Biohydrogen Production From Algae

C H A P T E R

9 Biohydrogen Production From Algae Man Kee Lam1, 2, Adrian Chun Minh Loy1, 2, Suzana Yusup1, 2, Keat Teong Lee3 1

Chemical Engineering Department, Universiti Teknologi PETRONAS, Seri Iskandar, Malaysia; 2Centre for Biofuel and Biochemical Research, Institute of Self-Sustainable Building, Universiti Teknologi PETRONAS, Seri Iskandar, Malaysia; 3School of Chemical Engineering, Universiti Sains Malaysia, Engineering Campus, Nibong Tebal, Malaysia

1. INTRODUCTION Heavy dependence on fossil fuels as a primary energy source has significantly damaged the Earth’s ecosystem. Fossil fuels are nonrenewable and will be exhausted under a century if new oil wells are not found [1]. Global industrialization and population growth are the main drivers that caused rapid energy exhaustion, which has consequently led to continual increases in the world price of petroleum and escalating tensions involving oil-producing countries [2,3]. In addition, the combustion of fossil fuels has created numerous environmental issues, such as greenhouse gas (GHG) effects, which are responsible for melting of arctic ice, rising sea levels, extreme heat waves, and frequent occurrences of droughts and desertification. Hence, identifying alternative renewable energy sources has become one of the key challenges of this century to attain a more sustainable and greener energy for the future. Renewable energy sources, such as solar, wind, hydro, and energy from biomass and waste, have been successfully developed and used by different nations to limit the use of fossil fuels [4]. Nevertheless, only energy produced from the combustion of biomass and wastes have made a significant contribution, indicating immense opportunities for biomass in the energy sector. In other words, research and development on green technologies using biomass to generate energy should be extensively promoted to strengthen this renewable source for diversification of energy supply in the global fuel market. Hydrogen (H2) is considered the cleanest renewable fuel because its combustion byproduct is only water vapor with no carbon dioxide (CO2) emitted to the atmosphere [5]. H2 has a higher specific energy content (142 MJ/kg) than methane (56 MJ/kg), natural gas (54 MJ/kg), and gasoline (47 MJ/kg) [6]. These characteristics make H2 a good candidate to be an important fuel in the future. To date, approximately 96% of the global H2 supply

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9. BIOHYDROGEN PRODUCTION FROM ALGAE

has been derived from nonrenewable and chemical-based processes, such as steam methane reforming (SMR), petroleum refining, and coal gasification [7,8]. Attention has been given to SMR because the process is cost-effective and currently available on a commercial-scale [9]. However, the SMR reaction is an endothermic reaction in which external energy (e.g., combustion of fuels) must be supplied to initiate the reaction to occur [10]. Furthermore, using methane for H2 production is not sustainable in the long-term because most methane is currently derived from fossil fuel, which is nonrenewable. On the contrary, biological H2 production from photosynthetic microorganisms has the potential to considerably reduce production costs and environmental impact because its production utilizes sunlight as the driving force to split water into H2 and O2 under controlled conditions [5,11]. Research on H2 metabolism by green microalgae was first explored by Gaffron and Rubin [12] and thereafter, for photosynthetic bacteria by Gest and Kamen [13]. The capability of these microorganisms to grow under both aerobic and anaerobic conditions has made them easier to shift within different cultivation modes to initiate H2 production [14]. Unlike biodiesel and bioethanol production from edible crops, biological H2 production does not compete with agricultural land for food production, which is an important consideration for the transition towards cleaner and greener energy development. There are several mechanisms for producing H2 from microalgae, including direct biophotolysis, indirect biophotolysis, photofermentation, and dark fermentation [15]. However, these methods must be improved through intensive research to make biological H2 production economically viable without requiring large energy inputs [2]. Thus the scope of this chapter is on the metabolic pathway for H2 production from microalgae and the steps taken to overcome production bottlenecks. The discussion also includes factors to be considered when scaling-up H2 production from microalgae and potential integrated system to enhance the overall life cycle energy balance of microalgae-based biohydrogen.

2. BIOHYDROGEN GENERATION BY MICROALGAE 2.1 Direct Biophotolysis Direct biophotolysis by microalgae is the most studied method for splitting water into H2 and O2 using sunlight as the energy source [2,16]. This is the simplest reaction for producing H2, involving only water and light with no emission of GHG [17]. When light is absorbed by photosynthetic microalgae, it enhances the oxidation of H2O molecules by Photosystem II (PS II), and the released electrons and protons are transported to the chloroplast ferredoxin (Fd) via PS I [14]. Reduced Fd acts as an electron donor to [FeFe]-hydrogenases [18], which reversibly facilitates the reduction of protons (Hþ) to H2 molecules as shown in Eq. (9.1) [11,14,19]. The [FeFe]-hydrogenases in microalgae are monomeric enzymes with molecular weights of approximately 48 kDa and have a unique prosthetic group (H-group) in the active center, which results in a 100-fold higher enzyme activity than [NiFe]-hydrogenases and metalfree hydrogenases [18,20]. However, the complicated reaction system for direct biophotolysis requires the input of a large amount of free energy (DG ¼ þ237 kJ/mol), resulted to slow H2 production rate by the microalgae [17]. A better understanding of H2 production by microalgae via direct biophotolysis pathway is illustrated in Fig. 9.1.

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2. BIOHYDROGEN GENERATION BY MICROALGAE

Starch

Light

CO2

+

H

NADPH NADP+

H2

Fp

[FeFe]-H2ase

H+

Light

e

-

+

Low H

PS II

e-

PQ(H2)

e-

Cyt b6f

e-

PS I

PC

e-

H+

eFD

ATP ATPase

ADP+Pi

High H+ H2O

O2 + H+

H+

FIGURE 9.1 H2 production pathway by photosynthetic microalgae. ATPase, ATP synthase; cyt b6f, cytochrome b6f complex; Fd, ferredoxin; Fp, ferredoxin-NADP reductase; H2ase, hydrogenase; PC, plastocyanin; PQ, plastoquinones; PQ(H2), reduced plastoquinones; PS, photosystem. Modified from I. Akkerman, M. Janssen, J. Rocha, R.H. Wijffels, Photobiological hydrogen production: photochemical efficiency and bioreactor design, Int. J. Hydrogen Energy 27 (11e12) (2002) 1195e1208; M.L. Ghirardi, A. Dubini, J. Yu, P.C. Maness, Photobiological hydrogen-producing systems, Chem. Soc. Rev. 38 (1) (2009) 52e61; Y.K. Oh, S.M. Raj, G.Y. Jung, S. Park, Current status of the metabolic engineering of microorganisms for biohydrogen production, Bioresour. Technol. 102 (18) (2011) 8357e8367.

