10-developments in hydrogen production through microbial processes; pakistan’s prospective

8/6/2019 10-Developments in Hydrogen Production through Microbial Processes; Pakistans Prospective

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June 2011, Volume 2, No.3International Journal of Chemical and Environmental Engineering

Developments in Hydrogen Production through

Microbial Processes; Pakistans Prospective

Abdul Waheed Bhutto1, , *

, Aqeel Ahmed Bazmi2,3

, Muhammad Nadeem Kardar2

and Muhammad Yaseen2,

Gholamreza Zahedi3

and Sadia Karim1Department of Chemical Engineering, Dawood College of Engineering

and Technology, M.A.Jinnah Road, Karachi-Pakistan2 Biomass Conversion Research Centre (BCRC), Department of Chemical Engineering,COMSATS Institute of

Information Technology, Defence Road, Off Raiwind Road, Lahore-Pakistan.3Process Systems Engineering Centre (PROSPECT), Chemical Engineering Department, Faculty of Chemical

Engineering, Universiti Teknologi Malaysia, Skudai 81310, Johor Bahru (JB), Malaysia.Affiliated member BCRC*Corresponding Author Email: [email protected]

Abstract

Currently, hydrogen (H2) is primarily used in the chemical industry as a reactant, but it is being proposed as future fuel. H 2 has greatpotential as an environmentally clean energy fuel and as a way to reduce reliance on imported energy sources. A combination of theneed to cut carbon dioxide emissions, the prospect of increasingly expensive oil and the estimated growth in the world's vehicle fleetindicates that only H2 can plug the gap. There are many processes for H 2 production. The key issue to make H2 an attractivealternative fuel is to optimize its production from renewable raw materials instead of the more common energy intensive processessuch as natural gas reforming or electrolysis of water. With such vision, this paper reviews developments in microbial processes for H 2

production followed by a road map to H2 economy in Pakistan. The H2 economy potentially offers the possibility to deliver a range ofbenefits for the country; however, significant challenges exist and these are unlikely to be overcome without serious efforts.

Keywords: At least five

1. IntroductionAt the start of the 21st century, we face significant

energy challenges. The concept of sustainable

development is evolved for a livable future where human

needs are met while keeping the balance with nature.

Driving the global energy system into a sustainable path

is progressively becoming a major concern and policy

objective.

At the present, worlds energy requirement is by large

being fulfilled by fossil fuels which serve as a primary

energy source. Fossil fuel has delivered energy andconvenience, in our homes, for transport and industry.

However, the overwhelming scientific evidence is that the

unfettered use of fossil fuels is causing the worlds

climate to change, with potential disastrous effect on our

planet. The dramatic increase in the price of petroleum

are also forcing for the search for new energy sources and

alternative ways. World is in search of convenient, clean,

safe, efficient and versatile energy source as well as

energy carrier that can be delivered to the end user.

Electricity is one of the energy carriers which is already

being used worldwide. Electricity is a convenient form of

energy, which can be produced from various sources and

transported over large distances. Hydrogen is another

clean energy source as well as energy carrier. H2

economy has often been proposed by researchers as

another clean, efficient and versatile renewable energy

sources as well as energy carrier [1-3], but the

transformation from the present fossil fuel economy to a

H2 economy will need the solution of numerous complex

scientific and technological issues. The provision of costcompetitive hydrogen in sufficient quantity and quality is

the groundwork of a hydrogen energy economy. Presently

H2 is not an alternative fuel but only an energy carrier

produced from H2-rich compounds. H2 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 foreign oil and

lower the carbon and criteria emissions from the

transportation sector as depicted in Table 1.


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Developments in Hydrogen Production through Microbial Processes; Pakistans Prospective

190

Table 1.Comparison of energy and emissions of combustible

fuels [4]

Fuel type Energy

per unit

mass

(MJ/kg)

Energy

per

volume

(MJ/l)(approx.)

Kg of

carbon

release per

kg of fuelused

H2 gas 120 2 0

H2 liquid 120 8.5 0

Coal(anthracite)

1519

0.5

Coal (sub-bituminous)

2730

0.7

Natural gas 3350 9 0.46

Petrol 4043 31.5 0.86

Oil 4245 38 0.84

Diesel 42.8 35 0.9

Bio-diesel 37 33 0.5

Ethanol 21 23 0.5

Charcoal 30 0.5

Agriculturalresidue

1017 0.5

Wood 15 0.5

H2 has some unique characteristics which make it

suitable for H2 economy, namely: H2 is one of the most

plentiful elements on Earth and in the Cosmos

Combustion of molecular H2 with oxygen produces heat.

H2 has the highest energy content per unit weight of any

known fuel (142 KJ /g or 61,000 Btu/lb) H2 can be

produced from and converted into electricity at a

relatively high efficiency. The only byproduct is water,

while burning of fossil fuels generates CO2 and a varietyof pollutants. H2 may be completely renewable fuel It can

be stored as liquid, gas It can be transported over large

distances using pipelines, tankers, or rail trucks. It can be

converted into other forms of energy in more ways and

more efficiently than any other fuel, i.e., in addition to

flame combustion (like any other fuel) H2 may be

converted through catalytic combustion, electro-chemical

conversion, and hydriding.

Some vehicle manufacturers have already

demonstrated that H2 can be used directly in an internal

combustion engine, and fuel cell-powered prototype cars

have also been constructed. H2 can be transported for

domestic/industrial consumption through conventional

means.

