skip to main content

Low-Density Floating PGlu–STY/EPS Immobilized Lipase Biocatalyst with Particle-Size-Controlled Architecture for Fatty Acid Ethyl Ester Production

1Sarasas Suvarnabhumi Institute of Technology, Thailand

2Department of Chemistry Faculty of Science Chulalongkorn University, Thailand

3Department of Chemistry, Faculty of Science, , Thailand

4 Chulalongkorn University, Thailand

View all affiliations
Received: 13 Mar 2026; Published: 17 Jul 2026.
Editor(s): H Hadiyanto
Open Access Copyright (c) 2025 The Author(s). Published by Centre of Biomass and Renewable Energy (CBIORE)
Creative Commons License This work is licensed under a Creative Commons Attribution-ShareAlike 4.0 International License.

Citation Format:
Abstract

Pseudomonas cepacia lipase was immobilized onto low-density polyglutaraldehyde–styrene-coated expandable polystyrene beads (PGlu–STY/EPS) and applied as a reusable biocatalyst for biodiesel production from soybean oil via ethanolysis. The catalyst system was developed through a progressive catalyst-engineering strategy involving floating support design, support-density engineering, particle-size optimization, and coating morphology refinement to may improve catalyst distribution, interfacial accessibility, and operational stability. The immobilized lipase exhibited a protein loading yield of 71.81% and catalytic activity of 26.12 U g⁻¹-support. Smaller EPS particle sizes and optimized PGlu–STY coating conditions improved enzyme immobilization efficiency and catalytic performance by enhancing substrate accessibility and reducing diffusion limitations. Under optimized transesterification conditions, including an oil-to-ethanol molar ratio of 1:5, temperature of 40 °C, reaction time of 24 h, and the use of absolute ethanol, the maximum fatty acid ethyl ester (FAEE) conversion reached 92.8%. Biodiesel conversion was quantified using ¹H NMR spectroscopy. The immobilized catalyst maintained substantial catalytic activity over more than 10 repeated reaction cycles, indicating favorable operational stability and reusability. FT-IR analysis suggested successful covalent immobilization through Schiff-base interactions between aldehyde groups of the support and amino groups of the enzyme. The floating low-density EPS architecture may contribute to improved catalyst distribution in the heterogeneous oil/alcohol reaction medium and reduced unfavorable sedimentation in glycerol-rich regions, thereby potentially enhancing interfacial transesterification behavior. Although advanced characterization techniques such as BET, XPS, and GC–MS were not available in the present study, the combined catalytic and morphological results demonstrate that PGlu–STY/EPS supports provide a potentially useful platform for reusable immobilized lipase systems for enzymatic transesterification applications.

Keywords: immobilized lipase; biodiesel; transesterification; Pseudomonas cepacia; fatty acid ethyl esters; enzyme immobilization

Article Metrics:

  1. A. Srivastava and R. Prasad, “Triglycerides-based diesel fuels,” Renewable and Sustainable Energy Reviews, vol. 4, no. 2, pp. 111–133, 2000
  2. P. Vignesh, A. R. Pradeep, N. S. Ganesh, V. Jayaseelan, and K. Sudhakar, “A review of conventional and renewable biodiesel production,” Chinese Journal of Chemical Engineering, vol. 40, pp. 1–17, 2021
  3. S. B. Monika and V. V. Vinayak, “Biodiesel production from waste cooking oil: Application of heterogeneous catalysts and sustainable perspectives,” Energy Conversion and Management, vol. 289, p. 117164, 2023
  4. M. C. Robert, Biodiesel Handling and Use Guidelines, 6th ed. National Renewable Energy Laboratory (NREL), 2006
  5. C. V. McNeff et al., “A continuous catalytic system for biodiesel production,” Applied Catalysis A: General, vol. 343, pp. 39–48, 2008
  6. C. Roman-Figueroa et al., “High-yield production of biodiesel by non-catalytic supercritical methanol transesterification of crude castor oil (Ricinus communis),” Energy, vol. 107, pp. 165–171, 2016
  7. Y. Liu and L. Wang, “Biodiesel production from rapeseed deodorizer distillate in a packed column reactor,” Chemical Engineering and Processing, vol. 48, pp. 1152–1156, 2009
  8. S. Bordoloi et al., “Biodiesel production from mixed oils: A sustainable approach toward industrial biofuel production,” Chemical Engineering Journal Advances, vol. 10, p. 100262, 2022
  9. S. F. Li, Y. H. Fan, R. F. Hu, and W. T. Wu, “Pseudomonas cepacia lipase immobilized onto electrospun PAN nanofibrous membranes for biodiesel production from soybean oil,” Journal of Molecular Catalysis B: Enzymatic, vol. 72, pp. 40–45, 2011
  10. J. H. C. Wancura, M. V. Tres, J. V. Oliveira, and M. A. Mazutti, “Enzymatic biodiesel production via lipase catalysis: Current advances and future perspectives,” Renewable Energy, vol. 210, pp. 1200–1215, 2023
  11. M. Alonazi et al., “Combined immobilized lipases for effective biodiesel production from spent coffee grounds,” Saudi Journal of Biological Sciences, vol. 30, p. 103713, 2023
  12. H. C. Shao and T. W. Wen, “Immobilization of Candida rugosa lipase on chitosan with activation of hydroxyl groups,” Biomaterials, vol. 25, pp. 197–204, 2004
  13. H. Noureddini, X. Gao, and R. S. Philkana, “Immobilized Pseudomonas cepacia lipase for biodiesel fuel production from soybean oil,” Bioresource Technology, vol. 96, pp. 769–777, 2005
  14. L. Fjerbaek, K. V. Christensen, and B. Norddahl, “A review of the current state of biodiesel production using enzymatic transesterification,” Biotechnology and Bioengineering, vol. 102, pp. 1298–1315, 2009
  15. Y. Watanabe et al., “Continuous production of biodiesel fuel from vegetable oil using immobilized Candida antarctica lipase,” Journal of the American Oil Chemists’ Society, vol. 77, pp. 355–360, 2000
  16. T. Tan, J. Lu, K. Nie, L. Deng, and F. Wang, “Biodiesel production with immobilized lipase: A review,” Biotechnology Advances, vol. 28, pp. 628–634, 2010
  17. Q. Wang et al., “Co-immobilization of lipases for biodiesel production,” International Journal of Molecular Sciences, vol. 24, no. 5, p. 4726, 2023
  18. A. El-Shafie, “Lipase-based enzymatic biodiesel production from waste cooking oil: Recent progress and industrial perspectives,” Fuel, vol. 365, p. 131245, 2024
  19. A. Moschona, A. Spanou, I. V. Pavlidis, A. J. Karabelas, and S. Patsios, “Optimization of enzymatic transesterification of acid oil for biodiesel production using a low-cost lipase: The effect of transesterification conditions and the synergy of lipases with different regioselectivity,” Applied Biochemistry and Biotechnology, vol. 196, no. 5, pp. 8168–8189, 2024
  20. A. H. Mat Saad et al., “Immobilized ancestral lipase for enzymatic FAEE production,” Applied Biochemistry and Biotechnology, vol. 198, pp. 655–679, 2026
  21. N. F. S. M. Nor et al., “Enzymatic transesterification using immobilized lipases,” Bulletin of Chemical Reaction Engineering & Catalysis, vol. 19, no. 1, pp. 45–58, 2024
  22. Y. Wang, H. Zhao, and J. Liu, “Immobilized lipase catalysts for enzymatic FAEE production: Challenges and opportunities,” Renewable Energy, vol. 235, p. 121456, 2025
  23. S. R. Sandler, W. Karo, J. A. Bonesteel, and E. M. Pearce, Polymer Synthesis and Characterization: A Laboratory Manual. New York, NY, USA: Academic Press, 2000
  24. F. Wang, Y. Xu, and D. Shan, “Immobilization of lipase on microporous resin and its application in synthesis of biodiesel,” Chinese Journal of Biotechnology, vol. 22, no. 1, pp. 114–118, 2006
  25. N. Rachmadona, D. A. Putri, and E. Prasetyo, “Activated carbon-supported lipase as biocatalyst for biodiesel synthesis from crude palm oil,” Current Applied Science and Technology, vol. 25, no. 2, pp. 145–156, 2025
  26. X. Chen, “Polymer-supported lipase catalysts for sustainable biodiesel synthesis,” Bioresource Technology, vol. 392, p. 130102, 2024
  27. E. Y. Park, M. Sato, and S. Kojima, “Fatty acid methyl ester production using lipase-immobilizing silica particles with different particle sizes and surface areas,” Enzyme and Microbial Technology, vol. 39, pp. 889–896, 2006
  28. F. Yagiz, D. Kazan, and A. N. Akin, “Biodiesel production from waste oils by using lipase immobilized on hydrotalcite and zeolites,” Chemical Engineering Journal, vol. 134, pp. 262–267, 2007
  29. F. Kanwal, N. Ahmad, and M. Malik, “Nanoparticle-immobilized lipase with enhanced catalytic stability for biodiesel synthesis,” Catalysts, vol. 15, no. 1, p. 85, 2025
  30. N. Dizge, B. Keskinler, and A. Tanriseven, “Biodiesel production from canola oil by using lipase immobilized onto hydrophobic microporous styrene-divinylbenzene copolymer,” Biochemical Engineering Journal, vol. 44, pp. 220–225, 2009
  31. M. Kaieda et al., “Effect of methanol on production of biodiesel fuel from plant oil catalyzed by various lipases in a solvent-free system,” Journal of Bioscience and Bioengineering, vol. 91, pp. 12–15, 2001
  32. J. Liu et al., “Improved catalytic stability of immobilized Candida antarctica lipase B on macroporous resin for biodiesel production,” Bioprocess and Biosystems Engineering, vol. 48, pp. 147–157, 2025
  33. H. Suo et al., “Immobilized lipase post-encapsulated in Cu-based MOFs with enhanced stability in biodiesel synthesis,” Process Biochemistry, vol. 147, pp. 440–447, 2024
  34. W. A. Alshehri et al., “Thermostable CaCO3-immobilized Bacillus subtilis lipase for enzymatic FAEE production from waste cooking oil,” Catalysts, vol. 14, no. 4, p. 245, 2024
  35. F. Kanwal et al., “Microalgae-based biodiesel: Integrating AI, CRISPR and nanotechnology for sustainable biofuel development,” Emerging Topics in Life Sciences, vol. 9, no. 1, pp. 55–72, 2025
  36. L. Giraldo et al., “Biodiesel production using MOF-lipase biocatalyst from Candida antarctica,” International Journal of Molecular Sciences, vol. 24, p. 10741, 2023

Last update:

No citation recorded.

Last update: 2026-07-25 13:26:16

No citation recorded.