2H þ þ 2FD4H2 þ 2FD

(9.1)

2.2 Limitations and Strategies to Improve H2 Evolution via Direct Photolysis 2.2.1 Sparging With Inert Gas One of the bottlenecks in direct biophotolysis is the very short period of H2 evolution by microalgae, typically ranging from several seconds to a few minutes [14]. This short period is caused by the simultaneous production of H2 and O2 molecules upon H2O oxidation with a theoretical ratio of H2:O2 ¼ 2:1 [14,21,22]. O2 is a powerful inhibitor of [FeFe]-hydrogenases enzymatic reaction and also a suppressor of [FeFe]-hydrogenases (HydA) gene expression [14,18]. Specifically, O2 is speculated to bind to an iron atom with the double Fe subcluster, which leads to the inhibition of the proton binding required for H2 generation [8]. This interferes with H2 metabolism in microalgae cells and resulted in low yield of H2 production [2,14,23]. To overcome this limitation, temporally or spatially removing O2 from the system is necessary to allow continuous activation of [FeFe]-hydrogenases in H2 metabolism [23]. This can be achieved by constantly sparging inert gas into the microalgae cultivation system to keep both H2 and O2 gases at low partial pressure [2]. The partial pressure of O2 should be maintained at below 0.1% or lower to effectively maintain the activity of [FeFe]-hydrogenases to produce H2 for long period. In reality, the sparging approach is impractical because a large quantity of inert gas is required to dilute the gas stream, leading to a high power input for gas transfer and consequently extra costs for the process [2,15]. In addition, the extra volume

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9. BIOHYDROGEN PRODUCTION FROM ALGAE

(photobioreactor) required to accommodate the additional quantity of inert gas will be large, causing the process to be economically infeasible for commercialization. The energy required to separate diluted H2 from O2 and inert gas is another issue that results in inert-gas sparging approach being unsustainable. 2.2.2 Sulfate Deprivation Another possible method for optimizing H2 production from microalgae via direct biophotolysis is to cultivate the microalgae under sulfate-deprived conditions [19]. This is a two-stage process, in which O2 and H2 production are temporally separated. Initially, microalgae are allowed to grow through normal photosynthesis with O2 generation while biofixing CO2 from the atmosphere. This stage allows the microalgae to attain high bulk density. The microalgae are then transferred to a new medium with minimal or no sulfate content to obtain low oxygen partial pressures and catalyze H2 evolution metabolism [14,19]. When microalgae are cultivated under limited sulfate conditions, synthesis of the PS II D1 polypeptide chain (32 kDa) is impeded, interfering the repair of PS II from photooxidative damage [14,24]. Thus the microalgae cells can maintain a high respiration rate exceeding the photosynthesis rate, creating an anaerobic condition that allows H2 production for 4e5 days [14,18]. In this situation, [FeFe]-hydrogenases are not significantly inhibited by O2 molecules because the O2 is quickly taken up during respiration. For example, when Chlamydomonas reinhardtii were deprived of sulfate, the photosynthesis rate was decreased by 5%e10% of the initial value after 15e20 h [24]. However, respiration activity was sustained over the first 70 h of sulfate deprivation and only slightly declined thereafter [19]. After 150 h, about 87% (v/v) H2 was recorded in the headspace of the photobioreactor, indicating the feasibility of sulfate deprivation for improving H2 production rate by microalgae [19]. Table 9.1 presents the H2 production rates under sulfate-deprived conditions for different microalgae strains. However, continuous evolution of H2 under sulfate-deprived conditions could not be sustained for a long period because substrates are utilized for catabolism [19,25]. In a study performed by Zhang et al. [25], the protein and starch content in microalgae cells were increased about 40% and 600%, respectively, from their initial values for the first 24 and 30 h of sulfate deprivation, but they gradually declined thereafter. This provides a clear evidence that substantial protein and starch catabolism occurs during H2 evolution under sulfate-deprived conditions [25]. At this point, the microalgae should be recultivated in a nutrient-rich medium to replace the needed metabolites before again being subjected to a sulfate-free medium for a subsequent cycle of H2 production [18]. The starch content in microalgae cells decreases because a substantial amount is consumed to support the cell’s mitochondrial respiration under anaerobic conditions in the cultivation system [25]. Mitochondrial respiration is responsible for absorbing the small amount of O2 released from residue photosynthetic activity, which helps to maintain the low partial pressure of O2 in the culture medium [26]. Microalgae are expected to decay under anaerobic conditions because of the absence of electron transport from H2O oxidation, which indirectly inhibits oxidative phosphorylation in mitochondria [14]. However, the expression of [FeFe]-hydrogenases in H2 evolution permits an alternative but slow rate of electron transport in the thylakoid membrane of photosynthesis, which is coupled to electron transport in mitochondria, leading to ATP generation in both organelles [14,25]. The ATP is probably used by the microalgae cells for sustaining basic life activities, prolonging their survival under sulfate-deprived conditions [14]. Fig. 9.2 illustrates the

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2. BIOHYDROGEN GENERATION BY MICROALGAE

TABLE 9.1

H2-Producing Microalgae by Means of Sulfate-Deprived Method

Microalgae Strain

H2 Production Rate (mL H2/L h)

Duration H2 (h)

References

Chlamydomonas reinhardtii

2

150

Melis et al. (2000) [19]

Chlamydomonas reinhardtii

4.3

552

Laurinavichene et al. (2006) [50]

Chlamydomonas reinhardtii

0.58

4000

Fedorov et al. (2005) [115]

Chlamydomonas reinhardtii

0.17e0.87

65e107

Tsygankov et al. (2006) [116]

Chlamydomonas reinhardtii

2.24

96

Jo et al. (2006) [117]

Chlamydomonas reinhardtii CC124

1.29

117

Faraloni et al. (2011) [118]

Chlamydomonas reinhardtii L159I-N230Y

1.81

285

Torzillo et al. (2009) [42]

Chlorella autotrophica IOAC689S

0.11

<7

He et al. (2012) [119]

Chlorella protothecoides IOAC038F

2.93

<6

He et al. (2012) [119]

Chlorella salina Mt

0.5

120

Chader et al. (2009) [120]

Chlorella sorokiniana Ce

1.35

222

Chader et al. (2009) [120]

Chlorella sp Pt6

0.8

120

Chader et al. (2009) [120]

Chlorella sp. IOAC085F

1.33

<6

He et al. (2012) [119]

Nannochloropsis sp. IOAC676S

0.018

7e19

He et al. (2012) [119]

Platymonas subcordiformis

0.04

5

Guan et al. (2004) [121]

Scenedesmus obliquus IOAC039F

0.052

<6

He et al. (2012) [119]

Tetraselmis striata IOAC707S

0.34

<7

He et al. (2012) [119]

electron transport pathways for photosynthesis and respiration by microalgae when deprived of sulfate. Hence, H2 production by microalgae cultivated with a limited sulfate medium involves a four-way coordinated interaction of oxygenic photosynthesis, mitochondrial respiration, catabolism of endogenous substrate, and electron transport via the [FeFe]hydrogenases pathway [14,25,26]. More fundamental research is urgently needed to address the relationship of these four processes and subsequently improve the stability of H2 production in green microalgae. 2.2.3 Mutagenesis and O2-Tolerant Hydrogenases Protein and genetic engineering is an alternative approach to enhance H2 production from microalgae [5,27]. As discussed previously, [FeFe]-hydrogenases play a vital role in biologically catalyzing H2 evolution from microalgae cells; however, this enzyme is extremely sensitive to the presence of O2 [14]. Therefore identifying and isolating O2-resistant [FeFe]hydrogenases from nature could be the solution to this problem [16]. In a study done by Ghirardi et al. [27], a classical random mutagenesis approach was conducted to identify O2-tolerance hydrogenases in the green microalgae C. reinhardtii. Initially, the mutagenized

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9. BIOHYDROGEN PRODUCTION FROM ALGAE

4 H+ Photophosphorylation

O2

2H2O

H2

Oxidative phosphorylation

2NADH

[starch]

