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Carbon dioxide fixation by microalgae cultivated in open bioreactors

Ana Priscila Centeno da Rosa, Lisiane Fernandes Carvalho, Luzia Goldbeck, Jorge Alberto Vieira Costa

Laboratory of Biochemical Engineering, College of Chemistry and Food, Federal University of Rio Grande (FURG), P.O. Box 474, Rio Grande, RS 96201-900, Brazil

a r t i c l e i n f o

Article history:

Received 7 April 2010Accepted 15 January 2011

Available online 20 May 2011

Keywords:

Carbon dioxideMicroalgaeRaceway

a b s t r a c t

The biofixation of carbon dioxide (CO2) by microalgae has been proven to be an efficient and economicalmethod, mainly due to the photosynthetic ability of these microorganisms to use this gas as a source of

nutrients for their development. The aim of this work was to study the growth of Spirulina LEB18 andChlorella kessleri microalgae, exposed to controlled and non-controlled conditions, with the injection ofdifferent concentrations of CO2. The cultures was carried out in 6 L open raceway ponds, under controlledconditions at 30 C and 39 lE m2 s1 and under non-controlled conditions, protected by a tunnel oftransparent film. The experiments were subjected to CO2 injections at concentrations of 0.038, 6, 12and 18% (v/v). The highest concentration of biomass (4.95 g L1) and maximum daily fixation(0.21 g g1 d1) were obtained for Spirulina LEB18 in culture that was prepared in non-controlled condi-tions with an injection of 6% (v/v) of CO2. C. kessleri had maximum (p < 0.0008) specific growth rate(0.84 d1) when grown with 18% (v/v) of CO2 in non-controlled conditions of cultivation.

2011 Elsevier Ltd. All rights reserved.

1. Introduction

The risk of irreversible effects on the worlds climate caused

by the increase in the greenhouse effect has stimulated thedevelopment of technologies for CO2 mitigation [1]. This reduc-tion can be achieved by increasing the efficiency of energy gen-eration [2], replacing fossil fuels with other sources of energy[3], growing trees to fix carbon [4] or by cultivating microalgae[5].

For the cultivation of microalgae in non-controlled conditions,open bioreactors can be used, which require low initial investment,are easy to build and operate [6], and use sunlight as the mainsource of energy [7].

The fixation of CO2 by microalgae exposed to non-controlledconditions may be one of the most effective alternatives for reduc-ing the emission of this gas. The use of CO2 from industrial gener-ation decreases the impact of CO2 on the environment and the cost

of producing biomass [8]. Microalgal biomass may be used as asource of bioactive compounds, such as polyunsaturated fattyacids, b-carotene and other pigments considered to be antioxidants[9,10], as well as biofuels [11].

The aim of this work was to study the growth and biofixationof CO2 by the Spirulina LEB 18 and Chlorella kessleri microalgae,kept at different conditions of cultivation and concentrations ofCO2.

2. Material and methods

2.1. Microorganisms and cultivation media

Spirulina LEB 18 [12] and C. kessleri [13] microalgae were usedand kept in Zarrouks medium [14] and MBM medium [15], respec-tively. The organisms were adapted for 7 d with 1% CO2 (v/v).

2.2. Bioreactors and cultivation conditions

The cultures were prepared in 6 L open raceway ponds, with a5 L useful volume and were agitate using a paddle-wheel rotatingat 18 revs min1. The experiments were conducted under con-trolled conditions (CC) and non-controlled (NC). The cultures un-der controlled conditions were placed in a growth chamber at30 C and illuminated with 39 lE m2 s1, provided by 40 W fluo-rescent lamps and a 12 h light/dark photoperiod. The cultures innon-controlled conditions were sited in an outdoor greenhouse,protected from UV light.

The CO2 that was added to the cultures came from industrialcylinders, through sprinklers made of sintered material, with aflow rate of 0.3 vvm (volume of air per volume of media per min-ute). The mixing of air and CO2 was performed at concentrations of6%, 12% and 18% (v/v), injected for 15 min h1, during the light per-iod of cultivation. For each experimental condition, a control testwas carried out without the addition of CO2.

In the cultures with Spirulina LEB 18, the CO2 replaced the car-bon source of the culture medium standard (16.8 g L1 of NaHCO3).For the C. kessleri, this substitution did not take place, because theonly source of carbon in its culture is derived from atmospheric

0196-8904/$ - see front matter 2011 Elsevier Ltd. All rights reserved.doi:10.1016/j.enconman.2011.01.008

Corresponding author. Tel.: +55 53 32338653; fax: +55 53 32338745.E-mail address: dqmjorge@furg.br (J.A.V. Costa).

Energy Conversion and Management 52 (2011) 30713073

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CO2. The cultures were discontinuous, with initial biomass concen-tration of 0.15 g L1, and lasted for 20 d. The volume of the cultureswas kept constant by the daily replacement of water lost byevaporation.