Production of H2 from petroleum product or natural

gas does not offer any advantage over the direct use of

such fuels while Production from coal by gasification

techniques with capture and sequestration of CO2 could

be an interim solution [5]. The key issue to make H2 an

attractive alternative fuel particularly for the

transportation sector is to optimize the production process

from renewable raw materials instead of the more

common energy intensive processes Water splitting by

artificial photosynthesis, photobiological methods based

on algae, and high temperatures obtained by nuclear or

concentrated solar power plants are promising approaches

[5]. The H2 economy is an inevitable energy system of the

future where the renewable sources will be used to

generate H2 and electricity as energy carriers, which are

capable of satisfying all the energy needs of human

civilization. However nearly all H2 produced today for the

industrial sector, is largely by thermal processes with

natural gas as the H2 feedstock. Thus the development of

alternative and renewable pathways for producing H2

fuels is of utmost importance. The purpose of this paper is

to provide a brief summary of significant current and

developing biological H2 production technologies. A

vision for H2 economy in Pakistan is also discussed.

2. Industrial Applications of HydrogenApproximately 49% of hydrogen produced is used for

the manufacture of ammonia, 37% for petroleum refining,

8% for methanol production and about 6% for

miscellaneous smaller-volume uses [6]. It is also used in

the petrochemical manufacturing, glass purification,

semiconductor industry and for the hydrogenation of

unsaturated fats in vegetable oil [7]. In metallurgical

processes, hydrogen mixed with N2, is used for heat

treating applications to remove O2 as O2 scavenger. The

future widespread use of hydrogen is likely to be in thetransportation sector, where it will help reduce pollution.

Vehicles can be powered with hydrogen fuel cells, which

are three times more efficient than a gasoline-powered

engine [8, 9].

3. Current Hydrogen ProductionWorldwide, H2 is being considered as a fuel for the

future. It is an environmentally benign replacement for

gasoline, diesel, heating oil, natural gas, and other fuels in

both the transportation and non-transportation sectors.

Although abundant on earth as an element, H2 combines

readily with other elements and is almost always found as

part of some other substances, such as water, biomass and

hydrocarbons like petroleum and natural Gas. Currently

500 billion cubic meters H2 are produced annually

worldwide. Presently, 40 % H2 is produced from natural

gas, 30 % from heavy oils and naphtha, 18 % from coal,

and 4 % from electrolysis and about 1 % is produced from

biomass [8, 10] Currently, the most developed and most

used technology is the reforming of natural gas/

hydrocarbon fuels [11]. Each method of H2 production

requires a source of energy, i.e., thermal or electrolytic.


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The merits and demerits of the biomass processes are

discussed in Table 2.

Table 2. Advantages and disadvantages of different H2

production processes from biomass [7, 12]Process Advantages Disadvantages

Thermochemical

gasification

(i)Maximum

conversion can beachieved

(i)Significant gas

conditioning isrequired(ii)Removal of tar

Pyrolysis (i)Produces

carbonaceous

material along

with bio-oil,

(ii)chemicals and

minerals

(i)Chances of

catalyst deactivation

Solar gasification (i)Good H2 yield (i)Required

effective collector

platesSupercritical

conversion

(i)Can process

sewage sludge,

which is difficult

to gasify

(i)Selection of

supercritical

medium

4.Biological H2 production processesProducing H2 using conventional methods defeats the

purpose of using H2 as a clean alternative fuel. The

production of H2 from non-fossil fuel sources has

becomes central for better transition to H2 economy.

Certain microorganisms can produce enzymes that can

produce H2 provides an attractive option to produce

hydrogen through microbial process. A large number of

microbial species, including significantly different

taxonomic and physiological types, can produce H2.

Diversity in microbial physiology and metabolism means

that there are a variety of different ways in which

microorganisms can produce H2, each one with seeming

advantages, as well as problematic issues [13]. From an

engineering perspective, they all potentially offer the

advantages of lower cost catalysts (microbial cells) and

less energy intensive reactor operation (mesophilic) than

the present industrial process for making hydrogen (steamreformation of methane) [14].

The H2 metabolism of green algae was first discovered

in the early 1940s by Hans Gaffron. He observed that

green algae (under anaerobic conditions) can either use

H2 as an electron donor in the CO2-fixation process or

evolve H2 in both dark and the light [15-17]. Although the

physiological significance of H2 metabolism in algae is

still a matter of basic research, the process of

photohydrogen production by green algae is of interest

because it generates H2 gas from the most plentiful

resources, light and water [18-21].

All microbial conversions can be carried out at ambient

conditions, however lower rate of H2 production and low

yield are chief drawbacks. All processes are controlled by

the hydrogen-producing enzymes, such as hydrogenase

and nitrogenase. Hydrogenases exist in most of the

photosynthetic microorganisms and they can be classified

into two categories: (i) uptake hydrogenases and (ii)

reversible hydrogenases. Uptake hydrogenases, such as

NiFe hydrogenases and NiFeSe hydro genases, act as

important catalysts for hydrogen consumption. Reversible

hydrogenases, as indicated by its name, have the ability to

produce H2 as well as consume hydrogen depending on

the reaction condition. The major components of

nitrogenase are MoFe protein and Fe protein. Nitrogenase

has the ability to use magnesium adenosine triphosphate

(MgATP) and electrons to reduce a variety of substrates(including protons). This chemical reaction yields

hydrogen production by a nitrogenase-based system

where ADP and Pi refer to adenosine diphosphate and

inorganic phosphate, respectively

2e- + 2H+ + 4ATP ---->H2 + 4ADP + 4Pi

The processes of biological H2 production can be

broadly classified into following distinct approaches for

include: 1) Direct biophotolysis 2) Indirect biophotolysis

3) Photofermentation 4) Dark fermentation 5) Microbial

fuel cell (MFC) (bioelectrohydrogenesis )

4.1.Direct Biophotolysis

The process of photosynthetic H2-production with

electrons derived from H2O [18, 22] entails H2O-

oxidation and a light-dependent transfer of electrons to

the [Fe]-hydrogenase, leading to the synthesis of

molecular H2. The concerted action of the two

photosystems of plant-type photosynthesis to split water

with absorbed photons and generate reduced ferredoxin to

drive the reduction of protons to hydrogen, is carried out

by some green algae and some cyanobacteria as shown in

(Fig. 1). The two photosynthetic systems responsible for

photosynthesis process are: (i) photo system I (PSI) which

produces reductant for CO2 and (ii) photo system II (PSII)

which splits water to evolve O2. The two photons

obtained from the splitting of water can either reduce CO2

by PSI or form H2 in the presence of hydrogenase. In

plants, due to the lack of hydrogenase, only CO2

reduction takes place. On the contrary, green algae and

cyanobacteria (blue-green algae) contain hydrogenase and

thus have the ability to produce H2 [23]. In these

organisms, electrons are generated when PSII absorbs

light energy, which is then transferred to ferredoxin. A

reversible hydrogenase accepts electrons directly from the


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reduced ferredoxin to generate H2 in the presence of

hydrogenase.

