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1.
PLoS One ; 15(7): e0227529, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32730337

RESUMO

The pretreatment of biomass remains a critical requirement for bio-renewable fuel production from lignocellulose. Although current processes primarily involve chemical and physical approaches, the biological breakdown of lignin using enzymes and microorganisms is quickly becoming an interesting eco-friendly alternative to classical processes. As a result, bioprospection of wild fungi from naturally occurring lignin-rich sources remains a suitable method to uncover and isolate new species exhibiting ligninolytic activity. In this study, wild species of white rot fungi were collected from Colombian forests based on their natural wood decay ability and high capacity to secrete oxidoreductases with high affinity for phenolic polymers such as lignin. Based on high activity obtained from solid-state fermentation using a lignocellulose source from oil palm as matrix, we describe the isolation and whole-genome sequencing of Dictyopanus pusillus, a wild basidiomycete fungus exhibiting ABTS oxidation as an indication of laccase activity. Functional characterization of a crude enzymatic extract identified laccase activity as the main enzymatic contributor to fungal extracts, an observation supported by the identification of 13 putative genes encoding for homologous laccases in the genome. To the best of our knowledge, this represents the first report of an enzymatic extract exhibiting laccase activity in the Dictyopanus genera, offering means to exploit this species and its enzymes for the delignification process of lignocellulosic by-products from oil palm.


Assuntos
Agaricales/genética , Genoma Fúngico , Lignina/metabolismo , Óleo de Palmeira/metabolismo , Agaricales/classificação , Agaricales/enzimologia , Biomassa , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Concentração de Íons de Hidrogênio , Lacase/genética , Lacase/metabolismo , Oxirredução , Filogenia , Temperatura , Sequenciamento Completo do Genoma
2.
Curr Pharm Biotechnol ; 21(2): 169-178, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31612825

RESUMO

BACKGROUND: Melanin protects the skin against the harmful effects of ultraviolet irradiation. However, melanin overproduction can result in several aesthetic problems, including melasma, freckles, age spots and chloasma. Therefore, development of anti-melanogenic agents is important for the prevention of serious hyperpigmentation diseases. Sesamolin is a lignan compound isolated from sesame seeds with several beneficial properties, including potential for melanin inhibition. OBJECTIVE: The aim of this study was to evaluate the anti-melanogenic effect of sesamolin in cell culture in vitro and the underlying mechanism of inhibition using molecular docking simulation. METHODS: Melanogenesis was induced by 3-isobutyl-1-methylxanthine in B16F10 melanoma cells, and the inhibitory effects of sesamolin were evaluated using zymography, a tyrosinase inhibitory activity assay, western blotting, and real-time reverse transcription-polymerase chain reaction analysis. Docking simulations between sesamolin and tyrosinase were performed using Autodock vina. RESULTS: Sesamolin significantly inhibited the expression of melanogenesis-related factors tyrosinase, and tyrosinase-related proteins 1 and 2 at the mRNA and protein levels. Treatment of melanoma cells with 50 µM sesamolin demonstrated the strongest inhibition against intercellular tyrosinase and melanin synthesis without exerting cytotoxic effects. Sesamolin significantly reduced mushroom tyrosinase activity in a dose-dependent manner via a competitive inhibition mechanism. Tyrosinase docking simulations supported that sesamolin (-6.5 kcal/mol) bound to the active site of tyrosinase more strongly than the positive control (arbutin, -5.7 kcal/mol). CONCLUSION: Sesamolin could be developed as a melanogenesis inhibiting agent owing to its dual function in blocking the generation of melanogenesis-related enzymes and inhibiting the enzymatic response of tyrosinase.


