is reduced to the minimum, and better hydrogen production capacity is achieved; the compound bacterial liquid produced by the synergistic fermentation of the two bacterial strains can effectively promote the hydrogen production of algae; the carbon dioxide generated by the bacterial liquid can meet the requirement of the rapid growth of algae cells, and further the biomass of algae is improved.
Finally, it should be noted that: the above examples are only intended to illustrate the technical solution of the present invention, but not to limit it; although the present invention has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical solutions described in the foregoing embodiments may still be modified, or some technical features may be equivalently replaced; and such modifications or substitutions do not depart from the spirit and scope of the corresponding technical solutions of the embodiments of the present invention.

Claims (6)

1. The process for producing hydrogen by using scenedesmus obliquus and fungi in a synergistic manner is characterized by comprising the following steps of:
step 1) preparing a compound bacterial liquid, namely respectively culturing enterococcus faecium and rhodopseudomonas palustris according to a conventional method to obtain 1 × 108Mixing the CFU/ml seed solution according to the volume ratio of 1:2 to obtain a mixed seed solution, transferring the mixed seed solution into a fermentation culture medium according to the inoculation amount of 10%, and culturing for 12 hours at 30 ℃ to obtain a compound bacterial solution;
step 2) preparing an algae solution: inoculating Scenedesmus obliquus in logarithmic growth phase into TAP culture solution according to the inoculation amount of 5-7% for culturing for 2-3 days to obtain algae solution;
step 3) preparing hydrogen production culture solution: adding sodium glycerophosphate and linoleic acid glyceride into the TAP culture solution to obtain a hydrogen production culture solution;
step 4), producing hydrogen cooperatively: transferring the hydrogen-producing culture solution into a reactor, and introducing N2Maintaining an anaerobic environment; then inoculating the algae liquid into the hydrogen production culture solution according to the volume ratio of 8-10%, performing dark culture for 12h, then inoculating the compound bacteria liquid into the hydrogen production culture solution according to the volume ratio of 0.5-1%, and performing synergistic culture for 72-96 h.
2. The process according to claim 1, wherein the fermentation medium comprises the following components in percentage by mass: molasses 3%, corn steep liquor 2.5%, soybean meal 2%, K2HPO40.1％、KH2PO40.1％、CaCO30.01％、FeSO40.005％、MnSO40.005% and the balance water, pH 7.0.
3. The process according to claim 1, wherein in the step 2), the culture conditions are as follows: the illumination intensity is 100 mu mol.m-2·s-1The ratio of light to dark is 12h to 12h, the water temperature is 25 ℃, and the pH value is 8.
4. The process of claim 1, wherein the concentration of sodium glycerophosphate is 0.5 mg/L.
5. The process according to claim 1, wherein the glycerol linoleate concentration is 1 mg/L.
6. The process according to claim 1, wherein in the step 4), the co-culture conditions are as follows: the illumination intensity is 100 mu mol.m-2·s-1The light-dark ratio is 12h:12 h.
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