微藻生物质暗发酵和光发酵耦合产氢气以及联产甲烷的机理研究
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摘要
化石燃料的过度利用导致了日趋严重的能源危机和环境污染。氢气的能量密度高、燃烧产物清洁,是一’种理想的二次能源载体。利用可再生的生物质为原料通过发酵的方法制取氢气已经成为国内外制氢领域的研究热点。微藻具有光合作用效率高、生长迅速、分布广泛等特点,是一种具有大规模能源化应用潜力的生物质原料。本文以微藻生物质为研究对象,对其暗发酵和光发酵耦合产氢气以及联产甲烷进行机理研究,实现生物质成分的高效分级利用,大幅度提高氢气产率和能量转化效率。
本文以微藻生物质中的典型蛋白质组分谷氨酸为原料,对谷氨酸的暗发酵和光发酵耦合产氢气联产甲烷的可行性进行了实验研究。在暗发酵、光发酵和甲烷发酵中使用的菌种均分离和富集于厌氧消化污泥,分别为混合产氢细菌、混合光合细菌和混合产甲烷细菌。在暗发酵阶段,谷氨酸可以被产氢细菌有效的利用和发酵产生大量的可溶代谢产物但是却很难产生氢气。暗发酵尾液中的主要代谢产物为乙酸、丁酸以及铵离子,由于高浓度的铵离子会显著抑制后续的光发酵产氢气,需要在光发酵之前去除尾液中的铵离子。沸石是一种廉价、可再生的天然资源,可以通过高效的离子交换去除溶液中的铵离子。经过沸石处理之后,尾液中的铵离子浓度由处理之前的36.7mM显著降低至3.2mM,铵离子去除率为91.1%。处理后的尾液接种光合细菌进行光发酵,得到最大氢气产率为292.9ml H2/g谷氨酸。光发酵尾液接种产甲烷细菌进行甲烷发酵,得到最大甲烷产率为102.7ml CH4/g谷氨酸。通过暗发酵和光发酵耦合产氢气联产甲烷,谷氨酸的能量转化效率由单纯产氢气的18.9%显著提高至氢气和甲烷联产的40.9%。
本文以微藻生物质中的典型碳水化合物成分海藻糖为原料,对海藻糖的暗发酵和光发酵耦合产氢气联产甲烷的可行性进行了实验研究。海藻糖是一种非还原性二糖,性质稳定很难水解,只有经过预处理水解为单糖以后才能高效的发酵产氢气。海藻糖经过微波加热辅助稀酸预处理之后接种产氢细菌进行暗发酵,得到最大氢气产率为396.2ml H2/g海藻糖。暗发酵尾液接种光合细菌进行光发酵,得到最大氢气产率为335.1ml H2/g海藻糖。光发酵的尾液接种产甲烷细菌进行甲烷发酵,得到最大甲烷产率为116.9ml H2/g海藻糖。通过暗发酵和光发酵耦合产氢气联产甲烷,海藻糖的能量转化效率由单纯产氢气的47.2%显著提高至氢气和甲烷联产的72.2%。
本文以钝顶节旋藻为原料,讨论和比较了微藻的两种暗发酵产氢气模式：利用外加产氢细菌的[FeFe]氢酶通过异相暗发酵产氢气和利用节旋藻的[NiFe]氢酶通过自相暗发酵产氢气。在异相暗发酵中,经过超声波破碎和酶水解之后的节旋藻可以被产氢细菌高效利用,在节旋藻浓度为20g/l得到最大的氢气产率为92.0ml H2/g DW。在自相暗发酵中,当节旋藻浓度由1g/l提高到20g/l时,最大氢气产率由51.4ml H2/g DW大幅度降低至11.0ml H2/gDW。在较高的节旋藻浓度条件下(20g/l),异相暗发酵的产氢峰值速率和最大氢气产率是自相暗发酵的110.0倍和8.4倍。因此在后续的实验中,微藻生物质应选用异相暗发酵产氢气的模式。节旋藻经过微波加热辅助稀酸预处理和酶水解可以有效的促进生物质的水解和强化暗发酵产氢气,随后利用沸石去除暗发酵尾液中的铵离子可以实现高效的光发酵产氢气。通过暗发酵和光发酵耦合产氢气,节旋藻的最大氢气产率大幅度提高至337.0mlH2/g DW。
本文以海洋微拟球藻为原料,讨论和比较了三种微藻生物质发酵产氢气和甲烷的方法：(1)暗发酵和光发酵耦合产氢气联产甲烷；(2)暗发酵产氢气联产甲烷；(3)直接发酵产甲烷。微拟球藻通过微波加热辅助稀硫酸预处理之后接种产氢细菌进行暗发酵产氢气。在暗发酵中产氢细菌对大部分经过预处理水解生成的氨基酸的消耗时间大约是还原糖消耗时间的两倍。微拟球藻通过暗发酵和光发酵耦合产氢气联产甲烷得到的最大氢气产率为161.3ml H2/g TVS,最大甲烷产率为183.9ml CH4/g TVS,整体能量转化效率为暗发酵产氢气联产甲烷的1.7倍、直接产甲烷的1.3倍。
本文以蛋白核小球藻为原料,研究了多种预处理方式对微藻生物质暗发酵产氢气的影响。经过蒸汽加热辅助稀酸和微波加热辅助稀酸预处理能够显著促进小球藻的水解和暗发酵产氢气。通过暗发酵和光发酵耦合产氢气联产甲烷,小球藻的最大氢气产率为198.3ml H2/g TVS,最大甲烷产率为186.2ml CH4/g TVS,整体能量转化效率为34.0%。在批次实验的基础之上,进行了小球藻的半连续流发酵产氢气的实验研究。通过长时间小球藻驯化得到的复杂产氢细菌菌群比之前通过葡萄糖驯化得到的简单菌群更能适应和高效利用小球藻生物质的各个成分,可以实现连续稳定的发酵产氢气。为了进一步提高生物质发酵的能量转化效率,以小球藻和木薯淀粉为混合原料,研究了碳氮摩尔比对发酵产氢气的影响。混合生物质在碳氮摩尔比为25.3的条件下得到的最大暗发酵氢气产率为276.2ml H2/gTVS,分别是单纯用小球藻和木薯淀粉为原料最大氢气产率的3.7倍和1.8倍。通过暗发酵和光发酵耦合产氢气联产甲烷,混合生物质的最大氢气产率和甲烷产率分别为664.2ml H2/g TVS和126.0ml CH4/g TVS,整体能量转化效率达到67.2%。
