RESUMO
Aflatoxins B1 (AFB1) and G1 (AFG1) are carcinogenic mycotoxins that contaminate crops such as maize and groundnuts worldwide. The broadly accepted method to assess chronic human aflatoxin exposure is by quantifying the amount of aflatoxin adducted to human serum albumin. This has been reported using ELISA, HPLC, or LC-MS/MS to measure the amount of AFB1-lysine released after proteolysis of serum albumin. LC-MS/MS is the most accurate method but requires both isotopically labelled and unlabelled AFB1-lysine standards, which are not commercially available. In this work, we report a simplified synthetic route to produce unlabelled, deuterated and 13C6 15N2 labelled aflatoxin B1-lysine and for the first-time aflatoxin G1-lysine. Additionally, we report on the stability of these compounds during storage. This simplified synthetic approach will make the production of these important standards more feasible for laboratories performing aflatoxin exposure studies.
Assuntos
Aflatoxina B1/síntese química , Aflatoxinas/síntese química , Lisina/química , Micotoxinas/síntese química , Cromatografia Líquida de Alta Pressão , Ensaio de Imunoadsorção Enzimática , Espectrometria de Massas em TandemRESUMO
Aspirochlorine is an unusual antifungal cyclopeptide produced by Aspergillus oryzae, an important mold used for food fermentation. Whereas its structure suggested that a non-ribosomal peptide synthetase assembles the cyclopeptide from phenylalanine and glycine building blocks, labeling studies indicated that one Phe moiety is transformed into Gly after peptide formation. By means of genetic engineering, heterologous expression, biotransformations, and inâ vitro assays, we dissected and reconstituted four crucial steps in aspirochlorine biosynthesis, which involve two cytochrome P450 monooxygenases, (AclL and AclO), a methyltransferase (AclU), and a halogenase (AclH). We found that the installation of the N-methoxylation of the peptide bond sets the stage for a retro-aldol reaction that leads to the Phe-to-Gly conversion. The substrate scopes of the dedicated enzymes as well as bioassays revealed that the peptide editing has evolved to optimize the antifungal action of the natural product.
Assuntos
Aldeídos/química , Amidas/química , Aminoácidos/química , Antifúngicos/síntese química , Micotoxinas/síntese química , Peptídeo Sintases/química , Compostos de Espiro/síntese química , Antifúngicos/farmacologia , Aspergillus fumigatus/efeitos dos fármacos , Candida albicans/efeitos dos fármacos , Cromatografia Líquida de Alta Pressão/métodos , Espectrometria de Massas/métodos , Testes de Sensibilidade Microbiana , Micotoxinas/farmacologia , Schizosaccharomyces/efeitos dos fármacos , Compostos de Espiro/farmacologia , Relação Estrutura-AtividadeRESUMO
α-Amanitin is an extremely toxic bicyclic octapeptide isolated from the death-cap mushroom, Amanita phalloides. As a potent inhibitor of RNA polymerase II, α-amanitin is toxic to eukaryotic cells. Recent interest in α-amanitin arises from its promise as a payload for antibody-drug conjugates. For over 60 years, A. phalloides has been the only source of α-amanitin. Here we report a synthesis of α-amanitin, which surmounts the key challenges for installing the 6-hydroxy-tryptathionine sulfoxide bridge, enantioselective synthesis of (2 S,3 R,4 R)-4,5-dihydroxy-isoleucine, and diastereoselective sulfoxidation.
