Ozonolysis Pretreatment of Wheat Straw for Enhanced Delignification: Applying RSM Technique for Modeling and Optimizing Process

Document Type : Research Paper


1 Department of Agrotechnology, College of Aburaihan, University of Tehran, Pakdasht, Tehran, Iran

2 Department of Agrotechnology, College of Abouraihan, University of Tehran, Tehran, Iran

3 Department of Animal and Poultry Science, College of Abouraihan, University of Tehran, Tehran, Iran

4 Department of Entomology and Plant Pathology, College of Abouraihan, University of Tehran, Tehran, Iran


Wheat straw is widely used for animal feed and biorefinery to sugar production. However, cellulose, which is the major source of sugar, is protected by lignin. Ozone is a powerful oxidizer that can deconstruct lignin and makes cellulose accessible to enzymatic digestion. The aim of this study is the delignification of wheat straw using ozonolysis technology as a green, environmentally friendly, and energy-efficient process. Modeling and optimization were performed by response surface methodology and the effects of five factors including ozone production rate (1, 2, and 3 g/h), reaction time (15, 30, and 45 min), the flow rate of ozone/oxygen (0, 3, and 6 L/min), moisture content (100, 200, and 300% w/w) and urea (0, 1.5, and 3% w/w) were investigated. The results of this study showed that the highest delignification (50%) was observed in the highest levels of ozone amount, time, flow rate, urea, and the lowest level of moisture content. The flow rate and moisture content factors were the most contributing factors in the delignification process with 36 and 20%, respectively. Under optimal conditions, 49.8% of delignification was obtained at an ozone amount (3 g/h), time (45 min), flow rate (5.7 L/min), moisture content (100% w/w), and urea amount (3% w/w) with a desirability index of 0.99. The response surface method creates a desirable fitness between the experimental and predicted responses with R2=0.90. The results showed that the ozonolysis process using the proposed factors can be used to delignification of wheat straw.


Ai, P., Zhang, X., Dinamarca, C., Elsayed, M., Yu, L., Xi, J. & Mei, Z. (2019). Different effects of ozone and aqueous ammonia in a combined pretreatment method on rice straw and dairy manure fiber for enhancing biomethane production. Bioresource Technology, 282, 275-284.
Al jibouri, A.K.H. (2012). Effect of intermediate washing on ozonolysis delignification and enzymatic hydrolysis of wheat straw. Master Thesis, Ryerson University, Toronto, Canada.
Al jibouri, A.K.H., Turcotte, G., Wu, J. & Cheng, C.H. (2015). Ozone pretreatment of humid wheat straw for biofuel production. Energy Science and Engineering, 3(6), 541-548.
ASAE Standards: ASAE S319.3. (2006). Method of determining and expressing fineness of feed materials by sieving, (pp, 601-605).
ASAE Standards: ASAE S358.2. (2003). Moisture measurement-forages, (pp, 607-608).
Baig, K.S., Wu, J., Turcotte, G. & Doan, H.D. (2015). Novel ozonation technique to delignify wheat straw for biofuel production. Energy and Environment, 26(3), 303-318.
Barrera-Martínez, I., Guzman, N., Pena, E., Vazquez, T., Ceron-Camacho, R., Folch, J., Salazar, J.A.H. & Aburto, J. (2016). Ozonolysis of alkaline lignin and sugarcane bagasse: Structural changes and their effect on saccharification. Biomass and Bioenergy, 94, 167-172.
Ben’ko, E.M., Chukhchin, D.G. & Lunin, V.V. (2017). Ozone pretreatment and fermentative hydrolysis of wheat straw. Zhurnal Fizicheskoi Khimii, 91(11), 1851-1857.
Ben’ko, E.M. & Lunin, V.V. (2020). Patterns of the ozone pretreatment of lignocellulosic biomass for subsequent fermentation into sugars. Russian Journal of Physical Chemistry A, 94(9), 1943-1948.
Ben’ko, E.M., Chukhchin, D.G. & Lunin, V.V. (2020). Changes in wheat straw cell walls during ozone pretreatment. Holzforschung, 74(12), 1157-1167.
Binder, A., Pelloni, L. & Fiechter, A. (1980). Delignification of straw with ozone to enhance biodegradability. European journal of applied microbiology and biotechnology, 11, 1-5.