Michondrion

O2

[FeFe]-hydrogenases

Chloroplast

2H2O

4 H+

FIGURE 9.2 Coordinated photosynthetic and respiratory electron transport coupled with phosphorylation during H2 production by microalgae. Electrons generated from oxidation of H2O through photosynthesis are delivered to [FeFe]-hydrogenases, leading to photophosphorylation and H2 evolution. The O2 generated by this process is responsible to drive the coordinate oxidative phosphorylation during mitochondrial respiration. At this point, electrons are derived upon endogenous substrate (starch) catabolism, generating reductant (NADH) and CO2. Production of H2 by the chloroplast permits the sustained operation of this coordinated photosynthesis-respiration function in green microalgae and allows the generation of ATP by the two bioenergetic organelles in the cell. Modified from A. Melis, T. Happe, Hydrogen production. Green algae as a source of energy, Plant Physiology 127 (3) (2001) 740e748.

microalgae cells were subjected to two different selective pressures. The first pressure was selected for the cells to metabolize H2 while the second pressure was selected for the cells to release H2 in the presence of O2. The surviving microalgae cells were subsequently subjected to additional screening using a chemochromic sensor that detects H2 evolution activity in microalgae [27,28]. Finally, mutated microalgae cells that recorded a high tolerance towards O2 were isolated for H2 production on a larger scale. Through this mutagenesis and isolation procedures, the selected mutants had 2e9 times higher O2 tolerance than the wild-type, indicating the possibility of sustaining H2 evolution by microalgae under aerobic conditions for longer periods [27,28]. However, more significant breakthroughs and the identification of O2-tolerant [FeFe]-hydrogenases from different microalgae strains for higher H2 production rate are required to accelerate its commercial development. In addition to isolating O2-tolerance hydrogenases, genetic engineering can be used to modify the electron transport pathway in H2 evolution system, so that the electrons are directed to the biophotolysis system, which enhances the H2 production rate [16,29]. Lee and Greenbaum [29] discovered a new competitive pathway for background O2 sensitivity in C. reinhardtii, which was 10 times more sensitive than the classic O2 sensitivity of [FeFe]-hydrogenases. From the study, O2 that apparently serves as a terminal electron acceptor was in competition with the H2 production pathway for photosynthetically generated electrons from direct biophotolysis (water splitting). Subsequently, this O2-sensitive H2 production electron transport pathway was inhibited by 3[3,4-dichlorophenyl]1,1dimethylurea (DCMU). DCMU is a strong chemical inhibitor that blocks the transport of electrons acquired from PS II water splitting to PS I [29]. Therefore genetic insertion of a programmed polypeptide protein channel with a hydrogenase promoter into the thylakoid

2. BIOHYDROGEN GENERATION BY MICROALGAE

225

membrane of microalgae cells could potentially dissipate the proton gradient across the membrane without ATP formation (ATP is produced during proton transfer) [17,29]. Through this strategy, the proton gradient that hinders electron transfer from water to [FeFe]hydrogenases could be avoided and simultaneously eliminate the newly discovered O2-sensitive pathway [29]. In another study done by Kruse et al. [30], mutated C. reinhardtii (with given name Stm6) was constructed by random gene insertion. This strain had a modified respiratory metabolism and was unable to perform cyclic electron transport around PS I. This led to the accumulation of starch inside the cells that enhanced substrate availability for a prolonged H2 production period (see Section 2.2.2). The starch accumulation was probably caused by inhibition of energy consumption by mitochondrial respiration under anaerobic conditions during illumination [30]. In addition, this mutated strain showed low levels of dissolved O2 concentration (30%e40% of parental strain), which helped to reduce the inhibitory effects of O2 towards [FeFe]-hydrogenases activity [30]. Hence, H2 production by Stm6 was sustained for a longer period with a production rate of 5e13 times higher than the parental strain. 2.2.4 Enhanced Light Capturing Efficiency Under an ideal environment, microalgae can absorb and use photon energy with nearly 100% efficiency to generate H2 from sunlight and water [14,31,32]. Unfortunately, under real conditions, the light intensity is high while the absorption capacity of photosynthetic pigments (chlorophyll a) in microalgae cells is limited; thus the photon energy could not be efficiently converted to chemical energy. Instead, up to 80% of the energy is wasted by dissipation in the form of fluorescence and heat [33e35]. Furthermore, the shading effect created by dense cultures and low light penetration into the cultivation system might bring down the efficiency to less than 1% and often closer to 0.1% [35]. As a consequence, upscaling a microalgae cultivation system for H2 production would be problematic, as the cultivation would probably take place in an enclosed photobioreactor and cultivated at outdoor with abundant sunlight [14]. To overcome this problem, a possible strategy is to engineer microalgae mutant with a limited amount of chlorophyll (smaller light-harvesting antenna size) is necessary to permit greater transmittance of irradiance through high-density culture as shown in Fig. 9.3 [14,33,34,36e38]. Hence, less photon energy would be wasted at the surface of the culture and sunlight could penetrate deeper into the culture, leading to an overall increase in biomass productivity and H2 production [39]. In a study done by Polle et al. [33], DNA insertional and chemical mutagenesis of C. reinhardtii was carried out to isolate tla1, a stable transformant that has truncated light-harvesting chlorophyll antenna size. From the biochemical analyses, this mutated strain was extremely deficient in chlorophyll content (but not photosystem), being 50% and 65% lower in functional chlorophyll antenna size of PS II and PS I, respectively [14,33,40,41]. Therefore the tla1 strain exhibited higher saturating light intensity for photosynthesis and greater solar conversion efficiency than the wild-type. This unique characteristic permits mass cultivation of microalgae under direct sunlight and thus enhances energy sustainability in the cultivation system. However, a high photosynthetic conversion efficiency does not necessarily translate to a high H2 production rate because of the sensitivity

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9. BIOHYDROGEN PRODUCTION FROM ALGAE

H2

H2

H2 H2

H2

Sunlight Sunlight

Waste heat

(A)

(B)

FIGURE 9.3 Sunlight absorption and dissipation by green microalgae. (A) Dark green microalgaedonly individual cells on the surface absorb the sunlight (more than can be utilized by photosynthesis) and dissipated in the form of waste heat and resulted in low productivity of H2. (B) Truncated chlorophyll antenna sizedindividual cells have limited light absorbing capacity; thus permits greater light penetration and H2 production by cells deeper in the culture. Modified from A. Melis, Solar energy conversion efficiencies in photosynthesis: minimizing the chlorophyll antennae to maximize efficiency, Plant Sci. 177 (4) (2009) 272e280.

of [FeFe]-hydrogenases towards O2 [35]. The microalgae must still be deprived of sulfate to initiate the anaerobic conditions for subsequent H2 production. In another study carried out by Kosourov et al. [34], a tla1 mutant (CC-4169) was immobilized within thin alginate films (w300 mm) and cultivated under sulfate-deprived conditions. Under limited light conditions (light intensity of 19 mE/m2 s), CC-4169 demonstrated an extremely low yield of H2, being four-times lower than the parental strain. This result was expected because CC-4169 had smaller light-harvesting antenna; thus lowering the photosynthetic efficiency. However, when the light intensity was increased to 285 and 350 mE/m2 s, the mutant exhibited four-times and more than six-times H2 production than the parental strain, respectively. Therefore the hypothesis was confirmed that truncating light-harvesting antenna in microalgae cells could enhance light conversion efficiency under high light intensity and result in a higher H2 production per unit of illuminated culture surface area [34]. A similar result was reported by Torzillo et al. [42], in which C. reinhardtii D1 mutant with the substitution of double amino acids exhibited positive result in H2 production because of low chlorophyll content, higher photosynthetic capacity and conversion efficiency, and a high respiration rate under anaerobic conditions. Therefore H2 production rate from this mutant was higher compared to the wild-type, 1.81 mL H2/L h versus 0.30 mL H2/L h, and H2 production was able to proceed for much longer, 285 h versus 53 h [42]. In short, microalgae with truncated light-harvesting chlorophyll antenna play a key role in improving H2 production and have a greater chance of mass cultivation under direct sunlight [14,42].

