1 2 Influence of bioflocculation parameters on harvesting Chlorella salina and its 3 optimization using response surface methodology 4 Surendhiran Duraiarasan Q1 , Vijay Mani * 5 Bioelectrochemical Laboratory, Department of Chemical Engineering, Annamalai University, Annamalainagar, Tamilnadu 608002, India 6 7 Introduction 8 Increasing demand and forthcoming depletion of fossil fuel 9 resources lead to the search for new and clean energy sources. 10 Biodiesel is one of the new types of renewable energy which is 11 considered to be a possible substitute of conventional diesel 12 because of its biodegradable nature, non-toxic, renewable and 13 reduced emission level of CO, SO 2 [1–4]. Nowadays microalgae are 14 focused by many researchers all over world for its source of 15 biodiesel-convertible lipids. They are a group of diverse photosyn- 16 thetic organisms that can accumulate substantial amounts of 17 lipids-up to 20–50% of dry cell weight in certain species [5–7]. 18 Moreover, microalgae have advantages of high growth rate over 19 traditional biodiesel feedstock (able to double their biomass within 20 a period of 24 h) [8,9]. 21 The microalgal size is very small (i.e., 3–30 mm in diameter) and 22 possesses similar density to water [10,11] hence the efficient 23 separation of cells from culture broth as well as to maintain their 24 viability is found to be difficult in large scale production of 25 microalgae. Moreover, harvesting of biomass represents one of the 26 significant cost factors in the production of biodiesel. Therefore, 27 microalgae harvesting process became a challenging task and 28 commercial production of biodiesel from microalgae is economi- 29 cally unfeasible. 30 Different studies showed that the harvesting cost of algal 31 production in open ponds accounts for more than 20–30% of the 32 total cost of biodiesel production [7]. Many separation methods, 33 such as centrifugation, gravity sedimentation, (ultra)filtration and 34 screening, flocculation, flotation and ultra sound waves have been 35 developed for microalgae recovery [8,10,12,13]. However, each has 36 its disadvantages that affect the overall economics of the process. 37 Centrifugation requires high energy input and initial capital cost 38 [8] and the process involves exposing cells to high gravitational 39 and shear forces which damage the cell structure. Second, the 40 processing of large culture volumes can be time-consuming [14]. 41 Filtration and screening require regular replacement of filters, 42 screens, membranes and can be very time consuming. Gravity 43 sedimentation is a low process and electroflotation requires 44 replacement of worn electrodes that have been consumed and a 45 high cost of electricity [8,15]. Thus, to minimize the energy 46 consumption of harvesting microalgae, an integrated approach is 47 needed [16]. Among several harvesting methods, flocculation 48 found to be an easy and effective method for harvesting of 49 microalgae [17]. During flocculation, the dispersed microalgal cells 50 aggregate and form flocs with higher sedimentation rate [18]. 51 Widely used chemical flocculants are non biodegradable and also 52 their intermediate degraded products are harmful to humans 53 [18,19]. For example, aluminum salts were verified as strong agent 54 that causes Alzheimer’s disease [20]. To resolve this problem, Journal of Environmental Chemical Engineering xxx (2013) xxx–xxx A R T I C L E I N F O Article history: Received 7 May 2013 Received in revised form 10 July 2013 Accepted 12 August 2013 Keywords: Bioflocculant Biodiesel Flocculation efficiency Response surface methodology Cell viability A B S T R A C T The present investigation deals with a cost effective harvesting of microalga, Chlorella salina, for biodiesel production with microbial flocculant, an exopolysaccharide (EPS) from marine Bacillus subtilis. The process with five independent variables namely temperature, pH, flocculation time, bioflocculant size and cationic inducer (ZnCl 2 ) concentration, were evaluated using one factor at a time and were statistically optimized by response surface methodology (RSM). Bioflocculation was enhanced by adding ZnCl 2 as the cationic inducer, which worked under the principle of ‘‘divalent cation bridging (DCB)’’ theory. Additionally ZnCl 2 did not distort the cell’s structural integrity. Using RSM, a maximum efficiency was found to be more than 98.66% with flocculation parameters as temperature (30.63 8C), pH (10.4), flocculation time (6.2 h), bioflocculant size (0.34 ml) and