The effect of shaking regime on the rate and extent of enzymatic hydrolysis of cellulose

Abstract

In an attempt to elucidate the effect of mixing on the rate and extent of enzymatic hydrolysis of cellulosic substrates, α-cellulose was hydrolysed using a commercial cellulase preparation at varying levels of substrate concentration (2.5, 5 and 7.5% (w/v)) and by using three shaking regimes: continuous at low-speed (25 rpm), continuous at high-speed (150 rpm) and an intermittent regime comprised of high and low-speed shaking intervals. The continuous, high-speed shaking produced the highest conversion yields, whereas the intermittent and low-speed shaking regimes resulted in lower conversions. After 72 h, at all shaking regimes (150 rpm, 25 rpm and intermittent), using a low substrate concentration (2.5%) produced conversion yields (82, 79 and 80%) higher than those obtained at high (7.5%) substrate concentration (68, 63 and 68%). As the substrate concentration increased, the conversion yields at intermittent shaking gradually approached those resulting from high-speed shaking. Thus, it appears that intermittent shaking could be a beneficial process option as it can reduce the mixing energy requirements while producing reasonably high conversion yields.

Introduction

Production of fuel ethanol from lignocellulosic feedstocks, such as hardwoods, softwoods and herbaceous materials, may involve an enzymatic hydrolysis sub-process during which the feedstocks' cellulose fraction is converted to glucose monomers, which can subsequently be fermented to ethanol by ethanologenic micro-organisms (Zacchi and Galbe, 1998). While the acid hydrolysis reaction is faster and more effective, enzymatic saccharification remains a favoured unit operation for the bioconversion process as it requires milder conditions and does not produce sugar degradation products that are inhibitory to fermentative micro-organisms (Hsu, 1996).

The rate and extent of cellulose hydrolysis by cellulase enzymes is influenced by many substrate and enzyme related factors (Esteghlalian et al., 1999) including the heterogeneity of the reactants, a liquid enzyme acting upon a solid substrate. Therefore, adequate mixing is required to ensure sufficient contact between the substrate and enzymes and to promote heat and mass transfer within the reaction vessel. It has been shown, however, that excessive mixing can deactivate the enzymes and reduce the conversion yield. This effect has been attributed to the shear force generated by the mixer and the entrapment of air bubbles into the medium at the air–liquid surface (Reese and Ryu, 1980, Mukataka et al., 1983). Mukataka et al. (1983) have shown that excessively high mixing speeds (>200 rpm) could lower the extent of cellulose conversion (Avicel and paper pulp) while moderate mixing speeds (100–200 rpm) provide a good combination of fast initial hydrolysis rates and high conversion yields. Studies conducted by Tengborg showed that while mixing speeds (paddle mixer) as high as 340 rpm enhanced the conversion of steam-pretreated spruce wood, higher mixing speeds (340–510 rpm) only increased the initial rate of hydrolysis and not the final conversion yield (Tengborg, 2000). Others also found that increasing the agitation speed (magnetic stirring) from 380 to 1500 rpm increased the conversion of soybean hulls (Enayati and Parulekar, 1995).

Intermittent mixing regimes that combine intervals of no-mixing with short periods of high or low speed mixing can benefit the enzymatic hydrolysis process by reducing energy consumption and limiting enzyme inactivation due to lower shearing of the reaction mixture, while providing reasonably high conversion yields (Nguyen, 1998). In this study we evaluated the effectiveness of an intermittent shaking regime, in comparison with continuous shaking at high or low speed, for saccharification of cellulose under various substrate concentrations.