Chemical Engineering Journal 252 (2014) 104111

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Chemical Engineering Journal

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Dynamic photosynthetic response of the microalga

Scenedesmus obtusiusculus to light intensity perturbations
Juan Cabello a, Marcia Morales b, Sergio Revah b,
a
b

Doctorado en Biotecnologa, Universidad Autnoma Metropolitana-Iztapalapa, San Rafael Atlixco 186, C.P. 09340 Mxico, DF, Mexico
Departamento de Procesos y Tecnologa, Universidad Autnoma Metropolitana-Cuajimalpa, Av. Vasco de Quiroga 4871, colonia Santa Fe Cuajimalpa, C.P. 05300 Mxico, DF, Mexico

h i g h l i g h t s
 The microalga Scenedesmus obtusiusculus acclimates fast to irradiance changes.
 Dynamic experiments allow rapid analysis of short-term photosynthetic response.
 Temperature variations may induce an apparent hysteresis in photosynthetic response.
 N-starved cells show slow re-directing of metabolic uxes to lipid accumulation.

a r t i c l e

i n f o

Article history:
Received 3 January 2014
Received in revised form 1 April 2014
Accepted 17 April 2014
Available online 30 April 2014
Keywords:
Photo-acclimation
Photosynthetic response
Scenedesmus
Photobioreactors
Algal oil
Dynamic model

a b s t r a c t
Unsteady state experiments in microalgal cultivation are important to evaluate the inuence of the irradiance on the dynamic photosynthetic response and to develop dynamic models for the design of photobioreactors. In this paper, the short-term effect of incident light uctuations on the oxygen production by
the microalga Scenedesmus obtusiusculus cultivated in a 20 L air-lift photobioreactor was performed at different operation times under nitrogen-replete or nitrogen-starved conditions. It was possible to reach
steady states in the oxygen production indicating short-term photosynthetic acclimation and the highest
1
for light intenvalues during the incremental light step-changes were between 103 and 207 mgO2 g1
b h
2 1
1
sities between 141 and 505 lmol m s at 0.13 gb L . The photosynthetic response was not symmetric
in the increase/decrease light step-changes due to temperature variations caused by the illumination system. Moreover, in nitrogen-starved conditions the dynamic photosynthetic response was slower than in
nitrogen-replete levels declining to 70% at 0.5 gb L1 at the maximum light intensity. Furthermore, a
mathematical model was developed to estimate the dynamic oxygen response and the biomass productivity. The simulations predicted the highest O2 concentrations at 35 C and irradiances above
600 lmol m2 s1 and the highest biomass productivity was 0.78 gb L1 d1.
2014 Elsevier B.V. All rights reserved.

1. Introduction
Microalgal cultivation is a promising technology for the biological xation of carbon dioxide and the generation of high valueadded products including renewable fuels [1]. As an example, oils
from algae can yield biodiesel through transterication and gasoline
or jet fuel through distillation and cracking. Different photobioreactor congurations can be used for cultivation, including tubular,
panel and raceway ponds. Light availability inuences decisively
the photosynthesis process, the specic growth rate and
consequently biomass productivity. Light is absorbed and scattered
by the cells [2] and therefore is spatially distributed; and most of the
Corresponding author. Tel.: +52 55 5814 6536.
E-mail address: srevah@correo.cua.uam.mx (S. Revah).
http://dx.doi.org/10.1016/j.cej.2014.04.073
1385-8947/ 2014 Elsevier B.V. All rights reserved.

cultivation systems are limited by light availability. The inuence of

light on the photosynthesis rate can be obtained from the photosynthesisirradiance, PI curve [3], which is particular for each strain
and must be measured under diluted culture to assure that cells
are exposed to the same irradiance. For spatial light distribution
most of the works consider an average irradiance dened as the
average of local irradiance values inside the culture [4] but some
others represent the radial decrease of light intensities [2,5] and
the mixing effects [6,7]. Furthermore, the knowledge of photosynthesis dynamics under uctuating light conditions and the determination of physical and biological parameters is important and
needed for: (a) the design, modeling and simulation of the process,
(b) the scale up of laboratory experiments to industrial level and (c)
the development of control systems that can be implemented for
measuring or estimating key variables in the process unit to