2.3 Indirect Biophotolysis For indirect biophotolysis, the problem of [FeFe]-hydrogenases inactivation by O2 is potentially eliminated through the temporal or spatial separation of O2 and H2 evolution [15,39,43]. In the first stage, microalgae are allowed to grow under normal cultivation conditions with light as the driving energy source to fix CO2 into carbohydrate [39]. During this process,

2. BIOHYDROGEN GENERATION BY MICROALGAE

227

microalgae are allowed to reproduce as much as possible to increase the overall total carbohydrate in the cells while producing O2 as by-product [2]. In the second stage, commonly referred as anaerobic dark fermentation, carbohydrate in microalgae cells is used as a substrate by the nitrogenase enzyme (nitrogen fixation) to produce H2 [15,39,44]. The overall reactions for both stages are shown in Eqs. (9.2) and (9.3), respectively. Compared to direct biophotolysis, indirect biophotolysis has the advantages of using water as the electron donor and inorganic carbon as the carbon source, but O2 inhibition during H2 production is not significant [5,16]. However, because of the multiple steps involved in converting solar energy to H2 and the requirement for ATP by nitrogenase enzyme, the maximum conversion efficiency of light to H2 via indirect biophotolysis is only 16.3% [44,45]. It should be noted that the twostage process using sulfate deprivation to create anaerobic conditions for H2 production (Section 2.2.2) is considered to be direct biophotolysis and not indirect biophotolysis, as photosynthesis is still functioning inside the cells and delivering electrons to the hydrogenases during the anaerobic phase [5,15,46]. 12H2 O þ 6CO2 /C6 H12 O6 þ 6O2

(9.2)

C6 H12 O6 þ 12H2 O/12H2 þ 6CO2

(9.3)

Cyanobacteria (or blueegreen algae) are well-known for H2 production via indirect biophotolysis [11,23,44,47]. Cyanobacteria strains that can produce H2 are mostly filamentous and containing specialized nitrogen-fixing cells known as heterocysts [11]. In the heterocyst, nitrogenase is protected from O2 by a thick cell wall that has low diffusivity of O2. With a high respiration rate, O2 is normally utilized, creating an ideal anaerobic environment for higher H2 production [47]. On the other hand, the lack of heterocysts in green microalgae caused it to be inefficient in fixing nitrogen under anaerobic dark conditions, which results in limited amounts of H2 generated through indirect biophotolysis. In a study done by Ohta et al. [48], green microalgae Chlamydomonas MGA 161 was initially grown using light intensity of 25 W/m2 with continuous sparging with air containing 5% CO2. Once the microalgae growth reached the stationary phase, the culture was transferred to a medium that was flushed with pure N2 gas and cultivated under dark environment to initiate anaerobic fermentation. After 12 h of incubation, 1.5 mmol H2/g dry wt was detected with degradation of starch at a rate of 0.08 mmol glucose/g dry wt/h. Apart from H2, other fermentative byproducts were also produced, such as CO2, acetate, ethanol, formate, and glycerol with mole ratios of 0.9, 0.73, 0.33, 0.13, and 0.05 for every 1 mol of H2 produced, respectively [48].

2.4 Integrated System To date, neither direct nor indirect biophotolysis of microalgae has been performed at a commercial-scale for sustainable H2 production even though significant breakthroughs and improvements have been achieved. This is mainly because several technical problems still remain unsolved, such as low yields of H2, sensitivity of hydrogenases towards O2, high cost of photobioreactor, and technological challenges in genetic and molecular engineering [18]. In contrast, dark fermentation is superior for H2 production, but the process is limited

228

9. BIOHYDROGEN PRODUCTION FROM ALGAE

to fermentative bacteria (e.g., Escherichia coli, Clostridium, and Sporolactobacillus) and rarely to green microalgae [5]. For instance, H2 production rate through the dark fermentation of E. coli SR 13 was 300 L H2/L h [49] whereas direct biophotolysis of C. reinhardtii only yielded 0.012 L H2/L h [50]. Nevertheless, bacteria required substrates, such as glucose and sucrose, during dark fermentation as energy source to grow and the cost of these substrates may prevent the implementation of this technology at a commercial-scale for H2 production [51]. To overcome this problem, the use of low-cost substrates, such as industrial food wastes or wastewater, appears to be a potential solution; however, several other problems may surface, such as the need to pretreat the low-cost substrates to provide a suitable environment for the bacteria to grow, possibility of contamination and competition for substrates utilization by other bacteria that do not produce H2, and variations and inconsistencies in substrates concentrations. Integrating direct biophotolysis by microalgae and dark fermentation by fermentative bacteria could circumvent the disadvantages of each individual process and enhance the overall H2 production process. As shown in Fig. 9.4, microalgae are initially cultivated in the first stage by utilizing light energy to produce H2 through direct biophotolysis. After the microalgae have stopped producing H2, the biomass is utilized as a substrate by bacteria to continue H2 production via dark fermentation process. During dark fermentation, CO2 is emitted along with H2 as shown in Eq. (9.4). If a gas separation unit is added into the system to separate CO2 from H2, CO2 could be used as a carbon source to grow microalgae in the first stage. This strategy not only creates a symbiotic relationship between indirect biophotolysis and dark fermentation, but also improves the energy conversion efficiency and CO2 utilization in this integrated system. Furthermore, including a photofermentation process using purple nonsulfur bacteria, such as Rhodospirillum rubrum, Rhodopseudomonas palustris, Rhodobacter sphaeroides, and Rhodobacter capsulatus in the integrated system appears to be another potential approach to enhance solar energy utilization and H2 productivity [11,18]. Dark fermentation : C6 H12 O6 þ 2H2 O/2CH3 COOH þ 2CO2 þ 4H2

(9.4)

Although the integrated system is still far from commercialization, several recent studies have revealed the potential of using microalgae starch as a substrate for fermentative bacteria

light

H2

Microalgae

H2 biomass

Fermentative bacteria CO2

Direct biophotolysis

FIGURE 9.4

Dark fermentation

Schematic diagram of integrated biological H2 production processes. Modified from E. Eroglu, A. Melis, Photobiological hydrogen production: recent advances and state of the art, Bioresour. Technol. 102 (18) (2011) 8403e8413; M.L. Ghirardi, L. Zhang, J.W. Lee, T. Flynn, M. Seibert, E. Greenbaum, A. Melis, Microalgae: a green source of renewable H2. Trends Biotechnol. 18 (12) (2000) 506e511.

