Optimizing Bioethanol Production Via Consolidated Bioprocessing: The Potential of Aspergillus niger B2484

https://doi.org/10.55230/mabjournal.v54i4.3230

Authors

  • Mona Fatin Syazwanee Mohamed Ghazali ASASIpintar Program, Pusat PERMATA@Pintar Negara, Universiti Kebangsaan Malaysia, 43600 UKM Bangi, Selangor
  • Muskhazli Mustafa Department of Biology, Faculty of Science, Universiti Putra Malaysia, 43400 Serdang, Selangor
  • Nur Ain Izzati Mohd Zainudin Department of Biology, Faculty of Science, Universiti Putra Malaysia, 43400 Serdang, Selangor
  • Nor Azwady Abd Aziz Department of Biology, Faculty of Science, Universiti Putra Malaysia, 43400 Serdang, Selangor

Keywords:

Bioethanol, Consolidated Bioprocessing, OFAT analysis, Single culture, Response Surface Methodology

Abstract

Consolidated bioprocessing (CBP) integrates enzyme secretion, hydrolysis, and fermentation into a single-step process, eliminating the need for costly separate enzyme production in bioethanol manufacturing. While CBP aims to utilize naturally occurring cellulolytic microbes, no single microorganism has been identified to efficiently perform all required processes. One of the key challenges in CBP is optimizing culture conditions to maximize bioethanol yield. This study investigates the potential of Aspergillus niger B2484 as a single-culture bioethanol producer and optimizes the physicochemical parameters for converting pretreated paddy straw into bioethanol through CBP. Key parameters, including saccharification and fermentation duration, temperature, substrate loading, and medium composition, were evaluated using the One-Factor-At-a-Time (OFAT) method and further optimized via Response Surface Methodology (RSM). The optimal conditions were determined to be 66.7 hr of saccharification at 29.8°C, followed by 32.3 hr of fermentation at 30.2°C, with a substrate loading of 2.6% (w/v) and a medium level of 14.8% (v/v). The actual ethanol yield (0.63 g/L) closely matched the RSM-predicted yield (0.61 g/L), confirming the reliability of the optimization model. This study demonstrates the feasibility of A. niger B2484 as a single-culture bioethanol producer in CBP, highlighting its potential for commercial application either as a standalone microbial agent or as part of a customized enzymatic system to enhance bioethanol yield.

Downloads

Download data is not yet available.

References

Aggarwal, N.K., Goyal, V., Saini, A., Yadav, A. & Gupta, R. 2017. Enzymatic saccharification of pretreated rice straw by cellulases from Aspergillus niger BK01. 3 Biotech, 7: 158. DOI: https://doi.org/10.1007/s13205-017-0755-0

Ahmad, F. & Usman, B. 2018. Production of bioethanol from rice husk using Aspergillus niger and Trichoderma harzianum. Bayero Journal of Pure and Applied Sciences, 10(1): 280-284. DOI: https://doi.org/10.4314/bajopas.v10i1.56S

Alabdalall, A.H., Almutari, A.A., Aldakeel, S.A., Albarrag, A.M., Aldakheel, L.A., Alsoufi, M.H., Alfuraih, L.Y. & Elkomy, H.M. 2023. Bioethanol production from lignocellulosic biomass using Aspergillus niger and Aspergillus flavus hydrolysis enzymes through immobilized S. cerevisiae. Energies, 16(2): 823. DOI: https://doi.org/10.3390/en16020823

Ali, S.S., Nugent, B., Mullins, E. & Doohan, F.M. 2016. Fungal-mediated consolidated bioprocessing: The potential of Fusarium oxysporum for the lignocellulosic ethanol industry. AMB Express, 6: 13. DOI: https://doi.org/10.1186/s13568-016-0185-0

Amiri, H., Karimi, K. & Zilouei, H. 2014. Organosolv pretreatment of rice straw for efficient acetone, butanol, and ethanol production. Bioresource Technology, 152: 450–456. DOI: https://doi.org/10.1016/j.biortech.2013.11.038

Arifa, T. & Sarwar S. 2012. Effect of cultural condition on production of ethanol from rotten apple waste by Saccharomyces cerevisiae straining. Canadian Journal of Applied Sciences, 2: 12-21. DOI: https://doi.org/10.21065/1925-7430.2.12

Bušić, A., Marđetko, N., Kundas, S., Morzak, G., Belskaya, H., Ivančić Šantek, M., Komes, D., Novak, S. & Šantek, B. 2018. Bioethanol production from renewable raw materials and its separation and purification: A review. Food Technology and Biotechnology, 56(3): 289-311. DOI: https://doi.org/10.17113/ftb.56.03.18.5546

