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1.
Phytochem Anal ; 35(3): 445-468, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38069552

RESUMO

INTRODUCTION: The genus Clusia L. is mostly recognised for the production of prenylated benzophenones and tocotrienol derivatives. OBJECTIVES: The objective of this study was to map metabolome variation within Clusia minor organs at different developmental stages. MATERIAL AND METHODS: In total 15 organs/stages (leaf, flower, fruit, and seed) were analysed by UPLC-MS and 1H- and heteronuclear multiple-bond correlation (HMBC)-NMR-based metabolomics. RESULTS: This work led to the assignment of 46 metabolites, belonging to organic acids(1), sugars(2) phenolic acids(1), flavonoids(3) prenylated xanthones(1) benzophenones(4) and tocotrienols(2). Multivariate data analyses explained the variability and classification of samples, highlighting chemical markers that discriminate each organ/stage. Leaves were found to be rich in 5-hydroxy-8-methyltocotrienol (8.5 µg/mg f.w.), while flowers were abundant in the polyprenylated benzophenone nemorosone with maximum level detected in the fully mature flower bud (43 µg/mg f.w.). Nemorosone and 5-hydroxy tocotrienoloic acid were isolated from FL6 for full structural characterisation. This is the first report of the NMR assignments of 5-hydroxy tocotrienoloic acid, and its maximum level was detected in the mature fruit at 50 µg/mg f.w. Seeds as typical storage organ were rich in sugars and omega-6 fatty acids. CONCLUSION: To the best of our knowledge, this is the first report on a comparative 1D-/2D-NMR approach to assess compositional differences in ontogeny studies compared with LC-MS exemplified by Clusia organs. Results derived from this study provide better understanding of the stages at which maximal production of natural compounds occur and elucidate in which developmental stages the enzymes responsible for the production of such metabolites are preferentially expressed.


Assuntos
Clusia , Clusia/química , Frutas/química , Cromatografia Líquida , Espectrometria de Massas em Tandem , Benzofenonas/análise , Benzofenonas/química , Benzofenonas/metabolismo , Flores/química , Folhas de Planta/química , Metabolômica/métodos , Sementes/química , Açúcares/análise
2.
J Nat Prod ; 86(6): 1373-1384, 2023 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-37306303

RESUMO

Three previously undescribed azepino-indole alkaloids, named purpurascenines A-C (1-3), together with the new-to-nature 7-hydroxytryptophan (4) as well as two known compounds, adenosine (5) and riboflavin (6), were isolated from fruiting bodies of Cortinarius purpurascens Fr. (Cortinariaceae). The structures of 1-3 were elucidated based on spectroscopic analyses and ECD calculations. Furthermore, the biosynthesis of purpurascenine A (1) was investigated by in vivo experiments using 13C-labeled sodium pyruvate, alanine, and sodium acetate incubated with fruiting bodies of C. purpurascens. The incorporation of 13C into 1 was analyzed using 1D NMR and HRESIMS methods. With [3-13C]-pyruvate, a dramatic enrichment of 13C was observed, and hence a biosynthetic route via a direct Pictet-Spengler reaction between α-keto acids and 7-hydroxytryptophan (4) is suggested for the biosynthesis of purpurascenines A-C (1-3). Compound 1 exhibits no antiproliferative or cytotoxic effects against human prostate (PC-3), colorectal (HCT-116), and breast (MCF-7) cancer cells. An in silico docking study confirmed the hypothesis that purpurascenine A (1) could bind to the 5-HT2A serotonin receptor's active site. A new functional 5-HT2A receptor activation assay showed no functional agonistic but some antagonistic effects of 1 against the 5-HT-dependent 5-HT2A activation and likely antagonistic effects on putative constitutive activity of the 5-HT2A receptor.