The extensive utilization of fossil fuels has resulted in serious energy crisis and environmental pollution. Hydrogen is considered as an ideal carbon-free secondary energy carrier with high energy density and clean combustion product. Hydrogen production from renewable biomass through fermentation is increasingly attracting worldwide attention. Microalgae biomass is a potential feedstock for fermentative hydrogen production because of its high photosynthetic efficiency, fast growth, and global distribution. In this study, microalgae biomass was used as feedstock to cogenerate hydrogen and methane through a novel three-stage method comprising dark-fermentation, photo-fermentation, and methanogenesis. The components in microalgae biomass were efficiently used through the three-stage method, therefore hydrogen yield and energy conversion efficiency (ECE) were significantly increased.
Glutamic acid, a typical amino acid degraded from protein components in microalgae biomass, was used as feedstock to investigate the feasibility of cogeneration of hydrogen and methane through the three-stage method comprising dark-fermentation, photo-fermentation, and methanogenesis. Hydrogen-producing bacteria (HPB), photosynthetic bacteria (PSB), and methane-producing bacteria (MPB) were used as the inocula during dark-fermentation, photo-fermentation, and methanogenesis, respectively. HPB can efficiently ferment glutamic acid to abundant soluble metabolite products (SMPs) and little hydrogen during dark-fermentation. The residual solution of dark-fermentation mainly contained acetate, butyrate, and ammonium. Because high concentration of ammonium (36.7mM) in the residual solution of dark-fermentation can significantly inhibit the activities of PSB in sequential photo-fermentation, a modified zeolite were used to extract ammonium by ion exchange to reduce the ammonium concentration to3.2mM (91.1%of ammonium removal efficiency). After ammonium removal, the treated solution was inoculated with PSB, exhibiting the maximum hydrogen yield of292.9ml H2/g glutamic acid during photo-fermentation. The residual solution from photo-fermentation was reused by MPB to produce the maximum methane yield of102.7ml CH4/g glutamic acid. The ECE from glutamic acid to gas fuels significantly increased from18.9%in hydrogen fermentation to40.9%in combined hydrogen fermentation and methanogenesis.
