Accepted Manuscript

Review

Biosequestration of atmospheric CO2 and flue gas-containing CO2 by microal-

gae

Wai Yan Cheah, Pau Loke Show, Jo-Shu Chang, Tau Chuan Ling, Joon Ching
Juan

PII: S0960-8524(14)01628-9
DOI: http://dx.doi.org/10.1016/j.biortech.2014.11.026
Reference: BITE 14232

To appear in: Bioresource Technology

Received Date: 30 August 2014

Revised Date: 7 November 2014
Accepted Date: 9 November 2014

Please cite this article as: Cheah, W.Y., Show, P.L., Chang, J-S., Ling, T.C., Juan, J.C., Biosequestration of
atmospheric CO2 and flue gas-containing CO2 by microalgae, Bioresource Technology (2014), doi: http://dx.doi.org/
10.1016/j.biortech.2014.11.026

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 A manuscript submitted to Bioresource Technology Special Issue on
"Advances in Biofuels and Chemicals from Algae"

Biosequestration of atmospheric CO2 and flue gas-containing CO2 by microalgae

Wai Yan Cheaha, Pau Loke Showb,c, Jo-Shu Changd,e,f, Tau Chuan Linga, Joon Ching Juang*,h

a
Institute of Biological Sciences, Faculty of Science, University of Malaya, 50603 Kuala
Lumpur, Malaysia.
b
Department of Chemical and Environmental Engineering, Faculty of Engineering, University of
Nottingham Malaysia Campus, Jalan Broga, 43500 Semenyih, Selangor Darul Ehsan,
Malaysia.
c
Manufacturing and Industrial Processes Division, Faculty of Engineering, Centre for Food and
Bioproduct Processing, University of Nottingham Malaysia Campus, Jalan Broga, 43500
Semenyih, Selangor Darul Ehsan, Malaysia.
d
University Center for Bioscience and Biotechnology, National Cheng Kung University, Tainan
701, Taiwan.
e
Department of Chemical Engineering, National Cheng Kung University, Tainan 701, Taiwan
f
Research Center for Energy Technology and Strategy, National Cheng Kung University, Tainan
701, Taiwan.
g
Nanotechnology & Catalyse Research Centre (NANOCAT), University of Malaya, 50603 Kuala
Lumpur, Malaysia.
h
Laboratory of Advanced Catalysis and Environmental Technology, School of Science,
Malayssia.

*Corresponding address:
Joon Ching Juan
Nanotechnology & Catalysis Research Centre (NANOCAT), University of Malaya, 50603 Kuala
Lumpur, Malaysia
Laboratory of Advanced Catalysis and Environmental Technology, School of Science, Malaysia
Tel: +603-79676267
Email: jcjuan@um.edu.my (J.C. Juan).

1
Abstract

The unceasing rise of greenhouse gas emission has led to global warming and climate change.

Global concern on this phenomenon has put forward the microalgal-based CO2 sequestration

aiming to sequester carbon back to the biosphere, ultimately reducing greenhouse effects.

Microalgae have recently gained enormous attention worldwide, to be the valuable feedstock for

renewable energy production, due to their high growth rates, high lipid productivities and the

ability to sequester carbon. The photosynthetic process of microalgae uses atmospheric CO2 and

CO2 from flue gases, to synthesise nutrients for their growth. In this review article, we will

primarily discuss the efficiency of CO2 biosequestration by microalgae species, factors

influencing microalgal biomass productions, microalgal cultivation systems, the potential and

limitations of using flue gas for microalgal cultivation as well as the bio-refinery approach of

microalgal biomass.

Keywords

Biomass production, CO2 sequestration, Fixation efficiency, Flue gas, Microalgae

1. Introduction

The anthropogenic activities such as burning of fossil fuel, deforestation and energy

generation have caused intensive greenhouse gases emission. United States Environmental

Protection Agency (USEPA) has stated that energy production and consumption, mainly from

transportation, has contributed 71% of greenhouse gas emission world-wide in 2010. This

emission has increased by 35% from 1990 to 2010 (Wilbanks & Fernandez, 2014). Carbon

dioxide (CO2), which is a major greenhouse gas, has risen in its atmospheric concentration,

which is from 280 to 390 ppm since pre-industrial revolution till 2013 (Rahaman et al., 2011;

Singh & Ahluwalia, 2013). Though CO2 is essential for survival of photoautotrophs, however

2
this copious and unceasing rise concentration has contributed largely to global warming, as CO2

retains with atmospheric lifetime of 50 to 200 years (Van Den Hende et al., 2012). At present,

CO2 is contributing approximately 52% in total global warming (Wilbanks & Fernandez, 2014).

An example of effort can be seen under the Kyoto Protocol, 37 industrialised countries

and European community have agreed to reduce the greenhouse gases emission by 18% below

1990 level within the period of 2013 to 2020 (Yoo & Al-Attiyah, 2013). Attempts have been

made to mitigate the emissions, for instance, the carbon sequestration of biomass feedstock,

microalgae. Microalgae have been long recognised as the promising alternatives for biofuel

production, aiming to replace the utilisation of fossil fuel, and serves as the major feedstock

towards greenhouse gases mitigation. This approach of using microalgae is predominantly due to

their photosynthetic efficiency in bio-converting carbon dioxide (CO2), high biomass

productivity, high lipid accumulation, and their valuable non-fuel co-products (Xie et al., 2014).

The photosynthetic or autotrophic microalgae have high capabilities to fix CO2, which is 10-50

times more efficiently than terrestrial plants (Cheng et al., 2013; Lam et al., 2012). Terrestrial

plants are only expected to contribute only 3-6% reduction in global CO2 emissions (Ho et al.,

2011; Kao et al., 2014). During photosynthetic process, the CO2 is utilised as carbon source, to

be converted into organic compounds by using solar energy. Carbon is the most important

element for microalgae nutrition, followed by nitrogen and phosphorus (Lam & Lee, 2011).

Dried microalgal biomass contains approximately 50% of carbon, which are all derived from

CO2 (Lam et al., 2012). Approximately 1.83 kg of CO2 can be fixed in every 1 kg of microalgae

biomass production (Jiang et al., 2013). These studies have revealed the fact that microalgae

serve as the strong agent in CO2 biosequestration. There are many studies on the microalgal-

based CO2 bio-mitigation presently but there is lacking of review literature on the latest

3
technologies on microalgal cultivation, which mainly towards successful CO2 bio-sequestration

of atmospheric CO2 and flue gas-containing CO2. Therefore, this review attempts to summarise

and discuss the CO2 bioconversion efficiency of microalgae species, factors influencing

microalgal biomass productions, microalgal cultivation systems, the potential and limitation of

flue gas utilisation for microalgae cultivation, and the bio-refinery of microalgal biomass.

2. Microalgae: The agent for carbon sequestration

Microalgae are photosynthetic microorganisms, the basic plants present in abundance in

the nature. Algae are divided into five main groups, namely Chlorophyceae, Rhodophyceae,

Phaeophycea, Cynophyceae and Bacillariophyceae. Characteristics of each algae group are

summarised in Table 1. Microalgae able to duplicate very rapidly of its cell biomass, and achieve

100 times faster than terrestrial plants (Lam et al., 2012). Microalgae though lacking in roots,

stems and leaves, but has chlorophyll as their photosynthetic pigment. They are able convert the

major carbon source which is the atmospheric CO2, to glucose for their growth (Ho et al.,

2014a). As shown in Fig. 1, photosynthesis reaction is categorised into light dependent and light

independent stage. The light dependent is the first stage whereby microalgae capture and store

energy from sunlight, converting ADP and NADP+ into energy-carrying molecules ATP and

NADPH. In the second stage of photosynthesis, which is the light independent reaction,

microalgae will capture CO2 producing organic compounds by Calvin-Benson cycle, with the

previously generated ATP and NADPH molecules (Zhao & Su, 2014). The high CO2

bioconversion efficiency and high growth rate of microalgae, allowing it to become the agent of

carbon sequestration.

