Method for producing hydrogen by anaerobic fermentation of blue-green algae based on pre-treatment synergism

Abstract

This invention discloses a method for hydrogen production through anaerobic fermentation of cyanobacteria based on pretreatment enhancement, belonging to the fields of environmental protection technology and biogas production technology. Persulfate and iron reagent are added to the cyanobacterial liquid, followed by ultrasonic oscillation-assisted treatment and oxidation treatment to obtain a pretreated cyanobacterial liquid. This pretreated cyanobacterial liquid is then subjected to anaerobic dark fermentation for hydrogen production. The process design of this invention enables cyanobacterial pretreatment to be completed under ambient temperature and pressure with stirring, and continuous and stable hydrogen production in a closed system. It truly achieves the dual benefits of waste-to-waste treatment and the conversion of waste into high-calorific-value clean energy, greatly reducing the environmental pressure caused by eutrophication of water bodies.

Description

Technical Field

[0001] This invention belongs to the fields of environmental protection technology and biogas production technology, and in particular relates to a method for hydrogen production by anaerobic fermentation of cyanobacteria based on pretreatment enhancement. Background Technology

[0002] With the increasing eutrophication of water bodies, cyanobacterial blooms are frequently occurring in lakes and reservoirs worldwide. To control these blooms, mechanical harvesting is commonly used, generating massive amounts of waste cyanobacterial biomass rich in moisture and organic matter. Developing effective strategies for the harmless disposal of cyanobacterial waste and improving its resource recovery rate is a major challenge for water environmental protection. Among numerous resource recovery pathways, anaerobic dark fermentation for hydrogen production has attracted considerable attention due to its ability to generate high-calorific-value and clean green hydrogen energy.

[0003] Current mainstream cyanobacterial anaerobic fermentation technologies typically face bottlenecks such as low substrate conversion rates and poor hydrogen production efficiency. This is mainly because cyanobacterial cells are encased in robust cell walls, hindering the release of soluble organic matter such as carbohydrates and proteins rich in the cells. Traditional direct fermentation or simple pretreatment methods often fail to completely break down the cell walls. Furthermore, during anaerobic fermentation, hydrogen-consuming microorganisms (such as methanogens) prevalent in the system rapidly consume the generated hydrogen, resulting in a low final hydrogen yield. Therefore, there is an urgent need to develop novel, highly efficient pretreatment technologies that can effectively suppress hydrogen-consuming bacteria in the fermentation system while breaking down the cyanobacterial cell walls to release carbon sources.

[0004] Persulfate / iron advanced oxidation technology is a commonly used method in water treatment, effectively disrupting cell structure through the free radicals generated during activation. Zero-valent iron (ZFI) is not only an excellent persulfate activator, but its reducing properties also provide a suitable environment for anaerobic fermentation. However, if not properly controlled, traditional persulfate / iron systems can easily mineralize organic matter into carbon dioxide and water, leading to a severe loss of valuable dissolved carbon sources required for fermentation. Simultaneously, conventional fermentation hydrogen production processes often require continuous addition of acids and alkalis to adjust pH, increasing engineering costs and operational complexity. Currently, there is a lack of a new method that eliminates the need for pH adjustment, precisely controls the persulfate to ZFI ratio to achieve efficient cell wall disruption and carbon release from cyanobacteria, and seamlessly integrates with long-term anaerobic hydrogen production. Further research and development of this safe, efficient, harmless, and high-value resource utilization technology is urgently needed. Summary of the Invention

[0005] To address the above problems, this invention provides a method for hydrogen production through anaerobic fermentation of cyanobacteria based on pretreatment enhancement.

[0006] To achieve the above-mentioned objectives, the present invention adopts the following technical solution: In a first aspect, the present invention provides a method for pretreatment of cyanobacteria, comprising the following steps: adding persulfate and iron reagent to cyanobacteria solution, followed by ultrasonic oscillation-assisted treatment and oxidation treatment to obtain pretreated cyanobacteria solution.

