RESUMO
A comprehensive linear gradient solvent system for centrifugal partition chromatography (CPC) was developed for the bioassay-guided isolation of natural compounds. The gradient solvent system consisted of three different ternary biphasic solvents types: n-hexane-acetonitrile-water (10:2:8, v/v), ethyl acetate-acetonitrile-water (10:2:8, v/v), and water-saturated n-butanol-acetonitrile-water (10:2:8, v/v). The lower phase of the n-hexane-acetonitrile-water (10:2:8, v/v) was used as the stationary phase, while its upper phase, as well as ethyl acetate-acetonitrile-water (10:2:8), and water-saturated n-butanol-acetonitrile-water (10:2:8, v/v) were pumped to generate a linear gradient elution, increasing the mobile phase polarity. We used the gradient CPC to identify antioxidant response elements (AREs), inducing compounds from Centipeda minima, using an ARE-luciferase assay in HepG2 cells, which led to the purification of the active molecules 3-methoxyquercetin and brevilin A. The developed CPC solvent systems allow the separation and isolation of compounds with a wide polarity range, allowing active molecule identification in the complex crude extract of natural products.
Assuntos
Asteraceae/química , Cromatografia Líquida/métodos , Distribuição Contracorrente/métodos , Extratos Vegetais/análise , Solventes/química , 1-Butanol/química , Acetatos/química , Acetonitrilas/química , Elementos de Resposta Antioxidante/efeitos dos fármacos , Bioensaio , Sobrevivência Celular/efeitos dos fármacos , Cromatografia Líquida/instrumentação , Distribuição Contracorrente/instrumentação , Crotonatos/isolamento & purificação , Genes Reporter/efeitos dos fármacos , Células Hep G2 , Hexanos/química , Humanos , Luciferases/química , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Quercetina/análogos & derivados , Quercetina/isolamento & purificação , Sesquiterpenos/isolamento & purificação , Água/químicaRESUMO
ABSTRACT This study aims to identify special metabolites in polar extracts from Urochloa humidicola (synonym Brachiaria humidicola) that have allelopathic effects and induce secondary photosensitization in ruminants. The compounds were isolated and identified via chromatographic and spectroscopic techniques. The compounds 4-hydroxy-3-methoxy-benzoic acid, trans-4-hydroxycinnamic acid, and p-hydroxy-benzoic acid; the flavonols isorhamnetin-3-O-β-d-glucopyranoside and methyl quercetin-3-O-β-d-glucuronate; and kaempferitrin, quercetin-3-O-α-l-rhamnopyranoside, and tricin were identified in the extract from the leaves of Urochloa humidicola. Two furostanic saponins, namely, dioscin and 3-O-α-l-rhamnopyranosyl-(1-4)-[α-l-rhamnopyranosyl-(1-2)]-β-d-glucopyranosyl-penogenin, as well as catechin-7-O-β-d-glucopyranoside were identified in the methanolic extract obtained from the roots of this plant. This species features a range of metabolites that may be toxic for animals if used in food and may interfere with the growth medium, thereby inhibiting the development of other species.
Assuntos
Flavonoides/isolamento & purificação , Extratos Vegetais/química , Brachiaria/química , Parabenos/isolamento & purificação , Parabenos/química , Saponinas/química , Ácido Vanílico/química , Flavonoides/química , Crotonatos/isolamento & purificação , Crotonatos/química , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Cromatografia em Camada Fina , Glicosídeos/isolamento & purificação , Glicosídeos/químicaRESUMO
Honaucins A-C were isolated from the cyanobacterium Leptolyngbya crossbyana which was found overgrowing corals on the Hawaiian coast. Honaucin A consists of (S)-3-hydroxy-γ-butyrolactone and 4-chlorocrotonic acid, which are connected via an ester linkage. Honaucin A and its two natural analogs exhibit potent inhibition of both bioluminescence, a quorum-sensing-dependent phenotype, in Vibrio harveyi BB120 and lipopolysaccharide-stimulated nitric oxide production in the murine macrophage cell line RAW264.7. The decrease in nitric oxide production was accompanied by a decrease in the transcripts of several proinflammatory cytokines, most dramatically interleukin-1ß. Synthesis of honaucin A, as well as a number of analogs, and subsequent evaluation in anti-inflammation and quorum-sensing inhibition bioassays revealed the essential structural features for activity in this chemical class and provided analogs with greater potency in both assays.
