Microalgae have recently emerged as an attractive source of products for the food, energy and pharmaceutical industries due to their high biomass productivity, metabolic versatility and environmental sustainability. This work focuses on the effect of flocculation on the harvesting of microalgae Nannochloropsissp. Through a three-factor experimental design. The most suitable conditions for flocculation were determined based on flocculant (AlCl3) dosage, pH and culture time. In addition, flocculant adding-pH adjustment methods were studied and harvesting was performed according to the method that provided the best results. The pH adjustment after flocculant addition showed higher flocculation efficiency (above 90%) compared to the reverse process. It was also found that the microalgae cells exhibited spontaneous sedimentation, suggesting that the use of flocculant is not required for biomass collection

Barajas, Paola Andrea Sanguino

Díaz, Manuel Alejandro Ortegón

González-Delgado, Ángel Darío

Lavecchia, Roberto

López, Nelson Osvaldo Mena

Solano, Andrés Fernando Barajas

Zuorro, Antonio

English

López, Nelson Osvaldo Mena

Díaz, Manuel Alejandro Ortegón

González-Delgado, Ángel Darío

Solano, Andrés Fernando Barajas

Archivio della ricerca- Università di Roma La Sapienza

                                VOL. 13, NO. 15, AUGUST 2018                                                                                                     ISSN 1819-6608 ARPN Journal of Engineering and Applied Sciences ©2006-2018 Asian Research Publishing Network (ARPN). All rights reserved.  www.arpnjournals.com      4475 EVALUATION OF THE EFFECT OF FLOCCULATION ON HARVESTING OF MICROALGAE NANNOCHLOROPSIS SP  Nelson Osvaldo Mena López1, Manuel Alejandro Ortegón Díaz1, Ángel Darío González-Delgado2, Paula Andrea Sanguino Barajas3, Andrés Fernando Barajas Solano3, Roberto Lavecchia4 and Antonio Zuorro4 1Industrial University of Santander, Colombia 2University of Cartagena, Colombia 3Universidad Francisco de Paula Santander UFPS, Colombia 4Sapienza University, Department of Chemical Engineering, Materials and Environment, Roma, Italy E-Mail: agonzalezd1@unicartagena.edu.co  ABSTRACT Microalgae have recently emerged as an attractive source of products for the food, energy and pharmaceutical industries due to their high biomass productivity, metabolic versatility and environmental sustainability. This work focuses on the effect of flocculation on the harvesting of microalgae Nannochloropsissp. Through a three-factor experimental design. The most suitable conditions for flocculation were determined based on flocculant (AlCl3) dosage, pH and culture time. In addition, flocculant adding-pH adjustment methods were studied and harvesting was performed according to the method that provided the best results. The pH adjustment after flocculant addition showed higher flocculation efficiency (above 90%) compared to the reverse process. It was also found that the microalgae cells exhibited spontaneous sedimentation, suggesting that the use of flocculant is not required for biomass collection.   Keywords: microalgae, harvesting, flocculation, sedimentation, pH.  INTRODUCTION Current industrial society depends on conventional energy sources like coal, natural gas, oil and uranium, which lead to serious environmental problems and depletion of resources(Kalidasan et al., 2015). Environmental and economic sustainability issues strongly support the use of renewable resources (El-Mekkawi et al., 2016). Microalgae are attracting enormous attention from the scientific community worldwide as renewable feedstock for energy production (Sunar et al., 2016). Microalgae are single-celled or colonial photosynthetic organisms that are naturally present in different aquatic/humid environments, including rivers, lakes, oceans and soils (Brasil et al., 2016). They have the advantages of higher productivities, compared to oil seeds, no competition with food crops and ability to grow under extreme conditions (Collotta et al., 2017). Microalgae can also be regarded as an attractive source for producing valuable bioproducts, such as polyunsaturated fatty acids, carotenoids, phycobiliproteins, polysaccharides and phycotoxin (Sathasivam et al., 2017). As energy source, the high yield of biofuel produced from microalgae relies on the large amounts of neutral lipids accumulated in microalgae cells, which can further be increased by acting on growth conditions (Zakariah et al., 2017).  Biofuel production from microalgae involves biomass recovery from the culture medium, followed by the extraction of lipids or other components. Many strategies have been proposed for harvesting, including centrifugation(Knuckey et al., 2006), filtration (Zhang et al., 2010), flotation(Chen, Liu, & Yih-Hsu, 1998) and flocculation(Sari & Purnawan, 2016). Similarly, the recovery of intracellular components by efficient techniques is the subject of intense research (Zuorro et al., 2016a and b; Maffei et al., 2018). Flocculation is considered the most suitable option for concentration of the biomass because it allows the collection oflarge amounts of biomass at low cost and can be easily scale scaled(Borges et al., 2011). The mechanism of flocculation involves charge neutralization, in which positively charged flocculants are attracted to the negatively charged colloids via electrostatic interaction, followed by the formation and growth of flocs and final agglomeration (Kim et al., 2017). Flocculants are classified into inorganic flocculants, as polyaluminum chloride (PAC), synthetic organic flocculants, as polyacrylamide derivatives, biomaterials, as chitosan, and nanoparticlessuch as nanomagnetite (Sari & Purnawan, 2016). For more practical applications, flocculation using metallic coagulant such as aluminum and ferric salts, commonly known as inorganic