Assuntos
Agaricales/química , Alfa-Amanitina/síntese química , Micotoxinas/síntese química , Alfa-Amanitina/química , Alfa-Amanitina/farmacologia , Animais , Células CHO , Cricetulus , Relação Dose-Resposta a Droga , Modelos Moleculares , Conformação Molecular , Micotoxinas/química , Micotoxinas/farmacologia , Relação Estrutura-AtividadeRESUMO
The analysis of the nephrotoxic mycotoxin citrinin in food, feed, and physiological samples is still challenging. Nowadays, liquid chromatography coupled with mass spectrometry is the method of choice for achieving low limits of detection. But matrix effects can present impairments for this method. Stable isotope dilution analysis can prevent some of these problems. Therefore, a stable isotopically labeled standard of citrinin for use in stable isotope dilution analysis was synthesized on large scale. The improved diastereoselective total synthetic strategy offered the possibility to introduce three 13C-labels in two steps by ortho-toluate anion chemistry. This led to a mass difference of 3 Da, sufficient for preventing spectral overlap. Additionally, a stable isotopically labeled form of dihydrocitrinone, the main urinary metabolite of citrinin, was synthesized with the same mass difference. This was achieved by a sequence of cyclisation, oxidation, deprotection, and carboxylation reactions starting from a protected intermediate of the labeled citrinin synthesis. Thus, this method also offers a complete way to synthesize dihydrocitrinone from citrinin on large scale.
Assuntos
Isótopos de Carbono , Citrinina/análogos & derivados , Citrinina/síntese química , Citrinina/metabolismo , Isótopos de Carbono/química , Cromatografia Líquida de Alta Pressão , Citrinina/análise , Citrinina/química , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Estrutura Molecular , Micotoxinas/análise , Micotoxinas/síntese química , Micotoxinas/química , Micotoxinas/metabolismo , Coloração e Rotulagem , Espectrometria de Massas em TandemRESUMO
Cyclic peptoids have recently emerged as important examples of peptidomimetics for their interesting complexing properties and innate ability to permeate biological barriers. In the present contribution, experimental and theoretical data evidence the intricate conformational and stereochemical properties of five novel hexameric peptoids decorated with N-isopropyl, N-isobutyl, and N-benzyl substituents. Complexation studies by NMR, in the presence of sodium tetrakis[3,5-bis(trifluoromethyl)phenyl]borate (NaTFPB), theoretical calculations, and single-crystal X-ray analyses indicate that the conformationally stable host/guest metal adducts display architectural ordering comparable to that of the enniatins and beauvericin mycotoxins. Similarly to the natural depsipeptides, the synthetic oligolactam analogues show a correlation between ion transport abilities in artificial liposomes and cytotoxic activity on human cancer cell lines. The reported results demonstrate that the versatile cyclic peptoid scaffold, for its remarkable conformational and complexing properties, can morphologically mimic related natural products and elicit powerful biological activities.
Assuntos
Micotoxinas/farmacologia , Peptidomiméticos/farmacologia , Peptoides/farmacologia , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cristalografia por Raios X , Relação Dose-Resposta a Droga , Humanos , Modelos Moleculares , Micotoxinas/síntese química , Micotoxinas/química , Peptidomiméticos/síntese química , Peptidomiméticos/química , Peptoides/síntese química , Peptoides/química , Conformação Proteica , Teoria Quântica , Relação Estrutura-AtividadeRESUMO
The stereoselective synthesis of the phomopsin A tripeptide side chain was achieved by using methyl 2-(((benzyloxy)carbonyl)amino)-2-(diphenoxyphosphoryl)acetate as a common synthetic precursor for the synthesis of E-Δ-dehydroisoleucine and E-Δ-aspartate.
Assuntos
Aminoácidos/química , Ésteres/química , Micotoxinas/síntese química , Oligopeptídeos/síntese química , Micotoxinas/química , Espectroscopia de Prótons por Ressonância MagnéticaRESUMO
Recently, deoxynivalenol-3-sulfate (DON-3-sulfate) was proposed as a major DON metabolite in poultry. In the present work, the first LC-MS/MS based method for determination of DON-3-sulfate, deepoxy-DON-3-sulfate (DOM-3-sulfate), DON, DOM, DON sulfonates 1, 2, 3, and DOM sulfonate 2 in excreta samples of chickens and turkeys was developed and validated. To this end, DOM-3-sulfate was chemically synthesized and characterized by NMR and LC-HR-MS/MS measurements. Application of the method to excreta and chyme samples of four feeding trials with turkeys, chickens, pullets, and roosters confirmed DON-3-sulfate as the major DON metabolite in all poultry species studied. Analogously to DON-3-sulfate, DOM-3-sulfate was formed after oral administration of DOM both in turkeys and in chickens. In addition, pullets and roosters metabolized DON into DOM-3-sulfate. In vitro transcription/translation assays revealed DOM-3-sulfate to be 2000 times less toxic on the ribosome than DON. Biological recoveries of DON and DOM orally administered to broiler chickens, turkeys, and pullets were 74%-106% (chickens), 51%-72% (roosters), and 131%-151% (pullets). In pullets, DON-3-sulfate concentrations increased from jejunum chyme samples to excreta samples by a factor of 60. This result, put into context with earlier studies, indicates fast and efficient absorption of DON between crop and jejunum, conversion to DON-3-sulfate in intestinal mucosa, liver, and possibly kidney, and rapid elimination into excreta via bile and urine.
