NMR spectroscopy: An invaluable tool to identify lignin structural modifications induced by oxidative enzymes

This chapter provides a comprehensive guide to the acquisition, processing, and analysis of NMR data for investigating the behaviour of lignins in enzymatic treatment. The necessary pre-treatments to obtain purified lignin samples, free from enzymes and contaminants, are described prior to the spectroscopic analyses. Detailed instructions for using quantitative ¹³C NMR with an internal standard are then given to determine the abundance of lignin bonding patterns, expressed both as mmol/g of lignin and normalised on the basis of an aromatic unit. Quantitative ³¹P NMR is the favoured analytical tool used to distinguish and quantify hydroxyl groups, which are key targets for oxidases. Finally, both qualitative and quantitative HSQC analyses (QQ-HSQC and HSQC₀) are described to monitor overall changes in lignin bonding patterns.