Bioresource Technology 145 (2013) 307312

Contents lists available at SciVerse ScienceDirect

Bioresource Technology
journal homepage: www.elsevier.com/locate/biortech

Engineering strategies for simultaneous enhancement of C-phycocyanin

production and CO2 xation with Spirulina platensis
Chun-Yen Chen a,, Pei-Chun Kao b, Chia-Jung Tsai b, Duu-Jong Lee c, Jo-Shu Chang a,b,d,1
a
University Center for Bioscience and Biotechnology, National Cheng Kung University, Tainan, Taiwan
b
Department of Chemical Engineering, National Cheng Kung University, Tainan, Taiwan
c
Department of Chemical Engineering, National Taiwan University, Taipei, Taiwan
d
Research Center for Energy Technology and Strategy, National Cheng Kung University, Tainan, Taiwan

h i g h l i g h t s

" Indigenous Spirulina platensis shows high potential as an C-phycocyanin producer.

" Using the at-type PBR, the S. platensis biomass production markedly enhanced.
" Optimal illumination condition and medium composition were identied to enhance CO2 xation and C-PC production.

a r t i c l e i n f o a b s t r a c t

Article history: Spirulina platensis produces nutraceutical product C-phycocyanin (C-PC) and simultaneously mitigates
Available online 18 January 2013 CO2 emissions during its growth. Using a designed at-type photobioreactor, the S. platensis biomass pro-
duction was markedly enhanced, leading to a CO2 removal rate and a biomass concentration of 0.23 g/L/d
Keywords: and 2.25 g/L, respectively. The cell growth, CO2 xation rate and C-PC production of S. platensis were
Spirulina platensis investigated when it was cultivated under different irradiation conditions. As the light intensity increased
C-phycocyanin from 100 to 700 lmol/m2/s, the overall biomass productivity, CO2 consumption rate and maximal C-PC
Photobioreactor
productivity increased signicantly to 0.74, 1.53 and 0.11 g/L/d, respectively. After determining the suit-
Light intensity
CO2 xation
able light intensity, the nitrogen concentration was also adjusted to further enhance the performance of
CO2 xation and C-PC production. The results show that with an optimal nitrogen concentration of
0.045 M, the CO2 consumption rate and maximal C-PC productivity were further increased to 1.58 and
0.13 g/L/d, respectively.
2013 Elsevier Ltd. All rights reserved.

1. Introduction achieved by the process of photosynthesis that is carried out by

Climate change is an increasing concern, and CO2 emissions
associated with human energy consumption is one of the most sig-
nicant sources of this greenhouse gas (Chen et al., 2011; Matondo
et al., 2004). Based on Antarctica ice core data, there was only a
slightly change in atmospheric CO2, from 280 to 295 ppmv, over
the thousand years before the start of 20th century. However,
atmospheric CO2 levels increased to 315 ppmv in 1958 and
377 ppmv in 2004, as detected in Hawaii (Song, 2006). Biological
ways to reduce CO2 levels by converting it into organic compounds
have thus attracted considerable attention, and this can be
Corresponding author. Address: University Center for Bioscience and Biotech-
nology, National Cheng Kung University, Tainan 701, Taiwan.
E-mail addresses: ccy.ncku@gmail.com (C.-Y. Chen), changjs@mail.ncku.edu.tw
(J.-S. Chang).
1
Address: Department of Chemical Engineering, National Cheng Kung University,
Tainan 701, Taiwan
plants and photosynthetic microorganisms (Ho et al., 2012; Ja-
cob-Lopes et al., 2009). For example, the CO2 removal efciency
of microalgae and cyanobacteria is 10 times higher than that of ter-
restrial plants, due to their faster cell growth rate (Skjanes et al.,
2007). In addition, algae biomass can also be used as feedstock to
produce a variety of downstream products, including biofuels, car-
bohydrates, human food, animal feed, cosmetics and medicines.
There are thus a number of benets associated with microalgae-
based CO2 mitigation (Chen et al., 2010; Ho et al., 2010, 2011;
Yeh et al., 2012). Also, it is environmentally sustainable to mitigate
CO2 with microalgae when combined with other environment pro-
tection processes, such as heavy metal removal (Jacome-Pilco et al.,
2009) or wastewater treatment (Mallick, 2002). As a result, the
microalgae-based CO2 xation has raised much attention around
the world.
Spirulina are cyanobacteria which are considered a good candi-
date for bioxation of CO2, owing to its fast cell growth rate. In
addition, Spirulina platensis can grow under alkaline conditions