Assuntos
Galinhas/metabolismo , Micotoxinas/farmacocinética , Tricotecenos/farmacocinética , Perus/metabolismo , Animais , Biotransformação , Fezes/química , Feminino , Jejuno/química , Jejuno/metabolismo , Espectroscopia de Ressonância Magnética , Masculino , Micotoxinas/síntese química , Micotoxinas/toxicidade , Reprodutibilidade dos Testes , Sulfatos/metabolismo , Distribuição Tecidual , Tricotecenos/síntese química , Tricotecenos/toxicidadeRESUMO
Conjugation of deoxynivalenol (DON) with sulfur compounds is recognized as a significant reaction pathway, and putative DON-glutathione (DON-GSH) conjugates have been reported in planta. To understand and control the reaction of trichothecenes with biologically important thiols, we studied the reaction of DON, T-2 tetraol, and de-epoxy-DON with a range of model thiols. Reaction conditions were optimized for DON with 2-mercaptoethanol. Major reaction products were identified using HRMS and NMR spectroscopy. The results indicate that thiols react reversibly with the double bond (Michael addition) and irreversibly with the epoxide group in trichothecenes. These reactions occurred at different rates, and multiple isomers were produced including diconjugated forms. LC-MS analyses indicated that glutathione and cysteine reacted with DON in a similar manner to the model thiols. In contrast to DON, none of the tested mercaptoethanol adducts displayed toxicity in human monocytes or induced pro-inflammatory cytokines in human macrophages.
Assuntos
Micotoxinas/química , Compostos de Sulfidrila/química , Tricotecenos/química , Linhagem Celular , Citocinas/imunologia , Humanos , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Micotoxinas/síntese química , Micotoxinas/farmacologia , Tricotecenos/síntese química , Tricotecenos/farmacologiaRESUMO
The stereoselective synthesis of a variety of functionalized hydroindoles suitable as building blocks for thiodiketopiperazine natural products such as rostratins B-D is reported. The key precursor for all transformations is a previously reported hexahydroindole compound. All functional groups were installed with the desired stereochemistry and the feasibility of the synthetic strategy was exemplified by dimerization of two hydroindole units to form the pentacyclic C2 -symmetric scaffold of rostratinâ C.
Assuntos
Produtos Biológicos/síntese química , Indóis/síntese química , Micotoxinas/síntese química , Produtos Biológicos/química , Indóis/química , Micotoxinas/química , Piperazinas , EstereoisomerismoRESUMO
Some hybrids of vinca alkaloids and phomopsin A, linked by a glycine pattern, have been synthesized in one or two steps, by an insertion reaction and shown to inhibit microtubule assembly. These compounds have been elaborated in order to interact with both the "vinca site" and the "peptide site" of the vinca domain in tubulin. Two out of three hybrids are potent inhibitors of microtubules assembly and they present good cytotoxicity against different cell lines. Molecular modelling studies show that they could bind, within the vinca domain, in similar spatial regions as those of vinca and phomopsin thanks to the flexibility provided by the glycine linker used to elaborate these hybrids.
