Primary Separation of Antioxidants (Unsaponifiables) the Wet Biomass Microalgae Chlamydomonas sp. and Production of the Biodiesel
Author(s)
Gisel Chenard Díaz1,
Yordanka Reyes Cruz1,
Mariana Monteiro Fortes1,
Carolina Vieira Viegas1,
René González Carliz1,
Nelson C. Furtado2,
Donato A. Gomes Aranda1
Affiliation(s)
1 Greentec Laboratory, School of Chemistry, Federal University of Rio de Janeiro, Rio de Janeiro, Brazil. 2 Brazilian Center for Physical Researches, Rio de Janeiro, Brazil.
1 Greentec Laboratory, School of Chemistry, Federal University of Rio de Janeiro, Rio de Janeiro, Brazil. 2 Brazilian Center for Physical Researches, Rio de Janeiro, Brazil.
ABSTRACT
This work studies the saponification which directs the wet biomass of algae Chlamydomonas sp. like a previous stage to production of biodiesel. This stage allows the obtainment of fatty acids to produce biodiesel, instead of the gross lipid fraction. In addition of the fatty acids, utilizing the same process one can also obtain the fraction unsaponifiable, these are soluble in apolar solvents and contain mainly carotenoids that can take action as antioxidants and photoprotectors, as they reduce the oxidation of unsaturated fatty acids. The saponification direct and extraction of fatty acids from biomass is faster and reduces the time and cost of operation. The separation of unsaponifiable matter from the biomass humid of microalgae Chlamydomonas sp., was held according to the method AOCS (Ca 6a-40), using extraction Liquid-liquid with hexane as solvent. Subsequently, phase hydroalcoholic or from soap, containing fatty acids, was acidified by addition of H2SO4 and the fatty acids were recovered by the addition of hexane. After acidulation of the soap, necessary for obtaining of the fatty acids was performed the stage of esterification for obtaining of biodiesel. The operating conditions were: molar ratio fatty acid:methanol (1:10), as catalyst 8% H2SO4 calculated in relation to the mass of fatty acid, 200℃ and reaction time of 90 minutes. The content of methyl esters was 96.8% determined by gas chromatography according to standard EN14103. The quality of biodiesel produced from wet biomass of Chlamydomonas sp. is according to the specification established by standard EN 14214 and RANP No. 14. For the identification of the composition the unsaponifiable fraction was used the method of High Performance Liquid Chromatography (HPLC). The composition of the material unsaponifiable found was of: Carotenoids total (0.76%); Lutein (0.45%); Zeaxanthin (0.07%); α-carotene (0.05%); β-carotene (0.11%); 13 cisβ-carotene (0.05%) and 9-cisβ-carotene (0.03%).
This work studies the saponification which directs the wet biomass of algae Chlamydomonas sp. like a previous stage to production of biodiesel. This stage allows the obtainment of fatty acids to produce biodiesel, instead of the gross lipid fraction. In addition of the fatty acids, utilizing the same process one can also obtain the fraction unsaponifiable, these are soluble in apolar solvents and contain mainly carotenoids that can take action as antioxidants and photoprotectors, as they reduce the oxidation of unsaturated fatty acids. The saponification direct and extraction of fatty acids from biomass is faster and reduces the time and cost of operation. The separation of unsaponifiable matter from the biomass humid of microalgae Chlamydomonas sp., was held according to the method AOCS (Ca 6a-40), using extraction Liquid-liquid with hexane as solvent. Subsequently, phase hydroalcoholic or from soap, containing fatty acids, was acidified by addition of H2SO4 and the fatty acids were recovered by the addition of hexane. After acidulation of the soap, necessary for obtaining of the fatty acids was performed the stage of esterification for obtaining of biodiesel. The operating conditions were: molar ratio fatty acid:methanol (1:10), as catalyst 8% H2SO4 calculated in relation to the mass of fatty acid, 200℃ and reaction time of 90 minutes. The content of methyl esters was 96.8% determined by gas chromatography according to standard EN14103. The quality of biodiesel produced from wet biomass of Chlamydomonas sp. is according to the specification established by standard EN 14214 and RANP No. 14. For the identification of the composition the unsaponifiable fraction was used the method of High Performance Liquid Chromatography (HPLC). The composition of the material unsaponifiable found was of: Carotenoids total (0.76%); Lutein (0.45%); Zeaxanthin (0.07%); α-carotene (0.05%); β-carotene (0.11%); 13 cisβ-carotene (0.05%) and 9-cisβ-carotene (0.03%).
