Microalgae for biodiesel production and other applications: A review
Teresa M. Mata, Anto
´
nio A. Martins, Nidia. S. Caetano
Keywords:
Microalgae
Biodiesel
Photo-bioreactors
Open ponds
Environmental applications
High-value chemical compounds
ABSTRACT
Sustainable production of renewable energy is being hotly debated globally since it is increasingly understood that first generation
biofuels, primarily produced from food crops and mostly oil seeds are limited in their ability to achieve targets for biofuel production,
climate change mitigation and economic growth. These concerns have increased the interest in developing second generation biofuels
produced from non-food feedstocks such as microalgae, which potentially offer greatest opportunities in the longer term. This paper
reviews the current status of microalgae use for biodiesel production, including their cultivation, harvesting, and processing. The
microalgae species most used for biodiesel production are presented and their main advantages described in comparison with other
available biodiesel feedstocks. The various aspects associated with the design of microalgae production units are described, giving an
overview of the current state of development of algae cultivation systems (photo-bioreactors and open ponds). Other potential
applications and products from microalgae are also presented such as for biological sequestration of CO
2
, wastewater treatment, in
human health, as food additive, and for aquaculture.
1. Introduction
The transportation and energy sectors are the major anthro-
pogenic sources, responsible in European Union (EU) for more than
20% and 60% of greenhouse gas (GHG) emissions, respectively [1].
Agriculture is the third largest anthropogenic source, representing
about 9% of GHG emissions, where the most important gases are
nitrous oxide (N
2
O) and methane (CH
4
) [2]. It is expected that with
the development of new growing economies, such as India and
China, the global consumption of energy will raise and lead to more
environmental damage [3].
GHG contributes not only to global warming (GW) but also to
other impacts on the environment and human life. Oceans absorb
approximately one-third of the CO
2
emitted each year by human
activities and as its levels increase in the atmosphere, the amount
dissolved in oceans will also increase turning the water pH
gradually to more acidic. This pH decrease may cause the quick loss
of coral reefs and of marine ecosystem biodiversity with huge
implications in ocean life and consequently in earth life [4].
As GW is a problem affecting different aspects of human life and
the global environment, not only a single but a host of solutions is
needed to address it. One side of the problem concerns the
reduction of crude oil reserves and difficulties in their extraction
and processing, leading to an increase of its cost [5]. This situation
is particularly acute in the transportation sector, where currently
there are no relevant alternatives to fossil fuels.
To find clean and renewable energy sources ranks as one of the
most challenging problems facing mankind in the medium to long
term. The associated issues are intimately connected with
economic development and prosperity, quality of life, global
stability, and require from all stakeholders tough decisions and
long term strategies. For example, many countries and regions
around the world established targets for CO
2
reduction in order to
meet the sustainability goals agreed under the Kyoto Protocol.
Presently many options are being studied and implemented
in practice, with different degrees of success, and in different
phases of study and implementation. Examples include solar
energy, either thermal or photovoltaic, hydroelectric, geother-
mal, wind, biofuels, and carbon sequestration, among others
[6,7]. Each one has its own advantages and problems and,
depending on the area of application, different options will be
better suited. One important goal is t o take measures for
transportation emissions reduction, such as the gradual replace-
ment of fossil fuels by renewable energy sources, where biofuels
are seen as real contributors to reach those goals, particularly in
the short term.
Biofuels production is expected to offer new opportunities to
diversify income and fuel supply sources, to promote employment
in rural areas, to develop long term replacement of fossil fuels, and
to reduce GHG emissions, boosting the decarbonisation of
transportation fuels and increasing the security of energy supply.
The most common biofuels are biodiesel and bio-ethanol,
which can replace diesel and gasoline, respectively, in today cars
with little or none modifications of vehicle engines. They are
mainly produced from biomass or renewable energy sources and
contribute to lower combustion emissions than fossil fuels per
equivalent power output. They can be produced using existing
technologies and be distributed through the available distribution
system. For this reason biofuels are currently pursued as a fuel
alternative that can be easily applied until other options harder to
implement, such as hydrogen, are available.
Although biofuels are still more expensive than fossil fuels their
production is increasing in countries around the world. Encour-
aged by policy measures and biofuels targets for transport, its
global production is estimated to be over 35 billion liters [8].
