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A NaBH4 Coupled Ninhydrin-Based Assay for the Quantification of Protein/Enzymes During the Enzymatic Hydrolysis of Pretreated Lignocellulosic Biomass

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Abstract

Accurate protein quantification is necessary in many of the steps during the enzymatic hydrolysis of pretreated lignocellulosic biomass, from the fundamental determination of enzyme kinetics to techno-economic assessments, such as the use of enzyme recycling strategies, evaluation of enzyme costs, and the optimization of various process steps. In the work described here, a modified, more rapid ninhydrin-based protein quantification assay was developed to better quantify enzyme levels in the presence of lignocellulosic biomass derived compounds. The addition of sodium borohydride followed by acid hydrolysis at 130 °C greatly reduced interference from monosaccharides and oligosaccharides and decreased the assay time 6-fold. The modified ninhydrin assay was shown to be more accurate as compared to various traditional colorimetric protein assays when commercial cellulase enzyme mixtures were quantified under typical pretreated lignocellulosic biomass enzymatic hydrolysis conditions. The relatively short assay time and microplate-reading capability of the modified assay indicated that the method could likely be used for high-throughput protein determination.

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Acknowledgments

We want to thank our colleague, Dr. Jinguang Hu, for all of his invaluable help. The Natural Sciences and Engineering Research Council of Canada (NSERC) and the Bioconversion network financially supported this work. We also thank our colleagues at Novozymes for their generous donation of enzyme preparations.

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Authors and Affiliations

  1. Forest Products Biotechnology/Bioenergy Group, Faculty of Forestry, University of British Columbia, 2424 Main Mall, Vancouver, BC, V6T 1Z4, Canada

    Yiu Ki Mok & Jack N. Saddler

  2. Department of Biotechnology, Lorena School of Engineering, University of São Paulo, CP 116, Rod. Itajubá—Lorena, km 74,5 Área I, Estrada Municipal do Campinho S/N 2602-810, Lorena, SP, Brazil

    Valdeir Arantes

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  1. Yiu Ki Mok
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Correspondence to Jack N. Saddler.

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Mok, Y.K., Arantes, V. & Saddler, J.N. A NaBH4 Coupled Ninhydrin-Based Assay for the Quantification of Protein/Enzymes During the Enzymatic Hydrolysis of Pretreated Lignocellulosic Biomass. Appl Biochem Biotechnol 176, 1564–1580 (2015). https://doi.org/10.1007/s12010-015-1662-7

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  • DOI: https://doi.org/10.1007/s12010-015-1662-7

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