Applications of immobilized Bacillus subtilis in biobutanol production purification and fuel characteristic analysis
DOI:
https://doi.org/10.70130/CAST.2025.8101Keywords:
Bio-butanol , Liquid liquid extraction , Immobilized cells, Distillation , Fuel parametersAbstract
The need for eco-friendly energy sources has led to a surge in biofuel research, where bio-butanol is emerging as a potential alternative to fossil fuels. The study delves into bio-butanol purification and fuel parameter analysis using immobilized Bacillus cells. A maximum amount of bio-butanol was recovered by integrating liquid-liquid extraction with distillation techniques. Bio-butanol production was boosted to more than 80 % compared to free cells. The immobilized cells of the potent Bacillus subtilis strain (T1) demonstrated remarkable efficiency, elevating bio-butanol production from 8.16 g/L (with free cells) to a substantial 15.03 g/L (immobilized cells). The recovered bio-butanol was suitable as a direct fuel source and a blend with standard petrol. The immobilization of the cells further solved one of the limitations in bio-butanol production. This breakthrough highlights bio-butanol potential as a sustainable and eco-friendly fuel, contributing to a more sustainable energy future. The findings provide a promising pathway toward reducing our dependence on conventional fossil fuels and mitigating environmental impact, making strides toward a greener and more energy-efficient future.
References
Aakko-Saksa, P. T., Lehtoranta, K., Kuittinen, N., Järvinen, A., Jalkanen, J.-P., Johnson, K., Jung, H., Ntziachristos, L., Gagné, S., Takahashi, C., Karjalainen, P., Rönkkö, T., & Timonen, H. (2023). Reduction in greenhouse gas and other emissions from ship engines: Current trends and future options. Progress in Energy and Combustion Science, 94, 101055. https://doi.org/10.1016/j.pecs.2022.101055
Algayyim, S. J. M., Yusaf, T., Hamza, N. H., Wandel, A. P., Fattah, I. M. R., Laimon, M., & Rahman, S. M. A. (2022). Sugarcane biomass as a source of biofuel for internal combustion engines (ethanol and acetone-butanol-ethanol): A review of economic challenges. Energies, 15(22), 8644. https://doi.org/10.3390/en15228644
Amin, M. A., Shukor, H., Shoparwe, N. F., Makhtar, M. M. Z., Abdul Hamid, A. A., & Rongwong, W. (2024). Medium optimization for Bio-butanol Production from Palm Kernel Cake (PKC) hydrolysate by Clostridium saccharoperbutylacetonium N1-4. Malaysian Applied Biology, 53(1), 67–81. https://doi.org/10.55230/mabjournal.v53i1.2869
Aragão Börner, R. A., Zaushitsyna, O., Berillo, D., Scaccia, N., Mattiasson, B., & Kirsebom, H. (2014). Immobilization of Clostridium acetobutylicum DSM 792 as macroporous aggregates through cryogelation for butanol production. Process Biochemistry, 49(1), 10–18. https://doi.org/10.1016/j.procbio.2013.09.027
Buday, Z., Linden, J. C., & Karim, M. N. (1990). Improved acetone-butanol-ethanol fermentation analysis using subambient HPLC column temperature. Enzyme and Microbial Technology, 12(1), 24–27. https://doi.org/10.1016/0141-0229(90)90175-P
Chairul, C., Ridho, M. H., Zultiniar, Z., & Hendra, A. (2023). Utilization of sago solid waste for bioethanol production with variation of sago solid waste solvent ratio and pretreatment time. Conference paper. 9th International Conference on Sustainable Agriculture, Food and Energy. SAFE.
Chen, Y., Zhou, T., Liu, D., Li, A., Xu, S., Liu, Q., Li, B., & Ying, H. (2013). Production of butanol from glucose and xylose with immobilized cells of Clostridium acetobutyllicum. Biotechnology and Bioprocess Engineering, 18(2), 234–241. https://doi.org/10.1007/s12257-012-0573-5
Chen, Y. Q., Garg, N. P., Luo, H., Kontogeorgis, G. M., & Woodley, J. M. (2021). Ionic liquid-based in situ product removal design exemplified for an acetone-butanol-ethanol fermentation. Biotechnology and Bioprocess Engineering, 5, e3183.
Chutke, N., Phadke, R. V., Revatkar, L. B., & Ramteke, G. M. (2016). Analysis of petrol: A clarification for purity of Petrol. ESR [Journal], 1(1).
