Algal Fame production with a novel surfactant based catalyst in a reactive extraction. NE1 7RU, United Kingdom.
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1 Algal Fame production with a novel surfactant based catalyst in a reactive extraction Kamoru A Salam a*, Sharon B Velasquez-Orta a, Adam P Harvey a a School of Chemical Engineering and Advanced Materials (CEAM), Newcastle University, NE1 7RU, United Kingdom. *Corresponding author kamoru.salam@ncl.ac.uk, kamar725@yahoo.com Sharon.velasquez-orta@ncl.ac.uk, adam.harvey@ncl.ac.uk INTRODUCTION During plant photosynthesis, bio-oils are produced as triglycerides which have a high combustion heat. One way of utilising this energy is through transesterification. However, the use of refined oil for transesterification is not economical as ~ 88% of total production cost of this conventional two steps biodiesel production is ascribed to refined oil feedstock [4]. Similarly biodiesel production from non-food oil crops is not sustainable. Microalgae are one of the sustainable biofuel feed stocks because of their compelling advantages. They are non-food biofuel feed stocks. They can be cultivated on non-arable land. Algae can be grown on the sea or wastewater. They can also be used for capture highly concentrated CO 2. Production of FAME from micro-algae via in situ transesterification is simple and potentially more economical since ~90% of the process energy is accounted for by the solvent extraction and drying steps in the conventional transesterification [5]. Previous studies have demonstrated the feasibility of obtaining greater FAME conversion from such in situ transesterification than from a conventional two-step approach [8, 9]. The major drawback of this method is the requirement of a large amount of methanol. This is necessary since methanol plays a dual role: it acts as an oil extractor and as a reactant. Moreover, the need to recover methanol from the products streams may add additional cost to the process. In this report the application of a synthesised zirconium dodecyl sulphate (ZDS) (a surfactant catalyst) in a reactive extraction of FAME from microalgae was investigated. The inclusion of sodium dodecyl sulphate (SDS) in H 2 SO 4 catalyst for enhancing FAME production during in situ transesterification of Nannochloropsis occulata and Chlorella vulgaris was also studied. We further investigated the potential of utilising the residual protein and carbohydrates for animal feeds or bioethanol production.
2 Procedure for in situ transesterification MATERIALS AND METHOD This is a modification of Velasquez-orta et al. [3]. The following process conditions were maintained in the in situ transesterification: 100 milligram of microalgae, temperature: 60 o C, mixing rate: 450 rpm, molar ratio of catalyst to oil: 8.5:1, molar ratio of methanol to oil: 600:1. Microalgae was first added to the reaction tube, followed by methanol and the catalyst (Either the surfactant catalyst, Sodium dodecyl sulphate and H 2 SO 4 or H 2 SO 4 catalyst). The mixture was then transferred to a shaking incubator (IKA KS 4000i control) whose temperature has been pre-set to 60 o C. The in situ transesterification was run for time ranged from 30minutes - 36hours. After each completed reaction the tube was kept in a freezer for at least 6 hours to stop the reaction. The algae residues were separated from the bulk liquid by centrifugation. The bulk liquid mixture of methanol, FAME and by-products was transferred to a pre-weighed tube. The final weight of the bulk liquid was recorded for each tube and the FAME concentration was measured by gas chromatography. Analytical techniques British standard procedure (BS EN 14103:2003) [11] was used for the FAME concentration analysis after the in situ transesterification. The gas chromatography GC was set to the following conditions: carrier gas: helium, 7psi; air pressure, 32psi; hydrogen pressure, 22psi; a capillary column head pressure was adjusted to 4.5psi. Samples of 200mg were mixed with 0.1 ml of an internal standard solution (C17:0 Sigma Aldrich, UK, 10 mg/ ml of methanol) in 2 ml vials. 