Microalgae as a Feedstock for Biofuel Production: Current Status and Future Prospects

Abstract

The wide use of fossil fuels is gradually documented as unsus-tainable due to the significant reduction of supplies and emission of greenhouse gases (GHGs) into the atmosphere. Alternative and renewable energy sources are required to fulfill the ever increasing energy demands. Microalgae are the alternative renewable energy source as it has the potential to generate high amount of biomass which can be used for the production of third-generation biofuels. Moreover, utilization of biofuels also helps to preserve the atmosphere by many means like wastewater treatment, reduction in greenhouse gas emissions.

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Top 5 Contributions in Energy Research and Development: 3rd Edition
Chapter 02
Microalgae as a Feedstock for Biofuel
Production: Current Status and Future
Prospects
Muhammad Rizwan Javed1*, Muhammad Junaid Bilal1, Muhammad
Umer Farooq Ashraf1, Aamir Waqar2, Muhammad Aamer Mehmood1,
Maida Saeed3 and Naima Nashat3
1Department of Bioinformatics and Biotechnology, Government College
University Faisalabad (GCUF), Pakistan
2Department of Biotechnology, University of Sargodha (UOS), Pakistan
3Department of Biochemistry, University of Agriculture Faisalabad (UAF),
Pakistan
*Corresponding Author: Dr. Muhammad Rizwan Javed, Assistant Profes-
sor, Department of Bioinformatics and Biotechnology, Government Col-
lege University Faisalabad (GCUF), Allama Iqbal Road, Faisalabad, 38000,
Pakistan, Email: rizwan@gcuf.edu.pk
First Published November 29, 2019
Copyright: © 2019 Muhammad Rizwan Javed, et al.
This article is distributed under the terms of the Creative Commons
Attribution 4.0 International License
(http://creativecommons.org/licenses/by/4.0/), which permits
unrestricted use, distribution, and reproduction in any medium,
provided you give appropriate credit to the original author(s) and
the source.

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Abstract
e wide use of fossil fuels is gradually documented as unsus-
tainable due to the signicant reduction of supplies and emission
of greenhouse gases (GHGs) into the atmosphere. Alternative and
renewable energy sources are required to fulll the ever increasing
energy demands. Microalgae are the alternative renewable energy
source as it has the potential to generate high amount of biomass
which can be used for the production of third-generation biofuels.
Moreover, utilization of biofuels also helps to preserve the atmosphere
by many means like wastewater treatment, reduction in greenhouse
gas emissions.
Background
For many years, fossil fuels (a fuel formed by natural processes,
such as anaerobic decomposition of buried dead organisms, contain-
ing energy and originating in ancient times) have become the most
important part of human life. Fossil fuels contain high percentages
of carbon and include petroleum, coal, and natural gas. Our daily life
is now totally dependent on fossil fuels, from transportation to en-
ergy production, for electricity, and for running factories. Fossil fuels
are present in a limited amount on earth and they are non-renew-
able sources that will be eliminated one day [1]. Due to growth of
human population and industrialization in every part of world, the
whole world is facing the fuel crisis [2]. Other than that, the world is
also facing many environmental issues because of fossil fuel usage as
energy source. Fossil fuels release high amount of CO2 in the atmos-
phere thus contributing considerably to greenhouse eects and global
warming [3]. Such concerns necessities the need for alternative and
renewable energy sources.
In this century, renewable energy sources are a major challenge
for the world. Energy sources based on wind, hydro, geothermal, so-

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lar and biomass wastage have been successfully developed and used
by dierent nations to limit the use of fossil fuels. However, all these
sources have their limits and provide only limited energy, so we have
to search for alternative renewable and vital energy sources [1]. In this
context biofuels produced by the organic producers, could be used as
energy resource since they are environmentally friendly and renew-
able so could play a vital role in this challenging situation [4].
Biofuel: A Renewable Energy Source
Renewable energy sources are those resources that could be used
to produce energy again and again like wind energy, solar energy, geo-
thermal energy, biomass energy [5]. Biomass energy technologies use
plants or waste materials for energy production with low greenhouse
eect than fossil fuels [6]. Agricultural products of starchy and ce-
real crops like corn, wheat, sugarcane, sorghum, and beets are used
to produce bioethanol. Oil- or tree-seeds of sunower, palm, soya,
coconut, rapeseed or jatropha are also used for production of bioetha-
nol. Now-a-days, bioethanol is the mostly used biofuel globally for
transport. Almost 60 percent of worldwide bioethanol yield is ob-
tained from sugarcane and 40 percent from other crops [7]. Brazil is
leading world producer with 15 billion liters distilled from sugarcane,
equivalent to 38 percent of global production. e United States is
the 2nd largest producer & consumer, accounting for 32 percent of
world’s bioethanol production (in 2004). Bioethanol production was
started from corn in the 1970s, but its use is increasing currently [8].
Biodiesel production was started in 1990s and since then production
has been increasing. Worldwide biodiesel production reached a re-
cord 1.8 billion liters in 2003. Compared to bioethanol, however, total
biodiesel production is fairly small. e European Union is the main
producer of biodiesel, which accounts for about 95 percent of world-
wide production. About 80-85 percent of European Union’s produc-
tion comes from rapeseed oil, which is equal to 20 percent of the total
European Union’s rapeseed production [9]. Worldwide production of
bioethanol in 2011 was almost 10 billion liters and in 2020 it will in-
crease up to 281.5 billion liters [10]. e energy demand in develop-