Box, G.E.P. & Hunter, J.S. (1957). Multi-factor experimental design for exploring response surfaces. Annals of Mathematical Statistics, 28, 195-241.
Bule, M.V., Gao, A.H., Hiscox, B. & Chen, S. (2013). Structural modification of lignin and characterization of pretreated wheat straw by ozonation. Journal of Agricultural and Food Chemistry, 61, 3916-3925.
Chen, X., Du, W. & Liu, D. (2008). Response surface optimization of biocatalytic biodiesel production with acid oil. Biochemical Engineering Journal, 40, 423-429.
Choi, H., Lim, H.N., Kim, J., Hwang, T.M. & Kang, J.W. (2002). Transport characteristics of gas phase ozone in unsaturated porous media for in-situ chemical oxidation. Journal of Contaminant Hydrology, 57, 81-98.
Contreras, S. (2002). Degradation and biodegradability enhancement of nitrobenzene and 2,4-dichlorophenol by means of advanced oxidation processes based on ozone. Ph. D Thesis, University of Barcelona.
Das, A., Chanchal, M. & Roy, S. (2015). Pretreatment methods of ligno-cellulosic biomass: a review. Journal of Engineering Science and Technology Review, 8(5), 141-165.
Derringer, G. & Suich, R. (1980). Simultaneous optimization of several response variables. Journal of Quality Technology, 12, 214-219.
Domański, J., Marchut-Mikołajczyk, O., Polewczyk, A. & Januszewicz, B. (2017). Ozonolysis of straw from Secale cereale L. for anaerobic digestion. Bioresource Technology, 245, 394-400.
FAO. (2020). Crop Prospects and Food Situation - Quarterly Global Report No. 1, March 2020, Rome.
García-Cubero, M.T., Coca, M., Bolado, S. & González-Benito, G. (2010). Chemical oxidation with ozone as pre-treatment of lignocellulosic materials for bioethanol production. Chemical Engineering Transactions, 21, 1273-1278.
García-Cubero, M.T., González-Benito, G., Indacoechea, I., Coca, M. & Bolado, S. (2009). Effect of ozonolysis pretreatment on enzymatic digestibility of wheat and rye straw. Bioresource Technology, 100, 1608-1613.
García-Cubero, M.T., Palacn, L.G., Gonzlez-Benito, G., Bolado, S., Lucas, S. & Coca, M. (2012). An analysis of lignin removal in a fixed bed reactor by reaction of cereal straws with ozone. Bioresource Technology, 107, 229-234.
Ghorbani, M., Aboonajmi, M., Ghorbani Javid, M. & Arabhosseini, A. (2017). Optimization of ultrasound-assisted extraction of ascorbic acid from fennel (Foeniculum vulgare) seeds and evaluation its extracts in free radical scavenging. Agricultural Engineering International: CIGR Journal, 19(4), 209-218. (In Farsi)
Ghorbani, M., Kianmehr, M.H., Arabhosseini, A., Sarlaki, E., Asadi Alamouti, A. & Sadeghi, R. (2020a). A review on ozone: properties, effects, reaction mechanisms, environmental and safety aspects in food processing. In: Proceedings of 12th National Congress on Biosystems Engineering and Agricultural Mechanization. 5-7 February, Shahid Chamran University of Ahvaz, Iran. (In Farsi)
Ghorbani, M., Kianmehr, M.H., Arabhosseini, A., Sarlaki, E., Asadi Alamouti, A. & Sadeghi, R. (2020b). Fundamental of generating, applying and controlling systems of plasma ozone and its applications in the food industry. In: Proceedings of 12th National Congress on Biosystems Engineering and Agricultural Mechanization. 5-7 February, Shahid Chamran University of Ahvaz, Iran. (In Farsi)
Ghorbani, M., Kianmehr, M.H., Arabhosseini, A., Sarlaki, E., Asadi Alamouti, A. & Sadeghi, R. (2020c). Ozonolysis: a novel and effective oxidation technique for lignocellulosic biomass pretreatment. In: Proceedings of 12th National Congress on Biosystems Engineering and Agricultural Mechanization. 5-7 February, Shahid Chamran University of Ahvaz, Iran. (In Farsi)
Gitifar, V., Eslamloueyan, R. & Sarshar, M. (2013). Experimental study and neural network modeling of sugarcane bagasse pretreatment with H2SO4 and O3 for cellulosic material conversion to sugar. Bioresource Technology, 148, 47-52.