Figure-1. Direct Biophotolysis (green algae cyanobacteria) [14]

Since hydrogenase is sensitive to oxygen, it is

necessary to maintain the oxygen content at a low level

(under 0.1 %) so that the hydrogen production can be

sustained [13]. This process results in the simultaneous

production of O2 and H2 with a H2: O2 = 2:1 ratio [24].

This mechanism holds the promise of generating

hydrogen continuously and efficiently through the solar

conversion ability of the photosynthetic apparatus. In theabsence of provision for the active removal of oxygen,

this mechanism can operate only transiently, as molecular

oxygen is a powerful inhibitor of the enzymatic reaction

and a positive suppressor of [Fe]-hydrogenase gene

expression. At present, this direct mechanism has

limitations as a tool of further research and for practical

application, mainly due to the great sensitivity of the [Fe]-

hydrogenase to O2, which is evolved upon illumination by

the water-oxidizing reactions of PSII [25]. Nevertheless,

such H2 co-production can be prolonged under conditions

designed to actively remove O2

from the reaction mixture.

Even though photosynthetic hydrogen production is a

theoretically perfect process with transforming solar

energy into hydrogen by photosynthetic bacteria, applying

it to practice is difficult due to the low utilization

efficiency of light and difficulties in designing the

reactors for hydrogen production [26-28].

4.2.PhotofermentationPhotofermentation also requires input of light energy

for hydrogen production from various substrates, in particular organic acids, by photosynthetic bacteria (Fig.2). Photosynthetic bacteria have long been studied fortheir capacity to produce hydrogen through the action oftheir nitrogenase system. Fermentative hydrogen production has the advantages of rapid hydrogen production rate and simple operation. Photosynthetic bacteria have long been studied for their capacity toproduce significant amounts of hydrogen due to their highsubstrate conversion efficiencies and ability to degrade awide range of substrates.

The photosynthetic bacteria have been shown to

produce hydrogen from various organic acids and food

processing and agricultural wastes [13]. Although pure

substrates have usually been used in model studies, some

success in using industrial wastewater as substrate has

been shown [29, 30]. In general, rates of hydrogen

production by photoheterotrophic bacteria are higher

when the cells are immobilized in or on a solid matrix,

than when the cell is free-living.

However, pre-treatment may be needed prior to

photosynthetic biohydrogen gas production due to either

the toxic nature of the effluent, or its color/ opaqueness.

Figure-2. Photofermentation (Photosynthetic bacteria) [14]

4.3.Dark fermentationIn dark fermentation, H2 production is inherently more

stable since it takes place in the absence of oxygen. The

oxidation of the substrate by bacteria generates electrons

which need to be disposed off in order to maintain the

electrical neutrality. Under the aerobic conditions O2


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serves as the electron acceptor while under the anaerobic

or anoxic conditions other compounds, such as protons,

act as the electron acceptor and are reduced to molecular

H2.

Hydrogen can be produced by anaerobic bacteria,

grown in the dark on carbohydrate-rich substrates.

While direct and indirect photolysis systems produce

pure H2, dark-fermentation processes produce a mixed

biogas containing primarily H2 and carbon dioxide (CO2),

but which may also contain lesser amounts of methane

(CH4), CO, and/or hydrogen sulfide (H2S). The gas

composition presents technical challenges with respect to

using the biogas in fuel cells. In order for hydrogen

production by dark fermentation to be economically

feasible and sustainable, a two-step/hybrid biological

hydrogen production process would be necessary.Higher overall substrate conversion efficiency is

possible by combining the anaerobic and photosynthetic

steps, as shown in Fig. 3. The photosynthetic microbes

can degrade the soluble metabolites from the fermentative

step using sunlight to overcome the energy barrier.

Dark fermentation reactions can be operated at

mesophilic (25 40C), thermophilic (4065C), extreme

thermophilic (6580C), or hyperthermophilic (80C)

temperatures.

Biohydrogen production by dark fermentation is

highly dependent on the process conditions such as

temperature, pH, mineral medium formulation, type of

organic acids produced, hydraulic residence time (HRT),

type of substrate and concentration, hydrogen partial

pressure, and reactor configuration [31].

Since organic substrates are the ultimate source of

hydrogen in photofermentations or indirect biophotolysis

processes, it can be argued that it should be simpler and

more efficient to extract the hydrogen from such

substrates using a dark fermentation process [13].