3. Perspectives of carbon dioxide capture for microalgae cultivation

4
 Carbon dioxide is readily available in the atmosphere in concentration of 0.03-0.06%

(v/v). Another elevated source of CO2 is originated from zero cost flue gas, which may give 6-

15% (v/v) of CO2 (Rahaman et al., 2011). These are the major sources of CO2 used for

microalgae cultivation, so as to continuously growing microalgae, for carbon sink production.

Fig. 2 shows the relationship between microalgal-CO2 sequestration and biomass production. The

CO2 fixation and biomass production vary distinctly depending on the characteristics of

microalgae species, effects of physicochemical process and effects of cultivation systems. The

following sections summaries the effects of microalgal species and growth conditions in relation

to microalgal biomass production and bioconversion efficiency.

3.1 Biomass production – Effects of microalgal species

Biomass production is the most significant indicator to observe when research into CO2

bio-sequestration by microalgae cultivation. The selection of appropriate microalgae species for

microalgae cultivation, determines the success in CO2 bioconversion for biomass production.

The ideal algal species should have high sinking capacity, high tolerance to CO2 concentration,

toxic pollutants concentrations, temperature, nutrients limitation and pH effect. As stated by

Singh and Ahluwalia (2013), microalgae species Scenedesmus obliquus, Botryococcus braunii,

Chlorella vulgaris and Nannochloropsis oculata are the most promising species for carbon

sequestration as well as for biofuel production. Chlorella sp. was identified as the species which

possesses higher biomass production up to 1.06 g L-1 d-1 as compared to Cyanophytes and

Chrysophyte species (Zhao & Su, 2014). As shown in Table 2, Chlorella sp., Scenedesmus sp.

and Spirulina sp. (Table 2, no.5-20, 23-26 & 27-35) possess high biomass yield. It should be

noted that the high performance of mentioned microalgae is obtained under different

experimental conditions, such as light intensity, CO2 concentration and photobioreactor design.

5
This variation may affect the microalgal bio-fixation efficiency and biomass productions. Table

2 summarises literature survey on microalgal species that have been discussed in latest research

on CO2 bioconversion and biomass production, rather than strict comparison on performance

among these species. In exact, the comparative study among microalgae species is still lacking to

date, due to the variation in parameters and cultivation conditions. The indication of best

microalgae species is obtained by considering the tolerance level of the microalgae. For instance,

a study reported that Scenedesmus dimorphus exhibited biomass of 4.51 and 3.82 g L-1

respectively under 10% (v/v) and 20% (v/v) of CO2, but achieved maximum biomass of 5.17 g L-
1
with 2% (v/v) CO2 (Jiang et al., 2013). This indicated that Scenedesmus sp. has high tolerance

level against CO2 concentration, making it able to grow within the range of 10-20% (v/v)

although the best CO2 concentration is only 2% (v/v).

3.2 Biomass production – Effects of physicochemical process

3.2.1 Effects of CO2 concentrations

Apart from typical microalgae species characteristics, the microalgae biomass production

is greatly affected by cultivation conditions. Chlorella sp. was reported to be able to grow up in

40% (v/v) CO2, at pH 5.5-6.0, 30°C (Chen et al., 2014a; Rahaman et al., 2011). Conversely, Lam

et al. (2012) stated that Chlorella sp. could only grow in less than 2% (v/v) of CO2 ; further

increase of CO2 will inhibit their growth. These have shown that though the same microalgae

species may has strong adaptation to tolerate, but the growth conditions determine the efficiency

in CO2 bioconversion for biomass production. The growth rate Nannochloropsis sp. was

increased for 58%, which is from 0.33 d-1 to 0.52 d-1 under cultivation of 15% (v/v) CO2 (Jiang

et al., 2011). High concentration of CO2 promotes photosynthetic efficiency of microalgae to

reproduce within a shorter time. Nevertheless, CO2 concentration above 5% (v/v) is considered

6
as toxic to microalgae growth (Ramanan et al., 2010; Zhao & Su, 2014). Microalgae growth was

inhibited by continuous injection of increasing CO2-containing flue gas concentration to the

medium (Lee et al., 2000). It was found that this flue gas inhibition is mainly occurred due to the

presence of toxic pollutants NOx and SO2, which acidified the cultivation medium (Ho et al.,

2014b; Ho et al., 2013; Rahaman et al., 2011; Zhao & Su, 2014).

3.2.2 Effects of nutrients

Carbon, nitrogen and phosphorus are the three essential nutrients for biomass growth.

Apart from carbon which can be obtained from atmospheric air or CO2 sparging, microalgae

assimilate sufficient nitrogen and phosphorus from medium for their metabolic activities.

Nitrogen present in the form ammonium is the primary nitrogen source for microalgae

assimilation (Kumar et al., 2010; Razzak et al., 2013). Moreover, phosphorus is the element

required for photosynthesis, metabolisms, formation of DNA, ATP and cell membrane.

Phosphorus is available in the medium in the form phosphate and normally supplied in excess as

it is not readily bioavailable. Other inorganic salts and trace element like metals and vitamins are

usually added into the medium for effective photosynthetic activity.

Microalgae serve as the important agent for bioremediation due to their ability to

assimilate the organics and nutrients from wastewaters for their growth. They are the dominant

biotics used in oxidation ponds in wastewater treatment plant and sewage treatment ponds. Algal

pond could remove pollutants especially N and P, more efficiently compared to conventional

activated sewage process (Zhou et al., 2014). Chlorella vulgaris, Pseudokirchneriella

subcapitata, Synechocystis salina and Microcystis aeruginosa achieved nitrogen reduction

percentage of 100% in wastewater, under high light supply condition (Gonçalves et al., 2014).

Despite nutrient-rich wastewaters can be used as the medium, challenges are still present

7
including the C/N and N/P ratio. A suitable nutrients profile of wastewater is required for

successful microalgae cultivation for CO2 biosequestration. Typically, Chlorella sp. and

Scenedesmus sp. are the most notable microalgae in assimilating nutrients in the wastewaters,

subsequently producing high biomass yield (Bhatt et al., 2014).

3.2.3 Effects of pH

MoMicroalgae species grow well in optimal pH ranges. Synechococcus sp. and Spirulina

platensis grow at optimal pH 6.8 and pH 9, respectively, meanwhile Chlorella sp. can tolerate to

pH below 4 (Zeng et al., 2011; Zhao & Su, 2014). Flue gas usually contains high concentrations

of NOx and SO2, which reduce the pH of culture medium. Dissolved NO from NOx is considered

as an alternative nitrogen source for cell growth. It is expected that NOx in the flue gas will not

bring significant negative impact to the microalgae growth. Conversely, flue gas with SO2 above

60 ppm is inappropriate to be used for microalgae cultivation (Lam et al., 2012). With the

presence of 100-250 ppm of SO2 in flue gas, the pH may reduce to 2.5-3.5, due to the formation

of bisulfite (HSO3-), sulfite (SO32-) and sulfate (SO42-) in the medium (Lam et al., 2012; Zhao &

Su, 2014). Nevertheless, B. braunii was able to utilised S-source of bisulphite in medium with

concentration of less than 104 ppm sodium bisulphite (Van Den Hende et al., 2012). This could

be due to the high adaptatibilty of B. braunii species to grow in low pH medium.

Furthermore, if 10-20% (v/v) of CO2 from flue gas is supplied, pH of medium can be

reduced reaching 5.5 (Chen et al., 2014b; Zhao & Su, 2014). To certain extent, this can be

counterbalanced by CO2 uptake of microalgae which will undoubtedly cause pH rising. Yet,

some microalgae species are unable to withstand the acidic condition resulted from carbonic acid

formed by CO2 dissolution in medium (Lam & Lee, 2011). Sodium hydroxide and calcium

8
carbonate are usually used to adjust pH reaching its optimal range, aiming to provide excellent

CO2 bioconversion and biomass production (Rahaman et al., 2011).

