Figure 6.

Use of Thioacidolysis to Reveal Lignin Labeling in the ¹³C Monolignol Feeding Experiments.

(A) Labeling pattern, structures, and masses of monolignol fragments obtained after thioacidolysis from lignin extracted from tissues fed with labeled ¹³C₆-caffeyl alcohol or ¹³C₆-coniferyl alcohol. The thiol group (−SEt) displaces the α-hydroxyl, α-ether, and β-aryl groups during thioacidolysis. The bar indicates the cleavage at the C7-C8 bond during the MS analysis, resulting in the primary fragments at m/z 327 and 269 from the benzylic cation of the trimethylsilylated (TMS) derivatives of unlabeled monolignols and at m/z 333 and 275 of the TMS derivatives of ¹³C₆-labeled monolignols.

(B), (C), (F), and (G) EICs showing ¹³C-labeled and unlabeled monolignol thioacidolysis products after separation during GC. Double peaks in the EICs represent the presence of threo- and erythro-isomers of the tri-thioetherates shown in (A). EICs of transgenic Medicago hairy root line L6 are deployed here as an example. The no-monolignol–fed sample ([B] and [F]) showed a clear doublet corresponding to the m/z of the unlabeled thioacidolysis products, with no doublet at the m/z of the labeled products. Lignin from samples that were fed with ¹³C₆-caffeyl (C) or ¹³C₆-coniferyl (G) alcohol and could incorporate labeled precursors exhibited both M+6 and unlabeled monolignol-derived products.

(D), (E), (H), and (I) Traces showing the relative abundance of ¹³C-labeled and unlabeled monolignol thioacidolysis products after fragmentation during MS. Traces correspond to the monolignol thioacidolysis product peaks shown in (B), (C), (F), and (G). Samples that were fed with ¹³C₆-caffeyl (E) or ¹³C₆-coniferyl (I) alcohols showed an increased ratio of M+6 monolignol-derived fragmentation products to the unlabeled ions.