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229

to produce H2 during dark fermentation [52e54]. In a study done by Nguyen et al. [52], starch accumulated in C. reinhardtii cells was utilized by the hyperthermophilic bacterium, Thermotoga neapolitana as an alternative source of substrate for H2 production. This bacteria is rod-shaped, nonspore-forming and able to ferment at high temperature, which means that it is less prone to interference by contaminants, as other bacteria may not grow at high temperature [52]. However, for microalgae biomass to be effectively used as substrate during dark fermentation, two important pretreatment steps must be taken: (1) microalgae cell wall is rigid and disruption of the cell wall is required to release the starch and (2) starch is a macromolecule and thus hydrolyzing the starch into simple reducing sugar is required to enhance its utilization. When the microalgae biomass was directly mixed with the hyperthermophilic bacteria without any pretreatment, no H2 was produced under dark fermentation conditions. Nevertheless, when microalgae cells were disrupted with methanol, H2 evolution was detected at a production rate of 53 mL/h L. In addition to that, when the disrupted microalgae cells were subjected to enzymatic hydrolysis (a-amylase, Termamyl 120L) and subsequently proceeded to dark fermentation, the H2 production rate was dramatically increased to 227 mL/h L. It is clear that pretreatment of the microalgae biomass played a crucial role in allowing fermentative bacteria to access and convert intracellular starch for H2 production. However, it should be noted that the expensive pretreatment steps may significantly increase the overall H2 production costs and make the process economically unsustainable. Another example of an integrated microalgae H2 system was demonstrated by Mussgnug et al. [55]. In that study, green microalgae C. reinhardtii was allowed to produce H2 under sulfate-deprived conditions as established by Melis et al. [19]. After the microalgae had stopped producing H2, the microalgae cells were harvested and used as a substrate in a subsequent anaerobic fermentation process. However, the targeted fermentative product was methane, which is an alternative biogas to H2. Through this integrated system, methane yield was increased to 123% compared to the control [55]. When C. reinhardtii was cultivated under sulfate-deprived conditions, fermentative compounds, such as starch, increased within the cells as a response to the induction of the H2 production cycle [14,25,55]. Simultaneously, the risk of emitting H2S during anaerobic fermentation will also be reduced, resulting in a higher purity of methane. Nevertheless, allowing microalgae to produce H2 in the first stage for a prolonged period exposed the risk of decreasing starch content in the microalgae cells due to mitochondrial respiration under anaerobic conditions [25]. As a result, limited starch would be available for the second-stage fermentation, leading to a possible low yield of biogas production. Hence, more studies are needed to address the possibilities of this integrated system, typically in term of energy balance and cost-effectiveness.

3. LIFE CYCLE ASSESSMENT OF H2 PRODUCTION FROM MICROALGAE Although H2 production from microalgae has the potential to significantly contribute in diversifying the global renewable energy resources, but, there is still a question mark of the sustainability of the process for long-term operation. Up to date, hydrogen evolution from microalgae has primarily been reported on a lab-scale [30,56], while pilot- and

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commercial-scale are rarely found in the literature. As a result, a critical understanding of the overall process operation is very limited, mainly in term of energy balance and global warming potential. Microalgae H2 production involves not only H2 evolution during cultivation, but also includes other processing stages, such as nutrients sources and preparation, biomass harvesting and drying, water purification and reuse, and H2 purification and compression, which are equally important in addressing its feasibility for scaling-up [57e61]. In this regard, performing a life cycle assessment (LCA) provides a platform to benchmark the environmental repercussions of H2 production from microalgae through an in-depth inventory and impact assessments for each processing stage [56]. LCA is currently accepted as an effective tool for researchers and policymakers to understand the real environmental potential of a particular product. It can also be used to indicate if the production of a particular product can lead to negative environmental phenomena, such as eutrophication, global warming, ozone depletion, human and marine toxicity, land competition, and photochemical oxidation, so that precautionary steps can be considered to reduce negative impacts [58,62e64]. In addition, an energy balance can also be calculated to determine the energy hotspot for all stages within the system boundary of the LCA [65]. Romagnoli et al. [56] recently carried out an LCA study of H2 production from C. reinhardtii when scaled-up to a commercial level. In that study, H2 production through direct biophotolysis coupled with sulfate deprivation was used as a baseline to benchmark GHG emissions compared to natural gas, heavy fuel oil, and coal. It was estimated that the H2 production yield from C. reinhardtii would be 109.5 tonne/year or equivalent to 15,549 GJ/year, assuming the higher heating value of H2 was 142 MJ/kg. The CO2 emission from the combustion of natural gas, heavy fuel oil, and coal, which could be avoided, was 8.4, 10.3, and 25.5 tonne CO2/year, respectively. This important finding has opened a new direction in the microalgae biofuel industry and provides an excellent platform for the evaluation of a stronger and wider H2 market strategy for policymakers and investors [56]. However, it is still unclear if H2 production from microalgae can have a positive energy balance. Several recent LCA studies of microalgae biofuel production showed that intensive energy was consumed especially in the cultivation system (e.g., nutrients production, air and water pumping) and in biomass processing (e.g., harvesting and drying) [66e69].

3.1 Nutrients Sources According to LCA study done by Clarens et al. [66], about 50% of the overall energy use and GHG emissions were associated with the use of chemical (inorganic) fertilizers. Although chemical fertilizers have the advantage of reducing contamination in the cultivation medium and promote healthy growth of microalgae, nevertheless, their long-term usage is definitely unsustainable and not beneficial to the environment. On the contrary, using wastewater as an alternative source of nutrients to cultivate microalgae seems to be more practical and economically viable. Secondary and tertiary wastewaters normally contain significant amounts of nitrate and orthophosphate, which are not removed during primary treatment. Removing these compounds typically requires an additional 60%e80% of the energy consumed in a typical wastewater treatment plant [66,70]. Thus cultivating microalgae in wastewater offers both an inexpensive wastewater treatment technology, as well as a means of reducing the need of chemical fertilizers and their associated life cycle burden [66].

3. LIFE CYCLE ASSESSMENT OF H2 PRODUCTION FROM MICROALGAE

231

In view of the potential of H2 evolution from C. reinhardtii, cultivating this microalgae strain using wastewater could synergize a better energy output and improve the sustainability of the microalgae cultivation system. Kong et al. [71] reported the possibility of cultivating C. reinhardtii in wastewater discharged from industrial facilities and households. It was found that the microalgae could not grow exponentially when cultivated with treated wastewater (effluent), which contains low concentrations of nitrogen and phosphorus. On the contrary, when the microalgae were supplied with concentrated wastewater that contains high level of nutrients source, their growth rate improved significantly, with a 2.5-fold increment in biomass quantity [71]. It is necessary to enhance microalgae growth with a nutrient-rich medium to obtain a higher cell division rate before the microalgae are subjected to sulfatedeprived medium for H2 production. Using wastewater as nutrients source is ultimately a winewin approach to obtain a high density of microalgae biomass while they play an important role in purifying the wastewater before discharge. Issues that would need to be addressed prior to commercializing this strategy include contamination by fungus and bacteria in wastewater that may annihilate the entire population of microalgae, possible inconsistency in the nutrient content of the wastewater that may affect the growth rate of microalgae and the wastewater after microalgae cultivation need to be monitored before being discharged to water sources to avoid microalgae bloom. After being grown to a high density, the microalgae cells are ready to be transferred to a new medium with limited sulfate concentration for H2 production. In this regard, wastewater is no longer suitable to be used as the culture medium unless sulfate is prior removed from the wastewater. Depending on the discharge sources, wastewater normally contains a high concentration of sulfate, as it is true for pharmaceutical wastewater [72], textile wastewater [73], and molasses wastewater [74]. However, chemical processes for removing sulfate from wastewater, such as precipitation by iron (or other metals), is relatively expensive and generates large amounts of solid wastes [75,76]. This makes the process impractical for integration with the microalgae H2 production system. In fact, there is no supporting evidence to date that wastewater with sulfate removed is effective in enhancing H2 evolution from microalgae. Apart from that, most works with microalgae have used chemical fertilizers and were conducted under controlled environments. Hence, more research on this subject is needed, particularly on the effects of fungal and bacterial contamination and inconsistent nutrient compositions in wastewater towards H2 evolution rate of microalgae.