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ing countries is estimated to rise upto 84%, and about 1/9th of this fuel
is typically acquired from other renewable sources like biofuels [11].
Biofuels play role as the major source of energy for more than half of
the total population globally, that accounts upto 90% of consumption
of energy in less developed nations [12].
Types of Biofuels
Mainly two types of biofuels are used; the rst one is biodiesel
and second one is bioethanol, that are obtained from seeds, lignocel-
lulose, and vegetable oils. Worldwide biofuel production is expected
to be over 35 billion liters [12]. Biodiesel is a substitute for petroleum
diesel, that is renewable, non-toxic and biodegradable that can be
used with or as an alternate of fuel [13]. Biodiesel is obtained from
vegetable oil but concern is that biodiesel production as feedstock
from vegetable oil may compete with food supply. at’s why recently
scientists have focused to nd bearing plants that produce non-edible
oil for biodiesel production [14].
Bioethanol is obtained from lignocellulosic biomass such as
trees, grass and waste materials [15]. Bioethanol production from lig-
nocellulosic biomass comprises the following main steps; (1) Hydrol-
ysis of cellulose and hemicellulose, (2) Sugar fermentation, (3) Sepa-
ration of lignin and (4) Recovery and purifying the bioethanol [16].
Production of bioethanol from sowood is also important, which is
a dominant source of lignocellulosic material in the Northern hemi-
sphere. Sowood such as pine and spruce contains around 43 to 45%
cellulose, 28% lignin, 20 to 23% hemicellulose. Hemicellulose is made
up of mannose (a hexose) that can be fermented by normal baker’s
yeast. Around 410 liters of bioethanol is produced using only hexose
fraction [17].
A third important type of biofuel is biogas. Production of biogas
from wastes, residues and energy crops will play vital role in future.
Europe policy estimates that 25% of all bioenergy can be derived from
biogas [18]. It is also a renewable energy source that can be the re-
placement of fossil fuels in power and heat production. Biogas pro-

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duction through anaerobic digestion oers signicant benets over
the other forms of bioenergy production [19]. One factor that aects
the process of biogas production is temperature regime; ermo-
philic anaerobic digestion at 55-70 ºC has a rate-advantage over MPD
(mesophilic digestion). However, anaerobic digestion performing
microorganisms are too much sensitive to change in the temperature
that aects the biogas production. Another factor that directly aects
the anaerobic digestion process and products is pH. An ideal range
of pH for anaerobic digestion is 6.8 to 7.4. C/N ratio that shows the
nutrient levels of anaerobic digestion substrate is also very important
for anaerobic digestion. A high C/N ratio leads to lower production
process of biogas [18]. For the production of Bio-hydrogen gas from
renewable sources, also known as ‘’green technology’’, the major fac-
tors that should be considered for the selection of waste materials to
be used are cost, availability, carbohydrate content, and biodegrada-
bility. Major bioprocesses that are employed for the production of bio-
hydrogen are; Bio-photolysis of water by algae, Dark-fermentative
hydrogen production during acidogenic phase of anaerobic digestion
of organic matter and Two stage dark/photo fermentative production
of hydrogen [20,21].
Classication of Biofuels Based on Generations
On the basis of feedstock being used for biofuel production, the
following generations can be categorized (Figure 1):
First Generation Biofuels
Bioethanol and biodiesel are classied among the rst genera-
tion biofuels which are produced from biomass that is mostly edible
using transesterication or yeast fermentation [22].
Bioethanol
Bioethanol is produced generally by fermentation of glucose
(C6) using genetically modied organisms (GMO) such as Saccha-
romyces cerevisiae. Corn is considered as an important source for the
production of bioethanol [23]. Corn which is suitable for the produc-