cationic inducer size (0.031 mM). This flocculation study concluded that chemical flocculation could be disadvantageous due to cell disintegration, toxicity and cost consumption, whereas a bioflocculation is environmental friendly as well as cost prohibitive process. ß 2013 Published by Elsevier Ltd. * Corresponding author. Tel.: +91 9443227891. E-mail address: [email protected](V. Mani). G Model JECE 130 1–6 Please cite this article in press as: S. Duraiarasan, V. Mani, Influence of bioflocculation parameters on harvesting Chlorella salina and its optimization using response surface methodology, J. Environ. Chem. Eng. (2013), http://dx.doi.org/10.1016/j.jece.2013.08.016 Contents lists available at ScienceDirect Journal of Environmental Chemical Engineering jou r n al h o mep ag e: w ww .elsevier .co m /loc ate/jec e 2213-3437/$ – see front matter ß 2013 Published by Elsevier Ltd. http://dx.doi.org/10.1016/j.jece.2013.08.016
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Journal of Environmental Chemical Engineering xxx (2013) xxx–xxx
G Model
JECE 130 1–6
Influence of bioflocculation parameters on harvesting Chlorella salina and itsoptimization using response surface methodology
Surendhiran Duraiarasan , Vijay Mani *
Bioelectrochemical Laboratory, Department of Chemical Engineering, Annamalai University, Annamalainagar, Tamilnadu 608002, India
A R T I C L E I N F O
Article history:
Received 7 May 2013
Received in revised form 10 July 2013
Accepted 12 August 2013
Keywords:
Bioflocculant
Biodiesel
Flocculation efficiency
Response surface methodology
Cell viability
A B S T R A C T
The present investigation deals with a cost effective harvesting of microalga, Chlorella salina, for biodiesel
production with microbial flocculant, an exopolysaccharide (EPS) from marine Bacillus subtilis. The
process with five independent variables namely temperature, pH, flocculation time, bioflocculant size
and cationic inducer (ZnCl2) concentration, were evaluated using one factor at a time and were
statistically optimized by response surface methodology (RSM). Bioflocculation was enhanced by adding
ZnCl2 as the cationic inducer, which worked under the principle of ‘‘divalent cation bridging (DCB)’’
theory. Additionally ZnCl2 did not distort the cell’s structural integrity. Using RSM, a maximum efficiency
was found to be more than 98.66% with flocculation parameters as temperature (30.63 8C), pH (10.4),
flocculation time (6.2 h), bioflocculant size (0.34 ml) and cationic inducer size (0.031 mM). This
flocculation study concluded that chemical flocculation could be disadvantageous due to cell
disintegration, toxicity and cost consumption, whereas a bioflocculation is environmental friendly as
well as cost prohibitive process.
� 2013 Published by Elsevier Ltd.
Contents lists available at ScienceDirect
Journal of Environmental Chemical Engineering
jou r n al h o mep ag e: w ww .e lsev ier . co m / loc ate / jec e
28293031323334353637383940414243444546474849
Introduction
Increasing demand and forthcoming depletion of fossil fuelresources lead to the search for new and clean energy sources.Biodiesel is one of the new types of renewable energy which isconsidered to be a possible substitute of conventional dieselbecause of its biodegradable nature, non-toxic, renewable andreduced emission level of CO, SO2 [1–4]. Nowadays microalgae arefocused by many researchers all over world for its source ofbiodiesel-convertible lipids. They are a group of diverse photosyn-thetic organisms that can accumulate substantial amounts oflipids-up to 20–50% of dry cell weight in certain species [5–7].Moreover, microalgae have advantages of high growth rate overtraditional biodiesel feedstock (able to double their biomass withina period of 24 h) [8,9].
The microalgal size is very small (i.e., 3–30 mm in diameter) andpossesses similar density to water [10,11] hence the efficientseparation of cells from culture broth as well as to maintain theirviability is found to be difficult in large scale production ofmicroalgae. Moreover, harvesting of biomass represents one of thesignificant cost factors in the production of biodiesel. Therefore,microalgae harvesting process became a challenging task and
Please cite this article in press as: S. Duraiarasan, V. Mani, Influence ooptimization using response surface methodology, J. Environ. Chem
2213-3437/$ – see front matter � 2013 Published by Elsevier Ltd.
http://dx.doi.org/10.1016/j.jece.2013.08.016
commercial production of biodiesel from microalgae is economi-cally unfeasible.