chemical flocculation, is considered the most promising method because of its simplicity and low energy requirement (Kim et al., 2017). It has been reported that flocculation by metallic salts can be carried out under acidic or alkaline conditions (Garzón-Sanabria et al., 2012). According to Uduman et al. (2010), metallic salts of iron or aluminum are preferred for acidic pH conditions. In addition, the effectiveness of sedimentation with this type of flocculant depends on the initial biomass concentration and dose (Garzón-Sanabria et al., 2012). Rwehumbiza et al. (2012) evaluated the effects of aluminum as flocculant on the extraction of lipids and methyl esters of fatty acids for producing biodiesel from Nannochloropsissp., obtaining efficiencies between 79 and 99%. After extraction and conversion of lipids to biodiesel, with an initial biomass concentration of 20 g/L, it was found that aluminum was not detectable and at lower initial biomass concentrations it decreased two to three times.                                 VOL. 13, NO. 15, AUGUST 2018                                                                                                     ISSN 1819-6608 ARPN Journal of Engineering and Applied Sciences ©2006-2018 Asian Research Publishing Network (ARPN). All rights reserved.  www.arpnjournals.com      4476 In order to provide further insight into the role and impact of flocculation on microalgae processing, this work investigated the effects of flocculation with metallic AlCl3 on biomass collection from Nannochloropsis sp.    METHODOLOGY   Culture methods  Nannochloropsis sp. was isolated from Gulf of Morrosquillo (Sucre-Córdoba, Colombia), which was cultivated in Bold Basal medium. Main components of this medium are (mg/L): NaNO3 (2.94), MgSO4·7H2O (3.04×10–1) NaCl (4.28×10–1), K2HPO4 (4.31×10–1), KH2PO4 (1.29), CaCl2·2H2O (1.70×10–1), H3BO3 (1.85×10–1), EDTA (1.71×10–1), KOH (5.53×10–1) and FeSO4·7H2O (1.79×10–2). The microalgae were kept growing in 12 L glass reactor equipped with a bubble aeration system and light-dark cycle 12:12 h.  Experimental design Experimental design was performed using STATISTICA 7.0 program based on the procedure described by Garzón-Sanabria et al. (2012). Factors considered in this study are shown in Table-1: culture time (5, 10 and 15 days), pH (4, 7 and 10) and concentration of flocculant (50, 100 and 150 mg/L).  Table-1. Experimental design layout.  Test Culture time (days) Dosage of flocculant (mg/L) pH 1 1.63 100 7 2 5 50 4 3 5 50 10 4 5 150 4 5 5 150 10 6 10 16.3 7 7 10 100 1.9 8 10 100 7 9 10 100 7 10 10 100 7 11 10 100 12 12 10 183.6 7 13 15 50 4 14 15 50 10 15 15 100 4 16 15 100 10 17 18.3 100 7  Determination of optimal conditions for flocculant adding-pH adjustment  Preliminary experiments were made to evaluate the optimal flocculation procedure (pH adjustment before or after adding flocculant). A solution of 40 g/L of flocculant agent (AlCl3) was prepared and then diluted to obtain aliquots of 50, 100 and 150 mg/L, which were added to 100 mL of sample. The pH adjustment was performed with 1M HCl and 1 M NaOHto get values of 4, 7 and 10. Samples were stirred at 500 rpm for 2 min, followed by slower mixing at 60 rpm for 15 minin order to allow the formation of aggregate and hydrolysis of flocculant. Finally, the pH was measured and samples of 10 mL were collected at 1-h intervals during 5 h to quantify the optical density of Nannochloropsis sp. at 750 nm.  Flocculation tests Microalgae were cultivated with an inoculum of 2.5 g/L of Nannochloropsis sp. wet biomass. Tests were carried out as shown in Figure-1. An optimal method was used to determine the initial and final absorbances, with the efficiency calculated according to Equation-1.   Efficiency (%) = (1 −final absorbanceinitial absorbance)  × 100            (1)    Figure-1. Experimental procedure.  RESULTS   Addition of AlCl3-pH adjustment Figure-2 shows the trends of pH with time for evaluating the best conditions for flocculant addition and pH adjustment with the two methods previously described. As can be seen, the pH remained approximately constant when flocculation occurred; therefore, the method chosen for developing the experimental design was the one providing a higher efficiency.                                    VOL. 13, NO. 15, AUGUST 2018                                                                                                     ISSN 1819-6608 ARPN Journal of Engineering and Applied Sciences ©2006-2018 Asian Research Publishing Network (ARPN). All rights reserved.  www.arpnjournals.com      4477   Figure-2. pH behaviour as a function of time: pH adjustment before (a) and after (b) addition of AlCl3.  Figure-3 shows that the flocculation efficiency was higher (values above 90%) when adjusting the pH after addition of flocculant. Other studies have also reported high efficiencies for harvesting Nannochloropsis sp. with different flocculants assummarized in Table-2. The maximum flocculation efficiency obtained in this work is similar to that reported by Garzon-Sanabria et al. (2013), who also used an aluminum salt at different flocculant dosages.The optimal flocculant dosage for achieving the highest efficiency on removal of the microalgal cells from the culture was 100 mg/L. The variations of efficiency with flocculant dosage are due to the microalgal cell surface charge, which affects the stability of the cell suspension. Cationic flocculants such as aluminum salts neutralize the surface charge on the cells to facilitate the formation of flocs; hence, the dosage of flocculant required to produce effective flocculation depends on surface charge(Chatsungnoen & Chisti, 2016).     Figure-3. Comparison of flocculation efficiency for two methods of flocculant addition and pH adjustment.  Table-2. Summary of different flocculants successfully used for harvesting Nannochloropsis sp.  