Goede, A. & van de Sanden, R. (2016). The need for basic energy research. Europhysics News, 47: 22-26.
Graves, C., Ebbesen, S.D., Mogensen, M. & Lackner, K.S. (2011). Sustainable hydrocarbon fuels by recycling CO2 and H2O with renewable or nuclear energy. Renewable and Sustainable Energy Reviews, 15(1), 1-23.
Harrington, E.C. (1965). The Desirability Function. Industrial Quality Control, 21, 494-498.
Heiske, S., Schultz-Jensen, N., Leipold, F. & Schmidt, J.E. (2013). Improving anaerobic digestion of wheat straw by plasma-assisted pretreatment. Journal of Atomic and Molecular Physics, 2013, 7 pages.
Huang, H., Wang, Z., Pan, S.C., Shoup, L.M., Felix, T.L., Perkins, J.B., May, O. & Singh, V. (2017). Fungal pretreatment to improve digestibility of corn stover for animal feed. Transactions of the ASABE, 60(3), 973-979.
Kádár, Z., Schultz-Jensen, N., Jensen, J.S., Hansen, M.A.T., Leipold, F. & Bjerre, A.B. (2015). Enhanced ethanol production by removal of cutin and epicuticular waxes of wheat straw by plasma assisted pretreatment. Biomass and Bioenergy, 81, 26-30.
Kaur, U., Oberoi, H.S., Bhargav, V.K., Sharma-Shivappa, R. & Dhaliwal, S.S. (2012). Ethanol production from alkali- and ozone-treated cotton stalks using thermotolerant Pichia kudriavzevii HOP-1. Ind. Industrial Crops and Products, 37, 219-226.
Kumar, P., Barrett, D.M., Delwiche, M.J. & Stroeve, P. (2009). Methods for pretreatment of lignocellulosic biomass for efficient hydrolysis and biofuel production. Industrial & Engineering Chemistry Research, 48(8), 3713-3729.
Li, C., Wang, L., Chen, Z., Li, Y.Y., Wang, R., Luo, X., Cai, G., Yu, Q. & Lu, J. (2015). Ozonolysis pretreatment of maize stover: the interactive effect of sample particle size and moisture on ozonolysis process. Bioresource Technology, 183, 240-247.
Maia, E.P. & Colodette, J.L. (2003). Efeito do conteúdo e da natureza da lignina residual na eficiência e na seletividade do branqueamento com ozônio. Revista Árvore, 27, 217-232.
Mamleeva, N.A., Autlov, S.A., Bazarnova, N.G. & Lunin, V.V. (2009). Delignification of softwood by ozonation. Pure and Applied Chemistry, 81, 2081-2091.
Miura, T., Lee, S.H., Inoue, S. & Endo, T. (2012). Combined pretreatment using ozonolysis and wet-disk milling to improve enzymatic saccharification of Japanese cedar. Bioresource Technology, 126, 182-186.
Moore-Landecker. E. (1996). Fundamentals of the fungi. Prentice Hall. Simon and Schuster/Aviacom company, (pp, 376-383).
Mourabet, M., El Rhilassi, A., El Boujaady, H., Bennani-Ziatni, M. & Taitai, A. (2017). Use of response surface methodology for optimization of fluoride adsorption in an aqueous solution by Brushite. Arabian Journal of Chemistry, 10, S3292-S3302.
Mussatto, S.I. (2016). Biomass fractionation technologies for a lignocellulosic feedstock based biorefinery. Publisher: John Fedor. ISBN: 978-0-12-802323-5, 674 Pages.
Napier-Munn, T.J. (2000). The central composite rotatable design JKMRC. Brisbane: The University of Queensland.
Nayan, N., Sonnenberg, A., Hendriks, W. & Cone, J. (2018). Screening of white‐rot fungi for bioprocessing of wheat straw into ruminant feed. Journal of Applied Microbiology, 125, 468-479.
Nayan, N., van Erven, G., Kabel, M.A., Sonnenberg, A.S., Hendriks, W.H. & Cone, J.W. (2019). Improving ruminal digestibility of various wheat straw types by white-rot fungi. Journal of the Science of Food and Agriculture, 99(2), 957-965.