Chang, Y.H., Chang, K.S., Chen, C.Y., Hsu, C.L., Chang, T.C. & Jang, H.D. 2018. Enhancement of the efficiency of bioethanol production by Saccharomyces cerevisiae via gradually batch-wise and fed-batch increasing the glucose concentration. Fermentation, 4: 45. DOI: https://doi.org/10.3390/fermentation4020045

Conesa, C., Seguí, L. & Fito, P. 2017. Hydrolytic performance of Aspergillus niger and Trichoderma reesei cellulases on lignocellulosic industrial pineapple waste intended for bioethanol production. Waste and Biomass Valorization, 9(8): 1359–1368. DOI: https://doi.org/10.1007/s12649-017-9887-z

Cunha, J.T., Soares, P.O., Baptista, S.L., Costa, C.E. & Domingues, L. 2020. Engineered Saccharomyces cerevisiae for lignocellulosic valorization: A review and perspectives on bioethanol production. Bioengineered, 11(1): 883-903. DOI: https://doi.org/10.1080/21655979.2020.1801178

Cutzu, R. & Bardi, L. 2017. Production of bioethanol from agricultural wastes using residual thermal energy of a cogeneration plant in the distillation phase. Fermentation, 3: 24. DOI: https://doi.org/10.3390/fermentation3020024

Dahir, A.A. & Al-Dossary, M.A.W. 2023. Bioethanol production from corn and barley wastes by Aspergillus flavus. Marsh Bulletin, 18(1): 29–42.

Dempfle, D., Kröcher, O. & Studer, M.H.P. 2021. Techno-economic assessment of bioethanol production from lignocellulose by consortium-based consolidated bioprocessing at industrial scale. New Biotechnology, 65: 53-60. DOI: https://doi.org/10.1016/j.nbt.2021.07.005

Devi, A., Bajar, S., Kour, H., Kothari, R., Pant, D. & Singh, A. 2022. Lignocellulosic biomass valorization for bioethanol production: A circular bioeconomy approach. Bioenergy Research, 15: 1820–1841. DOI: https://doi.org/10.1007/s12155-022-10401-9

Dinarvand, M., Rezaee, M. & Foroughi, M. 2017. Optimizing culture conditions for production of intra and extracellular inulinase and invertase from Aspergillus niger ATCC 20611 by response surface methodology (RSM). Brazilian Journal of Microbiology, 48: 427-441. DOI: https://doi.org/10.1016/j.bjm.2016.10.026

Du, Y., Zou, W., Zhang, K., Ye, G. & Yang, J. 2020. Advances and applications of Clostridium co-culture systems in biotechnology. Frontiers in Microbiology, 11: 560223. DOI: https://doi.org/10.3389/fmicb.2020.560223

Duarah, P., Haldar, D., Patel, A.K., Dong, C.D., Singhania, R.R. & Purkait, M.K. 2022. A review on global perspectives of sustainable development in bioenergy generation. Bioresource Technology, 348: 126791. DOI: https://doi.org/10.1016/j.biortech.2022.126791

Duncker, K.E., Holmes, Z.A. & You, L. 2021. Engineered microbial consortia: Strategies and applications. Microbial Cell Factories, 20: 211. DOI: https://doi.org/10.1186/s12934-021-01699-9

Gawande, S.B. & Patil, I.D. 2018. Experimental investigation and optimization for production of bioethanol from damaged corn grains. Materials Today: Proceedings, 5(1): 1509–1517. DOI: https://doi.org/10.1016/j.matpr.2017.11.240

Ghazal, M.A., Ibrahim, H.A.H., Shaltouta N.A. & Ali, A.E. 2016. Biodiesel and bioethanol production from Ulva fasciata delie biomass via enzymatic pretreatment using marine-derived Aspergillus niger. International Journal of Pure & Applied Bioscience, 4(5): 1-16. DOI: https://doi.org/10.18782/2320-7051.2274

Gupta, A., Stead, T.S. & Ganti, L. 2024. Determining a meaningful R-squared value in clinical medicine. Academic Medicine & Surgery. DOI: https://doi.org/10.62186/001c.125154

Hashem, M., Alamri, S.A., Asseri, T.A.Y., Mostafa, Y.S., Lyberatos, G., Ntaikou, I. 2021. On the optimization of fermentation conditions for enhanced bioethanol yields from starchy biowaste via yeast co-cultures. Sustainability, 13: 1890. DOI: https://doi.org/10.3390/su13041890

Humbird, D. & Fei, Q. 2016. Scale-up considerations for biofuels. In: Biotechnology for biofuels production and optimization. Eckert, C.A. & Trinh, C.T. (Eds.), Elsevier, Amsterdam, Netherlands. 513-537. DOI: https://doi.org/10.1016/B978-0-444-63475-7.00020-0