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tion of ethanol requires cropland and it raises thoughtful ethical and
economical issues like the cost e.g. ethanol production from corn and
molasses in several countries is about $0.75 and $0.74 per liter [24].
Sugarcane is used as an alternative feedstock for the production of
biofuels. It is also noted that greenhouse gases are emitted during the
production of ethanol from sugarcane. Total GHG emission of 436
kg CO2 eq m-3 ethanol was observed for anhydrous ethanol produc-
tion during 2005/2006 that is thought to be decreased to 345 kg CO2
eq m-3 in 2020 [25]. Ethanol is also produced using lignocellulose as
a feedstock. e lignocellulose is composed of lignin, cellulose, and
hemicellulose. e cost of ethanol production from starch crops and
sugars like sugarcane is very high as compared to lignocellulose [17].
Metabolically engineered microorganisms such as Saccharomyces
cerevisiae are deployed to convert lignocellulosic sugars into ethanol.
Due to recalcitrant nature of lignocellulose, it requires hydrolysis pri-
or to its fermentation [26].
Biodiesel
Biodiesel is prepared by the trans-esterication of lipids with al-
cohols having lower molecular weight. Biodiesel can be synthesized
by using waste greases, vegetable oils and animal fat [27]. Biodiesel
has some benets which are as follows: No discharge of sulphur dur-
ing the process of combustion; polycyclic aromatic hydrocarbons do
not produce while combustion is done; CO2 which is biomass-derived
can be recycled during the emission of fuel [28]. A process was devel-
oped by using waste cooking oil to reduce the cost of biodiesel. is
process has some characteristic steps like trans-esterication, settling,
glycerin washing, methanol recovery, hexane extraction, etc [29].
Second Generation Biofuels
Second-generation biofuels, also known as advanced biofuels,
are fuels that can be manufactured from various types of non-food
biomass. e biomass used for the production of second-generation
biofuels is categorized into three main groups: Non-homogeneous
feed stock that include low value municipal solid waste; Homogene-

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ous feed stock that include white wood chips which have price value
between US$100 to US$120/t; and Quasi-homogeneous feed stock
that include forest and agriculture residues and have price value of
US$60 and US$80/t [30]. Cellulosic ethanol and biomass-to-liquids
conversion using the Fischer-Tropsch process are examples of 2nd
generation biofuels [22]. e process of conversion of biomass to
second generation biofuels is done either by following the biologi-
cal or thermo-chemical pathways. Gasication is included in the
thermo-conversion in which the conversion of carbonaceous to gas-
eous products through a process of partial oxidation is carried out.
In gasication of biomass, the lignocellulosic structure of biomass is
broken down using temperature into carbon dioxide, carbon monox-
ide, and hydrogen which are the main components but some amount
of methane is also produced along with dierent types of gases. e
second method is biological conversion in which organisms that are
hydrogenogenic carboxydotrophs (e.g. Calderihabitans maritimus,
Carboxydocella thermautotrophica) are used to produce hydrogen by
the oxidation of CO and energy for this method is provided by the
transfer of the electron from CO to water [31].
Lignocellulosic biomass, particularly agricultural residues,
is non-food related and abundantly available without geographi-
cal limitation. e consumption of lignocellulosic biomass at large
scales does not compete for food with people, directly or indirectly
through diverting the use of arable land as that might occur in fuel
ethanol production from sugar and grains, making it a sustainable
feedstock for producing second generation fuel. Major components
in lignocellulosic biomass are cellulose, hemicelluloses, and lignin,
which are entangled together to form lignin-carbohydrate complexes
(LCCs). LCCs are recalcitrant to degradation, and thus pretreatment
is needed for their destruction to separate the cellulose component
from hemicelluloses and lignin so that the enzymatic hydrolysis can
be performed to release glucose for ethanol fermentation [32].

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It is thought that the emission of carbon and greenhouse gas-
es is reduced by using second generation biofuels while eciency
of energy is increased [33]. However, there are some drawbacks of
second-generation biofuels such as that they cannot be produced on
large scale at commercial level due to high cost and depending on raw
material there is heterogeneity in the production process [34].
Third Generation Biofuels
Among third generation biofuels; aquatic biomasses such as sea-
weed, caltrop diatoms, hyacinth, duckweed, salvinia. kelp and algae
are used as renewable sources for biofuel production [31]. e fuel
that is produced from seaweed is considered as a third-generation
biofuel. In seaweeds, lignin is not present, and level of lipid content
& cellulose is low while high solid content is there [35,36]. e third
generation of biofuel is an advanced biofuel due to advanced pro-
cesses and feedstocks used for its production [37]. e most accu-
rate statement about third generation biofuels is that they would be
obtained from the algal biomass that has the most admirable growth
and yield rates (algae could give 1-7 g/L production of biomass at op-
timum conditions) as compared to other organisms and crops. How-
ever, some geographical and technical issues related to algal biomass
are also present [3].
Fourth Generation Biofuels
Fourth generation biofuels (FBGs) are important due to the
synthetic biology of cyanobacteria and algae [38-40]. FBGs will be
based on raw materials that are low cost and easily available. Both
third generation biofuels and FGBs are synthesized by photosynthetic
microorganisms like microalgae, cyanobacteria, yeast, and fungi to
synthesize renewable fuels [41]. CO2 is converted into energy by using
these microorganisms [42]. FGBs are mainly based on metabolically
engineered algae. It uses recombinant, bioengineering and biological
techniques for modications of properties by introducing or modify-
ing algal metabolic pathways to increase biofuel production. FBGs in-

