LC/MS profiling of flavonoid glycoconjugates isolated from hairy roots, suspension root cell cultures and seedling roots of Medicago truncatula

Abstract

Hairy roots and suspension cell cultures are commonly used in deciphering different problems related to the biochemistry and physiology of plant secondary metabolites. Here, we address about the issue of possible differences in the profiles of flavonoid compounds and their glycoconjugates derived from various plant materials grown in a standard culture media. We compared profiles of flavonoids isolated from seedling roots, hairy roots, and suspension root cell cultures of a model legume plant, Medicago truncatula. The analyses were conducted with plant isolates as well as the media. The LC/MS profiles of target natural products obtained from M. truncatula seedling roots, hairy roots, and suspension root cell cultures differed substantially. The most abundant compounds in seedlings roots were mono- and diglucuronides of isoflavones and/or flavones. This type of glycosylation was not observed in hairy roots or suspension root cell cultures. The only recognized glycoconjugates in the latter samples were glucose derivatives of isoflavones. Application of a high-resolution mass spectrometer helped evaluate the elemental composition of protonated molecules, such as [M + H]+. Comparison of collision-induced dissociation MS/MS spectra registered with a quadrupole time-of-flight analyzer for tissue extracts and standards allowed us to estimate the aglycone structure on the basis of the pseudo-MS3 experiment. Structures of these natural products were described according to the registered mass spectra and literature data. The analyses conducted represent an overview of flavonoids and their conjugates in different types of plant material representing the model legume, M. truncatula.Electronic supplementary materialThe online version of this article (doi:10.1007/s11306-011-0287-2) contains supplementary material, which is available to authorized users.

Expression Analysis of Phenylalanine Ammonia Lyase Gene and Rosmarinic Acid Production in Salvia officinalis and Salvia virgata Shoots Under Salicylic Acid Elicitation

“…However, their result was inconsistent with the results of Dong et al, who reported that the SA-induced phenolic compound accumulation was correlated to the PAL activity in S. miltiorrhiza cell cultures [19]. It was claimed that this inconsistency was apparently due to differences in plant materials used in the researches, as the secondary metabolite profiles are varied among the different plant material types of the same plant species [34].…”

Mass spectrometric profiling of flavonoid glycoconjugates possessing isomeric aglycones

“…Therefore, numerous innovative MS‐based methods have been developed for profiling flavonoid glycoconjugates. A widely used approach relies on the evaluation of characteristic neutral losses occurring as a result of sequential cleavage of saccharide subunits from the intact flavonoid glycoconjugate . In addition to these so‐called top‐down profiling methods, bottom‐up and MRM‐based HPLC‐MS methods have also been recently reported for the screening of flavonoids in plants.…”

“…According to a recent study from the Stobiecki group, difficulties with the interpretation of registered data in HPLC‐MS analyses were summarized as follows: (1) the presence of compounds with the same nominal molecular weight and different elemental compositions, which appear as isobars in low‐resolution MS and (2) the isomers of natural products with different substitutions of sugar moieties, various patterns of acylating groups on the sugar moieties, and isomerization of the aglycones (e.g. flavones and isoflavones) . The problem of isobaric compounds with different elemental formulas can be handled via the enhanced selectivity of high‐resolution mass spectrometers .…”

“…A widely used approach relies on the evaluation of characteristic neutral losses occurring as a result of sequential cleavage of saccharide subunits from the intact flavonoid glycoconjugate. [13,[19][20][21] In addition to these so-called top-down profiling methods, bottom-up [22,23] and MRM-based [24][25][26] HPLC-MS methods have also been recently reported for the screening of flavonoids in plants. These published papers mainly presented examples of structural MS analyses focusing on the elucidation of the type of glycan, the substitution pattern of the glycan(s) on an aglycone or the placement of an interglycosidic bond.…”

Metabolomics in Plants and Humans: Applications in the Prevention and Diagnosis of Diseases

“…LC-MS is more suitable than GC-MS for labile compounds as well as for those that are difficult to derivatize. The application of LC-MS in plant metabolomics includes the description of the tomato metabolome database (MoTo DB) [79], the identification of the accumulation of oxylipins after wound-induced stress in Arabidopsis [80], the identification of flavonoid and isoflavonoid compounds in Medicago truncatula [81], the untargeted large-scale analysis of plant metabolome [82], the characterization of metabolites that respond to stress [83, 84], and the identification of flavonoid glycoconjugates in roots form Medicago truncaluta [85]. …”