Different studies showed that the harvesting cost of algalproduction in open ponds accounts for more than 20–30% of thetotal cost of biodiesel production [7]. Many separation methods,such as centrifugation, gravity sedimentation, (ultra)filtration andscreening, flocculation, flotation and ultra sound waves have beendeveloped for microalgae recovery [8,10,12,13]. However, each hasits disadvantages that affect the overall economics of the process.Centrifugation requires high energy input and initial capital cost[8] and the process involves exposing cells to high gravitationaland shear forces which damage the cell structure. Second, theprocessing of large culture volumes can be time-consuming [14].Filtration and screening require regular replacement of filters,screens, membranes and can be very time consuming. Gravitysedimentation is a low process and electroflotation requiresreplacement of worn electrodes that have been consumed and ahigh cost of electricity [8,15]. Thus, to minimize the energyconsumption of harvesting microalgae, an integrated approach isneeded [16]. Among several harvesting methods, flocculationfound to be an easy and effective method for harvesting ofmicroalgae [17]. During flocculation, the dispersed microalgal cellsaggregate and form flocs with higher sedimentation rate [18].Widely used chemical flocculants are non biodegradable and alsotheir intermediate degraded products are harmful to humans[18,19]. For example, aluminum salts were verified as strong agentthat causes Alzheimer’s disease [20]. To resolve this problem,
f bioflocculation parameters on harvesting Chlorella salina and its. Eng. (2013), http://dx.doi.org/10.1016/j.jece.2013.08.016
S. Duraiarasan, V. Mani / Journal of Environmental Chemical Engineering xxx (2013) xxx–xxx2
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oflocculant has attracted considerable attention and promisingbstitute for the chemical flocculants because of their biodegrad-ility, high efficiency, non-toxicity and safety for ecosystems1,22].Bioflocculants are polymers produced by microorganisms like
cteria, actinomycetes, yeast, fungi and algae during their growth3], with flocculating activities that are dependent on thearacteristics of the flocculants since bioflocculants are primarilyade up of exopolysaccharide (EPS), which plays an important role
bioflocculation process. The biotechnological process has not yeten able to overcome the chemical flocculation because theantity of EPS produced by the bacterial culture is lesser and
ould be insufficient when harvesting in large scale i.e. thetracellular product from the bacterial cell is cost consuming.nce this study involves the use of whole live culture as the
oflocculant for harvesting of Chlorella salina for biodieseloduction. Though bioflocculants are environment friendly thanemical flocculants, the flocculation efficiency is lower than the
tter. This is due to the similar net negative charges of EPS and theicroalgal cell wall. In order to enhance bioflocculation efficiency,e addition of divalent cations was employed as an inducer in theoflocculation process.
In this paper, bioflocculation using the bioflocculant (wholells) Bacillus subtilis was investigated. The effects of variousrameters namely temperature, pH, cationic inducer, biofloccu-
nt size and flocculation time were studied and optimizedatistically by response surface methodology (RSM). As to best
our knowledge, these are scanty reports available on use ofhole culture as bioflocculant, thus this would be one of the firstports on bioflocculation using live cells.
aterials and methods
lture condition
C. salina, obtained from Central Marine and Fisheries Researchstitute (CMFRI), Tuticorin, Tamilnadu (India), was grown inerile Walne’s medium. The filtered sterilized sea water wasriched with required quantity of Walne’s medium containing
l�1): NaH2PO4�2H2O, 20.0; Na2EDTA, 4.0; H3BO3, 33.6;nCl2�4H2O, 0.36; FeCl3�6H2O, 13.0; vitamin B12, 0.001; vitamin, 0.02; and NaSiO3, 6.6. The trace metal solution contained
l�1): ZnSO4�7H2O, 4.4; CoCl2 �6H2O, 2.0; (NH4)6Mo7O24�H2O, 0.9;d CuSO4�5H2O, 2.0. The medium was adjusted to pH 8 andtoclaved at 1218 C for 20 min. The filter sterilized vitamins wereded after cooling. The contents were later introduced into a0 ml Erlenmeyer flask and finally transferred to 20 l photo-
oreactor (PBR). Mixing was provided by sparging air from thettom of the PBR and lighting was supplied by cool-whiteorescent light with an intensity of 5000 lux under 12/12 light/rk cycle. End of the log phase culture was used for thecculation experiments.