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volve algal biodiesel, general algae gene transformation, high carbon
alcohols, algal ethanol and gaseous biofuels [43].
Figure 1: A schematic representation of the types and generation of biofuels.
Classication of Algae
Algae are divided into two categories based on cell size and com-
position [44]. (a) Macroalgae (b) Microalgae (Figure 2).
Macroalgae
Macroalgae (seaweed) is the sub-class of algae, having multicel-
lular organization and have developed anatomical arrangements that
resemble stems, roots, and leaves of higher plants. Macroalgae such
as seaweeds possess nuclei and organelles which grow to large in size,
their dierent nature has made them abundant on earth [44]. Lami-
naria, Saccorhiza, Alaria are common macroalgal genera which be-
long to brown algal group. Laminarin and manitol are main reserved
food materials [45]. Macroalgae is usually cultivated following three
dimensional approaches such as in seas and aquaculture, not in two

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dimensions such as on the surface of the earth [46]. Macroalgae pro-
duces 3.1 million dry metric tons per year in the world by aquacul-
ture [47]. For the utilization of macroalgae various thermochemical
options include liquefaction, pyrolysis, gasication, and combustion.
ese conversion processes of macroalgae have attained very low at-
tention [48]. e macroalgal biomass production converted to carbon
ranges from 1 to 3.4 kg carbon m-2 per year [49]. Communities of
seaweeds of North Atlantic coasts have annual production of approxi-
mately 2 kg carbon m-2 that is higher than production of grasslands
(less than 1 kg carbon m-2 per year) or plantation of temperate trees
[50]. Macroalgae can be cultured in open seas [51]. Seaweed farming
does not rely upon the fresh water and land areas are not occupied
[52]. e lower body parts of macroalgae are more important since
they do not possess well recognized leaves, stems and roots. ey
can grow faster, and their size can reach up to 10 meters in length.
Approximately, 10,000 species of seaweed exist in nature, which are
used worldwide, while only few are completely cultivated like Undaria
pinnatida [53]. In green macroalgae chlorophyll b and chlorophyll a
are the main pigments. Green macroalgae also produce starch which
is photosynthetic products. Cellulose and pectin are involved in the
formation of cell walls of green macroalgae [54]. Manitol and lamina-
rin are the food materials that are used for the production of ethanol
[55]. High amounts of carbohydrates are also accumulated by some
species of macroalgae that are used in microbial conversion process
as substrate.
Macroalgae like Gelidium amansii, Gracilaria spp. and Laminar-
ia spp. are candidates for the production of bioethanol [54]. Gelidium
amansii is a red alga that is made up of galactan, cellulose, and glucan
that can also be used as feed stock for bioconversion to ethanol [56].
Bioethanol production from terrestrial plant leaves has a large impact
on environment and human beings due to ecotoxicity, eutrophication,
and acidication [57]. Conversion of macroalgal biomass to ethanol
instead of using biomass of terrestrial plants has some benets like
there is no negative pressure on food security. e sugar contents are

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high and lignin contents are low then lignocelluloses that promote
high mass production [46]. Oxygen is supplied by macroalgae to the
sea and helps to decrease the CO2 accumulation in the atmosphere
[58]. Dierent kinds of supplementary products are produced by us-
ing algal biomass such as plastics and pigments [56]. Heavy metals
are removed by using many algal species which can be benecial to
the environment [54]. e bioethanol feed stock is obtained by using
macroalgae which have high biomass yield with better production in
comparison to dierent terrestrial crops [59].
Microalgae
Microalgae (microphytes) are single celled organisms that are
able to convert radiant energy emitted by the sun into chemical en-
ergy through photosynthesis. Microalgae contain bioactive com-
pounds and used for the production of many valuable compounds
such as carotenoids, antioxidants, fatty acids, enzymes, polymers,
peptides, toxins, and sterols. Microalgae were used by Chinese scien-
tists about 2000 years ago [60]. Microalgae have evolved from large
light-harvesting complexes to achieve maximum light absorption
in low light in which microalgae live [61]. Microalgae cultivation is
only a few decades old [62]. In early 1950s due to increase in world
population and insucient supply of protein, search for new and al-
ternative protein sources was initiated. e high content of protein
in dierent microalgae species is main reason to choose them as un-
conventional sources of proteins [63]. Microalgae biotechnology has
been recently developed for dierent purposes and commercial ap-
plications. Microalgae also contain chlorophyll which can be used as
food and in cosmetics [62]. Some species of microalgae are capable
to absorb heavy metal and x nitrogen and phosphorus [64]. Many
valuable compounds form microalgae like fatty acids (arachidonic)
[65], pigments (carotenoids), biochemically stable isotopes [66] and
vitamins C, E [67], etc have been extracted. In aquaculture, microal-
gae are live food of marine sh and lter feeding invertebrates such as
rotifer [68]. Microalgae biomass has almost y percent of carbon on