0960-8524/$ - see front matter 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.biortech.2013.01.054
308 C.-Y. Chen et al. / Bioresource Technology 145 (2013) 307312
(Carvalho et al., 2004), and can absorb CO2 more efciently in a cul-
turing system. Furthermore, the biomass of Spirulina is rich in C-
phycocyanin (C-PC), which is a phycobiliprotein that is generally
present in cyanobacteria, rhodophytes and cryptomonads (Glazer,
1994). Phycobiliproteins act as naturally occurring light harvesting
pigments in algae, and can efciently absorb parts of the wave-
lengths that are poorly utilized by chlorophyll (Hemlata and Fat-
ma, 2009). As a result, the existence of phycobiliproteins can
enhance the photosynthetic efciency of microalgae. In other
words, light intensity and light quality (e.g., wavelength distribu-
tion) may be critical factors that inuence the accumulation of
phycobiliproteins in microalgae (Babu et al., 1991). Among the var-
ious phycobiliproteins, C-phycocyanin is the major pigment pre-
senting in microalgal cells, while APC (allophycocyanin) is a
minor component (Moraes et al., 2010). C-phycocyanin has thus
been widely investigated with regard to its characteristics and
commercial potential. The application of C-PC has been examined
in a wide range of elds. First, C-phycocyanin has been developed
as the natural food colorant in food industries replacing the toxic
synthetic pigments due to its unique color (Leema et al., 2010). It
is also used as colorant in cosmetic products like eyeliner and lip-
stick (Sarada et al., 1999). In addition, the uorescence characteris-
tics of C-PC has been applied to uorescence microscopy,
uorescent probes, and be used as label for antibodies and recep-
tors proteins (Eriksen, 2008). Moreover, C-PC also nds its applica-
tions in pharmaceutical and cosmetic industries due to its
antioxidant, anti-inammatory and neuroprotective properties
(Romay et al., 2003). Therefore, using Spirulina sp. (such as S. plat-
ensis) to assimilate CO2 as the carbon source with simultaneous
production of valuable C-phycocyanin products gives great
benets.
There are many environmental factors that might affect cell
growth and pigment production in microalgal cultivation. During
the autotrophic growth of microalgae, light energy is utilized to
carry out photosynthesis in order to produce the energy needed
for cell growth and for xing CO2 in the Calvin cycle. Therefore,
light intensity is one of the most important factors for cell growth
and CO2 mitigation (Soletto et al., 2008). Further, the composition
of the medium is also important not only for cell growth, but also
for the cost involved in mass production of Spirulina (Raoof et al.,
2006). Nitrate is an important component in the medium, which
directly inuences the growth and composition of microalgal bio-
mass, because it is highly associated with the syntheses of protein
and chlorophyll within the microalgae (Carvalho et al., 2006). In
addition, C-phycocyanin has a secondary role as nitrogen source
in cells that are degraded, as these are exposed to nitrogen deple-
tion conditions (Boussiba and Richmond, 1980). In other words, the
content of C-phycocyanin would be strongly inuenced by the
nitrogen source concentration in the culture medium. This study
is undertaken to identify the optimal illumination condition and
medium composition to enhance biomass production, CO2 xation
ability and C-phycocyanin production simultaneously.
2. Methods
2.1. Microalgae strain and culture medium
The microalgal strain (i.e., S. platensis) used in this work was iso-
lated from a freshwater area located in southern Taiwan. The med-
