18 research outputs found
Isolation of nuclear proteins from flax (Linum usitatissimum L.) seed coats for gene expression regulation studies
<p>Abstract</p> <p>Background</p> <p>While seed biology is well characterized and numerous studies have focused on this subject over the past years, the regulation of seed coat development and metabolism is for the most part still non-elucidated. It is well known that the seed coat has an essential role in seed development and its features are associated with important agronomical traits. It also constitutes a rich source of valuable compounds such as pharmaceuticals. Most of the cell genetic material is contained in the nucleus; therefore nuclear proteins constitute a major actor for gene expression regulation. Isolation of nuclear proteins responsible for specific seed coat expression is an important prerequisite for understanding seed coat metabolism and development. The extraction of nuclear proteins may be problematic due to the presence of specific components that can interfere with the extraction process. The seed coat is a rich source of mucilage and phenolics, which are good examples of these hindering compounds.</p> <p>Findings</p> <p>In the present study, we propose an optimized nuclear protein extraction protocol able to provide nuclear proteins from flax seed coat without contaminants and sufficient yield and quality for their use in transcriptional gene expression regulation by gel shift experiments.</p> <p>Conclusions</p> <p>Routinely, around 250 ÎŒg of nuclear proteins per gram of fresh weight were extracted from immature flax seed coats. The isolation protocol described hereafter may serve as an effective tool for gene expression regulation and seed coat-focused proteomics studies.</p
Nettle (<i>Urtica dioica</i> L.) as a source of antioxidant and anti-aging phytochemicals for cosmetic applications
Transcriptional regulation of lignan biosynthesis in flax (Linum usitatissimum L.)
Plante de grande culture aux multiples applications, le lin accumule dans ses graines des mĂ©tabolites spĂ©cialisĂ©s appelĂ©s lignanes aux propriĂ©tĂ©s phytooestrogĂšne et antioxydante, bĂ©nĂ©fiques en santĂ© humaine et, dans ses feuilles, des lignanes de nature toxique. Pour lâutilisation et lâĂ©tude de ces composĂ©s phĂ©noliques issus de la voie des monolignols, une technique dâĂ©co-extraction assistĂ©e par les ultrasons a Ă©tĂ© dĂ©veloppĂ©e et appliquĂ©e Ă un criblage de diffĂ©rentes variĂ©tĂ©s pour leur contenu en lignanes confĂ©rant une haute valeur ajoutĂ©e aux extraits et produits. Les potentiels verrous mĂ©taboliques de la voie de biosynthĂšse de ces composĂ©s ont fait lâobjet dâinvestigations au niveau de la rĂ©gulation transcriptionnelle des gĂšnes responsables de la formation des Ă©nantiomĂšres opposĂ©s de sĂ©coisolaricirĂ©sinol dans la graine et les parties aĂ©riennes. La famille multigĂ©nique des protĂ©ines dirigeantes (DIR) du lin, premiers acteurs de cette voie, a Ă©tĂ© criblĂ©e au niveau gĂ©nomique et transcriptionnel afin de dĂ©terminer des candidates pour fonctionner avec les pinorĂ©sinol laricirĂ©sinol rĂ©ductases (PLR), enzymes bifonctionnelles catalysant la formation du sĂ©coisolaricirĂ©sinol. LâĂ©tude de la rĂ©gulation fine de lâexpression des deux isoformes des gĂšnes PLR a mis en Ă©vidence des profils dâinductibilitĂ© trĂšs contrastĂ©s et a conduit Ă lâidentification dâĂ©lĂ©ments cis-rĂ©gulateurs ainsi que de facteurs de transcription impliquĂ©s dans ces voies de rĂ©gulation. Lâensemble des rĂ©sultats convergent vers un rĂŽle de dĂ©fense des lignanes in planta et a permis de construire une vue dâensemble des mĂ©canismes complexes de rĂ©gulation de leur biosynthĂšse et enfin de proposer des pistes pour lâamĂ©lioration de la production de ces molĂ©cules naturelles par une stimulation de leur accumulation et lâaugmentation des rendements dâextraction par chimie verte.As a crop with multiple purposes, flax accumulates lignans, specialized metabolites with health benefits, known for their phytoestrogenic and antioxidant properties in its seed, and toxic lignans in its leaves. In order to use and study these phenolic compounds derived from monolignols, an eco-friendly method based on ultrasound-assisted extraction was developed and applied to the screening of cultivars for their lignan content which confers high value to extracts and flaxseed by-product. Regulation of the lignan biosynthetic pathway was investigated at the transcriptional level for the genes responsible for the formation of secoisolariciresinol in seeds and aerial parts. The multigene family of dirigent proteins (DIR), first actors of this pathway, was explored by genomic and transcriptional approaches in order to select candidates to operate with pinoresinol lariciresinol reductases (PLR), bifunctional enzymes catalyzing secoisolariciresinol biosynthesis. The study of the transcriptional regulation of PLR genes evidenced very contrasting expression profiles and led to the identification of transcription factors acting as master regulators of this biosynthesis and their cis-regulatory target elements. Results allowed first to reinforce the hypothesis of the lignans defensive role in planta, second to afford an overall view of complex mechanisms occurring in the regulation of lignan natural production and finally to suggest leads to improve lignan yield by a stimulation of natural production and an enhancement of extraction yield using green chemistry methods
