16 research outputs found
Etude de la spéciation chimique de la collection nationale de violettes et mise en place d'un agro-raffinage de la violette de Toulouse
Le projet « Viola Tolosa » a pour objectif de valoriser une plante produite en Occitanie, la violette et plus particuliĂšrement lâemblĂ©matique violette de Toulouse, pour des domaines essentiellement non alimentaires tels que la chimie des substances naturelles et la cosmĂ©tique. Les violettes appartiennent au genre Viola qui comprend plus de 500 espĂšces. Aujourdâhui, leurs usages sont principalement limitĂ©s Ă des aspects ornementaux et culinaires. NĂ©anmoins, lâintĂ©rĂȘt croissant de la part des acteurs de la filiĂšre (industriels, cultivateurs et acadĂ©miques) a conduit la rĂ©gion Occitanie Ă mettre en place le projet Viola Tolosa intitulĂ© « SpĂ©ciation chimique de la collection nationale des violettes et mise en place dâun agro-raffinage de la violette de Toulouse ». Il comporte quatre aspects interdisciplinaires associant aspects fondamentaux et applicatifs. La caractĂ©risation de la centaine de plants de la collection de violettes dĂ©tenue par les serres municipales de Toulouse, identifiĂ©e Ă 80% par des noms de cultivars ou vernaculaires, a Ă©tĂ© rĂ©alisĂ©e par lâintermĂ©diaire dâĂ©tudes gĂ©nĂ©tique et chimiotaxonomique. Une premiĂšre Ă©tude gĂ©nĂ©tique basĂ©e sur les sĂ©quences des espaces internes transcrits a permis de classer 58% de la collection au rang dâespĂšce. Cette Ă©tude phylogĂ©nĂ©tique a Ă©tĂ© complĂ©tĂ©e par une Ă©tude chimiotaxonomique Ă lâaide des profils chimiques des fractions volatiles des fleurs et non-volatiles des parties aĂ©riennes de la collection. Une projection orthogonale de structures latentes a permis dâindexer 96% de lâensemble des plants par un nom dâespĂšce. LâĂ©tude des mĂ©tabolites secondaires non volatils des feuilles a Ă©tĂ© entreprise dans le but dâĂ©tudier le potentiel biologique des violettes, notamment les activitĂ©s antioxydante, antifongique et inductrice des rĂ©ponses immunitaires des plantes. LâĂ©tude dĂ©taillĂ©e dâun extrait hydroalcoolique de la violette de Toulouse a permis dâidentifier huit composĂ©s antioxydants de la famille des flavonoĂŻdes et des coumarines, dont trois ont Ă©tĂ© caractĂ©risĂ©s par RMN 1D et 2D et deux de novo dĂ©rĂ©pliquĂ©s par rĂ©seau molĂ©culaire. Lâapplication sur lâensemble de la collection a ensuite permis dâidentifier six composĂ©s antioxydants, dont deux coumarines et quatre flavonoĂŻdes, prĂ©pondĂ©rants chez deux espĂšces. Une relation espĂšce-activitĂ© a donc Ă©tĂ© mise en Ă©vidence. Au niveau des activitĂ©s antifongiques, rĂ©alisĂ©es sur cinq souches de champignons, et de dĂ©fenses vĂ©gĂ©tales, par lâintermĂ©diaire de lâĂ©tude de lâexpression du gĂšne marqueur « pathogenesis-related protein 1 », les rĂ©sultats sont plus ambigus. Cependant, certaines espĂšces ont prĂ©sentĂ© une activitĂ© plus prononcĂ©e que les autres et ce criblage a permis de poser une hypothĂšse forte quant Ă lâimplication des cyclotides. Finalement, lâensemble de ces travaux a permis dâobtenir une carte dâidentitĂ© des violettes de la collection (identification gĂ©nĂ©tique, profil chimique, potentiel biologique) et une description semi-quantitative de lâensemble des groupes chimiques est proposĂ©e par combinaison des donnĂ©es chromatographiques du dĂ©tecteur Corona (CAD) et des donnĂ©es spectrales. DiffĂ©rentes mĂ©thodes dâextraction (Ă©lectroporation, micro-ondes, CO2 supercritique et extraction hydroalcoolique) rĂ©pondant aux prĂ©ceptes de la chimie verte ont ensuite Ă©tĂ© comparĂ©es afin de sĂ©lectionner celle prĂ©sentant le meilleur compromis entre le cahier des charges cosmĂ©tiques et lâenrichissement en molĂ©cules dâintĂ©rĂȘt, en vue dâun transfert technologique