Neely, W.C. (1984). Factors affecting the pretreatment of biomass with gaseous ozone. Biotechnology and Bioengineering, 26, 59-65.
Osuna-Laveaga, D.R., García-Depraect, O., Vallejo-Rodríguez, R., López-López, A. & León-Becerril, E. (2020). Integrated ozonation-enzymatic hydrolysis pretreatment of sugarcane bagasse: Enhancement of sugars released to expended ozone ratio. Processes, 8(10), 1274.
Panneerselvam, A., Sharma-Shivappa, R.R., Kolar, P., Ranney, T. & Peretti, S. (2013). Potential of ozonolysis as a pretreatment for energy grasses. Bioresource Technology, 148, 242-248.
Pedersen, J.F., Vogel, K.P. & Funnell, D.L. (2005). Impact of reduced lignin on plant fitness. Crop Science, 45, 812-819.
Rosen, Y., Mamane, H. & Gerchman, Y. (2019). Short ozonation of lignocellulosic waste as energetically favorable pretreatment. BioEnergy Research, 12, 292-301.
Sarlaki, E., Sharif Paghaleh, A., Kianmehr, M.H. & Mirsaeedghazi, H. (2017). Effect of processing temperature on membrane ultrafiltration of lignite coals-derived humic alkaline extracts, membrane performance and humic acid purity. Iranian Journal of Biosystems Engineering, 48, 475-489. (In Farsi)
Sarlaki, E., Sharif Paghaleh, A., Kianmehr, M.H. & Asefpour Vakilian, K. (2019a). Extraction and purification of humic acids from lignite wastes using alkaline treatment and membrane ultrafiltration. Journal of Cleaner Production, 235, 712-723.
Sarlaki, E., Sokhandan Toomaj, M., Sharif Paghaleh, A, Kianmehr, M.H. & Nikousefat, O. (2019b). Extraction of humic acid from lignite coals using stirred tank reactors (STRs): Assessment of process parameters and final product charaterization. Iranian Journal of Soil and Water Research, 50, 1111-1125. (In Farsi)
Sarlaki, E., Sharif Paghaleh, A., Kianmehr, M.H. & Asefpour Vakilian, K. (2020). Chemical, spectral and morphological characterization of humic acids extracted and membrane purified from lignite. Chemistry and Chemical Technology, 14(3), 353-361.
Sarlaki, E., Sharif Paghaleh, A., Kianmehr, M.H. & Asefpour Vakilian, K. (2021). Valorization of lignite wastes into humic acids: Process optimization, energy efficiency and structural features analysis. Renewable Energy, 163, 105-122.
Schultz-Jensen, N., Kádár, Z., Thomsen, A., Bindslev, H. & Leipold, F. (2011). Plasmaassisted pretreatment of wheat straw for ethanol production. Applied Biochemistry and Biotechnology, 165, 1010-1023.
Schultz-Jensen, N., Thygesen, A., Leipold, F., Thomsen, S.T., Roslander, C., Lilholt, H. & Bjerre, A.B. (2013). Pretreatment of the macroalgae Chaetomorpha linum for the production of bioethanol-comparison of five pretreatment technologies. Bioresource Technology, 140, 36-42.
Sen, R.K. (1997). Response surface optimization of the critical media components for the production of surfactin. Journal of Chemical Technology and Biotechnology, 68, 263-270.
Severe, J. & ZoBell, D.R. (2012). Review: Technical aspects for the utilization of small grain straws as feed energy sources for ruminants: Emphasis on beef cattle. AG/BeefCattle/2012-03, UtahState University Cooperative Extension from www. extension.usu.edu.
Shariat Panahi, H.K., Dehhaghi, M., Aghbashlo, M., Karimi, K. & Tabatabaei, M. (2020). Conversion of residues from agro-food industry into bioethanol in Iran: An under-valued biofuel additive to phase out MTBE in gasoline. Renewable Energy, 145, 699-710.
Sharif Paghaleh, A., Sarlaki, E., Kianmehr, M.H. & Shakiba, N. (2017). Study of spectral, structural and chemical characteristics of humic acids isolated from coalfield of Iran. Iranian Journal of Soil and Water Research, 48, 1145-1158. (In Farsi)
Shaw, M. & Tabil, L. (2006): Mechanical properties of selected biomass grinds. In: ASABE Annual International Meeting. July 9-12, Portland.