2µL of the mixture was injected to the GC and data was collected using Data Apex Clarity software, UK. The column used was CP WAX 52 CB 30m 0.32mm (0.25µm) (Agilent). The mass of FAME obtained in the biodiesel rich phase from the experiments was calculated by multiplying the weight of the final biodiesel mixture obtained times the FAME concentration measured by GC. FAME yield was calculated by dividing the mass of FAME obtained by the maximum FAME available in the lipids which was measured with Garces and Mancha [6]. Where: [ ] [ ] 100% A = Total peak areas of methyl esters = Area of Internal standard = Weight of the sample (mg) = Concentration of internal standard (mg/ml) = Volume of internal standard (ml)
3 FAME YIELD % ( w/w) RESULTS AND DISCUSSION FAME yield profiles obtained during in situ transesterification of freeze dried cells of the Nannochloropsis occulata is shown in figure 1. As can be seen from the figure the FAME yields by each of the catalyst increased with increase in reaction time. This is because the rate of FAME production increased as the reactants were consumed. However, a drop in the FAME yield after 12 hours was observed with the SDS and H 2 SO 4 catalyst. This could be due to a consumption of FAME via a side reaction. A FAME reduction during acid catalysed in situ transesterification of activated sludge above 60 o C has been reported by Revellame et al. [10]. Similarly, acid catalysed oligomerisation of oleic and different saturated fatty acid at 45 o C or 55 o C has been reported by Cermark and Isbell [12]. It was also observed that the FAME yield produced by SDS and H 2 SO 4 was ~ 70% greater than that of H 2 SO 4 at 12 hour. This shows that inclusion of SDS in H 2 SO 4 catalyst leads to a significant increase in FAME yield in this species. FAME yield produced by the zirconium dodecyl sulphate (a surfactant catalyst) was also more than that of H 2 SO 4 catalyst between hours. Nannochloropsis H2SO4 H2SO4+SDS ZDS In situ transesterification time (Hr) Fig.1: Reactive Extracted FAME Yield Profiles of the Nannochloropsis Process conditions: Molar ratio of methanol to oil = 600 (470µL methanol), molar ratio of H 2 SO 4 to oil = 8.5 (8.7 µl H 2 SO 4 ), agitation = 450rpm, temperature = 60 o C, mass of microalgae = 100 mg, mass of SDS (sodium dodecyl sulphate) = 9mg, ZDS: zirconium dodecyl sulphate.
4 FAME YIELD (% w/w) Figure 2 shows the FAME yield profiles obtained during in situ transesterification of spray dried cells of Chlorella vulgaris. It was observed that the FAME yields by the three catalysts increased with increase in reaction time. However, after 4 hours the FAME yield by the ZDS catalyst decreased with increase in time. It is interesting to note that a relatively low performance of the ZDS catalyst in terms of FAME production was observed in this strain. We found in one of our experiments (the data is not shown here) that the ZDS produced greater cell wall disruption with the Nannochloropsis occulata than in Chlorella vulgaris and there was no significant difference between the transesterifiable lipids of the two microalgae. This difference in cell wall disruption property shown by the ZDS could explain why there was difference in the FAME yield produced by the catalyst in the two species. The inclusion of SDS with H 2 SO 4 also enhanced the FAME yield obtained from this species when compared to only homogeneous H 2 SO 4 catalyst. At 24 hours ~24% increase in FAME yield was obtained by sodium dodecyl sulphate and H 2 SO 4 than that of H 2 SO 4 catalyst. Chlorella 80 H2SO4 H2SO4+SDS ZDS In situ transesterification time (Hr) Fig2. : Reactive Extracted FAME Yield Profiles of the Chlorella vulgaris Process conditions: Molar ratio of methanol to oil = 600 (470µL methanol), molar ratio of H 2 SO 4 to oil = 8.5 (8.7 µl H 2 SO 4 ), agitation = 450rpm, temperature = 60 o C, mass of microalgae = 100 mg, mass of SDS (sodium dodecyl sulphate) = 9mg, ZDS: zirconium dodecyl sulphate.