Crabbe, E., Nolasco-Hipolito, C., Kobayashi, G., Sonomoto, K., & Ishizaki, A. (2001). Biodiesel production from crude palm oil and evaluation of butanol extraction and fuel properties. Process Biochemistry, 37(1), 65–71. https://doi.org/10.1016/S0032-9592(01)00178-9
Dadgar, A. M., & Foutch, G. L. (1988). Improving the acetone‐butanol fermentation process with liquid‐liquid extraction. Biotechnology Progress, 4(1), 36–39. https://doi.org/10.1002/btpr.5420040107
Dalle Ave, G., & Adams, T. A. (2018). Techno-economic comparison of acetone-butanol-ethanol fermentation using various extractants. Energy Conversion and Management, 156, 288–300. https://doi.org/10.1016/j.enconman.2017.11.020
Demichelis, F., Laghezza, M., Chiappero, M., & Fiore, S. (2020). Technical, economic and environmental assessement of bioethanol biorefinery from waste biomass. Journal of Cleaner Production, 277, 124111. https://doi.org/10.1016/j.jclepro.2020.124111
Ester, A. M. A. L. M., & Pelarut, S. (2015). Optimization study for butanol extraction from butanol-water using fatty acid methyl Ester (FAME) as solvent. Malaysian Journal of Analytical Sciences, 19(1), 1–7.
Feng, R., Yang, J., Zhang, D., Deng, B., Fu, J., Liu, J., & Liu, X. (2013). Experimental study on SI engine fuelled with butanol–gasoline blend and H2O addition. Energy Conversion and Management, 74, 192–200. https://doi.org/10.1016/j.enconman.2013.05.021
Frick, C., & Schügerl, K. (1986). Continuous acetone-butanol production with free and immobilized Clostridium acetobutylicum. Applied Microbiology and Biotechnology, 25(3), 186–193. https://doi.org/10.1007/BF00253646
González-Peñas, H., Eibes, G., Lu-Chau, T. A., Moreira, M. T., & Lema, J. M. (2020). Altered Clostridia response in extractive ABE fermentation with solvents of different nature. Biochemical Engineering Journal, 154, 107455. https://doi.org/10.1016/j.bej.2019.107455
Groot, W. J., Soedjak, H. S., Donck, P. B., Van der Lans, R. G. J. M., Luyben, K. C. A. M., & Timmer, J. M. K. (1990). Butanol recovery from fermentations by liquid–liquid extraction and membrane solvent extraction. Bioprocess Engineering, 5(5), 203–216. https://doi.org/10.1007/BF00376227
Häggström, L., & Molin, N. (1980). Calcium alginate immobilized cells of Clostridium acetobutyllicum for solvent production. Biotechnology Letters, 2(5), 241–246. https://doi.org/10.1007/BF01209440
Hönig, V., Kotek, M., & Mařík, J. (2014). Use of butanol as a fuel for internal combustion engines. Agronomy Research, 12(2), 333–340.
Husin, H., Ibrahim, M. F., Kamal Bahrin, E., & Abd‐Aziz, S. (2019). Simultaneous saccharification and fermentation of sago hampas into bio-butanol by Clostridium acetobutylicum ATCC 824. Energy Science and Engineering, 7(1), 66–75. https://doi.org/10.1002/ese3.226
Jenkins, R. W., Munro, M., Nash, S., & Chuck, C. J. (2013). Potential renewable oxygenated biofuels for the aviation and road transport sectors. Fuel, 103, 593–599. https://doi.org/10.1016/j.fuel.2012.08.019
Jiménez-Bonilla, P., Zhang, J., Wang, Y., Blersch, D., de-Bashan, L. E., Guo, L., Li, X., Zhang, D., & Wang, Y. (2022). Polycationic surfaces promote whole cell immobilization and induce microgranulation of Clostridium saccharoperbutylacetonicum N1-4 for enhanced biobutanol production. ACS Applied Materials and Interfaces, 14(44), 49555–49567. https://doi.org/10.1021/acsami.2c14888
Johnravindar, D., Elangovan, N., Gopal, N. O., Muthaiyan, A., & Fei, Q. (2019). Bio-butanol production from cassava waste residue using Clostridium sp. AS3 in batch culture fermentation. Biofuels, 1–8.
Johnson, P., & Brown, L. (2019). Sago spent waste as a potential feedstock for bio-butanol production. Bioresource Technology, 271, 400–408.
