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The stability and activity of the low redox potential Melanocarpus albomyces laccase (MaL) in various aqueous organic (acetone, ethanol, propylene glycol, diethylene glycol monomethyl ether) solvent systems was studied spectrophotometrically using 2,6-dimethoxyphenol (2,6-DMP) as substrate. In addition, reactivity of the enzyme with two lignans; matairesinol (MR) and 7-hydroxymatairesinol (HMR), was examined by oxygen consumption measurements in the most potential aqueous organic solvent systems. Polymerization of the lignans by MaL was verified by matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) and size exclusion chromatography (SEC). Polymerization of the higher molecular weight lignin model compound, dehydrogenation polymers (DHPs), was studied by SEC. The solubilities of industrial softwood and hardwood kraft lignins were evaluated as parameters for investigation of enzymatic modification in aqueous organic solvent systems. The functioning of MaL in different aqueous organic media was excellent. Propylene glycol and diethylene glycol monomethyl ether were better solvents than ethanol or acetone in enzymatic oxidations. Even though they were the best solvents for enzyme oxidation, ethanol and propylene glycol were selected for further tests because of their different physicochemical properties. The results obtained in this study for the use of laccase-catalysed reactions in organic solvents to improve the efficiency of lignin oxidation may be exploited in several applications and areas in which the solubility of the reactants or products is a limiting factor.

International audience; In plant cell walls, lignin polymerization occurs in concentrated polysaccharide gel. The effect of high polysaccharide concentrations on the structure of lignin-like polymers (DHPs = dehydrogenation polymers), were investigated by running lignification-like polymerization under three reaction conditions in which the concentrations of all reactants (xylan/coniferyl alcohol (CA)/oxidising system) were gradually increased. Control experiments were also run in similar conditions but without polysaccharides. DHPs showed increased solubility with increased concentrations of reactants in the presence of xylans but were mostly insoluble in buffer control experiments. The structures of DHPs were characterized by thioacidolysis and size exclusion chromatography (SEC). Results indicated that the frequency of [beta]-alkyl aryl ether bonds and DHP molecular weight increased with increasing concentration of the reaction mixture in the presence of xylans whereas those of control DHPs decreased slightly under the same conditions. This emphasizes the role of the pre-existing polysaccharide gel and high concentrations existing in the cell wall during construction of the lignin polymer.

International audience; In order to mimic the first steps of lignification, pectin/synthetic lignin (dehydrogenation polymers (DHPs)) interactions were investigated using reconstructing approaches. In artificially lignified bacterial cellulose(BC)/pectin composites, DHP nodules display a more dispersed distribution than in BC mats. Occurrence of covalent linkages between pectin and DHP was investigated by solid state NMR. Non-covalent bonds seem to govern the associative behaviour and stability of the complexes. Studied by small angle neutron scattering, a compaction process of DHP/pectin clusters occurs during polymerization and phase separation of the two polymers was shown at the nanometer scale.

International audience; The properties of monolayers of the dioxan lignin and of model dehydrogenation polymers spread at the air-water interface were investigated by the following techniques: surface pressure, surface potential, ellipticity, rheological analysis and molecular modeling. Information about the state of the monolayer was obtained from Surface pressure-area and surface potential-area isotherms. At small deposited quantities (1-2 mgm(-2)) a quasi-monolayer (2D) structure is formed. The values obtained for the mean area per monomer and for the mean dipole moment Lire coherent with previously reported data. as well as with the proposed Molecular model. At higher deposited quantities (3-6 mgm(-2)) a complex 3D network is built. These statements are supported by the results obtained by means of ellipsometry and surface rheology. Ellipsometry provides data for the stability of the 2D structure and for the slightly unstable 3D network. By using an original rheological approach. both structures Lire distinguished by their mechanical response when a dilatational stress is applied. The quasi-monolayer structure behaves as in elastic 2D medium during the compression, while two relaxation processes with characteristic time of 10 and 100 s were observed. The 3D network behaves as a Maxwell viscoelastic body during, the compression. while three relaxation processes with characteristic times of 1.10 and 100 s were detected.

International audience; The mechanism of lignin carbohydrate complex formation by addition of polysaccharides on quinone methide (QM) generated during lignin polymerisation was investigated using a model approach. Dehydrogenation polymers (DHPs, lignin model compounds) were synthesized from coniferyl alcohol in the presence of a glucuronoarabinoxylan (GAX) extracted from oat spelts, by Zutropfverfahren (ZT) and Zulaufverfahren (ZL) methods. The methods ZT and ZL differed in their distribution of QM over the reaction period but generated roughly the same QM amount. Steric exclusion chromatography of the ZT and ZL reaction products showed that only the ZT reaction produced high molar mass compounds. Covalent linkages in the ZT reaction involving ether bonds between GAX moiety and alpha carbon of the lignin monomer were confirmed by C-13 NMR and xylanase-based fractionation. The underlying phenomena were further investigated by examining the interactions between GAX and DHP in sorption experiments. GA! X and DHPs were shown to interact to form hydrophobic aggregates. In the ZT process, slow addition permitted polymer reorganisation which led to dehydration around the lignin-like growing chains thereby limiting the addition of water on the quinone methide formed during polymerisation and thus favoured lignin-carbohydrate complex (LCC) formation.

International audience; Dehydrogenation polymers (DHPs. lignin model compounds) were synthesized in the presence of increasing pectin concentrations using two different methods. The first method ('Zutropfverfahren'. ZT) consists in the slow adding of monomers whereas in the second method ('Zulaufverfahren', ZL) all the reactants are added simultaneously. DHPs solubility increases with the pectin concentration in the ZT experiments and remains stable in the ZL experiments. Covalent bonds between pectin and DHP are formed during ZT polymerization resulting in lignin carbohydrate complex (LCC) which keeps the unbound DHPs in solution by the formation of aggregate or micelle-like structures. In contrast LCC are not formed during the ZL process which behave like the DHP reference. The ZT DHP molar masses increase observed is attributed to the reactivity of the high molar mass polymer solubilized by the: LCC whereas ZL higher molar mass polymers are precipitated out of the solution and cannot react further.