Trehalose, a typical carbohydrate component in microalgae biomass, was used as feedstock to investigate the feasibility of cogeneration of hydrogen and methane through the three-stage method comprising dark-fermentation, photo-fermentation, and methanogenesis. As a stable non-reducing sugar, trehalose was not easily used by HPB for efficient hydrogen production. Trehalose was first pretreated by microwave heating with dilute acid, and then was inoculated with HPB to produce hydrogen during dark-fermentation. The residual solution of dark-fermentation was reused by PSB during photo-fermentation. The residual solution of photo-fermentation was reused by MPB during methanogenesis. Overall, the maximum hydrogen yield of731.3ml H2/g trehalose and methane yield of116.9ml CH4/g trehalose were achieved. The sequential generation of hydrogen and methane from trehalose remarkably enhanced the ECE from47.2%in hydrogen fermentation to72.2%in combined hydrogen fermentation and methanogenesis.
Hydrogen production from Arthrospira platensis biomass through dark-heterofermentation by the [FeFe] hydrogenase of HPB and dark-auto fermentation by the [NiFe] hydrogenase of A. platensis was discussed. A. platensis biomass pretreated by ultrasonication and enzymatic hydrolysis was inoculated with HPB to produce hydrogen during dark-heterofermentation. The maximum hydrogen yield of92.0ml H2/g dry weight (DW) was obtained at20g/l of A. platensis biomass. In dark-autofermentation, hydrogen yield decreased from51.4ml H2/g DW to11.0ml H2/g DW with increasing substrate concentration from1g/1to20g/1. The hydrogen production peak rate and maximum hydrogen yield at20g/1of A. platensis biomass in dark-heterofermentation showed110.0-and8.4-fold increases, respectively, relative to those in dark-autofementation. Therefore, dark-heterofermentation was selected for the further investigation of fermentative hydrogen production from microalgae biomass. A. platensis biomass was pretreated by microwave heating with dilute acid to improve saccharification during enzymatic hydrolysis and hydrogen production during dark-fermentation. The residual solution of dark-fermentation was treated by zeolite to reduce ammonium concentration before photo-fermentation. The maximum hydrogen yield from A. platensis biomass was significantly increased to337.0ml H2/g DW through combined dark-fermentation and photo-fermentation.
Three methods for hydrogen and methane production from Nannochloropsis oceanica biomass were discussed as the following:(1) three-stage method comprising dark-fermentation. photo-fermentation, and methanogenesis;(2) two-stage comprising dark-fermentation and methanogenesis;(3) single-stage methanogenesis. N. oceanica pretreated by microwave heating with dilute acid was inoculated with HPB to produce hydrogen during dark-fermentation.The consumption time of most amino acids was about2times as long as that of most reducing sugars during dark-fermentation. The total ECE from N. oceanica biomass to gas fuels through the three-stage method showed1.7-and1.3-fold increases, respectively, compared with those through the two-stage and single-stage methods.
Effects of pretreatment methods on biomass saccharification and hydrogen fermentation from Chlorella pyrenoidosa were investigated. The steam heating with dilute acid and microwave heating with dilute acid can remarkably enhance the biomass hydrolysis and hydrogen fermentation. The maximum hydrogen yield of198.3H2ml/g total volatile solids (TVS) and methane yield of186.2ml H2/g TVS were achieved through the three-stage method comprising dark-fermentation, photo-fermentation, and methanogenesis. Semi-continuous fermentation of C. pyrenoidosa biomass was carried out based on batch fermentation. Compared with the simple microbial community formed at earlier stages of fermentation, the complex microbial community formed at later stages of fermentation was more adaptable to C. pyrenoidosa biomass and can utilize C. pyrenoidosa biomass more efficiently, thereby resulting in efficient and stable hydrogen fermentation. In order to enhance the ECE from C. pyrenoidosa biomass, cassava starch was mixed with C. pyrenoidosa biomass to optimize the carbon/nitrogen (C/N) molar ratio for efficient dark-fermentation. The maximum dark hydrogen yield of276.2ml H2/g TVS from the mixed biomass at C/N molar ratio of25.3showed3.7-and1.8-fold increases, respectively, compared with those from only C. pyrenoidosa biomass and only cassava starch. The maximum hydrogen yield of664.2H2ml/g TVS and methane yield of126.0ml H2/g TVS corresponding to the total ECE of67.2%were achieved through the three-stage method comprising dark-fermentation, photo-fermentation, and methanogenesis.