Assuntos
Cortinarius , Serotonina , Masculino , Humanos , Serotonina/metabolismo , Serotonina/farmacologia , Receptor 5-HT2A de Serotonina , Alcaloides Indólicos/farmacologia , Cortinarius/química , Cortinarius/metabolismo
3.
Plant J ; 105(5): 1309-1325, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33617106

RESUMO

Secretions from glandular trichomes potentially protect plants against a variety of aggressors. In the tomato clade of the Solanum genus, glandular trichomes of wild species produce a rich source of chemical diversity at the leaf surface. Previously, 7-epi-zingiberene produced in several accessions of Solanum habrochaites was found to confer resistance to whiteflies (Bemisia tabaci) and other insect pests. Here, we report the identification and characterisation of 9-hydroxy-zingiberene (9HZ) and 9-hydroxy-10,11-epoxyzingiberene (9H10epoZ), two derivatives of 7-epi-zingiberene produced in glandular trichomes of S. habrochaites LA2167. Using a combination of transcriptomics and genetics, we identified a gene coding for a cytochrome P450 oxygenase, ShCYP71D184, that is highly expressed in trichomes and co-segregates with the presence of the zingiberene derivatives. Transient expression assays in Nicotiana benthamiana showed that ShCYP71D184 carries out two successive oxidations to generate 9HZ and 9H10epoZ. Bioactivity assays showed that 9-hydroxy-10,11-epoxyzingiberene in particular exhibits substantial toxicity against B. tabaci and various microorganisms including Phytophthora infestans and Botrytis cinerea. Our work shows that trichome secretions from wild tomato species can provide protection against a wide variety of organisms. In addition, the availability of the genes encoding the enzymes for the pathway of 7-epi-zingiberene derivatives makes it possible to introduce this trait in cultivated tomato by precision breeding.


Assuntos
Hemípteros/metabolismo , Sesquiterpenos Monocíclicos/metabolismo , NADPH-Ferri-Hemoproteína Redutase/metabolismo , Solanum/metabolismo , Animais , Botrytis/efeitos dos fármacos , Botrytis/patogenicidade , Hemípteros/genética , Hemípteros/microbiologia , Sesquiterpenos Monocíclicos/toxicidade , NADPH-Ferri-Hemoproteína Redutase/genética , Phytophthora infestans/efeitos dos fármacos , Phytophthora infestans/patogenicidade , Solanum/genética
4.
Molecules ; 27(8)2022 Apr 13.
Artigo em Inglês | MEDLINE | ID: mdl-35458711

RESUMO

A rare dihydoxyflavan-epicatechin proanthocyanidin, entcassiflavan-(4ß→8)-epicatechin, was isolated from Dalbergia monetaria, a plant widely used by traditional people from the Amazon to treat urinary tract infections. The constitution and relative configuration of the compound were elucidated by HR-MS and detailed 1D- and 2D-NMR measurements. By comparing the experimental electronic circular dichroism (ECD) spectrum with the calculated ECD spectra of all 16 possible isomers, the absolute configuration, the interflavan linkage, and the atropisomers could be determined.


Assuntos
Catequina , Dalbergia , Dicroísmo Circular , Eletrônica , Humanos , Estrutura Molecular
5.
Plant J ; 102(3): 569-581, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-31837062

RESUMO

Black pepper (Piper nigrum L.) is known for its high content of piperine, a cinnamoyl amide derivative regarded as largely responsible for the pungent taste of this widely used spice. Despite its long history and worldwide use, the biosynthesis of piperine and related amides has been enigmatic up to now. In this report we describe a specific piperic acid CoA ligase from immature green fruits of P. nigrum. The corresponding enzyme was cloned and functionally expressed in E. coli. The recombinant enzyme displays a high specificity for piperic acid and does not accept the structurally related feruperic acid characterized by a similar C-2 extension of the general C6-C3 phenylpropanoid structure. The enzyme is also inactive with the standard set of hydroxycinnamic acids tested including caffeic acid, 4-coumaric acid, ferulic acid, and sinapic acid. Substrate specificity is corroborated by in silico modelling that suggests a perfect fit for the substrate piperic acid to the active site of the piperic acid CoA ligase. The CoA ligase gene shows its highest expression levels in immature green fruits, is also expressed in leaves and flowers, but not in roots. Virus-induced gene silencing provided some preliminary indications that the production of piperoyl-CoA is required for the biosynthesis of piperine in black pepper fruits.