3.2 Photobioreactor Photobioreactor design is equally as important as supplemental nutrients in optimizing H2 production from microalgae, especially through the direct biophotolysis method. A good photobioreactor should have the following characteristics: (1) effective illumination area, (2) optimal gaseliquid transfer rate, (3) easy to operate, (4) low potential for contamination, (5) low capital and production costs, and (6) minimal land area requirements [77]. There are various type of photobioreactors designed mainly for microalgae biomass production, such as (1) open system, which is normally known as raceway pond, and (2) closed system, which include tubular, bubble column, flat plate, and fermentor [78]. Table 9.2 summarizes the design characteristics of these photobioreactors.

232 TABLE 9.2

9. BIOHYDROGEN PRODUCTION FROM ALGAE

Different Photobioreactor Designs for Microalgae Cultivation [77,79,85,115,116] Raceway Pond

Flat-Plate

Tubular

Vertical-Column

Bioreactor with rectangular shape

Consists of an array of straight, coiled, or looped transparent tubes

Bioreactor with vertical arranged cylindrical column

The flat-plate are usually made of transparent plastic or glass

The tubes are usually made of transparent plastic or glass

The columns are usually made of transparent plastic or glass

Mixing and circulation Usually coupled are provided by with gas sparger paddle wheels

Usually coupled with a pump or airlift technology

Usually coupled with a pump or airlift technology

The depth of the pond is usually 0.2e0.5 m to ensure microalgae receive adequate exposure to sunlight

The light path (depth) is dependent on microalgae strain; range between 1.3 and 10 cm

Tube diameter is limited (0.1 m) to increase the surface/ volume ratio

Optimum column diameter is 0.2 m with 4 m height

Easy to construct and operate

Large illumination surface area gives maximum utilization of solar energy

Large illumination surface area

High mass transfer rate with good mixing

Low energy input and low-cost

Low concentration of dissolved oxygen

Relatively higher biomass productivity

Compact, easy to operate, and relatively low-cost

Can be position vertically or inclined at an optimum angle facing the sun

Potential of cell damage is minimized if airlift system is used

Lower power consumption

Conceptual design

Design Consist of closed-loop characteristics recirculation channel (oval shape) Usually built using concrete or compacted earth-lined pond with white plastic

Advantages

Lower power consumption Disadvantages Water loss due to high evaporation rate

Scale-up require many Require large land area compartments and because long tubes support materials are used

Small illumination surface area

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3. LIFE CYCLE ASSESSMENT OF H2 PRODUCTION FROM MICROALGAE

TABLE 9.2

Different Photobioreactor Designs for Microalgae Cultivation [77,79,85,115,116]dcont'd Raceway Pond

Flat-Plate

Tubular

Vertical-Column

Difficulty in controlling the temperature and pH

Difficulty in controlling culture temperature

Potential in accumulating high concentration of O2 (poison to microalgae) in culture medium if tubes are too long

Cell sedimentation may occur if airlift system is not used

Susceptible to contamination

Decreasing CO2 concentration along the tube may cause the microalgae deprived of carbon source Mixing is problematic in extended tubes

To date, the raceway pond appears to be the most feasible method for mass cultivation of microalgae biomass. The system consists of a closed-loop recirculation channel (oval shape) in which mixing is provided by paddle wheels. The depth of the pond is usually 0.2e0.5 m to ensure the microalgae receive sufficient exposure to sunlight [79e81]. The system is relatively low-cost compared to a closed system and requires a minimal energy input to operate. However, it is impossible to create an anaerobic environment in the raceway pond for H2 evolution from microalgae [78]. Even if H2 evolution is possible, H2 collection would become problematic because of the large cultivation area involved. In this case, an extremely high energy input to operate air pump is expected, which could potentially result in a net negative energy balance in the life cycle of microalgae H2 production. A closed photobioreactor is an alternative for scaling-up H2 production from microalgae. Generally, a closed photobioreactor offers several advantages compared to raceway ponds, including better contamination control, higher microalgae biomass yield per unit reactor volume, and the possibility of using single-strain culture for a prolonged duration [79,80]. A closed photobioreactor has a high surface area to volume (A/V) ratio that allows higher solar energy conversion efficiency by microalgae to H2 through direct biophotolysis. Among all the closed photobioreactors, the horizontal tubular/air-lift, helical tubular, and a-shaped photobioreactor exhibited a relatively higher A/V ratio and is currently being tested under labscale conditions for H2 production from cyanobacteria and microalgae [78]. However, recent LCA studies have shown that closed photobioreactors used substantial amounts of energy and resulted in an unsustainable practice for producing biofuel from microalgae [67,69]. For example, it was estimated that the energy input to operate air-lift photobioreactor was around 350% higher compared to raceway pond [65]. Furthermore, an energy assessment carried out by Stephenson et al. [67] indicated that using an air-lift photobioreactor to cultivate microalgae for biodiesel production could lead to net negative energy balance in the associated life cycle boundary. Most of the energy is used for powering pumps so that sufficient mixing and optimum gaseliquid transfer is achieved. Based on current technology maturity, an air-lift photobioreactor is not up to the commercialization stage yet, unless proper modifications are made to reduce the overall operating energy consumption.

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To overcome the limitations of open and closed photobioreactors, integrating both types of photobioreactors into one system may be a possible solution for commercial H2 production from microalgae. For the first stage, microalgae are cultivated in a nutrient-rich medium in raceway pond to promote the growth of microalgae until a certain quantity of cells is achieved. In the subsequent stage, the microalgae cells are harvested and immediately transferred to a sulfate-deficient medium in a closed photobioreactor to permit H2 production under anaerobic conditions. To achieve anaerobic conditions, the cultivation must be initially flushed with N2 and incubated in a dark environment for several hours before exposing to light sources to initiate H2 evolution. Mixing of microalgae culture utilizing air bubble is no longer possible in the second stage because of the need to have an anaerobic environment. Thus a mechanical agitator in the closed photobioreactor is needed to provide homogenous mixing and to allow microalgae to receive sufficient light [82,83]. In this regard, a vertical column-type photobioreactor is the most plausible option, as it allows easy installation of a mechanical agitator in the center of the column and facilitates H2 collection at the headspace. Hence, choosing a suitable agitator and optimizing the agitation speed are important in minimizing the shear stress on microalgae cells that may subsequently affect the H2 production rate [84,85]. Other types of closed photobioreactors, such as flat-plate, helical tubular, and a-shaped, are also suitable for cultivating microalgae, but is expected to consume more energy because of the difficulty of installing a mechanical agitator in these photobioreactors, resulting in the need for a heavy-duty water pump to circulate the culture to avoid from settling.

