Page Header

Influence of Sulfuric Acid Pretreatment and Inhibitor of Sugarcane Bagasse on the Production of Fermentable Sugar and Ethanol

Elizabeth Jayex Panakkal, Malinee Sriariyanun, Jakaphan Ratanapoompinyo, Patchanee Yasurin, Kraipat Cheenkachorn, Wawat Rodiahwati, Prapakorn Tantayotai


Improper disposal of agricultural waste after harvesting season has posed serious health and environmental issues. Alternative methods to utilize agricultural waste to produce a value-added product, especially biofuel, have become the focus of research and industrial stakeholders. To make the process feasible, the maximum conversion should be achieved with the optimum operational condition. This research applied Response Surface Methodology (RSM) with the Box-Behnken design (BBD) to optimize sulfuric acid pretreatment of sugarcane bagasse by varying three pretreatment factors namely, acid concentration (0.5–3.5%), temperature (60–140℃), and time (20–100 min). Pretreated biomass was enzymatically hydrolyzed, and the effectiveness of pretreatment was examined according to the reducing sugar concentration. However, inhibitors namely, acetic acid, 5-hydroxymethylfurfural (5-HMF), and furfural were produced during pretreatment, which was analyzed through GC-MS analysis. The Box-Behnken design could optimize and correlate the effect of pretreatment parameters on the hydrolysis of sugarcane bagasse. The optimum pretreatment condition was predicted at an acid concentration of 3.50%, the temperature of 136.08℃, and the time of 75.36 min to obtain the maximum sugar production. Sugarcane bagasse pretreatment at optimum condition could produce a reducing sugar of 180.15 mg/g-sugarcane bagasse, which is 3.06 folds higher than untreated sugarcane bagasse. However, ethanol yield from pretreated biomass was less than unpretreated biomass because of the inhibitor formation. This study provides a new insight into utilizing agricultural waste in a more efficient and eco-friendly manner.


[1] R. Janta, K. Sekiguchi, R. Yamaguchi, K. Sopajaree, B. Plubin, and T. Chetiyanukornkul, “Spatial and temporal variations of atmospheric PM10 and air pollutants concentration in upper Northern Thailand during 2006–2016,” Applied Science and Engineering Progress, vol. 13, no. 3, 2020, doi: 10.14416/j.asep.2020.03.007.

[2] N. Pasukphun, “Environmental health burden of open burning in northern Thailand: A review,” PSRU Journal of Science and Technology, vol. 3, no. 3, pp.11–28, 2018.
[3] K. Y. Foo and B. H. Hameed, “Transformation of durian biomass into a highly valuable end commodity: Trends and opportunities,” Biomass and Bioenergy, vol. 35, no. 7, pp. 2470–2478, 2011.

[4] S. Sanjaya, “The application of durian rind as a burning stimulant of coal briquettes,” Journal Ilmu dan Teknologi Kayu Tropis, vol. 13, no. 1, pp. 80–87, 2015.

[5] M. Sriariyanun and K. Kitsubthawee, “Trends in lignocellulosic biorefinery for production of value-added biochemicals,” Applied Science and Engineering Progress, vol. 13, no. 4, doi: 10.14416/j.asep.2020.02.005, 2020.

[6] M. Sriariyanun, J. H. Heitz, P. Yasurin, S. Asavasanti, P. Tantayotai, “Itaconic acid: A promising and sustainable platform chemical?,” Applied Science and Engineering Progress, vol. 12, no. 2, pp. 75–82, 2019, doi: 10.14416/j. asep.2019.05.002.

[7] P. Rachmontree, T. Douzou, K. Cheenkachorn, M. Sriariyanun, and K. Rattanaporn, “Furfural: A sustainable platform chemical and fuel,” Applied Science and Engineering Progress, vol. 13, no. 1, pp. 3–10, 2020, doi: 10.14416/j. asep.2020.01.003.

[8] S. C. Yat, A. Berger, and D. R. Shonnard, “Kinetic characterization of dilute surface acid hydrolysis of timber varieties and switchgrass,” Bioresource Technology, vol. 99, pp. 3855–3863, 2008.

[9] Y. Sun and J. Cheng, “Hydrolysis of lignocellulosic materials for ethanol production: A review,” Bioresource Technology, vol. 83, pp. 1–11, 2002.