J. Cabello et al. / Chemical Engineering Journal 252 (2014) 104111

105

Nomenclature
Cb
CG,O2
CG,O20
CL,O2
Csat
L,O2
CL,O2,i
CCO2,s
CCO2,e
Csat
L,CO2
DO2
DCO2
Daz
dt
Ea
Ed
FG
H
H
hg
hL
I0
Iav

biomass concentration, g L1

oxygen bulk-gas concentration, mg L1
initial oxygen concentration, mg L1
oxygen bulk-liquid concentration, mg L1
saturation oxygen concentration, mg L1
oxygen concentration by each pseudo-steady state,
mg L1
CO2 concentration in gas phase at the inlet of the reactor, mg L1
CO2 concentration in gas phase at the outlet of the reactor, mg L1
saturation carbon dioxide concentration, mg L1
diffusion coefcient for oxygen, m2 s1
diffusion coefcient for carbon dioxide, m2 s1
axial dispersion coefcient, m2 s1
tube diameter, m
activation energy, kcal mol1
deactivation energy, kcal mol1
gas ow, L min1
partition coefcient from Henry law, dened by H = H/
RT, dimensionless
Henry coefcient, atm-m3 mol1
gas hold-up, m3 m3
liquid hold-up, m3 m3
incident light intensity on reactor surface, lmol m2 s1
average light intensity within the reactor, lmol m2 s1

increase growth and biomass yield [811]. Temperature is another

important environmental variable, which affects both the structure
of cell components and the reaction rates. Recently, Bchet et al.
[11] presented a complete review of the models used to describe
microalgal growth kinetics.
Photobioreactor design requires precise information on microalgal growth. Biomass evolution results from the combined and
cumulative effects of physical and biological phenomena including
photosynthesis, uid dynamics, mass transfer and irradiance
which are spatially and diachronically distributed in the photobioreactors [12,13]. The models describing growth generally lump
these effects and consequently it is not generally possible to analyze them independently. Additionally, experiments are generally
lengthy and several assays with different initial and operating conditions are required. An alternative is the determination of the oxygen production rate [3,9], which contrary to the growth rate, has a
dynamic response in the time scale of minutes. It is proportional to
the growth rate according to the stoichiometry of the overall photosynthetic reaction [3] and it provides information about the
photo-acclimation phenomena generated in the photosystem II
as a response to uctuations in the light intensity [14,15]. Furthermore, the time needed for determination of the photosynthesisirradiance curves and the associated kinetic parameters is reduced
and some models [8,9,11,1618] have been developed for predicting O2 production. However, photosynthesis rate curves under Nreplete conditions obtained in short unsteady state experiments
of incident light uctuating have not been quantitatively determined yet.
Nitrogen starvation conditions have so far been the most commonly employed approach for directing metabolic uxes to lipid
accumulation of microalgae. However, the effect of incident light
perturbations on the oxygen dynamic responses under nitrogen
N-starved conditions has not been studied. This could provide
information about the changes in the metabolic regulation, which
would be achieved by the adjustment of PSI:PSII stoichiometry and
depends on the novo protein synthesis [19].

k0, k1
Ka
Kd
KI
KLaO2
KLaCO2
Ks
L
Pb
PO2,max
PPFR
rO2,exp
rO2,intr
rCO2,exp
T
t
uGe
uLe
Yb/O2
VR
z

1
frequency factors of Arrhenius, mgO2 g1
b s
biomass light absorption coefcient, m2 g1
b
metabolic coefcient, h1
inhibition constant for light intensity, lmol m2 s1
volumetric mass transfer coefcient for oxygen, h1
volumetric mass transfer coefcient for carbon dioxide,
h1
irradiation constant, lmol m2 s1
reactor length, m
biomass productivity, gb L1 h1
1
maximum photosynthesis rate, mgO2 g1
b s
photosynthetic photon uence rate, lmol m2 s1
experimental oxygen production rate per biomass unit,
1
mgO2 g1
b h
intrinsic oxygen production rate per biomass unit,
1
mgO2 g1
b h
experimental carbon dioxide uptake rate per biomass
unit, mgCO2 gb1 h1
temperature, K
time, min
effective gas velocity, m s1
effective liquid velocity, m s1
biomass yield on oxygen, gb g1
O2
reactor volume, m3
axial direction, m