Figure 3. Dark fermentation (Clostridia, Enterobacteracae) [14]

4.4.Microbial fuel cell (MFC)It is based on the concept and practice of a microbial

fuel cell (MFC). Fact the idea is to add a little electrical

potential to that generated by a microbial fuel cell, thus

reaching a sufficient force to reduce protons to hydrogen,

in a process that can be called bioelectrohydrogenesis. A

MEC consists of four parts: first, the anodic chamber with

the anode; second, the cathodic chamber with cathode;

third, an external electrical power source; and fourth, an

electronic separator [32, 33] as shown in Fig. 4.Thus the

cell could be called a microbial electrohydrogenesis cell

(MEC). Acetate is typically used as the electron donor

and it is oxidized according to the following reaction [34]:

Acetate - + 4H2O 2HCO3- + 9H+ +9e-

The pH at the anode surface has a strong tendency to

decrease, as one proton is produced per electro transferred

[35, 36]. At the cathode the hydrogen evolution reactiontakes place, in which protons and electrons are combined

to form hydrogen:

2H2 + 2e H2

The reaction can be catalyzed by microorganisms or

by a chemical catalyst like platinum or nickel. When

microorganisms are used as catalyst these reactions are

essentially anaerobic respirations where the external

electron acceptor is an electrode instead of the more usual

oxidized compound (nitrate, TMAO, fumurate, etc.). Thus

bioelectrohydrogenesis utilizes electrochemically active

micro-organisms which, with a small to moderate voltage


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input, convert dissolved organic matter into hydrogen

inside an electrochemical cell/microbial fuel cell via

coupled anode-cathode reactions. Expressed per amount

of organic matter, the MEC can achieve much higher

hydrogen yields (80100%) [37] compared to

fermentative hydrogen production (


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(stage 1) are temporally separated from the consumption

of cellular metabolites and concomitant H2 production

(stage 2) [18, 42]. H2 evolution strongly depended upon

the duration of anaerobic incubation, deprivation of

sulphur (S) from the medium and the medium pH [43].

It has been reported that inhibition of the hydrogenase

by oxygen can be partially overcome by cultivation of

algae under sulfur deprivation for 23 days to provide

anaerobic conditions under the light [26, 44]. Melis et al.

[42] and Ghirardi et al. [25] devised a mechanism to

partially inactivate PSII activity to a point where all the

O2 evolved by photosynthesis is immediately taken up by

the respiratory activity of the culture. This mechanism is

based on a two-step process. The steps, growth mode and

H2 production mode, are initiated by cycling between

sulfur-containing and sulfur-free culture medium. Thisresults in a temporal separation of net O2- and H2-

evolution activities in the green alga Chlamydomonas

reinhardtii. This discovery eliminates the need for a purge

gas, but introduces the need for careful sulfate controls in

the aqueous medium.

The absence of sulfur nutrients from the growth

medium of algae acts as a metabolic switch, one that

selectively and reversibly inhibits photosynthetic O2

production. Thus, in the presence of S, green algae do

normal photosynthesis (H2O-oxidation, O2-evolution and

biomass accumulation) [45].

In 2002, NREL researchers developed a system using

two continuous-flow reactors for producing H2

continuously for periods of up to several weeks [46]. The

continuous H2 production process involves using two

continuously-stirred tanks. Fig.6 shows the tank

configuration. In Reactor 1, cells are cultured in media

containing minimal levels of sulfur. PS-II is slowed and

oxygen production remains lower than oxygen

consumption for cellular respiration, but by bubbling the

solutions with carbon dioxide and a small amount of

oxygen, the cells are able to remain in Reactor 1

indefinitely, obtaining some energy from photosynthesisand some energy through respiration of acetate in

solution.

Cells from Reactor 1 are transferred to Reactor 2,

which is maintained under anaerobic conditions. Cells

entering Reactor 2 already have suppressed PS-II

systems, so they will not cause Reactor 2 to go aerobic.

Any residual oxygen is quickly consumed by the algae in

Reactor 2. Finding themselves under anaerobic

conditions, the cells will start producing hydrogenase and

subsequently, H2. The transition step that consumes the

oxygen in solution in the batch system is avoided by

having Reactor 2 already anaerobic. At the same time,

some cells are continuously removed from Reactor 2. The

effect is that the cells are removed from Reactor 2 before

they completely stop producing H2. Successful operation

has been shown with a dilution rate of 0.5/day, which is

equivalent to an average residence time of 2 days for the

cells. Because Reactor 2 is a continuously-stirred reactor(like Reactor 1), the average residence time is 2 days, but

some individual cells removed from the reactor may have

been there longer or shorter times. With an average

residence time of 2 days, one would expect a H 2

production rate lower than the initial production rate of

the batch system, but higher than the production rate at

the end of a batch production cycle.

Figure 6.Continuous H2 Production

The merits and demerits of each biological process are

discussed in Table 3.


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Table 3. Comparison of important biological H2 production processes [12, 47]Process General reaction Advantages Disadvantages Maximum

reported rate

(mmol H2 /L h)

Reference

Directbiophotolysis

2 H2O + light 2 H2 + O2 -Can produce H2 directlyfrom water and sunlight-Solar conversion energyincreased by ten folds ascompared to trees, crops

-Requires high intensity of light-Simultaneous production of O2and H2. O2 can be dangerous forthe system-Hydrogenase (green algae) ishighly sensitive to evenmoderately low concentrations ofO2-Lower photochemical efficiency

0.07 [48, 49]

Indirectbiophotolysis

(a) 6H2O + 6CO2 + lightC6H12O6 + 6O2(b) C6H12O6 + 2H2O 4H2 +2CH3COOH + 2CO2(c) 2CH3COOH + 4H2O +light 8 H2 + 4CO2Overall reaction

12H2O + light 12 H2 + 6O2

-Cyanobacteria canproduce H2 from water-Has the ability to fix N2from atmosphere

-Uptake hydrogenase enzymes areto be removed to stop degradationof H2-About 30% O2 present in gasmixture

0.36 [50, 51]

Photo-fermentation

CH3COOH + 2H2O + light 4H2 + 2CO2

-A wide spectral lightenergy can be used bythese bacteria-Can use different organicwastes-High substrateconversion efficiencies-Degrade a wide range of

substrates.