Assuntos
Alcaloides/metabolismo , Benzodioxóis/metabolismo , Coenzima A Ligases/metabolismo , Frutas/metabolismo , Piper nigrum/metabolismo , Piperidinas/metabolismo , Alcamidas Poli-Insaturadas/metabolismo , Coenzima A Ligases/genética , Frutas/genética , Inativação Gênica , Piper nigrum/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo
6.
Int J Mol Sci ; 22(23)2021 Nov 24.
Artigo em Inglês | MEDLINE | ID: mdl-34884518

RESUMO

Fungal species of genus Sepedonium are rich sources of diverse secondary metabolites (e.g., alkaloids, peptaibols), which exhibit variable biological activities. Herein, two new peptaibols, named ampullosporin F (1) and ampullosporin G (2), together with five known compounds, ampullosporin A (3), peptaibolin (4), chrysosporide (5), c(Trp-Ser) (6) and c(Trp-Ala) (7), have been isolated from the culture of Sepedonium ampullosporum Damon strain KSH534. The structures of 1 and 2 were elucidated based on ESI-HRMSn experiments and intense 1D and 2D NMR analyses. The sequence of ampullosporin F (1) was determined to be Ac-Trp1-Ala2-Aib3-Aib4-Leu5-Aib6-Gln7-Aib8-Aib9-Aib10-GluOMe11-Leu12-Aib13-Gln14-Leuol15, while ampullosporin G (2) differs from 1 by exchanging the position of Gln7 with GluOMe11. Furthermore, the total synthesis of 1 and 2 was carried out on solid-phase to confirm the absolute configuration of all chiral amino acids as L. In addition, ampullosporin F (1) and G (2) showed significant antifungal activity against B. cinerea and P. infestans, but were inactive against S. tritici. Cell viability assays using human prostate (PC-3) and colorectal (HT-29) cancer cells confirmed potent anticancer activities of 1 and 2. Furthermore, a molecular docking study was performed in silico as an attempt to explain the structure-activity correlation of the characteristic ampullosporins (1-3).


Assuntos
Antifúngicos/farmacologia , Antineoplásicos/farmacologia , Ésteres/química , Ácido Glutâmico/química , Hypocreales/fisiologia , Neoplasias/tratamento farmacológico , Peptaibols/farmacologia , Ascomicetos/efeitos dos fármacos , Botrytis/efeitos dos fármacos , Humanos , Neoplasias/patologia , Peptaibols/química , Phytophthora infestans/efeitos dos fármacos , Células Tumorais Cultivadas
7.
Molecules ; 26(6)2021 Mar 21.
Artigo em Inglês | MEDLINE | ID: mdl-33800987

RESUMO

Phytochemical investigation of the methanolic extract obtained from the aerial parts of Lagochilus setulosus (Lamiaceae) afforded the new compound 1-methoxy-3-O-ß-glucopyranosyl-α-l-oliose (1) together with five known glycosides, namely sitosterol-3-O-ß-glucoside (2), stigmasterol-3-O-ß-glucoside (3), pinitol (4), 6ß-hydroxyl-7-epi-loganin (5), and chlorotuberoside (6). The structures of these compounds were elucidated by extensive spectroscopic analyses, especially HR-MS, 1D and 2D NMR spectroscopy. The in vitro cytotoxic activity of the methanolic extract and the isolated compounds was assessed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and crystal violet (CV) staining assays. In addition, the antifungal activities of the components were evaluated against Botrytis cinerea, Septoria tritici, and Phytophthora infestans. The anthelmintic potential was determined against Caenorhabditis elegans nematodes. Neither the extract nor the isolated compounds showed promising activity in all the bioassays.


Assuntos
Anti-Helmínticos , Antifúngicos , Glicosídeos , Lamiaceae/química , Extratos Vegetais/química , Animais , Anti-Helmínticos/química , Anti-Helmínticos/isolamento & purificação , Anti-Helmínticos/farmacologia , Antifúngicos/química , Antifúngicos/isolamento & purificação , Antifúngicos/farmacologia , Ascomicetos/crescimento & desenvolvimento , Botrytis/crescimento & desenvolvimento , Caenorhabditis elegans/crescimento & desenvolvimento , Glicosídeos/química , Glicosídeos/isolamento & purificação , Glicosídeos/farmacologia , Phytophthora infestans/crescimento & desenvolvimento
8.
Molecules ; 26(3)2021 Feb 02.
Artigo em Inglês | MEDLINE | ID: mdl-33540661