[10] J. Perez, J. M. Dorado, T. D. De la Rubia, and J. Martinez, “Biodegradation and biological treatment of cellulose, hemicellulose, and lignin: An overview,” International Journal of Microbiology, vol. 5, pp. 53–63, 2002.

[11] S. S. Maleki, K. Mohammadi, and K. Ji, “Characterization of cellulose synthesis in plant cells,” The Scientific World Journal, vol. 2016, doi: 10.1155/2016/8641373.

[12] M. Ptashnyk and B. Seguin, “The impact of microfibril orientations on the biomechanics of plant cell walls and tissues,” Bulletin of Mathematical Biology, vol. 78, no. 11, pp. 2135– 2164, 2016, doi: 10.1007/s11538-016-0207-8.

[13] A. Zoghlami and G. Paës, “Lignocellulosic biomass: Understanding recalcitrance and predicting hydrolysis,” Frontiers in Chemistry, vol. 7, p. 874, 2019, doi: 10.3389/fchem.2019. 00874.

[14] P. Beguin and J. P. Aubert, “The biological degradation of cellulose,” FEMS Microbiology Reviews, vol. 13, pp. 25–58, 1994.

[15] P. Chandranupap and P. Chandranupap, “Enzymatic deinking of xerographic waste paper with non-ionic surfactant,” Applied Science and Engineering Progress, vol. 13, no. 2, pp. 136–145, 2020, doi: 10.14416/j.asep.2020.01.007.

[16] N. S. Mosier, C. Wyman, B. Dale, R. Elander, Y. Y. Lee, M. Holtzapple, and M. Ladisch, “Features of promising technologies for pretreatment of lignocellulosic biomass,” Bioresource Technology, vol. 96, pp. 673–686, 2005.

[17] W. Rodiahwati and M. Sriariyanun, “Lignocellulosic biomass to biofuel production: Integration of chemical and extrusion (screw press) pretreatment,” King Mongkut's University of Technology North Bangkok International Journal of Applied Science and Technology, vol. 9, no. 4, pp. 289–298, 2016, doi: 10.14416/j.ijast.2016.11.001.

[18] M. Sriariyanun, Q. Yan, I. Nowik, K. Cheenkachorn, T. Phusantisampan, and M. Modigell, “Efficient pretreatment of rice straw by combination of screw press and ionic liquid to enhance enzymatic hydrolysis,” Kasetsart Journal (Natural Science), vol. 49, no. 1, pp. 146–154, 2015.

[19] K. Cheenkachorn, T. Douzou, S. Roddecha, P. Tantayotai, and M. Sriariyanun, “Enzymatic saccharification of rice straw under influence of recycled ionic liquid pretreatments,” Energy Procedia, vol. 100, pp. 160–165, 2016.

[20] K. Rattanaporn, S. Roddecha, M. Sriariyanun, and K. Cheenkachorn, “Improving saccharification of oil palm shell by acetic acid pretreatment for biofuel production,” Energy Procedia, vol. 141C, pp. 146–149, 2017.

[21] K. Rattanaporn, P. Tantayotai, T. Phusantisampan, P. Pornwongthong, and M. Sriariyanun, “Organic acid pretreatment of oil palm trunk: Effect on enzymatic saccharification and ethanol production,” Bioprocess and Biosystem Engineering, vol. 41, pp. 467–477, 2018.

[22] Y. S. Cheng, Z. Y. Wu, and M. Sriariyanun, “Evaluation of Macaranga tanarius as a biomass feedstock for fermentable sugars production,” Bioresource Technology, vol. 294, 2019, Art. no. 122195.

[23] A. Boontum, J. Phetsom, W. Rodiahwati, K. Kitsubthawee, and T. Kuntothom, “Characterization of diluted-acid pretreatment of water hyacinth,” Applied Science and Engineering Progress, vol. 12, no. 4, pp. 253–263, 2019, doi: 10.14416/j. asep.2019.09.003.

[24] P. Tantayotai, K. Rattanaporn, S. Tepaamorndech, K. Cheenkachorn, and M. Sriariyanun, “Analysis of an ionic liquid and salt tolerant microbial consortium which is useful for enhancement of enzymatic hydrolysis and biogas production,” Waste and Biomass Valorization, vol. 10, no. 6, pp. 1481–1491, 2019.

