Luca Zoia

Structural characterization and antioxidant activity evaluation of lignins from rice husk.

In recent years, lignin and extractives from herbaceous plants and crops are receiving increasing attention for their renewability and large annual biomass stock. It is worth noting that only a few studies deal with the chemical characterization of rice husk, a side product of one of the most important crops with regard to human nutrition. Thus, in this study lignin from rice husk was isolated and characterized. Two different extraction procedures were optimized and tested: acidolysis and alkaline enzymatic (AE). The different lignins isolated were fully characterized by means of gravimetric, chromatographic (GPC), and spectroscopic (31P NMR, 2D-HSQC-NMR) analyses with the aim to compare yields, sample purity, and chemical properties, recognized as key parameters for future development. Notwithstanding the extraction procedure, the results highlighted that rice husk lignin is mainly formed by guaiacyl and p-hydroxyphenyl units. The acidolytic approach showed an appreciable lignin recovery and high purity, whereas the AE lignin sample was found to be rich in residual polysaccharides and oxidized functionalities. Moreover, different rice husk extracts, along with acidolysis lignin and AE lignin specimens, were assayed for their antioxidant activity by means of a DPPH radical scavenging test.

In Situ Determination of Lignin Phenolics and Wood Solubility in Imidazolium Chlorides Using 31P NMR

Corn stover, Norway spruce, and Eucalyptus grandis were pulverized to different degrees. These samples were subjected to quantitative analyses, upon the basis of predissolution into the imidazolium chloride-based ionic liquids [amim]Cl and [bnmim]Cl followed by labeling of hydroxyl groups as phosphite esters and quantitative (31)P NMR analysis. Analysis of different pulverization degrees provided semiempirical data to chart the solubility of Norway spruce in these ionic liquids. Further method refinment afforded an optimized method of analysis of the lignin phenolic functionalities, without prior isolation of the lignin from the fiber. The lignin in these samples was further enriched using cellulase and acidolysis treatments, allowing for comparison with the fibrous samples. Analysis of all samples charts the polymerized-monomer availability for each stage of the treatment. Conditions required for adequate signal-to-noise ratios in the (31)P NMR analysis were established with a notable improvement observed upon the lignin enrichment steps.

Molecular weight distributions and linkages in lignocellulosic materials derivatized from ionic liquid media.

A novel and reproducible method is described for accurately determining the molecular weight distribution by size exclusion chromatography (SEC) of whole lignocellulosic materials. This approach offers the opportunity to compare the molecular weight distributions of intact milled woods and its component fractions, lignins and holocelluloses, all from the same source, thus highlighting the potential of the technique and the contributions of the individual components to the chromatogram. The method is based on the dissolution of the ball-milled samples in the ionic liquid 1-allyl-3-methylimidazolium chloride ([amim]Cl). Under these homogeneous ionic liquid media, a derivatization reaction was performed with benzoyl chloride in the presence of pyridine. The thoroughly benzoylated wood with its associated carbohydrate and lignin components was found to be completely soluble in the THF SEC eluent with marked UV detector sensitivity. This methodology, when applied to the individually isolated holocellulose and lignin (enzymatic mild acidolysis lignin; EMAL) materials from Norway spruce ( Eucalyptus grandis ) wood and corn stover, offered a better understanding as to the possible ways the lignin and the carbohydrates may interact within these three different species. Finally, the applicability of the methodology is shown for a series of pure cellulosic samples under intense mechanical defibration conditions, offering a visualization of the molecular weight distribution changes induced during the production of nanofibrillated cellulose.

Chromatographic Detection of Lignin–Carbohydrate Complexes in Annual Plants by Derivatization in Ionic Liquid

The opportunity for detecting the presence and the amount of lignin-carbohydrate complexes (LCCs) in renewable feedstocks is a major issue for the complete utilization of biomass. Indeed, LCCs are known to shield cellulose from enzymatic hydrolysis, reducing the efficiency of the digestion processes needed for the production of biobased products. This study is focused on the chromatographic characterization of lignocellulose from agricultural residues (rice husk, wheat straw) and herbaceous energy crops ( Arundo donax , Miscanthus sinesis ) and their fractionation products (hemicellulose, cellulose, and lignin). Exploiting alternative chemical derivatizations on the aforementioned samples, it was possible to discern the connectivity among the various lignocellulosic components. The complete acetylation and benzoylation of the milled native substrates in ionic liquid media, and the systematic comparison between their GPC-UV chromatograms collected at different wavelengths has revealed itself as a straightforward technique in the detection of LCCs. This novel approach proved an extensive connectivity between the lignin and the hemicellulosic for all the analyzed specimens, whereas the cellulosic fraction was conceived as a substantially unbound moiety, accounting for the sample composition at higher molecular weights. Moreover, the collected lignin fractions were extensively characterized by means of (31)P NMR and 2D-HSQC techniques.

Laccase-initiated cross-linking of lignocellulose fibres using a ultra-filtered lignin isolated from kraft black liquor.