Assuntos
Glicina/química , Micotoxinas/síntese química , Tubulina (Proteína)/química , Alcaloides de Vinca/síntese química , Alcaloides/química , Apoptose , Sítios de Ligação , Linhagem Celular , Guanosina Trifosfato/química , Humanos , Células K562 , Microtúbulos/metabolismo , Modelos Moleculares , Micotoxinas/química , Peptídeos/química , Estrutura Terciária de Proteína , Transdução de Sinais , Vimblastina/análogos & derivados , Vimblastina/química , Alcaloides de Vinca/química , VinorelbinaRESUMO
Enantiocontrolled construction of B-E rings of penitrem E was accomplished from 4-iodoindole in 13 steps with an overall yield of 1.7%. Diastereoselective Tf2NH-catalyzed (2+2)-cycloaddition between silyl enol ether and methyl acrylate furnished a tetracyclic product possessing the characteristic cyclobutane ring bearing a hydroxyl group.
Assuntos
Micotoxinas/síntese química , Técnicas de Química Sintética , Cristalografia por Raios X , Estrutura Molecular , Micotoxinas/química , EstereoisomerismoRESUMO
Methylthiodeoxynivalenol (MTD), a novel derivative of the trichothecene mycotoxin deoxynivalenol (DON), was prepared by applying a reliable procedure for the formal Michael addition of methanethiol to the conjugated double bond of DON. Structure elucidation revealed the preferred formation of the hemiketal form of MTD by intramolecular cyclisation between C8 and C15. Computational investigations showed a negative total reaction energy for the hemiketalisation step and its decrease in comparison with theoretical model compounds. Therefore, this structural behaviour seems to be a general characteristic of thia-Michael adducts of type B trichothecenes. MTD was shown to be less inhibitory for a reticulocyte lysate based in vitro translation system than the parent compound DON, which supports the hypothesis that trichothecenes are detoxified through thia-adduct formation during xenobiotic metabolism.
Assuntos
Micotoxinas/síntese química , Micotoxinas/toxicidade , Reticulócitos/metabolismo , Tricotecenos/síntese química , Tricotecenos/toxicidade , Animais , Biotransformação , Sistema Livre de Células , Ciclização , Genes Reporter , Luciferases/genética , Luciferases/metabolismo , Micotoxinas/metabolismo , Biossíntese de Proteínas/efeitos dos fármacos , Coelhos , Reticulócitos/citologia , Reticulócitos/efeitos dos fármacos , Compostos de Sulfidrila/química , Termodinâmica , Tricotecenos/química , Tricotecenos/metabolismoRESUMO
Hybrids of vinca alkaloids and phomopsin A have been elaborated with the aim of interfering with the "vinca site" and the "peptide site" of the vinca domain in tubulin. They were synthesized by an efficient one-pot procedure that directly links the octahydrophomopsin lateral chain to the velbenamine moiety of 7'-homo-anhydrovinblastine. In their modeled complexes with tubulin, these hybrids were found to superimpose nicely on the tubulin-bound structures of vinblastine and phomopsin A. This good matching can account for the fact that two of them are very potent inhibitors of microtubules assembly and are cytotoxic against four cancer cell lines.
Assuntos
Antineoplásicos/síntese química , Micotoxinas/síntese química , Vimblastina/análogos & derivados , Vimblastina/síntese química , Antineoplásicos/química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Modelos Moleculares , Micotoxinas/química , Micotoxinas/farmacologia , Tubulina (Proteína)/química , Moduladores de Tubulina/síntese química , Moduladores de Tubulina/química , Moduladores de Tubulina/farmacologia , Vimblastina/química , Vimblastina/farmacologiaRESUMO
Penitrems are indole diterpene alkaloids best known for their BK channel inhibition and tremorgenic effects in mammals. In a previous study, penitrems A-F (1-5), their biosynthetic precursors, paspaline (6) and emindole SB (7), and two brominated penitrem analogs 8 and 9 demonstrated promising in vitro antiproliferative, antimigratory, and anti-invasive effects in the MTT (MCF-7 and MDA-MB-231), wound-healing, and Cultrex BME cell invasion (MDA-MB-231) assays, respectively. The study herein reports the novel ability of penitrem A to suppress total ß-catenin levels in MDA-MB-231 mammary cancer cells. Nine new penitrem analogs (10-18) were semisynthetically prepared, in an attempt to identify pharmacophores correlated with BK channel inhibition and tremorgenicity of penitrems and decrease their toxicity. The degree of BK channel inhibition was assessed using the nematode Caenorhabditis elegans, and in vivo tremorgenic EC50 was calculated using CD-1 male mice following an Up-and-Down Procedure (UDP). Although new analogs were generally less active than parent compound 1, some showed no BK channel inhibition or tremorgenicity and retained the ability of penitrem A (1) to suppress total ß-catenin levels in MDA-MB-231 cells. Paspaline (6) and emindole SB (7), both lacking BK channel inhibition and tremorgenicity, represent the simplest indole diterpene skeleton that retains the antiproliferative, antimigratory and total ß-catenin suppressing effects shown by the more complex penitrem A (1).
Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Diterpenos/farmacologia , Alcaloides Indólicos/farmacologia , Micotoxinas/farmacologia , Proteínas Wnt/antagonistas & inibidores , beta Catenina/antagonistas & inibidores , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Caenorhabditis elegans/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Diterpenos/síntese química , Diterpenos/química , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Humanos , Alcaloides Indólicos/síntese química , Alcaloides Indólicos/química , Células MCF-7 , Masculino , Camundongos , Estrutura Molecular , Micotoxinas/síntese química , Micotoxinas/química , Relação Estrutura-Atividade , Proteínas Wnt/metabolismo , beta Catenina/metabolismoAssuntos
Produtos Biológicos/síntese química , Fungos/química , Micotoxinas/síntese química , Aflatoxinas/síntese química , Aflatoxinas/isolamento & purificação , Antraquinonas/síntese química , Antraquinonas/isolamento & purificação , Produtos Biológicos/isolamento & purificação , Citrinina/síntese química , Citrinina/isolamento & purificação , Citocalasinas/síntese química , Citocalasinas/isolamento & purificação , Alcaloides de Claviceps/síntese química , Alcaloides de Claviceps/isolamento & purificação , Fumonisinas/síntese química , Fumonisinas/isolamento & purificação , Fungos/metabolismo , Lactonas/síntese química , Lactonas/isolamento & purificação , Micotoxinas/isolamento & purificação , Ocratoxinas/síntese química , Ocratoxinas/isolamento & purificação , Patulina/síntese química , Patulina/isolamento & purificação , Tricotecenos/síntese química , Tricotecenos/isolamento & purificação , Xantonas/síntese química , Xantonas/isolamento & purificaçãoRESUMO
4-Deoxynivalenol is one of the most prevalent mycotoxins in grain-based food and feed products worldwide. Conjugation of deoxynivalenol to glucuronic acid and elimination via the urine appears to be the major metabolism pathway, although with differing efficiency in different species. In order to make pure deoxynivalenol glucuronides for analytical methodologies available we intended to enzymatically synthesize glucuronides of deoxynivalenol using rat and human liver microsomes supplemented with uridine 5'-diphosphoglucuronic acid and alamethicin as detergent. Three glucuronides were isolated and purified using solid-phase extraction of microsomal incubations and subsequent semipreparative hydrophilic interaction chromatography. NMR spectra were obtained for all three compounds from solutions in methanol, showing that deoxynivalenol 3-O-ß-D-glucuronide and deoxynivalenol 15-O-ß-D-glucuronide were the major products from incubations of deoxynivalenol with rat and human liver microsomes, respectively. The NMR spectra of a third glucuronide showed replacement of the C-8 carbonyl by a ketal carbon. This glucuronide was finally identified as deoxynivalenol 8-O-ß-D-glucuronide. The present study provides unequivocal structural evidence for three glucuronides of deoxynivalenol formed by liver enzymes.