[25] P. Tantayotai, P. Pornwongthong, C. Muenmuang, T. Phusantisampan, and M. Sriariyanun, “Effect of cellulase-producing microbial consortium on biogas production from lignocellulosic biomass,” Energy Procedia, vol. 141C, pp. 180–183, 2017.

[26] M. Sriariyanun, P. Tantayotai, P. Yasurin, P. Pornwongthong, and K. Cheenkachorn, “Production, purification and characterization of an ionic liquid tolerant cellulase from Bacillus sp. isolated from rice paddy field soil,” Electronic Journal of Biotechnology, vol. 19, pp. 23–28, 2016.

[27] P. Tantayotai, P. Rachmontree, W. Rodiahwati, K. Rattanaporn, and M. Sriariyanun, “Production of ionic liquid-tolerant cellulase produced by microbial consortium and its application in biofuel production,” Energy Procedia, vol. 100, pp. 155–159, 2016.

[28] P. Kumar, D. M. Barrett, M. J. Delwiche, and P. Stroeve, “Methods for pretreatment of lignocellulosic biomass for efficient hydrolysis and biofuel production,” Industrial and Engineering Chemistry Research, vol. 48, no. 8, pp. 3713– 3729, 2009.

[29] Y. Sun and J. Cheng, “Hydrolysis of lignocellulosic materials for ethanol production: A review,” Bioresource Technology, vol. 83, pp. 1–11, 2002.

[30] D. F. Root, J. F. Saeman, and J. F. Harris, “Kinetics of the acid-catalyzed conversion of xylose to furfural,” Forest Products Journal, vol. 158, p. 165, 1959.

[31] A. Esteghlalian, A. G. Hashimoto, J. J. Fenske, and M. H. Penner, “Modeling and optimization of the dilute-sulfuric-acid pretreatment of corn stover, poplar, and switchgrass,” Bioresource Technology, vol. 59, pp. 129–136, 1997.

[32] N. S. Mosier, C. Wyman, B. Dale, R. Elander, Y. Y. R. Lee, M. Holtzapple, and M. R. Ladisch, “Features of promising technologies for pretreatment of lignocellulosic biomass,” Bioresource Technology, vol. 96, pp. 673–686, 2005.

[33] J. D. McMillan, M. E. Himmel, J. O. Baker, and R. P. Overend, “Pretreatment of lignocellulosic biomass,” in Enzymatic Conversion of Biomass for Fuels Production. Washington, DC: American Chemical Society, 1994, pp. 292–324.

[34] C.-G. Liu, K. Li, Y. Wen, B-Y. Geng, Q. Liu, and Y-H. Lin, “Bioethanol: New opportunities for an ancient product,” in Advances in Bioenergy. Amsterdam, Netherlands: Elsevier, 2019.

[35] J. Xu, “Microwave pretreatment,” in Pretreatment of Biomass. Amsterdam, Netherlands: Elsevier, 2015.

[36] A. Brangule, R. Šukele, and D. Bandere, “Herbal medicine characterization perspectives using advanced FTIR sample techniques– diffuse reflectance (DRIFT) and photoacoustic spectroscopy (PAS),” Frontiers in Plant Science, vol. 11, 2020, doi: 10.3389/fpls.2020.00356.

[37] P. J. Whitcomb and M. J. Anderson, RSM Simplified: Optimizing Processes Using Response Surface Methods for Design of Experiments. Florida: CRC Press, 2004.

[38] G. L. Miller, “Use of dinitrosalicylic acid reagent for determination of reducing sugar,” Analytical Chemistry, vol. 31, no. 3, pp. 426–428, 1959.

[39] Forage Fiber Analyses (Apparatus, Reagents, Procedures, and Some Applications). Agriculture Handbook no. 379, Agriculture Research Service USDA, Washington (DC), 1970, pp. 20.

[40] M. Sriariyanun, P. Mutrakulcharoen, S. Tepaamorndech, K. Cheenkachorn, and K. Rattanaporn, “A rapid spectrophotometric method for quantitative determination of ethanol in fermentation products,” Oriental Journal of Chemistry, vol. 35, no. 2, 2019, doi: 10.13005/ ojc/350234.

