Rapid High-Resolution Analysis of Polysaccharide-Lignin Interactions in Secondary Plant Cell Walls Using Proton-Detected Solid-State NMR

The plant secondary cell wall, a complex matrix composed of cellulose, hemicellulose, and lignin, is crucial for the mechanical strength and water-proofing properties of plant tissues, and serves as a primary source of biomass for biorenewable energy and biomaterials. Structural analysis of these polymers and their interactions within the secondary cell wall has been heavily relying on ¹³C-based solid-state NMR techniques. In this study, we explore the application of ¹H-detected solid-state NMR techniques for rapid, high-resolution structural characterization of polysaccharides and lignin, demonstrated on the stems of hardwood eucalyptus. We explored the use of synthesized 2D spectra to resolve central ¹H resonances and the combined application of 3D hCCH and hCHH experiments for complete resonance assignment and unambiguous identification of lignin-carbohydrate interactions. Our findings emphasize the central role of acetylated 3-fold xylan conformers, rather than 2-fold, in stabilizing the carbohydrate-lignin interface, with glucuronic acid side chains in eucalyptus glucuronoxylan colocalizing with lignin. We also observed cellulose-lignin interactions involving uncoated microfibril surfaces and detected pectin-lignin interactions indicative of early stage lignification. These results present a novel approach for rapid structural analysis of lignocellulosic biomaterials without the need for solubilization or extraction.