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Kinetics of Enzymatic High-Solid Hydrolysis of Lignocellulosic Biomass Studied by Calorimetry

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Abstract

Enzymatic hydrolysis of high-solid biomass (>10% w/w dry mass) has become increasingly important as a key step in the production of second-generation bioethanol. To this end, development of quantitative real-time assays is desirable both for empirical optimization and for detailed kinetic analysis. In the current work, we have investigated the application of isothermal calorimetry to study the kinetics of enzymatic hydrolysis of two substrates (pretreated corn stover and Avicel) at high-solid contents (up to 29% w/w). It was found that the calorimetric heat flow provided a true measure of the hydrolysis rate with a detection limit of about 500 pmol glucose s−1. Hence, calorimetry is shown to be a highly sensitive real-time method, applicable for high solids, and independent on the complexity of the substrate. Dose–response experiments with a typical cellulase cocktail enabled a multidimensional analysis of the interrelationships of enzyme load and the rate, time, and extent of the reaction. The results suggest that the hydrolysis rate of pretreated corn stover is limited initially by available attack points on the substrate surface (<10% conversion) but becomes proportional to enzyme dosage (excess of attack points) at later stages (>10% conversion). This kinetic profile is interpreted as an increase in polymer end concentration (substrate for CBH) as the hydrolysis progresses, probably due to EG activity in the enzyme cocktail. Finally, irreversible enzyme inactivation did not appear to be the source of reduced hydrolysis rate over time.

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Acknowledgements

This work was supported by the Danish Agency for Science, Technology and Innovation (grant # 2104-07-0028 to PW) and the Carlsberg Foundation. A special thanks to Leigh Murphy for productive comments on the manuscript and help with control experiments.

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Authors and Affiliations

  1. Roskilde University, NSM, Biomaterials, Universitetsvej 1, DK-4000, Roskilde, Denmark

    Søren N. Olsen, Erik Lumby & Peter Westh

  2. Novozymes Inc., 1445 Drew Avenue, Davis, CA, 95-618, USA

    Kc McFarland

  3. Novozymes A/S, Krogshøjvej 36, DK-2880, Bagsværd, Copenhagen, Denmark

    Kim Borch

Authors
  1. Søren N. Olsen
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  2. Erik Lumby
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  3. Kc McFarland
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  4. Kim Borch
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  5. Peter Westh
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Correspondence to Peter Westh.

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Olsen, S.N., Lumby, E., McFarland, K. et al. Kinetics of Enzymatic High-Solid Hydrolysis of Lignocellulosic Biomass Studied by Calorimetry. Appl Biochem Biotechnol 163, 626–635 (2011). https://doi.org/10.1007/s12010-010-9068-z

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  • DOI: https://doi.org/10.1007/s12010-010-9068-z

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