3.2.4 Effects of temperature

The optimal temperature for microalgae growth is ranging from 15 till 26 °C (Zhao & Su,

2014). Low temperature of culture medium is unfavourable for the enzyme activity of ribulose

bisphosphate carboxylase oxygenase (RuBisCo), leading to reduction in photosynthesis rate. In

contrast, high temperature inhibits microalgal metabolic rate and reduces the CO2 solubility

(Rahaman et al., 2011; Zhao & Su, 2014). Low CO2 solubility causes photorespiration whereby

the RuBisCo enzyme will bind with O2 rather than CO2, in consequence reducing carbon

bioconversion rate by 20-30% (Zeng et al., 2011). The deviation of culture temperature from

optimum range has resulted in lengthening of log phase. Flue gas in high temperature after the

cooling and scrubber treatment is usually found to be suitable for cultivating thermo-tolerant

microalgae like Chlorella sp. T-1 and Chlorella KR-1. They were able to grow at temperature of

35°C and 40°C, respectively (Zhao & Su, 2014). Chlorella sp. can survive and grow in hot

springs at temperature 42°C with more than 40% (v/v) CO2 (Razzak et al., 2013).

3.2.5 Effects of light intensity

The optimal light intensities used for microalgae cultivation is ranging from 10 to 30

mmol m2 s-1 (Razzak et al., 2013). Additionally, the metabolic rate of phototrophic microalgal

cells can be enhanced by increasing the light intensity to 400 mmol m2 s-1 (Zeng et al., 2011).

Algae with light-harvesting component (phycobilisomes) and most dinoflagellates prefer low (10

mmol m2 s-1) and high light intensities (60-100 mmol m2 s-1), respectively (Razzak et al., 2013).

Chlorella and Scenedesmus sp. were grown under light intensity of 200 mmol m2 s-1. Light

intensity is essential to be delivered to all the cells within the culture. High light intensity permits

9
light penetration to the high-density culture, allowing high photosynthetic rate due to less cell

damage of photosynthetic components (photoinhibition) as the light-harvesting components

absorb less light (Fernandes et al., 2014; Zeng et al., 2011). Conversely, excessive light intensity

will cause photoinhibition. The highest photosynthetic efficiency is usually achieved by low and

medium light intensity incorporated with appropriate temperature at a particular culturing stage.

3.3 Biomass production – Effects of cultivation strategies

Microalgae cultivation is to date, applying open pond systems and closed systems. The

cultivation system is designed according to the principle of achieving high surface area per

volume ratio, so as to give more surface area for the light penetration and CO2 gaseous transfer.

The design, scale up and the operation systems considering CO2 transfer, aeration system, and

light provision are vital to maximise energy efficiency for biomass production.

3.3.1 Mixing and aeration

CO2 has low solubility in the medium, meaning the mass transfer from gaseous to liquid

phases is low. These resulted in lack of CO2 distribution in the medium, which may limit the

microalgae growth. Optimal mixing is required to enhance CO2 distribution; simultaneously O2

stripping to eliminate O2 inhibition towards photosynthesis. Various cultivation associated with

mixing strategies have applied (i) mechanical stirring systems like paddle wheel and baffles; (ii)

gas injection like bubble diffuser; (iii) membrane-sparged device in order to (i) raise the gas and

liquid interface areas; (ii) improve nutrients distribution; (iii) prevent self-shading area and

phototoxicity for high density culture; (iv) control pH by ensuring CO2 dissolution; (v) stripping

off dissolved O2 to reducing its toxicity to microalgae. With optimised aeration and stirring

system, high performance Chlorella sp. achieved CO2 bioconversion efficiency of 58%, 27%,

20% and 16% under the CO2 concentration of 2%, 5%, 10% and 15% (v/v), respectively (Zhao

10
and Su, 2014). The mixing systems though contributing in CO2 gaseous transfer suffered some

drawbacks. Drawbacks include (i) loss of CO2 to the atmosphere; (ii) bio-fouling of membrane

and diffusers; (iii) shear damage to cells; (iv) large energy input and (v) poor mass transfer to

relatively low interfacial surface area. The appropriate flow and mixing has to be determined, so

as to elevate the CO2 fixation performance by maximising the pros and minimising the cons.

3.3.2 Light provision

Effective utilisation of solar energy is vital towards economically viable microalgal

cultivation for CO2 sequestration. Practically, sunlight and artificial light are the light sources

used in current cultivation systems. The light penetration is expressed as percentage of

irradiation invading on the culture surface. There are light zone and dark zone which describe the

illuminated volume supplied with light and the non-illuminated dark volume without supporting

photosynthesis, respectively (Razzak et al., 2013). In microalgae cultivation, the light supplied

throughout the whole culturing process has to be regulated according to the changing algae

population density. Neochloris oleobundans has shown doubling of its biomass in sequential

change of light intensity comparing to a constant light supply (Pires et al., 2012). The higher the

microalgal density, the smaller the light path, leads to limitation in light penetration and cell

shading. Effective mixing can prevent cell shading, which resulted from less light supply to

certain zone within the cultivation system. Moreover, light flux can be increased with decreasing

distance from irradiated source and receiving surface, and increasing the surface area for light

absorption (Ho et al., 2011; Pires et al., 2012). Optical fibers can also be used to guide sunlight

into photobioreactor, enhancing illumination for microalgal cultivation (Pires et al., 2012).

The photoperiods of light/dark periods required are typically ranging from 12/12 to 16/8

hours. These light/dark periods are important as the photo-induced damage caused by over-

11
illumination of excessive photon flux can be repaired during the dark period. Scenedesmus

obliquus has shown high photosynthetic rate with increasing light/dark frequencies (Pires et al.

(2012). The light/dark of 10 Hz frequency cycles for Spirulina platensis and Scenedesmus

dimorphus cultivation resulted in microalgae productivity to increase by 43% and 38%,

respectively (Razzak et al., 2013). Hence, high flow rate of medium can be incorporated as this

provides shorter light/dark cycle, greater frequency fluctuations, subsequently improving light

utilisation. Additionally, microalgae do not acclimate to a definite light/dark period. This is

dependent on the nature of microalgae species, its acclimated state, frequency of changing

light/dark period, and the duration of exposure (Fernandes et al., 2014; Pires et al., 2012).

Genetic engineering can be incorporated by shortening the size of photosynthetic

antenna, therefore boosting the microalgal photosynthetic efficiency (Ho et al., 2011). Placement

of lighting device like LEDs or optical fiber inside the reactor can maximise light absorbance for

microalgae utilisation (Kumar et al., 2010). The factors to be considered in bioreactor design and

configurations include density of culture medium, optical depth of light, the amount of radiation

being absorbed or scattered through medium, mixing efficiency, diameter of vessels to obtain

increase illuminated surface area/volume of culture and integration with internal illumination.

With these, light distribution can be enhanced to improve the microalgal photosynthetic

efficiency and subsequently bringing effective CO2 sequestration.

3.4 Carbon dioxide fixation efficiency

Photosynthesis is a natural way for CO2 recycling. Annually, 500 billion tonnes of CO2 is

bio-mitigated by terrestrial vegetation worldwide yet this fixation is considered as inadequate

(Xie et al., 2014). Microalgae have greater fixation ability than terrestrial plants, by bio-

sequestration of atmospheric CO2 and CO2 from flue gas, into organic compounds. The

12
assimilation of CO2 is highly dependent on the five main factors namely (i) characteristics of the

microalgae strain; (ii) CO2 concentration; (iii) cultivation system and photobioreactor design (iv)

operating conditions (v) environmental factors (Ho et al., 2014c; Razzak et al., 2013). Biomass

measurements and growth rate evaluations are crucial tools in evaluating the efficiency of

microalgae in bio-sequestrating CO2. The latest data on CO2 fixation rate, removal percentage

and biomass yield of various microalgae species are illustrated in Table 2. It has shown that

microalgae species posed various CO2 fixation rates and removal percentages under different

CO2 concentrations and cultivation methods.