The aim of this work was to study the dynamic response of the
photosynthetic activity of the microalga Scenedesmus obstusiusculus to different light intensities in an air-lift photobioreactor under
N-replete and N-starved conditions. The rst case could be closely
related to the short-term photo-acclimation and the second case to
the long-term metabolic adaptation. Furthermore, a mathematical
model which predicts the oxygen concentration and biomass productivity using parameters obtained from experiments was validated considering the hydrodynamics and mass transfer within
the system and a kinetic expression for intrinsic oxygen production
rate representing the photosynthesis activity as function of temperature, light intensity and biomass concentration under Nreplete conditions.
2. Materials and methods
2.1. Model development
A transient one-dimensional model was used to represent oxygen concentration as overall product of the photosynthesis process
within an air-lift photobioreactor. The following assumptions were
considered:
(1) The gas and liquid-microalga phases are homogeneously
distributed inside the reactor and their respective volume
fraction is conserved, i.e. hL + hG = 1.
(2) There is a homogeneous distribution of rising bubbles
throughout the air-lift riser.
(3) The gas phase is represented by plug ow.
(4) The liquid phase is represented by an isothermal and axial
dispersion model.
(5) The physical and transport properties of the culture media
such as viscosity, density and axial dispersion coefcient
are considered similar to those of water.
(6) Light distribution is considered within a homogenous isotropic medium.

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J. Cabello et al. / Chemical Engineering Journal 252 (2014) 104111

Fig. 1. Schematic diagram of the experimental air-lift photobioreactor used for growth of the microalga S. obtusiusculus.

(7) Biomass response is based on the average light in the photobioreactor and is independent on the short light cycles found
in the air-lift conguration.
(8) The biomass concentration does not change during the
dynamic experiments.
(9) The concentrations of carbon dioxide and nutrients do
not limit oxygen production during the dynamic
experiments.
Based on these assumptions, Eq. (1) shows the macroscopic
oxygen mass balance in the liquid phase to estimate the oxygen
bulk-liquid concentration CL,O2.


@C L;O2
@ 2 C L;O2
@C L;O2
C G;O2
hL
hL Daz

h
u

K
a
C

rO2 C b 1
L
Le
L
O
L;O
2
2
@t
@z2
@z
H

I av

I0
1  expdt K a C b 
dt K a C b

Moreover, pO2,max is the maximum O2 production rate for the

microalga under the culture conditions. It can be related to temperature through the Arrhenius expression given in Eq. (5) [21];

pO2 ;max k0 exp





Ea
Ed
 k1 exp
TR
TR

The mass balance for the oxygen bulk-gas concentration in gas

phase CG,O2 is:

hg



@C G;O2
@C G;O2
C G ; O2
uGe hg
K L aO2 C L;O2 
@t
@z
H

With the following initial and boundary conditions:

with the following initial and boundary conditions (Fig. 1):

t 0; C L;O2 C sat
L;O2
L;

z 0; C L;O2 C sat
L;O2

@C L;O2
0
@z

Daz @C L;O2
uLe @z

t 0; C G;O2 C 0G;O2

z
2

In Eq. (1), rO2,intr is the intrinsic O2 production rate per biomass

unit dened as:

0
r O2 ;intr pO2 ;max @

I av
2

K s Iav IKavI

where Iav is the average light intensity inside the air-lift photobioreactor and is represented, [20], by:

z L;

@C G;O2
0
@z

Biomass productivity Pb is associated to the rO2,intr and to the

metabolic coefcient (Kd) [11,17] as:

dC b
Pb r O2 ;intr C b Y b=O2  K d C b
dt

The numerical solution of the Eqs. (1)(7) was carried out in

FlexPDE software 6.06 student version. It uses the nite element
method for solving partial differential equations. The hydrodynamics of the photobioreactor was solved using the Bubbly Flow application in the Chemical Engineering module of the commercial
software COMSOL Multiphysics, (Burlington MA, USA).

