Home / Current Issue / Paper 1714827
Production Optimization of Lipopeptide Biosurfactant by Bacillus Subtilis Isolated from Soil Using Palm Oil Mill Effluent
Subject area: Biological & Medical Sciences · Area of research: Microbiology
Abstract
Microbial biosurfactants are amphiphilic compounds with diverse industrial, biomedical, and environmental applications, yet studies on optimized lipopeptide production by Bacillus subtilis remain limited. This study aimed to isolate, identify, and characterize microbial species from soil and evaluate lipopeptide biosurfactant production by B. subtilis. Soil samples were collected aseptically, serially diluted, heat-treated to enrich spore-forming bacteria, and cultured on selective media. Pure isolates were characterized morphologically, biochemically, and molecularly using 16S rRNA gene sequencing (for bacteria) and ITS region analysis (for fungi), followed by BLAST comparison and phylogenetic analysis. Lipopeptide production was optimized by varying pH, temperature, carbon and nitrogen sources, and incubation time. Extracted biosurfactants were characterized via Thin Layer Chromatography (TLC), High-Performance Liquid Chromatography (HPLC), and Liquid Chromatography–Mass Spectrometry (LC-MS). Functional activities, including emulsification, surface tension reduction, antimicrobial activity, and minimum inhibitory concentration (MIC), were assessed. Results confirmed the identity of B. subtilis, Escherichia coli, Staphylococcus aureus, and Candida albicans. B. subtilis produced surfactin, iturin, and fengycin most efficiently at pH 7.0, 37 °C, glucose and peptone as substrates, and 48-hour incubation. The biosurfactant demonstrated strong emulsifying ability, reduced surface tension to 28–35 mN/m, and exhibited broad-spectrum antimicrobial activity, with greater effectiveness against S. aureus. In conclusion, B. subtilis-derived lipopeptide biosurfactants possess potent bioactive properties suitable for industrial, biomedical, and environmental applications. It is recommended that industrial production, therapeutic evaluation, environmental application, and advanced molecular studies be pursued to fully exploit their potential.
Keywords
Optimization, Lipopeptide Biosurfactant, Bacillus Subtilis, Palm Oil Mill Effluent
References
[1] Abdullahi, M., Bello, A., & Adamu, H. (2022). Combined effects of pH and substrate concentration on lipopeptide biosurfactant production by Bacillus subtilis using palm oil mill effluent. Journal of Environmental Biotechnology, 18(2), 45–54.
[2] Adeyemi, O. A., Adekunle, O. C., & Olorode, O. A. (2020). Biochemical identification and antimicrobial susceptibility of Escherichia coli isolated from environmental samples. African Journal of Microbiology Research, 14(6), 312–319.
[3] Afolabi, T., & Adewale, S. (2022). Evaluation of treated palm oil mill effluent for lipopeptide biosurfactant production by soil-derived Bacillus subtilis. International Journal of Bioprocessing, 9(1), 12–21.
[4] Arima, K., Kakinuma, A., & Tamura, G. (2018). Surfactin, a crystalline peptide lipid surfactant produced by Bacillus subtilis: Isolation, characterization and its inhibition of fibrin clot formation. Biochemical and Biophysical Research Communications, 31(3), 488–494.
[5] Bello, M., & Sadiq, A. (2023). Effect of nutrient supplementation on lipopeptide biosurfactant production by Bacillus subtilis cultivated in POME-based medium. Journal of Industrial Microbiology & Biotechnology, 50(3), 215–227.
[6] Boateng, R., Mensah, E., & Adjei, K. (2020). Biosurfactant production by soil-isolated Bacillus subtilis using palm oil processing waste. Journal of Surfactants and Detergents, 23(5), 1123–1132.
[7] Cappuccino, J. G., & Welsh, C. T. (2017). Microbiology: A laboratory manual (11th ed.). Pearson Education.
[8] Cheesbrough, M. (2018). District laboratory practice in tropical countries (Part 2, 2nd ed.). Cambridge University Press.
[9] Clarridge, J. E. (2004). Impact of 16S rRNA gene sequence analysis for identification of bacteria on clinical microbiology and infectious diseases. Clinical Microbiology Reviews, 17(4), 840–862.
[10] Dubey, R. C., & Maheshwari, D. K. (2010). Bacillus subtilis: A potential producer of lipopeptide biosurfactants. Journal of Microbiology and Biotechnology, 20(2), 301–310.