[0007] Furthermore, the total solids concentration of cyanobacteria in the cyanobacterial solution is 1~60 g / L.

[0008] Furthermore, the persulfate is selected from sodium persulfate, potassium persulfate, or ammonium persulfate, and the iron reagent is zero-valent iron powder. The zero-valent iron acts as an activator for the persulfate during the pretreatment stage.

[0009] Furthermore, the amount of persulfate added is 50 mmol per g of TSS cyanobacteria; the molar ratio of persulfate to zero-valent iron powder is 1:(1~2).

[0010] Furthermore, the molar ratio of the persulfate and zero-valent iron powder is 1:2.

[0011] Furthermore, the oxidation treatment is carried out at room temperature and pressure, with stirring for 2 to 48 hours.

[0012] This invention utilizes waste cyanobacteria as a substrate and persulfate and zero-valent iron (ZVFe) as advanced oxidation pretreatment reagents to obtain a pretreated cyanobacteria solution for anaerobic fermentation. On one hand, ZVFe activates persulfate to generate sulfate and hydroxyl radicals, efficiently disrupting the robust cell wall structure of cyanobacteria and promoting the release of abundant carbohydrates and protein-soluble organic matter within the cells. On the other hand, the pretreatment system not only effectively inactivates intolerant hydrogen-consuming methanogens and enriches tolerant hydrogen-producing bacteria, but also constructs a suitable reducing environment through anaerobic fermentation. This combined persulfate / ZVFe advanced oxidation dark fermentation process ultimately achieves the harmless treatment of harmful cyanobacteria blooms and the simultaneous production of high-value green energy.

[0013] Secondly, the present invention provides a pretreated cyanobacteria solution obtained by the aforementioned cyanobacteria pretreatment method.

[0014] Thirdly, the present invention provides a method for hydrogen production by anaerobic fermentation of cyanobacteria based on pretreatment enhancement, comprising the following steps: performing anaerobic dark fermentation of the pretreated cyanobacteria liquid to produce hydrogen.

[0015] Furthermore, the conditions for anaerobic dark fermentation hydrogen production are as follows: under sealed conditions, hydrogen production is carried out by shaking fermentation at 35~45°C.

[0016] Furthermore, the anaerobic dark fermentation hydrogen production time is 8 to 30 days.

[0017] The anaerobic dark fermentation hydrogen production is carried out under anaerobic conditions by pretreating the microbial community contained in the cyanobacteria solution itself. The cyanobacteria pretreatment method provided by this invention simultaneously inhibits the activity of hydrogen-consuming methanogenic bacteria in cyanobacteria and enriches a highly tolerant hydrogen-producing bacterial community.

[0018] This invention utilizes persulfate and zero-valent iron pretreatment technology to achieve deep cell wall disruption of cyanobacteria and precisely targets and screens the microbial community within the system, obtaining a pretreated cyanobacterial broth for direct hydrogen production through fermentation. The pretreated cyanobacterial broth prepared by this invention releases abundant soluble carbohydrates and proteins, providing ample substrate for hydrogen-producing bacteria. More importantly, the 2-72 hour pretreatment completely inactivates hydrogen-intolerant microorganisms while simultaneously promoting the enrichment and dominance of resistant hydrogen-producing bacteria, thus ensuring efficient hydrogen production in the subsequent fermentation system.

[0019] Compared with the prior art, the present invention has the following advantages and technical effects: 1. This invention provides a method for pretreatment of cyanobacteria, which achieves deep cell wall disruption of cyanobacteria using simple components and processes, without the need for complex and energy-intensive equipment, and realizes the recovery of green hydrogen energy while harmlessly treating harmful algal bloom waste.

[0020] 2. This invention uses simple components and processes to achieve a large release of organic matter from cyanobacteria cells. The pretreated cyanobacteria solution obtained after treatment has a significant increase in soluble carbohydrates and proteins, providing a sufficient high-quality carbon source for the long-term fermentation of hydrogen-producing bacteria.