Assuntos
Antineoplásicos/farmacologia , Dioxóis/farmacologia , Oxirredutases Intramoleculares/antagonistas & inibidores , Melaninas/biossíntese , Monofenol Mono-Oxigenase/antagonistas & inibidores , Oxirredutases/antagonistas & inibidores , 1-Metil-3-Isobutilxantina , Agaricales/enzimologia , Animais , Antineoplásicos/isolamento & purificação , Linhagem Celular Tumoral , Dioxóis/isolamento & purificação , Relação Dose-Resposta a Droga , Humanos , Melanoma Experimental/patologia , Simulação de Acoplamento Molecular
3.
Food Chem ; 303: 125310, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31473456

RESUMO

Tyrosinase is a metalloenzyme involved in o-hydroxylation of monophenols and oxidation of o-diphenols to o-quinones, with formation of brown or black pigments (melanines). Tyrosinase inhibitors are of great interest in medicine and cosmetics (skin whitening compounds), but also in food and beverage industry (antibrowning agents). Here we report on the activity as mushroom tyrosinase inhibitors of a series of hydroxyphenyl thiosemicarbazones (1-5): one of them revealed an inhibitory activity stronger than kojic acid, used as reference. Enzymatic inhibition activity was confirmed by colorimetric measurements on small wheels of Fuji apples treated with the hydroxyphenyl thiosemicarbazones. The mechanism of action of compounds 1-5 was investigated by molecular modelling and by studying in solution their speciation with Cu(II) ions, the ions in the active site of the enzyme. Finally, compounds 1-5 were tested on human fibroblasts: they are not cytotoxic and they do not activate cells in a pro-inflammatory way.


Assuntos
Agaricales/enzimologia , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Reação de Maillard/efeitos dos fármacos , Monofenol Mono-Oxigenase/antagonistas & inibidores , Tiossemicarbazonas/química , Tiossemicarbazonas/farmacologia , Domínio Catalítico , Humanos , Cinética , Monofenol Mono-Oxigenase/química , Monofenol Mono-Oxigenase/metabolismo , Oxirredução/efeitos dos fármacos
4.
Eur J Med Chem ; 184: 111738, 2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31655335

RESUMO

The anti-melanogenic principle of peony (Paeonia officinalis subsp. officinalis) seeds was elucidated via activity-guided isolation. Resveratroloside (trans-resveratrol-4'-O-beta-d-glucopyranoside) was found to be the main metabolite of P. officinalis subsp. officinalis seeds and its tyrosinase inhibiting activity was confirmed via an enzymatic assay. Furthermore, the in vitro activity and the therapeutic window were studied employing the murine melanoma cell line B16F10. The results from the conducted stability assay and the high content of resveratroloside in the seeds (i.e. 10.4% dw) motivated us to push the extract forward to an in vivo tolerance assay. A clinical study with forty Caucasian participants proofed a good skin-tolerance with high moisture effect and reduction of pores.


Assuntos
Cosmecêuticos/farmacologia , Descoberta de Drogas , Monofenol Mono-Oxigenase/antagonistas & inibidores , Paeonia/química , Extratos Vegetais/farmacologia , Adulto , Agaricales/enzimologia , Idoso , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cosmecêuticos/química , Cosmecêuticos/isolamento & purificação , Relação Dose-Resposta a Droga , Feminino , Humanos , Luz , Masculino , Camundongos , Pessoa de Meia-Idade , Estrutura Molecular , Monofenol Mono-Oxigenase/metabolismo , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Sementes/química , Pele/efeitos dos fármacos , Pele/patologia , Relação Estrutura-Atividade
5.
J Agric Food Chem ; 67(38): 10744-10755, 2019 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-31525900

RESUMO

We previously reported that ß-glucosidase BGL1 at low concentration (15 µg mL-1) from Coprinopsis cinerea exhibited hydrolytic activity only toward laminarioligosaccharides but not toward cellooligosaccharides and gentiobiose. This study shows that BGL1 at high concentration (200 µg mL-1) also hydrolyzed cellobiose and gentiobiose, which accounted for only 0.83 and 2.05% of its activity toward laminaribiose, respectively. Interestingly, BGL1 at low concentration (1.5 µg mL-1) showed transglycosylation but BGL1 at high concentration (200 µg mL-1) did not. BGL1 utilizes only laminarioligosaccharides but not glucose, gentiobiose, and cellobiose to synthesize the higher oligosaccharides. BGL1 transferred one glucosyl residue from substrate laminarioligosaccharide to another laminarioligosaccharide as an acceptor in a ß(1 → 3) or ß(1 → 6) fashion to produce higher laminarioligosaccharides or 3-O-ß-d-gentiobiosyl-d-laminarioligosaccharides. The BGL1-digested laminaritriose exhibited approximately 90% enhancement in the anti-oxidant activity compared to that of untreated laminaritriose, implying a potential application of BGL1-based transglycosylation for the production of high value-added rare oligosaccharides.


