International Journal of Hydrogen Energy 33 (2008) 258 – 263

www.elsevier.com/locate/ijhydene

Biohydrogen as a renewable energy resource—Prospects and potentials

Shireen Meher Kotay, Debabrata Das ∗
Department of Biotechnology, Indian Institute of Technology, Kharagpur 721302, INDIA

Received 18 June 2007; accepted 9 July 2007

Available online 4 September 2007

Abstract
Biohydrogen holds the promise for a substantial contribution to the future renewable energy demands. It seems particularly suitable for
relatively small-scale, decentralized systems, integrated with agricultural and industrial activities or waste processing facilities. Biohydrogen
is considered as an important key to a sustainable world power supply and is currently being seen as the versatile fuel of the future, with the
potential to replace fossil fuels. It has the key prospective to become the ideal means among the range of renewable H2 production technologies
presently existing. This review attempts to delineate the prospects and potentials of biohydrogen as renewable energy resource.
䉷 2007 International Association for Hydrogen Energy. Published by Elsevier Ltd. All rights reserved.

Keywords: Biohydrogen; Renewable energy; Waste-to-energy; Bioremediation

1. Introduction closely related to the increased use of hydrogen in refineries

Hydrogen holds the promise as a dream fuel of the future with
many social, economic and environmental benefits to its credit.
It has the long-term potential to reduce the dependence on for-
eign oil and lower the carbon and criteria emissions from the
transportation sector. Only in the last decade that the idea of a
post-fossil fuel hydrogen-based economy started to gain main-
stream interest. Molecular H2 has the highest energy content
per unit weight among the known gaseous fuels (143 GJ ton−1 )
[1] and is the only carbon-free fuel which ultimately oxidizes
to water as a combustion product. Therefore burning hydrogen
not only has the potential to meet a wide variety of end use
applications but also does not contribute to greenhouse emis-
sion, acid rain or ozone depletion. The use of hydrogen will
contribute to significant reduction of these energy-linked envi-
ronmental impacts.
H2 can be used either as the fuel for direct combustion in an
internal combustion engine or as the fuel for a fuel cell. The
largest users of H2 , however, are the fertilizer and petroleum
industries with, respectively, 50% and 37%. Sales of hydrogen
have increased by 6% annually in the last five years, which is
∗ Corresponding author. Tel.: +91 3222 278053, +91 3222 283758;
fax: +91 3222 255303.
E-mail address: ddas@hijli.iitkgp.ernet.in (D. Das).
0360-3199/$ - see front matter 䉷 2007 International Association for Hydrogen Energy. Published by Elsevier Ltd. All rights reserved.
doi:10.1016/j.ijhydene.2007.07.031
as a result of stricter standards for fuel quality [2]. The Global
Hydrogen Vision envisions hydrogen as a flexible, safe, afford-
able, domestic energy resource to be used in all sectors of the
economy and all regions of the world. Hydrogen will become
world’s “clean energy choice”, joining electricity as a primary
energy carrier and providing the foundation for a globally sus-
tainable energy system [3]. It has a wide variety of applications,
including fuel for automobiles; distributed and central electric-
ity and thermal energy generation (Fig. 1).
2. Barriers for biohydrogen production
Technical challenges in achieving a hydrogen economy in-
clude lowering the cost of hydrogen production, delivery, stor-
age, conversion, and end use applications. Although hydrogen
is the most abundant element in the universe, it must be pro-
duced from other hydrogen-containing compounds such as fos-
sil fuels, biomass, or water. Each method of production requires
a source of energy, i.e., thermal (heat), electrolytic (electricity),
or photolytic (light) energy (Table 1). In other words they are
highly energy intensive and not always environmentally benign
[4]. Moreover, there is a growing concern arising out of the
projection of fossil fuel shortfall towards the middle of twenty-
first century. Under this perspective, biological hydrogen
 S. Meher Kotay, D. Das / International Journal of Hydrogen Energy 33 (2008) 258 – 263 259

Fig. 1. Chronology of fuel usage by mankind.