-Production rate of H2is slow-O2 has an inhibitory effect onnitrogenase-Light conversion efficiency isvery low, only 15%-Pre-treatment may be needed dueto either the toxic nature of the

substrate (effluent), or itscolor/opaqueness.-Large reactor surface areasrequirement -Expensive equipment

0.16 [52]

DarkFermentation

C6H12O6 + 2H2O 2CH3COOH + 4H2 + 2CO2

-Simpler, less expensive,and produce hydrogen atmuch higher rate-It can produce H2 all day

long without light-A variety of carbonsources can be used assubstrates

-It produces valuablemetabolites such as

butyric, lactic and aceticacids as by products-It is anaerobic process,so there is no O2

limitation problem

-O2 is a strong inhibitor ofhydrogenase-Relatively lower achievable yieldsof H2-As yields increase H2fermentation becomesthermodynamically unfavorable-Product gas mixture contains CO2

which has to be separated

75.60

64.50

[53, 54]

Microbial

fuel cell(MFC)

C6H12O6 + 2H2O 4H2 +

2CO2 + 2CH3COOH

Anode: CH3COOH + 2H2O 2CO2 + 8e- + 8H+ (15)

Cathode: 8H+ + 8e- 4H2

-Energy available in

waste streams can bedirectly recovered aselectricity (MFC) orhydrogen (MEC).

promising futureapproach to hydrogen

generation fromwastewater, especially foreffluents with low organiccontent.

-Metabolic pathways involved are

not clear-MEC studies have been carriedout only with mixed cultures, oftenusing those already enriched and

active in microbial fuel cells(MFC).

-Power densities at the electrodesurface are low, which translatesinto low volumetric hydrogen

production.-Higher yields require increasedvoltage, adversely affecting energyefficiency.

Two-stage

fermentation(dark +

photo)

51.20

47.92

[15, 55]


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5. Barriers for biohydrogen productionThe diffuse nature of solar energy and the consequent

low energy density places severe economic restrictions on

potential light-driven processes for biological conversionof solar energy to hydrogen [13].

Major challenges need to be overcome for the smooth

transition from the fossil fuel based economy to the H2

energy based economy and may be outlined as follows

[56]:

The yield of H2 from any of the processes definedabove is low for commercial application.

The pathways of H2 production have not beenidentified and the reaction remains energetically

unfavorable.

There is no clear contender for a robust, industriallycapable microorganism that can be metabolicallyengineered to produce more than 4 mol H2/mol of

glucose.

Several engineering issues need to be addressed which

include the appropriate bioreactor design for H2

production, difficult to sustain steady continuous H2

production rate in the long term, scale-up, preventing

interspecies H2 transfer in non sterile conditions and

separation/purification of H2.

Sensitivity of hydrogenase to O2 and H2 partial pressure

severely disrupts the efficiency of the processes and adds

to the problems of lower yields. Insufficient knowledge

on the metabolism of H2 producing bacteria and the levels

of H2 concentration tolerance of these bacteria.

Other barriers to microbial based, large-scale

production of H2 include [57] [Maness el al 2009] (a)

inherent properties of the microbes that preclude

continuity and efficiency of H2 production; (b) underlying

limitations of photosynthetic efficiency; and (c)

limitations of the hydrogenase catalytic function.

Scientific and technical barriers for biohydrogen

production have been summarized in Table IV.

6. ImmobilizationOne of the largest challenges of optimizing molecular

H2 production by Chlamydomonas reinhardtii cells is the

transfer of the cells from sulfur deficient conditions to

sulfur rich conditions (for regenerative purposes) and then

back to sulfur deficient conditions (for further H2

production). Recent research in immobilization has provided a new technique to eliminate this challenge.

Prior to the development of immobilizations, cells were

suspended in aqueous media with either sulfur rich or

deficient conditions present. This posed a problem for

scientists because the cells had to be filtered out of the

media to be transferred to the next media in the cycle of

molecular H2 production. The filtration process was very

time consuming and so was not feasible on an industrial

scale. Another dilemma that plagued the free suspension

in liquid media technique was the inability to make the

media with cells very concentrated. This restricted the

amount of light that could interact with the cellsdecreasing the overall yield of molecular H2. To avoid

difficulties with media transition or cellular concentration

immobilization techniques were developed [58].

Table 4. Scientific and technical barriers for biohydrogen production [7]

Type of barrier Barrier Putative Solution

Basic science

Organism

Bacteria do not produce more than 4mol H2/mol glucose naturally

Isolate more novel microbes and combinationalscreen for H2 production rates yields, anddurability. Genetic manipulation of established

bacteria.

Enzyme(hydrogenase)

Hydrogenase over expression notstableO2 sensitivityH2 feed back inhibition

Greater understanding of the enzyme regulationand expression.Mutagenic studies.Low H2 partial pressure fermentation.

FermentativeFeedstock

High cost of suitable feedstock(glucose)Low yield using renewable biomass

Renewable biomass as feedstock.Co-digestion/use of microbial consortia which canincrease the yield

StrainLack of industrial-suitable strain Development of industrially viable

strain(s)/consortia


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Process

Commercially feasible product yieldIncomplete substrate utilizationSustainable processSterilization

Hybrid system (photo + dark fermentation)Link fermentation to a second process that makes

both economically possibleApplication and utilization of fermentation toolssuch as continuous culture

Development of low-cost stream sterilizationtechnology/process that can bypass sterilization

Engineering

Reactor

Lack of kinetics/appropriate reactordesign for H2 production Lightintensity in case of photo-bioreactor

Incorporation of process engineering concepts todevelop a suitable reactor for the definedstrain/process, flat panel or hollow tube reactorcan be employed

ThermodynamicThermodynamic barrier

NAD(P)H H2 (+4.62 kJ/mol)Reverse electron transport to drive H2 production

past barrier

H2 H2 purification/separation StorageSelection absorption of CO2 /H2SBasic studies on H2 storage

7. Maximum possible yield of H2 by green algaeEven though the catalytic activity of the various

enzymes differs enormously, there is no evidence for thequantity of hydrogen-producing enzyme being the

limiting factor. Indeed, in many microbial systems,

potential catalytic activity far surpasses the amount of

hydrogen produced, suggesting that other metabolic

factors are limiting [13].