ium used to cultivate the pure culture of S. platensis consisted of
(per liter): 16.8 g NaHCO3, 0.5 g K2HPO4, 2.5 g NaNO3, 1 g K2SO4,
1 g NaCl, 0.2 g MgSO42H2O, 0.04 g CaCl42H2O, 0.01 g FeSO47H2O,
0.08 g EDTA and 1 ml of trace metal solution. The trace metal solu-
tion consisted of (per liter): 2.86 g H3BO3, 1.81 g MnCl44H2O,
0.222 g ZnSO44H2O, 0.0177 g Na2MoO4, 0.079 g CuSO45H2O. The
S. platensis culture was grown at 28 C under a light intensity of
approximately 100 lmol/m2/s (illuminated by TL5).
2.2. Photobioreactor design
The photobioreactors (PBRs) used were a 1-L conventional ask
photobioreactor and a at-type photobioreactor quipped with a
novel aeration diffuser and illuminated with an external light
source (14 W TL5 tungsten lament lamps; Philips Co., Taipei, Tai-
wan) mounted on both sides of the PBR. The light intensity on the
vessel wall of the PBRs was adjusted to ca. 50 lmol/m2/s. The pre-
cultured S. platensis were inoculated into the two PBRs with an
inoculum size of 50 mg/l. The PBRs were operated under 28 C,
pH 9.0, and an agitation rate of 300 rpm. Aeration was carried
out with 2.5% CO2 gas at the rate of 0.2 vvm. In addition, the
glass-made gas diffusers (pore size: 80100 lm) were utilized to
decrease the bubble size of CO2 to enhance the mass diffusion ef-
ciency of CO2. Gas samples were taken from sampling port with a
gas syringe at specic time intervals to measure the gas composi-
tion. The liquid sample was also collected from the sealed glass
vessel with respect to time to determine microalgae cell concentra-
tion, pH and residual NaHCO3 concentration.
2.3. Analysis
Liquid samples (5 ml each) were regularly collected from the
photobioreactor by a syringe for the determination of the cell con-
centration of the culture via optical density measurement at a
wavelength of 680 nm (i.e., OD680) using a spectrophotometer
(model U-2001, Hitachi, Tokyo, Japan). Prior to OD680 analysis,
the samples were diluted with deionized water, as the dilution fac-
tor varied depending on the cell concentration. The TOC (ELEMEN-
TAR) machine was used to determine the dissolved carbon
concentration (including organic and inorganic carbon) in the cul-
turing medium. The light intensity on the reactor wall was mea-
sured with a LI-250 light meter with a LI-200SA pyranometer
sensor (LI-COR, Inc., Lincoln, Nebraska, USA). This light meter gives
a unit of W/m2 for the measured light intensity.
2.4. Kinetic parameter calculation of CO2 xation
The gas input and gas output of the photobioreactor were de-
tected using gas chromatography (450-GC, Varian, California,
USA) to measure the CO2 content within the gas. The CO2 removal
efciency (gCO2) was determined based on Eq. (1).
gCO2 yCO2;in  yCO2;out  100%=yCO2;in 1
where yCO2,in and yCO2,out represent the CO2 content (%) of the gas
input and output, respectively. The CO2 removal rate (g CO2/L/d)
was calculated based on Eq. (2) (Yun et al., 1997).
C c  DX  M CO2
FACO2 2
M C  Dt
where CC represents the average carbon content of dry biomass de-
tected by the elemental analyzer; DX represent the variation of bio-
mass concentration within the cultivation time of Dt; and MCO2 and
MC represent the molecular weight of CO2 and elemental carbon,
respectively.
2.5. Extraction and spectroscopic measurement of phycocyanin
concentration
A xed amount (0.1 g dry weight) of the S. platensis was sus-
pended into 10 ml of 0.15 M phosphate buffer (pH = 7.0), and the
operating temperature was maintained at 4 C. After operating
for 1521 h, 100% recovery of C-PC extraction could be achieved.
 C.-Y. Chen et al. / Bioresource Technology 145 (2013) 307312 309