Table 1
Scientific and technical barriers for biohydrogen production

Type of barrier Barrier Putative solution

Basic science
Organism Bacteria do not produce more than 4 mol Isolate more novel microbes and combinational screen for H2
H2 /mol glucose naturally production rates yields, and durability
Genetic manipulation of established bacteria
Enzyme (hydrogenase) Hydrogenase overexpression not stable Greater understanding of the enzyme regulation and expression
O2 sensitivity Mutagenic studies
H2 feed back inhibition Low H2 partial pressure fermentation

Fermentative
Feedstock High cost of suitable feedstock (glucose) Renewable biomass as feedstock
Low yield using renewable biomass Co-digestion/use of microbial consortia which can increase the
yield
Strain Lack of industrial-suitable strain Development of industrially viable strain(s)/consortia
Process Commercially feasible product yield Hybrid system (photo + dark fermentation)
Incomplete substrate utilization Link fermentation to a second process that makes both economi-
cally possible
Sustainable process Application and utilization of fermentation tools such as continuous
culture
Sterilization Development of low-cost stream sterilization technology/process
that can bypass sterilization

Engineering
Reactor Lack of kinetics/appropriate reactor design Incorporation of process engineering concepts to develop a suitable
for H2 production reactor for the defined strain/process
Light intensity in case of photo-bioreactor Flat panel or hollow tube reactor can be employed
Thermodynamic Thermodynamic barrier NAD(P)H → H2 Reverse electron transport to drive H2 production past barrier
(+4.62 kJ/mol)
Hydrogen H2 purification/separation Selection absorption of CO2 /H2 S
Storage Basic studies on H2 storage