[41] L. Canilha, V. T. Santos, G. J. Rocha, J. B. A. e Silva, M. Giulietti, S. S. Silva, and W. Carvalho, “A study on the pretreatment of a sugarcane bagasse sample with dilute sulfuric acid,” Journal of Industrial Microbiology and Biotechnology, vol. 38, no. 9, pp. 1467–1475, 2011.

[42] S. G. Rueda, R. A. Rafael, G. S. Carlos, C. C. Aline, and R. M. Filho, “Pretreatment of sugar cane bagasse with phosphoric and sulfuric diluted acid for fermentable sugars production by enzymatic hydrolysis,” Chemical Engineering Transactions, vol. 20, pp. 321–326, 2010.

[43] I. B. Soares, K. C. S. Mendes, M. Benachour, and C. A. M. Abreu, “Evaluation of the effects of operational parameters in the pretreatment of sugarcane bagasse with diluted sulfuric acid using analysis of variance,” Chemical Engineering Communications, vol. 204, no. 12, pp. 1369– 1390, 2017.

[44] N. Sritrakul, S. Nitisinprasert, and S. Keawsompong, “Evaluation of dilute acid pretreatment for bioethanol fermentation from sugarcane bagasse pith,” Agriculture and Natural Resources, vol. 51, no. 6, pp. 512–519, 2017.

[45] R. Timung, N. Naik Deshavath, V. V. Goud, and V. V. Dasu, “Effect of subsequent dilute acid and enzymatic hydrolysis on reducing sugar production from sugarcane bagasse and spent citronella biomass,” Journal of Energy, vol. 2016, no. 4, pp. 1–12, 2016.

[46] S. Youssefi, Z. Emam-Djomeh, and S. M. Mousavi, “Comparison of artificial neural network (ANN) and response surface methodology (RSM) in the prediction of quality parameters of spray-dried pomegranate juice,” Drying Technology, vol. 27, no. 7–8, pp. 910–917, 2009.
[47] P. Amnuaycheewa, R. Hengaroonprasan, K. Rattanaporn, S. Kirdponpattara, K. Cheenkachorn, and M. Sriariyanun, “Enhancing enzymatic hydrolysis and biogas production from rice straw by pretreatment with organic acids,” Industrial Crops and Products, vol. 84, pp. 247–254, 2016.

[48] P. Tantayotai, P. Mutrakulchareon, A. Tawai, S. Roddecha, and M. Sriariyanun, “Effect of organic acid pretreatment of water hyacinth on enzymatic hydrolysis and biogas and bioethanol production,” IOP Conference Series: Earth and Environmental Science, vol. 346, 2019, doi: 10.1088/1755-1315/346/1/012004.

[49] S. Tiwari, J. Yadav, R. Gaur, and J. S. Yadav, “Assessment of a novel pretreatment techniques for enhancing the enzymatic saccharification of sugarcane bagasse: Structural and chemical analysis,” Research Square, 2020, doi: 10.21203/

[50] R. I. S. L-Azar, T. Morgan, G. P. M-Alfenas, and V. M. Guimaraes, “Inhibitors compounds on sugarcane bagasse saccharification: Effects of pretreatment methods and alternatives to decrease inhibition,” Applied Biochemistry and Biotechnology, vol. 188, pp. 29–42, 2019.

[51] S. Niju and M. Swathika, “Delignification of sugarcane bagasse using pretreatment strategies for bioethanol production,” Biocatalysis and Agriculture Biotechnology, vol. 20, 2019, Art. no. 101263.

[52] S. A. Allen, W. Clark, M. McCaffery, Z. Cai, A. Lanctot, P. J, Slininger, Z. L. Liu, and S. W. Gorsich, “Furfural induces reactive oxygen species accumulation and cellular damage in Saccharomyces cerevisiae,” Biotechnology for Biofuels, vol. 3, no. 2, 2010, doi: 10.1186/1754- 6834-3-2.

[53] L. J. Jönsson, B. Alriksson, and N. Nilvebrant, “Bioconversion of lignocellulose: Inhibitors and detoxification,” Biotechnology for Biofuels, vol. 6, no. 1, pp. 16–26, 2013.

[54] N. Sjulander and T. Kikas, “Origin, impact and control of lignocellulosic inhibitors in bioethanol production- A review,” Energies, vol. 13, no. 18, 2020, doi: 10.3390/en13184751.

