The document discusses hydrogen production through microbial biomass conversion, emphasizing the capabilities of microorganisms to digest biomass and generate hydrogen via fermentation and microbial electrolysis cells (MECs). It highlights strategies to optimize production rates and yields, as well as the potential for scaling up these technologies using various feedstocks. Additionally, specific examples of hydrogen-producing bacteria and research on enhancing efficiency and production methods are mentioned.

1. Hydrogen Production: Microbial Biomass Conversion
Dr. Naveen Gaurav
Associate Professor and Head
Department of Biotechnology
Shri Guru Ram Rai University
Dehradun

2. Hydrogen Production: Microbial Biomass Conversion
Microbial biomass conversion processes take advantage of the ability of microorganisms to
consume and digest biomass and release hydrogen. Depending on the pathway, this
research could result in commercial-scale systems in the mid- to long-term timeframe that
could be suitable for distributed, semi-central, or central hydrogen production scales,
depending on the feedstock used.
HOW DOES IT WORK?
In fermentation-based systems, microorganisms, such as bacteria, break down organic
matter to produce hydrogen. The organic matter can be refined sugars, raw biomass
sources such as corn stover, and even wastewater. Because no light is required, these
methods are sometimes called "dark fermentation" methods.
In direct hydrogen fermentation, the microbes produce the hydrogen themselves. These
microbes can break down complex molecules through many different pathways, and the
byproducts of some of the pathways can be combined by enzymes to produce hydrogen.
Researchers are studying how to make fermentation systems produce hydrogen faster
(improving the rate) and produce more hydrogen from the same amount of organic matter
(increasing the yield).
Microbial electrolysis cells (MECs) are devices that harness the energy and protons
produced by microbes breaking down organic matter, combined with an additional small
electric current, to produce hydrogen. This technology is very new, and researchers are
working on improving many aspects of the system, from finding lower-cost materials to
identifying the most effective type of microbes to use.

3. WHY IS THIS PATHWAY BEING CONSIDERED?
Biomass is an abundant domestic resource, and many microbes have evolved to efficiently
break down biomass to produce hydrogen and other products. Fermentation has already
been used as an industrial technology to generate biofuels and other products, and many
of the challenges to scaling up systems have been addressed for different products,
allowing hydrogen researchers to focus on the challenges unique to hydrogen production.
MEC-based systems have the potential to produce hydrogen from resources that otherwise
can’t be used for fuel production, and could reduce the large amount of energy normally
needed for wastewater treatment while producing a valuable fuel in the form of hydrogen.
These two pathways can be combined to maximize the hydrogen yield from the starting
biomass feedstock.

4. Process: Hydrogen production by microbes
hydrogen-producing bacteria and algae: Clostridium, Enterobacter,
Klebsiella, Citrobacter, Bacillus and Cyanobacteria, Anabaena

5. RESEARCH FOCUSES ON OVERCOMING CHALLENGES
1. Improving the rates and yields of hydrogen production from fermentation processes
through a number of methods such as microbial strain improvements, reactor system
optimization, and identifying feedstock sources and processing methods with the highest
yields.
2. Developing MEC systems that can be scaled up to commercially relevant sizes while
maintaining the production rates and system efficiencies seen at the bench scale and
minimizing the costs of the reactor components.
Examples:

6. Nitrogenase-dependent hydrogen production
Benemann and Weare demonstrated that a nitrogen-fixing cyanobacterium, Anabaena
cylindrica, produced hydrogen and oxygen gas simultaneously in an argon atmosphere for
several hours. Nitrogenase is responsible for nitrogen-fixation and is distributed mainly
among prokaryotes, including cyanobacteria, but does not occur in eukaryotes, under which
microalgae are classified. Molecular nitrogen is reduced to ammonium with consumption of
reducing power (e' mediated by ferredoxin) and ATP. The reaction is substantially irreversible
and produces ammonia:
N2 + 6H1+ + 6e- =2HN3
12ATP ↔ 12(ADP+Pi)
However, nitrogenase catalyzes proton reduction in the absence of nitrogen gas (i.e. in an
argon atmosphere).
2H+ + 2e- = H2
4ATP ↔ 4(ADP+Pi)
Hydrogen production catalyzed by nitrogenase occurs as a side reaction at a rate of one-third
to one-fourth that of nitrogen-fixation, even in a 100% nitrogen gas atmosphere.
Nitrogenase itself is extremely oxygen-labile. Unlike in the case of hydrogenase, however,
cyanobacteria have developed mechanisms for protecting nitrogenase from oxygen gas and
supplying it with energy (ATP) and reducing power. The most successful mechanism is the
localization of nitrogenase in the heterocysts of filamentous cyanobacteria Vegetative cells
(ordinary cells) in filamentous cyanobacteria carry out oxygenic photosynthesis. Organic
compounds produced by CO; reduction are transferred into heterocysts and are decomposed to
provide nitrogenase with reducing power. ATP can be provided by PSI-dependent and
anoxygenic photosynthesis within heterocysts.

7. Investigations into prolongation and optimization of hydrogen production, revealed that
the hydrogen-producing activity of cyanobacteria was stimulated by nitrogen starvation.
The presence and physiological roles of hydrogenases in nitrogen-fixing cyanobacteria
remains controversial, but 'uptake' hydrogenase appears to consume and re-use hydrogen
gas, resulting in a decrease in net hydrogen production. Asada and Kawamura reported
aerobic hydrogen production by a nitrogen-fixing Anabaena sp., believed to be an uptake
hydrogenase-deficient strain. After being cultured for 12 days, the strain accumulated
approximately 10% hydrogen and 70% oxygen gas within the gas phase of the vessel, by the
nitrogenase side reaction, even in the presence of air.
Miyamoto et al. conducted outdoor experiments on hydrogen production by Anabaena
cylindrica, in California. A nitrogen-starved culture of the cyanobacteria was continuously
sparged by argon-based gas, while the hydrogen content of the effluent gas was measured.
The average conversion efficiency over a period of one month (combustion energy of
hydrogen gas produced by cyanobacteria/incident solar energy into the photo-bioreactor
area) was approximately 0.2%.
Mitsui and co-workers extensively screened cyanobacteria for their hydrogen-producing
ability, and tested Miami BG-7, one of the most potent hydrogen-producing cyanobacteria,
in an outdoor culture. These workers also isolated a unicellular aerobic nitrogen
fixer, Synechococcus sp. Miami BG043511, and with the use of synchronous culture
techniques, discovered a new mechanism for protecting and driving oxygen-labile
nitrogenase in non-heterocystous and oxygen-evolving cells. This strain is also a potent
hydrogen-producer, having an estimated conversion efficiency of 3.5% based on PAR using
an artificial light source.

8. Hydrogen production by photosynthetic bacteria
Photosynthetic bacteria undergo anoxygenic photosynthesis with organic compounds
or reduced sulfur compounds as electron donors. Some non-sulfur photosynthetic
bacteria are potent hydrogen producers, utilizing organic acids such as lactic, succinic
and butyric acids, or alcohols as electron donors. Since light energy is not required for
water oxidation, the efficiency of light energy conversion to hydrogen gas by
photosynthetic bacteria, is in principle much higher than that by cyanobacteria.
Hydrogen production by photosynthetic bacteria is mediated by nitrogenase activity,
although hydrogenases may be active for both hydrogen production and hydrogen
uptake under some conditions. Miyake and Kawamura demonstrated a maximum
energy conversion efficiency (combustion energy of hydrogen gas produced/incident
light energy) of 6 to 8% using Rhodobacter sp. in laboratory experiments
Thank you
References: Online notes, notes from research papers
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