Closed photobioreactor cultivation has been studied widely due to the ease of regulating

microalgal growth parameters. Several CO2 concentrations are usually supplied to evaluate the

microalgal behavior in determining the CO2 sequestration efficiency. Some microalgae species

require high CO2 concentration in the medium, indicating their ability to bio-convert high CO2

concentration. The CO2 removal efficiency by microalgae can be determined as the difference in

CO2 concentration of the influent and effluent. This difference was due to the removal by

microalgae. The removal efficiency (%) can be determined using the following formula:

CO concentration of inluent − CO concentration of efluent

× 100%
CO concentration of inluent

Carbon dioxide fixation rate can be determined by the carbon content in the algal cell.

The CO2 fixation rate (g CO2 m-3 h-1) is estimated using the formula as follow:

MCO
RCO = C × μ
Mc
Where R CO2 and µ L are the CO2 fixation rate and the volumetric growth rate (g dry weight m-3

h-1) respectively, in the linear growth phase. MCO2 and MC represented the molar mass of CO2

and elemental carbon, respectively, CC is average carbon content of microalgal cells (% w/w).

13
The average carbon content measured by an elemental analyzer (CC measured by elemental

analyzer) (CHNS-932, Leco), was 0.507 g carbon per g dry cell weight. The algal growth rate

was determined in the linear growth phase because most of the algal growth occurred during this

phase. (Anjos et al., 2013; Razzak et al., 2013).

Chlorella sp., Scenedesmus sp. and Spirulina sp. have been identified as the favourable

microalgae strains to bio-sequester CO2. This could be due to their high CO2 fixation ability,

growth rate, tolerance level towards adverse environmental effects and rich in protein,

carbohydrate and lipid. With refers to Table 2 (no. 16-20), the increase in CO2 concentration for

Chlorella sp. cultivation led to increase in CO2 fixation rate, regardless of the cultivation systems

applied which are either lab scale methods, photobioreactor or open pond system. Additionally,

Chlorella KR-1 showed good growth rate at 10-50% CO2 (v/v) (Singh et al., 2014). Ho et al.

(2011) and Li et al. (2011) (Table 2, no. 24-25) have worked on the CO2 sequestration by

Scenedesmus obliquus using glass made vessel and air lift photobioreactor respectively. The

decrease in CO2 concentration gave better growth rate and CO2 removal efficiency (Ho et al.,

2011). Conversely, this was contradicted to the results obtained from Li et al. (2011). This could

be due to the variation in cultivation system and growth conditions. The growth rate, bio-fixation

rate and removal percentage of Spirulina sp. (Table 2, no.27-35) has shown inversely

proportional to the CO2 concentration supplied (De Morais & Costa, 2007; Knudsen et al., 2009;

Ramanan et al., 2010; Sydney et al., 2010). High CO2 concentration may increase the CO2 mass

transfer; however pH reduction may also inhibit to the microalgae growth. The elevated CO2

concentration supplied may lead to the decrease of pH, subsequently causing lengthening in log

phase and reduction in growth rate (Rinanti et al., 2014).

14
 CO2 diffuses into liquid phase 104 times slower than through gaseous medium (McGinn

et al., 2011). This has caused insufficient CO2 supply to the microalgae strains for growth, as

atmospheric CO2 is only in approximately 360 ppm (Lam & Lee, 2011). Costly CO2 sparging

like air-pumping is necessary to improve the retention time of CO2 in the medium. The CO2 flow

rate of 5 L min-1 was significantly contributes to the effective CO2 distribution (Rinanti et al.,

2014). Apart from CO2 sparging, emitted flue gases can be the carbon source to grow algae.

Some researchers have found out the possibility of direct CO2 fixation of Chlorella KR-1, by

using actual flue gases generated from combustion process (Singh et al., 2014).

4. Microalgae cultivation system

Successful microalgae cultivation requires supply of adequate sunlight, CO2 and

nutrients. Comprehensive design of cultivation system is aimed to provide the optimal growth

conditions for microalgal growth, thus maximising the biomass production. The selection

microalgal cultivation systems are still depending on these factors like (i) cost (ii) CO2 capture

source (iii) type of target products and (iv) the nutrient sources. The types of microalgae

cultivation systems used at present are summarised in the following sections.

4.1 Open pond system

Open pond system is the most commonly applied for large scale microalgae cultivation

due to its low cost and ease in operation and maintenance. It is commonly used for industrial

application, to produce significant amount of products for commercial purposes at relatively low

cost. The open pond is commonly designed as 0.25 m in width, with area of 0.2-0.5 hectares for

commercial purpose microalgal productions (Zhao & Su, 2014). Open pond system possesses

high surface area per volume ratio allowing the high CO2 mitigation. Additionally, if the nutrient

sources used is wastewater, incorporated with CO2 supplied from flue gas, the open pond system

is usually applied. It provides effective wastewater treatment concurrently. Nevertheless, open

15
pond system suffered some drawbacks whereby it requires significant land area, the culture is

subjected to high risk of contamination or predators and the evaporative water lost can be

significant due to the open structure. Thus, some ponds are covered with transparent material to

promote growing period of microalgae, prevent evaporation loss and facilitate CO2 distribution.

4.1.1 Raceway ponds

Raceway pond, an open pond system looks like race track, usually shallow with 15 to 25

cm in depth. It is equipped with paddle wheel agitation, so as to ensure good circulation and

nutrients homogenisation. In addition, the flow of culture is controlled and guided with baffles

placed in the flow channel (Ho et al., 2010; Lam & Lee, 2011). This promotes the liquid velocity

of the ponds are to be operated with more than 30 cm per second (Razzak et al., 2013). Raceway

ponds are presently the most commonly used large scale cultivation system for commercial scale.

It is used for commercial culturing of Chlorella sp., Spirulina platensis, Hematococcis sp. and

Dunaliella salina. If compared to closed photobioreactor system, raceway cultivation produced

low biomass productivity of Chlorella sp. and Spirulina sp. due to carbon limitation, as only 5%

of carbon is directly transferred by atmospheric air (De Godos et al., 2014). In order to achieve

outstanding carbon sequestration, the microalgal farms can be placed surrounding the industrial

plant, so as to utilise CO2 in the flue gas efficiently (McGinn et al., 2011).

4.1.2 Multi-layer bioreactor

Multi-layer bioreactor system has been considered as feasible and cost-effective

cultivation system in treating wastewater. It comprises of several tier of tanks, arranging one on

top to the other. The pilot scale of multi-layer ponds has been achieved up to 2000 L to 40000 L

capacity (Zhou et al., 2014). It has been successfully used for microalgae cultivation in centrate

and animal manure wastewaters (Zhou et al., 2014). The medium are flowing from tank at the

16
top to the bottom tank by gravity, undergo mixing and pump up to circulate medium back to the

top tank again. Apart from sunlight radiation, artificial light can also be installed at the top of

every layer with tank, so as to ensure sufficient light supply. The benefits of the cultivation

systems include less space requirement as it arrange in tiers, easy to scale up and cost-effective,

while the limitations are similar to the other types of open pond systems.

4.2 Closed system

Closed photobioreactors (PBR) have gained much interest by researchers due to better

control of cultivation parameters and capability to satisfy carbon requirement. It was also evident

that PBR cultivation has achieved high photosynthetic efficiency and biomass productions

compared to open pond system (Chen et al., 2014a). These advantages are even more important

if the desired microalgae are used for pharmaceutical purposes or highly selective products

applications (Fernandes et al., 2014). PBRs are designed in configuration to maximise

photosynthetic efficiency and CO2 mass transfer efficiency; minimise cultivation dark zone and

power consumption (Ho et al., 2012). The PBRs used are summarised in the following sections.

