Quantification of lignin-carbohydrate linkages with high-resolution NMR spectroscopy.

A quantitative approach to characterize lignin-carbohydrate complex (LCC) linkages using a combination of quantitative ¹³C NMR and HSQC 2D NMR techniques has been developed. Crude milled wood lignin (MWLc), LCC extracted from MWLc with acetic acid (LCC-AcOH) and cellulolytic enzyme lignin (CEL) preparations were isolated from loblolly pine (Pinus taeda) and white birch (Betula pendula) woods and characterized using this methodology on a routine 300 MHz NMR spectrometer and on a 950 MHz spectrometer equipped with a cryogenic probe. Structural variations in the pine and birch LCC preparations of different types (MWL, CEL and LCC-AcOH) were elucidated. The use of the high field NMR spectrometer equipped with the cryogenic probe resulted in a remarkable improvement in the resolution of the LCC signals and, therefore, is of primary importance for an accurate quantification of LCC linkages. The preparations investigated showed the presence of different amounts of benzyl ether, γ-ester and phenyl glycoside LCC bonds. Benzyl ester moieties were not detected. Pine LCC-AcOH and birch MWLc preparations were preferable for the analysis of phenyl glycoside and ester LCC linkages in pine and birch, correspondingly, whereas CEL preparations were the best to study benzyl ether LCC structures. The data obtained indicate that pinewood contains higher amounts of benzyl ether LCC linkages, but lower amounts of phenyl glycoside and γ-ester LCC moieties as compared to birch wood.

Quantitative characterization of a hardwood milled wood lignin by nuclear magnetic resonance spectroscopy.

The structure of Eucalyptus grandis milled wood lignin (MWL) was investigated by 2D 1H-13C HSQC, HMQC, and 1H-1H TOCSY correlation NMR techniques and by quantitative 13C NMR as well as by the permanganate oxidation degradation technique. The combination of 2D NMR and quantitative 13C NMR spectroscopy of nonacetylated and acetylated lignin preparations allowed reliable identification and calculation of the amount of different lignin structures. About 85% of side-chain moieties were estimated on the structural level. This information was substantiated by data on the quantity of various functional groups and interunit linkages as a whole. A modified method for calculation of the h:g:s ratio has been suggested and compared with previously suggested approaches. E. grandis MWL has been determined to have an h:g:s ratio of 2:36:62. The amounts of various phenolic/etherified noncondensed/condensed guaiacyl and syringyl moieties were approximately estimated. E. grandis MWL contained approximately 0.60/Ar of beta-O-4 moieties along with small amounts of other structural units such as pino/syringyresinol (0.03/Ar), phenylcoumaran (0.03/Ar), and spirodienone (0.05/Ar). The degree of condensation was estimated at approximately 21%; the main condensed structures are 4-O-5 moieties (approximately 0.09/Ar). The structure of E. grandis MWL was compared with those of other lignin preparations isolated from various hardwoods.

Rapid screening of wood chemical component variations using transmittance near-infrared spectroscopy.

A rapid transmittance near-infrared (NIR) spectroscopy method was developed to predict the variation in chemical composition of solid wood. The effect of sample preparation, sample quantity (single versus stacked multiple wood wafers), and NIR acquisition time on the quantification of alpha-cellulose and lignin content was investigated. Strong correlations were obtained between laboratory wet chemistry values and the NIR-predicted values. In addition to the experimental protocol and method development, improvements in calibration error associated with utilizing stacked multiple wood wafers as opposed to single wood wafers are also discussed.

Reaction mechanisms in delignification of pine Kraft-AQ pulp with hydrogen peroxide using Mn(IV)-Me4DTNE as catalyst.

Pine Kraft-AQ pulp was bleached with hydrogen peroxide catalyzed by [LMn(IV)(2) (mu-O)(3)](ClO(4))(2) at 80 degrees C for 120 min under optimum reaction conditions. The resulting bleached pulp was hydrolyzed with cellulase to obtain insoluble and soluble residual lignins. The alkaline effluent from the bleaching was acidified to precipitate alkaline soluble lignin. These lignin preparations were purified, and then analyzed by 2D HMQC NMR spectroscopic techniques. The results showed that biphenyl (5-5) and stilbene structures are preferentially degraded in the bleaching process, while beta-O-4, beta-5, and beta-beta structures undergo degradation only to a lesser extent. This implies that hydrogen peroxide bleaching using the catalyst is more effective in delignification of softwood pulps than hardwood pulps. The possible reaction mechanisms for the delignification of residual lignin in the pine Kraft-AQ pulp in the bleaching process are discussed on the basis of the 2D HMQC NMR spectroscopic data and the model compound experiments.

A comprehensive approach for quantitative lignin characterization by NMR spectroscopy.

A detailed approach for the quantification of different lignin structures in milled wood lignin (MWL) has been suggested using a combination of NMR techniques. 1H-13C heteronuclear multiple quantum coherence and quantitative 13C NMR of nonacetylated and acetylated spruce MWL have been found to have a synergetic effect, resulting in significant progress in the characterization of lignin moieties by NMR. About 80% of side chain moieties, such as different beta-O-4, dibenzodioxocin, phenylcoumaran, pinoresinol, and others, have been identified on the structural level. The presence of appreciable amounts of alpha-O-alkyl and gamma-O-alkyl ethers has been suggested. Although the quantification of various condensed moieties was less precise than for side chain structures, reliable information can be obtained. Comparison of the calculated results with known databases on spruce MWL structure shows that the suggested approach is rather informative and comparable with the information obtained from the combination of various wet chemistry methods. Discrepancies between the results obtained in this study and those previously published are discussed.

