For the first time, the bonding patterns of specific carbon atoms in woody plant lignin have been identified in situ. This was accomplished by administering and incorporating into the lignin fraction of Leucaena leucocephala, a tropical hardwood, ferulic acid enriched with ¹³C at either the 1-, 2-, or 3-C atom of the side chain. The plants were grown hydroponically over extended periods of time (28 days) under aseptic conditions in media containing the ferulic acid precursor, and then the tissues were examined by solid-state ¹³C NMR spectroscopy. Consequently, resonances due to the bonding patterns of the specific carbon atoms were determined. These resonances differ substantially from similarly labelled synthetic dehydrogenatively polymerized (DHP) lignin in both spectral profile and relative peak intensities. Subsequent studies using phenylalanine as precursor showed that it was better translocated into the aerial portions of the plant, and that its uptake did not result in distortion of lignification in those tissues, both in amount and monomeric composition. Consequently, the difference spectra obtained by ¹³C NMR analyses of phenylalanine-treated plants confirmed and extended the results obtained with ferulic acid. Evidence for the conversion of both precursors to the monolignols was shown by the difference spectra of [1-¹³C]-precursor-fed tissues, where the dominant resonance at 61-63 ppm is consistent with substructures containing the hydroxymethyl functionality. The spectrum obtained with roots administered [1-¹³C] ferulic acid showed the presence of a minor resonance (170-174 ppm) attributable to carboxylic acids/esters. By allowing the plant to undergo further metabolism by growing in hydroponic media without the precursor, these signals disappeared from the resulting spectrum. The first direct evidence for the dominance of the β-O-4’ linkage of lignin in situ was shown by the appearance of the resonance at 83 ppm corresponding to this substructure in both [2-¹³C] ferulic acid-treated roots and [2-¹³C] phenylalanine-treated roots and stems. Evidence for the occurrence of α-O-carbohydrate or α-O-aryl linkage in intact plant tissues was obtained in the spectra of tissues administered [3-¹³C] ferulic acid and [3-¹³C] phenylalanine. The effect of horseradish peroxidase/H₂O₂ in organic medium (dioxane/aqueous acetate buffer, pH 5, 95:5) on dehydrogenatively polymerized (DHP) lignin was reinvestigated. We found no evidence for vigorous depolymerization of DHP lignin under these conditions, contrary to claims made by Dordick, Marletta and Klibanov (1986, Proc. Natl. Acad. Sci. USA 83:6255-6257). Furthermore, we did not detect ferulic acid as a degradation product following treatment of DHP lignin with HRP/H₂O₂. Both coniferyl alcohol and DHP lignin were used in incubation experiments to determine effects of lignin peroxidase from the white-rot fungus Phanerochaete chrysosporium and H₂O₂ on these substrates. Gel filtration chromatography showed that polymeric materials of high molecular weights were the result of these treatments. Incubation of [1-¹³C], [2-¹³C] and [3-¹³C] coniferyl alcohol with lignin peroxidase/H₂O₂ resulted in products similar to-DHP lignins prepared by horseradish peroxidase/H₂O₂ with respect to occurrence of identical resonances in corresponding solution-state ¹³C NMR spectra. Consequently, the role of polymerization of low molecular weight phenolics as a mechanism for detoxification was ascribed to these fungal peroxidases. / Ph. D.