Targeted Dereplication of Microbial Natural Products by High-Resolution MS and Predicted LC Retention Time
A new strategy for the identification of known compounds in Streptomyces extracts that can be applied in the discovery of natural products is presented. The strategy incorporates screening a database of 5555 natural products including 5098 structures from Streptomyces sp., using a high-throughput LCMS data processing algorithm that utilizes HRMS data and predicted LC retention times (tR) as filters for rapid identification of known compounds in the natural product extract. The database, named StrepDB, contains for each compound the structure, molecular formula, molecular mass, and predicted LC retention time. All identified compounds are annotated and color coded for easier visualization. It is an indirect approach to quickly assess masses (which are not annotated) that may potentially lead to the discovery of new or novel structures. In addition, a spectral database named MbcDB was generated using the ACD/Spectrus DB Platform. MbcDB contains 665 natural products, each with structure, experimental HRESIMS, MS/MS, UV, and NMR spectra. StrepDB was used to screen a mutant Streptomyces albus extract, which led to the identification and isolation of two new compounds, legonmaleimides A and B, the structures of which were elucidated with the aid of MbcDB and spectroscopic techniques. The structures were confirmed by computer-assisted structure elucidation (CASE) methods using ACD/Structure Elucidator Suite. The developed methodology suggests a pipeline approach to the dereplication of extracts and discovery of novel natural products
Dereplication of natural products from complex extracts by regression analysis and molecular networking: case study of redox-active compounds from Viola alba subsp. dehnhardtii
Introduction : In natural product research, bioassay-guided fractionation was previously widely employed but is now judged to be inadequate in terms of time and cost, particularly if only known compounds are ultimately isolated. The development of metabolomics, along with improvements in analytical tools, allows comprehensive metabolite profiling. This enables dereplication to target unknown active compounds early in the purification workflow.
Objectives :Starting from an ethanolic extract of violet leaves, this study aims to predict redox active compounds within a complex matrix through an untargeted metabolomics approach and correlation analysis.
Methods : Rapid fractionation of crude extracts was carried out followed by multivariate data analysis (MVA) of liquid chromatographyâhigh resolution mass spectrometry (LCâHRMS) profiles. In parallel, redox active properties were evaluated by the capacity of the molecules to reduce 2,2-diphenyl-1-picrylhydrazyl (DPPH·) and superoxide (O2 ·â) radicals using UVâVis and electron spin resonance spectroscopies (ESR), respectively. A spectral similarity network (molecular networking) was used to highlight clusters involved in the observed redox activities.
Results : Dereplication on Viola alba subsp. dehnhardtii highlighted a reproducible pool of redox active molecules. Polyphenols, particularly O-glycosylated coumarins and C-glycosylated flavonoids, were identified and de novo dereplicated through molecular networking. Confirmatory analyses were undertaken by thin layer chromatography (TLC)âDPPHâMS assays and nuclear magnetic resonance (NMR) spectra of the most active compounds.
