Corrigendum: Quantification and Localization of Formylated Phloroglucinol Compounds (FPCs) in Eucalyptus Species.

[This corrects the article DOI: 10.3389/fpls.2019.00186.].

Quantification and Localization of Formylated Phloroglucinol Compounds (FPCs) in Eucalyptus Species.

The Eucalyptus genus is a hyper-diverse group of long-lived trees from the Myrtaceae family, consisting of more than 700 species. Eucalyptus are widely distributed across their native Australian landscape and are the most widely planted hardwood forest trees in the world. The ecological and economic success of Eucalyptus trees is due, in part, to their ability to produce a plethora of specialized metabolites, which moderate abiotic and biotic interactions. Formylated phloroglucinol compounds (FPCs) are an important class of specialized metabolites in the Myrtaceae family, particularly abundant in Eucalyptus. FPCs are mono- to tetra-formylated phloroglucinol based derivatives, often with an attached terpene moiety. These compounds provide chemical defense against herbivory and display various bioactivities of pharmaceutical relevance. Despite their ecological and economic importance, and continued improvements into analytical techniques, FPCs have proved challenging to study. Here we present a simple and reliable method for FPCs extraction, identification and quantification by UHPLC-DAD-ESI-Q-TOF-MS/MS. The method was applied to leaf, flower bud, and flower samples of nine different eucalypt species, using a small amount of plant material. Authentic analytical standards were used to provide high resolution mass spectra and fragmentation patterns. A robust method provides opportunities for future investigations into the identification and quantification of FPCs in complex biological samples with high confidence. Furthermore, we present for the first time the tissue-based localization of FPCs in stem, leaf, and flower bud of Eucalyptus species measured by mass spectrometry imaging, providing important information for biosynthetic pathway discovery studies and for understanding the role of those compounds in planta.

Ozone and Wounding Stresses Differently Alter the Temporal Variation in Formylated Phloroglucinols in Eucalyptus globulus Leaves.

Formylated phloroglucinol compounds (FPCs) are a class of plant specialized metabolite present in the Myrtaceae family, especially in the genus Eucalyptus. FPCs are widely investigated due to their herbivore deterrence properties and various bioactivities of pharmaceutical relevance. Despite the increasing number of studies elucidating new FPCs structures and bioactivity, little is known about the role of those compounds in planta, and the effects of environmental stresses on FPC concentration. Ozone (O3) and wounding are key stress factors regularly confronted by plants. In this study, we investigated how O3, wounding, and their combination affected individual and total FPC foliar concentration of the economically important species Eucalyptus globulus. Six individual FPCs, including five macrocarpals and one sideroxylonal, showed different response patterns to the single and combined stresses. Total macrocarpals only increased under single O3 treatment, whereas total sideroxylonals only increased in response to wounding treatment, suggesting different physiological roles played by the two groups of FPCs predominantly existing in E. globulus foliage. Total FPCs increased significantly under individual wounding and O3 treatments but not under the combined treatment. A principal component analysis indicated that all different treatments had unique FPC fingerprints. Total phenolic contents increased in all O3 and wounding treatments, and a marginally positive correlation was found between total FPCs and total phenolic contents. We suggest that, depending on the concentration and composition, FPCs play multiple physiological roles in planta, including serving as antioxidants to scavenge the reactive oxygen species brought about by O3 and wounding stresses.

Metabolomics approaches for the discrimination of disease suppressive soils for Rhizoctonia solani AG8 in cereal crops using 1H NMR and LC-MS.

The suppression of soilborne crop pathogens such as Rhizoctonia solani AG8 may offer a sustainable and enduring method for disease control, though soils with these properties are difficult to identify. In this study, we analysed the soil metabolic profiles of suppressive and non-suppressive soils over 2 years of cereal production. We collected bulk and rhizosphere soil at different cropping stages and subjected soil extracts to liquid chromatography-mass spectrometry (LC-MS) and proton nuclear magnetic resonance spectroscopy (1H NMR) analyses. Community analyses of suppressive and non-suppressive soils using principal component analyses and predictive modelling of LC-MS and NMR datasets respectively, revealed distinct biochemical profiles for the two soil types with clustering based on suppressiveness and cropping stage. NMR spectra revealed the suppressive soils to be more abundant in sugar molecules than non-suppressive soils, which were more abundant in lipids and terpenes. LC-MS features that were significantly more abundant in the suppressive soil were identified and assessed as potential biomarkers for disease suppression. The structures of a potential class of LC-MS biomarkers were elucidated using accurate mass data and MS fragmentation spectrum information. The most abundant compound found in association with suppressive soils was confirmed to be a macrocarpal, which is an antimicrobial secondary metabolite. Our study has demonstrated the utility of environmental metabolomics for the study of disease suppressive soils, resulting in the discovery of a macrocarpal biomarker for R. solani AG8 suppressive soil which can be further studied functionally in association with suppression pot trials and microbial isolation studies.

Macrocarpal C isolated from Eucalyptus globulus inhibits dipeptidyl peptidase 4 in an aggregated form.

Dipeptidyl peptidase 4 (DPP-4) inhibitors are used for the treatment of type-2 diabetes mellitus. Various synthetic inhibitors have been developed to date, and plants containing natural DPP-4 inhibitors have also been identified. Here, 13 plant samples were tested for their DPP-4 inhibitory activity. Macrocarpals A-C were isolated from Eucalyptus globulus through activity-guided fractionation and shown to be DPP-4 inhibitors. Of these, macrocarpal C showed the highest inhibitory activity, demonstrating an inhibition curve characterised by a pronounced increase in activity within a narrow concentration range. Evaluation of macrocarpal C solution by turbidity, nuclear magnetic resonance spectroscopy and mass spectrometry indicated its aggregation, which may explain the characteristics of the inhibition curve. These findings will be valuable for further study of potential small molecule DPP-4 inhibitors.

A water availability gradient reveals the deficit level required to affect traits in potted juvenile Eucalyptus globulus.

Background and aims: Drought leading to soil water deficit can have severe impacts on plants. Water deficit may lead to plant water stress and affect growth and chemical traits. Plant secondary metabolite (PSM) responses to water deficit vary between compounds and studies, with inconsistent reports of changes to PSM concentrations even within a single species. This disparity may result from experimental water deficit variation among studies, and so multiple water deficit treatments are used to fully assess PSM responses in a single species. Methods: Juvenile Eucalyptus globulus were grown for 8 weeks at one of ten water deficit levels based on evapotranspiration from control plants (100 %). Treatments ranged from 90 % of control evapotranspiration (mild water deficit) to 0 % of control evapotranspiration (severe water deficit) in 10 % steps. Plant biomass, foliar abscisic acid (ABA) levels, Ψ leaf , leaf C/N, selected terpenes and phenolics were quantified to assess responses to each level of water deficit relative to a control. Key Results: Withholding ≥30 % water resulted in higher foliar ABA levels and withholding ≥40 % water reduced leaf water content. Ψ leaf became more negative when ≥60 % water was withheld. Plant biomass was lower when ≥80 % water was withheld, and no water for 8 weeks (0 % water) resulted in plant death. The total oil concentration was lower and C/N was higher in dead and desiccated juvenile E. globulus leaves (0 % water). Concentrations of individual phenolic and terpene compounds, along with condensed tannin and total phenolic concentrations, remained stable regardless of water deficit or plant stress level. Conclusions: These juvenile E. globulus became stressed with a moderate reduction in available water, and yet the persistent concentrations of most PSMs in highly stressed or dead plants suggests no PSM re-metabolization and continued ecological roles of foliar PSMs during drought.