The use of light attenuation devices that transfer

sunlight into the depths of a dense algal culture is an

approach to overcoming the light saturation effect in light

driven processes. The simplest approach is to arrange

photobioreactors in vertical arrays to reduce direct

sunlight. Of course, this arrangement also proportionally

increases the area of required photobioreactors, which is

the limiting economic factor in any photobiological fuel-

production process.

Another alternative is the use of optical fiber

photobioreactors, in which light energy is collected by

large concentrating mirrors and piped into small

photobioreactors with optical 1bers [13].

Application of the two-stage photosynthesis and H2

production protocol to a green alga mass culture could

provide a commercially viable method of renewable H2

generation.Table 5 provides preliminary estimates of maximum

possible yield of H2 by green algae, based on the

luminosity of the sun and the green algal photosynthesis

characteristics. Calculations were based on the integrated

luminosity of the sun during a cloudless spring day. In

mid-latitudes at springtime, this would entail delivery of

approximately 50 mol photons m 2 d 1 (Table 5). It is

generally accepted that electron transport by the two

photosystems and via the hydrogenase pathway for the

production of 1 mol H2 requires the absorption and

utilization of a minimum of 5 mol photons in the

photosynthetic apparatus (Table 5). On the basis of these

optimal assumptions, it can be calculated that green

algae could produce a maximum 10 mol (20 g) H2 per m2

culture area per day. If yields of such magnitude could be

approached in mass culture, this would constitute a viable

and profitable method of renewable H2 production.

Table 5.Yield of H2 photoproduction by green algae (Estimates are based on maximum possible daily integrated irradiance and algal

photosynthesis characteristics.) [20, 59-61]Photoproduction Characteristics Comments on Assumptions Made

Maximum photosynthetically active radiation, 50 mol photons m 2d 1 (based on a Gaussian solar intensity profile in which the peaksolar irradiance reaches 2,200 mol photons m 2 s 1)

Daily irradiance can vary significantly depending on season and cloudcover. It can be greater than 50 mol photons m 2 d 1 in the summer andmuch less than that on cloudy days and in the winter. [29].

Theoretical minimum photon requirement for H2 production in greenalgae: 5 mol photons/mol H2

Based on the requirement of 10 photons for the oxidation of two watermolecules and the release of four electrons and four protons in

photosynthesis [30, 31]

Theoretical maximum yield of H2 production by green algae: 10 molH2 m 2 d 1 (20 g H2 m 2 d 1; ~80 kg H2 acre 1 d 1)

Assuming that all incoming photosynthetically active radiation will beabsorbed by the green algae in the culture and that it will be converted intostable charge separation.


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8. Optical properties of light absorption bygreen algae

Light absorption by the photosynthetic apparatus is

essential for the generation of H2 gas. However theoptical properties of light absorption by green algae

impose a limitation in terms of solar conversion

efficiency in the algae chloroplast. This is because wild-

type green algae are equipped with a large size light-

harvesting chlorophyll antenna to absorb as much sunlight

as they can. Under direct and bright sunlight, they could

waste up to 60% of the absorbed irradiance [47, 62]. This

evolutionary trait may be good for survival of the

organism in the wild, where light is often limiting, but it

is not good for the photosynthetic productivity of a green

algal mass culture. This optical property of the cells could

further lower the productivity of a commercial H2production farm.

The analysis up to this point has shown that H2

production can be limited by the photons available or the

capacity of algae to process the photons into H2. Another

observation is that the number of photons absorbed is

much higher than the algaes ability to process the

photons. By reducing the number of excess photons

absorbed and let them reach deeper into the liquid, it may

be possible to produce more H2. By reducing the size of

the algaes light collecting antennae, but not affecting the

organisms ability to process the photons to produce H2,

one gets deeper light penetration for the same cell

concentration, which means more photons are available at

the lower depths for H2 production.

While regular green algae absorb most of the light

falling on them, algae engineered to have less chlorophyll

let some light left through. In University of California,

Berkeley, Melis and his colleagues are designing algae

that have less chlorophyll so that they absorb less sunlight

[63]. When grown in large, open bioreactors in dense

cultures, the chlorophyll-deficient algae will let sunlight

penetrate to the deeper algae layers and thereby utilize

sunlight more efficiently [64].The critical enzymatic component of this

photosynthetic reaction is the reversible hydrogenase

enzyme, which reduces protons with high potential

energy electrons to form H2. During normal

photosynthesis, algae focus on using the suns energy to

convert carbon dioxide and water into glucose, releasing

oxygen in the process. Only about 3 to 5 percent of

photosynthesis leads to H2. Because hydrogenase is

sensitive to oxygen, this H2 production must be carried

out in an anaerobic environment

Photosynthetic H2 production by green algae involves

water splitting to produce H2 and oxygen. Unfortunately,

H2 production by this process is quite ineffective since it

simultaneously produces oxygen, which inhibits the

hydrogenase enzyme. Thus, during light reaction, H2

evolution ceases due to an accumulation of oxygen.

Therefore the prerequisite for photohydrogen production

by green algae is that they have to adapt to an anaerobic

condition.

By exposing the cells to specific conditions scientists

are able to modify photosynthesis so that oxygen will not

act as the final electron carrier of the electron transport

chain; rather H2 will allow the cells to release molecular

H2 as opposed to molecular oxygen.

Melis [45] estimates that, if the entire capacity of the

photosynthesis of the algae could be directed toward H2 production, 80 kilograms of H2 could be produced

commercially per acre per day. The yield of H2

production currently achieved in the laboratory

corresponds to only 15 to 20% of the measured capacity

of the photosynthetic apparatus for electron transport

[63].

In a laboratory, Melis [45] worked with low-density

cultures and have thin bottles so that light penetrates from

all sides. Because of this, the cells use all the light falling

on them. But in a commercial bioreactor, where dense

algae cultures would be spread out in open ponds under

the sun, the top layers of algae absorb all the sunlight but

can only use a fraction of it [63].

