Enrichment and Identification of Lignin–Carbohydrate Complexes in Softwood Extract

Lignin–carbohydrate complexes (LCCs) are hybrid structures containing covalently linked moieties of lignin and carbohydrates. The structure and behavior of LCCs affect both industrial processes and practical applications of lignocellulosic biomass. However, the identification of phenylglycoside, benzylether, and gamma (γ)-ester LCC bonds in lignocellulosic biomass is limited due to their relatively low abundance compared to plain carbohydrate and lignin structures. Herein, we enriched the LCC bonds in softwood galactoglucomannan (GGM)-rich extract fractionated by (1) a solvent (ethanol), (2) enzymes, and (3) physical techniques. Two-dimensional nuclear magnetic resonance (NMR) spectroscopy analysis was used to identify the LCC bonds. Phenylglycoside and benzylether bonds were concentrated in the ethanol-soluble GGM fractions. A benzylether bond was concentrated into GGM fractions containing larger molecules (>500 Da) through physical techniques. The γ-ester bond was identified in all studied GGM fractions, which is explained by its stability and possible presence in residual xylan. In summary, we demonstrated the potential of the suggested techniques to enrich LCC bonds in softwood extract and improve LCC identification. Such techniques may also enable further studies on the structure and functionality of LCC bonds and open new prospects in the engineering of biomolecules.