Abstract
Brown rice, unlike white rice, retains its bran and germ layers, making it rich in dietary fiber and physiologically active compounds. However, these components lower the efficiency of starch hydrolysis, making brown rice less suitable for fermentation. In this study, we aimed to optimize the brown rice liquefaction and saccharification processes using response surface methodology. In a Box–Behnken design, pH, temperature, and time served as independent variables, while soluble solid, reducing sugar, total sugar, maltose, and glucose concentrations as dependent variables. The liquefaction process demonstrated a good fit and reliability, with P < 0.05 and the coefficient of determination (R2) between 0.9290 and 0.9903 for all models. However, for the saccharification process, only the reducing sugar, maltose, and glucose models showed P < 0.05 and R2 ≥ 0.9. We then subjected the sugar syrup prepared under the optimized saccharification conditions (pH 3.5, 65 °C, 4.8 h) to yeast fermentation, achieving an ethanol content of 62.77 mg/mL. Our study was successful in improving brown rice fermentation efficiency through enzymatic liquefaction and saccharification processes. The results may serve as crucial foundational data for the fermented beverage industry.
Similar content being viewed by others
Data availability
Data will be made available on request from the corresponding author.
References
Jaswal, D., Bhushan, K., Kocher, G. S. & Singh, A. Optimization of pretreatment of potato waste for release of fermentable sugars for vodka production: Response surface methodology approach. Biomass Convers. Biorefin. 15, 27593–27605 (2024).
Jeon, H. J., Yu, J. C., Kim, G. W. & Kong, H. S. Quality characteristics of Takju by yeast strain type. Korean J. Food Nutr. 27, 971–978 (2014).
Baek, C. H. et al. Quality characteristics of brown rice makgeolli produced under differing conditions. Korean J. Microbiol. Biotechnol. 41, 168–175 (2013).
Lee, S. W. et al. Quality characteristics of brown rice vinegar by different yeasts and fermentation condition. J. Korean Soc. Food Sci. Nutr. 39, 1366–1372 (2010).
Pino, A. et al. Formulation of germinated brown rice fermented products functionalized by probiotics. Innov. Food Sci. Emerg. Technol. 80, 103076 (2022).
Woo, S. M. et al. Effect of α-amylase treatment of brown rice (Goami) alcohol fermentation by-product. Food Sci. Preserv. 14, 617–623 (2007).
Spinosa, W. A., Santos Júnior, V. D., Galvan, D., Fiorio, J. L. & Gomez, R. J. H. C. Syrup production via enzymatic conversion of a byproduct (broken rice) from rice industry. Acta Sci. Technol. 38, 13–22 (2016).
Espinosa-Ramírez, J., Pérez-Carrillo, E. & Serna-Saldívar, S. O. Maltose and glucose utilization during fermentation of barley and sorghum lager beers as affected by β-amylase or amyloglucosidase addition. J. Cereal Sci. 60, 602–609 (2014).
Mondal, P., Sadhukhan, A. K., Ganguly, A. & Gupta, P. Optimization of process parameters for bio-enzymatic and enzymatic saccharification of waste broken rice for ethanol production using response surface methodology and artificial neural network–genetic algorithm. 3 Biotech. 11, 28 (2021).
Gong, E. S. et al. Phytochemical profiles and antioxidant activity of brown rice varieties. Food Chem. 227, 432–443 (2017).
Saleh, A. S. M., Wang, P., Wang, N., Yang, L. & Xiao, Z. Brown rice versus white rice: Nutritional quality, potential health benefits, development of food products, and preservation technologies. Compr. Rev. Food Sci. Food Saf. 18, 1070–1096 (2019).
Seo, W. D. et al. Comparative analysis of physicochemicals and antioxidative properties in new red rice (Oryza sativa L. cv. Gunganghongmi). J. Crop Sci. Biotechnol. 16, 63–68 (2013).
Vo Minh, H. & Nguyen Duc, T. Response surface optimization of enzymatic hydrolysis of germinated brown rice for higher reducing sugar production. Viet J. Food Control. 5, 645–657 (2022).
Zahra, N. & Jabeen, S. Brown rice as useful nutritional source. Pak J. Agric. Res. 33, 445 (2020).
Miller, G. L. Use of dinitrosalicylic acid reagent for determination of reducing sugar. Anal. Chem. 31, 426–428 (1959).
DuBois, M., Gilles, K. A., Hamilton, J. K., Rebers, P. A. & Smith, F. Colorimetric method for determination of sugars and related substances. Anal. Chem. 28, 350–356 (1956).
Klein, H. & Leubolt, R. Ion-exchange high-performance liquid chromatography in the brewing industry. J. Chromatogr. A. 640, 259–270 (1993).
Abedi, E., Kaveh, S., Mohammad, B. & Hashemi, S. M. B. Structure-based modification of a-amylase by conventional and emerging technologies: comparative study on the secondary structure, activity, thermal stability and amylolysis efficiency. Food Chem. 437, 137903 (2024).
Kim, H. R., Kwon, Y. H., Jo, S. J., Kim, J. H. & Ahn, B. H. Characterization and volatile flavor components in glutinous rice wines prepared with different yeasts of Nutuks. Korean J. Food Sci. Technol. 41, 296–301 (2009).
Cho, J. H. Effects of growing climatic environment on growth, soluble sugar, and enzyme activity changes in Asian pear fruits. PhD Thesis (Chonnam Natl Univ., 2023) 4–8.