Assuntos
Agaricales/enzimologia , Dissacarídeos/metabolismo , Proteínas Fúngicas/química , Oligossacarídeos/metabolismo , beta-Glucosidase/química , Agaricales/química , Agaricales/genética , Sequência de Aminoácidos , Dissacarídeos/química , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Glicosilação , Hidrólise , Cinética , Estrutura Molecular , Oligossacarídeos/química , Especificidade por Substrato , beta-Glucosidase/genética , beta-Glucosidase/metabolismo
6.
Int J Mol Sci ; 20(19)2019 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-31561511

RESUMO

The organogermanium compound 3-(trihydroxygermyl)propanoic acid (THGP) has various biological activities. We previously reported that THGP forms a complex with cis-diol structures. L-3,4-Dihydroxyphenylalanine (L-DOPA), a precursor of melanin, contains a cis-diol structure in its catechol skeleton, and excessive melanin production causes skin darkening and staining. Thus, the cosmetic field is investigating substances that suppress melanin production. In this study, we investigated whether THGP inhibits melanin synthesis via the formation of a complex with L-DOPA using mushroom tyrosinase and B16 4A5 melanoma cells. The ability of THGP to interact with L-DOPA was analyzed by 1H-NMR, and the influence of THGP and/or kojic acid on melanin synthesis was investigated. We also examined the effect of THGP on cytotoxicity, tyrosinase activity, and gene expression and found that THGP interacted with L-DOPA, a precursor of melanin with a cis-diol structure. The results also showed that THGP inhibited melanin synthesis, exerted a synergistic effect with kojic acid, and did not affect tyrosinase activity or gene expression. These results suggest that THGP is a useful substrate that functions as an inhibitor of melanogenesis and that its effect is enhanced by combination with kojic acid.


Assuntos
Agaricales/enzimologia , Levodopa/química , Levodopa/farmacologia , Melaninas/biossíntese , Monofenol Mono-Oxigenase/antagonistas & inibidores , Compostos Organometálicos/química , Compostos Organometálicos/farmacologia , Animais , Vias Biossintéticas/efeitos dos fármacos , Melanoma Experimental , Camundongos
7.
Drug Des Devel Ther ; 13: 2169-2178, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31371919

RESUMO

Background and aim: Tyrosinase (EC 1.14.18.1) is responsible for enzymatic browning in fruits and vegetables. Its inhibitors may be applied to efficiently treat hyperpigmentation and are widely used in pharmaceutical and cosmetic products, food supplements and insecticides. Previous studies have shown that heterocyclic compounds with an amino group can inhibit tyrosinase activity. The present study aims to evaluate the inhibitory effect of some novel 2,6-diamino-4-chloropyrimidine derivatives (1a-e) and 2,4,6-triaminopyrimidine (2a-e) including bioactive aniline moiety on the activity of the mushroom tyrosinase. Methods: In practice, the azo salt was initially synthesized from aniline derivatives and combined subsequently with the 2,4,6-triaminopyrimidine and 2,6-diamino-4 chloropyrimidine followed by crystallization. The structures of resulting compounds were confirmed by FT-IR, 13C NMR, and 1H NMR. The derivatives (0-100 µM) were evaluated for their inhibitory effect on tyrosinase activity using l-3,4-dihydroxyphenylalanine (l-DOPA) as substrate. Results: All compounds showed inhibitory effects against the activity of the enzyme. About 23.72-55.08% inhibition was observed in the presence of 30 µM of each compound. The IC50 values of the synthesized compounds were measured, and their inhibition properties were also visualized by zymography. Based on the results, the compounds 1a-e and 2a-e showed moderate inhibitory activities. Notably, pyrimidine derivatives 1a (IC50=24.68) and 1d (IC50=24.45) also exhibited similar inhibitory activities when compared with the positive control, kojic acid (IC50=25.24 µM). Kinetic studies indicated that the type of inhibition was noncompetitive. Conclusion: All results suggest that pyrimidine derivatives, especially 1d and 1a, can be considered as safe and efficient tyrosinase inhibitors.