J. Cabello et al. / Chemical Engineering Journal 252 (2014) 104111

107

2.2. Experimental
2.2.1. Inoculum
The microalga Scenedesmus obtusiusculus [22], a promissory
strain for dioxide carbon xation and lipid storage was used. It
was initially cultivated for 2 weeks in a 3 L bubble column reactor
with mineral medium BG-11 at 30 C and a pH of 7.5 under continuous uorescent light illumination of 96 lmol m2 s1. The reactor
was fed with air containing 3.8% CO2 at a supercial velocity of
0.014 m s1. The air-lift photobioreactor was inoculated with this
culture in the exponential growth phase. N-replete medium experiments refer to the BG-11 medium with normal nitrogen concentration and N-starved is when medium does not contain nitrogen
source.
2.2.2. Description of the instrumented air-lift photobioreactor
Fig. 1 shows a schematic representation of the internal loop airlift photobioreactor. The acrylic column has a coaxial section with
internal diameter of 12.7 cm and a height of 110 cm and a degassing zone of 40 cm with a diameter of 20 cm. The concentric tube
has an inner diameter of 8.3 cm and an effective height of
105 cm, it was located 5 cm above the bottom of the column. The
gas phase was distributed from the bottom through a sparger with
60 holes of 0.56 mm internal diameter.
The external articial illumination system consisted of high
intensity white light LEDs (10 m of LED 5050 strip lights, illuminate LEDs, China) and also four 54 Watts uorescent lamps
(400700 nm, MAGG, Mexico). The intensity of light incident on
the surface of photobioreactor was measured with a quantum sensor
2p (407026sp, Extech, USA) and the photosynthetic photon uence
rate, PPFR, in the center of reactor with a spherical micro quantum
sensor 4p (US-SQS/L, Heinz Walz GmbH, Germany). The pH was
measured with an electrochemical sensor (27003-20, Cole-Parmer,
USA). A polarographic sensor (SN-29020-10, Cole-Parmer, USA)
was used to determine the dissolved oxygen concentration in the
range of 0200%; dissolved CO2 was measured with an electrochemical sensor (SN-29000-01, Cole Parmer, USA). CO2 in the gas
phase was monitored with an infrared detector (9500, Omega
Alpha, USA). The signals of the sensors located were continuously
recorded on-line by a data acquisition module (CompactDAQmx,
NI, USA) connected to a computer equipped with a NI LabVIEW
software for data logging.
The photobioreactor was operated in batch mode with 16.8 L of
N-replete medium and 1.8 L of S. obtusiusculus inoculum and
exposed to a continuous light intensity of 117 lmol m2 s1. Air
containing 3.8% CO2 was continuously supplied at a gas supercial
velocity of 0.0104 m s1 (based on the cross sectional area of the
riser, 3.4 L min1). The air-lift photobioreactor was also operated
in N-starved conditions with a biomass concentration of around
0.5 g L1. Biomass was determined every 24 h by dry weight after
ltration with 0.4 lm and lipid content was evaluated at the end
of the experiment with Nile Red [22].
2.2.3. Dynamic experiments
The dynamic experiments to determine the O2 evolution at different light intensities and at different growth stages of the microalga S. obtusiusculus were made from the rst day of operation and
sequentially as indicated in Fig. 2. The dynamic experiments
started with a light intensity of 117 lmol m2 s1 followed by step
increases to 141, 180, 336 and 505 lmol m2 s1 for periods of
30 min each. Once the highest light intensity was reached, it was
sequentially decreased to the same light intensities also for
30 min down to the normal conditions (117 lmol m2 s1). These
experiments were done at different operation times when biomass
contents were 0.13, 0.3, 0.4, 0.5, 1.7 and 1.9 g L1. At these conditions, the variables reached different pseudo-steady states and at

Fig. 2. Experimental data of N-replete biomass growth, dissolved O2, dissolved CO2,
pH and temperature evolution during the normal operation conditions of the air-lift
photobioreactor, and the periods in which the perturbations in the light intensity
were carried out (DI0).

this point the observable rate of O2 production per biomass unit

was determined for each dynamic experiment by the equation:

rO2 ;exp K L aO2 C L;O2 i  C sat

L;O2

considering the O2 saturation concentration, Csat

L,O2 and the O2 volumetric mass transfer, KLaO2, obtained in abiotic conditions.
For each dynamic experiment, the CO2 uptake rate was determined based on the gas velocity and the difference of the CO2 concentration in the gas phase at the inlet and outlet of the reactor for
each pseudo-steady state, using Eq. (10).

