The hydrolysis is carried out into two different photosynthesis stages: photosystem I (PSI) and photosystem II (PSII).
In photosystem I (PSI), production of the reductant of CO 2
taken place whereas in photosystem II (PSII) split water into
H 2 and O 2 (Bolatkhan et al. 2019). In direct photolysis, various
green algae (such as Chlamydomonas reinhardtii, Chlorococcumlittorale, Chlorella fusca, Platymonassubcordiformis,
Scenedesmus obliquusetc.) (Fan et al. 2016; Guan et al. 2004)
and cyanobacteria (Anabaena cylindrical, Oscillatoria brevis,
Nostocmuscorum, etc.) are widely used for hydrogen production (Das and Veziroglu 2008; Dutta et al. 2005).
Water is used as the primary feedstock in the direct
biophotolysis method which is inexpensive and available
everywhere. On the other side, hydrogen production is
prohibited by the suppressive effect of oxygen as a
by-product of photosynthesis and enzymatic catalysis which
is the major drawback of this method (Table 3) (Sun et al.
2019). Moreover, this process yields less hydrogen and
cost-inefficient in the industrial-scale production (Sakurai
and Masukawa 2007).
5.2.2 Indirect Biophotolysis
Indirect biophotolysis is carried out in two steps: photosynthesis and fermentation. Firstly, the synthesis of carbohydrates takes place under the light (Eq. 4). Secondly, the
hydrogen is produced from carbohydrates via anaerobic dark
fermentation (Eq. 5) (Hallenbeck and Benemann 2002;
Kossalbayev et al. 2020).
6H 2 O + 6CO 2 Sun light ! C 6 H 12 O 6 + 6O 2
ð4Þ
C 6 H 12 O 6 þ 12H 2 O Sun light ! 12H 12 + 6CO 2
ð5Þ
Cyanobacteria play a major role in the production of
hydrogen in indirect biophotolysis processes. It possesses
major enzymes such as nitrogenase and hydrogenase which
helped in metabolic functions for the hydrogen (Hallenbeck
and Benemann 2002; Kossalbayev et al. 2020).
5.2.3 Photo-Fermentation
In this process, lignocellulosic feedstocks are decomposed
into hydrogen and carbon dioxide by using photosynthetic
microorganisms such as Rhodobacter sp. in the presence of
sunlight and organic acids. Photo-fermentation occurs under
oxygen deficient condition with the optimal temperature of
30–35 °C and pH 7.0 (Eq. 6) (Argun and Kargi 2011). In
this process wide range of organic wastes such as fruits and
vegetable wastes or other lignocellulosic wastes can be used
as substrate for the production of biohydrogen (Özgür et al.
2010; Fascetti and Todini 1995).
CH 3 COOH + 2H 2 O Sun light ! 4H 2 + 2CO 2
ð6Þ
5.2.4 Dark Fermentation
Dark fermentation is environmentally friendly and widely
used method for biohydrogen production from organic
feedstocks. This process is undertaken in a dark and anaerobic environment in which anaerobic bacteria convert
carbohydrate-rich substrates into hydrogen (Toledo-Alarcón
et al. 2018). This process is carried out by different groups of
bacteria such as Enteric and Clostridia sp. (Khanna and Das
2013). In the dark fermentation, the first step is the glycolysis process in which glucose is fermented to pyruvate.
Then, under the anaerobic environment, pyruvate is oxidized
to acetyl-CoA, CO 2, and H 2 (Li and Fang 2007). Compared
to other biological production methods, dark fermentation
process is cost-effective, higher hydrogen production rate,
and faster conversion efficiencies. This process can utilize a
wide range of organic feedstocks including municipal
wastes, agriculture, and forest residues (Ghimire et al. 2015).
6 Metabolic Pathway of Hydrogen
Production
Hydrogen can be produced via biophotolysis,
photo-fermentation, and dark fermentation (Ding et al.
2016). Although biophotolysis by green microalgae and
cyanobacteria is a highly desirable process, the photochemical efficiency is low due to oxygen inhibition on
hydrogenase (Oh et al. 2013). Microbial fermentation (including dark fermentation and photo-fermentation) could be
one of the potential alternatives to produce biohydrogen.
However, except for the glucose, lignocellulosic biomass has
not been studied extensively for the hydrogen production
due to structural complexity of plant biomass. Usually, a
pretreatment of lignocellulosic biomass is an essential step
before hydrolysis to convert a complex biopolymer into
fermentable sugars (glucose and xylose). Hydrolysis of
cellulosic biomass is catalyzed by synergistic effect of
cellulase-endoglucanase,
cellobiohydrolase,
and
b-glucosidase to form glucose, whereas the hemicellulosic
components are catalyzed by hemicellulolytic enzymes such
as endo-xylanase, exo-xylanase, and b-xylosidase to form
xylose (Sharma et al. 2019). These sugars can be utilized
further in hydrogen production. However, the hydrolysis of
lignocellulosic biomass often produces some inhibitory
compounds such as phenolic and other aromatic compounds,
levulinic acid, aliphatic acids, and furan aldehydes, etc.
which inhibit the microbial growth and hinder the downstream processing of bioproducts (Jönsson et al. 2013;
Jönsson and Martín 2016). Thus, a direct bioconversion
(without pretreatment) of cellulosic and hemicellulosic biomass for hydrogen production is gaining popularity due to its
environmental and economic benefit. Some thermophilic
274
J. R. Khatiwada et al.
In photosystem I (PSI), production of the reductant of CO 2
taken place whereas in photosystem II (PSII) split water into
H 2 and O 2 (Bolatkhan et al. 2019). In direct photolysis, various
green algae (such as Chlamydomonas reinhardtii, Chlorococcumlittorale, Chlorella fusca, Platymonassubcordiformis,
Scenedesmus obliquusetc.) (Fan et al. 2016; Guan et al. 2004)
and cyanobacteria (Anabaena cylindrical, Oscillatoria brevis,
Nostocmuscorum, etc.) are widely used for hydrogen production (Das and Veziroglu 2008; Dutta et al. 2005).