Assuntos
Enzimas/química , Glucuronídeos/síntese química , Micotoxinas/química , Tricotecenos/síntese química , Animais , Glucuronídeos/química , Humanos , Masculino , Microssomos Hepáticos/enzimologia , Estrutura Molecular , Micotoxinas/síntese química , Ratos , Ratos Wistar , Tricotecenos/químicaRESUMO
A unified synthetic strategy directed towards mycotoxins belonging to the thiodiketopiperazine family is reported. The building blocks for a number of natural products--including exserohilone, gliotoxin, the epicoccins, the epicorazines, rostratinâ A and aranotin--have been synthesised stereoselectively from a common precursor. This key intermediate was constructed through an efficient and highly diastereoselective [2+2] cycloaddition between a ketene and an enecarbamate derived from L-pyroglutamic acid. The annelation of the second ring was accomplished through ring-closing metathesis and enol ether-olefin ring-closing metathesis to provide both cis- and trans-annelated azabicyclic cyclohexenones, as well as an annelated seven-membered cyclic enol ether. A Pd-catalysed elimination of allyl acetate gave rise to the cyclohexadienol structure of gliotoxin. Dimerisation of one building block to afford the diketopiperazine core was demonstrated.
Assuntos
Produtos Biológicos/síntese química , Gliotoxina/síntese química , Piperazinas/química , Produtos Biológicos/química , Catálise , Cristalografia por Raios X , Reação de Cicloadição , Cicloexenos/química , Etilenos/química , Gliotoxina/química , Cetonas/química , Conformação Molecular , Micotoxinas/síntese química , Micotoxinas/química , Paládio/química , Piperazinas/síntese química , Ácido Pirrolidonocarboxílico/química , EstereoisomerismoRESUMO
Nine simplified vinca alkaloids and phomospin A hybrids, in which vindoline moiety has been replaced by a simpler scaffold, have been elaborated to evaluate their activity on the inhibition of tubulin polymerization. This article deals with the synthesis of various simplified vinca alkaloids, using a stereoselective coupling of catharantine with reactive aromatic compounds and methanol as well as their subsequent condensation with a large peptide chain mimicking those of phomopsin A. Biological evaluation and molecular modeling studies are also reported.
Assuntos
Micotoxinas/química , Vimblastina/análogos & derivados , Alcaloides de Vinca/química , Metanol/química , Micotoxinas/síntese química , Estereoisomerismo , Termodinâmica , Tubulina (Proteína)/química , Tubulina (Proteína)/metabolismo , Moduladores de Tubulina/síntese química , Moduladores de Tubulina/química , Vimblastina/química , Alcaloides de Vinca/síntese química , Alcaloides de Vinca/farmacologiaRESUMO
Longianone and the biosynthetically related mycotoxins isopatulin and (Z)-ascladiol were prepared following a divergent route from a readily available furan diol. The route toward longianone features an unprecedented TBAF-promoted intramolecular oxa-Michael reaction to a conjugated keto enoate, and the oxidation of dihydrolongianone to longianone with stabilized IBX. The route to isopatulin features a chemoenzymatic synthesis of (Z)-ascladiol, and the regioselective oxidation of (Z)-ascladiol to isopatulin with MnO(2).
Assuntos
Furanos/química , Micotoxinas/síntese química , Micotoxinas/isolamento & purificação , Furanos/síntese química , Furanos/isolamento & purificação , Compostos Heterocíclicos com 2 Anéis/síntese química , Compostos Heterocíclicos com 2 Anéis/química , Compostos Heterocíclicos com 2 Anéis/isolamento & purificação , Lactonas/síntese química , Lactonas/química , Lactonas/isolamento & purificação , Micotoxinas/química , Oxirredução , Compostos de Espiro/síntese química , Compostos de Espiro/química , Compostos de Espiro/isolamento & purificação , EstereoisomerismoRESUMO
We have isolated the small, highly strained carboxylic acid cycloprop-2-ene carboxylic acid from the Asian toxic mushroom Russula subnigricans. This compound is responsible for fatal rhabdomyolysis, a new type of mushroom poisoning that is indicated by an increase in serum creatine phosphokinase activity in mice. We found that polymerization of the compound at high concentrations via ene reaction abolishes its toxicity.