Cite this paper
Díaz, G. , Cruz, Y. , Fortes, M. , Viegas, C. , Carliz, R. , Furtado, N. and Aranda, D. (2014) Primary Separation of Antioxidants (Unsaponifiables) the Wet Biomass Microalgae Chlamydomonas sp. and Production of the Biodiesel. Natural Science, 6, 1210-1218. doi: 10.4236/ns.2014.615108.
Díaz, G. , Cruz, Y. , Fortes, M. , Viegas, C. , Carliz, R. , Furtado, N. and Aranda, D. (2014) Primary Separation of Antioxidants (Unsaponifiables) the Wet Biomass Microalgae Chlamydomonas sp. and Production of the Biodiesel. Natural Science, 6, 1210-1218. doi: 10.4236/ns.2014.615108.
References
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http://dx.doi.org/10.1016/S0167-7799(00)01433-5 [7] Pulz, O. (2004) Photobioreactors: Production Systems for Phototrophic Microorganisms. Applied Microbiology and Biotechnology, 57, 287-293. [8] Bligh, G. and Dyer, W. (1959) A Rapid Method for Total Lipid Extraction and Purification. Canadian Journal of Biochemistry and Physiology, 37, 911-917.
http://dx.doi.org/10.1139/o59-099 [9] Yoo, C., Jun, S.Y. and Lee, J.Y. (2010) Selection of Microalgae for Lipid Production under High Levels Carbon Dioxide. Bioresource Technology, 101, S71-S74.
http://dx.doi.org/10.1016/j.biortech.2009.03.030 [10] Pacheco, S. (2009) Preparo de padrões analíticos, estudo de estabilidade e parâmetros de validação para ensaio de carotenóides por cromatografia líquida. UFRRJ, Dissertação, Mestrado em Ciência e Tecnologia de Alimentos, Ciência dos Alimentos, Seropédica, 106 p. [11] Tsigankov, A.A., Kosourov, S.N., Tolstygina, I.V., Ghirardi, M.L. and Seibert, M. (2006) Hidrogen Production by Sulfur-Deprived Chlamydomonas reinhardtii under Photoautotrophic Conditions. Internacional Journal of Hidrogen Energy, 31, 1574-1584. [12] Mattox, K.R. and Stewart, K.D. (1984) Classification of the Green Algae: A Concept Based on Comparative Cytology. In: Irvine, D.E.G. and John, D., Eds., Systematics of the Green Algae, Academic Press, London, 41, 42, 57, 58. [13] Becker, W. (2004) Microalgae in Human and Animal Nutrition. In: Richmond, A., Ed., Handbook of Microalgal Culture: Biotechnology and Applied Phycology, Blackwell Science, London, 566 p. [14] Carolino, L. do R.V.C. (2011) Cultivo de microalgas unicelulares para determinação da produção lipídica e sequestro de carbono. ULisboa Faculdade de Ciências Departamento de Biologia Vegetal, Mestrado de Biologia Celular e Biotecnologia, 91. [15] Chisti, Y. (2007) Biodiesel from Microalgae. Biotechnology Advances, 25, 294-306.
http://dx.doi.org/10.1016/j.biotechadv.2007.02.001 [16] Gouveia, L. and Oliveira, A. (2009) Microalgae as Raw Materials for Biofuels Production. Journal of Industrial Microbiology and Biotechnology, 36, 269-274.
http://dx.doi.org/10.1007/s10295-008-0495-6 [17] Deng, X., Li, Y. and Fei, X. (2009) Microalgae: A Promising Feedstock for Biodiesel. African Journal of Microbiology Research, 3, 1008-1014. [18] Colla, L.M., Bertolini, T.E. and Costa, J.A. (2004) Fatty Acids Profile of Spirulina platensis Grown under Different Temperatures and Nitrogen Concentrations. Zeitschrift fur Naturforschung, 59, 55-59. [19] Deshnium, P., Paithoonrangsarid, K. and Suphatrakul, A. (2000) Temperature-Independent and Dependent Expression of Desaturase Genes in Filamentous Cyanobacterium Spirulina platensis strain C1 (Arthrospira sp. PCC 9438). FEMS Microbiology Letters, 184, 207-213.