Flocculant Maximum Recovery (%) Flocculant dosage Reference Chitosan 90 100 mg/L (Sadegh et al., 2013) Tanfloc 98 10 mg/L (Roselet et al., 2016) AlCl3 98 50 mg/L (Garzon-Sanabria et al., 2013) Bioflocculant from Solibacillussilvestris 90 0.1 % (w/v) (Wan et al., 2013) AlCl3 97 100 mg/L This work  Flocculation tests  According to the tests carried out for experimental design with Nannochloropsis sp., spontaneous flocculation occurred in all the experiments. For samples corresponding to 1.6 and 5 days, an attempt was made to measure the flocculation by spectrophotometry, however, this procedure was not possible to perform because cells already had agglomerated before starting samples preparation since precipitation occurred immediately. For 10, 15 and 18.3 days, tests were carried out according to experimental design and without flocculant as control; all the tests were photographed (not shown). The recording of control samples (Figure-4) shows that incorporating flocculant was not necessary and pH did not have a relevant influence on sedimentation process, since the precipitation was the same in all experiments.                                   VOL. 13, NO. 15, AUGUST 2018                                                                                                     ISSN 1819-6608 ARPN Journal of Engineering and Applied Sciences ©2006-2018 Asian Research Publishing Network (ARPN). All rights reserved.  www.arpnjournals.com      4478   (a)    (b)   (c)  Figure-4. Control samples for the experimental design. (a) 10 days of culture; (b) 15 days of culture; (c) 18.3 days of culture. (1) Freshly stirred sample; (2) Samples after some seconds of agitation.  It is evident that the performance of the biomass used for experimental design and for best flocculant adding-pH adjustment method is different. The growth was different for both strains, since in the first there was the formation of aggregates facilitating the sedimentation; thisis becausethe cultivation of this strain was carried out in fresh water and not in marine water, where this species of microalga usually grows.  CONCLUSIONS This work investigated the effects of flocculation on harvesting Nannochloropsis sp. High flocculation efficiencies (above 90%) were obtained, suggesting that flocculation is a suitable method for biomass recovery from the culture medium. However, microalgae culture exhibited spontaneous sedimentation; hence, the use of flocculant was not required for biomass collection. In addition, pH adjustment after adding flocculant indicated higher flocculation efficiencies compared to the reverse method.   ACKNOWLEDGMENT The authors thank the Industrial University of Santander, Universidad Francisco de Paula Santander, University of Cartagena and Sapienza University of Rome for supporting this research.    REFERENCES  Borges L., Morón–Villarreyes J., Montes D’Oca M. & Abreu P. 2011. Effects of flocculants on lipid extraction and fatty acid composition of the microalgae Nannochloropsis oculata and Thalassiosira weissflogii. Biomass and Bioenergy. 31, 4449-4454.  Brasil B.S.A., Silva F.C.P. & Siqueira F.G. 2016. Microalgae biorefineries: The Brazilian scenario in perspective. New Biotechnology.   Chatsungnoen T. & Chisti Y. 2016. Harvesting microalgae by flocculation-sedimentation. Algal Research. 13, 271-283.   Chen Y., Liu J. & Yih-Hsu J. 1998. Flotation removal of algae from water. Colloids and Surfaces. Biointerfaces. 12, 49-55.  Collotta M., Champagne P., Mabee W., Tomasoni G., Gustavo B., Busi L. & Alberti M. 2017. Comparative LCA of flocculation for the harvesting of microalgae for biofuels production. Procedia CIRP. 61, 756-760.   El-Mekkawi S.A., El-Ardy O.A., Abdelmonem N.M. & Elahwany A.H. 2016. A scope on microalgae as potential source of biofuel. ARPN Journal of Engineering and Applied Sciences. 11(19): 11421-11432.  Garzon-Sanabria A.J., Ramirez-Caballero S.S., Moss F.E.P. & Nikolov Z.L. 2013. Effect of algogenic organic matter (AOM) and sodium chloride on Nannochloropsissalina flocculation efficiency. Bioresource Technology. 143, 231-237.                                   VOL. 13, NO. 15, AUGUST 2018                                                                                                     ISSN 1819-6608 ARPN Journal of Engineering and Applied Sciences ©2006-2018 Asian Research Publishing Network (ARPN). All rights reserved.  www.arpnjournals.com      4479 Garzón-Sanabria A.J., Davis R.T. & Nikolov N.L. 2012. Harvesting Nannochloris oculata by inorganic electrolyte flocculation: Effect of initial cell density, ionic strength, coagulant dosage, and media pH. Bioresource Technology. 118, 418-424.  Kalidasan B., Srinivas T. & Shankar R. 2015. Experimental study on power generation using biomass based and solar based brayton cycles. ARPN Journal of Engineering and Applied Sciences. 10(9): 3987-3990.  Kim D., Lee K., Lee J., Lee Y., Han J., Park J. & Oh Y. 2017. Acidified-flocculation process for harvesting of microalgae: Coagulant reutilization and metal-free-microalgae recovery. Bioresource Technology. 239, 190-196.   Knuckey R., Brown M., Robert R. & Frampton D. 2006. Production of microalgal concentrates by flocculation and their assessment as aquaculture feeds. Agricultural Engineering. 35, 300-313.  Maffei G., Bracciale M.P., Broggi A., Zuorro A., Santarelli M.L. & Lavecchia R. 2018. Effect of an enzymatic treatment with cellulase and mannanase on the structural properties of Nannochloropsis microalgae.Bioresource Technology. 249, 592-598.  Roselet F., Burkert J. & Cesar P. 2016. Flocculation of Nannochloropsis oculata using a tannin-based polymer: Bench scale optimization and pilot scale reproducibility. 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Evaluation of the effect of flocculation on harvesting of microalgae Nannochloropsis sp