本文以微藻生物质中的典型蛋白质组分谷氨酸为原料,对谷氨酸的暗发酵和光发酵耦合产氢气联产甲烷的可行性进行了实验研究。在暗发酵、光发酵和甲烷发酵中使用的菌种均分离和富集于厌氧消化污泥,分别为混合产氢细菌、混合光合细菌和混合产甲烷细菌。在暗发酵阶段,谷氨酸可以被产氢细菌有效的利用和发酵产生大量的可溶代谢产物但是却很难产生氢气。暗发酵尾液中的主要代谢产物为乙酸、丁酸以及铵离子,由于高浓度的铵离子会显著抑制后续的光发酵产氢气,需要在光发酵之前去除尾液中的铵离子。沸石是一种廉价、可再生的天然资源,可以通过高效的离子交换去除溶液中的铵离子。经过沸石处理之后,尾液中的铵离子浓度由处理之前的36.7mM显著降低至3.2mM,铵离子去除率为91.1%。处理后的尾液接种光合细菌进行光发酵,得到最大氢气产率为292.9ml H2/g谷氨酸。光发酵尾液接种产甲烷细菌进行甲烷发酵,得到最大甲烷产率为102.7ml CH4/g谷氨酸。通过暗发酵和光发酵耦合产氢气联产甲烷,谷氨酸的能量转化效率由单纯产氢气的18.9%显著提高至氢气和甲烷联产的40.9%。
本文以微藻生物质中的典型碳水化合物成分海藻糖为原料,对海藻糖的暗发酵和光发酵耦合产氢气联产甲烷的可行性进行了实验研究。海藻糖是一种非还原性二糖,性质稳定很难水解,只有经过预处理水解为单糖以后才能高效的发酵产氢气。海藻糖经过微波加热辅助稀酸预处理之后接种产氢细菌进行暗发酵,得到最大氢气产率为396.2ml H2/g海藻糖。暗发酵尾液接种光合细菌进行光发酵,得到最大氢气产率为335.1ml H2/g海藻糖。光发酵的尾液接种产甲烷细菌进行甲烷发酵,得到最大甲烷产率为116.9ml H2/g海藻糖。通过暗发酵和光发酵耦合产氢气联产甲烷,海藻糖的能量转化效率由单纯产氢气的47.2%显著提高至氢气和甲烷联产的72.2%。
本文以钝顶节旋藻为原料,讨论和比较了微藻的两种暗发酵产氢气模式：利用外加产氢细菌的[FeFe]氢酶通过异相暗发酵产氢气和利用节旋藻的[NiFe]氢酶通过自相暗发酵产氢气。在异相暗发酵中,经过超声波破碎和酶水解之后的节旋藻可以被产氢细菌高效利用,在节旋藻浓度为20g/l得到最大的氢气产率为92.0ml H2/g DW。在自相暗发酵中,当节旋藻浓度由1g/l提高到20g/l时,最大氢气产率由51.4ml H2/g DW大幅度降低至11.0ml H2/gDW。在较高的节旋藻浓度条件下(20g/l),异相暗发酵的产氢峰值速率和最大氢气产率是自相暗发酵的110.0倍和8.4倍。因此在后续的实验中,微藻生物质应选用异相暗发酵产氢气的模式。节旋藻经过微波加热辅助稀酸预处理和酶水解可以有效的促进生物质的水解和强化暗发酵产氢气,随后利用沸石去除暗发酵尾液中的铵离子可以实现高效的光发酵产氢气。通过暗发酵和光发酵耦合产氢气,节旋藻的最大氢气产率大幅度提高至337.0mlH2/g DW。
本文以海洋微拟球藻为原料,讨论和比较了三种微藻生物质发酵产氢气和甲烷的方法：(1)暗发酵和光发酵耦合产氢气联产甲烷；(2)暗发酵产氢气联产甲烷；(3)直接发酵产甲烷。微拟球藻通过微波加热辅助稀硫酸预处理之后接种产氢细菌进行暗发酵产氢气。在暗发酵中产氢细菌对大部分经过预处理水解生成的氨基酸的消耗时间大约是还原糖消耗时间的两倍。微拟球藻通过暗发酵和光发酵耦合产氢气联产甲烷得到的最大氢气产率为161.3ml H2/g TVS,最大甲烷产率为183.9ml CH4/g TVS,整体能量转化效率为暗发酵产氢气联产甲烷的1.7倍、直接产甲烷的1.3倍。
本文以蛋白核小球藻为原料,研究了多种预处理方式对微藻生物质暗发酵产氢气的影响。经过蒸汽加热辅助稀酸和微波加热辅助稀酸预处理能够显著促进小球藻的水解和暗发酵产氢气。通过暗发酵和光发酵耦合产氢气联产甲烷,小球藻的最大氢气产率为198.3ml H2/g TVS,最大甲烷产率为186.2ml CH4/g TVS,整体能量转化效率为34.0%。在批次实验的基础之上,进行了小球藻的半连续流发酵产氢气的实验研究。通过长时间小球藻驯化得到的复杂产氢细菌菌群比之前通过葡萄糖驯化得到的简单菌群更能适应和高效利用小球藻生物质的各个成分,可以实现连续稳定的发酵产氢气。为了进一步提高生物质发酵的能量转化效率,以小球藻和木薯淀粉为混合原料,研究了碳氮摩尔比对发酵产氢气的影响。混合生物质在碳氮摩尔比为25.3的条件下得到的最大暗发酵氢气产率为276.2ml H2/gTVS,分别是单纯用小球藻和木薯淀粉为原料最大氢气产率的3.7倍和1.8倍。通过暗发酵和光发酵耦合产氢气联产甲烷,混合生物质的最大氢气产率和甲烷产率分别为664.2ml H2/g TVS和126.0ml CH4/g TVS,整体能量转化效率达到67.2%。