Secondary metabolite accumulation, antibacterial and antioxidant properties of in vitro propagated Clidemia hirta L. extracts are influenced by the basal culture medium
International audienceClidemia hirta L, a tropical shrub used for traditional medicine in numerous countries, could constitute a new resource of phytochemicals for cosmetic applications. In vitro micro propagation of C. hirta was used to evaluate the influence of different culture media on plant growth, production of phytochemicals, and the antioxidant and antibacterial properties of leaf extracts. Quoirin and Lepoivre medium and Lloyd and McCown's woody plant medium gave the best results. Both production of phytochemicals (i.e., flavonoids, phenolics and saponins) and biological activities were affected by the culture medium composition with the strongest effects for plants cultivated on Quoirin and Lepoivre medium. Strong correlations were shown between the antibacterial activity and the saponin content and between the antioxidant capacity and the flavonoid content. The present study shows how mineral nutrition influences the production of secondary metabolites in C hirta, thus modulating the biological activities of extracts with a view to their possible use in the cosmetic industry. (C) 2016 Academie des sciences. Published by Elsevier Masson SAS
Nettle (Urtica dioica L.) as a source of antioxidant and anti-aging phytochemicals for cosmetic applications
Nettle (Urtica dioica L.) is a herbaceous perennial that has been used for centuries in folk medicine. More recently, nettle extracts have also been used in cosmetics because of the many benefits of their topical application for skin health. Their potential anti-aging action is of particular interest and is primarily ascribed to their antioxidant capacity. Here, using an experimental design approach and a clustering analysis, we linked the phytochemical composition of nettle extracts to their biological activities. This approach confirmed the antioxidant capacity of nettle extracts as well as providing the first evidence of another mechanism for their anti-aging potential involving the inhibition of enzyme activities, such as elastase and collagenase. We attributed these inhibitory effects to ursolic acid and quercetin present in the nettle extracts. Our results also demonstrated the possibility of extracting ursolic acid, quercetin and other phenolic compounds differentially to obtain an extract with a strong antioxidant capacity and anti-aging activities toward both elastase and collagenase. This could be of particular interest for cosmetic applications of nettle extracts.L'ortie (Urtica dioica L.) est une plante herbacĂ©e vivace utilisĂ©e depuis des siĂšcles en mĂ©decine traditionnelle. Plus rĂ©cemment, les extraits d'ortie ont Ă©tĂ© Ă©galement utilisĂ©s dans des produits cosmĂ©tiques en raison des nombreux effets bĂ©nĂ©fiques de leurs applications topiques sur la santĂ© de la peau. En particulier, leur potentielle action anti-Ăąge prĂ©sente un fort intĂ©rĂȘt et a Ă©tĂ© essentiellement attribuĂ©e Ă leur fort pouvoir antioxydant. Dans cette Ă©tude, en utilisant un plan d'expĂ©rience et une analyse hiĂ©rarchisĂ©e, nous avons connectĂ© la composition phytochimique d'extraits d'ortie Ă leurs activitĂ©s biologiques. Cette approche nous a permis de confirmer la capacitĂ© antioxydante des extraits d'ortie, mais aussi de proposer la premiĂšre preuve d'un autre mĂ©canisme de leur potentiel anti-Ăąge impliquant l'inhibition d'activitĂ©s enzymatiques. En effet, nos travaux montrent que ce second mĂ©canisme implique l'inhibition d'enzymes de dĂ©gradation de la matrice extracellulaire telles que l'Ă©lastase et de la collagĂ©nase, et ces effets inhibiteurs ont Ă©tĂ© attribuables principalement Ă l'acide ursolique et la quercĂ©tine prĂ©sents dans les extraits d'ortie. Nos rĂ©sultats ont Ă©galement mis en Ă©vidence la possibilitĂ© d'extraire de maniĂšre diffĂ©rentielle l'acide ursolique, la quercĂ©tine et d'autres composĂ©s phĂ©noliques pour obtenir un extrait avec une forte capacitĂ© antioxydante et une activitĂ© anti-Ăąge au travers de l'inhibition Ă la fois de l'Ă©lastase et de la collagĂ©nase, ce qui pourrait ĂȘtre d'un intĂ©rĂȘt particulier pour les applications cosmĂ©tiques des extraits d'ortie