The cell debris was then removed by centrifugation at 13,000 rpm, 8 1.0
and the supernatant (in blue color) was collected. The absorbance (a) Biomass production

Maximal biomass production (g/L)

Overall biomass productivity (g/L/d)

Biomass productivity
of crude extract was measured with UVVis at wavelengths of 615
0.8
and 652 nm, and C-PC concentration of the extract was calculated 6
according to Eq. (3) (Patel et al., 2005).
0.6
OD615  0:474  OD652
C-PC concentration g=L 3 4
5:34
0.4

3. Results and discussion 2

0.2
3.1. Effects of photobioreactor type on microalgae growth and CO2
xation
0 0.0
16
(b) C-PC content 0.14

Maximal C-PC productivity (g/L/d)

In conventional photobioreactors (PBRs), the light intensity C-PC productivity

Maximal C-PC content (%)

tends to decrease rapidly due to the shielding effects arising from 14 0.12
increases in the microalgae biomass concentration. Due to the 12
problems and limitations associated with conventional PBRs, this 0.10
work aimed to develop a at-type PBR to improve illumination 10
0.08
area. The S. platensis was grown in a photobioreactor illuminated 8
with TL5 lament lamps at a one side light intensity of ca. 0.06
100 lmol/m2/s using CO2 as the carbon source. Table 1 presents 6
the microalgae growth and CO2 xation performance obtained 4
0.04
with different PBRs. The results show that a at-type PBR has a
0.02
higher biomass production, biomass productivity and CO2 removal 2
than those obtained from the control culture (using conventional 0 0.00
ask PBR). This might be due to the higher S/V ratio and short light 0 200 400 600 800 1000 1200 1400
path, which result in higher light penetration efciency, and thus a Light intensity (mol/m 2 /s)
high biomass concentration of S. platensis. Similar behavior was
also observed in other research, which found that a at-type reac- Fig. 1. Effects of light intensity on (a) maximal biomass production and overall
biomass productivity, (b) maximal C-PC content and C-PC productivity.
tor could reduce the light path, and thus enhance biomass produc-
tivity (Xu et al., 2009). The biomass production, overall biomass
productivity and CO2 removal rate with using a at-type photobi-
oreactor increased signicantly, up to about 2.1 times more than
that seen with a conventional ask PBR. Based on these results, a
2.0
at-type PBR is a more effective kind for the cultivation of S. plat-
Specific growth rate (d )
-1

ensis, and thus was used in the rest of the experiments.

1.5
3.2. Effects of light intensity on microalgae growth rate, CO2 xation
and C-PC production
1.0
The S. platensis strain was cultivated with the at-type PBR illu-
minated at different light intensities to examine the effects of this Specific growth rate
Monod model
0.5
on microalgae growth, CO2 xation and C-PC production perfor- Haldane model
mance. During the cultivation, the temperature were controlled
at 30 C and all the pH values were maintained at 9.0. The results
0.0
show that the maximum biomass production and biomass produc- 0 200 400 600 800 1000 1200 1400
tivity increased rapidly with increasing light intensity, from 100 to 2
One-side light intensity (mol/m /s)
700 lmol/m2/s, while they remained nearly unchanged when the
light intensity was higher than 900 lmol/m2/s (Fig. 1a). As the light Fig. 2. Light intensity-dependent growth kinetics and model simulations.
intensity increased to 1300 lmol/m2/s, a longer lag phase was ob-
served in the initial period of the growth curve (data not shown),
and this may be because excessive illumination would inhibit the
Table 2
biomass production and CO2 xation efciency (Soletto et al., Estimated kinetic parameters of Monod-type model and Haldane model based on the
2008). effect of light intensity on the growth of Spirulina platensis.

Model Estimated parameters

Table 1 lmax Ks KI R2
The growth and CO2 xation performance of Spirulina platensis cultivated in different
Monod 1.83 72.97 N.A 0.916
photobioreactors.
Haldane 4.31 387.4 820.8 0.982
Photobioreactor Biomass Biomass Average Average
type production productivity gCO2 (%) FACO2 (g/
(g/L) (g/L/d) L/d)
Fig. 1b depicts the effects of light intensity on C-PC production
Conventional 1.09 0.10 0.07 0.01 1.94 0.09 0.12 0.01 and the C-PC content of S. platensis. The cumulative C-PC produc-
ask type
tivity increased with increasing light intensity from 100 to
Flat type 2.25 0.283 0.14 0.02 2.74 0.23 0.23 0.03
700 lmol/m2/s (Fig. 1b). The maximal C-PC productivity reached
310 C.-Y. Chen et al. / Bioresource Technology 145 (2013) 307312

2.0 14 0.14
7
12 0.12

Biomass concentration (g/L)

Nitrate concentration (g/L)

C-PC productivity (g/L/d)

1.6 6

C-PC content (%)

10 0.10
5
1.2
8 0.08
4 C-PC productivity
C-PC content
Biomass 6 0.06
0.8 3
Nitrate

2 4 0.04
0.4
1 2 0.02

0.0 0 0 0.00
0 2 4 6 8 10
Time (d)

Fig. 3. Timecourse proles of cell growth, nitrogen source consumption, and C-PC production with Spirulina platensis.