rCO2 ;exp

C CO2 ;s  C CO2 ;e  F G
VR

10

These dynamic experiments were done for both N-replete and

N-starved conditions.
2.2.4. Determination of the model parameters
An independent experiment at 28 3 C, under N-replete
growth conditions, was made in the airlift photobioreactor without
light intensity perturbations to determine the stoichiometric yield
coefcient, Yb/O2 and the metabolic coefcient, Kd. The parameters
were calculated from O2 production and biomass evolution during
9 days of operation.
The axial dispersion coefcient, Daz, was experimentally evaluated in abiotic conditions from the residence time distribution
curve obtained by injection of a tracer (1 M NaOH, 15 ml) into
the liquid phase. The gas hold-up, hG, effective gas velocity, uGe,
and effective liquid velocity, uLe, in the air-lift photobioreactor
were estimated with the COMSOL software similarly to the work
reported by Heckmat et al. [10], using the momentum transport
equations (two-uid EulerEuler model) for bubble column ow
for the axial symmetry (2D) geometry (mesh of 12704 triangular
elements. The simulation was done with a gas supercial velocity
of 0.0104 m s1.
The volumetric mass transfer coefcient for O2, KLaO2, was
experimentally estimated from the overall gasliquid mass transfer coefcient of carbon dioxide. It was determined at pH of 4 by
the mass balance of the CO2 absorption in liquid phase, for a gas
supercial velocity of 0.0104 m s1. The CO2 mass balance and

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J. Cabello et al. / Chemical Engineering Journal 252 (2014) 104111

the correction for the diffusion coefcients were established as follows [23]:


1=2
dC L;CO2
DO2
K L aCO2 C sat
L;CO2  C L;CO2 ; K L aO2 K L aCO2
dt
DCO2

11

The photosynthesis-irradiance curves of S. obtusiusculus at different temperatures were obtained with a method similar to that
described by Brindley et al. [3]. Thus, the intrinsic oxygen production rate was determined in a 3.5 ml jacketed reactor with
mechanical agitation under controlled temperature and incident
light intensity conditions. The initial biomass was adjusted to
0.1 g L1. Temperatures studied were between 540 C and light
intensities between 9 and 2400 lmol m2 s1 in an arrangement
of LEDs and uorescence lamps set around the reactor. Dissolved
oxygen was continuously recorded on-line by a data acquisition
module for a 20 min period. The effect of the dynamic photosynthetic response to the temperature variations caused by the light
step-changes was evaluated in an agitated isothermal 100 ml reactor containing 80 ml of a 0.5 g L1 cell suspension.
Intrinsic kinetic parameters in Eq. (3), Ks, KI and pO2,max were
obtained from the photosynthetic-irradiance curves. The values
of the intrinsic oxygen production rate were expressed as a function of average irradiance, Iav, to which the cells were exposed,
and tted using a hyperbolic model with an inhibitory term
[11,24]. The values of Ea, Ed, k0 and k1 were obtained by tting
Eq. (5) to the maximum oxygen production, PO2,max, and the temperature. Intrinsic O2 production rate refers only to biological effect
assuming low resistance of transport phenomena, low light attenuation and uniform irradiance in the jacketed reactor.
The biomass light absorption coefcient in Eq. (4), Ka, was
determined by measuring the absorbance of cultures with different
biomass concentrations. Light intensity in the center of the 3.5 ml
jacketed reactor illuminated from all directions was measured
with the spherical micro quantum sensor 4p. The absorption coefcient was calculated with the BeerLambert law equation.

Fig. 3. Photosynthetic photon uence rate (PPFR) in the airlift photobioreactor

center as a function of biomass content and time (insert) during the normal
operation conditions.

0.5 g L1, the photon ux density in the center decreased fourfold

from 120 to 30 lmol m2 s1. In our case, for biomass concentrations above 1 g L1, the reactor operated with less than
17.2 lmol m2 s1 of PPFR in the annular section (laminar ow)
and a dark phase in the concentric section (bubble ow). Under
these growth conditions, the nal biomass content (close to
2.0 g L1) was lower than that previously found for this strain,
[22], in a bubble column with an inner diameter of 0.105 m, and