Assuntos
Agaricales/enzimologia , Inibidores Enzimáticos/farmacologia , Monofenol Mono-Oxigenase/antagonistas & inibidores , Pirimidinas/farmacologia , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Simulação de Acoplamento Molecular , Estrutura Molecular , Monofenol Mono-Oxigenase/metabolismo , Pirimidinas/síntese química , Pirimidinas/química , Relação Estrutura-Atividade
8.
Talanta ; 204: 525-532, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31357329

RESUMO

In this work, we present a smartphone-based multiplexed enzymatic biosensor utilizing the unique colorimetric properties of the poly(aniline-co-anthranilic acid) (ANI-co-AA) composite film coupled with horseradish peroxidase (HRP), glucose oxidase (GOx), horseradish peroxidase-glucose oxidase (GOx-HRP) and tyrosinase (Tyr) enzymes. The enzymes are immobilized on the composite polymer film by adsorption and they catalyze a reversible redox color change of the host polymer from green to blue in the presence of their substrate. A smartphone was applied as color detector, for image acquisition and data handling. A ColorLab® android application, free of charge software application, was used to enable easy and clear display of the sensors' response indicating remarkable changes in the optical features. The results were confirmed by the spectrophotometric measurements. The developed colorimetric enzymatic biosensors were studied and optimized in relation to different experimental parameters. Moreover, the colorimetric enzymatic biosensors were applied to food and pharmaceutical analysis. It has been shown by these studies that the colorimetric biosensors are promising as quick and simple tests for handheld analysis in various fields.


Assuntos
Catecóis/análise , Glucose/análise , Peróxido de Hidrogênio/análise , Agaricales/enzimologia , Armoracia/enzimologia , Técnicas Biossensoriais/métodos , Colorimetria/instrumentação , Colorimetria/métodos , Enzimas Imobilizadas/química , Sucos de Frutas e Vegetais/análise , Glucose Oxidase/química , Peroxidase do Rábano Silvestre/química , Limite de Detecção , Monofenol Mono-Oxigenase/química , Polímeros/química , Romã (Fruta)/química , Pyrus/química , Reprodutibilidade dos Testes , Smartphone , Vinho/análise
9.
Prep Biochem Biotechnol ; 49(9): 908-915, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31271327

RESUMO

Pentachlorophenol (PCP) is an organochlorine pesticide whose toxicity led it to be banned in several countries. In Brazil however, this compound is widely accessible, and its indiscriminate use leads to extensive soil contamination, which requires henceforth, the development of new approaches to manage PCP presence in environment. Considering that PCP is susceptible to undergo enzymatic degradation, this investigation is thence, aimed at the purification, and characterization, of a thermostable fungal enzyme (i.e. Laccase of Deconica castanella (Dc-Lac), and assess its role in PCP in vitro biodegradation. Results evidenced that, molecular mass of the partially purified Dc-Lac was estimated to be of 64 kDa, presenting apparent Km of 0.47 µmol, and Vmax of 11.56 U mg-1. The optimum temperature and pH were 55 °C and 2.5, respectively. The T½ verified at 55, 60, and 80 °C were 19 and 17 hr, and 47 min, respectively. The highest PCP biodegradation was of 23% at pollutant concentration of 100 µg L-1, which evidenced that Dc-Lac is a thermostable enzyme that acted directly in PCP degradation and may be a useful asset to remediate this pollutant.


Assuntos
Agaricales/enzimologia , Poluentes Ambientais/metabolismo , Lacase/metabolismo , Pentaclorofenol/metabolismo , Praguicidas/metabolismo , Agaricales/química , Agaricales/metabolismo , Biodegradação Ambiental , Estabilidade Enzimática , Lacase/química , Temperatura
10.
J Microbiol Biotechnol ; 29(8): 1204-1211, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31336432