5 Component (wt %) The protein and carbohydrate content of the algal residue was measured at maximum FAME yield and the results obtained are shown in the figure H2SO4 +SDS H2SO4 ZDS H2SO4 +SDS H2SO4 ZDS FAME Initial Carbohydrate Final carbohydrate Initial protein Final protein Nannochloropsis Chlorella Fig 3: Carbohydrate and protein content of the microalgae before and after in situ transesterification It can be seen from the figure that the residual biomass for both species maintained almost their original protein after the in situ transesterification. However a reduction in the carbohydrate content was observed in the residue for the two microalgae in all the catalysts investigated. This shows that part of the carbohydrate has been hydrolysed to simple sugars or other co associated products during the in situ transesterification. The protein left over could be utilised for animal feed supplement or co-products for biogas generation. The carbohydrate in the residue can also be used for bio-ethanol production. For instance, Harun et al. [7] has produced bioethanol from algae residue whose lipid has been extracted. Utilisation of the algae residue after the in situ transesterification for value added products could substantially improve the process economy.
6 CONCLUSION In this study FAME was produced from Chlorella vulgaris and Nannochloropsis occulata, via an in situ transesterification reactive extraction with methanol which was catalysed by an H 2 SO 4, a synthesised surfactant catalyst (zirconium dodecyl sulphate) or sodium dodecyl sulphate plus H 2 SO 4. The microalgae were also characterised in terms of carbohydrate and protein content before and after the reactive extraction. The following conclusions were made based on the results obtained from the study: The inclusion of sodium dodecyl sulphate in H 2 SO 4 significantly improved the FAME yields obtained from Nannochloropsis and Chlorella. A zirconium dodecyl sulphate catalyst performed better with Nannochloropsis than in Chlorella. The micro-algae maintain almost all the protein content and some of the carbohydrate after the in situ transesterification. The reaction time and yields were significant function of algae strain and catalyst.
7 REFERENCES [1] Gerhardt P., Murray R. G. E., Wood W. A., Krieg, N. R. (1994)." Methods for General and Molecular Bacteriology." ASM,Washington D. C, p518. [2] Lourenc, S.O, Barbarino, E., Lavin, P.L., Marquez, U.M.L., and Aidar, E. (2004) Distribution of intracellular nitrogen in marine microalgae: Calculation of new nitrogen-to-protein conversion factors Eur. J. Phycol., 39 (1): [3] Velasquez-orta, S. B., Lee, J. G. M. and Harvey, A. (2012). "Alkaline in situ transesterification of Chlorella vulgaris." Fuel, 94, [4] Haas, M. J., McAloon, A. J., Yee, W. C. and Foglia, T. A. (2006). A process model to estimate biodiesel production costs. Bioresource Technology, 97, [5] Lardon L., Sialve B., Steyer J. and Bernard O. (2009). Life-Cycle Assessment of Biodiesel Production from Microalgae. Environmental Science and Technology, 17, [6] Garces, R., Mancha, M. (1993). One-step lipid extraction and fatty acid methyl esters preparation from fresh plant tissues. Anal Biochem, 211, [7] Harun, R., Danquah, M.K and G. M. Forde, M.K (2010). Microalgal biomass as a fermentation feedstock for bioethanol production. J Chem Technol Biotechnol 2010; 85: [8] Lewis, T., Nichols, P. D. and McMeekin, T. A. (2000). "Evaluation of extraction methods for recovery of fatty acids from lipid-producing microheterotrophs." Journal of Microbiological Methods, 43(2), [9] Vicente, G., Bautista, L. F., Rodriguez, R., Gutierrez, J. F., Sadaba, I., Ruiz- Vazquez, R.M., Torres-Martinez, S. and Garre, V. (2009). "Biodiesel production from biomass of an oleaginous fungus." Biochemical Engineering Journal, 48 (1), [10] Revellame E, Hernandez R, French W, Holmes W, Alley E. (2010). Biodiesel from activated sludge through in situ transesterification. J Chem Technol Biotechnol; 85: [11] British standard: Fat and oil derivatives-fatty Acid Methyl Esters (FAME)- Determination of ester and linolenic acid methyl ester contents (BS EN 14103: 2003). [12] Cermak, S. C., Isbell, T.A. (2001). Synthesis of Estolides from Oleic and Saturated Fatty Acids. JAOCS, 78(6),
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