2.3. Analytical determinations

The concentration of biomass and the pH of cultures were mon-itored daily. The concentration of biomass was determined bymeasuring the optical density of the samples using a 670 nm spec-trophotometer (Femto 700-Plus, Brazil), using a standard curvethat relates the optical density of each microalgae with dry weightof biomass [16]. Digital pHmeter was used to measure pH (QuimisQ.400H, Brazil). The concentration of carbon, hydrogen, nitrogenand sulfur in the biomass was determined in duplicate for eachexperiment, using a CHNS Elementary Analyzer (PerkinElmer2400, USA). A certified cystine standard was used for the calibra-tion of the equipment (Perkin Elmer, USA).

2.4. Determination of the kinetic parameters

For each experiment was determined the maximum concen-tration of biomass (Xmx, g L

1), maximum specific growth rate(lmx, d

1) and maximum specific death rate (k, d1). The maxi-mum specific growth rate was obtained by regression in the log-arithmic phase of growth. The maximum specific death rate wasobtained by exponential regression in the phase of cellulardecline [17]. This study determined the daily fixation rates of

CO2 (FD, gCO2fixed g1

CO2injected d1) in the cultures, which is the

amount of fixed CO2 (g) in relation to the total injected (g) perday [18].

2.5. Statistical analysis

The kinetic responses were assessed by analysis of variance

(ANOVA) to compare the means with a 95% level of confidence(p < 0.05), using Tukeys test.

3. Results and discussion

The results of maximum concentration of biomass, maximumspecific growth rate and daily maximum CO2 fixation for the cul-tures under different conditions are presented in Table 1. Fig. 1shows the growth curves as a function of time for the SpirulinaLEB 18 and C. kessleri microalgae.

According to Fig. 1a, the Spirulina LEB 18 grown under non-controlled conditions (NC) had an adaptation phase of approxi-mately 3 d. At the end of the 20th day it was still not in a stage ofcellular decline as happened with C. kessleri, both under controlledconditions and in non-controlled (Fig. 1b). The C. kessleri enteredthecelldeathphaseafterapproximately11dofcultureinbothstud-ied conditions. Thespecific death rate (k) ofC. kessleri when exposedto non-controlledconditionswas 0.14,0.11 and0.09 d1 for cultureswith the addition of 6%, 12% and 18% CO2, respectively.

The test conducted in non-controlled conditions, in which therewas no added CO2 in the culture of C. kessleri, had no cell deathphase. The only source of carbon in this test was 0.038% CO2, towhich the microalgae was already adapted. Morais and Costa[13] used CO2 as a carbon source in cultures in 2 L tubular verticalphotobioreactors, with the C. kessleri, which also did not have a celldeath phase in the culture with the addition of 18% (v/v) of CO2.

For the Spirulina LEB 18 grown under controlled conditions,there was no difference in the maximum concentration of bio-

mass (p > 0.06) for the tests with the addition of 6%, 12% and18% (v/v) CO2 (0.81 g L

1, 0.78 g L1 and 0.64 g L1, respectively).According to Morais and Costa [5,13], when 4 L vertical tubularphotobioreactors were exposed to 39 lE m2 s1, a 12 h light/darkphotoperiod and 30 C, the maximum values of biomass concen-tration were achieved when Spirulina LEB 18 was grown with6% (v/v) (4.13 g L1) and Scenedesmus obliquus with 12% (v/v)(1.14 g L1) of CO2.

The test using Spirulina LEB 18, under non-controlled conditionsand 6% (v/v) CO2 had the highest concentration of biomass(4.95 g L1, p < 0.0002). For the C. kessleri, the highest values

Table 1

Maximum concentration of biomass (Xmx, g L1), maximum specific growth rate

(lmx, d1) and maximum daily fixation of CO2 (FDmx, gfixed CO2 g

1injetedCO2 d

1) for

Spirulina LEB 18 and C. kessleri grown under controlled conditions (CC) and non

controlled conditions (NC), exposed to various concentrations of CO2.

Spirulina LEB 18 C. kessleri

%CO2 Xmx lmx FDmx Xmx lmx FDmxCC 0.038 0.83a 0.12j 0.67v 0.51c,e 0.17k 1.09u

6 0.81a,b 0.12j 0.03t 0.52c,e 0.19l 0.02t

12 0.78a,b 0.13j 0.01t 0.54c,e 0.20l 0.01t

18 0.64b,c 0.11n 0.01t 0.47c 0.18k 0.01t

NC 0.038 4.82d 0.26o 4.66x 0.64b,e 0.08m 1.32u

6 4.95d 0.23p 0.21t 1.57i 0.08m 0.10t

12 4.15g 0.17k 0.18t 2.62f 0.82r 0.09t

18 3.60h 0.16q 0.10t 2.50f 0.84s 0.07t

The same letter for the same parameter, indicates no significant difference(p < 0.05) between the means Tukeys test.