The main alternative to diesel fuel in EU is biodiesel,
representing 82% of total biofuels production [9] and is still
growing in Europe, Brazil, and United States, based on political and
economic objectives.
Biodiesel is produced from vegetable oils (edible or non-edible)
or animal fats. Since vegetable oils may also be used for human
consumption, it can lead to an increase in price of food-grade oils,
causing the cost of biodiesel to increase and preventing its usage,
even if it has advantages comparing with diesel fuel.
The potential market for biodiesel far surpasses the availability
of plant oils not designated for other markets. For example, to fulfill
a 10% target in EU from domestic production, the actual feedstocks
supply is not enough to meet the current demand and the land
requirements for biofuels production, would be more than the
potential available arable land for bio-energy crops [10]. The
extensive plantation and pressure for land use change and increase
of cultivated fields may lead to land competition and biodiversity
loss, due to the cutting of existing forests and the utilization of
ecological importance areas [11]. Biodiesel may also be dis-
advantageous when replacing crops used for human consumption
or if its feedstocks are cultivated in forests and other critical
habitats with associated biological diversity.
Current policies at regional and national levels and the expected
cost and difficulties in obtaining fossil fuels will necessarily lead to
an increase in biodiesel production and of other types of renewable
energy. To become a more viable alternative fuel and to survive in
the market, biodiesel must compete economically with diesel. The
end cost of biodiesel mainly depends on the price of the feedstocks
that accounts for 60–75% of the total cost of biodiesel fuel [12].
In order to not compete with edible vegetable oils, the low-cost
and profitable biodiesel should be produced from low-cost
feedstocks such as non-edible oils, used frying oils, animal fats,
soap-stocks, and greases. However the available quantities of
waste oils and animal fats are not enough to match the today
demands for biodiesel. Thus transition to second generation
biofuels, such as microalgae, can also contribute to a reduction in
land requirements due to their presumed higher energy yields per
hectare as well as to their non-requirement of agricultural land.
Additionally, biodiesel needs to have lower environmental impacts
and ensure the same level of performance of existing fuels [13].
Albeit the growing interest and fast growth of this area, it is still
on its infancy. A large investment in research and development
(R&D) and correct policies and strategies are still needed, for all
stages of the biofuels value chain, from raw materials production to
delivery and final consumption. Among the various possibilities
currently being investigated and implemented at pilot scale or
even at industrial scale concerning potential feedstocks, the more
interesting ones are microalgae. Besides their cultivation is not
directly linked to human consumption, they have low space
requirements for its production.
This review focuses its attention on microalgae and how they can
be used for biodiesel production. Questions associated with
production and processing of microalgae are considered in detail,
not only those directly related with biofuels production but also the
possibilities of combiningit withpollution control,in particular with
biological sequestration of CO
2
emissions and other greenhouse
gases, or wastewater treatment. This work starts by describing
which microalgae are normally used for the production of biofuels
and their main advantages when compared with other available
feedstocks. Then, the current status of biodiesel production from
microalgae, concerning their growth, harvest, and processing is
reviewed. Other potential applications and how to combine them
with biodiesel production are also described.
2. Microalgae for biodiesel production
2.1. Viability of microalgae for biodiesel
2.1.1. What are microalgae?
Microalgae are prokaryotic or eukaryotic photosynthetic
microorganisms that can grow rapidly and live in harsh conditions
due to their unicellular or simple multicellular structure. Examples
of prokaryotic microorganisms are Cyanobacteria (Cyanophyceae)
and eukaryotic microalgae are for example green algae (Chlor-
ophyta) and diatoms (Bacillariophyta) [14,15]. A more in depth
description of microalgae is presented by Richmond [16].
Microalgae are present in all existing earth ecosystems, not just
aquatic but also terrestrial, representing a big variety of species
living in a wide range of environmental conditions. It is estimated
that more than 50,000 species exist, but only a limited number, of
around 30,000, have been studied and analyzed [16].
During the past decades extensive collections of microalgae
have been created by researchers in different countries. An
example is the freshwater microalgae collection of University of
Coimbra (Portugal) considered one of the world’s largest, having
more than 4000 strains and 1000 species. This collection attests to
the large variety of different microalgae available to be selected for
use in a broad diversity of applications, such as value added
products for pharmaceutical purposes, food crops for human
consumption and as energy source.