3. Results and discussion

Fig. 2 shows the evolution of biomass and operational conditions of S. obtusiusculus in the photobioreactor under N-replete
conditions. The discontinuities reect the periods where the
dynamic experiments comprising step changes of light intensities
were performed. The dissolved O2 concentration increased from
saturation, 6.4 mg L1, to 10.5 mg L1 when the biomass was
1.7 g L1. The dissolved CO2 concentration remained at around
95.6% of saturation with respect to the inlet gas CO2 and the pH
increased from 6.4 to 7.2 due to nitrate and CO2 uptake. The overall
results showed cyclic temperature variations due to environmental
conditions, which in turn affected the produced O2 due to changes
in the photosynthetic activity, the CO2 consumed through an initial
accumulation of intracellular inorganic carbon and further carboxylation reactions [25] and the pH by changes in the CO2 and bicarbonate ion in the liquid phase. The small reduction in activity,
observed at around day 11, was also related to the lower temperature. An algal concentration close to 2 g L1 at day 15 was consistent with the initial mineral medium added and a residual soluble
nitrogen concentration of 64 mg L1. The values of Yb/O2 of 0.65 gb
1
g1
were obtained
O2 and of the metabolic coefcient (Kd) of 0.005 h
in the experiment without light perturbation. The Yb/O2, was similar to that reported by Acin et al. [20] of 0.77 gb g1
O2 and the Kd to
that reported by Molina et al. [4] of 0.00385 h1.
Fig. 3 shows the relationship between the internal PPFR and the
biomass. The non-linear behavior responds to the light attenuation
caused by absorption and scattering associated to the biomass
increase [12]. For instance, for biomass contents of 0.25 and

Fig. 4. Dynamic oxygen response to the light step-changes experiments, (a) and the
effect of temperature for a biomass content of 0.5 g L1. (b) For biomass content of
0.3, 0.4, 0.5 and 1.7 g L1.

J. Cabello et al. / Chemical Engineering Journal 252 (2014) 104111

109

with an irradiance of 134 lmol m2 s1. Compared to air lift bioreactors, bubble columns induce a higher frequency of cells shifting
between the photic and dark zones due to mixing in all the
cross-sectional area of the column which may possibly explain
the nal yield difference.
3.1. Dynamic uctuation of light intensity
As seen in Fig. 4a for a biomass content of 0.5 g L1 under Nreplete conditions, O2 concentration varies initially sharply as a
photosynthetic response to the light change and then slower due
to small temperature changes caused by the heat generated by
the external lamps. The pseudo-steady state O2 concentration, for
each irradiance, was obtained from the average in the slow
response period with its corresponding temperature. The O2 concentration response was not symmetric in the increase/decrease
light step-changes. The pseudo-steady state O2 concentrations,
obtained during the decrementing steps in the light intensities,
were higher than those previously obtained in the incremental part
due to a higher photosynthetic activity at higher temperatures. The
relevance of temperature is shown in Fig. 4a for an irradiance of
336 lmol m2 s1, where the increment of about 3 C increased
the pseudo-steady state oxygen in the liquid phase approximately
21% with respect to saturation despite the fact that at this higher
temperature the O2 solubility is reduced by approximately 5%.
For this microalga, the maximum value of the photosynthesis
activity is reached by an optimum temperature of 35 C. A separate
dynamic irradiance experiment in the 100 ml reactor under isothermal conditions (30 C) yielded similar pseudo steady state dissolved O2 values on both increasing/decreasing irradiance
conrming that the asymmetric behavior in Fig. 4b was induced
by temperature. Experiments presented by Levy et al. [26] showed
that, for some algae, the photosynthetic activities may vary in daytime during the morning and the afternoon, at the same sunlight
intensities levels. This apparent hysteresis effect has been attributed to a delay between the irradiation and the temperature gradient that is generated [26].
Fig. 4b shows the results of the not symmetric photosynthetic
response at different operation times when biomass were between
0.3 and 1.7 g L1 to the step changes of light intensities. For biomass contents of 0.3, 0.4, 0.5 and 1.7 g L1, the highest values of
pseudo-steady state O2 concentrations were 10.4, 11.1, 11.3 and
12.6 g L1 for the increment in the light step-changes up to
505 lmol m2 s1. Each pseudo-steady state represents the
short-term photo-acclimation period of S. obtusiusculus. In this
case, the state transitions (excitation energy redistribution
between photosystems) and non-photochemical mechanism operate to adjust the amount of light energy delivered to photosystem
II on a time scale of minutes [14,15,27].
Furthermore, signicant changes in the optical properties of S.
obstusiusculus were observed in the N-starved dynamic experiments. Fig. 5 compares the dynamic responses of O2 concentration
under N-replete and N-starved conditions during the changes in
the light intensities. Nitrogen deprivation affected the mechanisms
that regulated short-term photosynthetic acclimation, resulting in
a reduction in the photosynthesis activity and, therefore, the O2
concentration yield declined to 29% (day 3 of N-starvation) and
to 70% (day 18 of N-starvation) of the N-replete levels due to
changes observed in cell pigmentation. N-limitation affects photosynthesis activity reducing the efciency of energy collection due
to the chlorophyll loss and to the adjustment of the relative quantities of non-photochemically active molecules such as lipids or
carotenoids [19,28].
The delay in the dynamic O2 responses under N-starved condition in Fig. 5 compared to the N-replete medium, may be attributed
to the slow adjustment of the PSI/PSII stoichiometry ratio [15] due