In this work, the effect of Trametes pubescens laccase (TpL) used in combination with a low-molecular-weight ultra-filtered lignin (UFL) to improve mechanical properties of kraft liner pulp and chemi-thermo-mechanical pulp was studied. UFL was isolated by ultra-filtration from the kraft cooking black liquor obtained from softwood pulping. This by-product from the pulp industry contains an oligomeric lignin with almost twice the amount of free phenolic moieties than residual kraft pulp lignin. The reactivity of TpL on UFL and kraft pulp was studied by nuclear magnetic resonance spectroscopy and size exclusion chromatography. Laccase was shown to polymerise UFL and residual kraft pulp lignin in the fibres, seen by the increase in their average molecular weight and in the case of UFL as a decrease in the amount of phenolic hydroxyls. The laccase initiated cross-linking of lignin, mediated by UFL, which gives rise to more than a twofold increase in wet strength of kraft liner pulp handsheets without loosing other critical mechanical properties. Hence, this could be an interesting path to decrease mechano-sorptive creep that has been reported to lessen in extent as wet strength is given to papers. The laccase/2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) mediator system showed a greater increase in wet tensile strength of the resulting pulp sheets than the laccase/UFL system. However, other mechanical properties such as dry tensile strength, compression strength and Scott Bond internal strength were negatively affected by the laccase/ABTS system.

Time-Resolved Optical Spectroscopy of Wood

We have proposed and experimentally demonstrated that picosecond time-resolved optical spectroscopy in the visible/near-infrared (NIR) region (700–1040 nm) is a useful technique for noninvasive characterization of wood. This technique has been demonstrated on both softwood and hardwood samples treated in different ways simulating the aging process suffered by waterlogged woods. In all the cases, alterations of absorption and scattering spectra were observed, revealing changes of chemical and structural composition.

Oxidation of isoeugenol by salen complexes with bulky substituents.

The catalytic properties of bulky water-soluble salen complexes in the oxidation of isoeugenol (2-methoxy-4-(1-propenyl) phenol) have been investigated in aqueous ethanol solutions in order to obtain a mixture of polymeric compounds through dehydrogenative polymerization. The average molecular weight of dehydrogenated polymers (DHPs) was monitored by GPC and correlated to reaction conditions such as time, concentration of substrate, concentration of catalyst, type of oxidation agent, etc. The DHP synthesized by adopting the best reaction conditions was characterized by different analytical techniques (GPC, 13C-NMR, 31P-NMR and LC-MS) to elucidate its structure. The lignin-like polymer resulting from isoeugenol radical coupling possesses valuable biological activity and finds applications in a variety of fields, such as packaging industry and cultural heritage conservation.

Microwave assisted lignin isolation using the Enzymatic Mild Acidolysis (EMAL) protocol

The use of microwaves is explored in an effort to further improve the recently developed lignin isolation protocol termed EMAL (enzymatic mild acidolysis lignin). Because the presence of the lignin-carbohydrate linkages seems to be rather pronounced within wood, a microwave reactor was used to replace traditional refluxing during the mild acidolysis step. This was done in an attempt to augment the selectivity of this step toward cleaving lignin-carbohydrate bonds as well as reducing the overall intensity of this step toward inducing changes in the lignin structure, thus affording lignin in greater yields and purities. Consequently, in this study the yields, purities, and structures of lignins isolated from spruce (softwood) by the EMAL protocol under various microwave conditions were examined. The variables studied included microwave power, microwave heating time, hydrochloric acid concentration and water content of the reaction medium. Microwave heating afforded EMAL samples of high purity (90%, comparable to the conventional protocol) but in significantly greater gravimetric yields. Quantitative (31)P NMR and SEC data confirmed that the structure of lignin was similar to that obtained by traditional EMALs, with comparable contents of beta-aryl ether bonds, phenolic hydroxyls (condensed and uncondensed), and carboxylic acids.

Colour removal and carbonyl by-production in high dose ozonation for effluent polishing

Experimental tests have been conducted to investigate the efficiency and the by-product generation of high dose ozonation (10-60 mg O3 L(-1)) for complete colour removal from a treated effluent with an important component of textile dyeing wastewater. The effluent is discharged into an effluent-dominated stream where no dilution takes place, and, thus, the quality requirement for the effluents is particularly strict. 30, 60 and 90 min contact times were adopted. Colour was measured as absorbance at 426, 558 and 660 nm wavelengths. pH was monitored throughout the experiments. The experimental work showed that at 50 mg L(-1) colour removal was complete and at 60 mg O3 L(-1) the final aldehyde concentration ranged between 0.72 and 1.02 mg L(-1). Glyoxal and methylglyoxal concentrations were directly related to colour removal, whereas formaldehyde, acetaldehyde, acetone and acrolein were not. Thus, the extent of colour removal can be used to predict the increase in glyoxal and methylglyoxal concentrations. As colour removal can be assessed by a simple absorbance measurement, in contrast to the analysis of specific carbonyl compounds, which is much longer and complex, the possibility of using colour removal as an indicator for predicting the toxic potential of ozone by-products for textile effluents is of great value.

Understanding the radical mechanism of lipoxygenases using 31P NMR spin trapping.

In this paper, we use our quantitative (31)P NMR spin trapping methods, already developed for simple oxygen- and carbon-centered radicals, to understand the radical intermediates generated by enzymatic systems and more specifically lipoxygenases. Our methodology rests on the fact that free radicals react with the nitroxide phosphorus compound, 5-diisopropoxy-phosphoryl-5-methyl-1-pyrroline-N-oxide (DIPPMPO), to form stable radical adducts, which are suitably detected and accurately quantified using (31)P NMR in the presence of a phosphorus containing internal standard. This system was thus applied to better understand the mechanism of enzymatic oxidation of linoleic acid by soybean lipoxygenases-1 (LOX). The total amount of radicals trapped by DIPPMPO was detected by (31)P NMR at different experimental conditions. In particular the effect of dioxygen concentration on the amount of radicals being trapped was studied. At low dioxygen concentration, a huge increase of radicals trapped was observed with respect to the amount of radicals being trapped at normal dioxygen concentrations.