Microalgae have recently emerged as an attractive source of products for the food, energy and pharmaceutical
industries due to their high biomass productivity, metabolic versatility and environmental sustainability. This work focuses
on the effect of flocculation on the harvesting of microalgae Nannochloropsissp. Through a three-factor experimental
design. The most suitable conditions for flocculation were determined based on flocculant (AlCl3) dosage, pH and culture
time. In addition, flocculant adding-pH adjustment methods were studied and harvesting was performed according to the
method that provided the best results. The pH adjustment after flocculant addition showed higher flocculation efficiency
(above 90%) compared to the reverse process. It was also found that the microalgae cells exhibited spontaneous
sedimentation, suggesting that the use of flocculant is not required for biomass collection

ZUORRO, Antonio

Barajas Solano, andres F

Mena, Nelson Osvaldo

González-Delgado, Angel Darío

Ortegón, Manuel Alejandro

Sanguino, Paola Andrea

Repositorio Universidad Francisco de Paula Santander

Evaluation of the effect of flocculation on harvesting of microalgaenannochloropsis SP

https://iris.uniroma1.it/bitstream/11573/1182057/1/Lopez_Evaluation-effect-flocculation_2018.pdf

Evaluation of the effect of flocculation on harvesting of microalgae Nannochloropsis sp

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Archivio della ricerca- Università di Roma La Sapienza

Repositorio Universidad Francisco de Paula Santander