lture for bioflocculation
The marine bacterial culture B. subtilis (MTCC 10619) was used the bioflocculant, obtained from the Department of Marine
ble 1ded values based on the factor at a time experiment for the 5 variables employed
ode Variables �2
1 Temperature (8C) 20
2 pH 6
3 Flocculation time (h) 2
4 Bioflocculant size (ml) 0.1
5 Cationic inducer concentration (Mm) 0.01
Please cite this article in press as: S. Duraiarasan, V. Mani, Influence
optimization using response surface methodology, J. Environ. Chem
biology, CAS in Marine biology, Parangipettai, Annamalai Univer-sity. The bacterial culture was cultivated for growth andbioflocculant production using Nutrient broth containing peptone– 5 g l�1, beef extract – 1.5 g l�1, yeast extract – 1.5 g l�1 and NaCl –5 g l�1, subcultured periodically and stored as stocks on nutrientagar slants at 4 8C.
Evaluation of bioflocculation experiment – one factor at a time design
A quantity of 50 ml of C. salina was used for optimization study.The effects of flocculation parameters, namely temperature, pH,time, bioflocculant concentration and cationic inducer size, wereindividually experimented by analyzing bioflocculation efficiency.For the effect of pH, the culture was divided in different test tube,and the pH was adjusted to fixed values by the addition of 1 M HCland 1 M NaOH, ranging from approximately 6.0–10. Likewise, foreffect of temperature the test tubes were incubated at desiredtemperatures.
After the set up of fixed parameters, each tube was kept inorbital shaker (Model-Technico, Honeywell Ltd., India) and stirringspeed was maintained at 250 rpm. The initial microalgal biomassconcentration in the tubes was estimated from the optical densityof 750 nm (OD750), using UV-VIS Spectrophotometer (Model-SL159, ELICO Ltd., India). At a time interval of every 30 min, theoptical density of the supernatant was measured at half the heightof the clarified culture. Culture broth without bioflocculant wasused as control and culture medium with appropriate quantity ofeach salt were used for blank to respective flocculants. Flocculationefficiency was calculated by [14,24]:
Flocculation Efficiency ð%Þ ¼ 1 � A
B
� �� 100;
where A = OD750 value of sample and B = OD750 value of control.
Response surface methodology-CCD
A central composite design (CCD) of the experiments wasformulated to investigate five flocculation parameters. Each 50 mlculture of C. salina was added into test tubes and the parameterswere set according to the orthogonal values of central compositedesign (CCD) (Table 1). RSM is known to evaluate the interactionbetween the significant factors of an experiment and optimizethem [25]. Five level factorial experiment set up, with 8 centralpoints, was designed using Design Expert Software version 8.0.7.1,Stat-Ease, Minneapolis, USA and the quality of analysis model wasbased on analysis of variance (ANOVA). The response variable (Y),representing the bioflocculation activity was fitted using a second-order polynomial equation given as,
S. Duraiarasan, V. Mani / Journal of Environmental Chemical Engineering xxx (2013) xxx–xxx 3
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where Y is the predicted response, b0 was the constant, X1 to X5
were the input variables, b1 to b5 were the linear coefficients, b12
to b45 were the second order interactive coefficients and b11 to b55
were the quadratic coefficients.The actual values of coded levels of different parameters:
Temperature (X1), pH (X2), flocculation time (X3), bioflocculant size(X4) and cationic inducer concentration (X5) is presented in Table 1and its influence on harvesting of microalgae by flocculation,represented as Y, the response variable, has been investigated. Theactual values of coded level ‘00 were fixed based on one-factor-at-a-time method.
Results and discussion
Variables influencing the bioflocculation process
Effect of temperature on bioflocculation
Effective process occurred at a temperature of 30 8C, as the cellsof marine bacterium, B. subtilis, was able to produce morebioflocculant i.e. exopolysaccharide (EPS) because of temperaturestress condition. A rapid decrease in efficiency was observed, whenthe temperature was raised above 30 8C, which was due to thesusceptibility of microalgae cells as well as molecular mobility athigher temperature. Thus collision occurred between bioflocculantand microalgal cells, which lead to cell distortion [8]. Moreover, asthe microalgae and the bioflocculant producing bacteria are frommarine sources, supplementation of additional medium compo-nents/nutrients may not be necessary.
Effect of pH on bioflocculation
pH is one of the most important factors for harvestingmicroalgae, hence the influence of pH on bioflocculation efficiencywas tested with pH ranging between 6 and 10. From the statisticalexperimental results, the effect of pH on flocculation efficiency washighly significant (p < 0.01) and the flocculation efficiency wasfound to be higher with increase in pH i.e. 10. This result is inagreement with previous studies [26]. As pH increases, thenegative charge of microalgal cells increase. This phenomenoncould be a major cause for flocculation at higher pH which is due todifference in protonation conformational changes and structuralalterations in flocs.