Further research and development aimed at increasing

rates of synthesis and final l yields of H2 are essential.

Optimization of bioreactor designs, rapid removal and

puri6cation of gases, and genetic modifcation of enzyme

pathways that compete with hydrogen producing enzyme

systems offer exciting prospects for biohydrogen systems

[48]. Increase in the rate of H2 would reduce bioreactor

size dramatically to overcome the engineering challenges

of scale up, and create new opportunities for practical

applications.9. H2 Economy

A typical energy chain for sustainable H2 comprises

the harvesting of sunlight into H2 as energy carrier, the

storage and distribution of this energy carrier to the end-

device where it is converted to power. The key market for

fuel cells has always assumed to be the automotive

industry. The great expectation that hydrogen fuel-cell

powered vehicles will displace gasoline and diesel

powered vehicles has not materialized for a variety of

reasons, but primarily because fuel cell technology has


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not yet matured and the infrastructure required for

hydrogen storage, transportation, and refueling has been

slow to develop. Consumer energy applications will

require delivery systems that can supply H2 as readily as

gasoline and natural gas are supplied today. Higher-

pressure gaseous storage and non-conventional storage

technologies will be used to meet the requirements of

transportation applications (storage at 350700 bar

compared to the 200 bar storage pressure commonly used

in normal merchant gas systems) [65].

Gas purity requirements are important for the H2

energy market. They very much depend on the energy

conversion device used, as well as on the storage

technology. Combustion systems are much less sensitive

to impurity levels, however, fuel cells are very sensitive

to CO and sulfur poisoning.The U.S. Department of Energy has developed a

multiyear plan with aggressive milestones and targets for

the development of H2 infrastructure, fuel cells, and

storage technologies. The targeted H2 cost is $24 kg-1

(energy equivalent of 1 gallon of gasoline) delivered [66,

67]

A rollout of such a sustainable H2 chain in developed

countries could go either gradually via a H2 economy

based on fossil fuels or discontinuously in the case of

inventions of disruptive technologies. For developing

countries the situation may be different. Introduction of

such H2 chains for their fast-growing primary energy

demands might enable them to skip the stage of

conventional, fossil fuel-based technologies and markets

and leapfrog directly to a sustainable H2 economy [68].

The salient features of a H2 economy will be as follows

[69]:

A H2-based energy system will increase the

opportunity to use renewable energy in the transport

sector. This will increase the diversity of energy sources

and reduce overall greenhouse gas emissions. H2 in the

transport sector can reduce local pollution, which is a

high priority in many large cities.The robustness and flexibility of the energy system

will be increased by the introduction of H2 as a strong

new energy carrier that can interconnect different parts of

the energy system. The targets for reducing vehicle noise

may be met by replacing conventional engines with H2-

powered fuel cells. Fuel cells for battery replacement and

backup power systems are niche markets in which price

and efficiency are relatively unimportant. Sales in this

market will drive the technology forward towards the

point at which fuel cells will become economic for the

introduction into the energy sector. H2 electrolysers/fuel

cells connected directly to wind turbines are a convenient

way to balance out local fluctuations in the availability of

wind power. The development of fuel cells and a H2

economy will provide new market opportunities and new

jobs. Present knowledge indicates that H2 as an energy

carrier will involve little environmental risk. All

renewable hydrogen production technologies face the

common challenge of integration with hydrogen

purification and storage [65].

10. Present energy scenario of PakistanPakistan is basically an energy deficient country.

Pakistans per capita energy consumption, 3894kWh as

against the world average of 17620kWh, gives it a

ranking of 100 amongst the nations of the world [70]. The

demand for primary energy in Pakistan has increasedconsiderably over the last few decades and the country is

facing serious energy shortage problems. The energy

supply is not increasing by any means to cope with the

rising energy demands. As a result the gap between the

energy demand and supply is growing every year. The

country is meeting about 86% of oil demand from imports

by spending around US$6.65 billion per annum [71].

Pakistans future energy system looks rather

uncertain. In recent years, the combination of rising oil

consumption and flat oil production in Pakistan has led to

rising oil imports from Middle East exporters. The

balance recoverable reserves of crude oil in the country ason January 1st 2010 have been estimated at 303.63

million barrels [72].

Natural gas accounts for the largest share of

Pakistans energy use, amounting to nearly 43.7 percent

of total energy consumption. As on January 1, 2010, the

balance recoverable natural gas reserves have been

estimated at 28.33 trillion cubic feet. The average

production of natural gas during July- March 2009-10

was 4,048.76 million cubic feet per day (mmcfd) [72]. As

the demand of natural gas exceeds the supply, country is

already facing shortage of natural gas and during the peak

demand most of the gas fired generating units are

shutdown while duel fuel units are fired by oil. Pakistan is

presently facing shortage of around 300-350 MMCDF of

natural gas which is likely to go up because of rising

needs and slowing down of supplies at home [73].

According to The Energy Security Action Plan of the

Planning Commission, Pakistan will be facing a shortfall

in gas supplies rising from 1.4 Billion Cubic Feet (BCF)

per day in 2012 to 2.7 BCF in 2015 and escalating to 10.3

BCF per day by the year 2025 [74]. It is therefore a matter


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of economic security to develop alternative H2 resources

to avoid mid century energy crises in the country.

Natural gas is used in general industry to prepare

consumer items, produce cement, fertilizer and generate

electricity. At present, the power sector is the largest user

of gas accounting for 33.5 percent share followed by the

industrial sector (23.8 percent), household (18.1 percent),

fertilizer (15.6 percent), transport (5.4 percent) and

cement (0.9 percent) [75]. Natural gas is used in the

transport sector in the form of CNG. There are about

3,116 established CNG stations in the country and

approximately 2 million vehicles are using CNG. Pakistan

has become the largest CNG consuming country among

Natural Gas Vehicle (NGV) countries. According to

Petroleum Policy 1997; the use of CNG in vehicles was

encouraged by Government to reduce pressure onpetroleum imports, to curb pollution and to improve the

environment [75].