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a dry weight basis [69]. ey have high rate of growth with duplicat-
ing their cells many times in a single day [70] such as Nannochloropsis
oculata, Phaeodactylum tricornutum, and Chlorella vulgaris.
Figure 2: A broad classication of the algae [71].
Potential of Microalgae as a Feedstock
As the demand for fuel resources increases by the growth of
population and global economy, new alternative resources are needed
[4]. e idea about using the microalgae as a feedstock for biofuel
production is not new, but now it is discussed seriously because of
reduction in fossil fuel sources and the harms of global warming as-
sociated with them [66]. Algae has become a viable alternate energy
resource as it employs photosynthesis to take CO2 and change it into
carbon source for biofuel. Now biofuel is produced by CO2, water, and
sunlight which are all renewable sources [4]. Algae have cells which
convert CO2 into biofuels, foods and bioactive compounds [66]. Al-
gae accumulate lipids (Table 1) that make algae a good feedstock for
biofuels such as for bioethanol and biodiesel production.

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Table 1: Lipid content of various microalgal strains.
Under optimum growth conditions, algae grow relatively fast
than other species and, in few hours, they double its number [72].
ey grow at very high rate about 100 times faster than other plants
and their biomass is doubled in less than a day [73]. Algal harvesting
time is very short (less than 10 days) and could be done continuously.
ey are not like conventional crops which can only be harvested one
or two times a year. Most algae need tropical environmental condi-
tions, however some algal species like Chlorella sp. that is isolated
form a block of arctic sea ice could grow at 4 to 32 ºC [72]. Algae
and some cyanobacteria like Spirulina platensis could grow in alkaline
environments [74]. Microalgae could provide many types of biofuels
including methane by digestion of algae anaerobically, biodiesel from
microalgal oil and gas produced during photosynthesis [66].
e potential advantages of using microalgae as a feedstock for
biofuels are followings [3,75]:
Microalgal Strain Algae Type Lipid Content
(%)
References
Chlamydomonas
biconvexa
Green 1.26-1.58 Santana et al., 2017
[129]
Scenedesmus sp. Green 12.50-13.40 Diniz et al., 2017 [120]
Chlorella vulgaris Green 21.69-28.07 Peng et al., 2016 [127]
Spirogyra orientalis Green 21 D’Alessandro and Filho,
2016 [118]
Botryococcus terribilis Green 18.5-40 Cabanelas et al., 2015
[116]
Chlorella sorokiniana Green 19 Li et al., 2015 [124]
Skeletonema costatum Diatoms 17.4 Maity et al., 2014 [125]
Botryococcus brauniiGreen 9.55-26 Cabanelas et al., 2013
[115]
Auxenochlorella proto-
thecoides
Green 39.3 De la Hoz Siegler et al.,
2012 [119]

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• Oil produced by algae does not have adverse eects on other
agriculture products because it does not compete with other
food crops for land.
• Algae can be grown in almost any environment, like sea-
water, etc. Algae grown in the aqueous environment need
much less quantity of water than other crops, so they reduce
the freshwater source load.
• Microalgae can also be harvested in saline water, therefore,
it does not experience land use changes.
• Algal growth can be helpful in the treatment of wastewater
as they remove phosphorous and carbon dioxide.
• Algae are also helpful to produce a broad range of biofuels
such as hydrogen, syngas, and methane.
• Microalgae could be grown round all the year, so the pro-
duction of oil exceeds the yield of the oilseed crops.
• Microalgae have an exponential growth rate. ey have oil
content ranging from 20-50% biomass (dry weight) and
they double their biomass in very short time (about in 3.5
hours).
• Algal cultivation does not need the application of any pesti-
cides or herbicides.
• Microalgae could also produce many useful co-products
like residual biomass from oil extraction and proteins which
are used as fertilizers and feed. Additionally, the CO2 xa-
tion, treatment of wastewater, bio-hydrogen production and
biofuel production are the potentials of microalgae to be
used as a feedstock.
Characteristics of Microalgae for Biofuel
Production
Microalgae accrue carbohydrates and lipid compounds which
make them the most favorable feedstock for biofuel production such