RESUMO

Legume sprouts are a fresh nutritive source of phytochemicals of increasing attention worldwide owing to their many health benefits. Nuclear magnetic resonance (NMR) was utilized for the metabolite fingerprinting of 4 major legume sprouts, belonging to family Fabaceae, to be exploited for quality control purposes. Thirty-two metabolites were identified belonging to different classes, i.e., fatty acids, sugars, amino acids, nucleobases, organic acids, sterols, alkaloids, and isoflavonoids. Quantitative NMR was employed for assessing the major identified metabolite levels and multivariate data analysis was utilized to assess metabolome heterogeneity among sprout samples. Isoflavones were detected exclusively in Cicer sprouts, whereas Trigonella was characterized by 4-hydroxyisoleucine. Vicia sprouts were distinguished from other legume sprouts by the presence of L-Dopa versus acetate abundance in Lens. A common alkaloid in all sprouts was trigonelline, detected at 8-25 µg/mg, suggesting its potential role in legume seeds' germination. Trigonelline was found at highest levels in Trigonella sprouts. The aromatic NMR region data (δ 11.0-5.0 ppm) provided a better classification power than the full range (δ 11.0-0.0 ppm) as sprout variations mostly originated from secondary metabolites, which can serve as chemotaxonomic markers.


Assuntos
Quimioinformática , Fabaceae/química , Fabaceae/metabolismo , Espectroscopia de Ressonância Magnética , Metabolômica
9.
Plant Physiol ; 180(3): 1598-1613, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31015300

RESUMO

In nature, plants interact with numerous beneficial or pathogenic soil-borne microorganisms. Plants have developed various defense strategies to expel pathogenic microbes, some of which function soon after pathogen infection. We used Medicago truncatula and its oomycete pathogen Aphanomyces euteiches to elucidate early responses of the infected root. A. euteiches causes root rot disease in legumes and is a limiting factor in legume production. Transcript profiling of seedlings and adult plant roots inoculated with A. euteiches zoospores for 2 h revealed specific upregulation of a gene encoding a putative sesquiterpene synthase (M. truncatula TERPENE SYNTHASE 10 [MtTPS10]) in both developmental stages. MtTPS10 was specifically expressed in roots upon oomycete infection. Heterologous expression of MtTPS10 in yeast led to production of a blend of sesquiterpenes and sesquiterpene alcohols, with NMR identifying a major peak corresponding to himalachol. Moreover, plants carrying a tobacco (Nicotiana tabacum) retrotransposon Tnt1 insertion in MtTPS10 lacked the emission of sesquiterpenes upon A. euteiches infection, supporting the assumption that the identified gene encodes a multiproduct sesquiterpene synthase. Mttps10 plants and plants with reduced MtTPS10 transcript levels created by expression of an MtTPS10-artificial microRNA in roots were more susceptible to A. euteiches infection than were the corresponding wild-type plants and roots transformed with the empty vector, respectively. Sesquiterpenes produced by expression of MtTPS10 in yeast also inhibited mycelial growth and A. euteiches zoospore germination. These data suggest that sesquiterpene production in roots by MtTPS10 plays a previously unrecognized role in the defense response of M. truncatula against A. euteiches.


Assuntos
Alquil e Aril Transferases/genética , Resistência à Doença/genética , Medicago truncatula/genética , Doenças das Plantas/genética , Proteínas de Plantas/genética , Raízes de Plantas/genética , Alquil e Aril Transferases/metabolismo , Aphanomyces/fisiologia , Perfilação da Expressão Gênica/métodos , Regulação Enzimológica da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Interações Hospedeiro-Patógeno , Medicago truncatula/enzimologia , Medicago truncatula/microbiologia , Doenças das Plantas/microbiologia , Proteínas de Plantas/metabolismo , Raízes de Plantas/enzimologia , Raízes de Plantas/microbiologia , Sesquiterpenos/metabolismo
10.
Nat Chem Biol ; 14(2): 171-178, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29291349