3.3 Harvesting Technology To promote H2 evolution through direct biophotolysis, microalgae need to be harvested from the nutrient-rich medium before transferring to the sulfate-limited medium. Harvesting microalgae is challenging because they are small (generally, 3e30 mm) and suspended in liquid [86]. Centrifugation and filtration are among the most effective methods for harvesting microalgae cells and are commonly applied at the lab-scale. However, it is not feasible to implement these two methods on a commercial-scale because of their high energy consumption, and high capital and maintenance costs [65]. A recent LCA study has highlighted that using filtration and centrifugation to harvest microalgae biomass contributed 88.6% and 92.7%, respectively, to the entire energy input of the system boundary [87]. This important finding should not be ignored, as it may have a significant impact on the overall energy balance in producing H2 from microalgae. Coagulation and flocculation offer an alternative and relatively low energy method to harvest microalgae cells. Microalgae cells always carry negative charges that cause them to repel each other and to remain suspended in liquid even if mixing is not provided. The negative charges surrounding the microalgae cells can be neutralized by introducing a positively charged coagulant into the culture medium. At the same time, flocculant can also be added to promote agglomeration by creating bridges between the neutralized cells, creating dense flocs that naturally settle due to gravity [88]. Nevertheless, high doses of conventional flocculants (e.g., multivalent salts) are required to achieve satisfactory results that may cause contamination of microalgae cells [89,90]. In addition, some of the microalgae cells might

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235

not survive when being transferred to a new medium with limited sulfate content and consequently, affecting the H2 production rate. Although no scientific research work or assessment has been carried out to justify this hypothesis, flocculant toxicity should not be ignored especially when the harvested microalgae cells are used for the second phase of cultivation. Nevertheless, there are a handful of researches that currently focused on the development of bioflocculants and immobilization technologies to reduce toxicity effects after harvesting and permit longer H2 evolution from microalgae [50,90e93]. Immobilization of microalgae for H2 production was described by Hahn et al. [91]. C. reinhardtii was immobilized onto fumed silica particles to produce H2 in a two-step cycle. Immobilized microalgae were initially grown in a sulfate-rich environment to promote cell growth in the silica particles, and then the silica particles were filtered and moved to a sulfate-free environment for H2 evolution. Since the silica particles are much denser than water and can be easily filtered, the microalgae cells were readily transferred to the sulfatedeficient medium without requiring much energy input. The results from the study indicated that H2 evolution from microalgae was not affected by immobilization, and the H2 production rate was nearly the same as that of free cells. In fact, this immobilization technology can be easily integrated into existing photobioreactor designs with little or no modification [91]. This technology could eliminate high-energy harvesting methods, such as centrifugation, leading to a better energy balance for H2 production from microalgae. Another important finding about the immobilization of microalgae for H2 production was reported by Kosourov and Seibert [93]. From their study, immobilization of C. reinhardtii within thin alginate films exhibited an extraordinary observation, in which the microalgae were reported to produce H2 even under atmospheric conditions (aerobic) instead of anaerobic conditions. This could be because the alginate films acted as shields that restrict O2 diffusion into the microalgae cells and thus protected the hydrogenases from deactivation [93,94]. However, it was found that when high density of cells was embedded within the alginate films, the H2 production rate was significantly reduced. One possible explanation for this observation was that a high population of microalgae cells increased the porosity of alginate films [93], resulting in more O2 diffusing into the alginate films and inhibits hydrogenases from producing H2. On the contrary, higher H2 production rates were observed in alginate films with low cell density, suggesting a more efficient light utilization per unit matrix area by the microalgae and the elimination of any shading effects.

3.4 Process Integration With Biodiesel Production From Microalgae To date, it has not been feasible to commercialize H2 production from microalgae because of the low volume of H2 produced, high costs, high energy input, and sensitivity of microalgae towards the surrounding environment for inducing or promoting H2 production. Therefore integration with an H2 production system from bacteria is suggested to enhance H2 productivity and to drive a positive energy balance for the system (see Section 2.4). It is also possible to improve the process energy balance for solely microalgae culture by expanding and diversifying biofuel production from the microalgae biomass. Apart from carbohydrates and proteins, certain microalgae strains are able to accumulate a significant amount of lipids inside their cells, in which these lipids can be extracted and

236

9. BIOHYDROGEN PRODUCTION FROM ALGAE

converted to biodiesel. Biodiesel is currently recognized as a green and alternative renewable diesel fuel that has attracted vast interest from researchers, governments, and local and international traders. Some of the advantages of using biodiesel instead of fossil diesel are that it is a nontoxic fuel, biodegradable, and has a lower emission of GHG when burned in diesel engine [95]. H2-producing microalgae, such as C. reinhardtii, could accumulate lipids up to a content of 25% [71,96], which makes it a potential microalgae strain for diversifying the types of biofuel that can be produced. In other words, after C. reinhardtii has stopped producing H2, the microalgae cells can be harvested, dried, and followed by subsequent lipid extraction for biodiesel production. Various lipid extraction methods have been reported, including mechanical press, chemical solvent extraction, and supercritical fluid extraction. However, only chemical solvent extraction (hexane, methanol, chloroform, and mixed solvent) has demonstrated the highest efficiency due to high selectivity and solubility of the solvent towards lipids. This allows even interlipids to be extracted out through diffusion across microalgae cell walls [97]. The extracted lipids are then subjected to transesterification reaction in which biodiesel is produced as the main product while glycerol is the by-product [4]. Through this strategy, the overall process chain to produce H2 from microalgae would become more sustainable and feasible in term of energy balance and production cost. Even though research on this integrated system is limited, there are a number of potential advantages to this process.

4. THERMOCHEMICAL CONVERSION METHOD 4.1 Thermochemical Conversion Technologies Despite of extensive studies into different aspects of microalgae to produce biofuel over the past few decades, the conversion of microalgae feedstock to syngas and biohydrogen via thermochemical conversion routes still received little attention from the researchers. Nevertheless, it is expected that thermochemical conversion of microalgae biomass to biohydrogen is a promising technology in the near future due to the simple cell structure of microalgae compared to lignocellulose biomass [98]. Generally, there are three types of thermochemical conversion methods to produce biohydrogen from microalgae, which are pyrolysis, gasification, and direct combustion [99]. During thermochemical conversion process, the microalgae biomass will be heated in the deficient conditions to produce syngas, which primarily consisted of biohydrogen and carbon monoxide [100]. Table 9.3 presents the microalgae species that are currently used to produce biohydrogen via thermochemical pathways.

4.2 Gasification Gasification is one of the promising thermochemical technology to convert organic biomass to fuel gases or syngas. Gasification usually occurs at high temperature 800e1000 C in partial oxidation of air, steam, or oxygen [101]. The mechanism of gasification starts with thermochemical degradation of biomass to H2, CO, CO2, and light hydrocarbons gaseous, such as CH4, C2H4, and C2H6, along with unwanted products, such as water and tar. The main target of gasification is to increase the yield of gases product from biomass and to

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4. THERMOCHEMICAL CONVERSION METHOD

TABLE 9.3

Potential Biohydrogen Production From Microalgae Biomass via Thermochemical Pathways

Microalgae Species

Reaction

Method

H2 Produced

References

Chlorella vulgaris

Gasification

Batch and continuous flow reactor with K2Co3 catalyst

53.00 vol%

Chakinala et al. (2010) [105]

Pyrolysis

Quartz tube reactor with activated carbon catalysts

37.11 vol%

Hu et al. (2014) [122]

Gasification

Batch reactor without catalysts

38.60 vol%

Brown et al. (2010) [123,124]

Pyrolysis

Fixed bed reactor with HZSM-5 zeolite catalysts

0.18 wt%

Scenedesmus almeriensis

Gasification

Conventional electrical furnace and in a single mode microwave oven without catalysts

48.00 vol%

Baneroso et al. (2013) [110]

Spirulina platensis

Gasification

Hydrothermal gasification and methanation reactor with Ru/C catalysts

29.3 vol%

Stucki et al. (2009) [125]

Nannochloropsis sp.

