Augmentation of ethanol production through statistically designed growth and fermentation medium using novel thermotolerant yeast isolates Richa Arora a, b, 2 , Shuvashish Behera a, 2 , Nilesh Kumar Sharma a, b , Sachin Kumar a, *, 1 a Biochemical Conversion Division, Sardar Swaran Singh National Institute of Bio-Energy, Kapurthala,144601, India b I.K Gujral Punjab Technical University, Kapurthala,144601, India article info Article history: Received 3 June 2016 Received in revised form 14 March 2017 Accepted 18 March 2017 Available online 21 March 2017 Keywords: Thermotolerant yeast Kluyveromyces marxianus Ethanologen Optimization Face-centered central composite design abstract Overproduction of metabolites, high product yield and process economics are greatly influenced by the media composition used for growth and fermentation. The main purpose of this study is to enhance the ethanol production through statistical tool of response surface methodology (RSM) by optimizing media components for the growth and fermentation of thermotolerant isolates Kluyveromyces marxianus NIRE- K1 and NIRE-K3. Five different salts were used in the Face-centered Central Composite Design (FCCD), with the responses of biomass formation and ethanol production for growth and fermentation, respectively. Yeast extract and K 2 HPO 4 were found to be the key media components for the growth and fermentation which is revealed from their interaction in both the yeast isolates. Further studies on batch fermentation kinetics using the optimized values of the medium composition for K. marxianus NIRE-K1 and NIRE-K3 resulted in final ethanol concentration of 17.73 (86.27% of theoretical ethanol yield) and 19.01 g l 1 (94.12% of theoretical ethanol yield), respectively. An increase in the ethanol yield and pro- ductivity by 11.36,10.42% and 2.0, 2.7% was revealed in NIRE-K1 and NIRE-K3, respectively, as compared to our previous study. © 2017 Elsevier Ltd. All rights reserved. 1. Introduction The drastic increase in the energy crisis, green house gas emis- sions and exhaustion of fossil fuel reserves have led to the develop- ment of renewable energy technologies [1e3]. However, one of the major challenges in the fuel production from renewable resources lies in the development of improved strains with efficient ethanol production [4,5]. Current research primarily focuses on the utiliza- tion of thermotolerant yeasts for efficient bioconversion of biomass to ethanol [6e9]. Thermotolerant ethanologenic fermentations are reported to be superior rather than the conventional mesophilic ones with higher bioconversion rates, continuous product recovery, economically viable processes due to lesser requirement of cooling and reduced risk of contamination [4,10e12]. Apart from the use of thermotolerant ethanologens, the commercialization of a bioprocess and its economics depends upon the cost for the cultivation of the culture and its subsequent ethanol yield. The biochemical and nutritional requirements of the bio- processing strains is highly influenced by carbon, nitrogen sources along with supplements like amino acids, vitamins, antibiotics, etc., which further aids in the cost [13]. Also, mineral salts are generally used in ethanol producing industries to supplement the fermen- tation media and provide acceptable yields [14]. Moreover, various medium components have strong interactions which may affect the competence of the process, both positively and negatively [15]. Thus, there is a need to develop a medium formulation for conve- nient, cost-effective and efficient bioprocess technology for bio- ethanol production. There are two methods for evaluating the optimal level, empirical method and statistical method. The former has several limitations because it involves substantial amount of time and la- bour taking OFAT (one-factor-at-a-time) approach into account. Moreover, it does not account for the interaction among the vari- ables which strongly influences the bioprocess [16]. On the other hand, the latter involves the statistical tools like response surface * Corresponding author. Biochemical Conversion Division, Sardar Swaran Singh National Institute of Bio-Energy, Jalandhar-Kapurthala Road, Wadala Kalan, Kapurthala, 144601, Punjab, India. E-mail address: sachin.biotech@gmail.com (S. Kumar). 1 Present Address: Department of Chemical & Biological Engineering, South Dakota School of Mines and Technology, Rapid City, SD, 57701, USA. 2 Equal contribution. Contents lists available at ScienceDirect Renewable Energy journal homepage: www.elsevier.com/locate/renene http://dx.doi.org/10.1016/j.renene.2017.03.059 0960-1481/© 2017 Elsevier Ltd. All rights reserved. Renewable Energy 109 (2017) 406e421