A bit all over the world, other algae collections attest for the
interest that algae have risen, for many different production
purposes. For example, the collection of the Goettingen University,
Germany (SAG), that started in the early 1920s and has about 2213
strains and 1273 species. About 77% of all the strains in the SAG
collection are green algae and about 8% cyanobacteria (61 genera
and 230 strains). Some of them are freshwater red algae and others
from saline environments.
The University of Texas Algal Culture Collection is another very
well known collection of algae cultures that was founded in 1953.
It includes 2300 different strains of freshwater algae (edaphic
green algae and cyanobacteria), but includes representatives of
most major algal taxa, including many marine macrophytic green
and red algae species.
In the Asian continent, the National Institute for Environmental
Studies Collection (NIES), in Ibaraki, Japan, holds a collection of
about 2150 strains, with around 700 species of different algae. The
CSIRO Collection of Living Microalgae (CCLM), in Australia, holds
about 800 strains of different algae, including representatives from
the majority of classes of marine and some freshwater microalgae,
being the majority of the strains isolated from Australian waters.
2.1.2. Advantages of using microalgae for biodiesel production
Many research reports and articles described many advantages
of using microalgae for biodiesel production in comparison with
other available feedstocks [14,15,17,21–27]. From a practical point
of view, they are easy to cultivate, can grow with little or even no
attention, using water unsuitable for human consumption and
easy to obtain nutrients.
Microalgae reproduce themselves using photosynthesis to
convert sun energy into chemical energy, completing an entire
growth cycle every few days [17]. Moreover they can grow almost
anywhere, requiring sunlight and some simple nutrients, although
the growth rates can be accelerated by the addition of specific
nutrients and sufficient aeration [18–20].
Different microalgae species can be adapted to live in a variety
of environmental conditions. Thus, it is possible to find species best
suited to local environments or specific growth characteristics,
which is not possible to do with other current biodiesel feedstocks
(e.g. soybean, rapeseed, sunflower and palm oil).
They have much higher growth rates and productivity when
compared to conventional forestry, agricultural crops, and other
aquatic plants, requiring much less land area than other biodiesel
feedstocks of agricultural origin, up to 49 or 132 times less when
compared to rapeseed or soybean crops, for a 30% (w/w) of oil
content in algae biomass [21]. Therefore, the competition for
arable soil with other crops, in particular for human consumption,
is greatly reduced.
Microalgae can provide feedstock for several different types of
renewable fuels such as biodiesel, methane, hydrogen, ethanol,
among others. Algae biodiesel contains no sulfur and performs as
well as petroleum diesel, while reducing emissions of particulate
matter, CO, hydrocarbons, and SO
x
. However emissions of NO
x
may
be higher in some engine types [28].
The utilization of microalgae for biofuels production can also
serve other purposes. Some possibilities currently being consid-
ered are listed below.
Removal of CO
2
from industrial flue gases by algae bio-fixation
[29], reducing the GHG emissions of a company or process while
producing biodiesel [30].
Wastewater treatment by removal of NH
4
+
,NO
3
,PO
4
3
, making
algae to grow using these water contaminants as nutrients [29].
After oil extraction the resulting algae biomass can be processed
into ethanol, methane, livestock feed, used as organic fertilizer
due to its high N:P ratio, or simply burned for energy co-
generation (electricity and heat) [29];
Combined with their ability to grow under harsher conditions,
and their reduced needs for nutrients, they can be grown in areas
unsuitable for agricultural purposes independently of the
seasonal weather changes, thus not competing for arable land
use, and can use wastewaters as the culture medium, not
requiring the use of freshwater.
Depending on the microalgae species other compounds may
also be extracted, with valuable applications in different
industrial sectors, including a large range of fine chemicals
and bulk products, such as fats, polyunsaturated fatty acids, oil,
natural dyes, sugars, pigments, antioxidants, high-value bioac-
tive compounds, and other fine chemicals and biomass
[14,15,31].
Because of t his variety of high-value biological derivatives,
with many possible commercial applications, microalgae can
potentially revolutionize a large number of biotechnology
areas including biofuels, cosmetics, pharmaceuticals, nutrition
and food additives, aquaculture, and pollution prevention
[25,31].
