Scale-up of microbial lipid and bioethanol production from oilcane.

Microbial oils are a sustainable biomass-derived substitute for liquid fuels and vegetable oils. Oilcane, an engineered sugarcane with superior feedstock characteristics for biodiesel production, is a promising candidate for bioconversion. This study describes the processing of oilcane stems into juice and hydrothermally pretreated lignocellulosic hydrolysate and their valorization to ethanol and microbial oil using Saccharomyces cerevisiae and engineered Rhodosporidium toruloides strains, respectively. A bioethanol titer of 106 g/L was obtained from S. cerevisiae grown on oilcane juice in a 3 L fermenter, and a lipid titer of 8.8 g/L was obtained from R. toruloides grown on oilcane hydrolysate in a 75 L fermenter. Oil was extracted from the R. toruloides cells using supercritical CO2, and the observed fatty acid profile was consistent with previous studies on this strain. These results demonstrate the feasibility of pilot-scale lipid production from oilcane hydrolysate as part of an integrated bioconversion strategy.

Tolerance of engineered Rhodosporidium toruloides to sorghum hydrolysates during batch and fed-batch lipid production.

BACKGROUND: Oleaginous yeasts are a promising candidate for the sustainable conversion of lignocellulosic feedstocks into fuels and chemicals, but their growth on these substrates can be inhibited as a result of upstream pretreatment and enzymatic hydrolysis conditions. Previous studies indicate a high citrate buffer concentration during hydrolysis inhibits downstream cell growth and ethanol fermentation in Saccharomyces cerevisiae. In this study, an engineered Rhodosporidium toruloides strain with enhanced lipid accumulation was grown on sorghum hydrolysate with high and low citrate buffer concentrations. RESULTS: Both hydrolysis conditions resulted in similar sugar recovery rates and concentrations. No significant differences in cell growth, sugar utilization rates, or lipid production rates were observed between the two citrate buffer conditions during batch fermentation of R. toruloides. Under fed-batch growth on low-citrate hydrolysate a lipid titer of 16.7 g/L was obtained. CONCLUSIONS: Citrate buffer was not found to inhibit growth or lipid production in this engineered R. toruloides strain, nor did reducing the citrate buffer concentration negatively affect sugar yields in the hydrolysate. As this process is scaled-up, $131 per ton of hydrothermally pretreated biomass can be saved by use of the lower citrate buffer concentration during enzymatic hydrolysis.

Pitfalls in the 3, 5-dinitrosalicylic acid (DNS) assay for the reducing sugars: Interference of furfural and 5-hydroxymethylfurfural.

Transformation of renewable biomass into value-added chemicals and biofuels has evolved to be a vital field of research in recent years. Accurate estimation of reducing sugars post pretreatment of lignocellulosic biomass has been very inconsistent. For a few decades, 3,5-dinitrosalicylic acid (DNS) assay has been widely employed for the estimation of reducing sugars derived from pretreatment of lignocellulosic biomass. This assay tests for the presence of free carbonyl group (C=O), the so-called reducing sugars. This involves the oxidation of the aldehyde functional group present to the corresponding acid while DNS is simultaneously reduced to 3-amino-5-nitrosalicylic acid under alkaline conditions. However, the presence of other active carbonyl groups can potentially also react with DNS leading to incorrect yields of reducing sugars. Therefore, a detailed study has been carried out to evaluate the influence of active carbonyl compounds like furfural and 5-hydroxymethylfurfural (5-HMF) in the overall estimation of reducing sugars (glucose, xylose and arabinose) by DNS assay. In addition to this, reducing sugars estimation in the presence of furans were also investigated, it reveals that reducing sugars estimation was found to be 68% higher than actual sugars. Therefore, current findings strongly indicate that the employment of DNS assay for quantifying the reducing sugars in the presence of furans is not appropriate.

Dilute acid pretreatment of sorghum biomass to maximize the hemicellulose hydrolysis with minimized levels of fermentative inhibitors for bioethanol production.

Conversion of lignocellulosic biomass into monomeric carbohydrates is economically beneficial and suitable for sustainable production of biofuels. Hydrolysis of lignocellulosic biomass using high acid concentration results in decomposition of sugars into fermentative inhibitors. Thus, the main aim of this work was to investigate the optimum hydrolysis conditions for sorghum brown midrib IS11861 biomass to maximize the pentose sugars yield with minimized levels of fermentative inhibitors at low acid concentrations. Process parameters investigated include sulfuric acid concentration (0.2-1 M), reaction time (30-120 min) and temperature (80-121 °C). At the optimum condition (0.2 M sulfuric acid, 121 °C and 120 min), 97.6% of hemicellulose was converted into xylobiose (18.02 mg/g), xylose (225.2 mg/g), arabinose (20.2 mg/g) with low concentration of furfural (4.6 mg/g). Furthermore, the process parameters were statistically optimized using response surface methodology based on central composite design. Due to the presence of low concentration of fermentative inhibitors, 78.6 and 82.8% of theoretical ethanol yield were attained during the fermentation of non-detoxified and detoxified hydrolyzates, respectively, using Pichia stipitis 3498 wild strain, in a techno-economical way.