Conclusion : Our dereplication strategy allowed the screening of leaf extracts to highlight new biologically active metabolites in few steps with a limited amount of crude material and reduced time-consuming manipulations. This approach could be applied to any kind of natural extract for the study of various biological activities
Study of the chemical speciation of the national violet collection and setting up of an agro-refining of the violet of Toulouse
Le projet « Viola Tolosa » a pour objectif de valoriser une plante produite en Occitanie, la violette et plus particuliĂšrement lâemblĂ©matique violette de Toulouse, pour des domaines essentiellement non alimentaires tels que la chimie des substances naturelles et la cosmĂ©tique. Les violettes appartiennent au genre Viola qui comprend plus de 500 espĂšces. Aujourdâhui, leurs usages sont principalement limitĂ©s Ă des aspects ornementaux et culinaires. NĂ©anmoins, lâintĂ©rĂȘt croissant de la part des acteurs de la filiĂšre (industriels, cultivateurs et acadĂ©miques) a conduit la rĂ©gion Occitanie Ă mettre en place le projet Viola Tolosa intitulĂ© « SpĂ©ciation chimique de la collection nationale des violettes et mise en place dâun agro-raffinage de la violette de Toulouse ». Il comporte quatre aspects interdisciplinaires associant aspects fondamentaux et applicatifs. La caractĂ©risation de la centaine de plants de la collection de violettes dĂ©tenue par les serres municipales de Toulouse, identifiĂ©e Ă 80% par des noms de cultivars ou vernaculaires, a Ă©tĂ© rĂ©alisĂ©e par lâintermĂ©diaire dâĂ©tudes gĂ©nĂ©tique et chimiotaxonomique. Une premiĂšre Ă©tude gĂ©nĂ©tique basĂ©e sur les sĂ©quences des espaces internes transcrits a permis de classer 58% de la collection au rang dâespĂšce. Cette Ă©tude phylogĂ©nĂ©tique a Ă©tĂ© complĂ©tĂ©e par une Ă©tude chimiotaxonomique Ă lâaide des profils chimiques des fractions volatiles des fleurs et non-volatiles des parties aĂ©riennes de la collection. Une projection orthogonale de structures latentes a permis dâindexer 96% de lâensemble des plants par un nom dâespĂšce. LâĂ©tude des mĂ©tabolites secondaires non volatils des feuilles a Ă©tĂ© entreprise dans le but dâĂ©tudier le potentiel biologique des violettes, notamment les activitĂ©s antioxydante, antifongique et inductrice des rĂ©ponses immunitaires des plantes. LâĂ©tude dĂ©taillĂ©e dâun extrait hydroalcoolique de la violette de Toulouse a permis dâidentifier huit composĂ©s antioxydants de la famille des flavonoĂŻdes et des coumarines, dont trois ont Ă©tĂ© caractĂ©risĂ©s par RMN 1D et 2D et deux de novo dĂ©rĂ©pliquĂ©s par rĂ©seau molĂ©culaire. Lâapplication sur lâensemble de la collection a ensuite permis dâidentifier six composĂ©s antioxydants, dont deux coumarines et quatre flavonoĂŻdes, prĂ©pondĂ©rants chez deux espĂšces. Une relation espĂšce-activitĂ© a donc Ă©tĂ© mise en Ă©vidence. Au niveau des activitĂ©s antifongiques, rĂ©alisĂ©es sur cinq souches de champignons, et de dĂ©fenses vĂ©gĂ©tales, par lâintermĂ©diaire de lâĂ©tude de lâexpression du gĂšne marqueur « pathogenesis-related protein 1 », les rĂ©sultats sont plus ambigus. Cependant, certaines espĂšces ont prĂ©sentĂ© une activitĂ© plus prononcĂ©e que les autres et ce criblage a permis de poser une hypothĂšse forte quant Ă lâimplication des cyclotides. Finalement, lâensemble de ces travaux a permis dâobtenir une carte dâidentitĂ© des violettes de la collection (identification gĂ©nĂ©tique, profil chimique, potentiel biologique) et une description semi-quantitative de lâensemble des groupes chimiques est proposĂ©e par combinaison des donnĂ©es chromatographiques du dĂ©tecteur Corona (CAD) et des donnĂ©es spectrales. DiffĂ©rentes mĂ©thodes dâextraction (Ă©lectroporation, micro-ondes, CO2 supercritique et extraction hydroalcoolique) rĂ©pondant aux prĂ©ceptes de la chimie verte ont ensuite Ă©tĂ© comparĂ©es afin de sĂ©lectionner celle prĂ©sentant le meilleur compromis entre le cahier des charges cosmĂ©tiques et lâenrichissement en molĂ©cules dâintĂ©rĂȘt, en vue dâun transfert technologique.The "Viola Tolosa" project aims to promote a plant produced in Occitanie region, the violet and especially the emblematic violet of Toulouse, essentially for non-food fields such as the chemistry of natural compounds and cosmetics. Violets belong to Viola genus including more than 500 species. Today, their uses are mainly limited to ornamental and culinary aspects. Nevertheless, the growing interest of the