RESUMO

Biosynthesis of the phytohormone jasmonoyl-isoleucine (JA-Ile) requires reduction of the JA precursor 12-oxo-phytodienoic acid (OPDA) by OPDA reductase 3 (OPR3). Previous analyses of the opr3-1 Arabidopsis mutant suggested an OPDA signaling role independent of JA-Ile and its receptor COI1; however, this hypothesis has been challenged because opr3-1 is a conditional allele not completely impaired in JA-Ile biosynthesis. To clarify the role of OPR3 and OPDA in JA-independent defenses, we isolated and characterized a loss-of-function opr3-3 allele. Strikingly, opr3-3 plants remained resistant to necrotrophic pathogens and insect feeding, and activated COI1-dependent JA-mediated gene expression. Analysis of OPDA derivatives identified 4,5-didehydro-JA in wounded wild-type and opr3-3 plants. OPR2 was found to reduce 4,5-didehydro-JA to JA, explaining the accumulation of JA-Ile and activation of JA-Ile-responses in opr3-3 mutants. Our results demonstrate that in the absence of OPR3, OPDA enters the ß-oxidation pathway to produce 4,5-ddh-JA as a direct precursor of JA and JA-Ile, thus identifying an OPR3-independent pathway for JA biosynthesis.


Assuntos
Arabidopsis/metabolismo , Ciclopentanos/metabolismo , Resistência à Doença , Isoleucina/análogos & derivados , Oxilipinas/metabolismo , Doenças das Plantas/prevenção & controle , Alelos , Alternaria , Animais , Proteínas de Arabidopsis/metabolismo , Bioensaio , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Homozigoto , Insetos , Isoleucina/metabolismo , Mutação , Reguladores de Crescimento de Plantas/metabolismo , Raízes de Plantas/metabolismo , Transdução de Sinais
11.
Planta Med ; 86(12): 858-866, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32483774

RESUMO

Dalbergia monetaria is an Amazonian plant whose bark is widely used to treat urinary tract infections. This paper describes a bio-guided study of ethanolic extracts from the bark and leaves of D. monetaria, in a search for metabolites active against human pathogenic bacteria. In vitro assays were performed against 10 bacterial strains, highlighting methicillin-sensitive Staphylococcus aureus and methicillin-resistant Staphylococcus aureus and Pseudomonas aeruginosa. Fractioning of the extracts was performed using instrumental and classical techniques, and samples were characterized by UHPLC-HRMS/MS. Ethyl acetate fractions from bark and leaves showed similar antibacterial activities. EAFB is enriched in isoflavone C-glucosides and EAFL enriched in proanthocyanidins. Subfractions from EAFL presented higher activity and showed a complex profile of proanthocyanidins constructed by (epi)-cassiaflavan and (epi)-catechin units, including dimers, trimers and tetramers. The fragmentation pattern emphasized the neutral loss of cassiaflavan units by quinone-methide fission. Fraction SL7-6, constituted by (ent)-cassiaflavan-(ent)-cassiaflavan-(epi)-catechin isomers, showed the lowest MIC against the S. aureus and P. aeruginosa with values corresponding to 64 and 32 µg/mL, respectively. Cassiaflavan-proanthocyanidins have not been found previously in another botanical genus, except in Cassia, and the traditional medicinal use of D. monetaria might be related to the antibacterial activity of proanthocyanidins characterized in the species.


Assuntos
Dalbergia , Staphylococcus aureus Resistente à Meticilina , Plantas Medicinais , Proantocianidinas , Antibacterianos , Cromatografia Líquida de Alta Pressão , Humanos , Testes de Sensibilidade Microbiana , Extratos Vegetais , Staphylococcus aureus
12.
Int J Mol Sci ; 21(2)2020 Jan 14.
Artigo em Inglês | MEDLINE | ID: mdl-31947682