Fig. 5. Comparison between the dynamic photosynthetic responses in N-replete

and N-starved conditions, for a biomass content of 0.5 g L1 and 505 lmol m2 s1.
Insert: relationship of the photosynthetic photon uence rate (PPFR) under Nstarved conditions; with high chlorophyll pigmentation (day 3) and low pigmentation (day 18).

to the decrease of the novo protein synthesis that directly affects

the PSII core proteins [19,27] due to nitrogen limitation. The Nstarved dynamic O2 response reached 95% of the response in
pseudo-steady state in about 6 h, (data not shown) indicating a
long-term photosynthetic adaptation of the cells [27].
Fig. 5 shows the light attenuation under N-starvation conditions
and the increase in the light availability due to changes in the optical properties of the cells. For instance, at the maximum incident

Fig. 6. Experimental data representing the short-term photosynthetic acclimation

of S. obtusiusculus under different incident light uctuations for biomass contents of
0.13(.), 0.3 (j), 0.4 (D), 0.5 (d), 1.7 ( ) and 1.9 g L1 (s) of (a) the specic O2
production rates; and (b) the specic CO2 uptake rates.

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J. Cabello et al. / Chemical Engineering Journal 252 (2014) 104111

light intensity, the PPFR values increased up to 3-fold in the center

of the reactor at day 18 of N-starvation in comparison to the day 3
of N-starvation. Under these conditions, the light penetration was
better and the cells were exposed to a larger quantity of light
energy, resulting in a higher metabolic ux generated from photosynthesis to be channeled to lipid accumulation from a content of
19% at day 3 to 42% at day 18.
Fig. 6a shows the O2 production rates for the incremental light
step-changes experiments that were determined by Eq. (9) for each
short-term photosynthetic photo-acclimation state (Fig. 4b) under
N-replete conditions. As can be seen, for biomass contents of
0.131.9 g L1 the photosynthetic activity increased linearly as the
incident light intensity increased from 141 to 505 lmol m2 s1,
indicating the absence of light saturation at these relatively low
light intensities. For the reactor used in this study, a biomass content of 1.9 g L1 produces an 87% reduction in O2 specic production rates due to the light attenuation. Fig. 6b shows the overall
rates of carbon dioxide consumption determined by Eq. (10), the
values increased with respect to the light intensity and diminished
as biomass content increased. Fig. 1SI in the supplementary information relates CO2 consumption and O2 production predicting an
1
endogenous respiration rate of 27.7 mg CO2 g1
.
b h
The light response curves (Fig. 6a and b) show that the overall
photosynthetic process is governed by light availability which in
turn denes the microalgal performance. The response to the light
depends strongly on the strain and the internal phenomena taking
place within photobioreactor. As far as we know, no analysis of this
type of photosynthesis response curves by changes in the light
intensity for Scenedesmus has been published yet. The behavior
of the light response curves can be compared qualitatively to those
obtained by Mazzuca et al. [29] and Rebolloso et al. [18] for a continuous pilot-scale tubular photobioreactor operated under cyclic
variation of solar radiation.

Fig. 7. Relation between simulated and experimental values obtained for the O2
concentrations at pseudo-steady state for biomass of 0.3 (j), 0.5 (d) and 1.7 g L1
( ).