actors of the sector (industrials, growers and academicals) led the Occitanie region to implement the Viola Tolosa project entitled "Chemical speciation of the national collection of violets et establishment of an agro-refining of the violet of Toulouse ". It comprises four interdisciplinary aspects associating fundamental and applicative aspects. The characterization of the 100 or so plants in the violet collection owned by the Toulouse municipal greenhouses, including 80% identified by cultivar or vernacular names, was carried out through genetic and chemotaxonomic studies. A first genetic study based on internal transcribed spacers conducted to classify 58% of the collection as a species. This phylogenetic study was completed by chemotaxonomic studies of chemical profiles of flowers volatile fractions and non-volatile aerial parts of the collection. Discriminant analysis of orthogonal projection to latent structure model finally allowed indexation of 96% of all plants with a species name. Study of non-volatile secondary metabolites of leaves has also been undertaken to study the biological potential of violets, including antioxidant, antifungal and defense inducer. The detailed study of a hydroalcoholic extract of the violet of Toulouse allowed the identification of eight antioxidant compounds belonging to flavonoids and coumarins. Three of them have been characterized by 1D and 2D NMR and two were de novo dereplicated through molecular network. The application to the whole collection conducted to highlight six antioxidant compounds, including two coumarins and four flavonoids, predominant in two species. A species-activity relationship was therefore highlighted. Regarding antifungal activities carried out on five fungal strains, and defense inducer through the study of pathogenesis-related protein 1, the results are more ambiguous. However, some species showed better activity than others and this screening led to a strong hypothesis regarding the involvement of cyclotides. Finally, all this work led to the establishment of an identity card of violets of the collection (genetic identification, chemical profiling, et biological potential) et a semi-quantitative description of all the species is considered by combining chromatographic data based on corona detector et spectral data. Different methods of extraction (electroporation, microwaves, supercritical CO2 et hydroalcoholic extraction) corresponding to green chemistry precepts were then compared in order to select the one presenting the best compromise between cosmetic specifications et enrichment in molecules of interest, for technological transfer
Comparative Transcriptomics Suggests Early Modifications by VintecÂź in Grapevine Trunk of Hormonal Signaling and Secondary Metabolism Biosynthesis in Response to Phaeomoniella chlamydospora and Phaeoacremonium minimum
International audienceGiven their well-known antifungal abilities, species of the genus Trichoderma are of significant interest in modern agriculture. Recent studies have shown that Trichoderma species can induce plant resistance against different phytopathogens. To further extend this line of investigation, we investigate herein the transcriptomic response of grapevine trunk to Vintec R , which is a Trichoderma atroviride SC1-based commercial formulation for biological control of grapevine trunk diseases and which reduces wood colonization. The aim of the study is to understand whether the biocontrol agent Vintec R modifies the trunk response to Phaeoacremonium minimum and Phaeomoniella chlamydospora, which are two esca-associated fungal pathogens. An analysis of transcriptional regulation identifies clusters of co-regulated genes whose transcriptomic reprogramming in response to infection depends on the absence or presence of Vintec R. On one hand, the results show that Vintec R differentially modulates the expression of putative genes involved in hormonal signaling, especially those involved in auxin signaling. On the other hand, most significant gene expression modifications occur among secondary-metabolism-related genes, especially regarding phenylpropanoid metabolism and stilbene biosynthesis. Taken together, these results suggest that the biocontrol agent Vintec R induces wood responses that counteract disease development
Chemotaxonomy vs Genotaxonomy of Viola genus: Can native volatile compounds of flowers be genuine tracers for violets differentiation?