4.2.1 Airlift photobioreactor

In airlift PBR, the liquid volume in the vessel is separated into two connected zones by

baffle. The liquid is moved in the circulatory flow caused by the CO2 supply at the bottom of the

reactor (Razzak et al., 2013). It gave the most fixation efficiency due to its relatively better mass

transfer and circulation (Ho et al., 2011). This provides the high cycling of medium with low

surface area exposed for light radiation, thus resulting in minimum photoinhibition. With

pressurised gas-liquid system incorporated to generate fine bubbles into the reactor, CO2

concentration can be regulated easily rather than using baffles as in open pond system.

Additionally, in optimised PBR, generated microbubbles exert higher surface area to volume

ratio, slow rising in the medium, leading to better dissolution of gas into liquid (Lam et al.,
17
2012). The microbubbles can rising gradually and collapse in the medium, rather than

macrobubbles rising rapidly and burst on the surface of medium to the atmosphere.

Airlift PBR is effectively in gas hold-up, yet it is difficult to scale up as it possess high

cell shear effect, difficult in temperature control, complex liquid flow pattern and high operation

cost. In order to address these limitations, split column airlift PBR was introduced recently, by

integrating temperature control system and light transport internally at the centre of the airlift

PBR (Fernandes et al., 2014). This central flat plate provides illuminated surface in the medium,

serves as the central baffle to prevent dark zone and functions as heat exchanger to ensure better

temperature control. Fernandes et al. (2014) reported that the biomass productivity of microalgae

cultivated in spit column airlift PBR was 15-36% higher than in conventional bubble column.

4.2.2 Tubular photobioreactor

Tubular PBR system is the most noticeable system for large scale outdoor cultivation (Ho

et al., 2011). Tubular PBRs are made up of transparent materials and placed in outdoors under

sunlight radiation. The microalgae are cultivated in the vessel, permitting the addition of air,

CO2, and nutrients into the medium; and O2 removal by reactor. The medium is circulated

through tubes and back to reservoir in high turbulent flow, by using mechanical pump. The flow

rate in the tube is ranging from 30 cm s-1 to 50 cm s-1, to ensure CO2 distribution, light/dark cycle

and prevent cell deposition (Razzak et al., 2013). A portion of microalgae is usually harvested

after it circulated through solar collection tubes. The tubes are generally 5-20 cm in diameter to

enabling sunlight penetration, thus producing higher microalgal productivities (Razzak et al.,

2013; Zhao & Su, 2014). Though it is often considered as the most suitable for microalgae

cultivation, the reactor size and length are limited to parameter control, O2 removal and CO2

depletion (Ho et al., 2011). To date, the maximum capacity it can achieve is about 20 L. Further

18
increase in concentration culture has resulted in the increase tube length and diameter. Thus, it is

difficultly to scale up and the only solution is to multiply the reactor units.

4.2.3 Flat plate photobioreactor

Flat plate PBR has large surface area per volume ratio allowing large irradiated zone, cost

effective, large volume culture and excellent biomass productivities. Flat plate PBR has achieved

short light path and steep light gradients, and can be further enhanced by addition of baffles for

aeration towards light gradient (Ho et al., 2011). The aeration rate using bubbling technique can

be expressed in gas volumetric flow rate per unit volumetric culture medium (vvm). The

optimum aeration rate of 0.023-1.000 vvm was proposed for 5% (v/v) or 10% (v/v) CO2 aeration

and 0.05 vvm is appropriate for flat-plate PBR (Zhao & Su, 2014). Flat plate PBR is scalable,

accommodating 1000 – 2000 L capacity. Limitations include difficulty in temperature control,

limited degree of growth at the near wall region and hydrodynamic stress (Razzak et al., 2013).

4.2.4 Bag photobioreactor

Bag PBR is a semi-continuous PBR, cultivating microalgae in transparent polyethylene

bags. The bags are hung and placed in the cage with multiple partitions, located under the

sunlight. The air is sparged from the bottom of the bags, together with sealing the bags in conical

shape at the bottom, to prevent settling of cells (Razzak et al., 2013). This is commonly used in

lab scale before proceeding to outdoor pilot plant.

4.2.5 Membrane photobioreactor

Membrane PBR is basically the integration of membrane into the air lift or tubular PBR

to produce fine bubbles for better CO2 mass transfer into the medium. The membrane exerts

large surface area, with microspores that will generate fine bubbles with diameter 5.5-10.1 mm

(Lam et al., 2012). An ideal membrane PBR should be (i) easy to install; (ii) good CO2

19
distribution while preventing O2 build up; (iii) membrane has to be resistance to alkaline and

acidic conditions; (iv) less membrane fouling; (v) high durability. Hollow fiber membrane was

integrated in airlift PBR to investigate the effects of membrane porosity on CO2 fixation rate. It

was observed that low porosity of membrane provides better CO2 mass transfer, resulting in

higher CO2 fixation rate (Lam et al., 2012). This is due to the generation of microbubbles by low

porosity membrane were collapse in the medium, providing better CO2 mass transfer. With

membrane system, lower CO2 gas pressures are sufficient to be implemented while producing

same biomass productivity (Merriman et al., 2014). CO2 bubbling is less effective if compared

with membrane PBR (Rahaman et al., 2011). Hydrophobic hollow fiber membrane even

contributed better CO2 transfer and exerts lesser dissolved oxygen in the medium (Merriman et

al., 2014). Yet, the dispersion of fine bubbles creating a cloudy condition inside the reactor

reduces in light penetration. The solution to address this limitation is by separating the gas and

light delivery systems into two individual systems (Lam et al., 2012; Rahaman et al., 2011).

4.2.6 Filtration photobioreactor

Filtration PBR is developed recently whereby microalgal cells are attached and grow on

the membrane in the cultivation tank. Apparently, the concept is originated by biofilm system,

allowing microalgae and bacteria to attach on growth media, entrapped by extracellular

polymeric substances excreted by themselves (Zhang et al., 2014). Filtration PBR comprises of a

transparent cultivation tank and a recycle tank, separated by a membrane between them. The

initial operation of filtration PBR requires inoculated microalgae cells suspended in the

cultivation tank. As medium feeding through the membrane, the microalgae cells are filtered and

attached on the membrane, forming biomass. Microalgal cells can be easily washed off by the

flowing liquid medium, subsequently viable cells can be harvested. The liquid medium is then

recycled from recycle tank and recirculated back to the cultivation tank continuously. This
20
system is easy to start up and enable elimination of the harvesting cost for the downstream

processing biomass applications. However, the flow rate of medium has to be controlled, so as to

create minimal disturbance to the even distribution and attachment of the microalgae. Chlorella
-2
sp. cultivated using filtration PBR showed highest productivity, 13.56 gm d-1 in 7.5% (v/v)

CO2, at 35°C (Zhang et al., 2014).

Furthermore, majority of PBR employ degasser to release dissolved oxygen (DO). In

order to ensure effective gas-liquid separation, the path to degasser is required to enable

sufficient time for the release of small bubbles from the medium (Kumar et al., 2010).

Additionally, gas bubbling in sodium sulfite solution before it return to the reactor is required, so

as to prevent DO accumulation. Rahaman et al. (2011) also reported on the utilisation of

selective polymer-membrane gas separation system to incorporate with the PBR, to capture the

atmospheric CO2. Polyethylenimine and ionic liquids which can be switched to polymer form

have shown good performance in CO2 capturing (Rahaman et al., 2011).

Overall, excellent microalgae cultivation system should be universal for cultivating

various microalgae species, able to supply permanent solar energy illumination, high CO2 mass

transfer, prevent contamination, low energy and low cost of operation. The life cycle assessment

is essential to address the energy consumption and CO2 balance for each process of the

microalgal cultivation system. Table 3 summarises the variation in properties of different

cultivation systems. Bag PBR (Table 3) seems to be the most potential cultivation system to be

applied, whereby it achieves most of the listed criteria. It needs low cost and low energy

consumption as in open pond systems, meanwhile retaining the advantages as in closed system.