Shi, F., Xiang, H. & Li, Y. (2015). Combined pretreatment using ozonolysis and ball milling to improve enzymatic saccharification of corn straw. Bioresource Technology, 179, 444-451.
Song, S., Xia, M., He, Z., Ying, H., Lü, B. & Chen, J. (2007). Degradation of p-nitrotoluene in aqueous solution by ozonation combined with sonolysis. Journal of Hazardous Materials, 144, 532-537.
Souza-Correa, J.A., Oliveira, C., Wolf, L.D., Nascimento, V.M., Rocha, J.G.M. & Amorim, J. (2013). Atmospheric pressure plasma pretreatment of sugarcane bagasse: the influence of moisture in the ozonation process. Applied Biochemistry and Biotechnology, 171, 104-116.
Staehelin, J. & Hoigne, J. (1985). Decomposition of ozone in water in the presence of organic solutes acting as promoters and inhibitors of radical chain reactions. Environmental Science and Technology, 19(12), 1206-1213.
Sundstol, F., Coxwoth, E. & Mowat, D.N. (1978). Improving the nutritive value of straw and other low-quality roughages by treatment with ammonia. World Animal Review, 26, 13-21.
Tajinia, R., Kianmehr, M.H., Sarlaki, E., Sharif Paghaleh A. & Mirsaeedghazi, H. (2020). Extracting humic acids from spend mushroom compost (SMC) by alkaline treatment and membrane ultrafiltration. Iranian Journal of Biosystems Engineering, 50:847-861. (In Farsi)
Talebnia, F., Karakashev, D. & Angelidaki, I. (2010). Production of bioethanol from wheat straw: an overview on pretreatment, hydrolysis and fermentation. Bioresource Technology, 101, 4744-4753.
Travaini, R., Marangon-Jardim, C., Colodette, J.L., Morales-Otero, M.D. & Bolado-Rodríguez, S. (2014). In: Pandey, A., Negi, S., Binod, P. and Larroche, C. (Eds.), Pretreatment of biomass-processes and technologies,(pp, 105-135), Academic Press, USA.
Travaini, R., Martín-Juárez, J., Lorenzo-Hernando, A. & Bolado-Rodríguez, S. (2016). Ozonolysis: An advantageous pretreatment for lignocellulosic biomass revisited. Bioresource Technology, 199, 2-12.
Travaini, R., Otero, M.D.M., Coca, M., Da-Silva, R. & Bolado, S. (2013). Sugarcane bagasse ozonolysis pretreatment: Effect on enzymatic digestibility and inhibitory compound formation. Bioresource Technology, 133, 332-339.
Van Kuijk, S.J.A., Sonnenberg, A.S.M., Baars, J.J.P., Hendriks, W.H., del Río, J.C., Rencoret, J., Gutiérrez, A., de Ruijter, N.C.A. & Cone, J.W. (2017). Chemical changes and increased degradability of wheat straw and oak wood chips treated with the white rot fungi Ceriporiopsis subvermispora and Lentinula edodes. Biomass and Bioenergy, 105, 381-391.
Van Soest, P.J., Robertson, J.B. & Lewis, B.A. (1991). Methods for dietary fiber, neutral detergent fiber, and nonstarch polysaccharides in relation to animal nutrition. Journal of Dairy Science, 74, 3583-3597.
Wu, J., Ein-Mozaffari, F. & Upreti, S. (2013). Effect of ozone pretreatment on hydrogen production from barley straw. Bioresource Technology, 144, 344-349.
Yang, Z.H., Huang, J., Zeng, G.M., Ruan, M., Zhou, C.S., Li, L. & Rong, Z.G. (2009). Optimization of flocculation conditions for kaolin suspension using the composite flocculant of MBFGA1 and PAC by response surface methodology. Bioresource Technology, 100(3), 4233-4239.
Yokota, S., Iizuka, K., Ishiguri, F., Abe, Z. & Yoshizawa, N. (2006). Ozone-dioxane delignification from the cell walls of Japanese cypress (Chamaecyparis obtuse Endl.). Journal of Material Cycles and Waste Management, 8, 140-144.
Yu, Z., Jameel, H., Chang, H.M. & Park, S. (2011). The effect of delignification of forest biomass on enzymatic hydrolysis. Bioresource Technology, 102, 9083-9089.