http://dx.doi.org/10.1111/j.1574-6968.2000.tb09015.x [20] Olguín, E., Galicia, S. and Angulo-Guerrero, O. (2001) The Effect of Low Light Flux and Nitrogen Deficiency on the Chemical Composition of Spirulina sp. (Arthrospira) Grown on Digested Pig Waste. Bioresource Technology, 77, 19-24.
http://dx.doi.org/10.1016/S0960-8524(00)00142-5 [21] Makulla, A. (2000) Fatty Acid Composition of Scenedesmus obliquus: Correlation to Dilution Rates. LimnologicaEcology and Management of Inland Waters, 30, 162-168.
http://dx.doi.org/10.1016/S0075-9511(00)80011-0 [22] Eonseon, J., Polle, J.E.W., Kumlee, H., Hyun, S.M. and Chang, M. (2003) Xanthophylls in Microalgae: From Biosynthesis to Biotechnological Mass Production and Application. Journal of Microbiology and Biotechnology, 13, 165-174. [23] Guerin, M., Huntley, M.E. and Olaizola, M. (2003) Haematococcus Astaxanthin: Application for Human Health and Nutrition. Trends in Biotechnology, 21, 210-216.
http://dx.doi.org/10.1016/S0167-7799(03)00078-7 [24] Del-Campo, J.A., Garcia-Gonzalez, M. and Guerrero, M.G. (2007) Outdoor Cultivation of Microalgae for Carotenoid Production: Current State and Perspectives. Applied Microbiology and Biotechnology, 74, 1163-1174.
http://dx.doi.org/10.1007/s00253-007-0844-9 [25] Brasil. Agência Nacional de Vigilância Sanitária. Resolução da Diretoria Colegiada-RDC No. 217, de 29 de julho de 2005. https://prosig.alvessilva.com.br/textos/5853.doc
[1] Hill, J., Nelson, E., Tilman, D., Polasky, S. and Tiffany, D. (2006) Environmental, Economic, and Energetic Costs and Benefits of Biodiesel and Ethanol Biofuels. PNAS, 103, 11206-11210.
http://dx.doi.org/10.1073/pnas.0604600103 [2] Sheehan, J., Dunahay, T., Benemann, J. and Roessler, P. (1998) A Look Back at the U.S. Department of Energy’s Aquatic Species Program—Biodiesel from Algae. National Renewable Energy Laboratory, Report NREL/TP-580-24190. [3] Patil, V., Tran, K.Q. and Giseirod, H.R. (2008) Towards Sustainable Production of Biofuels from Microalge. International Journal Molecular Sciences, 9, 1188-1195.
http://dx.doi.org/10.3390/ijms9071188 [4] Griffiths, M.J. and Harrison, S.T.L. (2009) Lipid Productivity as a Key Characteristic for Choosing Algal Species for Biodiesel Production. Journal of Applied Phycology, 21, 493-507.
http://dx.doi.org/10.1007/s10811-008-9392-7 [5] Spolaore, P., Joannis-Cassan, C., Duran, E. and Isambert, A. (2006) Commercial Applications of Microalgae. Journal of Bioscience and Bioengineering, 101, 87-96.
http://dx.doi.org/10.1263/jbb.101.87 [6] Lorenz, R.T. and Cysewski, G.R. (2003) Commercial Potential for Haematococcus microalga as a Natural Source of Astaxanthin. Trends in Biotechnology, 18, 160-167.
http://dx.doi.org/10.1016/S0167-7799(00)01433-5 [7] Pulz, O. (2004) Photobioreactors: Production Systems for Phototrophic Microorganisms. Applied Microbiology and Biotechnology, 57, 287-293. [8] Bligh, G. and Dyer, W. (1959) A Rapid Method for Total Lipid Extraction and Purification. Canadian Journal of Biochemistry and Physiology, 37, 911-917.
http://dx.doi.org/10.1139/o59-099 [9] Yoo, C., Jun, S.Y. and Lee, J.Y. (2010) Selection of Microalgae for Lipid Production under High Levels Carbon Dioxide. Bioresource Technology, 101, S71-S74.