2.1.3. Historic evolution of microalgae production systems
For the past 50 years, extensive research has been performed
on microalgae and how they can be used in a wide variety of
processes or to manufacture many practical and economic
important products. The first large-scale culture of microalgae
started in the early 1960s in Japan by Nihon Chlorella with the
culture of Chlorella [32]. The interest in using microalgae for
renewable energy increased in 1970s during the first oil crisis [32].
The U.S. National Renewable Energy Laboratory (NREL) through
the Aquatic Species Program (ASP), launched a specific R&D
Program dedicated to alternative renewable fuels, including
biodiesel from microalgae that lasted from 1978 to 1996 [17].
One of its main objectives was to study the biochemistry and
physiology of lipid production in oleaginous microalgae. From
1987 to 1990, an ‘‘Outdoor Test Facility’’ of two 1000 m
2
high-rate
ponds was operated in Roswell, New Mexico. It was concluded that
the use of microalgae for the low-cost production of biodiesel was
technically feasible, but still needs considerable long term R&D to
achieve the high productivities required. Other objective of this
NREL R&D program was to produce improved algae strains by
looking for genetic variability between algal isolates, attempting to
use flow cytometry to screen for naturally occurring high lipid
individuals, and exploring algal viruses as potential genetic
vectors. However in 1995 the Department of Energy reduced
the budget allocated to funding this program and it was
discontinued before these experiments could be carried out
beyond the preliminary stages [17].
The recent price volatility of crude oil and the expected future
price increase, tied with the urge to reduce pollutant emissions and
greenhouse gases, have created a new interest in the production of
biodiesel using microalgae. For example, several companies were
created or have entered this market niche, selling either entire
processes or key process units, such as photo-bioreactors with
optimized designs to cultivate microalgae for biodiesel production
and other applications [33–35]. Torrey [37] presents links to 37
companies that are currently exploring algae as a fuel source.
Nowadays, microalgae are seen as an alternative feedstock for
biodiesel production, being the target of a large number of
consortiums, private and public organizations’ investments in
R&D, aiming to use the most effective and cheap technology to
produce large amounts of oil. They are considered to be a second
generation feedstock together with other biomass sources, such as
Jatropha, lignocellulosic materials, agricultural residues, and
systematically grown energy crops, with high potential yields of
biofuels and that are not used as food source for human
consumption [36].
Though it is not cost effective yet to compete with fossil diesel
without additional support (for example government subsidies)
research is being done to turn it economically viable, both in
academia and in industry [35,38,39]. In a long term, as crude oil
reserves diminish and price per barrel increases in a daily basis,
other alternatives must become available, and thus, it is now the
time to search, develop and implement them.
Recent research efforts have concentrated on applying meta-
bolic engineering and genetic methods to microalgae in order to
develop organisms optimized for high productivity and energy
Table 1
Lipid content and productivities of different microalgae species.
