Downstream extraction process development for recovery of organic acids from a fermentation broth.

The present study focused on organic acids (OAs) recovery from an acidogenic fermentation broth, which is the main problem regarding the use of OAs for production of ester-based new generation biofuels or other applications. Specifically, 10 solvents were evaluated for OAs recovery from aqueous media and fermentation broths. The effects of pH, solvent/OAs solution ratios and application of successive extractions were studied. The 1:1 solvent/OAs ratio showed the best recovery rates in most cases. Butyric and isobutyric acids showed the highest recovery rates (80-90%), while lactic, succinic, and acetic acids were poorly recovered (up to 45%). The OAs recovery was significantly improved by successive 10-min extractions. Alcohols presented the best extraction performance. The process using repeated extractions with 3-methyl-1-butanol led to the highest OAs recovery. However, 1-butanol can be considered as the most cost-effective option taking into account its price and availability.

Economic evaluation of technology for a new generation biofuel production using wastes.

An economic evaluation of an integrated technology for industrial scale new generation biofuel production using whey, vinasse, and lignocellulosic biomass as raw materials is reported. Anaerobic packed-bed bioreactors were used for organic acids production using initially synthetic media and then wastes. Butyric, lactic and acetic acid were predominately produced from vinasse, whey, and cellulose, respectively. Mass balance was calculated for a 16,000L daily production capacity. Liquid-liquid extraction was applied for recovery of the organic acids using butanol-1 as an effective extraction solvent which serves also as the alcohol for the subsequent enzyme-catalyzed esterification. The investment needed for the installation of the factory was estimated to about 1.7million€ with depreciation excepted at about 3months. For cellulosics, the installation investment was estimated to be about 7-fold higher with depreciation at about 1.5years. The proposed technology is an alternative trend in biofuel production.

New generation biofuel: continuous acidogenesis of sucrose-raffinose mixture simulating vinasse is promoted by γ-alumina pellets.

BACKGROUND: This investigation comprises a contribution on the production of a new generation biofuel using the industrial liquid waste of bioethanol distilleries, known as vinasse. This study focuses on the exploitation of vinasse as an acidogenesis substrate for volatile fatty acids and simultaneous ethanol production. These products can be used for ester production, which is the new generation biofuel. Therefore, the aims of the present study are (i) to examine any promotional effect of γ-alumina on acidogenesis of a sucrose-raffinose mixture simulating vinasse, (ii) to study the operational stability of the continuous acidogenesis of sucrose and raffinose and subsequently of vinasse, and (iii) to determine the volatile fatty acid chemical composition and ethanol formation. RESULTS: Batch acidogenesis of sucrose and raffinose mixtures showed that γ-alumina promoted fermentation leading to an increase in the volatile fatty acid yield factor from 40% to 95% compared to free cells. The application of the system in continuous mode for more than 3 months showed that the continuous volatile fatty acid productivity obtained was higher than 7 g/L/day. Lactic acid was the predominant acid when sucrose and raffinose were used while butyric acid in the case of vinasse. The highest volatile fatty acid concentration reached was 19 g/L for vinasse. CONCLUSIONS: A promoting effect of γ-alumina in acidogenic fermentation of sucrose-raffinose and vinasse is reported. Continuous acidogenesis of sucrose-raffinose mixtures and vinasse using γ-alumina with immobilized cells showed high operational stability (more than 3 months). These findings result in easy scale up of the process that will produce a high annual added value of $11,000,000 in a daily bioethanol production plant of 50,000 L.

Continuous acidogenesis of sucrose, raffinose and vinasse using mineral kissiris as promoter.

The use of kissiris as promoter (culture immobilization carrier) in anaerobic acidogenesis of sucrose, raffinose and vinasse is reported. Initially, the effect of pH (4-8) and fermentation temperature (18-52 °C) on the accumulation of low molecular weight organic acids (OAs) during sucrose acidogenesis, was evaluated. The promoting effect of kissiris was confirmed compared to free cells, resulting in 80% increased OAs production. The optimum conditions (pH 8; 37 °C) were used during acidogenesis of sucrose/raffinose mixtures. A continuous system was also operated for more than 2 months. When sucrose and sucrose/raffinose mixtures were used, lactic acid type fermentation prevailed, while when vinasse was used, butyric acid type fermentation occurred. Total OAs concentrations were more than 14 g/L and ethanol concentrations were 0.5-1 mL/L. Culture adaptation in vinasse was necessary to avoid poor results. The proposed process is promising for new generation ester-based biofuel production from industrial wastes.

Tailoring structure-function properties of L-asparaginase: engineering resistance to trypsin cleavage.

Bacterial L-ASNases (L-asparaginases) catalyse the conversion of L-asparagine into L-aspartate and ammonia, and are widely used for the treatment of ALL (acute lymphoblastic leukaemia). In the present paper, we describe an efficient approach, based on protein chemistry and protein engineering studies, for the construction of trypsin-resistant PEGylated L-ASNase from Erwinia carotovora (EcaL-ASNase). Limited proteolysis of EcaL-ASNase with trypsin was found to be associated with a first cleavage of the peptide bond between Lys53 and Gly54, and then a second cleavage at Arg206-Ser207 of the C-terminal fragment, peptide 54-327, showing that the initial recognition sites for trypsin are Lys53 and Arg206. Site-directed mutagenesis of Arg206 to histidine followed by covalent coupling of mPEG-SNHS [methoxypoly(ethylene glycol) succinate N-hydroxysuccinimide ester] to the mutant enzyme resulted in an improved modified form of EcaL-ASNase that retains 82% of the original catalytic activity, exhibits enhanced resistance to trypsin degradation, and has higher thermal stability compared with the wild-type enzyme.