Comparative structural profiling of trichome specialized metabolites in tomato (Solanumlycopersicum) and S. habrochaites: acylsugar profiles revealed by UHPLC/MS and NMR.

Banibrata Ghosh,Thomas C. Westbrook,A. Daniel Jones

doi:10.1007/s11306-013-0585-y

Abstract

Many plants accumulate large quantities of specialized metabolites in secretory glandular trichomes (SGTs), which are specialized epidermal cells. In the genus Solanum, SGTs store a diverse collection of glucose and sucrose esters. Profiling of extracts from two accessions (LA1777 and LA1392) of Solanum habrochaites using ultra-high performance liquid chromatography–mass spectrometry (UHPLC/MS) revealed wide acylsugar diversity, with up to 11 isomers annotated for each individual elemental formula. These isomers arise from differences in ester chain lengths and their positions of substitution or branching. Since fragment ion masses were not sufficient to distinguish all isomers, 24 acylsucroses were purified from S. habrochaites accessions and cultivated tomato (Solanum lycopersicum M82) and characterized using NMR spectroscopy. Two-dimensional NMR spectra yielded assignments of positions of substitution of specific acyl groups, and locations of branching. The range of substitution was wider than reported earlier, and in contrast to previous reports, tetra- and penta-acylsucroses were substituted at position 2 with acyl groups other than acetate. Because UHPLC/MS fails to yield sufficient information about structure diversity, and quantitative NMR of acylsugar mixtures is confounded by structural redundancy, the strategic combination of NMR and UHPLC/MS provides a powerful approach for profiling a class of metabolites with great structural diversity across genotypes.Electronic supplementary materialThe online version of this article (doi:10.1007/s11306-013-0585-y) contains supplementary material, which is available to authorized users.

Highlights

The plant kingdom is estimated to synthesize a phenomenally diverse suite of bioactive specialized metabolites (Crozier et al 2006), many of which serve as bioactive chemical defenses and attractants for pollinators (Waterman 1992)
Mass spectra alone have not distinguished isomers differing in positions of acyl substitution or in acyl chain branching. To resolve such structural ambiguities, we demonstrate a combination of ultra-high performance liquid chromatography–mass spectrometry (UHPLC/MS) and NMR for comparative structural profiling of acylsugar metabolites from leaves from Solanum habrochaites accessions LA1777 and LA1392 and cultivated tomato Solanum lycopersicum M82 (LA3475)
This is consistent with the fatty acid synthase (FAS)-mediated acyl elongation pathway where branched even-carbon acids are synthesized from valine, straight chain acids are formed via normal fatty acid biosynthesis mechanisms, and odd-carbon acids by elongation of leucine or isoleucine, all with the increment of two carbons per elongation cycle (Kroumova and Wagner 2003; van der Hoeven and Steffens 2000). This investigation has documented far greater complexity and diversity in acylsucrose metabolite structures than was apparent in earlier reports, yet acylation patterns exhibit a remarkable selectivity that differentiates genotypes. These advances have relied in large part on the powerful combination of modern UHPLC/MS and high field NMR spectroscopy

Summary

Introduction

The plant kingdom is estimated to synthesize a phenomenally diverse suite of bioactive specialized metabolites (Crozier et al 2006), many of which serve as bioactive chemical defenses and attractants for pollinators (Waterman 1992). In the plant family Solanaceae, which encompasses tomato, potato, peppers, tobacco, nightshade, and petunia, specialized acylsugar metabolites accumulate in specific epidermal cells known as secretory glandular trichomes (SGTs) (McDowell et al 2011; Schilmiller et al 2008; Wagner 1991). SGTs are hair-like epidermal outgrowths, normally found on plant leaves and stems, and their accessibility facilitates investigation of gene expression and chemistry of individual cell types. These metabolites are believed to have important biological functions including plants’ self-defense against insects and herbivores (Stipanovic 1983; Weinhold and Baldwin 2011). In Solanum tissues, known acylsugars are based on glucose or sucrose cores esterified to 2–5 straight or branched chain aliphatic acids each with 2–12 carbon atoms (Arrendale et al 1990; Ghosh et al 2011; King et al 1990, 1993)

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