Effect of time on bioflocculation
As the time prolonged, deterioration was observed in bio-flocculation efficiency. Bioflocculants (EPS) are generally found tobe produced during late exponential phase or stationary phase ofthe bacterial growth [27], after which the concentration or theproduction of the polymer remains constant in the medium. Henceas the time increased beyond the production time, flocculationdecreased. Lower efficiency could also be experienced when cellsdue to the production of deflocculating enzymes, along withbioflocculants, beyond stationary phase [27].
Effect of bioflocculant size on bioflocculation
Significant effect was observed with bioflocculant size onflocculation of C. salina. More the bioflocculant, lesser was theinteraction between them and lower was the efficiency. Theflocculation mechanism between microalgal cells and bacterialcells happened as a series of interbridging between cells,neutralization of charges and precipitation enmeshment [25].Larger amount of bioflocculant might be detrimental, due to itsadsorption to the cells, reducing their surface potential thusdestabilizing the microalgal cells. Similar report was observed onharvesting microalgal cells using g-polyglutamic acid, which onoverdosing with 30 mg l�1, caused lower flocculation due to chargeneutralization and destabilization [25].
Please cite this article in press as: S. Duraiarasan, V. Mani, Influence ooptimization using response surface methodology, J. Environ. Chem
Role of cationic inducer
For this study, ZnCl2 was used as cationic inducer, as confirmedby cell viability test carried out in our previous study [28]. Traceamount of ZnCl2 added as cationic inducer significantly affectedbioflocculation. Addition of divalent cationic salts in the mediumenhances the flocculation at high pH by interlinking the cells,forming dense flocs [26] because the exopolymer produced by B.
subtilis was negatively charged as similar to the microalgal cellwall. This mechanism is known as ‘‘divalent cation bridging theory(DCB)’’ [29]. Zn2+ salt aides the process as a linker, whichneutralizes the residual negative charge of functional groups thusenhancing the bioflocculation process [21]. Higher concentrationof the inducer, led to the destruction of the conformation of thecells, and flocculation efficiency became lesser (Fig. 1).
Central composite design
Bioflocculation of marine microalgae, C. salina was carried outwith the bacterial cells of B. subtilis which is capable of producingbioflocculant, i.e. EPS (exopolysaccharide). Optimization of the fiveindependent variables was performed using central compositedesign (CCD) with 50 runs consisting of 8 central points. Theinteraction between the studied variables was tested by carryingout experiments using different concentrations. The higher levelswere fixed as in Table 1, above which the algal cells get disrupteddue to stress imposed on the cells by each variable at elevatedconditions and particular lower levels were taken because growthof cells would not occur.
Multiple linear regression analysis was carried out using asecond order polynomial equation that was fitted to the abovedata as,
where Ybiofloc is the response variable, X1 to X5 are the linear effectsof the independent variables, X1 X2 to X4, X5 are the interactiveterms of the variables and X2
1 to X25 squared effects of the
variables. A positive sign denoted that the effect of the variableson flocculation was greater at a higher concentration whereas anegative symbol represented that influence of variable onflocculation was greater at a lower concentration.
The goodness of fit of regression equation developed could bemeasured by determination coefficient. The R2 value of 0.9852 andadjusted R2 of 0.9447 showed that the model could be significantpredicting the response and explaining 95% of the variability in thedata. ANOVA table was illustrated (Table 2). The calculated F
value (103.06) and a low p value (p = 0.000) demonstrated that thequadratic model was highly significant. From the table it was alsoobserved that the coefficient of square, linear and interactioneffects were also highly significant (p = 0.000). Smaller theprobability (p) values, i.e. lesser than 0.05 (p < 0.05) and largerthe magnitude of ‘t’ value was highly significant. The coefficients ofthis response, viz., X1, X2, X3, X4, X5, X2
1 ; X22 ; X2
3 ; X24 ; X2
5 , X1X2, X1X5,X2X4, X2X5 and X3X4 were found to be the most significant of thismodel (p < 0.05).