0.11 g/L/d at 700 lmol/m2/s, while further increases in light inten-

sity did not lead to better C-PC production performance, and the 14 1.0
(a)

Overall biomass productivity (g/L/d)

Biomass production

Maximal biomass production (g/L)

cell growth rate slightly decreased when the culture was illumi- Biomass productivity
12
nated at high light intensities (Figs. 1a and 2). The CO2 consump- 0.8
tion rate (obtained when maximal C-PC productivity was
10
reached) increased from 0.23 to 1.53 g/L/d as the light intensity
was increased from 100 to 1100 lmol/m2/s. The trends of these re- 8
0.6
sults are quite reasonable, as a higher light intensity usually leads
to higher cell growth rate, which also increases the nitrogen con- 6
0.4
sumption and CO2 xation rates. As indicated in Table 2, the
Monod model and Haldene model were used to t specic growth 4
rates at different light intensities. The Haldane model seems to t 0.2
2
the experimental data with much better agreement (r2 = 0.982)
(Fig. 2), indicating that light intensity inhibition occurred. The val-
0 0.0
ues of the light intensity-based growth kinetic constants (lmax, Ks 16 (b) C-PC content 0.16

Maximal C-PC productivity (g/L/d)

and KI) according to Haldane model were 4.31 d1, 387.4 lmol/m2/ C-PC productivity
Maximal C-PC content (%)

14 0.14
s and 820.8 lmol/m2/s respectively (Table 2). The high KI value
(820.8 mg/l) indicates that S. platensis could tolerate the inhibition 12 0.12
effects of higher light intensity.
10 0.10

3.3. Effects of nitrogen concentration on microalgae growth rate, CO2 8 0.08

xation and C-PC production 6 0.06

Fig. 3 shows that the maximal C-PC content usually appeared 4 0.04

just before nitrogen starvation occurred, while the C-PC content 2 0.02
dramatically decreased after the nitrate was exhausted (Eriksen,
2008). Therefore, with the same initial nitrogen concentration, 0 0.00
0.02 0.04 0.06 0.08 0.10
the C-PC productivity would be higher if nitrogen starvation occurs
earlier. This could be the reason why the C-PC productivity in- Initial nitrate concentration (M)
creased initially with an increase in light intensity, because the cell Fig. 4. The effect of initial nitrogen source (NaNO3) concentration on (a) the
growth and nitrogen consumption rates both rose along with the maximal biomass production titer and the overall biomass productivity; (b) the
light intensity, until the latter increased beyond the inhibitory le- maximal C-PC content and C-PC productivity.
vel. These results indicate that illuminating at 700 lmol/m2/s
could lead to higher biomass productivity, C-PC productivity and
CO2 xation rate. Therefore, the following experiments exploring
the effects of carbon and nitrogen concentration on the perfor-
mance of cell growth and C-PC production with S. platensis were Table 3
conducted at a light intensity of 700 lmol/m2/s. Comparison of C-PC content and productivity obtained from this study with those
The literature shows that the nitrogen source plays key roles in reported in the literature.

microalgae growth, CO2 removal rate and C-PC productivity (Jin Microalgae C-PC content C-PC References
et al., 2006). In particular, the accumulation of C-PC appears to strain (%) productivity
be nitrogen concentration dependent. As indicated in Fig. 3, the (g/L/d)

C-PC content reached its maximum level just before the depletion Synechococcus sp. 18.5 0.024 Takano et al. (1995)
of the nitrogen source, and started to decompose under nitrogen Spirulina platensis 10.7 0.087 Chen and Zhang (1997)
Arthrospira platensis 4.8 0.014 Leema et al. (2010)
decient conditions, as at this point the cyanobacteria might begin
Spirulina platensis 16.8 0.068 Zeng et al. (2012)
to utilize phycobiliproteins as a nitrogen source (Eriksen, 2008). Spirulina platensis 12.6 0.125 This study
Therefore, the initial concentration of nitrogen that signicantly af-
 C.-Y. Chen et al. / Bioresource Technology 145 (2013) 307312 311

Table 4
Comparison of CO2 consumption rate obtained from this study with those reported in the literature.