Fig. 1. Curves of growth for Spirulina LEB 18 (a) and C. kessleri (b) grown under controlled conditions (closed symbol) and non-controlled conditions (open symbol), using theCO2 concentrations of 0.038% (d,s), 6% (j,h), 12% (N,4) e 18% (,}).

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(p < 0.0002) for biomass concentration achieved were 2.62 and2.50 g L1 in cultures under non-controlled conditions using 12and 18% (v/v) CO2, respectively. Chlorella has been studied as analternative method of absorption of atmospheric CO2 due, mainly,to its characteristic of maintaining growth when cultivated underhigh concentrations of CO2. When grown with 70% (v/v) of CO2[19], Chlorella ZY-1 had the maximum concentration of biomass

in the test with the addition of 10% (v/v) CO2.The maximum specific growth rate (lmx) for Spirulina LEB 18,grown under controlled conditions did not differ significantly(p > 0.48) when the concentration of CO2 went from 0.038%(0.12 d1) to 12% (v/v) (0.13 d1). On the other hand, the Synecho-coccus sp. CCMP1334 [20], grown in a closed photobioreactor and0.075% CO2 showed no significant difference regarding lmx whencompared with the culture with the addition of 0.038%.

In non-controlled conditions when CO2 was added to the cul-ture, the Spirulina LEB 18 had the highest lmx (0.23 d

1) in cul-tures with addition of 6% (v/v) CO2 (p < 0.0002). When cultivatedin 4 L tubular photbioreactors, Spirulina LEB 18 had a lmx of0.42 d1 in the test with 6% CO2 (v/v) [5]. For the Chlamydomonasacidophila [21] cultivated in closed photobioreactors, the lmxwas 0.52 d1, when 5% (v/v) of CO2 was used.

For C. kessleri the highest values oflmx under controlled condi-tions were achieved when 6% and 12% (v/v) CO2 (0.19 d

1 and0.20 d1, respectively) were injected. In non-controlled conditions,C. kessleri had the highest lmx (0.84 d

1) when injected 18% of CO2(p < 0.0008). When C. kessleri was grown at 6% and 12% (v/v) of CO2in 4 L tubular vertical photobioreactors at 30 C, 39 lE m2 s1 anda 12 h light/dark photoperiod [13], it had the highest lmx values(0.27 d1), showing that uncontrolled conditions are more favor-able for microalgal growth.

C. kessleri had the highest Xmx (2.62 and 2.50 g L1) and lmx

(0.82 and 0.84 d1) in tests conducted in non-controlled conditionswith the addition of 12% and 18% (v/v) of CO2, respectively. In thesetrials the light intensity reached maximum values of up to1040 lE m2 s1 and a temperature of 40 C and this may have

caused a greater need for the microalgae to consume the nutrients.There was no significant difference (p < 0.04) between the val-

ues of CO2 fixation (FDmx) for both microalgae and conditions ofcultivation, when CO2 was added. This indicates the possibility ofSpirulina LEB 18 and C. kessleri cultivation with the addition of thisgas at the studied proportions, even if these were not adapted tocarbon in their original cultivation media. The C. vulgaris grownwith the addition of 0.5% to 3% of CO2 [22] attained highest CO2fixation with 1% CO2 in the culture. Doucha and Lvansky [23]cultivated Chlorella sp. in 1000 L open bioreactors under non-controlled conditions with the injection of 0.15% (v/v) of CO2 andobtained 70% absorption of injected CO2.

When CO2 was added, the largest FDmx was obtained forSpirulina LEB 18 with 6% CO2 in CN (0.21 gCO2fixed g

1CO2injected

d

1). Morais and Costa [18] obtained an FDmx of 9.15%, when theycultivated Spirulina LEB 18 with 6% CO2 in 4 L photobioreactors andcontrolled conditions of cultivation.

4. Conclusions

The Spirulina LEB 18 and C. kessleri presented growth during the20 d of culture with concentrations of up to 18% (v/v) of CO 2 in theculture. The maximum values of the concentration of biomass andfixation of CO2 were 4.95 g L

1 and 0.21 gfixed CO2 g1

injected CO2 d1,

respectively, in cultures with Spirulina LEB 18 exposed to non-controlled conditions and an injection of 6% (v/v) CO 2. C. kessleri

had the highest specific growth rate (0.84 d1) when grown with18% (v/v) CO2 in non-controlled conditions of cultivation. There-fore, Spirulina LEB 18 and C. kessleri can be used in the fixation ofCO2 emitted from industrial activities, which contributes to areduction in global warming.

Acknowledgements

The authors thank Centrais Eltricas Brasileiras S.A (ELET-ROBRS) and the Companhia de Gerao Trmica de Energia Eltri-ca (CGTEE) for their financial support of this study.

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