The aim of this study was to present integrated mass spectrometric methods for the structural characterization and identification of flavonoid glycoconjugates. During the liquid chromatography/mass spectrometry analyses, TriVersa NanoMate chip-based system with nanoelectrospray ionization and fraction collection was combined to a quadrupole time-of-flight mass spectrometer. In the extract samples prepared from green leaves of wheat plantlets, 41 flavonoid derivatives were recognized. Part of the target natural products had the full structure being characterized after the registration of mass spectra, where m/z values for protonated [M + H](+) and deprotonated molecules [M - H](-) were annotated. MS(2) and pseudo-MS(3) experiments were performed for [M + H](+) or [M - H](-) and aglycone ions (Y0(+/-)-type), respectively. It should be underlined that pseudo-MS(3) mass spectra were registered for aglycone product ions in the mass spectra of O-glycosides present in the extract samples. In many cases, only tentative structural identification of aglycones was possible, mainly because of the presence of numerous C-monoglycoside or C-diglycoside in the samples. Acylation of the sugar moiety and/or methylation of the aglycone in the flavonoid glycosides under study was observed. The existence of isobaric and/or isomeric compounds was demonstrated in the extract studied. The collision-induced dissociation mass spectra registered for C,O-diglycosides and C,C-diglycosides did not permit to draw complete structural conclusions about the compounds studied. For the investigated class of natural products, unambiguous classification of sugar moieties linked to the aglycones from the recorded mass spectra was not possible. Registration of the positive and negative ion mass spectra did not lead to the precise conclusion about the glycosylation position at C-6 or C-8, and O-4' or O-7 atoms. It was possible, on the basis of the collected MS(2) spectra, to differentiate between O-glycosides and C-glycosides present in the samples analyzed.

To date, studies on volatile metabolites in cereal grain have focused mainly on a single species. In this paper, results are presented of the analysis of volatile compounds in five cereal grain species (spring wheat, durum wheat, triticale, rye, oats and barley) based on representative sampling of at least 15 cultivars of individual species. Profiles of volatile compounds were determined using solid phase microextraction (SPME) and GC-TOF (time of flight mass spectrometry). Many of the volatile compounds were only present in single samples; however, several dozen were found in over 50% of samples and 46 volatiles were found in all samples. Among them there were six alcohols, 10 aldehydes and ketones, six terpenes, seven hydrocarbons and 11 benzene derivatives. The highest concentrations of these compounds were found in durum wheat, while the lowest were observed in triticale and rye.

Ergosterol (ERG) is a major sterol constituent of most fungi. Its concentration is negligible in higher plants, but can be used as a chemical marker of the presence of fungal contaminations. In this study, ERG concentration was assessed in randomly collected samples of naturally contaminated grain (wheat, barley and oat) and in samples of grain (wheat, barley, triticale and oat) harvested after inoculation of heads with conidia of different Fusarium species. Wheat samples were analysed at three stages of grain development. The lowest ERG concentration was found in non-inoculated samples at the first stage of grain development. This concentration was increasing with grain ripening. In naturally contaminated samples collected after harvest, ERG concentration was lower in wheat than in barley and oat. ERG concentrations in inoculated samples varied significantly, but were always significantly higher than in naturally contaminated samples. In the above cereal samples it was much lower than the levels assayed in laboratory cultures inoculated with fungi from genus Fusarium. The content of ERG was also analyzed in milling products of small-grained cereals and other foodstuffs, where a considerable variation was observed. The lowest ERG amounts were assayed in flours with a high degree of purification, while the highest ones in case of flours and products with a low purification rate. The results indicate the potential application of HPLC combined with microwave-assisted extraction both when assaying samples with low ERG concentrations (naturally contaminated) and those characterized with high contents of fungal biomass (strongly infected, artificially inoculated). It also facilitates analyses of fungal biomass in technological processes, where results may be expected to vary considerably.

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Plant-derived compounds limiting mycotoxin contamination are currently of major interest in food and feed production. However, their potential application requires an evaluation of their effects on fungal secondary metabolism and membrane effects. In this study, different strains of Fusarium culmorum and F. graminearum sensu stricto were exposed to trans-cinnamic and chlorogenic acids on solid YES media. Fusaria produced phenolic acids, whose accumulation was lowered by exogenous phenolic compounds. In addition, fungi reduced exogenous phenolic acids, leading either to their conversion or degradation. trans-Cinnamic acid was converted to caffeic and ferulic acids, while chlorogenic acid was degraded to caffeic acid. The latter underwent further degradation to protocatechuic acid. Fungal-derived trans-cinnamic acid, as the first intermediate of the shikimate pathway, increased after chlorogenic acid treatment, presumably due to the further inhibition of the conversion of trans-cinnamic acid. Exogenous trans-cinnamic and chlorogenic acid displayed the inhibition of mycotoxin production by Fusaria, which appeared to be largely dependent on the phenolic compound and its concentration and the assayed strain. Exogenous phenolic acids showed different effects on ergosterol biosynthesis by fungi. It was found that the production of this membrane sterol was stimulated by trans-cinnamic acid, while chlorogenic acid negatively impacted ergosterol biosynthesis, suggesting that phenolic acids with stronger antifungal activities may upregulate ergosterol biosynthesis by Fusaria. This paper reports on the production of phenolic acids by Fusaria for the first time.