Three dimensional response surface plots and contour plots forthe bioflocculation efficiency were shown in Fig. 1. The shapes ofthe contour plots indicate the significance of the interaction
f bioflocculation parameters on harvesting Chlorella salina and its. Eng. (2013), http://dx.doi.org/10.1016/j.jece.2013.08.016
S. Duraiarasan, V. Mani / Journal of Environmental Chemical Engineering xxx (2013) xxx–xxx4
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tween the variables. An elliptical plot illustrates greaternificance of interaction whereas a circular contour plot indicates
at the interaction is negligible [30]. Mutual interactions andtimization of the tested variables could be conveniently studiedrough 3-D surface and contour plots. From the graphicalpresentation the effects of interactions were studied.
. 1. Response surface and contour plot representing the bioflocculation on variou
perature and incubation time (C), temperature and bioflocculant size (D), tempe
flocculant size (G), pH and cationic inducer concentration (H), incubation time an
Please cite this article in press as: S. Duraiarasan, V. Mani, Influence
optimization using response surface methodology, J. Environ. Chem
The response surface and contour plots indicated the mutualeffects of the studied variables on flocculating the microalgal cells.Effect of temperature along with other parameters namely pH,incubation time, bioflocculant size and cationic inducer concen-tration showed that at a maximum temperature of 30 8C (codedlevel 0), highest efficiency of 98.21% was obtained when other
s interactions between the five independent variables – temperature and pH (A),
rature and cationic inducer concentration (E), pH and incubation time (F), pH and
d bioflocculant size (I), bioflocculant size and cationic inducer concentration (J).
of bioflocculation parameters on harvesting Chlorella salina and its. Eng. (2013), http://dx.doi.org/10.1016/j.jece.2013.08.016
Table 3Comparison of different harvesting methods and their efficiencies.
Method Microalgae Harvesting References
S. Duraiarasan, V. Mani / Journal of Environmental Chemical Engineering xxx (2013) xxx–xxx 5
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parameters ranged between 6–10, 2–10 h, 0.1–0.5 ml and 0.01–0.05 mM respectively (Fig. 1A–D). The maximum flocculationefficiency was also obtained when pH was maintained at 10 whileother factors were at their respective ranges (Fig. 1E–G). Highestbioflocculation of 98.21% was noted when processing at 6 h ofincubation on maintaining a bioflocculant size of 0.3 ml andcationic inducer concentration of 0.03 mM (Fig. 1 H–J).
By analyzing the response surface plots and contour represen-tation, the optimal values of the tested variables for the highestbioflocculation efficiency were temperature 30.638 C, pH 10.4,
Table 2ANOVA table for response surface function on bioflocculation efficiency.
DF, degree of freedom; Seq SS, sequential sum of squares; Adj SS, adjusted sum of
squares; F, Fischer’s test; p, probability value
Please cite this article in press as: S. Duraiarasan, V. Mani, Influence ooptimization using response surface methodology, J. Environ. Chem
flocculation time 6.2 h, bioflocculant size 0.34 ml and cationicinducer concentration 0.031 mM.
This model also exhibited that the interaction betweenvariables were also highly significant. The coefficient X1X2, X1X5,X2X4, X2X5, X3X4 (p < 0.05) were found to be highly significant
efficiency (%)
Bioflocculation with
whole cell
Chlorella salina >98 Current
study
Flocculation with
cationic polymer
Chlorococcum sp. >89 [8]
Flocculation with AlCl3 Chlorella minutissima >90 [14]
Flocculation with
g-polyglutamic acid
Chlorella vulgaris and
Nannochloropsis oculata
>90 [25]
Increasing pH Chlorella vulgaris 95 [26]
Flocculation with
polyelectrolytes
Chaetoceros calcitrans >90 [31]
Flocculation with
chitosan
Thalassiosira pseudonana 90 [32]
Centrifugation Phaeodactylum tricornutum 94 [32]
Increasing pH Dunaliella tertiolecta 90 [33]
f bioflocculation parameters on harvesting Chlorella salina and its. Eng. (2013), http://dx.doi.org/10.1016/j.jece.2013.08.016
S. Duraiarasan, V. Mani / Journal of Environmental Chemical Engineering xxx (2013) xxx–xxx6
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odel terms. The validation of the model was done by carrying out triplicates under optimized conditions. The mean value obtainedas 98.66%, which was in good agreement with the predictedsponse, 98.53%.