Additionally, bag PBR is made up of transparent polyethylene bags, enabling the cells to obtain

light supply from all angles without any blockage that tends to cause cell shading. If to be

21
compared among the open pond systems which are more desirable for larger scale microalgal

cultivation, the multi-stage bioreactor will be the best choice. Although it involves energy

consumption for medium pumping and mixing, the land space used is very minimum compared

to other open pond systems. Moreover, unlike raceway pond, artificial lights are installed on top

of every tier of tanks allowing good light penetration into the medium. The flowing of medium

from the top tank to the lower tank by energy-free gravity force is also contributing to the CO2

gas transfer and ensures liquid medium homogenisation.

5. Inhibition of toxic pollutants in flue gas

Despite microalgae is efficient in CO2 bioconversion with cultivation strategies like

costly and energy required CO2 sparging, treated flue gas can become the potential CO2 source

for microalgal cultivation. The typical CO2 concentration in flue gas is at 15% (v/v), which is

approximately 400 times more concentrated if compared to atmospheric CO2 (Lam & Lee, 2011;

McGinn et al., 2011). This has resulted to the usage of flue gas for microalgae cultivation.

Chlorella KR-1 was observed to be successfully fixing the actual flue gas discharged from

boiler, without any treatment (Singh et al., 2014). This could be due to the characteristics of

Chlorella sp. which has strong adaptability on high temperature and low pH of flue gas (Razzak

et al., 2013; Zhao & Su, 2014). However, flue gas generated by combustion process of industrial

plants generally contains inhibitory toxic pollutants SOx and NOx, which may affect most of the

microalgal growth. The following sections describe the inhibition of these pollutants towards

microalgae growth.

5.1 Effects of SO2

Flue gas typically comprise of 9.5-16.5% (v/v) CO2, 2-6.5% (v/v) O2, CO, 100-300 ppm

NOx, 280-320 ppm SOX, heavy metals and particulate matter (Lam et al., 2012; Lee et al., 2000).

22
SO2 derived from SOx, hydrolysed in water forming hydrogen ions which tend to cause pH

reduction of the culture medium. SO42- and HSO4- derived from SO2 hydrolysis are also the

inhibition factors affecting microalgae growth. SO2 with concentration exceeding 100 ppm and

60 ppm, respectively, inhibit the growth of almost all kinds of microalgae species (Zhao and Su,

2014; Lam et al., 2012). The growth of Chlorella KR-1 were completely suppressed and

inhibited in 150 ppm SO2 condition (Lee et al., 2000). Apart from pollutants concentration, the

inhibition effect of SOx varies greatly depending on the source of flue gas. The flue gas

generated from different industries; exert differences in toxicity level (Lam et al., 2012). In

overall, the inhibition effects are depending on the characteristics of microalgae species, growth

conditions and pollutants’ concentrations with its toxicity.

Table 4 summarises the inhibition effects of microalgae species cultivated using flue gas

with SOx and NOx compounds. The Chlorella sp. and Scenedesmus sp. listed in Table 4 (no.3-9)

have shown to be the most prominent microalgae with only slight and no growth inhibition effect

towards the CO2, SOx and NOx concentrations. The growth of Chlorella sp. MTF-15 was slightly

inhibited by more than 24% (v/v) CO2 concentration (Kao et al., 2014). This was confirmed

when the biomass production of Chlorella sp. MTF-15 was optimised when cultivated using

diluted flue gas. In overall, this has proved that Chlorella sp. and Scenedesmus sp. are indeed

efficient in mitigation of CO2, SOx and NOx pollutants from emission, provided that CO2

concentration in flue gas is not exceeding 24% (v/v). Mixed culture grown in high rate algal

pond which is used for treating diluted swine manure, shown approximately 30% increase in

biomass productions when flue gas was supplied (De Godos et al., 2010). Apparently, microalgal

species are capable in bio-fixing CO2 and remove SO2, NO2 and VOC concurrently (Ho et al.,

2011; Kao et al., 2014; Van Den Hende et al., 2012). This process could be considered as

23
phytoremediation in industrial plant with its cost-effective way of pretreating flue gas before

direct emission.

Nevertheless, there are also microalgae species especially isolates, which can withstand

the inhibition of SO2 though undergoing longer lag phase for adaptability. Or else, pH adjustment

can be made to neutralise the medium back to its optimal range. Alternately, desulfurisation unit

can be installed to remove SO2 prior to flue gas supply for cultivation. There are dry and wet

desulfurization processes, which involve uses of CaO sorbent and limestone-gypsum method,

respectively (Lee et al., 2005).

5.2 Effects of NOX

The NOx in flue gas emitted from industrial plant typically contains 5-10% (v/v) of NO2

and 90-95% (v/v) of NO (Zhao and Su, 2014). In general, microalgae can assimilate nitrogen in

the form of NO3-, NO2-, NO, N2 and NH4+ (Van Den Hende et al., 2012). The major constituent

in NOx which is NO is unlike SO2, as it hardly possesses direct impact to the microalgae growth.

Tetraselmis sp. was able to grow under flue gases containing 185 ppm SOx, 125 ppm NOx and

14.1% (v/v) CO2 (Kumar et al., 2010). It can tolerate NO in high concentration which is up to

300 ppm because NO is not directly inhibiting the microalgal growth (Kumar et al., 2010). The

growth of Chlorella sp. can be affected only with above 300 ppm of NO supply (Lee et al.,

2000). In fact, the dissolved NO serves as an alternative nitrogen source, can easily being

absorbed by culture medium for microalgae growth. Conversely, the positive effect of NO

concentration is limited, whereby the increase in NO concentration may decrease the growth rate

of most strains, but not to the extent of total inhibition.

5.3 Potential and limitation of using flue gas

24
 Capturing CO2 from flue gas has been recognised mainly due to the low atmospheric CO2

concentration. CO2 concentration in the air though has been proven possible to be captured, but

in very low concentration. However, as seen in Table 2 (no. 5), the atmospheric concentration of

0.03% (v/v) CO2 without capturing managed to achieve 92.2% removal efficiency compared to

5% (v/v) CO2-sparged conditions (Lam et al., 2012). This could be due to the low solubility of

CO2, whereby most sparged CO2 was released back to atmosphere, before even dissolved in

medium. Condition is worsening by the evaporative CO2 loss if microalgae were to be cultivated

in open pond system (Chen et al., 2014b). The utilisation of flue gas-containing CO2 provides the

solution to the mentioned problem, more importantly contribute in CO2 sequestration.

Direct utilisation of flue gas serves as the alternatives for providing carbon source for

biomass production. Nevertheless, till date, the in situ research on using flue gas from

microalgae cultivation is still very limited. Though efforts have been made, yet there are multiple

existing challenges and limitations waiting to be resolved, such as high temperature of the flue

gas and presence of toxic pollutants. Microalgae species can be screened on their resistance

towards toxic pollutants especially on site, as flue gas compositions may vary depending on the

materials feed for combustion. High CO2 concentration in flue gas reduces pH of the medium,

thus acidophilic microalgae, which are potentially effective in CO2 sequestration should have

identified, especially under high loading rates of acidic flue gas compounds (Van Den Hende et

al., 2012). Integration of desulfurisation unit can also be installed to minimise the SO2 inhibition

effect. Alternative strategy will be encouraging the usage of low sulphur fuel for small scale

industrial plants. Moreover, flue gas cooling system and gas distribution system have to be

installed aiming to supply the optimal and homogenous CO2 to microalgae.