RESUMO

Fungal exopolysaccharides are important natural products having diverse biological functions. In this study, exopolysaccharides from Fomitopsis castanea mycelia (FEPS) were prepared, and the highest mushroom tyrosinase inhibitory activity was found. FEPS were prepared from cultivation broth by ethanol precipitation method. The extraction yield and protein concentration of FEPS were 213.1 mg/l and 0.03%, respectively. FEPS inhibited mushroom tyrosinase with the half maximal inhibitory concentration (IC50) of 16.5 mg/ml and dose-dependently inhibited cellular tyrosinase activity (63.9% at 50 µg/ml, and 83.3% at 100 µg/ml) in the cell-free extract of SK-MEL-5 human melanoma cell and α-melanocytestimulating hormone (α-MSH)-stimulated melanin formation in intact SK-MEL-5 human melanoma cell. The IC50 of FEPS against NO production from RAW264.7 macrophage cells was 42.8 ± 0.64 µg/ml. By in vivo study using a zebrafish model, exposure of FEPS at 400 µg/ml to dechorionated zebrafish embryos for 18 h decreased the pigment density, compared to that without FEPS-treated control.


Assuntos
Coriolaceae/metabolismo , Polissacarídeos Fúngicos/antagonistas & inibidores , Polissacarídeos Fúngicos/metabolismo , Melanoma/tratamento farmacológico , Micélio/metabolismo , Agaricales/enzimologia , Animais , Linhagem Celular Tumoral/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Modelos Animais de Doenças , Polissacarídeos Fúngicos/isolamento & purificação , Humanos , Concentração Inibidora 50 , Melaninas/metabolismo , Melanócitos/efeitos dos fármacos , Melanoma Experimental , Camundongos , Monofenol Mono-Oxigenase/efeitos dos fármacos , Células RAW 264.7 , Peixe-Zebra , alfa-MSH/efeitos dos fármacos
11.
Int J Biol Macromol ; 137: 469-474, 2019 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-31242449

RESUMO

Endopolygalacturonase (EndoPG) from Stereum purpureum was expressed as a soluble protein in Pichia pastoris GS115, where after 3 days methanol induction the enzyme activity in the culture supernatant was 40 U mL-1. After purification by IMAC, SDS-PAGE analysis showed that the molecular weight of EndoPG was approximately 60.0 kDa. The carbohydrate content of the recombinant enzyme was estimated to be 67.0% (w/w). The optimum temperature and pH of catalysis were 60-70 °C and pH of 4.5, respectively. The enzyme was highly stable over the pH range 6.0-8.0 and retained approximately 60% of its initial activity after incubation at 70 °C for 30 min. The enzyme showed a specific activity of 5040.0 ±â€¯217 U mg-1 and hydrolyzed citrus pectin with Vmax and a KM of 4947.10 ±â€¯393.63 U mg-1 and 2.45 ±â€¯0.23 mg mL-1, respectively, and showed a catalytic efficiency of 2052.90 ±â€¯193.54 mL mg-1 s-1. EndoPG alone reduced the viscosity of papaya juice by 20% after 30 min, and increased its transmittance about 50% with a concomitant reduction of the color by about 55% after 5 h of enzymatic treatment. For apple juice, the relative reduction of viscosity was 30% after 5 h, and the reduction of the color was 30% with a 12% increase in transmittance. Supplementation of a commercial enzymatic cocktail for lignocellulose saccharification with EndoPG increased total reducing sugar release by 8.6 ±â€¯2.1% against sugar cane bagasse, indicating improved access of the cellulolytic enzymes to the biomass polysaccharides.


Assuntos
Agaricales/enzimologia , Biotecnologia , Poligalacturonase/química , Poligalacturonase/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Carica/química , Parede Celular/metabolismo , Estabilidade Enzimática , Sucos de Frutas e Vegetais , Concentração de Íons de Hidrogênio , Hidrólise , Cinética , Malus/química , Poligalacturonase/genética , Proteínas Recombinantes/genética , Saccharum/citologia , Especificidade por Substrato , Temperatura
12.
Int J Biol Macromol ; 136: 1034-1041, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31233796