Our current findings indicated that the bioflocculation processder the optimized conditions gave the maximum efficiency as.66% whereas various other flocculation procedures produced
wer results (Table 3). Thus, our study concluded that biofloccu-tion using live whole bacterial cells producing exopolysaccharideoflocculant is highly efficient to harvest microalgal cells forodiesel production.
nclusion
An ecofriendly approach of flocculation was carried out usingicrobial source marine bacterium B. subtilis as bioflocculantent, for harvesting marine microalga, C. salina. Though bio-cculant helped in efficient flocculation, a chemical flocculantCl2, at a lower concentration of 0.031 mM was an essentialquirement as a cationic inducer to enhance the process. Therameters were successfully optimized using response surfaceethodology (RSM) and a maximum efficiency of 98.66% wastained with 0.34 ml of bioflocculant size. This experimentmonstrated that bioflocculant could be a natural and potentialurce and a cost effective method to harvest microalgae forodiesel production.
ferences
] X. Miao, Q. Wu, Biodiesel production from heterotrophic microalgal oil, Bioresour.Technol. 97 (2006) 841–846.
] S.T. Keera, S.M. El Sabagh, A.R. Taman, Transesterification of vegetable oil tobiodiesel fuel using alkaline catalyst, Fuel 90 (2011) 42–47.
] L. Gouveia, A.C. Oliveira, Microalgae as a raw material for biofuels production,J. Ind. Microbiol. Biotechnol. 36 (2009) 269–274.
] L. Xin, H. Hong-ying, Z. Yu-ping, Growth and lipid accumulation properties of afreshwater microalga Scenedesmus sp. under different cultivation temperature,Bioresour. Technol. 102 (2011) 3098–3102.
] S.A. Khan, Rashmi, Z. Mir Hussain, S. Prasad, U.C. Banerjee, Prospects of biodieselproduction from microalgae in India, Renew. Sustain. Rev. 13 (2009) 2361–2372.
] R. Halim, B. Gladman, M.K. Danquah, P.A. Webley, Oil extraction from microalgaefor biodiesel production, Bioresour. Technol. 102 (2011) 178–185.
] S. Salim, R. Bosma, M.H. Vermue, R.H. Wijffels, Harvesting of microalgae by bio-flocculation, J. Appl. Phycol. 23 (2011) 849–855.
] N. Uduman, Y. Qi, M.K. Danquah, A.F.A. Hoadley, Marine microalgae flocculationand focused beam reflectance measurement, Chem. Eng. J. 162 (2010) 935–940.
] A.L. Ahmed, N.H. Mat Yasin, C.J. Lim, Microalgae as a sustainable energy source forbiodiesel production: a review, Renew. Sustain. Energy Rev. 15 (2011) 584–593.
] D. Vandamme, I. Foubert, B. Meesschaert, K. Muylaert, Flocculation of microalgaeusing cationic starch, J. Appl. Phycol. 22 (2010) 525–530.
] X. Zhang, Q. Hu, M. Sommerfeld, E. Puruhito, Y. Chen, Harvesting algal biomassfor biofuels using ultrafiltration membranes, Bioresour. Technol. 101 (2010)5297–5304.
Please cite this article in press as: S. Duraiarasan, V. Mani, Influence
optimization using response surface methodology, J. Environ. Chem
[12] A.K. Lee, D.M. Lewis, P.J. Ashman, Microbial flocculation, a potentially low-costharvesting technique for marine microalgae for the production of biodiesel,J. Appl. Phycol. 21 (2009) 559–567.
[13] E. Poelman, N. De Pauw, B. Jeurissen, Potential of electrolytic flocculation forrecovery of micro-algae, Resour. Conserv. Recy. 19 (1997) 1–10.
[14] P. Papazi, P. Makridis, Divanach, Harvesting Chlorella minutissima using cellcoagulants, J. Appl. Phycol. 22 (2010) 349–355.
[15] L. Borges, J.A. Moron-Villarreyes, M.G. Montes D’Oca, P.C. Abreu, Effects offlocculants on lipid extraction and fatty acid composition of the microalgaeNannochloropsis oculata and Thalassiosira weissflogii, Biomass Bioenerg. 35(2011) 4449–4454.
[16] J.R. Benemann, CO2 mitigation with microalgae systems, Energy Conver. Manage.38 (Suppl.) (1997) S475–S479.
[17] R.M. Knuckey, M.R. Brown, R. Robert, D.M.F. Frampton, Production of microalgalconcentrates by flocculation and their assessment as aquaculture feeds, Aquacult.Eng. 35 (2006) 300–313.