http://dx.doi.org/10.1016/j.biortech.2009.03.030 [10] Pacheco, S. (2009) Preparo de padrões analíticos, estudo de estabilidade e parâmetros de validação para ensaio de carotenóides por cromatografia líquida. UFRRJ, Dissertação, Mestrado em Ciência e Tecnologia de Alimentos, Ciência dos Alimentos, Seropédica, 106 p. [11] Tsigankov, A.A., Kosourov, S.N., Tolstygina, I.V., Ghirardi, M.L. and Seibert, M. (2006) Hidrogen Production by Sulfur-Deprived Chlamydomonas reinhardtii under Photoautotrophic Conditions. Internacional Journal of Hidrogen Energy, 31, 1574-1584. [12] Mattox, K.R. and Stewart, K.D. (1984) Classification of the Green Algae: A Concept Based on Comparative Cytology. In: Irvine, D.E.G. and John, D., Eds., Systematics of the Green Algae, Academic Press, London, 41, 42, 57, 58. [13] Becker, W. (2004) Microalgae in Human and Animal Nutrition. In: Richmond, A., Ed., Handbook of Microalgal Culture: Biotechnology and Applied Phycology, Blackwell Science, London, 566 p. [14] Carolino, L. do R.V.C. (2011) Cultivo de microalgas unicelulares para determinação da produção lipídica e sequestro de carbono. ULisboa Faculdade de Ciências Departamento de Biologia Vegetal, Mestrado de Biologia Celular e Biotecnologia, 91. [15] Chisti, Y. (2007) Biodiesel from Microalgae. Biotechnology Advances, 25, 294-306.
http://dx.doi.org/10.1016/j.biotechadv.2007.02.001 [16] Gouveia, L. and Oliveira, A. (2009) Microalgae as Raw Materials for Biofuels Production. Journal of Industrial Microbiology and Biotechnology, 36, 269-274.
http://dx.doi.org/10.1007/s10295-008-0495-6 [17] Deng, X., Li, Y. and Fei, X. (2009) Microalgae: A Promising Feedstock for Biodiesel. African Journal of Microbiology Research, 3, 1008-1014. [18] Colla, L.M., Bertolini, T.E. and Costa, J.A. (2004) Fatty Acids Profile of Spirulina platensis Grown under Different Temperatures and Nitrogen Concentrations. Zeitschrift fur Naturforschung, 59, 55-59. [19] Deshnium, P., Paithoonrangsarid, K. and Suphatrakul, A. (2000) Temperature-Independent and Dependent Expression of Desaturase Genes in Filamentous Cyanobacterium Spirulina platensis strain C1 (Arthrospira sp. PCC 9438). FEMS Microbiology Letters, 184, 207-213.
http://dx.doi.org/10.1111/j.1574-6968.2000.tb09015.x [20] Olguín, E., Galicia, S. and Angulo-Guerrero, O. (2001) The Effect of Low Light Flux and Nitrogen Deficiency on the Chemical Composition of Spirulina sp. (Arthrospira) Grown on Digested Pig Waste. Bioresource Technology, 77, 19-24.
http://dx.doi.org/10.1016/S0960-8524(00)00142-5 [21] Makulla, A. (2000) Fatty Acid Composition of Scenedesmus obliquus: Correlation to Dilution Rates. LimnologicaEcology and Management of Inland Waters, 30, 162-168.
http://dx.doi.org/10.1016/S0075-9511(00)80011-0 [22] Eonseon, J., Polle, J.E.W., Kumlee, H., Hyun, S.M. and Chang, M. (2003) Xanthophylls in Microalgae: From Biosynthesis to Biotechnological Mass Production and Application. Journal of Microbiology and Biotechnology, 13, 165-174. [23] Guerin, M., Huntley, M.E. and Olaizola, M. (2003) Haematococcus Astaxanthin: Application for Human Health and Nutrition. Trends in Biotechnology, 21, 210-216.
http://dx.doi.org/10.1016/S0167-7799(03)00078-7 [24] Del-Campo, J.A., Garcia-Gonzalez, M. and Guerrero, M.G. (2007) Outdoor Cultivation of Microalgae for Carotenoid Production: Current State and Perspectives. Applied Microbiology and Biotechnology, 74, 1163-1174.
http://dx.doi.org/10.1007/s00253-007-0844-9 [25] Brasil. Agência Nacional de Vigilância Sanitária. Resolução da Diretoria Colegiada-RDC No. 217, de 29 de julho de 2005. https://prosig.alvessilva.com.br/textos/5853.doc