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as bioethanol and biodiesel. ey can be grown in every environment
and all year-around, which make them suitable for use as feedstock
[4]. Microalgae contain fatty acids and lipids as membrane compo-
nents, metabolites, sources of energy and storage products that make
microalgae a viable possibility as a feedstock for biofuel [76]. Micro-
algae use photosynthesis to convert the light into chemical energy and
as they can grow in every environment that is not possible for other
currently used feedstocks of biodiesel such as rapeseed, soybean, and
palm oil. e microalgal growth rate is higher than agricultural crops,
conventional forestry, and other aquatic plants. ey need less area
than other feedstocks, 49 to 132 times less than soybean crops [66].
Microalgae can play important role as feedstock for many dierent
kinds of renewable biofuels (Table 2) such as methane, hydrogen, bio-
diesel, and bioethanol. Microalgal biodiesel does not contain sulfur
and performs truly as petroleum fuel [77]. e most signicant char-
acter of microalgal oil is its biodiesel yield, according to an estimate
the yield of biodiesel from algae per acre is over 200 times more than
the yield of other crops and plants. If algal biodiesel is produced com-
mercially it would reduce the cost of oil per barrel from 100$ to 20$
per barrel only [76].
e application of microalgae for the production of biofuels can
also help in other purposes, some of them are the following: Dur-
ing biodiesel production, elimination of CO2 from industrial gases
using algal xation reduces the greenhouse gas eects [78]. During
treatment of wastewater for removal of NO3, PO4, NH3 using micro-
algae, it utilizes these water pollutants as nutrients. Aer extraction of
oil, resultant algal biomass could be used as a feedstock for methane,
ethanol, biofertilizers because of its high N:P ratio or can be burned
to energy for the production of heat and electricity [79]. Microalgae
can produce high value bio-derivatives, therefore, have potential to
transform number of biotechnology elds such as biofuels, pharma-
ceuticals, cosmetics, nutrition, food additives and pollution preven-
tion [78].

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Table 2: Biofuel productivity and types being produced by dierent microalgae.
Cultivation and Harvesting of Microalgae
e whole process of obtaining biofuels from microalgae is sum-
marized in Figure 3.
Microalgae Name Algae
Type
Biofuel Productivity
of Biofuel
References
Chlorococum sp. Green Biodiesel 10 g L-1 Harun and Dan-
quah., 2011 [122]
Arthrospira maxima Blue-
Green
Hydrogen,
Biodiesel
40-69% Ananyev et al., 2008
[113]
Chlorella sp. Green Ethanol 22.6 g L-1 Zhou et al., 2011
[134]
Haematococcus
pluvialis
Red Biodiesel 420 GJ/
ha/yr
Li et al., 2008 [123]
Spirulina platensis
UTEX 1926
Blue-
Green
Methane 0.40 m3 kg-1 Maity et al., 2014
[125]
Spirulina sp. LEB 18 Blue-
Green
Methane 0.79 g L-1 Brennan and
Owende., 2010 [3]
Dunaliella sp. Green Ethanol 11.0 mg g-1 Shirai et al., 1998
[131]
Chlorella protothe-
coides
Green Biodiesel 15.5 g L-1 Chen and Walker,
2011a [85]
Neochlorosis olea-
bundans
Green Biodiesel 56.0 g g-1 Gouveia and Olivei-
ra., 2009 [121]
Spirulina platensis Green Hydrogen 1.8 µmol
mg-1
Aoyama et al., 1997
[114]

18 www.avidscience.com
Top 5 Contributions in Energy Research and Development: 3rd Edition
Figure 3: Schematic presentation of biofuel production from microalgae. Microalgae
are cultivated by dierent methods e.g. photoautotrophic, heterotrophic, photohetero-
trophic and mixotrophic, then harvested by the bulking method in which microalgae
are isolated from suspension through oatation, occulation or gravity sedimentation.
ickening is the second stage used to concentrate the algal slurry aer bulking pro-
cess.
Cultivation
Microalgae can adjust themselves metabolically through physi-
ological and biochemical acclimation in response to environmental
changes, so dierent cultivation methods (Figure 4) can be employed
such as photoautotrophic, heterotrophic, photoheterotrophic and
mixotrophic [78,80].
Photoautotrophic Cultivation: e process uses the light source
as energy to produce chemical energy by photosynthesis reaction,
while inorganic carbon is utilized as carbon source [81]. Currently,
this method is the only process that is economically and technically
suitable for the viable production of algal biomass [3]. Two variants
have been developed for the photoautotrophic cultivation of algae,
(1) Open pond production, (2) Closed photobioreactors [82]. e
higher productivity of biomass can be obtained by the closed pho-

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toautotrophic bioreactors than open pond production of microalgae
[3]. A carbon dioxide rich environment can increase the productiv-
ity of biomass to a certain level [83]. However, the light penetration
decreases rapidly by the increment of broth turbidity thus creating
diculty to achieve the very large-scale productivity of biomass [84].
Lipid concentration has great importance as a parameter for biodiesel
production [76]. e lipid concentration (dry weight) varies in mi-
croalgae ranging from 5-68% in photoautotrophic cultivation [85].
A large content of lipids could be obtained in the environment in
which nitrogen is present in less amount and limited concentration.
However, the productivity of biomass that is achieved in this type of
stressed environment is usually much lower than that is obtained in
the normal conditions, which results in even lower productivity of
microalgal lipid [81].
Heterotrophic Cultivation: In this method, microalgae are
grown on organic sources. e carbon source is used as both carbon
and energy source [3]. e heterotrophic method avoids the limita-
tions that were present in the photoautotrophy, so higher productiv-
ity of biomass can be achieved. Lipid concentration in this method is
much higher than the photoautotrophic method [81]. is method
is light independent so simpler and smaller fermenters or reactors
are used for cultivation [3]. ese setups produce a higher level of
growth and also decrease the cost of harvesting because of high level
of obtained cell densities [86]. A wide range of organic substrates is
utilized in heterotrophy such as organic carbon, sugar sources [87].
However, some cheaper sources like glycerol and corn powder can
also provide some good yield. A large number of organic compounds
and fermenters are required for heterotrophic cultivation, so the cost
of this method is higher than that of photoautotrophy [81]. Another
disadvantage of heterotrophy is the susceptibility to be contaminated
with other microorganisms, thereby may lower the quantity and the
quality of the products [85].
Photoheterotrophic Cultivation: is mode is also known as
photoassimilation, photometabolism or photoganitrophy, the mode
in which the light is needed to use the organic compounds as the