RNAi-mediated pinoresinol lariciresinol reductase gene silencing in flax (Linum usitatissimum L.) seed coat: Consequences on lignans and neolignans accumulation
International audienceRNAi technology was applied to down regulate LuPLR1 gene expression in flax (Linum usitatissimum L.) seeds. This gene encodes a pinoresinol lariciresinol reductase responsible for the synthesis of (+)-secoisolariciresinol diglucoside (SDG), the major lignan accumulated in the seed coat. If flax lignans biological properties and health benefits are well documented their roles in planta remain unclear. This loss of function strategy was developed to better understand the implication of the PLR1 enzyme in the lignan biosynthetic pathway and to provide new insights on the functions of these compounds. RNAi plants generated exhibited LuPLR1 gene silencing as demonstrated by quantitative RT-PCR experiments and the failed to accumulate SDG. The accumulation of pinoresinol the substrate of the PLR1 enzyme under its diglucosylated form (PDG) was increased in transgenic seeds but did not compensate the overall loss of SDG. The monolignol flux was also deviated through the synthesis of 8-5' linked neolignans dehydrodiconiferyl alcohol glucoside (DCG) and dihydro-dehydrodiconiferyl alcohol glucoside (DDCG) which were observed for the first time in flax seeds
Measurement report: Comparison of airborne, in situ measured, lidar-based, and modeled aerosol optical properties in the central European background â identifying sources of deviations
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Microwave-Assisted Extraction of Herbacetin Diglucoside from Flax (Linum usitatissimum L.) Seed Cakes and Its Quantification using an RP-HPLC-UV System
Flax (Linum usitatissimum L.) seeds are widely used for oil extraction and the cold-pressed flaxseed (or linseed) cakes obtained during this process constitute a valuable by-product. The flavonol herbacetin diglucoside (HDG) has been previously reported as a constituent of the flaxseed lignan macromolecule linked through ester bonds to the linker molecule hydroxymethylglutaric acid. In this context, the development and validation of a new approach using microwave-assisted extraction (MAE) of HDG from flaxseed cakes followed by quantification with a reverse-phase HPLC system with UV detection was purposed. The experimental parameters affecting the HDG extraction yield, such as microwave power, extraction time and sodium hydroxide concentration, from the lignan macromolecule were optimized. A maximum HDG concentration of 5.76 mg/g DW in flaxseed cakes was measured following an irradiation time of 6 min, for a microwave power of 150 W using a direct extraction in 0.1 M NaOH in 70% (v/v) aqueous methanol. The optimized method was proven to be rapid and reliable in terms of precision, repeatability, stability and accuracy for the extraction of HDG. Comparison with a conventional extraction method demonstrated that MAE is more effective and less time-consuming
Investigation of the Lignan Content in Extracts from Linum, Callitris and Juniperus Species in Relation to Their In Vitro Antiproliferative Activities
Podophyllotoxin, a lignan still extracted from the rhizomes of Podophyllum hexandrum (Berberidaceae), is the starting molecule for the semisynthesis of widely used anticancer drugs such as etoposide. However, this source is threatened by the over-collection of P. hexandrum. Plants belonging to the Linaceae and Cupressaceae families could be attractive alternative sources with species that contain the lignan podophyllotoxin or its precursors and derivatives. Wild flax species, such as Linum flavum, as well as some Juniperus and Callitris species were investigated for their lignan content, and the in vitro antiproliferative capacity of their extracts was assayed on four tumor cell lines. Some of the lignans were detected by LC-HRMS for the first time in these extracts. In addition, lignans purified from these plants and compounds semisynthesized from commercially available podophyllotoxin were tested in terms of their in vitro antiproliferative activity. The genus Juniperus was the most promising given its in vitro antiproliferative effects, which were also observed with extracts from L. flavum and Callitris species. The in vitro antiproliferative effect of the plant extracts studied here appears to correlate well with the contents of the aryltetralin lignan podophyllotoxin and its glycoside as well as with deoxypodophyllotoxin and 6-methoxypodophyllotoxin. The strongest correlation between the lignan content of the extracts and the antiproliferative activity was observed for 6-methoxypodophyllotoxin. Regarding the possibility of producing large renewable amounts of 6-methoxypodophyllotoxin, this molecule could be of interest to produce new anticancer drugs and to bypass the resistance mechanisms against podophyllotoxin-derived drugs.SCOPUS: ar.jinfo:eu-repo/semantics/publishe