Eed, J. Factors affecting enzyme activity. Essai 10, 19 (2012). https://dc.cod.edu/essai/vol10/iss1/19
Dutta, T. K., Jana, M., Pahari, P. R. & Bhattacharya, T. The effect of temperature, pH, and salt on amylase in Heliodiaptomus viduus (Gurney) (Crustacea: Copepoda: Calanoida). Turk. J. Zool. 30, 187–195 (2006).
Kim, S. H., Oh, S. Y., Jeong, Y. J. & Kwon, J. H. Changes in characteristics of the ancient grain farro according to liquefaction and saccharification processes. Food Sci. Preserv. 32, 17–29 (2025).
Alenyorege, E. A. et al. Response surface methodology centred optimization of mono-frequency ultrasound reduction of bacteria in fresh-cut Chinese cabbage and its effect on quality. LWT 122, 108991 (2020).
Cai, B. Y., Ge, J. P., Ling, H. Z., Cheng, K. K. & Ping, W. X. Statistical optimization of dilute sulfuric acid pretreatment of corncob for xylose recovery and ethanol production. Biomass Bioenergy 36, 250–257 (2012).
Prasetiyo, H. et al. Optimizing the coating for masking conditions process with gum Arabic using Box-Behnken design (BBD) on the properties of masked Spirulina Powder. BIO Web Conf. 176, 02010 (2025).
Körbahti, B. K. & Rauf, M. A. Response surface methodology (RSM) analysis of photoinduced decoloration of toludine blue. Chem. Eng. J. 136, 25–30 (2008).
An, J. O., Chung, C. H. & Lee, S. J. Effect of rice pre-treatment on enzymatic saccharification in the brewing process. Microbiol. Biotechnol. Lett. 45, 277–283 (2017).
Kim, H. S. & Kang, Y. J. Optimal conditions of saccharification for a traditional malt syrup in Cheju. Korean J. Food Sci. Technol. 26, 659–664 (1994).
Maicas, S. The role of yeasts in fermentation processes. Microorganisms 8, 1142 (2020).
Park, J. H., Yeo, S. H., Choi, J. H., Jeong, S. T. & Choi, H. S. Production of makgeolli using rice treated with Gaeryang-Nuruk (for non-steaming process) extract. Food Sci. Preserv. 19, 144–152 (2012).
Serrano-Febles, J. et al. Optimization of enzymatic hydrolysis of corn starch to obtain glucose syrups by genetic algorithm [Optimización de la hidrólisis enzimática del almidón de maíz para obtener siropes de glucosa mediante algoritmos genéticos]. DYNA 92, 83–91 (2025).
Lee, J. S. et al. Exploration of optimum conditions for production of saccharogenic mixed grain beverages and assessment of anti-diabetic activity. J. Nutr. Health. 47, 12–22 (2014).
Nabatian, E., Dolatabadi, M. & Ahmadzadeh, S. Application of experimental design methodology to optimize acetaminophen removal from aqueous environment by magnetic chitosan@multi-walled carbon nanotube composite: Isotherm, kinetic, and regeneration studies. Anal. Method. Environ. Chem. J. 5, 61–74 (2022).
Bae, S. M. et al. Preparation and characterization of sweet persimmon wine. Appl. Biol. Chem. 45, 66–70 (2002).
Kim, J. Y. & Yi, Y. H. pH, acidity, color, amino acids, reducing sugars, total sugars, and alcohol in puffed millet powder containing millet takju during fermentation. Korean J. Food Sci. Technol. 42, 727 (2010).
Kim, M. J., Han, S. Y., Kang, D., Shin, J. H. & Lee, S. J. Functional insights into microbial communities of Korean traditional rice wine (makgeolli) during fermentation. LWT 210, 116826 (2024).
Lee, S., Yoo, M. & Shin, D. The identification and quantification of biogenic amines in Korean turbid rice wine, makgeolli by HPLC with mass spectrometry detection. LWT 62, 350–356 (2015).
Wong, B. et al. Characterisation of Korean rice wine (makgeolli) prepared by different processing methods. Curr. Res. Food Sci. 6, 100420 (2023).
Acknowledgements
This research was supported by the Korea Food Research Institute, funded by the Ministry of Science and ICT, Republic of Korea (Grant No. E0211400-05).
Author information
Authors and Affiliations
Contributions
SWJ: Writing-original draft, Methodology, Visualization, Formal analysis, Data curation, HHY: Methodology, Conceptualization, Writing-review and editing, Investigation, Data curation, JCK: Writing-review and editing, Project administration, MJ: Writing-review and editing, Supervision, Conceptualization.
Corresponding author
Ethics declarations
Competing interests
The authors declare no competing interests.
Additional information
Publisher’s note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Rights and permissions
Open Access This article is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License, which permits any non-commercial use, sharing, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if you modified the licensed material. You do not have permission under this licence to share adapted material derived from this article or parts of it. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by-nc-nd/4.0/.
About this article
Cite this article
Jang, SW., Yu, H.H., Kim, JC. et al. Optimizing brown rice liquefaction and saccharification using response surface methodology for grain ethanol production. Sci Rep (2026). https://doi.org/10.1038/s41598-026-40430-9
Received:
Accepted:
Published:
DOI: https://doi.org/10.1038/s41598-026-40430-9