Comparative studies on the delignification of pine kraft-anthraquinone pulp with hydrogen peroxide by binucleus Mn(IV) complex catalysis.

Pine kraft-anthraquinone (kraft-AQ) pulp was bleached in alkaline solution with hydrogen peroxide catalyzed by either [L(1)Mn(IV)(micro-O)(3)Mn(IV)L(1)](PF(6))(2)] (C1) or [LMn(IV)(2)(micro-O)(3)] (ClO(4))(2) (C2) at 60 and 80 degrees C for 120 min with a catalyst charge of 10 ppm on pulp. The resulting bleached pulp was hydrolyzed with cellulase to obtain insoluble and soluble residual lignins. The alkaline bleaching effluents were acidified to precipitate alkaline-soluble lignins. These lignin preparations were then characterized by 2D heteronuclear multiple-quantum coherence (HMQC) NMR spectroscopic techniques. The results showed that biphenyl (5-5) and stilbene structures of the residual lignin in the pulp are preferentially degraded in both the C1- and C2-catalyzed bleachings, whereas beta-O-4, beta-5, and beta-beta structures undergo degradation to a lesser extent. In both cases, the degradation of the residual lignin increased with the increase in reaction temperature from 60 to 80 degrees C. Thus, the result of C1-catalyzed delignification is not in agreement with the observed decrease in the disappearance rate for substrates in the C1-catalyzed oxidation of lignin model compounds with hydrogen peroxide when the reaction temperature is increased from 60 to 80 degrees C. In addition, the resulting residual lignins in the C2-catalyzed bleaching at 80 degrees C are less degraded than the corresponding lignins in the C1-catalyzed bleaching at both 60 and 80 degrees C. Thus, C1 is more effective than C2 as catalyst in the binucleus Mn(IV) complex-catalyzed bleaching of pine kraft-AQ pulp with hydrogen peroxide.

Elucidation of the structures of residual and dissolved pine kraft lignins using an HMQC NMR technique.

Comparative studies on the structures of residual and dissolved lignins isolated from pine kraft pulp and pulping liquor have been undertaken using the (1)H-(13)C HMQC NMR technique, GPC, and sugar analysis to elucidate the reaction mechanisms in kraft pulping and the lignin reactivity. A modified procedure for the isolation of enzymatic residual lignins has resulted in an appreciable decrease in protein contaminants in the residual lignin preparations (N content < 0.2%). The very high dispersion of HMQC spectra allows identification of different lignin moieties, which signals appear overlapped in 1D (13)C NMR spectra. Elucidation of the role of condensation reactions indicates that an increase in the degree of lignin condensation during pulping results from accumulation of original condensed lignin moieties rather than from the formation of new alkyl-aryl structures. Among aryl-vinyl type moieties, only stilbene structures are accumulated in lignin in appreciable amounts. Benzyl ether lignin-carbohydrate bonds involving primary hydroxyl groups of carbohydrates have been detected in residual and dissolved lignin preparations. Structures of the alpha-hydroxyacid type have been postulated to be among the important lignin degradation products in kraft pulping. The effect of the isolation method on the lignin structure and differences between the residual and dissolved lignins are discussed.

Lignin structural variation in hardwood species.

A comprehensive lignin structure analysis of ten industrially relevant hardwood species is presented. Milled wood lignin (MWL) was isolated from each species using a modified protocol and all milled wood lignin preparations were analyzed through quantitative (13)C NMR spectroscopy, elemental analysis, methoxyl analysis, sugar analysis, and nitrobenzene oxidation. Nitrobenzene oxidation and ozonation were carried out on extractive-free wood, alkali-extracted wood, milled wood lignin, and alkali-extracted lignin. Milled wood lignin isolated by the modified protocol was found to be representative of the total lignin in alkali-extracted wood. Significant variations in lignin structures, such as syringylpropane/guaiacylpropane ratio (S/G ratio), arylglycerol-ß-aryl ether (ß-O-4), degree of condensation, and elemental and methoxyl contents, were found among the hardwood species studied. These structural variations among species appear to be correlated to a single factor, the syringyl/guaiacyl ratio. A new method to predict the S/G ratio of total lignin in wood was developed, using a calibration line established by the syringaldehyde/vanillin (S/V) ratio (nitrobenzene oxidation) and the S/G ratio ((13)C NMR) of milled wood lignin (MWL).

Lignin changes after steam explosion and laccase-mediator treatment of eucalyptus wood chips.

Eucalyptus globulus chips were steam exploded followed by treatment with a laccase-mediator system (LMS) under different experimental conditions. Removal of hemicelluloses and, to a lesser extent, lignin was observed. Thermogravimetic analyses of whole meal obtained from chips before and after steam explosion indicated an increase in lignin degradation temperature due to lignin condensation. In contrast, application of LMS treatment caused a reduction in lignin and polysaccharide degradation temperatures. Lignins were isolated from wood samples before and after each treatment and analyzed by 2D NMR and (13)C NMR. An increase in carboxyl and phenolic hydroxyl groups and a significant decrease in ß-O-4 structures were found in steam-exploded samples. The most relevant changes observed after laccase treatment were increased secondary OH and degree of condensation.