production assumes paramount importance as an alternative and
renewable energy resource.
Biological processes, unlike their chemical or electrochemi-
cal counterparts are catalyzed by microorganisms in an aqueous
environment at ambient temperature and pressure. Furthermore,
these techniques are well suited for decentralized energy pro-
duction in small-scale installations in locations where biomass
or wastes are available, thus avoiding energy expenditure and
costs for transport. From thermodynamic perspective, as the
organic substrates dissolved and diluted in wastewater are in a
high entropy state, it is somewhat difficult to obtain their com-
bustion enthalpy by mechanical means [5]. On the contrary in
biological processes microorganisms can recover and concen-
trate the energy from high water content organic resources such
as, industrial wastewater and sludge in a usable form. Thus,
biohydrogen production, in a sense, is an entropy reducing
260 S. Meher Kotay, D. Das / International Journal of Hydrogen Energy 33 (2008) 258 – 263
process, which could not be realized by mechanical or chem-
ical systems. The aim is to develop a blue print for hydrogen
from biomass plant [4,5].
3. Biohydrogen as renewable energy
Interest in biohydrogen started getting prominence in early
90s, when it became apparent that atmospheric pollution by
fossil fuels is not only unhealthy locally, but might also cause
significant climate changes globally. As a result, biological hy-
drogen production became a focus of Governmental support,
particularly in Germany, the US and Japan, with meager efforts
in the other countries. Biological hydrogen production now has
gained such a tremendous impetus that more than 30 countries
have chosen to invest in the concerned research [3,4]. Realiza-
tion of practical processes for biohydrogen production would
result in a major, novel biological source of sustainable and
renewable energy, without greenhouse gas emissions or envi-
ronmental pollution. However, development of such practical
processes will require significant scientific and technological
advances, and relatively long-term (> 10 yr) basic and applied
R&D. For H2 to be renewable, it must come from renewable
sources.
Major resources in biomass include agricultural crops and
their waste byproducts, lignocellulosic products such as wood
and wood waste, waste from food processing and aquatic plants
and algae, and effluents produced in the human habitat. Energy
from water-containing biomass such as sewage sludge, agri-
cultural and livestock effluents as well as animal excreta are
recovered mainly by microbial fermentation. Biomass as en-
ergy source is characterized in the form of both flow and stock.
If these resources were used under appropriate control, they
would become the major source of energy in the future.
Biomass has the potential to become a significant source
of renewable hydrogen. In addition, many of the processes
which produce hydrogen from biomass are complementary to
those that produce biomaterials. Therefore, countries with large
agricultural economies have potential for significant economic
development through incorporation of bioenergy into bioindus-
try. While renewable hydrogen technologies that use low value
waste biomass as feedstock have great potential to become cost
competitive, it is currently more expensive to produce hydro-
gen from biomass than it is to derive it from natural gas. In-
dependent of the source of hydrogen, there are many logistical
and market challenges that must also be overcome before a hy-
drogen economy can become a reality:
biomass + O2 → CO + H2 + CO2 + energy.
The critical challenges of the hydrogen fermentation are low
hydrogen conversion efficiency and unstable hydrogen produc-
tion [7]. One of the important factors governing the hydrogen
conversion efficiency is process engineering, such as bioreac-
tor design and operating parameters. Improving reactor design
and optimizing the operating parameters such as pH, hydraulic
retention time, and temperature could enhance the hydrogen
conversion efficiency. The unstable hydrogen production is pos-
sibly attributed to the metabolic shift of hydrogen producing
bacteria, and this could be minimized by the study of microbial
growth.
4. Microbiology of biohydrogen production
Nature in the form of microorganisms has been using hy-
drogen as a primary fuel source for billions of years, and has
solved the problem of converting hydrogen to electricity by
means of the biocatalyst, hydrogenase (Table 2). Although they
catalyze a deceptively simple reaction, they have proved to be
some of the most complex and ingenious bioinorganic struc-
tures known. Microorganisms generate hydrogen for two prin-
ciple reasons, first to dispose of excess reducing equivalents or
as a byproduct in nitrogen fixation.
Microbial H2 production is an attractive process for sup-
plying a significant share of the H2 required for the near
future. Galore microbial species, belonging to the genera En-
terobacter, Citrobacter, Bacillus, and Clostridium are reported
(Table 3) to produce hydrogen through dark fermentation
[6]. Apart from pure cultures, various mixed micro-flora and
co-cultures have also been explored for hydrogen production
from carbohydrates [6–8]. Nevertheless, the quest for ideal mi-
crobe(s) for microbial H2 production has thrust the researchers
to screen various sources.
5. Biohydrogen production: conventional and novel
approaches
Biological processes are carried out largely at ambient tem-
peratures and pressures, and hence are less energy intensive than
chemical or electrochemical ones. A large number of microbial
species, including significantly different taxonomic and physi-
ological types, can produce hydrogen. Biological processes use
the enzyme hydrogenase or nitrogenase as hydrogen produc-
ing protein. This enzyme regulates the hydrogen-metabolism
of uncountable prokaryotes and some eukaryotic organisms in-
cluding green algae. The functioning of nitrogenase, as well as
hydrogenase, is linked with the utilization of the products of
photosynthetic reactions that generate reductants from water.
The processes of biological hydrogen production can be
broadly classified into two distinct groups. One is light depen-
dent and the other is light independent process. Light mediated
processes include direct or indirect biophotolysis and photofer-
mentation whereas dark fermentation is the major light inde-
pendent process (Fig. 2). Biophotolysis involves light-driven
decomposition of water in the presence of micro-algae or
cyanobacteria (Table 4 ). An attractive and even more favored
possibility is the direct splitting of water to generate hydrogen
by solar radiation [4]. This H2 O splitting can be achieved either
in photochemical cells where, for example, TiO2 is illuminated
as the catalyst, or by applying photovoltaics, which indirectly
utilize solar radiation for the electrolysis of water into H2 and
O2 [5]. Photosynthetic microorganisms are able to convert
directly solar energy to hydrogen from organic or inorganic
substrates or from water. Despite its relatively lower yields
of hydrogen, the fermentative route is a promising method of
biohydrogen production due to its higher rate of H2 evolution
 S. Meher Kotay, D. Das / International Journal of Hydrogen Energy 33 (2008) 258 – 263 261