[0021] 3. The cyanobacteria pretreatment method provided by this invention achieves efficient targeted screening and enrichment of microbial communities. The advanced oxidative stress lasting 2 to 72 hours completely inactivates hydrogen-consuming methanogenic bacteria and other microorganisms, while enabling highly tolerant hydrogen-producing bacteria to accumulate in large quantities after pretreatment. This fundamentally eliminates hydrogen-consuming pathways in the fermentation system and significantly increases the final cumulative hydrogen production.

[0022] 4. The process design of this invention can complete the pretreatment of cyanobacteria under normal temperature and pressure stirring, and continuously and stably produce hydrogen in a closed system, truly realizing the dual effects of treating waste with waste and converting waste into high-calorific-value clean energy, greatly reducing the environmental pressure caused by eutrophication of water bodies. Attached Figure Description

[0023] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0024] Figure 1 This is a SEM (scanning electron microscope) image of the original cyanobacteria in Example 1; Figure 2 This is a SEM (scanning electron microscope) image of cyanobacteria after persulfate / zero-valent iron pretreatment in Example 1; Figure 3 This is a comparison of the changes in LDH (lactate dehydrogenase) activity in the supernatant of the original cyanobacteria and the supernatant of the pretreated cyanobacteria in Example 1. Figure 4 This is a comparison of the soluble carbohydrate content in the supernatant of the original cyanobacteria and the supernatant of the pretreated cyanobacteria in Example 1. Figure 5 A comparison of the relative abundance of microbial communities at the genus level between the fermentation broth of step (3) in Example 1 and the fermentation broth of step (2) in Comparative Example 4. Figure 6 The cumulative hydrogen production curves for Examples 1-4 and Comparative Examples 1-4 during the 12-day anaerobic dark fermentation period are shown. Detailed Implementation

[0025] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0026] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Every smaller range between any stated value or intermediate value within a stated range, and any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0027] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0028] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be apparent to those skilled in the art. This specification and embodiments are merely exemplary.

[0029] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0030] This invention provides a method for pretreatment of cyanobacteria, comprising the following steps: adding persulfate and iron reagent to cyanobacteria solution, followed by ultrasonic oscillation-assisted treatment and oxidation treatment to obtain pretreated cyanobacteria solution.

[0031] In some preferred embodiments of the present invention, the total solids concentration of cyanobacteria in the cyanobacterial solution is 1~60 g / L.

[0032] In some preferred embodiments of the present invention, the persulfate is potassium persulfate, and the iron reagent is zero-valent iron powder.

[0033] In some preferred embodiments of the present invention, the amount of potassium persulfate added is 50 mmol per g of TSS cyanobacteria; the molar ratio of potassium persulfate to zero-valent iron powder is 1: (1~2).

[0034] In some preferred embodiments of the present invention, the molar ratio of potassium persulfate to zero-valent iron powder is 1:2.

[0035] In some preferred embodiments of the present invention, the oxidation treatment is carried out at room temperature and pressure with stirring for 2 to 48 hours.

[0036] This invention also provides a pretreated cyanobacteria solution obtained by the aforementioned cyanobacteria pretreatment method.

[0037] This invention also provides a method for hydrogen production by anaerobic fermentation of cyanobacteria based on pretreatment enhancement, comprising the following steps: performing anaerobic dark fermentation of the pretreated cyanobacteria liquid to produce hydrogen.

[0038] In some preferred embodiments of the present invention, the conditions for anaerobic dark fermentation to produce hydrogen are: under sealed conditions, the hydrogen is produced by shaking fermentation at 35~45°C.

[0039] In some preferred embodiments of the present invention, the anaerobic dark fermentation hydrogen production time is 8 to 30 days.

[0040] The cyanobacteria pretreatment method provided by this invention utilizes persulfate / zero-valent iron pretreatment. On one hand, this activates persulfate to generate free radicals, efficiently disrupting the robust cell wall structure of cyanobacteria and promoting the release of large amounts of soluble organic matter from the cyanobacterial cells. On the other hand, the pretreated system is enriched with a large number of hydrogen-producing bacteria, further enhancing hydrogen production. This combined cyanobacteria pretreatment and anaerobic fermentation process achieves the reduced-volume disposal of harmful cyanobacteria blooms and the efficient production of hydrogen as a green energy source, effectively improving the resource conversion rate of waste cyanobacteria and providing a new approach for the harmless and high-value utilization of cyanobacteria.