International audienceViolets are odorous flowers known in the Mediterranean basin since Antiquity and were a part of the first perfumes created. Toulouse has an historical relationship with this flower which is the symbol of the town and the home of the famous Viola Tolosa, Parma of Toulouse variety (Viola alba subsp. dehnhardtii). As a part of the regional Viola Tolosa project, a chemotaxonomy study of the hundred varieties/cultivars of violets belonging to the French Violet Conservatory hosted by the Toulouse Municipal Greenhouses will be set up. Based on headspace analysis of native volatile compounds emitted by flowers at maturity, dual chemical-sensory fingerprints were established for a rapid and efficient differentiation. A HS-SPME-GC/O method was used on two types of volatile samplings: an in-vivo and non-destructive analysis on the whole plant with the use of a bell-shaped glass covering to trap the volatiles as well as an ex-vivo analysis on the fresh cut flowers placed in a vial. First, experimental analytical conditions were optimized by experimental design methodology in order to determine the best suited protocols to obtain the richest volatile cocktails. Then optimized methods were applied to a sampling of ten plants of the collection selected according to the colour of flowers and their geographical origins. In parallel, genetic analyses were conducted on the same plants to confirm the putative varieties or cultivars. Finally, all the results were combined to match the chromatographic profiles with the genetic affiliation to a group. Acknowledgements: The Regional Council Languedoc-Roussillon Midi-Pyrénées (Project CLE ) for sponsoring 1
Opportunities and challenges in the adoption of new grape varieties by producers: a case study from the Northeastern United States
National audienc
Modification of Early Response of Vitis vinifera to Pathogens Relating to Esca Disease and Biocontrol Agent VintecÂź Revealed By Untargeted Metabolomics on Woody Tissues
International audienceEsca disease is one of the most destructive grapevine trunk diseases. Phaeoacremonium minimum and Phaeomoniella chlamydospora are two of the known fungal pathogens associated with this disease. Today, biocontrol agents against Esca are mainly based on the use of the strain of the mycoparasite fungal genus Trichoderma such as the Vintec Âź product. The aim of this study was to investigate early response of woody tissues to Esca pathogens and identify metabolites that could be correlated with a biocontrol activity within a complex woody matrix. An untargeted liquid chromatographyâhigh-resolution mass spectrometry metabolomic approach coupled to a spectral similarity network was used to highlight clusters of compounds associated with the plant response to pathogens and biocontrol. Dereplication highlighted the possible role of glycerophospholipids and polyphenol compounds, the latest mainly belonging to stilbenoids. Antifungal activity of some relevant biomarkers, evaluated in vitro on Phaeomoniella chlamydospora and Botrytis cinerea , suggests that some of these compounds can play a role to limit the development of Esca pathogens in planta
Phenolic profile of a Parma violet unveiled by chemical and fluorescence imaging
International audienceThe ability of phenolic compounds to autofluoresce upon illumination by UV or blue light was exploited to explore the nature and distribution of these metabolites within the flower petals, leaves and roots of the violet, Viola alba subsp. dehnhardtii. This was achieved through a dual complementary approach that combined fluorescence microscopy imaging of living intact tissues and chemical extraction of pulverized material. The blue to red fluorescence displayed by living tissues upon illumination was indicative of their richness in phenolic compounds. Phenolic acids were found in all tissues, while flavonoids characterized the aerial part of the plant, anthocyanidins being restricted to the petals. The chemical quantification of phenolics in plant extracts confirmed their tissue-specific distribution and abundance. A key finding was that the spectral signatures obtained through confocal microscopy of endogenous fluorophores in living tissues and their counterpart extracts share the same fluorescence patterns, pointing out the potential of fluorescence imaging of intact organs for a proper estimation of their phenolic content. In addition, this study highlighted a few distinct morphology cell types, in particular foliar-glandular-like structures, and jagged petal cell walls. Altogether, these data provide a comprehensive histochemical localization of phenolics in living tissues of a violet. Converting fluorescence imaging into a chemical imprint indicated that one can rely on fluorescence microscopy of intact living tissues as a rapid, non-destructive means to follow their phenolic imprint under various environmental conditions
Développement d'une approche métabolomique appliquée à  l'identification rapide de molécules bioactives: Etude de cas de la violette de Toulouse (Viola alba subsp. Dehnhardtii)
International audienc