Table 2
Physiological diversity of biohydrogen production

Heterotrophic Photoheterotrophic

Fermentative H2 production from biomass by heterotrophic bacteria Biophotolytic H2 production by green algae or cyanobacteria (water splitting)
(C6 H12 O6 + 2H2 O → 2CH3 COOH + 2CO2 + 4H2 ) (12H2 O → 12H2 + 6O2 )
H2 production from CO by photosynthetic bacteria Photoproduction of H2 from biomass by phototrophic bacteria
(CO + H2 O → H2 + CO2 ) (C6 H12 O6 + 6H2 O → 6CO2 + 12H2 )

Table 3
Imperative hydrogen producing microbial strains reported and their H2 yield

Organism Substrate Process Maximum References

yield of H2
(mol H2 /mol
substrate)

Enterobacter aerogenes HU-101 Glucose Batch, (blocking metabolites formation) 1.17 Mahyudin et al. [9]
(mutant AY-2)
Enterobacter aerogens Molasses Ar sparging, batch 1.58 Tanisho et al. [10]
Enterobacter aerogens Molasses Batch 0.52 Tanisho et al. [10]
Clostridium butyricum Glucose N2 sparging continuous 1.4–2.3 Kataoka et al. [11]
Enterobacter cloacae Glucose Continuous (immobilized bioreactor) 2.3 Kumar and Das [16]
IIT BT 08
Enterobacter cloacae DM11 Glucose Continuous (immobilized bioreactor) 3.8 Kumar et al. [17]
Citrobacter sp. Y19 Glucose Batch Ar sparging 2.49 Oh et al. [18]
Rhodopseudomonas Glucose Batch, with intermittent purging of Ar 2.76 Oh et al. [19]
palustris P4
Mixed culture (predominantly Glucose N2 sparging, continuous HRT:8.5 h 1.43 Mizuno et al. [12]
Clostridium sp.)
Mixed microflora Wheat starch co-product N2 sparging continuous 1.9 Hussy et al. [13]
Mixed microflora 0.75% soluble starch Chemostat HRT: 17 h 2.14 Lay [14]
Mixed microflora Sewage-sludge Anaerobic and acidogenic digestion 1.7 Lin and Chang [15]
(Mesophilic)

Fig. 2. Various approaches of biohydrogen production.

in the absence of any light source as well as the versatility of
the substrates used.
Anaerobic heterotrophic microorganisms can form hydro-
gen during the oxidation of organic substrates. A great advan-
tage of fermentation is fast degradation of solids and other
complex organics found in wastes and agricultural products.
On the other hand, fermentation today converts only about 15%
of the energy to hydrogen [7]. While fermentation is fast, it
is not yet efficient for capturing the energy value of biomass
to hydrogen. The goal is to increase the biohydrogen yield to
262 S. Meher Kotay, D. Das / International Journal of Hydrogen Energy 33 (2008) 258 – 263

Table 4
Comparison of important biological hydrogen production processes

Process Advantages Representative organism Maximum References

reported rate
(mmol H2 /L h)

Conventional
Direct biophotolysis Can produce H2 directly from water and sunlight Chlamydomonas reinhardtii 0.07 [8,9]
Solar conversion energy increased by tenfold as
compared to trees, crops
Indirect biophotolysis Can produce H2 from water Has the ability to fix Anabaena variabilis 0.36 [10,11]
N2 from atmosphere
Photo fermentation A wide spectral light energy can be used by these Rhodobacter spheroides 0.16 [12]
bacteria
Can use different waste materials like distillery ef-
fluents, waste, etc.
Dark fermentation It can produce H2 all day long without light Enterobacter cloacae DM 11 75.60 [13]
A variety of carbon sources can be used as substrates Clostridium sp. strain No. 2 64.50 [14]
It produces valuable metabolites such as butyric,
lactic and acetic acids as by products
It is anaerobic process, so there is no O2 limitation
problem
Novel
Two-stage fermentation Stoichiometric yield of 12 mol H2 per mol hexose Enterobacter cloacae 51.20 [15]
(dark + photo) represents the ultimate target for biohydrogen DM 11 + Rhodobacter
sphaeroidesOU 001
Mixed microbial flora + 47.92 [16]
Rhodobacter sphaeroides
OU 001