Izmirlioglu, G. & Demirci, A. 2016. Improved simultaneous saccharification and fermentation of bioethanol from industrial potato waste with co-cultures of Aspergillus niger and Saccharomyces cerevisiae by medium optimization. Fuel, 185: 684-691. DOI: https://doi.org/10.1016/j.fuel.2016.08.035

Jouzani, G.S. & Taherzadeh, M.J. 2015. Advances in consolidated bioprocessing systems for bioethanol and butanol production from biomass: A comprehensive review. Biofuel Research Journal, 5: 152-195. DOI: https://doi.org/10.18331/BRJ2015.2.1.4

Junior, A.B., Borges, D.G., Tardioli, P.W. & Farinas, C.S. 2014. Characterization of β-glucosidase produced by Aspergillus niger under solid-state fermentation and partially purified using MANAE-agarose. Biotechnology Research International, 2014: 317092. DOI: https://doi.org/10.1155/2014/317092

Kamei, I., Hirota, Y., Mori, T., Hirai, H., Meguro, S. & Kondo, R. 2012. Direct ethanol production from cellulosic materials by the hypersaline-tolerant white-rot fungus Phlebia sp. MG-60. Bioresource Technology, 112: 137–142. DOI: https://doi.org/10.1016/j.biortech.2012.02.109

Kricka, W., Fitzpatrick, J. & Bond, U. 2014. Metabolic engineering of yeasts by heterologous enzyme production for degradation of cellulose and hemicelluloses from biomass: A perspective. Frontiers in Microbiology, 5: 174: 1-11. DOI: https://doi.org/10.3389/fmicb.2014.00174

Lin, Y. & Wiegand, K. 2023. Low R2 in ecology: Bitter, or B-side?. Ecological Indicators, 153: 110406. DOI: https://doi.org/10.1016/j.ecolind.2023.110406

Madhuvanthi, S., Jayanthi, S., Suresh, S. & Pugazhendhi, A. 2022. Optimization of consolidated bioprocessing by response surface methodology in the conversion of corn stover to bioethanol by thermophilic Geobacillus thermoglucosidasius. Chemosphere, 304: 135242. DOI: https://doi.org/10.1016/j.chemosphere.2022.135242

Mauch, F., Mauch-Mani, B. & Boller, T. 1988. Antifungal hydrolases in pea tissue and inhibition of fungal growth by combinations of chitinase and β-1,3-glucanase. Plant Physiology, 88: 936-942. DOI: https://doi.org/10.1104/pp.88.3.936

Mbaneme-Smith, V. & Chinn, M. 2015. Consolidated bioprocessing for biofuel production: Recent advances. Energy and Emission Control Technologies, 3: 23-44. DOI: https://doi.org/10.2147/EECT.S63000

Olawale, B.I., Iliyasu, M.Y., Musa, B., Abdulrahman, A. & Umar, A.F. 2021. Production of bioethanol by co-culture of Aspergillus niger and Saccharomyces cerevisiae using watermelon peels as substrate. Path of Science, 7(10): 6001-6011. DOI: https://doi.org/10.22178/pos.75-10

Olguin-Maciel, E., Larqué-Saavedra, A., Lappe-Oliveras, P.E., Barahona-Pérez, L.F., Alzate-Gaviria, L., Chablé-Villacis, R., Domínguez-Maldonado, J., Pacheco-Catalán, D., Ruíz, H.A. & Tapia-Tussell, R. 2019. Consolidated bioprocess for bioethanol production from raw flour of Brosimum alicastrum seeds using the native strain of Trametes hirsuta Bm-2. Microorganisms, 7: 483. DOI: https://doi.org/10.3390/microorganisms7110483

Pang, J., Hao, M., Li, Y., Liu, J., Lan, H., Zhang, Y., Zhang, Q. & Liu, Z. 2018. Consolidated bioprocessing using Clostridium thermocellum and Thermoanaerobacterium thermosaccharolyticum co-culture for enhancing ethanol production from corn straw. BioResource, 13(4): 8209-8221. DOI: https://doi.org/10.15376/biores.13.4.8209-8221

Periyasamy, S., Isabel, J.B., Kavitha, S., Karthik, V., Mohamed, B.A., Gizaw, D.G., Sivashanmugam, P. & Aminabhavi, T.M. 2023. Recent advances in consolidated bioprocessing for conversion of lignocellulosic biomass into bioethanol – A review. Chemical Engineering Journal, 453(1): 139783. DOI: https://doi.org/10.1016/j.cej.2022.139783

Saini, J.K., Saini, R. & Tewari, L. 2015. Lignocellulosic agriculture wastes as biomass feedstocks for second-generation bioethanol production: Concepts and recent developments. 3 Biotech, 5: 337–353. DOI: https://doi.org/10.1007/s13205-014-0246-5

Sarabana, S.S.H., El-Gabry, K.I.M. & Eldin, A.M. 2015. Optimizing growth conditions provoked ethanol production by fungi grown on glucose. Middle East Journal of Applied Sciences, 5: 1222-1231.

