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.

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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.

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.