RESUMO

Nowadays, applying bio-organic fertilizer (e.g., chitosan, Ch) or integrating beneficial microorganisms (e.g., arbuscular mycorrhizal fungi, AMF) are among the successful strategies to promote plant growth. Here, the effect of two application modes of Ch (foliar spray or root treatment) and Ch-derived nanoparticles (NPs) on tomato plants colonized with the AMF Rhizophagus irregularis were analyzed, thereby focusing on plant biomass, flowering and mycorrhization. An increase of shoot biomass and flower number was observed in arbuscular mycorrhizal (AM) plants sprayed with Ch. The interaction with AMF, however, was reduced as shown by decreased mycorrhization rates and AM-specific gene expression. To get insights into Ch effect on mycorrhization, levels of sugars, jasmonates, abscisic acid, and the expression of two chitinase-encoding genes were determined in mycorrhizal roots. Ch had no effect on sugar and phytohormone levels, but the reduced mycorrhization was correlated with down- and upregulated expression of Chi3 and Chi9, respectively. In contrast, application of NPs to leaves and Ch applied to the soil did not show any effect, neither on mycorrhization rate nor on growth of mycorrhizal plants. Concluding, Ch application to leaves enhanced plant growth and flowering and reduced interaction with AMF, whereas root treatment did not affect these parameters.


Assuntos
Quitinases/genética , Quitinases/metabolismo , Quitosana , Fertilizantes , Micorrizas , Solanum lycopersicum/efeitos dos fármacos , Solanum lycopersicum/crescimento & desenvolvimento , Quitosana/química , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Nanopartículas
13.
Molecules ; 25(21)2020 Nov 04.
Artigo em Inglês | MEDLINE | ID: mdl-33158186

RESUMO

1H-NMR is a very reproducible spectroscopic method and, therefore, a powerful tool for the metabolomic analysis of biological samples. However, due to the high complexity of natural samples, such as plant extracts, the evaluation of spectra is difficult because of signal overlap. The new NMR "Pure Shift" methods improve spectral resolution by suppressing homonuclear coupling and turning multiplets into singlets. The PSYCHE (Pure Shift yielded by Chirp excitation) and the Zangger-Sterk pulse sequence were tested. The parameters of the more suitable PSYCHE experiment were optimized, and the extracts of 21 Hypericum species were measured. Different evaluation criteria were used to compare the suitability of the PSYCHE experiment with conventional 1H-NMR. The relationship between the integral of a signal and the related bin value established by linear regression demonstrates an equal representation of the integrals in binned PSYCHE spectra compared to conventional 1H-NMR. Using multivariate data analysis based on both techniques reveals comparable results. The obtained data demonstrate that Pure Shift spectra can support the evaluation of conventional 1H-NMR experiments.


Assuntos
Hypericum/metabolismo , Metaboloma , Metabolômica , Ressonância Magnética Nuclear Biomolecular
14.
Angew Chem Int Ed Engl ; 59(1): 259-263, 2020 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-31797518

RESUMO

In contrast to the myriad of methods available to produce α-helices and antiparallel ß-sheets in synthetic peptides, just a few are known for the construction of stable, non-cyclic parallel ß-sheets. Herein, we report an efficient on-resin approach for the assembly of parallel ß-sheet peptides in which the N-alkylated turn moiety enhances the stability and gives access to a variety of functionalizations without modifying the parallel strands. The key synthetic step of this strategy is the multicomponent construction of an N-alkylated turn using the Ugi reaction on varied isocyano-resins. This four-component process assembles the orthogonally protected turn fragment and incorporates handles serving for labeling/conjugation purposes or for reducing peptide aggregation. NMR and circular dichroism analyses confirm the better-structured and more stable parallel ß-sheets in the N-alkylated peptides compared to the non-functionalized variants.

15.
Bioconjug Chem ; 30(1): 253-259, 2019 01 16.
Artigo em Inglês | MEDLINE | ID: mdl-30575393

RESUMO

Peptide stapling is traditionally used to lock peptide conformations into α-helical structures using a variety of macrocyclization chemistries. In an endeavor to add a diversity-generating tool to this repertoire, we introduce a multicomponent stapling approach enabling the simultaneous stabilization of helical secondary structures and the exocyclic N-functionalization of the side chain-tethering lactam bridge. This is accomplished by means of a novel solid-phase methodology comprising, for the first time, the on-resin Ugi reaction-based macrocyclization of peptide side chains bearing amino and carboxylic acid groups. The exocyclic diversity elements arise from the isocyanide component used in the Ugi multicomponent stapling protocol, which allows for the incorporation of relevant fragments such as lipids, sugars, polyethylene glycol, fluorescent labels, and reactive handles. We prove the utility of such exocyclic reactive groups in the bioconjugation of a maleimide-armed lactam-bridged peptide to a carrier protein. The on-resin multicomponent stapling proved efficient for the installation of not only one, but also two consecutive lactam bridges having either identical or dissimilar N-functionalities. The easy access to helical peptides with a diverse set of exocyclic functionalities shows prospect for applications in peptide drug discovery and chemical biology.