RESUMO

Many skin disorders and diseases are related to tyrosinase activity, in particular, due to the vital role played by this enzyme in the melanogenic process. Although numerous natural and synthetic tyrosinase inhibitors have been published, substantial efforts have been made to understand the influence of tyrosinase inhibition on the viability of melanoma cells. Here, we assess the impact of two keto-derivatives: 2-acetyl-furan (F1), furfural-acetone (F2), and two carboxyl-derivatives: 2-furan-acrylic acid (F3), 5-methyl-2-furan-acrylic acid (F4), on the mushroom tyrosinase (mTYR) activity, by applying spectroscopic, kinetic and theoretical techniques. From an exploratory and theoretical point of view, results indicated that albeit all furans bind tightly to and inhibit mTYR very efficient, carboxyl-furan derivatives presented best inhibitory activities than keto- derivatives and performed the inhibition competitively and reversible. Moreover, we examined the influence of carboxyl derivative on the viability of melanoma cells. Results expose differential toxicity of these furan derivatives, which indicates a piece of evidence that furan inhibition activity may be related to its toxicity against B16F10 cells.


Assuntos
Antineoplásicos/farmacologia , Inibidores Enzimáticos/farmacologia , Furanos/farmacologia , Melanoma/patologia , Monofenol Mono-Oxigenase/antagonistas & inibidores , Agaricales/enzimologia , Animais , Antineoplásicos/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Inibidores Enzimáticos/metabolismo , Furanos/metabolismo , Humanos , Camundongos , Simulação de Acoplamento Molecular , Monofenol Mono-Oxigenase/química , Monofenol Mono-Oxigenase/metabolismo , Conformação Proteica
13.
Int J Mol Sci ; 20(10)2019 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-31130632

RESUMO

Neuromelanin (NM) is a dark brown pigment found in dopaminergic neurons of the substantia nigra (SN) and in norepinephrinergic neurons of the locus coeruleus (LC). Although NM is thought to be involved in the etiology of Parkinson's disease (PD) because its content decreases in neurodegenerative diseases such as PD, details are still unknown. In this study, we characterized the biosynthetic pathway of the oxidation of dopamine (DA) by tyrosinase in the presence of thiol peptides and proteins using spectroscopic and high-performance liquid chromatography (HPLC) methods and we assessed the binding of DA via cysteine residues in proteins by oxidation catalyzed by redox-active metal ions. To examine whether the protein-bound DA conjugates exhibit pro-oxidant activities, we measured the depletion of glutathione (GSH) with the concomitant production of hydrogen peroxide. The results suggest that the fate of protein-bound DA conjugates depends on the structural features of the proteins and that DA-protein conjugates produced in the brain possess pro-oxidant activities, which may cause neurodegeneration due to the generation of reactive oxygen species (ROS) and the depletion of antioxidants.


Assuntos
Dopamina/metabolismo , Estresse Oxidativo , Doença de Parkinson/metabolismo , Proteínas/metabolismo , Agaricales/enzimologia , Animais , Bovinos , Glutationa/metabolismo , Humanos , Lactoglobulinas/metabolismo , Melaninas/metabolismo , Monofenol Mono-Oxigenase/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Soroalbumina Bovina/metabolismo
14.
Appl Biochem Biotechnol ; 189(3): 855-870, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31131419

RESUMO

Condensed tannins (CTS) have been isolated and purified from leaves of Acanthus ilicifolius Linn. And their structures were investigated by three methods: 13C nuclear magnetic resonance (13C NMR), reversed-phase high-performance liquid chromatography (RP-HPLC), and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The results showed that the CTS were a mixture of catechin/epicatechin, galatechin/epicatechin, and amphicin/epigalin, and that the polymer chain lengths were 3-mers to 14-mers. Antityrosinase activities and antioxidant activities of the CTS from A. ilicifolius leaves were further studied. The IC50 of the CTS on mushroom tyrosinase activity was determined to be 19.7 ± 0.13 µg/mL, and inhibition type analyses indicated that the CTS were mixed type inhibitors and their inhibition CTS was reversible. The CTS from A. ilicifolius leaves also exhibited potential antioxidant activity. The IC50 of DPPH and ABTS scavenging activities were 104 ± 0.894 µg/mL and 86 ± 0.616 µg/mL, respectively. And the FRAP value was 758.28 ± 2.42 mg AAE/g. In addition, we found that the CTS from A. ilicifolius leaves had an excellent effect on preserving the quality of fresh-cut apples by preventing apples from browning through reducing polyphenol oxidase activities in apples.


