Low melting point pyridinium ionic liquid pretreatment for enhancing enzymatic saccharification of cellulosic biomass Uju a , Aya Nakamoto a , Yasuhiro Shoda a , Masahiro Goto a,b , Wataru Tokuhara c , Yoshiyuki Noritake c , Satoshi Katahira d , Nobuhiro Ishida d , Chiaki Ogino e , Noriho Kamiya a,b,⇑ a Department of Applied Chemistry, Graduate School of Engineering, Kyushu University, 744 Motooka, Fukuoka 819-0395, Japan b Center for Future Chemistry, Kyushu University, Japan c Bio Fuel Group, Organic Material Engineering Division, Toyota Motor Corporation, 1 Toyota-cho, Toyota, Aichi 471-8572, Japan d Biotechnology Laboratory, Toyota Central R&D Labs Inc., 41-1 Yokomichi, Nagakute, Aichi 480-1192, Japan e Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University, Kobe, Japan article info Article history: Available online 5 July 2012 Keywords: Biomass Cellulose Enzymatic saccharification Ionic liquid Pretreatment abstract The potential of 1-hexylpyridinium chloride ([Hpy][Cl]), to pretreat cellulosic feedstocks was investigated using microcrystalline cellulose (Avicel) and Bagasse at 80 °C or 100 °C. Short [Hpy][Cl] pretreatments, <30 min, at lower temperature accelerate subsequent enzymatic saccharification of Avicel. Over 95% conversion of pretreated Avicel to glucose was attained after 24 h enzymatic saccharification under opti- mal conditions, whereas regenerated Bagasse showed 1–3-fold higher conversion than untreated bio- mass. FT-IR analysis of both Avicel and Bagasse samples pretreated with [Hpy][Cl] or 1-ethyl-3- methyimidazolium acetate ([Emim][OAc]) revealed that these ionic liquids behaved differently during pretreatment. [Hpy][Cl] pretreatment for an extended duration (180 min) released mono- and disaccha- rides without using cellulase enzymes, suggesting [Hpy][Cl] has capability for direct saccharification of cellulosic feedstocks. On the basis of the results obtained, [Hpy][Cl] pretreatment enhanced initial reac- tion rates in enzymatic saccharification by either crystalline polymorphic alteration of cellulose or partial degradation of the crystalline cellulosic fraction in biomass. Ó 2012 Elsevier Ltd. All rights reserved. 1. Introduction A major challenge faced when using lignocellulosic biomass as a source for biofuel production is difficulty hydrolyzing cellulose because lignocellulose is covered by a robust and complex structure of lignin and hemicelluloses, which hampers the function of cellu- lase enzymes (Chang and Holtzapple, 2000; Yoshida et al., 2008). Furthermore, native cellulose has high crystallinity arising from inter- and intramolecular hydrogen bonding, making this material insoluble in a range of solvents, and it is thus difficult to set up a hydrolyzation process (Puri, 1984). Therefore, pretreatment pro- cesses before the saccharification step are essential. The aim of biomass pretreatment is to increase the accessibility of cellulase en- zymes to cellulose by removing lignin and hemicellulose as well as to reduce cellulose crystallinity (Chang and Holtzapple, 2000; Yoshida et al., 2008). Biological, physical, chemical, and physico- chemical pretreatment methods have been proposed for this purpose. Problems with pretreatment processes include slow reaction rates, high-energy requirements and significant costs associated with effluent disposal or recovery (Balat, 2011; Klein- Marcuschamer et al., 2011). Therefore, it is important to identify an effective pretreatment method. Recently, the use of ionic liquids (ILs) has provided significant scope as a cellulosic biomass pretreatment process because ILs are non-volatile, have low melting points and high thermal stabil- ity (Ohno and Fukaya, 2009) and can also be recycled and reused with little loss of effectiveness (Lee et al., 2009; Shill et al., 2011; Wu et al., 2011). ILs based on the alkylimidazolium cation with various anions have been developed for dissolving cellulose and removing lignin and hemicellulose from soft wood, hardwood and herbaceous matter (Mäki-Arvela et al., 2010). Comparison studies of IL pretreatments on lignocellulosic biomass have been reported. Li et al. (2010) reported that an IL pretreatment showed better performance on enhancing enzymatic saccharification than a dilute acid pretreatment. Although the cost of using ILs as pre- treatment agents must be addressed (Klein-Marcuschamer et al., 2011), IL pretreatments could reduce energy consumption com- pared to alkali and acid pretreatments (Yoon et al., 2011). The capability of ILs for dissolving cellulose possibly arises from disruption of the hydrogen bond network in cellulose by the 0960-8524/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.biortech.2012.06.096 ⇑ Corresponding author at: Department of Applied Chemistry, Graduate School of Engineering, Kyushu University, 744 Motooka, Fukuoka 819-0395, Japan. Tel.: +81 92 802 2807; fax: +81 92 802 2810. E-mail address: nori_kamiya@mail.cstm.kyushu-u.ac.jp (N. Kamiya). Bioresource Technology 135 (2013) 103–108 Contents lists available at SciVerse ScienceDirect Bioresource Technology journal homepage: www.elsevier.com/locate/biortech