6. Genetic engineering of microalgae species

25
 Apart from utilisation of elevated CO2 from flue gas to enhance microalgal cultivation,

genetic engineering approach can be involved to genetically modify microalgae towards

enhancing photosynthetic efficiency and CO2 fixation efficiency. There are researches developed

for transgene expression and gene knockdown for microalgae which pose greater interest in

industrial application (Zeng et al., 2011). Microalgae strains were modified by reducing the size

of light-harvesting chlorophyll antenna, so as to absorb more light for photosynthesis and reduce

photoinhibition (Lee et al., 2002; Ho et al., 2011; Zeng et al., 2011). As reported by Lee et al.

(2002), antenna-deficient strain Chlamydomonas sp. was achieved greater CO2 photoassimilation

if compared to a wild type species in a light saturated condition. Besides, the lipid synthesis gene

overexpression may improve microalgal lipid synthesis, but may cause cell division reduction

and lower biomass productions. Inducible promoter has to be activated at microalgal growth

stationary phase aiming in controlling lipid anabolism in microalgal cells (Zeng et al., 2011).

Additionally, knocking down of enzyme genes which causing lipid catabolism has been reported

as effective in increasing lipid accumulation in cells.

7. Downstream biomass applications

Successful microalgae cultivation systems, apart from CO2 sequestration, have also

contributing to microalgae biomass productions which benefiting the people. This bio-refinery

concept is introducing the application of microalgal biomass to produce biofuel, energy and

value added products like human food supplement and medical drugs (Zhou et al., 2014). The

utilisation of microalgae is mainly due to its cellular components which are rich in protein, lipids

and carbohydrates. For instances, fatty acids and microalgae extracts are used in cosmetics,

carotenoids used as dye pigments, poly-β-hydroxybutyrate used in plastics, polysaccharides used

as food thickening agent, eicosapentanoic acid used as medical drugs, Omega-3 fatty acids for

26
nutritional supplement and glycerol used in food (Kumar et al., 2010). The microalgae species

that commonly used for commercialised applications include Chlorella sp., Dunaliella salina,

Botryococcus braunii, Spirulina plantensis, Haematococcus pluvialis, Arthrospira sp. and

Nannochloropsis sp. (Ho et al., 2011). At present, thousand tons of biomass is produced yearly

for commercial application by using open pond cultivation system. Nevertheless, the application

of biomass produced by genetically modified microalgal species is greatly affected by the

purpose of application. For instances, toxicity tests are required if biomass is used to produce

cosmetic products and food supplement, but not necessarily required for biofuel production.

Biofuel is undeniably the energy product mostly associated with microalgal research.

Microalgae store significant amount of energy which can be converted into diesel, methane and

ethanol via thermal-chemical and biological methods (Ho et al., 2011). Biofuel in-used presently

are predicted to be insufficient to satisfy the world energy demand in the near future. The

diminishment of natural resources, such as petroleum and coal, is soon to be occurred (Bhatt et

al., 2014). Microalgae are known as the valuable feedstock for renewable energy production, due

to their high growth rates and high lipid productivities (McGinn et al., 2011; Rogers et al., 2014).

The cellular lipid contents of microalgae can reach up to 70% per cell (Pires et al., 2012). For

instance, Scenedesmus sp. is the promising microalgae for CO2 sequestration, bio-oil production,

carotenoids and pigments production for health food application (Ho et al., 2011). Additionally,

microalgae are also the feedstock used for bioethanol production, as their carbohydrates and

proteins serve as the carbon source for fermentation. Subsequently after the fermentation

process, the remaining microalgae biomass can be further used for bio-methane generation via

anaerobic digestion (Lam & Lee, 2011; Zhou et al., 2014).

27
 Microalgae gain elevated the CO2 supply by utilising of flue gas which could contains

significant amount of SOx and NOx pollutants. The production of biofuel using these microalgae

is allowable with no effect on combustion. Conversely, bio-refinery approach on microalgal

biomass produced from flue gas CO2 bio-conversion is still in a developing field especially for

human nutritious supplement and pharmaceutical products. Further efforts like drug toxicity test

and safety tests on animals are required to investigate the toxicity of microalgal biomass that

could be probably brought by these flue gas pollutants. Notably, the microalgal applications have

significantly improve the sustainability and economic feasibility of the microalgal cultivation

systems for effective CO2 sequestration.

8. Conclusions

Carbon sequestration through CO2 bioconversion is the natural mechanism for microalgae

cultivation. Microalgae are able to bio-convert atmospheric CO2 and in particularly CO2 from

flue gas, leading to reduction of greenhouse gas emission. This CO2 bio-mitigation will thus

minimising the greenhouse effect, leading to ecosystem preservation. Moreover, microalgae

contribute to renewable energy and valuables co-products production. Effective carbon

sequestration and bio-refinery approach can be achieved towards environmental sustainability

and economic feasibility. The innovation of microalgal-based CO2 bio-sequestration is required

to bring significant advancement in CO2 bio-mitigation aiming towards global warming solution.

Acknowledgments

This work is supported financially by SATU Joint Research Scheme (RU022E-2014) from

University of Malaya, Malaysia’s Fundamental Research Grant Scheme (FP054-2013B and

FRGS/1/2013/SG05/UNIM/02/1), Malaysia’s Ministry of Science, Technology, Innovation

(SF016-2013 and MOSTI-02-02-12-SF0256), Taiwan's Ministry of Science and Technology

28
(MOST 103-3113-E-006-006 and MOST 103-2221-E-006-190-MY3) and Taiwan's Ministry of

Education on Top University Grants.

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 Caption of Figures

Fig. 1 Light dependent and light independent stage during photosynthesis

Fig. 2 Flowchart relating microalgal-CO2 sequestration and biomass production

O2

CO2
NADPH
Sunlight
ATP
Light dependent stage
Calvin-
Photosystem I & II
Benson cycle
Electron transport
chain
ADP
Light independent
stage
Water NADP+
Glucose

Fig. 1. Light dependent and light independent stage during photosynthesis

34
 CO2 Sequestration Biomass Production

Atmospheric
Air CO2 Microalgae Microalgae biomass
• CO2 Cultivation - products
Capture Photosynthesis
• Bioenergy and
• Microalgae biofuel
species • Biogas
CO2 separation • Physiochemical • Co-products like
Flue gas and capture conditions food supplement
• Cultivation system

Toxic gas for treatment

Fig. 2. Flowchart relating microalgal-CO2 sequestration and biomass production

35
List of Tables

Table 1: The characteristics of algae groups

Table 2: Microalgae biomass yield and their CO2 fixation rate or removal percentage

Table 3: The variation in properties of different cultivation systems

Table 4: The inhibition effects of microalgae species cultivated using flue gas with

SOx and NOx compounds

Table 1
The characteristics of algae groups

Algae group Common name Characteristics

Chlorophyceae Green algae (i) Estimated 6000 – 8000 species

(ii) 90% live in freshwater rather than marine
(iii) Ranging from tiny unicellular and colonial
organisms to large macroscopic weeds
(iv) In monophyletic group as the terrestrial plants

Rhodophyceae Red algae (i) Estimated 4000 – 5000 species

(ii) 90% live in marine
(iii) Ranging from unicellular to macroscopic algae
often found on rocky shore
(iv) In monophyletic group as the terrestrial plants

Phaeophyceae Brown algae (i) Estimated 1500 – 2000 species

(ii) Almost all live in marine
(iii) Ranging from giant kelps to smaller intertidal
seaweeds
(iv) Grow in rocky intertidal zone

Cyanophyceae Blue-green (i) Prokaryotic cell

algae (ii) Present in almost all feasible habitats
(iii) CO2 and nitrogen fixers since billion years ago

Bacillariophyceae Diatoms (i) 12000 known species

(ii) Single celled with microscopic in size
(iii) Grow in seas, lakes and moist soils and out of
glass (silicon dioxide or silicon)