Marine and freshwater
microalgae species
Lipid content
(% dry weight biomass)
Lipid
productivity (mg/L/day)
Volumetric productivity
of biomass (g/L/day)
Areal productivity
of biomass (g/m
2
/day)
Ankistrodesmus sp. 24.0–31.0 – – 11.5–17.4
Botryococcus braunii 25.0–75.0 – 0.02 3.0
Chaetoceros muelleri 33.6 21.8 0.07 –
Chaetoceros calcitrans 14.6–16.4/39.8 17.6 0.04 –
Chlorella emersonii 25.0–63.0 10.3–50.0 0.036–0.041 0.91–0.97
Chlorella protothecoides 14.6–57.8 1214 2.00–7.70 –
Chlorella sorokiniana 19.0–22.0 44.7 0.23–1.47 –
Chlorella vulgaris 5.0–58.0 11.2–40.0 0.02–0.20 0.57–0.95
Chlorella sp. 10.0–48.0 42.1 0.02–2.5 1.61–16.47/25
Chlorella pyrenoidosa 2.0 – 2.90–3.64 72.5/130
Chlorella 18.0–57.0 18.7 – 3.50–13.90
Chlorococcum sp. 19.3 53.7 0.28 –
Crypthecodinium cohnii 20.0–51.1 – 10 –
Dunaliella salina 6.0–25.0 116.0 0.22–0.34 1.6–3.5/20–38
Dunaliella primolecta 23.1 – 0.09 14
Dunaliella tertiolecta 16.7–71.0 – 0.12 –
Dunaliella sp. 17.5–67.0 33.5 – –
Ellipsoidion sp. 27.4 47.3 0.17 –
Euglena gracilis 14.0–20.0 – 7.70 –
Haematococcus pluvialis 25.0 – 0.05–0.06 10.2–36.4
Isochrysis galbana 7.0–40.0 – 0.32–1.60 –
Isochrysis sp. 7.1–33 37.8 0.08–0.17 –
Monodus subterraneus 16.0 30.4 0.19 –
Monallanthus salina 20.0–22.0 – 0.08 12
Nannochloris sp. 20.0–56.0 60.9–76.5 0.17–0.51 –
Nannochloropsis oculata. 22.7–29.7 84.0–142.0 0.37–0.48 –
Nannochloropsis sp. 12.0–53.0 37.6–90.0 0.17–1.43 1.9–5.3
Neochloris oleoabundans 29.0–65.0 90.0–134.0 – –
Nitzschia sp. 16.0–47.0 8.8–21.6
Oocystis pusilla 10.5 – – 40.6–45.8
Pavlova salina 30.9 49.4 0.16 –
Pavlova lutheri 35.5 40.2 0.14 –
Phaeodactylum tricornutum 18.0–57.0 44.8 0.003–1.9 2.4–21
Porphyridium cruentum 9.0–18.8/60.7 34.8 0.36–1.50 25
Scenedesmus obliquus 11.0–55.0 – 0.004–0.74 –
Scenedesmus quadricauda 1.9–18.4 35.1 0.19 –
Scenedesmus sp. 19.6–21.1 40.8–53.9 0.03–0.26 2.43–13.52
Skeletonema sp. 13.3–31.8 27.3 0.09 –
Skeletonema costatum 13.5–51.3 17.4 0.08 –
Spirulina platensis 4.0–16.6 – 0.06–4.3 1.5–14.5/24–51
Spirulina maxima 4.0–9.0 – 0.21–0.25 25
Thalassiosira pseudonana 20.6 17.4 0.08 –
Tetraselmis suecica 8.5–23.0 27.0–36.4 0.12–0.32 19
Tetraselmis sp. 12.6–14.7 43.4 0.30 –
value, in order to achieve their full processing capabilities [25,31].
Since microalgae represent a much simpler system than plants,
usually with no cell differentiation, genetic manipulations to
increase its content of higher value compounds is very tempting.
Nevertheless, progress in the genetic engineering of algae was
extremely slow until recently. Also, these promising advances
should be viewed with caution because transgenic algae poten-
tially pose a considerable threat to the ecosystem and thus will
most likely be banned from outdoor cultivation systems [40].
2.1.4. Microalgae lipid content and productivities
Many microalgae species can be induced to accumulate
substantial quantities of lipids [17] thus contributing to a high
oil yield. The average lipid content varies between 1 and 70% but
under certain conditions some species can reach 90% of dry weight
[14,15,21,32].
Table 1 presents both lipid content and lipid and biomass
productivities of different marine and freshwater microalgae
species, showing significant differences between the various
species [15,16,20,21,24,32,41–68].
As shown in Table 1, oil content in microalgae can reach 75% by
weight of dry biomass but associated with low productivities (e.g.
for Botryococcus braunii). Most common algae (Chlorella, Crypthe-
codinium, Cylindrotheca, Dunaliella, Isochrysis, Nannochloris, Nanno-
chloropsis, Neochloris, Nitzschia, Phaeodactylum, Porphyridium,
Schizochytrium, Tetraselmis) have oil levels between 20 and 50%
but higher productivities can be reached.
Chlorella seems to be a good option for biodiesel production.
Yet, as other species are so efficient and productive as this one, the
selection of the most adequate species needs to take into account
other factors, such as for example the ability of microalgae to
develop using the nutrients available or under specific environ-
mental conditions. All these parameters should be considered
simultaneously in the selection of the most adequate species or
strains for biodiesel production.