around 85%. While hydrogen fermentations have been demon-
strated in the laboratory, yields have been low and it is uncer-
tain whether this technology can be developed to provide high
yields of hydrogen and become economically competitive with
gasoline or with alternative hydrogen production pathways. It
is speculated that a fermentation yield of 10 mol of hydrogen
per mole of glucose and a glucose cost of 5 cents per dry pound
will be required for this process to approach hydrogen costs
that are competitive with traditional fuels.
However, a combination of dark and photofermentation in a
two-stage hybrid system can improve the overall yield of hy-
drogen [20–22]. The synergy of the process lies in the maxi-
mum utilization of the substrate which otherwise fails to achieve
complete conversion due to thermodynamic limitation.
In the first stage, the biomass is fermented to acetate, carbon
dioxide and hydrogen in a thermophilic dark fermentation. In a
separate photobioreactor acetate is converted to hydrogen and
carbon dioxide [23,24]. The combination could be expected to
reach as close to the theoretical maximum production of 12 mol
of H2 (mol glucose)−1 equivalent as possible, according to the
following reactions:
(i) Stage I—dark fermentation (facultative anaerobes)
C6 H12 O6 + 2H2 O → 2CH3 COOH + 2CO2 + 4H2 .
(ii) Stage II—Photofermentation (photosynthetic bacteria)
2CH3 COOH + 4H2 O → 8H2 + 4CO2 .
Both basic and applied R&D activities on biological produc-
tion of hydrogen have drawn significant attention over past few
decades. Research progress for identifying the major techno-
economic bottlenecks of various bioprocesses for commercial
production of hydrogen appears promising. However, high hy-
drogen yield remains to be the ultimate goal and challenge
for the biohydrogen research and development. Enhancement
in hydrogen yield may be possible by using suitable micro-
bial strain, process modification, efficient bioreactor design and
also genetic and metabolic engineering technique, to redirect
metabolic pathway [20]. Hydrogen is currently more expensive
than other fuel options, so it is likely to play a major role in
the economy in the long run, if technology improvements suc-
ceed in bringing down costs. Biological hydrogen production,
employing renewable biomass may be a potential answer to
overcome some of the economic constraints to fulfill many of
our energy needs. There is scope to use sugarcane juice, mo-
lasses or distillery effluent as substrates, because they contain
sugar in significant quantities. Therefore, production as well
as unit energy cost of biohydrogen would be reduced drasti-
cally. However, a rigorous techno-economic analysis is nec-
essary to draw a cost-effective comparison between biologi-
cally produced hydrogen and the various other conventional
fossil fuels.
6. Conclusions
The vision for a hydrogen future is one based on clean sus-
tainable renewable energy supply of global proportions that
 S. Meher Kotay, D. Das / International Journal of Hydrogen Energy 33 (2008) 258 – 263
plays a key role in all sectors of the economy. If the technolog-
ical potential of hydrogen is realized, it will contribute to the
sustainable growth of the world economy by facilitating a sta-
ble supply of energy and by helping to reduce future emissions
of greenhouse gases. Research on Biogenic H2 has proclaimed
its promise and primacy over other routes. Conclusively, for
H2 to be renewable it must come from renewable sources and
biohydrogen production is the ideal means. However, extensive
research and development efforts are required for developing