20 www.avidscience.com
Top 5 Contributions in Energy Research and Development: 3rd Edition
source of carbon [78]. e key dierence between the photohetero-
trophic and mixotrophic is that, for the later, only light is used as an
energy source. Moreover, in photoheterotrophic cultivation, light
and other organic sources both are essential at the same time. Even
though the production of metabolites controlled by the light intensity
could be enhanced in photoheterotrophic cultivation, the purpose of
this type of cultivation for biodiesel production is very rare, as is the
mixotrophic cultivation. Both types of cultivation are constrained by
the risk of contamination and the presence of light may necessitate
a large-scale photobioreactor, resulting economical constrains [88].
Mixotrophic Cultivation: Mixotrophic cultivation is the method
in which organisms are capable of using either autotrophic or het-
erotrophic modes [3]. ereby, microalgae can utilize both organic
carbon and inorganic source [81]. Inorganic compounds are xed by
photosynthesis and organic sources are adjusted by aerobic respira-
tion that is aected by the amount of available organic carbon [89].
Main dierence between the mixotrophy and the photoheterotrophy
is that the photoheterotrophy needs light as the main energy source
(photoheterotrophy needs light and the organic compounds at the
same time), whereas the mixotrophy could use the organic compo-
nents to attain that and it is rarely adopted to produce the microal-
gal diesel [85]. Some studies show that the microalgal growth rate
in mixotrophic cultivation is almost the total sum of growth under
heterotrophic and photoautotrophic modes [90]. Since mixotrophic
cultivation utilizes the organic compounds, the growth or algae does
not solely dependent on photosynthesis; light is not a limiting factor
for growth of microalgae, therefore photo-inhibition or photo-limi-
tation can be decreased in the mixotrophic culture, especially when
the light levels are too high or too low [81]. Mixotrophic cultivation
can enhance the growth rate, reduce the growth cycle, decrease the
biomass loss when light is not present due to clean respiration, thus
can enhance the productivity of biomass [91]. Lipid content can be
increased as well, that can lead to higher productivity of lipid [92].
Furthermore, the carbon dioxide released via aerobic respiration of

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microalgae can be reused for further photosynthesis in mixotrophic
cultivation, which increases the productivity of lipids and biomass
[78]. e method can reduce the cost of cultivation of microalgae, so
this mode is benecial for the production of microalgal biodiesel [81].
Figure 4: Cultivation methods of microalgae: (A) Photoautotrophic cultivation: Open
pond (upper; https://algaeforbiofuels.com) or closed bioreactor (lower; https://www.
treehugger.com); (B) Heterotrophic cultivation [93]; (C) Photoheterotrophic cultiva-
tion (Author’s Lab Source); (D) Mixotrophic cultivation [94].
Harvesting
Microalgal harvesting costs about 20-30% of the total produc-
tion of biomass [95]. Dierent harvesting methods could be adopted
depending on the density, size, and value of the product [96]. Gener-
ally harvesting methods consist of two stages [1]; (1) Bulk harvesting
is the basic process to isolate microalgae from suspension through
oatation, occulation or gravity sediments (2) ickening, is the
second stage used to concentrate the algal slurry aer the application
of bulking technique. Methods that are mostly used at this stage are
centrifugation, ultrasonic aggregation, and ltration. So, this is more
concentrated stage than bulking [3].