[11] Eze, C., Udeh, P., & Okeke, N. (2021). Application of Plackett–Burman design to optimize lipopeptide biosurfactant production by Bacillus subtilis grown on palm oil mill effluent. Biocatalysis and Agricultural Biotechnology, 36, 102-123.
[12] Hamzah, R., Karim, M., & Yusof, N. (2020). Influence of aeration and agitation on lipopeptide biosurfactant production by soil-isolated Bacillus subtilis in POME medium. Bioresource Technology Reports, 10, 380.
[13] Hassan, M., Farooq, U., & Ahmad, S. (2020). Optimization of surfactin production by soil-derived Bacillus subtilis using waste oil substrates including POME. Applied Microbiology and Biotechnology, 104(15), 6755–6765.
[14] Janda, J. M., & Abbott, S. L. (2007). 16S rRNA gene sequencing for bacterial identification in the diagnostic laboratory: Pluses, perils, and pitfalls. Journal of Clinical Microbiology, 45(9), 2761–2764.
[15] Jawetz, E., Melnick, J. L., & Adelberg, E. A. (2020). Medical microbiology (28th ed.). McGraw-Hill Education.
[16] Kinsella, R., Raaijmakers, J. M., & van Elsas, J. D. (2009). Diversity and bioactivity of lipopeptides from Bacillus species. FEMS Microbiology Letters, 299(2), 119–127.
[17] Kiran, G. S., Priyadharsini, S., Selvin, J., & Sabarathnam, B. (2011). Optimization of lipopeptide production by Bacillus subtilis for environmental and industrial applications. Bioresource Technology, 102(17), 8675–8682.
[18] Kumar, P., & Singh, R. (2021). Optimization of lipopeptide biosurfactant production by Bacillus subtilis using waste lipid substrates. Journal of Environmental Chemical Engineering, 9(4), 106-120.
[19] Lawal, A., Balogun, S., & Ajayi, O. (2020). Optimization of surfactin production by Bacillus subtilis using agro-industrial wastes including POME. Bioprocess and Biosystems Engineering, 43(7), 1321–1331.
[20] Nwachukwu, E., & Okonko, I. O. (2019). Phenotypic characterization of enteric bacteria isolated from environmental and clinical sources. International Journal of Environmental Health Research, 29(3), 275–286.
[21] Odonkor, S. T., & Addo, K. K. (2018). Prevalence of multidrug-resistant Escherichia coli isolated from drinking water sources. International Journal of Microbiology, 2(1), 1–7.
[22] Ogunyemi, A., Olalekan, A., & Adekunle, J. (2021). Optimization of lipopeptide biosurfactant production by Bacillus subtilis using palm oil mill effluent. Biotechnology Reports, 30, 6-20.
[23] Okoro, C., & Nwankwo, C. (2023). Biosurfactant production by Bacillus subtilis using blended palm oil mill effluent and mineral salt medium. Environmental Technology & Innovation, 29, 102-119.
[24] Olatunji, O., Adeyemi, T., & Oladipo, F. (2022). Effect of fermentation duration and substrate concentration on biosurfactant production by Bacillus subtilis grown on palm oil mill effluent. Journal of Cleaner Production, 350, 131-141.
[25] Oyeleke, S. B., Dauda, B. E. N., & Oyewole, O. A. (2021). Biochemical characterization of bacteria isolated from food and water samples. Nigerian Journal of Basic and Applied Sciences, 29(2), 92–101.
[26] Patel, R. (2019). Identification of clinically relevant bacteria using molecular methods. Clinical Infectious Diseases, 68(2), 275–282.
[27] Peypoux, F., Bonmatin, J. M., & Wallach, J. (1999). Recent trends in the biochemistry of surfactin. Applied Microbiology and Biotechnology, 51(5), 553–563.
[28] Pratama, R., Putra, M., & Santoso, B. (2021). Optimization of lipopeptide biosurfactant production using treated palm oil mill effluent. Environmental Technology, 42(12), 1742–1752.
[29] Prescott, L. M., Harley, J. P., & Klein, D. A. (2017). Microbiology (10th ed.). McGraw-Hill Education.
[30] Raaijmakers, J. M., de Bruijn, I., & de Kock, M. J. D. (2010). Cyclic lipopeptide production by plant-associated Pseudomonas species: Diversity, activity, and biosynthesis. Molecular Plant-Microbe Interactions, 23(2), 167–177.
[31] Rahim, F., Khan, A., & Ali, S. (2021). Batch fermentation optimization of lipopeptide biosurfactant production by Bacillus subtilis using anaerobically treated POME. Biochemical Engineering Journal, 169, 107-119.
