| Lignins from cinnamyl alcohol dehydrogenase (CAD)-deficient pine and tobacco transgenic plants were examined by using 1D and 2D NMR, which provided evidence for the incorporation of hydroxycinnamyl aldehydes into the lignins. Horseradish peroxidase and hydrogen peroxide (H2O2) were used to radically couple hydroxycinnamyl aldehydes and produced dehydrodimers, with 8-O-4-, 8-5-, 8-8-, and 5-5-linkages.; Antisense-CAD-downregulated transgenic tobacco showed peaks for typical hydroxycinnamyl aldehyde and syringaldehyde end groups, 8-8-structures, and two types of 8-O-4 cross-coupled structures. All the peaks were significantly enhanced over the normal tobacco lignin. Two types of 8-0-4 peaks can be distinguished by NMR data of their side chains where G-G, S-G, G-S and S-S structures are possible.* However, the G-G structure was not involved in the CAD-downregulated transgenic tobacco lignin.; The CAD-deficient pine mutant showed coniferyl aldehyde end groups and significant vanillin unit peaks. The pine mutant did not have a detectable 8-8-structure peak analogous to that involved in transgenic tobacco lignin, but appeared to have the two weak peaks analogous to G-S or S- S 8-O-4-structures in tobacco. Since gymnosperm lignin does not contain syringyl units, these peaks are presumably attributable to 8-O-4 structures with alternatively 5-linked guaiacyl units (5-5 or 5-O-4). The pine mutant lignin also had significant levels of strikingly unexpected structures, such as dihydrocinnamyl alcohol (DHCA) and derived arylpropane-1,3-diol (APD) units, which are not significantly involved in CAD-downregulated transgenic tobacco lignin.; Valuable marker compounds can be derived by cleaving the new 8-O-4 structures of CAD-deficient transgenic lignins. The indene structures of marker compounds from thioacidolysis were elucidated by using 8-O-4 aldehyde model compounds.; Finally, we demonstrated that a commonly used aldehyde-specific color reaction, the Wiesner reaction, does not actually reflect aldehyde components in transgenics since major structures in CAD-downregulated angiosperms do not undergo the color reaction.; *G = guaiacyl, S = syringyl; S- G for example in sinapyl aldehyde 8-O-4-linked to a guaiacyl unit. |