Assuntos
Acanthaceae/química , Antioxidantes/química , Antioxidantes/farmacologia , Malus/efeitos dos fármacos , Taninos/química , Taninos/farmacologia , Agaricales/enzimologia , Antioxidantes/isolamento & purificação , Conservação de Alimentos , Cinética , Malus/metabolismo , Monofenol Mono-Oxigenase/antagonistas & inibidores , Folhas de Planta/química , Polimerização , Taninos/isolamento & purificação
15.
J Enzyme Inhib Med Chem ; 34(1): 990-998, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31072148

RESUMO

The novel kojic acid derivatives KAD1 and KAD2 have been demonstrated that they exhibited potent anti-melanogenesis activity in our previous report. In this study, we further study the inhibitory mechanism on mushroom tyrosinase. The inhibitory types of both KADs on diphenolase were classified as mixed type based on the results of the kinetic model. The interaction between KADs and tyrosinase was illustrated by fluorescence quenching, molecular docking and copper chelate activity. The KADs were also evaluated with respect to their antioxidant activities by DPPH and ABTS+ assays. The results showed that KADs have more potent antioxidant activities than kojic acid. Our study could provide new ideas for the development of new anti-tyrosinase and antioxidant agents.


Assuntos
Antioxidantes/farmacologia , Inibidores Enzimáticos/farmacologia , Simulação de Acoplamento Molecular , Monofenol Mono-Oxigenase/antagonistas & inibidores , Pironas/farmacologia , Agaricales/enzimologia , Antioxidantes/química , Benzotiazóis/antagonistas & inibidores , Compostos de Bifenilo/antagonistas & inibidores , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/química , Cinética , Monofenol Mono-Oxigenase/metabolismo , Picratos/antagonistas & inibidores , Pironas/química , Espectrometria de Fluorescência , Relação Estrutura-Atividade , Ácidos Sulfônicos/antagonistas & inibidores
16.
Mol Microbiol ; 112(2): 605-619, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31087720

RESUMO

Fungi defend their ecological niche against antagonists by producing antibiosis molecules. Some of these molecules are only produced upon confrontation with the antagonist. The basidiomycete Coprinopsis cinerea induces the expression of the sesquiterpene synthase-encoding gene cop6 and its two neighboring genes coding for cytochrome P450 monooxygenases in response to bacteria. We further investigated this regulation of cop6 and examined if the gene product is involved in the production of antibacterials. Cell-free supernatants of axenic cultures of the Gram-positive bacterium Bacillus subtilis were sufficient to induce cop6 transcription assessed using a fluorescent reporter strain. Use of this strain in a microfluidic device revealed that the cop6 gene was induced in all hyphae directly exposed to the supernatant and that induction occurred within less than one hour. Targeted replacement of the cop6 gene demonstrated the requirement of the encoded synthase for the biosynthesis of the sesquiterpene lagopodin B, a previously reported antibacterial compound from related species. Accordingly, lagopodin B from C. cinerea inhibited the growth of several Gram-positive bacteria including B. subtilis but not Gram-negative bacteria. Our results demonstrate that the C. cinerea vegetative mycelium responds to soluble compounds of a bacterial culture supernatant by local production of an antibacterial secondary metabolite.


Assuntos
Agaricales/metabolismo , Antibacterianos/metabolismo , Bacillus subtilis/fisiologia , Sesquiterpenos/metabolismo , Agaricales/enzimologia , Agaricales/genética , Antibacterianos/farmacologia , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Negativas/crescimento & desenvolvimento , Sesquiterpenos/farmacologia
18.
FEMS Microbiol Lett ; 366(7)2019 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-31004490

RESUMO

The glycoside hydrolase (GH) 16 family of Coprinopsis cinerea includes 15 members distributed in four subgroups (A1, A2, B and D) by phylogenetic analysis. The expression patterns match well with the requirement of wall-softening in the germination of basidiospores, hyphal growth and branching, primordium formation, stipe elongation, pileus expansion and autolysis. Remarkably, expression levels of different GH16 members varied with different morphogenetic events. Like orthologs of Aspergillus fumigatus GH16 glucanases (ENG2-5), which were expressed in the dormant conidia and conidiogenesis, and essential for segregation of conidia, some members such as ENG in the subgroup A1 in C. cinerea were also predominantly expressed in dormant basidiospores, primordia and maturing pilei during basidiosporogenesis. In contrast, other members in subgroup A2, subgroup B or D were dominantly expressed in the germinating basidiospores, the growing mycelia, and the elongating stipes. We did not find the members of the GH81 or GH55 family in C. cinerea genome, which was different from A. fumigatus. However, C. cinerea contains an extra three subgroups (A2, B and D) compared with A. fumigatus. These extra subgroups of GH16 family members may function as those endo-ß-1,3-glucanases belonging to other GH families in the development and growth of C. cinerea.