Microalgae strain CO2 content in the feed Biomass productivity (mg/ CO2 consumption rate (mg/ References
(%) L/d) L/d)
Spirulina platensis 5 N.D 318.2 Sydney et al. (2010)
Chlorella vulgaris 5 N.D 251.6 Sydney et al. (2010)
Botryococcus braunii 5 N.D 497.0 Sydney et al. (2010)
Dunaliella tertiolecta 5 N.D 272.4 Sydney et al. (2010)
Chlorella vulgaris 15 228 460.8 Jin et al. (2006)
Scenedesmus sp. 15 304 612 Jin et al. (2006)
Microcystis ichthyoblabe 15 258 520.8 Jin et al. (2006)
Microcystis aerusinosa 15 243 489.6 Jin et al. (2006)
Thermosynechococcus 20 90 170 Eberly and Ely (2012)
elongatus
Aphanothece microscopica 15 770 1440 Jacob-Lopes et al.
Nageli (2009)
Spirulina platensis 2.5 995 1650 This study

fects the biomass production performance and the cultivation time
required to reach nitrogen starvation should be adjusted to achieve
the highest C-PC productivity. In addition, the timing of collecting
microalgae cells with the highest C-PC content is also crucial for
better C-PC production. For this purpose, S. platensis was grown
at a NaNO3 concentration range of 0.030.09 M to examine the ef-
fects of this on microalgae growth and C-PC production using a
at-type PBR illuminated externally with TL5 lamps.
The results show that the overall biomass productivity in-
creased from 0.73 g/L to a peak value of 0.82 g/L at a NaNO3 con-
centration of 0.045 M, while it decreased slightly when the
NaNO3 concentration was higher than 0.045 M (Fig. 4a). Mean-
while, the maximal biomass production reached around 10 g/L
after increasing the nitrate concentration from 0.03 to 0.045 M,
and remained at a similar level as the nitrogen source concentra-
tion was further increased from 0.045 to 0.09 M (Fig. 4a). This indi-
cates that a sufcient nitrogen supply is necessary for microalgae
biomass production, but its enhancing effect on the biomass pro-
ductivity is insignicant when the NaNO3 concentration is higher
than 0.045 M. The C-PC productivity reached a maximum value
of 0.126 g/L/d at a NaNO3 concentration of 0.045 M (Fig. 4b).
Although the C-PC content obtained from this study was not the
highest among the values reported in the literatures, the attained
C-PC productivity is higher than most reported values thanks to
the high biomass productivity achieved by using the proposed cul-
tivation system (Table 3). Meanwhile, the CO2 consumption rate
obtained from this work reached 1680 mg/L/d, which is also supe-
rior to most of the reported values (Table 4).
The maximum C-PC content increased from 10% to nearly 12%
as the initial nitrogen source concentration rose from 0.03 to
0.045 M, while further increasing the NaNO3 concentration did
not lead to higher C-PC content, nor did it enhance C-PC productiv-
ity. The reason for this might be because when the nitrogen source
concentration was too high, a longer cultivation time was required
to reach nitrogen depletion, and a lower NaNO3 conversion rate
was achieved. This results in lower overall biomass productivity,
as well as lower C-PC productivity. Furthermore, using a nitrate
concentration of 0.045 M also led to the highest CO2 consumption
rate (FACO2), calculated at the culture time when the C-PC produc-
tivity reached its maximum level. This is because CO2 consump-
tion rate and biomass production rate are highly correlated, and
it also indicates that the maximum C-PC productivity occurred
when the biomass productivity and CO2 xation rate all reached
their highest levels. Therefore, the timing for harvesting the micro-
algae biomass with the highest C-PC production performance is
quite important for the commercial production of C-PC. The results
show that the optimal harvest timing is highly dependent on the
level of cell growth and the level of residual nitrogen
concentration.
4. Conclusions
This work demonstrates that C-phycocyanin production and