Assuntos
Lactamas/química , Lipídeos/química , Peptídeos/química , Polietilenoglicóis/química , Açúcares/química
16.
Metabolomics ; 15(5): 80, 2019 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-31087208

RESUMO

INTRODUCTION: The demand to develop efficient and reliable analytical methods for the quality control of nutraceuticals is on the rise, together with an increase in the legal requirements for safe and consistent levels of its active principles. OBJECTIVE: To establish a reliable model for the quality control of widely used Senna preparations used as laxatives and assess its phyto-equivalency. METHODS: A comparative metabolomics approach via NMR and MS analyses was employed for the comprehensive measurement of metabolites and analyzed using chemometrics. RESULTS: Under optimized conditions, 30 metabolites were simultaneously identified and quantified including anthraquinones, bianthrones, acetophenones, flavonoid conjugates, naphthalenes, phenolics, and fatty acids. Principal component analysis (PCA) was used to define relative metabolite differences among Senna preparations. Furthermore, quantitative 1H NMR (qHNMR) was employed to assess absolute metabolites levels in preparations. Results revealed that 6-hydroxy musizin or tinnevellin were correlated with active metabolites levels, suggesting the use of either of these naphthalene glycosides as markers for official Senna drugs authentication. CONCLUSION: This study provides the first comparative metabolomics approach utilizing NMR and UPLC-MS to reveal for secondary metabolite compositional differences in Senna preparations that could readily be applied as a reliable quality control model for its analysis.


Assuntos
Metabolômica , Senosídeos/metabolismo , Acetofenonas/metabolismo , Antracenos/metabolismo , Antraquinonas/metabolismo , Flavonoides/metabolismo , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Estrutura Molecular , Naftalenos/metabolismo , Fenóis/metabolismo , Análise de Componente Principal , Controle de Qualidade , Senosídeos/química
17.
Anal Chem ; 90(1): 649-656, 2018 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-29035042

RESUMO

NMR is a widely used analytical technique with a growing number of repositories available. As a result, demands for a vendor-agnostic, open data format for long-term archiving of NMR data have emerged with the aim to ease and encourage sharing, comparison, and reuse of NMR data. Here we present nmrML, an open XML-based exchange and storage format for NMR spectral data. The nmrML format is intended to be fully compatible with existing NMR data for chemical, biochemical, and metabolomics experiments. nmrML can capture raw NMR data, spectral data acquisition parameters, and where available spectral metadata, such as chemical structures associated with spectral assignments. The nmrML format is compatible with pure-compound NMR data for reference spectral libraries as well as NMR data from complex biomixtures, i.e., metabolomics experiments. To facilitate format conversions, we provide nmrML converters for Bruker, JEOL and Agilent/Varian vendor formats. In addition, easy-to-use Web-based spectral viewing, processing, and spectral assignment tools that read and write nmrML have been developed. Software libraries and Web services for data validation are available for tool developers and end-users. The nmrML format has already been adopted for capturing and disseminating NMR data for small molecules by several open source data processing tools and metabolomics reference spectral libraries, e.g., serving as storage format for the MetaboLights data repository. The nmrML open access data standard has been endorsed by the Metabolomics Standards Initiative (MSI), and we here encourage user participation and feedback to increase usability and make it a successful standard.


Assuntos
Bases de Dados de Compostos Químicos/normas , Espectroscopia de Ressonância Magnética/estatística & dados numéricos , Metabolômica/métodos , Software
18.
Z Naturforsch C J Biosci ; 73(1-2): 33-39, 2018 Jan 26.
Artigo em Inglês | MEDLINE | ID: mdl-28937966

RESUMO

Three new glucosylceramides (GluCers) named malusides I-III (1-3) were isolated from apple (cultivars of Malus domestica) pomace (fruit material remaining after juice extraction). An unusual oxo/hydroxy group pattern within the sphingadienine (d18:2) type sphingoid base was observed. All compounds contained the same α-hydroxylated fatty acid (h16:0) and a ß-D-glucose moiety. Their structures were assigned on the basis of one- and two-dimensional (1D and 2D) nuclear magnetic resonance (NMR) spectroscopic analyses and mass spectrometry (MS) measurements.