reduce the solid residue, such as tar. Apparently, researchers found out that biomass with low moisture content (<30 wt%) is suitable for gasification. Therefore microalgae were classified as the potential candidate for gasification process due to its low moisture content (3.99e10 wt%) after drying process [102,103]. There are mainly three phases in gasification of microalgae, which are removal of moisture, followed by devolatilization, and oxidation [100]. The removal of moisture content of microalgae biomass should take place in the temperature range of 50e200 C. When temperature reaches 450 C, microalgae biomass starts to decompose into combustible gases, such as H2, CO2, CO, and solid char or tar. Partial oxidation will occur at the last phase of gasification within the temperature range of 450e900 C, where the gasifying agent (steam/air) reacts with the combustible gases to enhance the yield of H2, CO, and CH4. In a previous study, Hirano et al. [104] had studied the volume composition yield of H2 in gasification process of microalgae biomass at different temperature. The study revealed that high temperature favored the yield of H2 and minimizes the solid remaining tar. Meanwhile, Chakinala et al. [105] studied the effect of catalytic supercritical water gasification process of Chlorella vulgaris for biohydrogen production. From the study, it was found that addition of nickel catalyst to the process could result higher H2 production with lower CO yield via enhanced wateregas-shift reaction. Previous study also studied the effect of temperature in catalytic gasification process of Cladophora fracta at a temperature range of 500e1000 C. It was reported that the syngas yield (H2 þ CO) had increased from 28 to 57 vol% with increase in temperature from 550 to 950 C [95]. Therefore it can be concluded that the presence of gasification agent (air or steam), presence of catalysts (e.g., dolomite, coal bottom ash, and nickel), and high reaction temperature (800e1000 C) play a vital role in gasification process to increase H2 yield [101].

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4.3 Pyrolysis Biomass pyrolysis is defined as a thermochemical process that undergoes either in complete absence of oxygen or in limited supply that gasification does not occur to an appreciable extent. Generally, pyrolysis process occurs at high temperature range of 600e900 C, low heating rates, and long residence time to obtain high yield of hydrogen-enriched syngas [106,107]. The operational conditions of pyrolysis are categorized into three different stages: slow pyrolysis, fast pyrolysis, and flash pyrolysis. Apparently, most of the reported studies were dedicated to biooil production from microalgae, such as slow pyrolysis of Tetraselmis chuii and Chlorella sp. to produce alkenes, phenol, and amides [108], and fast pyrolysis of Chlorella protothecoides for high yield of biooil production [109]. However, there are limited literature discussed about catalytic pyrolysis of microalgae for syngas production. Beneroso et al. [110] reported microwave induced pyrolysis of Scenedesmus almeriensis at a temperature range of 400e800 C. The study showed that more than 90 vol% of syngas and 50 vol% of hydrogen were attained in noncatalytic pyrolysis of microalgae. From Table 9.4, higher volume of H2 was obtained via microwave pyrolysis of microalgae as compared to other feedstocks. This observation could be due to high calorific value and low dissolved oxygen concentration of microwave biomass, which promoted better storage stability over other lignocellulose feedstock, which derived noncondensable gases (H2, CO, and CO2). Du et al. [111] reported that high-quality biooil and gases products could be attained in microwaveassisted pyrolysis using Chlorella sp. at the temperature of 500 C. The produced biooil had lower oxygen content with aromatic hydrocarbon meanwhile the gases product contained high volume concentration of hydrogen. This observation is due to the microplasmas and TABLE 9.4

Comparison of Microwave-Assisted Pyrolysis of Microalgae Biomass, Lignocellulosic, and Sludge for Syngas Production H2 Produced (vol%)

Syngas Produced (vol%)

References

Feedstock

Method

Rice husk

Microwave-assisted pyrolysis with char supported metallic catalysts

41.20

67.86

Zhang et al. (2015) [112]

Sawdust pellets

Microwave-assisted pyrolysis with corn stover biochar catalysts

20.43

43.03

Ren et al. (2014) [126]

Sewage sludge

Microwave-assisted pyrolysis without catalysts

45.00

87.90

Domínguez et al. (2008) [127]

Cornstalk

Microwave-assisted pyrolysis without catalysts

35.00

50.00

Zhao et al. (2013) [119]

Microalgae (Scenedesmus almeriensis)

Microwave-assisted pyrolysis without catalysts

50.00

90.00

Beneroso et al. (2013) [110]

5. CONCLUSIONS AND PERSPECTIVES

239

hotspots caused by microwave heating, which resulted in pseudocatalytic effect on the formation of hydrogen. Besides, high heating rate of microwave-assisted pyrolysis could speed up the mass transfer rate between microalgae biomass to perform thermal cracking, which enhanced the production of noncondensable gas (H2, CO, and CO2) [112].

4.4 Direct Combustion Direct combustion is a thermochemical technique in which the biomass is burned in open air or in the presence of excess air. In this process, the photosynthetically stored chemical energy of the biomass will be converted into gases. Generally, direct combustion is carried out inside a furnace, steam turbine, or boiler at a temperature range of 800e1000 C. This process is suitable for all types of biomass, which has low moisture content (<50%) [113]. The cost of energy production from direct combustion is slightly higher as compared to pyrolysis and gasification due to the need of pretreatment of biomass, such as dehydrating, cutting, and crushing, before introducing into the combustion chamber. There is another possibility for sustainable operation of microalgae biomass in direct combustion is coalealgae cofiring. Kadam [114] proposed that the idea of coalealgae cofiring could reduce the emission of CO2 by recycling the CO2 from the combustion process to microalgae cultivation. This leads to lower emission of GHGs into the atmosphere and a golden opportunity for carbon credit program [65]. However, there is still no extensive study that has been carried out to determine the feasibility of this technology.

5. CONCLUSIONS AND PERSPECTIVES Photosynthetic microalgae may prove to be an appealing alternative for biological H2 production because of their natural ability to capture solar energy and store it as chemical energy for water splitting. Significant breakthroughs have been made to improve the H2 productivity from microalgae, including genetic engineering and cultivation methods. C. reinhardtii appears to be the most promising strain to produce H2 through direct biophotolysis when cultivated in a sulfate-limited medium. However, the H2 production rate is still far lower compared to chemical-based processes, such as SME, which impedes the commercialization potential of this technology. Several issues and factors needed to be carefully considered before scaling-up the microalgae-based H2 production, such as nutrients source, photobioreactor design, and harvesting methods. Integrated systems with diversified biofuel (e.g., biodiesel) production from microalgae may be a possible solution to improve the overall life cycle energy balance and economic feasibility of this industry instead of solely producing H2.

Acknowledgments The authors would like to acknowledge the funding given by Universiti Sains Malaysia, through Short Term Grant with cost center 304/PJKIMIA/6315016, and the Ministry of Higher Education (MOHE) Malaysia, through Malaysia’s Rising Star Award 2016 (MRSA) with cost center 203/PJKIMIA/6071362. M.K. Lam would like to acknowledge the funding given by Universiti Teknologi PETRONAS (UTP), through YUTP-FRG with cost center 0153AA-H46.

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Further Reading [1] E. Sierra, F.G. Acién, J.M. Fernández, J.L. García, C. González, E. Molina, Characterization of a flat plate photobioreactor for the production of microalgae, Chem. Eng. J. 138 (1e3) (2008) 136e147. [2] T.M. Mata, A.A. Martins, N.S. Caetano, Microalgae for biodiesel production and other applications: a review, Renew. Sustain. Energy Rev. 14 (1) (2010) 217e232. [3] X. Zhao, Z. Song, H. Liu, Z. Li, L. Li, C. Ma, Microwave pyrolysis of corn stalk bale: a promising method for direct utilization of large-sized biomass and syngas production, J. Anal. Appl. Pyrol. 89 (1) (2010) 87e94.