CO2 xation of S. platensis were signicantly enhanced by using a
at-type photobioreactor equipped with a novel aeration device.
The results show that the best C-PC production performance was
obtained when the microalgae were grown under a light intensity
of 700 lmol/m2/s and an initial NaNO3 concentration of 0.045 M.
By using the at-type PBR and the optimal operation conditions
identied, the highest CO2 consumption rate and C-PC productivity
obtained were 1.58 and 0.126 g/L/d, respectively, which are 5.9-
and 5.3-fold improvement over those obtained from the conven-
tional PBR.
Acknowledgements
The authors gratefully acknowledge nancial support from Tai-
wans National Science Council under grant numbers NSC 101-
2221-E-006-209-MY3, NSC 101-3113-E-006-015 and NSC 101-
2221-E-006-094-MY3. This research was also supported in part
by funding from the Headquarters of University Advancement at
the National Cheng Kung University, which is sponsored by the
Ministry of Education, Taiwan, ROC.
References
Babu, T.S., Kumar, A., Varma, A.K., 1991. Effect of light quality on phycobilisome
components of the cyanobacterium Spirulina platensis. Plant Physiol. 95 (2),
492497.
Boussiba, S., Richmond, A.E., 1980. C-phycocyanin as a storage protein in the blue-
green-alga Spirulina platensis. Arch. Microbiol. 125 (12), 143147.
Carvalho, A.P., Pontes, I., Gaspar, H., Malcata, F.X., 2006. Metabolic relationships
between macro- and micronutrients, and the eicosapentaenoic acid and
docosahexaenoic acid contents of Pavlova lutheri. Enzyme Microb. Technol. 38
(34), 358366.
Carvalho, J.C.M., Francisco, F.R., Almeida, K.A., Sato, S., Converti, A., 2004. Cultivation
of Arthrospira (Spirulina) platensis (Cyanophyceae) by fed-batch addition of
ammonium chloride at exponentially increasing feeding rates. J. Phycol. 40 (3),
589597.
Chen, C.Y., Yeh, K.L., Aisyah, R., Lee, D.J., Chang, J.S., 2011. Cultivation,
photobioreactor design and harvesting of microalgae for biodiesel production:
a critical review. Bioresour. Technol. 102 (1), 7181.
Chen, C.Y., Yeh, K.L., Su, H.M., Lo, Y.C., Chen, W.M., Chang, J.S., 2010. Strategies to
enhance cell growth and achieve high-level oil production of a Chlorella vulgaris
isolate. Biotechnol. Prog. 26 (3), 679686.
Chen, F., Zhang, Y.M., 1997. High cell density mixotrophic culture of Spirulina
platensis on glucose for phycocyanin production using a fed-batch system.
Enzyme Microb. Technol. 20 (3), 221224.
Eberly, J., Ely, R., 2012. Photosynthetic accumulation of carbon storage compounds
under CO2 enrichment by the thermophilic cyanobacterium
Thermosynechococcus elongatus. J. Ind. Microbiol. Biotechnol. 39 (6), 843850.
Eriksen, N.T., 2008. Production of phycocyanin a pigment with applications in
biology, biotechnology, foods and medicine. Appl. Microbiol. Biotechnol. 80 (1),
114.
Glazer, A.N., 1994. Phycobiliproteins a family of valuable, widely used
uorophores. J. Appl. Phycol. 6 (2), 105112.
312 C.-Y. Chen et al. / Bioresource Technology 145 (2013) 307312
Hemlata, Fatma, T., 2009. Screening of cyanobacteria for phycobiliproteins and
effect of different environmental stress on its yield. Bull. Environ. Contam.
Toxicol. 83 (4), 509515.
Ho, S.-H., Chen, C.-Y., Chang, J.-S., 2012. Effect of light intensity and nitrogen
starvation on CO2 xation and lipid/carbohydrate production of an indigenous
microalga Scenedesmus obliquus CNW-N. Bioresour. Technol. 113, 244252.
Ho, S.-H., Chen, C.-Y., Lee, D.-J., Chang, J.-S., 2011. Perspectives on microalgal CO2-
emission mitigation systems a review. Biotechnol. Adv. 29, 189198.
Ho, S.H., Chen, W.M., Chang, J.S., 2010. Scenedesmus obliquus CNW-N as a potential
candidate for CO2 mitigation and biodiesel production. Bioresour. Technol. 101
(22), 87258730.
Jacob-Lopes, E., Revah, S., Hernandez, S., Shirai, K., Franco, T.T., 2009. Development
of operational strategies to remove carbon dioxide in photobioreactors. Chem.