The extensive utilization of fossil fuels has resulted in serious energy crisis and environmental pollution. Hydrogen is considered as an ideal carbon-free secondary energy carrier with high energy density and clean combustion product. Hydrogen production from renewable biomass through fermentation is increasingly attracting worldwide attention. Microalgae biomass is a potential feedstock for fermentative hydrogen production because of its high photosynthetic efficiency, fast growth, and global distribution. In this study, microalgae biomass was used as feedstock to cogenerate hydrogen and methane through a novel three-stage method comprising dark-fermentation, photo-fermentation, and methanogenesis. The components in microalgae biomass were efficiently used through the three-stage method, therefore hydrogen yield and energy conversion efficiency (ECE) were significantly increased.
Glutamic acid, a typical amino acid degraded from protein components in microalgae biomass, was used as feedstock to investigate the feasibility of cogeneration of hydrogen and methane through the three-stage method comprising dark-fermentation, photo-fermentation, and methanogenesis. Hydrogen-producing bacteria (HPB), photosynthetic bacteria (PSB), and methane-producing bacteria (MPB) were used as the inocula during dark-fermentation, photo-fermentation, and methanogenesis, respectively. HPB can efficiently ferment glutamic acid to abundant soluble metabolite products (SMPs) and little hydrogen during dark-fermentation. The residual solution of dark-fermentation mainly contained acetate, butyrate, and ammonium. Because high concentration of ammonium (36.7mM) in the residual solution of dark-fermentation can significantly inhibit the activities of PSB in sequential photo-fermentation, a modified zeolite were used to extract ammonium by ion exchange to reduce the ammonium concentration to3.2mM (91.1%of ammonium removal efficiency). After ammonium removal, the treated solution was inoculated with PSB, exhibiting the maximum hydrogen yield of292.9ml H2/g glutamic acid during photo-fermentation. The residual solution from photo-fermentation was reused by MPB to produce the maximum methane yield of102.7ml CH4/g glutamic acid. The ECE from glutamic acid to gas fuels significantly increased from18.9%in hydrogen fermentation to40.9%in combined hydrogen fermentation and methanogenesis.
