MALDI-MS Imaging of Urushiols in Poison Ivy Stem

Abstract

Urushiols are the allergenic components of Toxicodendron radicans (poison ivy) as well as other Toxicodendron species. They are alk-(en)-yl catechol derivatives with a 15- or 17-carbon side chain having different degrees of unsaturation. Although several methods have been developed for analysis of urushiols in plant tissues, the in situ localization of the different urushiol congeners has not been reported. Here, we report on the first analysis of urushiols in poison ivy stems by matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI). Our results show that the urushiol congeners with 15-carbon side chains are mainly localized to the resin ducts, while those with 17-carbon side chains are widely distributed in cortex and vascular tissues. The presence of these urushiols in stem extracts of poison ivy seedlings was confirmed by GC-MS. These novel findings provide new insights into the spatial tissue distribution of urushiols that might be biosynthetically or functionally relevant.

Keywords: MALDI-MSI; Toxicodendron radicans; in situ localization; poison ivy; urushiols.

Figure 1

Structures of poison ivy urushiol congeners.

Figure 2

GC-MS quantification of urushiols in poison ivy stem tissues extracts. Urushiols were extracted from both hypocotyls (A,B) and the first internodes (C,D) collected from four individual five-week-old plants using C15:0 alkylresorcinol as an internal standard and the different C15 and C17 urushiol congeners were quantified using GC-MS and specific fragments as described in Material and Methods. There is a considerable variability in the total urushiol content among the four individual plants, however the relative abundance of the different congeners is quite similar, where C15:1 and C15:2 are the most abundant among all C15 species, while C17:2 is the most abundant C17 congener.

Figure 3

Selected regions of MALDI-MS spectra containing urushiol sodium adducts. Identification of urushiol sodium adducts was made in MALDI-MS imaging datasets acquired for both poison ivy stem sections and the Japanese lacquer tree extract. Identified ions had very similar masses in both the sections and the extract with the mass difference between the theoretical and observed masses is less than 10 ppm for all selected ions. The MALDI-MSI data were collected in centroid and profile modes for the tissue sections and the Japanese lacquer tree extract, respectively. A broader mass range is depicted in supplementary Figures S1 and S2 for one of the Japanese lacquer tree extracts and one of the stem tissue sections, respectively, where the C15:1 urushiol congener was identified.

Figure 4

Spatial localization of urushiols in poison ivy hypocotyl sections. Sodium adduct of the C15:1 congener is localized to the resin ducts while that of the C17:3 is distributed predominantly in the cortex and vascular tissues of the hypocotyl. Distribution of the other congeners is not well-defined. Intensity of the selected ions was normalized to the total ion count before the images were generated.

Figure 5

Spatial localization of urushiols in poison ivy stem (first internode) sections. Sodium adducts of the C15:0, C15:1, and C15:2 congeners are localized to the resin ducts. In contrast, C17:2 and C17:3 congener sodium adducts are localized to the cortex and vascular tissues of the stem. Distribution of the other congeners is not well-defined. Intensity of the selected ions was normalized to the total ion count prior to image generation.

Figure 6

The C15 and C17 urushiol congeners have distinctly different localization within poison ivy hypocotyls and the first internodes. Combined images showing that the sodium adduct of the C15:1 congener (red) is mainly localized to the resin ducts, while that of the C17:3 congener (green) is more widely distributed in the cortex and vascular tissues in both the hypocotyl and the first internode regions of the stem. These images were generated by merging the individual images of the selected ions (m/z) presented in Figure 4 and Figure 5.