Also significant is the composition of fatty acids of the different
microalgae species, as they can have a significant effect on the
characteristics of biodiesel produced. These are composed of
saturated and unsaturated fatty acids with 12–22 carbon atoms,
some of them of
v
3 and
v
6 families. Thomas et al. [66] analyzed
the fatty acid compositions of seven fresh water microalgae species
showing that all of them synthesized C14:0, C16:0, C18:1, C18:2,
and C18:3 fatty acids. This author reported that the relative
intensity of other individual fatty acids chains is species specific,
e.g. C16:4 and C18:4 in Ankistrodesmus sp., C18:4 and C22:6 in
Isochrysis sp., C16:2, C16:3 and C20:5 in Nannochloris sp., C16:2,
C16:3, and C20:5 in Nitzschia sp.
Different nutritional and environmental factors, cultivation
conditions and growth phases may affect the fatty acid composi-
tion. For example, nitrogen deficiency and salt stress induced the
accumulation of C18:1 in all treated species and to some extent
C20:5 in B. braunii [66]. Other authors also reported a differentia-
tion between fatty acid composition of various algae species
[19,23,47,59,62,69].
Although the microalgae oil yield is strain-dependent it is
generally much greater than other vegetable oil crops, as shown in
Table 2 that compares the biodiesel production efficiencies and
land use of microalgae and other vegetable oil crops, including the
amount of oil content in a dry weight basis and the oil yield per
hectare, per year [21,65,70–80].
Table 2 shows that although the oil contents are similar
between seed plants and microalgae there are significant varia-
tions in the overall biomass productivity and resulting oil yield and
biodiesel productivity with a clear advantage for microalgae. In
terms of land use, microalgae followed by palm oil biodiesel are
clearly advantageous because of their higher biomass productivity
and oil yield.
2.2. Microalgae biodiesel value chain stages
Although in a simplistic view microalgae may seem to not
significantly differ from other biodiesel feedstocks, they are
microorganisms living essentially in liquid environments, and
thus with particular cultivation, harvesting, and processing
techniques that ought to be considered in order to efficiently
produce biodiesel.
All existing processes for biodiesel production from microalgae
include a production unit where cells are grown, followed by the
separation of the cells from the growing media and subsequent
lipids extraction. Then, biodiesel or other biofuels are produced in a
form akin to existing processes and technologies used for other
biofuels feedstocks. Recently other possibilities for biofuel
production are being pursued instead of the transesterification
reaction, such as the thermal cracking (or pyrolysis) involving the
thermal decomposition or cleavage of the triglycerides and other
organic compounds presented in the feedstock, in simpler
molecules, namely alkans, alkenes, aromatics, carboxylic acids,
among others [81–83].
Fig. 1 shows a schematic representation of the algal biodiesel
value chain stages, starting with the selection of microalgae
species depending on local specific conditions and the design and
implementation of cultivation system for microalgae growth.
Then, it follows the biomass harvesting, processing and oil
extraction to supply the biodiesel production unit.
In the next subsections issues related to each algal biodiesel
value chain stage are presented and discussed.
2.2.1. Algae and site selection
Currently a lot of research effort is being focused on the algal
cultivation unit, as in most cases it represents the key step that
Table 2
Comparison of microalgae with other biodiesel feedstocks.
Plant source Seed oil content
(% oil by wt in biomass)
Oil yield
(L oil/ha year)
Land use
(m
2
year/kg biodiesel)
Biodiesel productivity
(kg biodiesel/ha year)
Corn/Maize (Zea mays L.) 44 172 66 152
Hemp ( Cannabis sativa L.) 33 363 31 321
Soybean (Glycine max L.) 18 636 18 562
Jatropha (Jatropha curcas L.) 28 741 15 656
Camelina (Camelina sativa L.) 42 915 12 809
Canola/Rapeseed (Brassica napus L.) 41 974 12 862
Sunflower (Helianthus annuus L.) 40 1070 11 946
Castor (Ricinus communis) 48 1307 9 1156
Palm oil (Elaeis guineensis) 36 5366 2 4747
Microalgae (low oil content) 30 58,700 0.2 51,927
Microalgae (medium oil content) 50 97,800 0.1 86,515
Microalgae (high oil content) 70 136,900 0.1 121,104