Jones, D. T., Schulz, F., Roux, S., & Brown, S. D. (2023). Solvent-producing Clostridia Revisited. Microorganisms, 11(9), 2253. https://doi.org/10.3390/microorganisms11092253
Joseph, A. M., Tulasi, Y., Shrivastava, D., & Kiran, B. (2023). Techno-economic feasibility and exergy analysis of bioethanol production from waste. Energy Conversion and Management: X, 18, 100358. https://doi.org/10.1016/j.ecmx.2023.100358
Khedkar, M. A., Nimbalkar, P. R., Gaikwad, S. G., Chavan, P. V., & Bankar, S. B. (2020). Solvent extraction of butanol from synthetic solution and fermentation broth: Batch and continuous studies. Separation and Purification Technology, 249, 117058. https://doi.org/10.1016/j.seppur.2020.117058
Krasňan, V., Plž, M., Marr, A. C., Markošová, K., Rosenberg, M., & Rebroš, M. (2018). Intensified crude glycerol conversion to butanol by immobilized Clostridium pasteurianum. Biochemical Engineering Journal. M. and Rebroš, 134, 114–119. https://doi.org/10.1016/j.bej.2018.03.005
Kumar, K., Jadhav, S. M., & Moholkar, V. S. (2024). Acetone- butanol- ethanol (ABE) fermentation with clostridial coculture for enhanced bio-butanol production. Process Safety and Environmental Protection, 185, 277–285. https://doi.org/10.1016/j.psep.2024.03.027
Kumar, M., Saini, S., & Gayen, K. (2014). Acetone-butanol-ethanol fermentation analysis using only high performance liquid chromatography. Anal. Methods, 6(3), 774–781. https://doi.org/10.1039/C3AY41717D
Li, M., Li, R., & Wang, C. (2023). Characteristics of flame velocity and burning rate of large-scale continuous leakage spilling fire of n-butanol fuel. Fuel, 332, 126148. https://doi.org/10.1016/j.fuel.2022.126148
Lin, X., Fan, J., Wen, Q., Li, R., Jin, X., Wu, J., Qian, W., Liu, D., Xie, J., Bai, J., & Ying, H. (2014). Optimization and validation of a GC–FID method for the determination of acetone-butanol-ethanol fermentation products. Journal of Chromatographic Science, 52(3), 264–270. https://doi.org/10.1093/chromsci/bmt022
Linggang, S., Phang, L. Y., Wasoh, H., & Abd-Aziz, S. (2013). Acetone–butanol–ethanol production by Clostridium acetobutylicum ATCC 824 using sago pith residues hydrolysate. Bioenergy Research, 6(1), 321–328. https://doi.org/10.1007/s12155-012-9260-9
Liu, J., Fan, L. T., Seib, P., Friedler, F., & Bertok, B. (2004). Downstream process synthesis for biochemical production of butanol, ethanol, and acetone from grains: Generation of optimal and near-optimal flowsheets with conventional operating units. Biotechnology Progress, 20(5), 1518–1527. https://doi.org/10.1021/bp049845v
Liu, L., Wang, Y. C., Wang, N., Chen, X. M., Li, B. G., Shi, J. P., & Li, X. (2021). Process optimization of acetone-butanol-ethanol fermentation integrated with pervaporation for enhanced butanol production. Biochemical Engineering Journal, 173, 108070. https://doi.org/10.1016/j.bej.2021.108070
Liu, W., Zha, W., Yin, H., Yang, C., Lu, K., & Chen, J. (2023). Integration of rare earth element stimulation, activated carbon absorption and cell immobilization in ABE fermentation for promoting biobutanol production. Chemical Engineering and Processing: Process Intensification, 184, 109–306.