Trehalose, a typical carbohydrate component in microalgae biomass, was used as feedstock to investigate the feasibility of cogeneration of hydrogen and methane through the three-stage method comprising dark-fermentation, photo-fermentation, and methanogenesis. As a stable non-reducing sugar, trehalose was not easily used by HPB for efficient hydrogen production. Trehalose was first pretreated by microwave heating with dilute acid, and then was inoculated with HPB to produce hydrogen during dark-fermentation. The residual solution of dark-fermentation was reused by PSB during photo-fermentation. The residual solution of photo-fermentation was reused by MPB during methanogenesis. Overall, the maximum hydrogen yield of731.3ml H2/g trehalose and methane yield of116.9ml CH4/g trehalose were achieved. The sequential generation of hydrogen and methane from trehalose remarkably enhanced the ECE from47.2%in hydrogen fermentation to72.2%in combined hydrogen fermentation and methanogenesis.
Hydrogen production from Arthrospira platensis biomass through dark-heterofermentation by the [FeFe] hydrogenase of HPB and dark-auto fermentation by the [NiFe] hydrogenase of A. platensis was discussed. A. platensis biomass pretreated by ultrasonication and enzymatic hydrolysis was inoculated with HPB to produce hydrogen during dark-heterofermentation. The maximum hydrogen yield of92.0ml H2/g dry weight (DW) was obtained at20g/l of A. platensis biomass. In dark-autofermentation, hydrogen yield decreased from51.4ml H2/g DW to11.0ml H2/g DW with increasing substrate concentration from1g/1to20g/1. The hydrogen production peak rate and maximum hydrogen yield at20g/1of A. platensis biomass in dark-heterofermentation showed110.0-and8.4-fold increases, respectively, relative to those in dark-autofementation. Therefore, dark-heterofermentation was selected for the further investigation of fermentative hydrogen production from microalgae biomass. A. platensis biomass was pretreated by microwave heating with dilute acid to improve saccharification during enzymatic hydrolysis and hydrogen production during dark-fermentation. The residual solution of dark-fermentation was treated by zeolite to reduce ammonium concentration before photo-fermentation. The maximum hydrogen yield from A. platensis biomass was significantly increased to337.0ml H2/g DW through combined dark-fermentation and photo-fermentation.
Three methods for hydrogen and methane production from Nannochloropsis oceanica biomass were discussed as the following:(1) three-stage method comprising dark-fermentation. photo-fermentation, and methanogenesis;(2) two-stage comprising dark-fermentation and methanogenesis;(3) single-stage methanogenesis. N. oceanica pretreated by microwave heating with dilute acid was inoculated with HPB to produce hydrogen during dark-fermentation.The consumption time of most amino acids was about2times as long as that of most reducing sugars during dark-fermentation. The total ECE from N. oceanica biomass to gas fuels through the three-stage method showed1.7-and1.3-fold increases, respectively, compared with those through the two-stage and single-stage methods.
Effects of pretreatment methods on biomass saccharification and hydrogen fermentation from Chlorella pyrenoidosa were investigated. The steam heating with dilute acid and microwave heating with dilute acid can remarkably enhance the biomass hydrolysis and hydrogen fermentation. The maximum hydrogen yield of198.3H2ml/g total volatile solids (TVS) and methane yield of186.2ml H2/g TVS were achieved through the three-stage method comprising dark-fermentation, photo-fermentation, and methanogenesis. Semi-continuous fermentation of C. pyrenoidosa biomass was carried out based on batch fermentation. Compared with the simple microbial community formed at earlier stages of fermentation, the complex microbial community formed at later stages of fermentation was more adaptable to C. pyrenoidosa biomass and can utilize C. pyrenoidosa biomass more efficiently, thereby resulting in efficient and stable hydrogen fermentation. In order to enhance the ECE from C. pyrenoidosa biomass, cassava starch was mixed with C. pyrenoidosa biomass to optimize the carbon/nitrogen (C/N) molar ratio for efficient dark-fermentation. The maximum dark hydrogen yield of276.2ml H2/g TVS from the mixed biomass at C/N molar ratio of25.3showed3.7-and1.8-fold increases, respectively, compared with those from only C. pyrenoidosa biomass and only cassava starch. The maximum hydrogen yield of664.2H2ml/g TVS and methane yield of126.0ml H2/g TVS corresponding to the total ECE of67.2%were achieved through the three-stage method comprising dark-fermentation, photo-fermentation, and methanogenesis.
引文
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