BackgroundThe enzymatic hydrolysis of cellulose is still considered as one of the main limiting steps of the biological production of biofuels from lignocellulosic biomass. It is a complex multistep process, and various kinetic models have been proposed. The cellulase enzymatic cocktail secreted by Trichoderma reesei has been intensively investigated. β-glucosidases are one of a number of cellulolytic enzymes, and catalyze the last step releasing glucose from the inhibitory cellobiose. β-glucosidase (BGL1) is very poorly secreted by Trichoderma reesei strains, and complete hydrolysis of cellulose often requires supplementation with a commercial β-glucosidase preparation such as that from Aspergillus niger (Novozymes SP188). Surprisingly, kinetic modeling of β-glucosidases lacks reliable data, and the possible differences between native T. reesei and supplemented β-glucosidases are not taken into consideration, possibly because of the difficulty of purifying BGL1.ResultsA comparative kinetic analysis of β-glucosidase from Aspergillus niger and BGL1 from Trichoderma reesei, purified using a new and efficient fast protein liquid chromatography protocol, was performed. This purification is characterized by two major steps, including the adsorption of the major cellulases onto crystalline cellulose, and a final purification factor of 53. Quantitative analysis of the resulting β-glucosidase fraction from T. reesei showed it to be 95% pure. Kinetic parameters were determined using cellobiose and a chromogenic artificial substrate. A new method allowing easy and rapid determination of the kinetic parameters was also developed. β-Glucosidase SP188 (Km = 0.57 mM; Kp = 2.70 mM) has a lower specific activity than BGL1 (Km = 0.38 mM; Kp = 3.25 mM) and is also more sensitive to glucose inhibition. A Michaelis-Menten model integrating competitive inhibition by the product (glucose) has been validated and is able to predict the β-glucosidase activity of both enzymes.ConclusionsThis article provides a useful comparison between the activity of β-glucosidases from two different fungi, and shows the importance of fully characterizing both enzymes. A Michaelis-Menten model was developed, including glucose inhibition and kinetic parameters, which were accurately determined and compared. This model can be further integrated into a cellulose hydrolysis model dissociating β-glucosidase activity from that of other cellulases. It can also help to define the optimal enzymatic cocktails for new β-glucosidase activities.
Résumé -Ester d'acide gras en Europe : tendances de marché et perspectives technologiques -En raison d'un système initialement fortement incitatif mis en place par l'Union Européenne, la production d'ester d'acide gras (EAG) commence à s'étendre et à gagner le monde. Un examen approfondi du système complexe de compensations financières proposé par les états membres et une mise à jour annoncée en matière d'augmentation des capacités de production de biodiesel peuvent permettre de dégager les facteurs de succès de cette expansion aussi brutale que soudaine. Ce tissu de données économiques et législatives démontre l'existence de solutions technologiques alternatives pertinentes permettant de pérenniser ce succès. En effet, cette hausse de la production attendue d'EAG engage de plus en plus à se soucier de l'optimisation et de la rationalisation des unités de forte capacité pour qu'elles utilisent la meilleure technologie disponible visant des rendements élevés tout en veillant au respect de l'environnement, particulièrement au regard de la co-production de déchets. Cet article présente une technologie alternative permettant d'apporter une réponse originale à ce problème, technologie mettant en oeuvre un catalyseur hétérogène pour la transesterification des huiles végétales. Ce procédé s'appuie sur la réaction de méthanolyse des huiles végétales, réaction réalisée sur deux réacteurs successifs en lit fixe, suivie par des étapes de séparation du méthanol en excès et de la glycérine de l'ester méthylique formé. Ce procédé ne requiert ni section de récupération du catalyseur, ni traitement de rejet aqueux. Il montre un haut rendement en ester méthylique, proche de la limite thermodynamique. La conversion peut égale-ment être ajustée sans avoir recours à une consommation de catalyseur complémentaire. La glycérine co-produite possède directement une pureté de 98 % et est exempte de contaminants sous forme de sels ou de savons. L'absence de rejets et la qualité des produits fait que ce procédé est, au-delà de l'aspect 'environmentally friendly' inhérent à ses produits, un « procédé vert ».
Abstract -Fatty Acid Esters in
The demand for transportation fuels - gasoline (for cars), diesel (for trucks and cars), and kerosene (for aircraft) - is predicted to increase. The fastest growth will be observed for kerosene, in competition with diesel, inducing constraints on diesel. At the same time, all of these fuels are derived mainly from oil (more than 95 %), thus generating growing, uncontrolled CO2 emissions. Therefore, production of diesel derived from biomass (the so-called biodiesel) appears as a major objective. In this paper, we describe the existing industrial processes, discuss the possible improvements, and present the new routes (the "second-generation" processes) under development that will allow biodiesel to gain a significant percentage of the diesel (and maybe of middle distillates) pool.
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