3.2. Model validation

The mathematical model was validated by comparing experimental data (the increments of light step-changes in Fig. 4b) and
the predicted O2 concentration. The parameters used in the model
simulation are listed in Table 1. The values of the parameters are
consistent with those reported in the work by Fernndez et al.
[9] and a recent report by Bchet et al. [11]. Fig. 7 shows a good
agreement (5%) between experimental and predicted data and
therefore the model can be considered an adequate approximation
to describe the pseudo-steady state of the O2 concentration for biomass contents up to 1.7 g L1.

Table 1
Parameters used in the model.
Parameters

Value

Units

Daz
DCO2
DO2
Ea
Ed
hg
Ka
Kd
KI
KLaO2
KLaCO2
Ks
k0
k1
uGe
uLe
Yb/O2

0.027
1.9  105
2.7  105
16.1
30
0.02
0.096
0.005
4970
12.3
10.3
75.7
8.60  1013
3.63  1023
0.74
0.07
0.65

m2 s1
m2 s1
m2 s1
kcal mol1
kcal mol1
m3 m3
m2 g1
b
h1
lmol m2 s1
h1
h1
lmol m2 s1
1
gO2 Kg1
b h
1
gO2 Kg1
b h
m s1
m s1
gb g1
O2

Fig. 8. Model simulations of the effect of the temperature and light intensity on the
predicted O2 concentration in the liquid phase.

The simulations obtained through of the mathematical model

are shown in Fig. 8, under N-replete condition and for a biomass
of 0.5 g L1. The effect of the incident light and the temperature
on the O2 production was considered to obtain the best operation
conditions that could be used for the growth of the S. obtusiusculus
in the air-lift photobioreactor. The predicted O2 production was
normalized to better observe the inuence of the variables. The
model allowed describing the response curves of the irradiance
effect on photosynthesis, predicting between 72% and 85% of the
highest O2 concentration for light intensities between 600 and
980 lmol m2 s1. The effect of temperature on photosynthesis
estimated the maximum O2 concentration at 35 C and decreases
at temperatures between 38 and 45 C due to greater sensibility
of Ed in Eq. (5). Ed represents the enzymatic deactivation energy
of the photosynthesis apparatus of S. obtusiusculus. Validation of
the model with Yb/O2 and Kd values, (Eq. (8)), at 35 C with the
experimental data for biomass production at 117 lmol1 s1 is
shown in Fig. 2SI. The results do not show an inuence of the
dynamic experiments on the overall response of the microalga, this
may be due on one hand to the fact that the duration of the tests
was small, less that 10%, as compared to the experiment duration
(16 days) and on the other hand to the slow response of the

J. Cabello et al. / Chemical Engineering Journal 252 (2014) 104111

biomass growth as compared to O2 or CO2. The model is further

used to estimate concentration (Fig. 3SI) and productivity
(Fig. 4SI) at different light intensities obtaining values up to
0.78 gb L1 d1 at 610 lmol m2 s1.
4. Conclusions
The results showed that short unsteady state experiments on
microalgal cultivation allow gathering solid information that can
be used for a rapid analysis of the short-term oxygen response of
photosynthetic cells to changes in light uctuation avoiding long
experiments. A key nding in this study is that the dynamic
response indicated that the adaptation of the photosynthetic apparatus to light uctuations in N-starved cells is very slow for directing metabolic uxes to lipid accumulation and could represent the
current bottleneck for the production of microalgal oil.
The proposed model seems an efcient tool to predict the effect
of temperature and light on the dynamic O2 concentration and
consequently CO2 uptake and biomass productivity and could be
used in real-time control of photobioreactors and to select the best
operating conditions and optimize growth and productivity in microalgal production process.
Finally, dynamic changes in O2 production and CO2 uptake rates
could also be used to evaluate other critical operational variables
such as CO2 concentration in the gas phase or the effect of supercial gas velocity, which modies both the frequency of cell exposure to lightdark cycles, especially in air-lift reactors, and the
nutrient and gaseous transfer rates between the growth medium
and the microalga.
Acknowledgements
The authors thank to Alma Toledo, Teresa Lopez for their technical support and the scholarship provided by the National Council
of Science and Technology (CONACYT).
Appendix A. Supplementary material
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.cej.2014.04.073.
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