Singhania, R.R., Patel, A.K., Singh, A., Haldar, D., Soam, S., Chen, C.W., Tsai, M.L. & Dong, C.D. 2022. Consolidated bioprocessing of lignocellulosic biomass: Technological advances and challenges. Bioresource Technology, 354: 127153. DOI: https://doi.org/10.1016/j.biortech.2022.127153

Syazwanee, M.M.F., Shaziera, A.N., Izzati, M.N.A., Azwady, A.N. & Muskhazli, M. 2018. Improvement of delignification, desilication and cellulosic content availability in paddy straw via physicochemical pretreatments. Annual Research and Review Biology, 26: 1-11. DOI: https://doi.org/10.9734/ARRB/2018/40947

Syazwanee, M.M.F., Izzati, M.N.A., Azwady, A.N. & Muskhazli, M. 2019. Screening of lignocellulolytic fungi for hydrolyzation of lignocellulosic materials in paddy straw for bioethanol production. Malaysian Journal of Microbiology, 15: 379-386.

Syazwanee, M.M.F., Izzati, M.N.A., Azwady, A.N. & Muskhazli, M. 2021. Improvement of bioethanol production in consolidated bioprocessing (CBP) via consortium of Aspergillus niger B2484 and Trichoderma asperellum B1581. Pertanika Journal of Science and Technology, 29: 301–316. DOI: https://doi.org/10.47836/pjst.29.1.17

Syazwanee, M.M.F., Izzati, M.N.A., Azwady, A.N. & Muskhazli, M. 2022. Consolidated bioethanol production using Trichoderma asperellum B1581. Jordan Journal of Biological Sciences, 15: 621–627. DOI: https://doi.org/10.54319/jjbs/150410

Wilkinson, S., Smart, K.A., James, S. & Cook, D.J. 2017. Bioethanol production from brewers spent grains using a fungal consolidated bioprocessing (CBP) approach. Bioenergy Research, 10(1): 146-157. DOI: https://doi.org/10.1007/s12155-016-9782-7

Wu, Z. 2019. Mixed fermentation of Aspergillus niger and Candida shehatae to produce bioethanol with ionic-liquid-pretreated bagasse. 3 Biotech, 9(2): 41. DOI: https://doi.org/10.1007/s13205-019-1570-6

Wu, J., Elliston, A., Le Gall, G., Colquhoun, I.J., Collins, S.R.A., Wood, I.P., Dicks, J., Roberts, I.N. & Waldron, K.W. 2018. Optimising conditions for bioethanol production from rice husk and rice straw: Effects of pre-treatment on liquor composition and fermentation inhibitors. Biotechnology for Biofuels, 11: 62. DOI: https://doi.org/10.1186/s13068-018-1062-7

Zeghlouli, J., Christophe, G., Guendouz, A., El Modafar, C., Belkamel, A., Michaud, P. & Delattre, C. 2021. Optimization of bioethanol production from enzymatic treatment of argan pulp feedstock. Molecules, 26: 2516. DOI: https://doi.org/10.3390/molecules26092516

Zhao, X., Moates, G.K., Elliston, A., Wilson, D.R., Coleman, M.J. & Waldron, K.W. 2015. Simultaneous saccharification and fermentation of steam exploded duckweed: Improvement of the ethanol yield by increasing yeast titre. Bioresource Technology, 194: 263–269. DOI: https://doi.org/10.1016/j.biortech.2015.06.131

Zhao, J., Shi, D., Yang, S., Lin, H. & Chen, H. 2020. Identification of an intracellular β-glucosidase in Aspergillus niger with transglycosylation activity. Applied Microbiology and Biotechnology, 104: 8367–8380. DOI: https://doi.org/10.1007/s00253-020-10840-4

Zhu, J., Wang, J., An, Z., Shen, C., Dong, H., Wang, H., Peng, Z., Yang, B., Liu, J., Wang, X. & Fang, Z. 2025. Microbial community succession during tobacco fermentation reveals a flavor-improving mechanism. Frontiers in Bioengineering and Biotechnology, 13: 1627842. DOI: https://doi.org/10.3389/fbioe.2025.1627842

Published

21-12-2025

How to Cite

Mohamed Ghazali, M. F. S., Mustafa, M., Mohd Zainudin, N. A. I., & Abd Aziz, N. A. (2025). Optimizing Bioethanol Production Via Consolidated Bioprocessing: The Potential of Aspergillus niger B2484. Malaysian Applied Biology, 54(4), 89–97. https://doi.org/10.55230/mabjournal.v54i4.3230

Issue

Section

Research Articles