[0041] The room temperature / normal temperature in this invention refers to 20~25℃.

[0042] In this invention, atmospheric pressure refers to ambient atmospheric pressure, which is 101.325 kPa.

[0043] Example 1: A method for hydrogen production by anaerobic fermentation of cyanobacteria based on pretreatment enhancement (1) Take freshly harvested cyanobacteria (TSS), adjust its total solids concentration to 30 g / L, and obtain cyanobacteria solution; (2) Add potassium persulfate and zero-valent iron powder to the cyanobacteria solution in step (1), wherein the amount of potassium persulfate added is 50 mmol / g TSS and the amount of zero-valent iron added is 100 mmol / g TSS. After ultrasonic oscillation assisted treatment for 20 min, mechanical stirring at 300 rpm at room temperature and pressure is carried out for 48 h to obtain the pretreated cyanobacteria solution. (3) The pretreated cyanobacteria liquid obtained in step (2) is directly transferred to the anaerobic fermentation reactor without any external operation; the anaerobic fermentation reactor is sealed and placed in an air bath shaker at 40°C for dark fermentation to produce hydrogen for 12 days.

[0044] SEM (scanning electron microscope) observation of cyanobacteria after persulfate / zero-valent iron pretreatment in step (2) of Example 1, with the original cyanobacteria before treatment as a control, results are shown in [Figure 1]. Figure 1 SEM (scanning electron microscope) image of the original cyanobacteria in Example 1 and Figure 2 SEM (scanning electron microscope) image of cyanobacteria after persulfate / zero-valent iron pretreatment in Example 1.

[0045] Depend on Figure 1 and Figure 2 The SEM images show that the original cyanobacterial cells before treatment have an intact structure and a smooth surface. After pretreatment for 48 hours according to step (2) in Example 1, obvious holes and ruptures appeared on the surface of the cyanobacterial cells, and the cell structure was destroyed, indicating that the method can effectively break the strong cell wall of cyanobacteria.

[0046] The cyanobacterial solution obtained in step (1) was centrifuged to obtain the supernatant of the original cyanobacteria. The pretreated cyanobacterial solution obtained in step (2) was centrifuged to obtain the supernatant of the pretreated cyanobacteria. The LDH (lactate dehydrogenase) activity in the supernatants of the original and pretreated cyanobacteria was measured, with the supernatant of the original cyanobacteria (cyanobacteria) as a control. The results are shown in […]. Figure 3 Comparison of changes in LDH (lactate dehydrogenase) activity in the supernatant of the original cyanobacteria and the supernatant of the pretreated cyanobacteria in Example 1.

[0047] Depend on Figure 3 It can be seen that the LDH activity in the supernatant of pretreated cyanobacteria (PS / Fe pretreatment) increased sharply. Since LDH is a typical intracellular enzyme, its large-scale release into the supernatant further confirms the complete rupture of the cyanobacterial cell membrane.

[0048] The content of soluble carbohydrates in the supernatant of the original cyanobacteria and the supernatant of the pretreated cyanobacteria was determined, with the supernatant of the original cyanobacteria (cyanobacteria) as a control. The results are shown in [Figure number missing]. Figure 4 Comparison of soluble carbohydrate content in the supernatant of the original cyanobacteria and the supernatant of the pretreated cyanobacteria in Example 1.

[0049] Depend on Figure 4 It can be seen that the carbohydrate content in the supernatant of the pretreated cyanobacteria increased to 2.2 times that of the supernatant of the original cyanobacteria. This indicates that the cell wall disruption in step (2) promoted the release of a large amount of core nutrients from the cells, providing abundant high-quality substrates for hydrogen-producing bacteria.