[32] Schoch, C. L., Seifert, K. A., Huhndorf, S., Robert, V., Spouge, J. L., Levesque, C. A., … Fungal Barcoding Consortium. (2012). Nuclear ribosomal internal transcribed spacer (ITS) region as a universal DNA barcode marker for Fungi. Proceedings of the National Academy of Sciences, 109(16), 6241–6246.
[33] Singh, P., & Cameotra, S. S. (2004). Enhancement of surfactin production by Bacillus subtilis through optimization of culture conditions. Applied Microbiology and Biotechnology, 64(4), 428–435.
[34] Sulaiman, A., & Garba, M. (2022). Effect of incubation time and temperature on lipopeptide biosurfactant production by Bacillus subtilis using palm oil mill effluent. Journal of Applied Microbiology, 132(4), 2565–2576.
[35] Todar, K. (2012). Todar’s online textbook of bacteriology. University of Wisconsin–Madison.
[36] Verma, P., Sharma, S., & Singh, N. (2022). Industrial effluent-based media for biosurfactant production by Bacillus subtilis: Optimization using response surface methodology. Journal of Industrial Microbiology & Biotechnology, 49(6), 1–12.
[37] Vijayakumar, R., & Saravanan, V. S. (2015). Optimization of lipopeptide biosurfactant production by Bacillus subtilis. International Journal of Current Microbiology and Applied Sciences, 4(4), 456–466.
[38] Woo, P. C. Y., Lau, S. K. P., Teng, J. L. L., Tse, H., & Yuen, K. Y. (2008). Then and now: Use of 16S rDNA gene sequencing for bacterial identification and discovery of novel bacteria in clinical microbiology laboratories. Clinical Microbiology and Infection, 14(10), 908–934.
[39] Yakubu, M., & Mohammed, H. (2023). Kinetics of lipopeptide biosurfactant production by Bacillus subtilis using palm oil mill effluent-based fermentation. Biotechnology Reports, 38, 6-17.
How to cite this paper
@article{1714827,
author = {Chikere, Jessica Evi, Ndiokwere Chioma Gabriella, Thankyou, Saturday Okpabi},
title = {Production Optimization of Lipopeptide Biosurfactant by Bacillus Subtilis Isolated from Soil Using Palm Oil Mill Effluent},
journal = {Iconic Research And Engineering Journals},
year = {2026},
volume = {9},
number = {9},
pages = {565-576},
issn = {2456-8880},
url = {https://www.irejournals.com/formatedpaper/1714827.pdf},
abstract = {Microbial biosurfactants are amphiphilic compounds with diverse industrial, biomedical, and environmental applications, yet studies on optimized lipopeptide production by Bacillus subtilis remain limited. This study aimed to isolate, identify, and characterize microbial species from soil and evaluate lipopeptide biosurfactant production by B. subtilis. Soil samples were collected aseptically, serially diluted, heat-treated to enrich spore-forming bacteria, and cultured on selective media. Pure isolates were characterized morphologically, biochemically, and molecularly using 16S rRNA gene sequencing (for bacteria) and ITS region analysis (for fungi), followed by BLAST comparison and phylogenetic analysis. Lipopeptide production was optimized by varying pH, temperature, carbon and nitrogen sources, and incubation time. Extracted biosurfactants were characterized via Thin Layer Chromatography (TLC), High-Performance Liquid Chromatography (HPLC), and Liquid Chromatography–Mass Spectrometry (LC-MS). Functional activities, including emulsification, surface tension reduction, antimicrobial activity, and minimum inhibitory concentration (MIC), were assessed. Results confirmed the identity of B. subtilis, Escherichia coli, Staphylococcus aureus, and Candida albicans. B. subtilis produced surfactin, iturin, and fengycin most efficiently at pH 7.0, 37 °C, glucose and peptone as substrates, and 48-hour incubation. The biosurfactant demonstrated strong emulsifying ability, reduced surface tension to 28–35 mN/m, and exhibited broad-spectrum antimicrobial activity, with greater effectiveness against S. aureus. In conclusion, B. subtilis-derived lipopeptide biosurfactants possess potent bioactive properties suitable for industrial, biomedical, and environmental applications. It is recommended that industrial production, therapeutic evaluation, environmental application, and advanced molecular studies be pursued to fully exploit their potential.},
keywords = {Optimization, Lipopeptide Biosurfactant, Bacillus Subtilis, Palm Oil Mill Effluent},
month = {March},
doi = {https://doi.org/10.64388/IREV9I9-1714827}
}