[55] Y. Zha, B. Muilwijk, L. Coulier, and P. J. Punt, “Inhibitory compounds in lignocellulosic biomass hydrolysates during hydrolysate fermentation processes,” Journal of Bioprocessing and Biotechniques, vol. 2, no. 1, 2012, doi: 10.4172/2155-9821.1000112.

[56] M. Neureiter, H. Danner, C. Thomasser, B. Saidi, and R. Braun, “Dilute-acid hydrolysis of sugarcane bagasse at varying conditions,” Applied Biochemistry and Biotechnology, vol. 98, no. 1–9, pp. 49–58, 2002.

[57] Y. H. Jung and K. H. Kim, “Evaluation of the main inhibitors from lignocellulose pretreatment for enzymatic hydrolysis and yeast fermentation,” BioResources, vol. 12, no. 4, pp. 9348–9356, 2017.

[58] X-B. Zhao, L. Wang, and D-H. Liu, “Peracetic acid pretreatment of sugarcane bagasse for enzymatic hydrolysis: A continued work,” Journal of Chemical Technology and Biotechnology, vol. 83, pp. 950–956, 2008.

[59] P. P-Barahona, E. J. C-Barriga, J. M-Gil, and P. M-Ramos, “Sugarcane bagasse hydrolysis enhancement by Microwave-Assisted Sulfolane pretreatment,” Energies, vol. 12, no. 9, 2019, doi: 10.3390/en12091703.

[60] R. G. Hemansi, V. K. Aswal, and J. K. Saini, “Sequential dilute acid and alkali deconstruction of sugarcane bagasse for improved hydrolysis: Insight from small angle neutron scattering (SANS),” Renewable Energy, vol. 147, pp. 2091– 2101, 2020.

[61] L-Q. Wang, L-Y. Cai, and Y-L. Ma, “Study on inhibitors from acid pretreatment of corn stalk on ethanol fermentation by alcohol yeast,” Royal Society of Chemistry, vol. 10, pp. 38409–38415, 2020, doi: 10.1039/d0ra04965d.

[62] N. Sritrakul, S. Nitisinprasert, and S. Keawsompong, “Evaluation of dilute acid pretreatment for bioethanol fermentation from sugarcane bagasse pith,” Agriculture and Natural Resources, vol. 51, no. 6, pp. 512–519, 2017.

[63] E. S. Lopes, K. Dominices, M. Lopes, L. Tovar, M. R. Filho, “Enzymatic hydrolysis exploration and fermentation: Acid pretreatment and delignification in sugarcane bagasse for 2G ethanol production,” Chemical Engineering Transactions, vol. 57, pp. 151–156, 2017.
[64] K. J. Dussan, D. D. V. Silva, E. J. C. Moraes, P. V. Arruda, and M. G. A. Felipe, “Dilute-acid hydrolysis of cellulose to glucose from sugarcane bagasse,” Chemical Engineering Transactions, vol. 38, pp. 433–438, 2014, doi: 10.30303/ CET1438073.

[65] C. S. Lima, T. Neitzel, I. de Oliveira Pereira, S. C. Rabelo, J. L. Ienczak, I. C. Roberto, and G. J. M. Rocha, “Effect of the sugarcane bagasse deacetylation in the pentoses fermentation process,” BioEnergy Research, 2021, doi: 10. 1007/s12155-020-10243-3.

[66] P. Baral, L. Jain, A. K. Kurmi, V. Kumar, and D. Agrawal, “Augmented hydrolysis of acid pretreated sugarcane bagasse by PEG 6000 addition: A case study of cellic CTec2 with recycling and reuse,” Bioprocess and Biosystems Engineering, vol. 43, no. 3, pp. 473–482, 2020, doi: 10.1007/s00449- 019-02241-3.

[67] T. C. G. Oliveira, K. E. Hanlon, M. A. Interlandi, P. C. Torres-Mayanga, M. A. C. Silvello, D. Lachos-Perez, M. T. Timko, M. A. Rostagno, R. Goldbeck, and T. Forster-Carneiro, “Subcritical water hydrolysis pretreatment of sugarcane bagasse to produce second generation ethanol,” The Journal of Supercritical Fluids, 2020, doi:10.1016/j.supflu.2020.104916.

Full Text: PDF

DOI: 10.14416/j.asep.2021.07.006


  • There are currently no refbacks.