[0050] Example 2: A method for hydrogen production by anaerobic fermentation of cyanobacteria based on pretreatment enhancement (1) Take freshly harvested cyanobacteria and adjust its total solids concentration to 30 g / L to obtain cyanobacteria solution; (2) Add potassium persulfate and zero-valent iron powder to the cyanobacteria solution in step (1), wherein the amount of potassium persulfate added is 50 mmol / g TSS and the amount of zero-valent iron added is 50 mmol / g TSS. After ultrasonic oscillation assisted treatment for 20 min, mechanical stirring at 300 rpm at room temperature and pressure is carried out for 48 h to obtain the pretreated cyanobacteria solution. (3) The pretreated cyanobacteria liquid obtained in step (2) is directly transferred to the anaerobic fermentation reactor without any external operation; the anaerobic fermentation reactor is sealed and placed in an air bath shaker at 40°C for dark fermentation to produce hydrogen for 12 days.

[0051] Example 3: A method for hydrogen production by anaerobic fermentation of cyanobacteria based on pretreatment enhancement (1) Take freshly harvested cyanobacteria and adjust its total solids concentration to 60 g / L to obtain cyanobacteria solution; (2) Add potassium persulfate and zero-valent iron powder to the cyanobacteria solution in step (1), wherein the amount of potassium persulfate added is 50 mmol / g TSS and the amount of zero-valent iron added is 50 mmol / g TSS. After ultrasonic oscillation assisted treatment for 20 min, mechanical stirring at 300 rpm at room temperature and pressure is carried out for 48 h to obtain the pretreated cyanobacteria solution. (3) The pretreated cyanobacteria liquid obtained in step (2) is directly transferred to the anaerobic fermentation reactor without any external operation; the anaerobic fermentation reactor is sealed and placed in an air bath shaker at 45°C for dark fermentation to produce hydrogen for 30 days.

[0052] Example 4: A method for hydrogen production by anaerobic fermentation of cyanobacteria based on pretreatment enhancement (1) Take freshly harvested cyanobacteria and adjust its total solids concentration to 1 g / L to obtain cyanobacteria solution; (2) Add potassium persulfate and zero-valent iron powder to the cyanobacteria solution in step (1), wherein the amount of potassium persulfate added is 50 mmol / g TSS and the amount of zero-valent iron added is 50 mmol / g TSS. After ultrasonic oscillation assisted treatment for 20 min, mechanical stirring at 300 rpm under normal temperature and pressure for 2 h is carried out to obtain pretreated cyanobacteria solution. (3) The pretreated cyanobacteria liquid obtained in step (2) is directly transferred to the anaerobic fermentation reactor without any external operation; the anaerobic fermentation reactor is sealed and placed in an air bath shaker at 35°C for dark fermentation to produce hydrogen for 8 days.

[0053] Comparative Example 1 Same as Example 1, except that in step (2), the amount of zero-valent iron added is 25 mmol / g TSS.

[0054] Comparative Example 2 Same as Example 1, except that zero-valent iron powder is not added in step (2).

[0055] Comparative Example 3 Same as Example 1, except that potassium persulfate is not added in step (2).

[0056] Comparative Example 4 (1) Take freshly harvested cyanobacteria and adjust its total solids concentration to 30 g / L to obtain cyanobacteria solution; (2) Transfer the cyanobacteria liquid from step (1) to the anaerobic fermentation reactor without any external operation; seal the anaerobic fermentation reactor and place it in an air bath shaker at 40°C for dark fermentation to produce hydrogen for 12 days.

[0057] The fermentation broth after step (3) of Example 1 was subjected to 16S rRNA high-throughput sequencing to analyze the relative abundance of genus-level microbial communities. The control group (cyanobacteria) that was directly fermented without pretreatment in Comparative Example 4 was used as a reference. The results are shown in […]. Figure 5 The relative abundance of microbial communities at the genus level was compared between the fermentation broth of step (3) in Example 1 and the fermentation broth of step (2) in Comparative Example 4.