Water is used as the primary feedstock in the direct
biophotolysis method which is inexpensive and available
everywhere. On the other side, hydrogen production is
prohibited by the suppressive effect of oxygen as a
by-product of photosynthesis and enzymatic catalysis which
is the major drawback of this method (Table 3) (Sun et al.
2019). Moreover, this process yields less hydrogen and
cost-inefficient in the industrial-scale production (Sakurai
and Masukawa 2007).
5.2.2 Indirect Biophotolysis
Indirect biophotolysis is carried out in two steps: photosynthesis and fermentation. Firstly, the synthesis of carbohydrates takes place under the light (Eq. 4). Secondly, the
hydrogen is produced from carbohydrates via anaerobic dark
fermentation (Eq. 5) (Hallenbeck and Benemann 2002;
Kossalbayev et al. 2020).
6H 2 O + 6CO 2 Sun light ! C 6 H 12 O 6 + 6O 2
ð4Þ
C 6 H 12 O 6 þ 12H 2 O Sun light ! 12H 12 + 6CO 2
ð5Þ
Cyanobacteria play a major role in the production of
hydrogen in indirect biophotolysis processes. It possesses
major enzymes such as nitrogenase and hydrogenase which
helped in metabolic functions for the hydrogen (Hallenbeck
and Benemann 2002; Kossalbayev et al. 2020).
5.2.3 Photo-Fermentation
In this process, lignocellulosic feedstocks are decomposed
into hydrogen and carbon dioxide by using photosynthetic
microorganisms such as Rhodobacter sp. in the presence of
sunlight and organic acids. Photo-fermentation occurs under
oxygen deficient condition with the optimal temperature of
30–35 °C and pH 7.0 (Eq. 6) (Argun and Kargi 2011). In
this process wide range of organic wastes such as fruits and
vegetable wastes or other lignocellulosic wastes can be used
as substrate for the production of biohydrogen (Özgür et al.
2010; Fascetti and Todini 1995).
CH 3 COOH + 2H 2 O Sun light ! 4H 2 + 2CO 2
ð6Þ
5.2.4 Dark Fermentation
Dark fermentation is environmentally friendly and widely
used method for biohydrogen production from organic
feedstocks. This process is undertaken in a dark and anaerobic environment in which anaerobic bacteria convert
carbohydrate-rich substrates into hydrogen (Toledo-Alarcón
et al. 2018). This process is carried out by different groups of
bacteria such as Enteric and Clostridia sp. (Khanna and Das
2013). In the dark fermentation, the first step is the glycolysis process in which glucose is fermented to pyruvate.
Then, under the anaerobic environment, pyruvate is oxidized
to acetyl-CoA, CO 2, and H 2 (Li and Fang 2007). Compared
to other biological production methods, dark fermentation
process is cost-effective, higher hydrogen production rate,
and faster conversion efficiencies. This process can utilize a
wide range of organic feedstocks including municipal
wastes, agriculture, and forest residues (Ghimire et al. 2015).
6 Metabolic Pathway of Hydrogen
Production
Hydrogen can be produced via biophotolysis,
photo-fermentation, and dark fermentation (Ding et al.
2016). Although biophotolysis by green microalgae and
cyanobacteria is a highly desirable process, the photochemical efficiency is low due to oxygen inhibition on
hydrogenase (Oh et al. 2013). Microbial fermentation (including dark fermentation and photo-fermentation) could be
one of the potential alternatives to produce biohydrogen.
However, except for the glucose, lignocellulosic biomass has
not been studied extensively for the hydrogen production
due to structural complexity of plant biomass. Usually, a
pretreatment of lignocellulosic biomass is an essential step
before hydrolysis to convert a complex biopolymer into
fermentable sugars (glucose and xylose). Hydrolysis of
cellulosic biomass is catalyzed by synergistic effect of
cellulase-endoglucanase,
cellobiohydrolase,
and
b-glucosidase to form glucose, whereas the hemicellulosic
components are catalyzed by hemicellulolytic enzymes such
as endo-xylanase, exo-xylanase, and b-xylosidase to form
xylose (Sharma et al. 2019). These sugars can be utilized
further in hydrogen production. However, the hydrolysis of
lignocellulosic biomass often produces some inhibitory
compounds such as phenolic and other aromatic compounds,
levulinic acid, aliphatic acids, and furan aldehydes, etc.
which inhibit the microbial growth and hinder the downstream processing of bioproducts (Jönsson et al. 2013;
Jönsson and Martín 2016). Thus, a direct bioconversion
(without pretreatment) of cellulosic and hemicellulosic biomass for hydrogen production is gaining popularity due to its
environmental and economic benefit. Some thermophilic
274
J. R. Khatiwada et al.