[18] C.M.L. Lapa Teixeira, F.V. Kirsten, P.C.N. Teixeira, Evaluation of Moringa oleiferaseed flour as a flocculating agent for potential biodiesel producer microalgae,J. Appl. Phycol. 24 (2012) 557–563.
[19] H. Yokoi, T. Arima, J. Hirose, S. Hayashi, Y. Takasaki, Flocculation properties ofpoly(g-glutamic acid) produced by Bacillus subtilis, J. Ferment. Bioeng. 82 (1)(1996) 84–87.
[20] C. Ganesh Kumar, H.S. Joo, Rajesh Kumar, J.W. Choi, C.S. Chang, Characterization ofan extracellular biopolymer flocculant from a haloalkalophilic Bacillus isolate,World J. Microbiol. Biotechnol. 20 (2004) 837–843.
[21] Y. Zheng, Z.L. Ye, X.L. Fang, Y.H. Li, W.M. Cai, Production and characteristics ofa bioflocculant produced by Bacillus sp. F19, Bioresour. Technol. 99 (2008)7686–7691.
[22] Y. Pan, B. Shi, Y. Zhang, Research on flocculation property of bioflocculant PG. a21Ca, Mod. Appl. Sci. 3 (6) (2009) 106–112.
[23] C. Zhu, C. Chen, L. Zhao, Y. Zhang, J. Yang, L. Song, S. Yang, Bioflocculant producedby Chlamydomonas reinhardtii, J. Appl. Phycol. (2011), http://dx.doi.org/10.1007/s108-011-9769x.
[24] H.M. Oh, S.J. Lee, M.H. Park, H.S. Kim, H.C. Kim, J.H. Yoon, G.S. Kwon, B.D. Yoon,Harvesting of Chlorella vulgaris using a bioflocculant from Paenibacillus sp. AM49,Biotechnol. Lett. 23 (2001) 1229–1234.
[25] H. Zheng, Z. Gao, J. Yin, X. Tang, X. Ji, H. Huang, Harvesting of microalgae byflocculation with poly(g-glutamic acid), Bioresour. Technol. 112 (2012) 212–220.
[26] D. Vandamme, I. Foubert, I. Fraeye, B. Meesschaert, K. Muylaert, Flocculation ofChlorella vulgaris induced by high pH: Role of magnesium and calcium andpractical implications, Bioresour. Technol. 105 (2012) 114–119.
[27] D.G. Kim, H.J. La, C.Y. Ahn, Y.H. Park, H.M. Oh, Harvest of Scenedesmus sp. withbioflocculant and reuse of culture medium for subsequent high-density cultures,Bioresour. Technol. 102 (2011) 3163–3168.
[28] D. Surendhiran, M. Vijay, Effect of different flocculants on harvesting of haloto-lerant microalgae Chlorella salina for biodiesel production, Int. J. Green. Chem.Bioproc. 3 (1) (2013) 6–11.
[29] D.C. Sobeck, M.J. Higgins, Examination of three theories for mechanisms of cation-induced bioflocculation, Water Res. 36 (2002) 527–538.
[30] X. Song, X. Zhang, C. Kuang, L. Zhu, N. Guo, Optimization of fermentationparameters for the biomass and DHA production of Schizochytrium limacinumOUC88 using response surface methodology, Process Biochem. 42 (2007)1391–1397.
[31] Z.T. Harith, F.M. Yusoff, M.S. Mohamed, M.S. Mohamed Din, A.B. Ariff, Effect ofdifferent flocculants on the flocculation performance of microalgae, Chaetoceroscalcitrans, cells, Afr. J. Biotechnol. 8 (21) (2009) 5971–5978.
[32] M. Heasman, J. Diemar, W. O’Connor, T. Sushames, L. Foulkes, J.A. Nell, Develop-ment of extended shelf-life microalgae concentrate diets harvested by centrifu-gation for bivalve molluscs—a summary, Aquacult. Res. 31 (2000) 637–659.
[33] J.I. Horiuchi, I. Ohba, K. Tada, M. Kobayashi, T. Kanno, M. Kishimoto, Effective cellharvesting of the halotolerant microalga Dunaliella tertiolecta with pH control,J. Biosci. Bioeng. 95 (2003) 412415.
of bioflocculation parameters on harvesting Chlorella salina and its. Eng. (2013), http://dx.doi.org/10.1016/j.jece.2013.08.016