Transport sector is one of the major consumers of

commercial energy in Pakistan. It accounted for about

28% of the total final commercial energy consumed

(33.95 MTOE) and 55.8% of the total petroleum products

consumed (15 MTOE) in the country.

11.H2 Production in PakistanIn Pakistan H2 is largely produced in the fertilizer

industry from natural gas, which is used for the

production of anhydrous ammonia. All urea plants in thecountry are based on natural gas as feedstock. On an

average, the fertilizer sector consumes 15.6 per cent of

natural gas produced in country. The government

provides an indirect subsidy to fertilizer manufacturers by

selling feedstock gas at rates ranging up to $1.0 against

commercial rates of $4.0 per MMBTU. The return on

paid-up capital in the fertilizer industry is about 80-100

per cent per annum [73]. The current energy scenario in

the country, already discussed above , identifies the

transport sector and fertilizer sector as key sectors where

the H2 gas can be immediately employed as substitute to

fossil fuel.

Mirza et al. [76] has presented complete road map to

H2 economy in Pakistan. They have concluded that the H2

economy potentially offers the possibility to deliver a

range of benefits for the country including reducing

dependence on oil imports, environmental sustainability

and economic competitiveness. In medium term advent of

H2 will bring about technological developments in many

fields, including power generation, agriculture, the

automotive industry, and other as yet unforeseen

applications. It will increase employment, stimulate the

economy, and will have a positive impact on the

environment in which atmospheric pollution is all but

alleviated and the so-called greenhouse effect is

mitigated.

To ensure a sustainable energy future for Pakistan, it

is necessary that the energy sector be accorded a high

priority. In Pakistan efforts to reduce reliance on fossil

fuels through increasing the share of renewable energy in

the energy supply systems have met with little success so

far. Mirza et al. [77] and Sahir and Qureshi [78] have

discussed the barriers to development of renewable

energy. Mirza, et al. [77] has broadly classified these

barriers as policy and regulatory barriers, institutional

barriers, fiscal and financial barriers, market-related

barriers, technological barriers and information and social

barriers. They have also suggested better coordinationamong various stakeholders and indigenization of

renewable energy technologies to overcome these

barriers.

Sahir, and Qureshi [78] has suggested an integrated

energy planning approach, consistency in government

policies and rational policy instruments to deal with the

techno-economic and socio-political barriers are the pre-

requisites for long-term sustainable development of the

renewable energy technologies.

There is little doubt that power production by

renewable energies, energy storage by H2, and electric

power transportation and distribution by smart electric

grids will play an essential role in phasing out fossil fuels.

12.ConclusionsConcerns about global warming and environmental

pollution due to the use of fossil fuels, combined with

projections of potential fossil fuel shortfall toward the

middle of the 21st century, make it imperative to develop

alternative energy sources that would clean, renewable,

and environmentally friendly.

It is important to note that hydrogen can be produced

from a wide variety of feed stocks available almost

anywhere. There are many processes under development

which will have a minimal environmental impact.

Development of these technologies may decrease the

worlds dependence on fuels that come primarily from

unstable regions. The in house hydrogen production

may increase both national energy and economic security.

The ability of hydrogen to be produced from a wide

variety of feedstocks and using a wide variety of

processes makes it so that every region of the world may

be able to produce much of their own energy. It is clear


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that as the technologies develop and mature, hydrogen

may prove to be the most ubiquitous fuel available.

The vision for a H2 future is one based on clean

sustainable renewable energy supply of global

proportions that plays a key role in all sectors of the

economy.

Microbial Processes provides an attractive option to

produce H2 at ambient conditions. A large number of

microbial species, including significantly different

taxonomic and physiological types, can produce H2,

Diversity in microbial physiology and metabolism means

that there are a variety of different ways in which

microorganisms can produce H2, each one with seeming

advantages, as well as problematic issues. Lower rate of

H2 production and low yield are chief drawbacks. From

an engineering perspective, they all potentially offer theadvantages of lower cost catalysts (microbial cells) and

less energy intensive reactor operation (mesophilic) than

the present industrial process for making hydrogen.

The recently developed single-organism, two-stage

photosynthesis and H2 production protocol with green

algae is of interest because significant amounts of H2 gas

can be generated essentially from sunlight and water.

Further, this method does not entail the generation of any

undesirable, harmful, or polluting byproducts and it may

even offer the advantage of value-added products as a

result of the mass cultivation of green algae.

However, several biological and engineering

challenges must be overcome before this promising

technology becomes a practical reality. Foremost, the

cellular metabolism and basic biochemistry that support

this process must be well understood and much

fundamental research on the mechanism of H2 production

by S- deprivation remains to be done.

For Pakistan the indigenous H2 production may

increase both national energy and economic security. The

ability of H2 to be produced from a wide variety of

feedstocks and using a wide variety of processes makes it

so Pakistan may be able to produce much of her ownenergy. Fertilizer sector is key area where the H2 gas can

be immediately employed as substitute to natural gas.

The advent of H2 will bring about technological

developments in many fields, including power generation,

agriculture, the automotive industry, and other as yet

unforeseen applications. It will increase employment,

stimulate the economy of all nations on earth, and will

have a positive impact on the environment in which

atmospheric pollution is all but alleviated and the so-

called greenhouse effect is mitigated.

Non-incorporation of renewable energy issues in the

regulatory policy and lack of awareness among regulators

restrict technology penetration. There is a lack of

financial resources and proper lending facilities,

particularly for small-scale projects in country. In

addition, the absence of a central body for overall

coordination of energy sector activities results in

duplication of R&D activities. Unfortunately private

sector especially transports and fertilizer sector has made

no contributions to promote research activities to produce

H2 from renewable resources.

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