Fermentation of xylose by the thermotolerant yeast strains Kluyveromyces marxianus IMB2, IMB4, and IMB5 under anaerobic conditions Mark R. Wilkins a, * , Michael Mueller a , Sabine Eichling b , Ibrahim M. Banat c a Department of Biosystems and Agricultural Engineering, Oklahoma State University, 224 Agricultural Hall, Stillwater, OK 74078, USA b University of Applied Sciences at Bingen, 109 Berlinstrasse, Bingen 55411, Germany c School of Biomedical Sciences, University of Ulster, Coleraine, Northern Ireland BT52 1SA, UK Received 12 September 2007; received in revised form 18 October 2007; accepted 19 December 2007 Abstract The effects of temperature and initial pH on anaerobic xylose utilization by the thermotolerant yeast Kluyveromyces marxianus IMB2, IMB4, and IMB5 were evaluated. Ethanol yield from xylose was greatest with IMB4, a temperature of 40 8C and an initial pH of 5.5. Xylitol yield was greatest with yeast strains IMB2 and IMB5, a temperature of 45 8C and an initial pH of 4.5 or 5.0. Additional fermentations of xylose at 40 and 45 8C, pH 5.5, and 20 times more cells were done using IMB4 since it exhibited the greatest ethanol yield. Ethanol and xylitol yields from xylose were greater at 40 8C than at 45 8C and volumetric production rates for ethanol and xylitol were 0.02 and 0.08 g/(l h), respectively. IMB4 xylitol yields were comparable to other xylitol producers, but with lower productivity. These strains are not good candidates for utilization of xylose under anaerobic conditions and other strategies should be explored. # 2007 Elsevier Ltd. All rights reserved. Keywords: Xylose; Fermentation; Thermotolerant; Yeast; Biofuels; Bioproducts 1. Introduction Ethanol for transportation fuel is primarily produced from fermentation of sugar crops and enzymatically hydrolyzed grain starch; however, to meet future demand, efforts to produce ethanol from cellulosic biomass are being pursued. Cellulosic biomass refers to grasses, wood, crop residues, and waste materials that contain cellulose, a polymer of beta- glucose. Cellulosic biomass usually contains polysaccharides other than cellulose, particularly various hemicelluloses that contain both hexoses and pentoses. The primary sugars found in cellulosic biomass are D-glucose and D-xylose, though other sugars such as L-arabinose, mannose, galactose, and rhamnose are also present [1]. Wild-type strains of Saccharomyces cerevisiae, the predominant organism used for commercial ethanol production, are unable to utilize xylose, though several researchers have genetically modified S. cerevisiae strains to produce ethanol from xylose [2,3]. There are also native yeast species that ferment xylose to produce ethanol, including several Pichia and Candida species as well as some strains of Kluyveromyces marxianus [4,5]. Five strains of K. marxianus were previously isolated in an Indian distillery and characterized for growth on a number of substrates, including xylose, at 45 8C [6,7]. These strains, known as the IMB strains, showed the ability to consume xylose and produce ethanol in an oxygen-limited environment in shake flasks, with a maximum yield of 23% theoretical by K. marxianus IMB4 at 45 8C and an initial pH of 5.5 [7]. The K. marxianus IMB strains utilize a wider range of substrates and exhibit greater thermotolerance than does S. cerevisiae. One of the major applications previously proposed for these strains is simultaneous saccharification and fermentation (SSF) of cellulosic biomass [8–10]. The temperature optimum of cellulases used in SSF is 45–50 8C, while most yeast cannot survive at these temperatures [9]. The use of the K. marxianus IMB strains would allow SSF temperatures to be increased, resulting in faster cellulose hydrolysis. The overall goal of this study was to determine the yields of ethanol and xylitol K. marxianus IMB2, IMB4, and IMB5 can produce under anaerobic conditions and temperatures similar to www.elsevier.com/locate/procbio Process Biochemistry 43 (2008) 346–350 * Corresponding author. Tel.: +1 405 744 8416; fax: +1 405 744 6059. E-mail address: mark.wilkins@okstate.edu (M.R. Wilkins). 1359-5113/$ – see front matter # 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.procbio.2007.12.011