Over the past 20 years, our group has looked at steam pretreatment (SP) with regard to its suitability for pretreating a range of lignocellulosic sub­strates, the subsequent ease of enzymatic hydrolysis of the cellulosic stream and the recovery of most of the hemicellulose sugars and lignin in a use­able form. SP is an attractive pretreatment process as it makes limited use of chemicals, requires relatively low levels of energy and, depending on the conditions employed, results in the recovery of most of the original cellulose — and hemicellulose-derived carbohydrates in a fermentable form [23-28]. As will be discussed in more detail, SP disrupts the lignin barrier [29] and facili­tates access of cellulases to the cellulose fibers [30]. Previous work has shown that SO2-catalyzed SP is an effective pretreatment for softwood [26-34], hard­wood [35-37] and agricultural residues [22] and that impregnation of SO2 prior to pretreatment results in lower treatment temperatures and shorter reaction times, thereby improving hemicellulose recovery and reducing the
formation of sugar degradation products [23]. It has also been shown that SO2 impregnation prior to SP enhanced the carbohydrate hydrolysis rate by increasing the accessibility of cell walls via the formation of fractures and the removal of hemicellulose during the steaming of the substrate [28], while re­ducing the DP of the oligomers and increasing the proportion of monomers

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in the water-soluble stream [31,38-40]. The “severity” of SP can be summa­rized by a single factor called Ro (Ro = t exp(T — 100)/14.75) which links the effects of time (t, min) and temperature (T, °C) [41]. Due to the high tem­perature and acidic conditions employed during the SP process, depending on the “severity” (temperature, time, pressure, catalyst dosage) of the treat­ment, a portion of the hemicellulose-derived sugars and solubilized lignin fragments can be degraded or transformed into compounds such as furfural and 5-hydroxymethylfurfural (5-HMF); aliphatic acids, such as acetic, formic, and levulinic acid; and phenolic compounds [42]. It is known that these compounds can inhibit both downstream hydrolysis by cellulases [43] and fermentation of the liberated sugars to ethanol [44]. Therefore, compromised SP conditions have to be defined that provide an easily hydrolyzable cellu — losic substrate, good recovery of the hemicellulose-derived sugars, ideally in a monomeric form, while minimizing the formation of inhibitors. Ideally, a reactive lignin stream, with a higher economic application than its intrinsic fuel value should also be obtained.

It is apparent that the nature of the substrate and the pretreatment method used has at least as much influence on the ease of enzymatic hydrolysis as does the nature and efficiency of the enzyme system used to conduct hydrolysis. As illustrated in Fig. 1, the efficiency of enzymatic hydrolysis of a given lignocellu — losic substrate is the result of interplay of various factors. Although it is evident that substrates such as agricultural residues are generally less recalcitrant than softwood residues, it is recognized that enzyme — or substrate-related factors that govern effective hydrolysis can be controlled to an appreciable extent by the type and conditions of the pretreatment employed.

In the sections below we will describe how pretreatment, specifically SP, in­fluences the characteristics of the substrate and the subsequent recovery of the cellulose, hemicellulose and lignin components. Recent progress in eluci­dating the role of substrate properties such as crystallinity, DP, pore volume, and available surface area in enzymatic hydrolysis will also be discussed using wood pulps as “model substrates”. The final section offers concluding re­marks and outlines the remaining challenges associated with understanding the progress of enzymatic hydrolysis during the bioconversion process.

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