Ni, B.-J., Rittmann, B. E., & Yu, H.-Q. (2011). Soluble microbial products and their implications in mixed culture biotechnology. Trends in Biotechnology, 29(9), 454–463. https://doi.org/10.1016/j.tibtech.2011.04.006
A, O. U. (2007). Cassava wastes: Treatment options and value addition alternatives. African Journal of Biotechnology, 6(18), 2065–2073. https://doi.org/10.5897/AJB2007.000-2319
Owens, J. E., & Lowe, L. E. (2023). Headspace versus direct injection for the quantitation of methanol, ethyl acetate, and fusel oils in distilled spirits by gas chromatography – Flame ionization detection. MethodsX, 11, 102387. https://doi.org/10.1016/j.mex.2023.102387
Potter, T. L. (1996). Analysis of petroleum‐contaminated water by GC/FID with direct aqueous injection. Groundwater Monitoring & Remediation, 16(3), 157–162. https://doi.org/10.1111/j.1745-6592.1996.tb00145.x
Qureshi, N., & Blaschek, H. P. (1999). Production of acetone butanol ethanol (ABE) by a hyper‐producing mutant strain of Clostridium beijerinckii BA101 and recovery by pervaporation. Biotechnology Progress, 15(4), 594–602. https://doi.org/10.1021/bp990080e
Qureshi, N., Maddox, I. S., & Friedl, A. (1992). Application of continuous substrate feeding to the ABE fermentation: Relief of product inhibition using extraction, perstraction, stripping, and pervaporation. Biotechnology Progress, 8(5), 382–390. https://doi.org/10.1021/bp00017a002
Rakopoulos, D. C., Rakopoulos, C. D., Papagiannakis, R. G., & Kyritsis, D. C. (2011). Combustion heat release analysis of ethanol or n-butanol diesel fuel blends in heavy-duty DI diesel engine. Fuel, 90(5), 1855–1867. https://doi.org/10.1016/j.fuel.2010.12.003
Rasyid, T. H., Kusumawaty, Y., & Hadi, S. (2020). The utilization of sago waste: Prospect and challenges. IOP Conference Series: Earth and Environmental Science. IOP Conference Series, 415(1). https://doi.org/10.1088/1755-1315/415/1/012023
Roffler, S. R., Blanch, H. W., & Wilke, C. R. (1987). In-situ recovery of butanol during fermentation. Bioprocess Engineering, 2(1), 1–12. https://doi.org/10.1007/BF00369221
Ruban, P., Sangeetha, T., & Indira, S. (2013). Starch waste as a substrate for amylase production by sago effluent isolates Bacillus subtilis and Aspergillus niger. American–Eurasian Journal of Agricultural and Environment Science, 13(1), 27–31.
Salwa, N., Gunawan, C. E. P., Danisa, F., Nilnalmuna, R., & Rohmah, N. (2024). Bioethanol production from sago waste as renewable energy: A review. Cleaning Technology, 1(1), 29–38.
Saraswat, M., & Chauhan, N. R. (2018). Assessment study of butanol-gasoline blends in variable compression ratio spark ignition engine.
Schoutens, G. H., Nieuwenhuizen, M. C. H., & Kossen, N. W. F. (1985). Continuous butanol production from whey permeate with immobilized Clostridium beyerinckii LMD 27.6. Applied Microbiology and Biotechnology, 21(5), 282–286. https://doi.org/10.1007/BF00252705
Shafei, M. S., Allam, R. F., & Sallam, L. A. R. (2002). Physiological factors affecting the production of acetone and n-butanol by immobilized Clostridium acetobutylicum culture. Egyptian Journal of Biotechnology, 11, 206–216.
Shalinimol, C. R. (2016). A study on optimization of microbial alpha-amylase production. DJ International Journal of Advances in Microbiology & Microbiological Research, 1(1), 22–27. https://doi.org/10.18831/djmicro.org/2016011004
Srinorakutara, T., Kaewvimol, L., & Saengow, L. A. (2006). Approach of cassava waste pretreatments for fuel ethanol production in Thailand. J. Sci. Res. Chula. Univ., 31(1), 77–84.
Tsai, T.-Y., Lo, Y.-C., Dong, C.-D., Nagarajan, D., Chang, J.-S., & Lee, D.-J. (2020). Biobutanol production from lignocellulosic biomass using immobilized Clostridium acetobutylicum. Applied Energy, 277, 115531. https://doi.org/10.1016/j.apenergy.2020.115531
Tsuey, L. S., Ariff, A. B., Mohamad, R., & Rahim, R. A. (2006). Improvements of GC and HPLC analyses in solvent (acetone-butanol-ethanol) fermentation by Clostridium saccharobutylicum using a mixture of starch and glycerol as carbon source. Biotechnology and Bioprocess Engineering, 11(4), 293–298. https://doi.org/10.1007/BF03026243
Van der Merwe, A. B., Cheng, H., Görgens, J. F., & Knoetze, J. H. (2013). Comparison of energy efficiency and economics of process designs for bio-butanol production from sugarcane molasses. Fuel, 105, 451–458. https://doi.org/10.1016/j.fuel.2012.06.058
Wang, Y., & Zhang, L. (2018). Response surface methodology for optimization of bio-butanol production: A comprehensive review. Bioengineering, 5(2), 28.
Xue, C., Yang, D., Du, G., Chen, L., Ren, J., & Bai, F. (2015). Evaluation of hydrophobic micro-zeolite-mixed matrix membrane and integrated with acetone–butanol–ethanol fermentation for enhanced butanol production. Biotechnology for Biofuels, 8(1), 105. https://doi.org/10.1186/s13068-015-0288-x
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