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Top 5 Contributions in Energy Research and Development: 3rd Edition
Depending on the density and size of the microalgae, four key
methods that are used for harvesting with respect to biofuel produc-
tion are; belt ltering, sedimentation, otation, and mircrostaining.
Microstrainers are striking method for harvesting because of their
mechanical simplicity and availability of large size units. e new and
better accessibility polyester screens have revitalized the interest of
their use in harvesting microalgae. Recent studies show that it would
be essential to occulate the cells before the mircrostaining process
[95]. Till now centrifugation and ltration are not feasible modes for
microalgal harvesting at commercial level, because they need high
energy and high level of maintenance cost, which make them un-
favorable for long term use. While on the other hand, occulation
requires less energy for microalgal harvesting. is happens because
the microalgal cells carry -ve charge, therefore repels each other and
could be suspended in the medium for long period when mixing is
not forced to them. By adding +vely charged coagulant into the me-
dium, the charge of microalgal cells would be neutralized. Meanwhile
occulent could be neutralized and form dense ocs that settle under
the natural gravity [1].
The Bottleneck of Microalgae as a Biofuel
Source
Microalgae are the unexploited resources having more than
25,000 species from which only a few are in use [97]. Currently the
commercial production of microalgal fuels is limited [98]. e main
hurdles to the microalgal fuel production include: Lack of Informa-
tion and knowledge at the major scale to grow algae eciently; Large
land and equipment are required for open pond cultivation method;
Cost of harvesting and oil extraction from algae is quite high; Large
concentration of alcohol is required for transesterication [75]. e
process of dewatering of microalgae is also a major barrier to the in-
dustrial scale production of biofuels from microalgae. e watery na-

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ture of the harvested microalgae causes a huge cost during the process
of dewatering so this makes the algal fuel less attractive economically.
No other technique at present is available as an alternative to dewater-
ing; consequently the production of algal fuels causes high cost [99].
Moreover, algal species diversity, cultivation conditions (Photobiore-
actors, Open ponds, etc.) and the use of dierent biomass are the rea-
sons for microalgal product heterogenity [100]. e large scale open
ponds had lesser productivity (due to low temperature during night
in areas where these ponds are established) than required to make the
process economically feasible [101]. e complex processing and har-
vesting procedures collectively with inadequately produced dry mass
of algae are the restrictive factors for biofuel production [102].
e exploitation of existing biofuel production processes needs
lipid material free from water and fatty acids that also leads to higher
cost of the process [103]. e algal biofuel production costs about 50
€/L, which is far away to appeal the commercial production of biofu-
els from algae [104]. ere are many limitations in the photosynthetic
growth of microalgae that include; biosynthesis rates, temperature,
CO2 limitation, nutrient limitations, self-shading, light saturation and
photo-inhibition [44]. Another bottleneck for algal fuel production
is the low mass concentration in the algal culture due to limited light
penetration, which in association with small algal cells also makes the
process of harvesting more costly. e higher costs and exhaustive
care required for microalgal farming in comparison with the agricul-
tural farms are among major factors that obstruct the commercial op-
eration of the algal biofuel strategy [105].
Future Prospects
Energy demand has increased dramatically particularly in devel-
oping countries where energy is required for economic growth [106].
Projected world demand for primary energy by 2050 is expected to be
in 600-1000 EJ per year [107]. Biomass can be used for hydrogen pro-
duction but this technology needs more development. It is believed
that in future biomass will be an important source of hydrogen. e

24 www.avidscience.com
Top 5 Contributions in Energy Research and Development: 3rd Edition
share of hydrogen produced from biomass in automotive fuel mar-
ket will be high in next decade [108]. Biomass is harvested and pro-
cessed to make triacylglycerides (TAGs) that can by trans-esteried to
produce biodiesel with methanol, to yield methyl esters of fatty acids
[109]. e ever increasing policy driven demands for biofuels in com-
bination with concerns that available technologies are falling short of
expectations have asserted more pressure for production of second
and third generation biofuels [110]. Microalgal ponds have 1-4% e-
ciency of solar energy conversion under normal operating conditions,
while higher eciencies can be achieved with closed photo-bioreac-
tor systems [111]. Microalgal biofuel systems are independent of soil
fertility, therefore can contribute to the fulllment of global demand
without increasing pressure on arable land. Mostly microalgae can be
grown in saline water and capable to produce vast variety of feed-
stock for biofuel production. Economic analysis suggests that today’s
biofuel system is dependent on the development of photoautotrophic
production systems for the production of co-products for protable-
ness [112]. Development of dierent technologies including micro-
algal biomass harvesting, advances in photobioreactor design may
lead to enhanced cost-eectiveness. In future, genetic modication
of microalgae for simultaneous high biomass productivity and lipid
content may also contribute considerably. Due to high tolerance level
of CO2 and use of carbon as energy source, microalgae can be utilized
to clean industrial gas emissions, thereby reducing GHGs emissions.
Microalgae can use nutrients form variety of wastewater sources, so
providing useful bioremediation of wastewater for economic and en-
vironmental benets.
Conclusion
Algae are an economical option for biofuel production as a feed-
stock without competing with the cultivation area as compared to
other available feedstock. e oil production rate and lipid productiv-
ity from microalgae are far higher than from other feedstock. Open
ponds used for algal cultivation are economically suitable for micro-

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algal production alongside wastewater treatment. Whereas, closed
ponds are resistant against environmental uctuations such as rain-
fall, evaporation, and temperature. Another key benet of microalgae
is the potential to capture greenhouse gases such as CO2 from fossil
power plants as well as from atmosphere. erefore, microalgae are
very ecofriendly to eliminate greenhouse gas emissions and useful for
the large scale production of biofuels.
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