biohydrogen production methods for commercialization. Coop-
erative efforts among nations can help speed effective progress
towards these goals.
References
[1] Boyles D. Bioenergy technology—thermodynamics and costs. New York:
Wiley; 1984.
[2] Elam CC, Gregoire Padro CE, Sandrock G, Luzzi A, Lindblad P, Hagen
E-F. Realizing the hydrogen future: the International Energy Agency’s
efforts to advance hydrogen energy technologies. Int J Hydrogen Energy
2003;28:601–7.
[3] Benemann JR. Hydrogen biotechnology: progress and prospects. Nat
Biotechnol 1996;14:1101–3.
[4] Momirlan M, Veziroglu TN. Current status of hydrogen energy.
Renewable Sustainable Energy Rev. 2002;6:141–79.
[5] Rao KK, Hall DO. Hydrogen production by cyanobacteria: potential,
problems and prospects. J Mar Biotechnol 1996;4:10–5.
[6] Nandi R, Sengupta S. Microbial production of hydrogen: an overview.
Crit Rev Microbiol 1998;24:61–84.
[7] Das D, Verziroglu TN. Hydrogen production by biological processes: a
survey of literature. Int J Hydrogen Energy 2001;26:13–28.
[8] Levin DB, Pitt L, Love M. Biohydrogen production: prospects and
limitations to practical application. Int J Hydrogen Energy 2004;29:
173–85.
[9] Mahyudin AR, Furutani Y, Nakashimada Y, Kakizono T, Nishio N.
Enhanced hydrogen production in altered mixed acid fermentation
of glucose by Enterobacter aerogenes. J Ferment Bioeng 1997;83(4):
358–363.
263
[10] Tanisho S, Kuromoto M, Kadokura N. Effect of CO2 removal
on hydrogen production by fermentation. Int J Hydrogen Energy
1998;23:559–63.
[11] Kataoka N, Miya A, Kiriyama K. Studies on hydrogen production by
continuous culture system of hydrogen producing anaerobic bacteria.
Water Sci Technol 1997;36:41–7.
[12] Mizuno O, Dinsdale R, Hawkes FR, Hawkes DL, Noike T. Enhancement
of hydrogen production from glucose by nitrogen gas sparging.
Bioresource Technol 2000;73:59–65.
[13] Hussy I, Hawkes FR, Dinsdale R, Hawkes DL. Continuous fermentative
hydrogen production from a wheat starch co-product by mixed
microflora. Biotech Bioeng 2003;84(6):619–26.
[14] Lay J-J. Modelling and optimization of anaerobic digested
sludge converting starch to hydrogen. Biotech Bioeng 2000;68(3):
269–78.
[15] Lin C-Y, Chang R-C. Hydrogen production during anaerobic acidogenic
conversion of glucose. J Chem Tech Biotech 1999;74(6):496–500.
[16] Kumar N, Das D. Enhancement of hydrogen production by Enterobacter
cloacae IIT- BT 08. Process Biochem 2000;35:589–93.
[17] Kumar N, Ghosh A, Das D. Redirection of biochemical pathways for
the enhancement of H2 production by Enterobacter cloacae. Biotechnol
Lett 2001;23:537–41.
[18] Oh Y-K, Seol E-H, Kim JR, Park S. Fermentative biohydrogen production
by a new chemoheterotrophic bacterium Citrobacter sp. Y19. Int J
Hydrogen Energy 2003;28:1353–9.
[19] Oh Y-K, Seol E-H, Yeol Lee E, Park S. Fermentative hydrogen production
by a new chemolithotrophic bacterium Rhodopseudomonas palustris P4.
Int J Hydrogen Energy 2002;27:1373–9.
[20] Nath K, Das D. Improvement of fermentative hydrogen production:
various approaches. Appl Microbiol Biotechnol 2004;65:520–9.
[21] Hawkes FR, Dindale R, Hawkes DL, Hussy I. Sustainable fermentative
biohydrogen: challenges for process optimization. Int J Hydrogen Energy
2002;27:1339–47.
[22] Momirlan M, Veziroglu TN. Recent directions of world hydrogen
production. Renewable Sustainable Energy Rev 1999;3:219–31.
[23] Nath K, Das D. Amelioration of biohydrogen production by a two-stage
fermentation process. Ind Biotechnol 2006;2:44–7.
[24] Eroglu E, Eroglu I, Gunduz U, Turker L, Yucel M. Biological hydrogen
production from olive mill wastewater with two-stage processes. Int J
Hydrogen Energy 2006;31(11):1527–35.