[0058] Depend on Figure 5 The genus-level microbial abundance plot shows that the microbial community in the control group (cyanobacteria) was relatively chaotic, with a high proportion of hydrogen-consuming microorganisms. However, after the advanced oxidation pretreatment (PS / Fe pretreatment) of Example 1 and 12 days of fermentation, the relative abundance of Enterobacteriaceae, which have typical hydrogen-producing capabilities, increased significantly, becoming the dominant bacterial group, while the abundance of hydrogen-consuming microorganisms dropped to extremely low levels. This indicates that the 48-hour persulfate / zero-valent iron stress exerted a powerful directional screening effect, successfully enriching highly resistant hydrogen-producing bacteria and completely disrupting the hydrogen-consuming pathway.

[0059] The cumulative hydrogen production during 12 days of fermentation in Examples 1-4 and Comparative Examples 1-4 was measured, and the results are shown in the figure. Figure 6 Cumulative hydrogen production variation curves of Examples 1-4 and Comparative Examples 1-4 during 12 days of anaerobic dark fermentation.

[0060] Depend on Figure 6 As shown in the cumulative hydrogen production graph, the gas production process of all fermentation systems is mainly concentrated in the first 6 days of fermentation, after which the gas production rate gradually weakens and eventually stabilizes. The final cumulative hydrogen production efficiency of each group shows extremely significant differences, and the order of their final hydrogen production is as follows: Example 1 (PS / Fe = 1:2) > Example 3 (PS / Fe = 1:1) > Example 4 (PS / Fe = 1:1) > Example 2 (PS / Fe = 1:1) > Comparative Example 1 (PS / Fe = 1:0.5) = Comparative Example 2 (PS alone) = Comparative Example 3 (Fe alone) = Comparative Example 4 (cyanobacteria).

[0061] The above are merely preferred embodiments of the present invention, but the scope of protection of the present invention is not limited thereto. Any variations or substitutions that can be easily conceived by those skilled in the art within the scope of the technology disclosed in the present invention should be included within the scope of protection of the present invention.

Claims

1. A method for pretreating cyanobacteria, characterized in that, Includes the following steps: Persulfate and iron reagent were added to the cyanobacterial solution, and after ultrasonic oscillation-assisted treatment, oxidation treatment was carried out to obtain pretreated cyanobacterial solution.

2. The cyanobacteria pretreatment method according to claim 1, characterized in that, The total solids concentration of cyanobacteria in the cyanobacterial solution is 1~60 g / L.

3. The cyanobacteria pretreatment method according to claim 1, characterized in that, The persulfate is selected from sodium persulfate, potassium persulfate, or ammonium persulfate, and the iron reagent is zero-valent iron powder.

4. The cyanobacteria pretreatment method according to claim 3, characterized in that, The amount of persulfate added is 50 mmol per g of TSS cyanobacteria; the molar ratio of persulfate to zero-valent iron powder is 1:(1~2).

5. The cyanobacteria pretreatment method according to claim 4, characterized in that, The molar ratio of persulfate to zero-valent iron powder is 1:

2.

6. The cyanobacteria pretreatment method according to claim 1, characterized in that, The oxidation treatment is a stirring reaction at room temperature and pressure for 2 to 48 hours.

7. A pretreated cyanobacteria solution obtained by the cyanobacteria pretreatment method according to any one of claims 1 to 6.

8. A method for hydrogen production by anaerobic fermentation of cyanobacteria based on pretreatment enhancement, characterized in that, Includes the following steps: Hydrogen is produced by anaerobic dark fermentation of the pretreated cyanobacteria liquid according to claim 7.

9. The method for hydrogen production by anaerobic fermentation of cyanobacteria based on pretreatment enhancement according to claim 8, characterized in that, The conditions for anaerobic dark fermentation hydrogen production are as follows: under sealed conditions, hydrogen production is carried out by shaking fermentation at 35~45℃.

10. The method for hydrogen production by anaerobic fermentation of cyanobacteria based on pretreatment enhancement according to claim 8, characterized in that, The anaerobic dark fermentation hydrogen production time is 8 to 30 days.

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