Eng. J. 153 (13), 120126.
Jacome-Pilco, C.R., Cristiani-Urbina, E., Flores-Cotera, L.B., Velasco-Garcia, R., Ponce-
Noyola, T., Canizares-Villanueva, R.O., 2009. Continuous Cr(VI) removal by
Scenedesmus incrassatulus in an airlift photobioreactor. Bioresour. Technol. 100
(8), 23882391.
Jin, H.F., Lim, B.R., Lee, K., 2006. Inuence of nitrate feeding on carbon dioxide
xation by microalgae. J. Environ. Sci. Health, Part A 41 (12), 28132824.
Leema, J.T.M., Kirubagaran, R., Vinithkumar, N.V., Dheenan, P.S., Karthikayulu, S.,
2010. High value pigment production from Arthrospira (Spirulina) platensis
cultured in seawater. Bioresour. Technol. 101 (23), 92219227.
Mallick, N., 2002. Biotechnological potential of immobilized algae for wastewater N,
P and metal removal: a review. Biometals 15 (4), 377390.
Matondo, J.I., Peter, G., Msibi, K.M., 2004. Evaluation of the impact of climate change
on hydrology and water resources in Swaziland: Part II. Phys. Chem. Earth 29,
11931202.
Moraes, C.C., Burkert, J.F.D., Kalil, S.J., 2010. C-phycocyanin extraction process for
large-scale use. J. Food Biochem. 34, 133148.
Patel, A., Mishra, S., Pawar, R., Ghosh, P.K., 2005. Purication and characterization of
C-phycocyanin from cyanobacterial species of marine and freshwater habitat.
Protein Expression Purif. 40 (2), 248255.
Raoof, B., Kaushik, B.D., Prasanna, R., 2006. Formulation of a low-cost medium for
mass production of Spirulina. Biomass Bioenergy 30 (6), 537542.
Romay, C., Gonzalez, R., Ledon, N., Remirez, D., Rimbau, V., 2003. C-phycocyanin: a
biliprotein with antioxidant, anti-inammatory and neuroprotective effects.
Curr. Protein Pept. Sci. 4 (3), 207216.
Sarada, R., Pillai, M.G., Ravishankar, G.A., 1999. Phycocyanin from Spirulina sp.:
inuence of processing of biomass on phycocyanin yield, analysis of efcacy of
extraction methods and stability studies on phycocyanin. Process Biochem. 34
(8), 795801.
Skjanes, K., Lindblad, P., Muller, J., 2007. BioCO2 a multidisciplinary, biological
approach using solar energy to capture CO2 while producing H-2 and high value
products. Biomol. Eng. 24 (4), 405413.
Soletto, D., Binaghi, L., Ferrari, L., Lodi, A., Carvalho, J.C.M., Zilli, M., Converti, A.,
2008. Effects of carbon dioxide feeding rate and light intensity on the fed-batch
pulse-feeding cultivation of Spirulina platensis in helical photobioreactor.
Biochem. Eng. J. 39 (2), 369375.
Song, C.S., 2006. Global challenges and strategies for control, conversion and
utilization of CO2 for sustainable development involving energy, catalysis,
adsorption and chemical processing. Catal. Today 115 (14), 232.
Sydney, E.B., Sturm, W., de Carvalho, J.C., Thomaz-Soccol, V., Larroche, C., Pandey, A.,
Soccol, C.R., 2010. Potential carbon dioxide xation by industrially important
microalgae. Bioresour. Technol. 101 (15), 58925896.
Takano, H., Arai, T., Hirano, M., Matsunaga, T., 1995. Effects of intensity and quality
of light on phycocyanin production by a marine cyanobacterium Synechococcus
sp. Nkbg-042902. Appl. Microbiol. Biotechnol. 43 (6), 10141018.
Xu, L., Weathers, P.J., Xiong, X.R., Liu, C.Z., 2009. Microalgal bioreactors: challenges
and opportunities. Eng. Life Sci. 9 (3), 178189.
Yeh, K.-L., Chen, C.-Y., Chang, J.-S., 2012. pH-stat photoheterotrophic cultivation of
indigenous Chlorella vulgaris ESP-31 for biomass and lipid production using
acetic acid as the carbon source. Biochem. Eng. J. 64, 17.
Yun, Y.S., Lee, S.B., Park, J.M., Lee, C.I., Yang, J.W., 1997. Carbon dioxide xation by
algal cultivation using wastewater nutrients. J. Chem. Technol. Biotechnol. 69
(4), 451455.
Zeng, X.H., Danquah, M.K., Zhang, S.D., Zhang, X., Wu, M.Y., Chen, X.D., Ng, I.S., Jing,
K.J., Lu, Y.H., 2012. Autotrophic cultivation of Spirulina platensis for CO2 xation
and phycocyanin production. Chem. Eng. J. 183, 192197.