Browsing by Author "Amraoui, Yassin"
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Item Open Access Enhanced 2,3-Butanediol production by mutant Enterobacter ludwigii using Brewers’ spent grain hydrolysate: process optimization for a pragmatic biorefinery loom(Elsevier, 2021-06-18) Amraoui, Yassin; Prabhu, Ashish A.; Narisetty, Vivek; Coulon, Frederic; Chandel, Anuj Kumar; Willoughby, Nicholas; Jacob, Samuel; Koutinas, Apostolis; Kumar, Vinod2,3-Butanediol (BDO) is a fossil-based versatile bulk chemical with a multitude of applications. BDO can also be synthesized using microbial cell factories harnessing renewable feedstocks. However, the high cost of the substrate via microbial route impedes commercial manufacturing of BDO. Therefore, identification of cheaper substrates could make bio-based BDO production more cost-competitive. Brewers’ spent grain (BSG), a major by-product of breweries, is an inexpensive source of fermentable sugars and proteins. In the present study, we have attempted the bioproduction of BDO by Enterobacter ludwigii using BSG as feedstock. A random E. ludwigii mutant obtained after treatment with ethyl methane sulfonate (EMS) resulted in a BDO titer (9.5 g/L), ~30% higher in comparison to the wild type strain with a yield of 0.48 gBDO/gGlucose approaching the theoretical yield of 0.50 gBDO/gGlucose. The enzymatic hydrolysis of microwave-assisted alkali pretreated BSG was optimized using the statistical Taguchi design. The BSG hydrolysis under optimal conditions (pH: 6.0; temperature: 50 °C; BSG: 10% w/v; enzyme loading: 2% v/v) resulted in a glucose yield of 0.25 gGlucose/gBiomass. The uncontrolled pH was found to be more beneficial for BDO accumulation from BSG hydrolysate in batch bioreactor cultivation as compared with controlled one. The fed-batch cultivation with forced pH fluctuations at an aeration rate of 2.0 vvm resulted in BDO accumulation of 118.5 g/L from glucose-rich BSG hydrolysate with the yield and productivity of 0.43 g/g and 1.65 g/L.h, respectively. To the best of our knowledge, this is the first study on BDO production from BSG.Item Open Access Enhanced xylitol production using non-detoxified xylose rich pre-hydrolysate from sugarcane bagasse by newly isolated Pichia fermentans(BioMed Central / Springer Verlag, 2020-12-29) Prabhu, Ashish A.; Bosakornranut, Ekkarin; Amraoui, Yassin; Agrawal, Deepti; Coulon, Frederic; Vivekanand, Vivekanand; Thakur, Vijay Kumar; Kumar, VinodBackground Integrated management of hemicellulosic fraction and its economical transformation to value-added products is the key driver towards sustainable lignocellulosic biorefineries. In this aspect, microbial cell factories are harnessed for the sustainable production of commercially viable biochemicals by valorising C5 and C6 sugars generated from agro-industrial waste. However, in the terrestrial ecosystem, microbial systems can efficiently consume glucose. On the contrary, pentose sugars are less preferred carbon source as most of the microbes lack metabolic pathway for their utilization. The effective utilization of both pentose and hexose sugars is key for economical biorefinery. Results Bioprospecting the food waste and selective enrichment on xylose-rich medium led to screening and isolation of yeast which was phylogenetically identified as Pichia fermentans. The newly isolated xylose assimilating yeast was explored for xylitol production. The wild type strain robustly grew on xylose and produced xylitol with > 40% conversion yield. Chemical mutagenesis of isolated yeast with ethyl methanesulphonate (EMS) yielded seven mutants. The mutant obtained after 15 min EMS exposure, exhibited best xylose bioconversion efficiency. This mutant under shake flask conditions produced maximum xylitol titer and yield of 34.0 g/L and 0.68 g/g, respectively. However, under the same conditions, the control wild type strain accumulated 27.0 g/L xylitol with a conversion yield of 0.45 