Assuntos
4-Butirolactona/análogos & derivados , Antibacterianos/química , Anti-Inflamatórios/química , Crotonatos/química , Cianobactérias/química , Percepção de Quorum/efeitos dos fármacos , 4-Butirolactona/química , 4-Butirolactona/isolamento & purificação , 4-Butirolactona/farmacologia , Animais , Antibacterianos/isolamento & purificação , Antibacterianos/farmacologia , Anti-Inflamatórios/isolamento & purificação , Anti-Inflamatórios/farmacologia , Linhagem Celular , Crotonatos/isolamento & purificação , Crotonatos/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos , Dados de Sequência Molecular , Óxido Nítrico/imunologia , Relação Estrutura-Atividade , Vibrio/efeitos dos fármacos , Vibrio/fisiologia , Vibrioses/tratamento farmacológicoRESUMO
Deamination of UV filters, such as kynurenine (KN), in the human lens results in protein modification. Thermal reactions of the product of kynurenine deamination, 4-(2-aminophenyl)-4-oxocrotonic acid (CKA), with amino acids (histidine, lysine, methionine, tryptophan, tyrosine, cysteine) and antioxidants (ascorbate, NADH, glutathione reduced) were studied. The rate constants of the reactions under physiological conditions were measured. The rate constants of CKA addition to cysteine k(Cys)=36+/-4M(-1)s(-1) and to glutathione k(GSH)=2.1+/-0.2M(-1)s(-1) are 4-5 orders of magnitude higher than the rate constants of CKA reactions with the other amino acids and antioxidants. The Arrhenius parameters for k(Cys) and k(GSH) were determined: A(GSH)=(1.8+/-0.7)x10(5)M(-1)s(-1), E(GSH)=29.2+/-5.6kJmol(-1), A(Cys)=(2.7+/-0.9)x10(8)M(-1)s(-1), E(Cys)=40.4+/-5.7kJmol(-1). The large difference in frequency factors for k(Cys) and k(GSH) is attributed to steric hindrance, peculiar to the bulky GSH molecule.
Assuntos
Aminoácidos/química , Antioxidantes/química , Crotonatos/química , Proteínas do Olho/química , Cristalino/química , Cromatografia Líquida de Alta Pressão/métodos , Crotonatos/isolamento & purificação , Desaminação , Humanos , Cinuramina/química , Ligação Proteica , Espalhamento de Radiação , Raios UltravioletaAssuntos
Carcinógenos , Crotonatos/toxicidade , Diterpenos/toxicidade , Ácidos Graxos Monoinsaturados/toxicidade , Irritantes , Plantas/análise , Animais , Carcinógenos/isolamento & purificação , Fenômenos Químicos , Química , Crotonatos/isolamento & purificação , Diterpenos/isolamento & purificação , Ésteres , Hemiterpenos , Irritantes/isolamento & purificação , Camundongos , Estrutura MolecularRESUMO
The structure of a glyoxalase I inhibitor (I), isolated from a cultured broth of Streptomyces griseosporeus, was found to be 2-crotonyloxymethyl-4,5,6-trihydroxy-cyclohex-2-enone by chemical studies. Stereochemistry and absolute configuration were determined to be 4R, 5R and 6R by X-ray crystallographic analysis of a bromine-containing crystalline derivative. The crotonyloxy group of I shows a surprising proclivity to be displaced by SH-compounds. This property is shown to be the basis for its biological activity.