Assuntos
Glucosilceramidas/química , Malus/química , Glucosilceramidas/análise
19.
Mar Drugs ; 15(7)2017 Jul 04.
Artigo em Inglês | MEDLINE | ID: mdl-28677625

RESUMO

Sarcophyton sp. soft corals are rich in cembranoid diterpenes, which represent the main chemical defense of corals against their natural predators in addition to their myriad biological effects in humans. Quantitative NMR (qNMR) was applied for assessing the diterpene variation in 16 soft coral specimens in the context of their genotype, origin, and growing habitat. qNMR revealed high diterpene levels in Sarcophyton sp. compared to Sinularia and Lobophyton, with (ent)sarcophines as major components (17-100 µg/mg) of the coral tissues. Multivariate data analysis was employed to classify samples based on the quantified level of diterpenes, and compared to the untargeted NMR approach. Results revealed that qNMR provided a stronger classification model of Sarcophyton sp. than untargeted NMR fingerprinting. Additionally, cytotoxicity of soft coral crude extracts was assessed against androgen-dependent prostate cancer cell lines (PC3) and androgen-independent colon cancer cell lines (HT-29), with IC50 values ranging from 10-60 µg/mL. No obvious correlation between the extracts' IC50 values and their diterpene levels was found using either Spearman or Pearson correlations. This suggests that this type of bioactivity may not be easily predicted by NMR metabolomics in soft corals, or is not strongly correlated to measured diterpene levels.


Assuntos
Antozoários/química , Antineoplásicos/farmacologia , Diterpenos/farmacologia , Animais , Antineoplásicos/química , Linhagem Celular Tumoral , Diterpenos/química , Humanos , Concentração Inibidora 50 , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Análise Multivariada
20.
J Proteome Res ; 15(4): 1274-87, 2016 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-26892921

RESUMO

Marine life has developed unique metabolic and physiologic capabilities and advanced symbiotic relationships to survive in the varied and complex marine ecosystems. Herein, metabolite composition of the soft coral genus Sarcophyton was profiled with respect to its species and different habitats along the coastal Egyptian Red Sea via (1)H NMR and ultra performance liquid chromatography-mass spectrometry (UPLC-MS) large-scale metabolomics analyses. The current study extends the application of comparative secondary metabolite profiling from plants to corals revealing for metabolite compositional differences among its species via a comparative MS and NMR approach. This was applied for the first time to investigate the metabolism of 16 Sarcophyton species in the context of their genetic diversity or growth habitat. Under optimized conditions, we were able to simultaneously identify 120 metabolites including 65 diterpenes, 8 sesquiterpenes, 18 sterols, and 15 oxylipids. Principal component analysis (PCA) and orthogonal projection to latent structures-discriminant analysis (OPLS) were used to define both similarities and differences among samples. For a compound based classification of coral species, UPLC-MS was found to be more effective than NMR. The main differentiations emanate from cembranoids and oxylipids. The specific metabolites that contribute to discrimination between soft corals of S. ehrenbergi from the three different growing habitats also belonged to cembrane type diterpenes, with aquarium S. ehrenbergi corals being less enriched in cembranoids compared to sea corals. PCA using either NMR or UPLC-MS data sets was found equally effective in predicting the species origin of unknown Sarcophyton. Cyclopropane containing sterols observed in abundance in corals may act as cellular membrane protectant against the action of coral toxins, that is, cembranoids.


Assuntos
Antozoários/química , Diterpenos/isolamento & purificação , Metabolômica , Sesquiterpenos/isolamento & purificação , Esteróis/isolamento & purificação , Animais , Antozoários/metabolismo , Biodiversidade , Cromatografia Líquida de Alta Pressão/métodos , Análise Discriminante , Diterpenos/metabolismo , Egito , Oceano Índico , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Análise de Componente Principal , Metabolismo Secundário , Sesquiterpenos/metabolismo , Esteróis/metabolismo
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