g/g. Improved performance of the mutant was attributed to 34.6% activity enhancement in xylose reductase with simultaneous reduction of xylitol dehydrogenase activity by 22.9%. Later, the culture medium was optimized using statistical design and validated at shake flask and bioreactor level. Bioreactor studies affirmed the competence of the mutant for xylitol accumulation. The xylitol titer and yield obtained with pure xylose were 98.9 g/L and 0.67 g/g, respectively. In comparison, xylitol produced using non-detoxified xylose rich pre-hydrolysate from sugarcane bagasse was 79.0 g/L with an overall yield of 0.54 g/g. Conclusion This study demonstrates the potential of newly isolated P. fermentans in successfully valorising the hemicellulosic fraction for the sustainable xylitol productionItem Open Access High yield recovery of 2,3-butanediol from fermented broth accumulated on xylose rich sugarcane bagasse hydrolysate using aqueous two-phase extraction system(Elsevier, 2021-06-26) Narisetty, Vivek; Amraoui, Yassin; Abdullah, Alamri; Ahmad, Ejaz; Agrawal, Deepti; Parameswaran, Binod; Pandey, Ashok; Goel, Saurav; Kumar, VinodDownstream processing of chemicals obtained from fermentative route is challenging and cost-determining factor of any bioprocess. 2,3-Butanediol (BDO) is a promising chemical building block with myriad applications in the polymer, food, pharmaceuticals, and fuel sector. The current study focuses on the recovery and purification of BDO produced (68.2 g/L) from detoxified xylose-rich sugarcane bagasse hydrolysate by a mutant strain of Enterobacter ludwigii. Studies involving screening and optimization of aqueous-two phase system (ATPS) revealed that 30% w/v (NH4)2SO4 addition to clarified fermentation broth facilitated BDO extraction in isopropanol (0.5 v/v), with maximum recovery and partition coefficient being 97.9 ± 4.6% and 45.5 ± 3.5, respectively. The optimized protocol was repeated with unfiltered broth containing 68.2 g/L BDO, cell biomass, and unspent protein, which led to the partitioning of 66.7 g/l BDO, 2.0 g/L xylose and 9.0 g/L acetic acid into organic phase with similar BDO recovery (97%) and partition coefficient (45).Item Open Access Integrated fermentative production and downstream processing of 2,3-butanediol from sugarcane bagasse-derived xylose by mutant strain of Enterobacter ludwigii(American Chemical Society, 2021-07-16) Amraoui, Yassin; Narisetty, Vivek; Coulon, Frederic; Agrawal, Deepti; Chandel, Anuj Kumar; Maina, Sofia; Koutinas, Apostolis; Kumar, VinodIn this study, a mutant strain of Enterobacter ludwigii developed in our previous work, was evaluated to utilize pure xylose as the sole carbon and energy source for 2,3-butanediol (BDO) production. Later, this strain was also investigated on detoxified and nondetoxified xylose-rich hydrolysate obtained from hydrothermally pretreated sugarcane bagasse (SCB) for BDO production. Supplementing the fermentation medium with 0.2% w/v yeast extract improved cell growth (31%), BDO titer (43%), and yield (41%) against the synthetic medium devoid of any complex nitrogen source. The fed-batch culture with cyclic control of pH resulted in a BDO production of 71.1 g/L from pure xylose with overall yield and productivity of 0.40 g/g and 0.94 g/L·h, respectively. While BDO titer, yield, and productivity of 63.5 g/L, 0.36 g/g, and 0.84 g/L·h, were acheived with detoxified hydrolysate, respectively. In contrast, 32.7 g/L BDO was produced from nondetoxified hydrolysate with a conversion yield of 0.33 g/g and a productivity of 0.43 g/L·h. BDO accumulated on pure xylose and detoxified SCB hydrolysate was separated by aqueous two-phase system (ATPS) method using (NH4)2SO4 as salting-out agent and isopropanol as an extractant, resulting in the BDO recovery of more than 85%. The results achieved in the current work exemplify a step toward industrial BDO production from cost-effective hemicellulosic hydrolysates by E. ludwigii.