1
Table 2
Microalgae biomass yield and their CO2 fixation rate or removal percentage

No Microalgae species Initial CO2 CO2 bio- Removal Biomass Cultivation References
(%)(v/v) fixation rate percentage yield (g L-1) system
(g L-1 d-1) (%)

1 Anabaena sp. ~ 0.03 (Air) 1.45 - - Bubble column (Lopez et al., 2009)
2 Anabaena sp. 10 1.01 - - Bubble column (Chiang et al.,
2011)
3 Botryococcus braunii 5 0.5 - 3.11 Fermenter (Sydney et al.,
2010)
4 Botryococcus braunii 10 - 3.05 Bioreactor (Yoo et al., 2010)
5 Chlorella vulgaris 0.03 (Air) - 92.2 ~0.315 Sequential (Lam et al., 2012)
bioreactor
6 Chlorella vulgaris 0.09 (Air) 3.45 - 0.9 Membrane- (Fan et al., 2008)
sparged helical
tubular
bioreactor
7 Chlorella vulgaris 2 0.43 - 2.03 Vertical tubular (Yeh and Chang,
bioreactor 2011)
8 Chlorella vulgaris 5 0.25 - 1.94 Fermenter (Sydney et al.,
2010)
9 Chlorella vulgaris 5 - 1.5 ~0.73 Sequential (Lam et al., 2012)
bioreactor
10 Chlorella vulgaris 6 2.22 - 10.02 Glass bubble (Anjos et al., 2013)
column
11 Chlorella sp. 0.038 - 60 0.4 Lab scale (Ramkrishan et al.,

2
 photobioreactor 2014)
12 Chlorella sp. 0.106 - 80 0.7 Lab scale (Ramkrishan et al.,
photobioreactor 2014)
13 Chlorella sp. 1 - 59 - Lab scale flask (Ramanan et al.,
method 2010)
14 Chlorella sp. 2 0.857 - - Bubble column (Chiu et al., 2008)
15 Chlorella sp. 5 0.7 - 2.02 Vertical tubular (Ryu et al., 2009)
bioreactor
16 Chlorella sp. 5 - 51 - Lab scale flask (Ramanan et al.,
method 2010)
17 Chlorella sp. 10 - 46 2.25 Lab scale flask (Ramanan et al.,
method 2010)
18 Chlorella sp. 10 - 63 5.15 Air lift (Chiu et al., 2009a)
photobioreactor
19 Chlorella sp. 15 - 85.6 0.95 Sequential (Cheng et al.,
bioreactor 2013)
20 Chlorella sp. 10 - 46 2.25 Open race-way (Ramanan et al.,
pond 2010)
21 Dunaliella tertiolecta 5 0.27 - 2.15 Fermenter (Sydney et al.,
2010)
22 Nannochloropsis oculata 2-15 - 11-47 0.246-1.32 1 Cylindrical glass (Chiu et al., 2009b)
photobioreactor
23 Scenedesmus obliquus 10 - 40.2 0.653 Air lift (Li et al., 2011)
photobioreactor
24 Scenedesmus obliquus 10 0.55 - 3.51 Glass-made (Ho et al., 2010)
vessel
25 Scenedesmus obliquus 20 0.39 - 2.63 Glass-made (Ho et al., 2010)
vessel
26 Geneticly modified Scenedesmus 20 - 61.8 0.948 Air lift (Li et al., 2011)
obliquus photobioreactor

3
27 Spirulina platensis 1 - 53 - Lab scale flask (Ramanan et al.,
method 2010)
28 Spirulina platensis 5 - 41 - Lab scale flask (Ramanan et al.,
method 2010)
29 Spirulina platensis 5 0.32 - 2.18 Fermenter (Sydney et al.,
2010)
30 Spirulina platensis 10 - 39 2.91 Lab scale flask (Ramanan et al.,
method 2010)
31 Spirulina platensis 15 0.92 - 2.13 Hollow fiber (Knudsen et al.,
membrane 2009)
photobioreactor
32 Spirulina sp. 6 - 53.29 3.40 Serial tubular (De Morais &
photobioreactor Costa, 2007)
33 Spirulina sp. 12 - 45.61 3.50 Serial tubular (De Morais &
photobioreactor Costa, 2007)
34 Spirulina obliquus 6 - 28.08 1.58 Serial tubular (De Morais &
photobioreactor Costa, 2007)
35 Spirulina obliquus 12 - 13.56 1.60 Serial tubular (De Morais &
photobioreactor Costa, 2007)

36 Mixed culture of Chlorella 5 0.98 59.80 4.90 Vertical (Rinanti et al.,

sp., Scenedesmus sp. and photobioreactor 2014)
Ankistrodesmus sp.
37 Mixed culture of Chlorella sp., 10 0.85 63.10 5.80 Vertical (Rinanti et al.,
Scenedesmus sp. and photobioreactor 2014)
Ankistrodesmus sp.
1
Data for 2% (v/v) CO2 concentration, growth inhibited for CO2 concentration of higher than 2% (v/v)

4
 Table 3
The variation in properties of different cultivation systems

Cultivation system Cost Scale-up Energy Space Gas Light Growth

consumption transfer efficiency rate

Raceway pond Low Easy Low High Fair Low Low

Multi-layer Middle Easy Low Low Fair Middle High
bioreactor
Airlift High Difficult Middle Low Good High High
photobioreactor
Tubular High Difficult High Low Good High High
photobioreactor
Flat plate High Middle Middle Low Good High High
photobioreactor
Bag photobioreactor Low Middle Middle Low Good High High
Membrane High Difficult Middle Low Good High High
photobioreactor
Filtration High Middle Middle Low Good High High
photobioreactor

5
Table 4
The inhibition effects of microalgae species cultivated using flue gas with SOx and NOx compounds

No Microalgal species CO2 NOx SOx Source Inhibitory Cultivation References

% (ppm) (ppm) effect system

(v/v)

1 Nannochloropsis 10 - 25 Real flue gas from rice Inhibited Bubble (Ronda et al.,
limnetica husk emission column 2014)

2 Nannochloropsis 3 - 11 Real flue gas from rice Inhibited Bubble (Ronda et al.,
limnetica husk emission column 2014)

3 Chlorella sp. 8-10.2 38 3.8 Real flue gas from co- No Bubble (Kaštanek et
generator units inhibition column al., 2010)

4 Chlorella sp. 6-8 37 - Real flue gas from No Open thin (Doucha et al.,
combustion of natural inhibition layer PBR 2005)
gas from boiler

5 Chlorella sp. 23 78 87 Real flue gas from coke No Double set (Chiu et al.,
oven of steel plant inhibition PBR 2011)

6
6 Chlorella sp.MTF-15 25 70-80 80-90 Real flue gas from coke Slight Column- (Kao et al.,
oven of steel plant inhibition type glass- 2014)
fabricated
PBR

7 Chlorella sp. MTF-15 26 8-10 15-20 Real flue gas from hot Slight Column- (Kao et al.,
stove of steel plant inhibition type glass- 2014)
fabricated
PBR

8 Chlorella sp. MTF-15 24 25-30 15-20 Real flue gas from Slight Column- (Kao et al.,
power plant of steel inhibition type glass- 2014)
plant fabricated
PBR

9 Scenesdemus sp. 18 150 200 Real flue gas from No Airlift (Li et al.,
combustion chamber of inhibition 2011)
coke oven

10 Mixed culture of 7.5 77 - Real flue gas from No High rate (De godos, et
Scenesdemus sp., combustion of natural inhibition algal pond al., 2010)
Chlorella sp., Nitzschia gas in manure-drying
sp., Chamydomonas sp., motors
Oocystis sp. &
Protoderma sp.

7
Highlights

• Microalgae are the agent for carbon biosequestration

• CO2 from flue gas is potential carbon source for microalgae cultivation

• The effect of inhibition derived by toxic pollutants in flue gas toward microalgae.

• The advantages and disadvantages of microalgae cultivation system

44