Assuntos
Agaricales/enzimologia , Proteínas Fúngicas/genética , Regulação Enzimológica da Expressão Gênica , Glicosídeo Hidrolases/genética , Agaricales/classificação , Agaricales/genética , Proteínas Fúngicas/metabolismo , Regulação Fúngica da Expressão Gênica , Glicosídeo Hidrolases/metabolismo , Família Multigênica , Filogenia , Esporos Fúngicos/classificação , Esporos Fúngicos/enzimologia , Esporos Fúngicos/genética
19.
Med Chem ; 15(7): 715-728, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30892163

RESUMO

BACKGROUND: Tyrosinase is involved in the melanin biosynthesis and the abnormal accumulation of melanin pigments leading to hyperpigmentation disorders. Controlling the melanogenesis could be an important strategy for treating abnormal pigmentation. METHODS: In the present study, a series of amide derivatives (3a-e and 5a-e) were synthesized aiming to inhibit tyrosinase activity and melanin production. All derivatives were screened for tyrosinase inhibition in a cell-free system. The possible interactions of amide derivatives with tyrosinase enzyme and effect of these interactions on tyrosinase structure were checked by molecular docking in silico and by Circular Dichroism (CD) studies, respectively. The most potent amide derivative (5c) based on cell-free experiments, was further tested for cellular ROS inhibition and for tyrosinase activity using mouse skin melanoma (B16F10) cells. RESULTS: The tyrosinase inhibitory concentration (IC50) for tested compounds was observed between the range of 68 to 0.0029 µg/ml with a lowest IC50 value of compound 5c which outperforms the reference arbutin and kojic acid. The cellular tyrosinase activity and melanin quantification assay demonstrate that 15µg/ml of 5c attenuates 36% tyrosinase, 24% melanin content of B16F10 cells without significant cell toxicity. Moreover, the zebrafish in vivo assay reveals that 5c effectively reduces melanogenesis without perceptible toxicity. Furthermore, the molecular docking demonstrates that compound 5c interacts with copper ions and multiple amino acids in the active site of tyrosinase with best glide score (-5.387 kcal/mol), essential for mushroom tyrosinase inhibition and the ability to diminish the melanin synthesis in-vitro and in-vivo. CONCLUSION: Thus, we propose compound 5c as a potential candidate to control tyrosinase rooted hyperpigmentation in the future.


Assuntos
Amidas/farmacologia , Antineoplásicos/farmacologia , Inibidores Enzimáticos/farmacologia , Melanoma/tratamento farmacológico , Monofenol Mono-Oxigenase/antagonistas & inibidores , Agaricales/enzimologia , Amidas/síntese química , Amidas/química , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Cinética , Melaninas/análise , Melanoma/metabolismo , Melanoma/patologia , Camundongos , Simulação de Acoplamento Molecular , Estrutura Molecular , Monofenol Mono-Oxigenase/metabolismo , Espécies Reativas de Oxigênio/análise , Espécies Reativas de Oxigênio/metabolismo , Relação Estrutura-Atividade , Peixe-Zebra
20.
Biosens Bioelectron ; 132: 279-285, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-30884314

RESUMO

Highly sensitive phenol biosensor was developed by using well-dispersed carbon nanotubes (CNTs) in enzyme solution and adding CNTs in enzyme electrodes. First, the intact CNTs were dispersed in aqueous tyrosinase (TYR) solution, and TYR molecules were precipitated and crosslinked to prepare the sample of enzyme adsorption, precipitation and crosslinking (EAPC). EAPC exhibited 10.5- and 5.4-fold higher TYR activity per mg of CNTs as compared to enzyme adsorption (EA) and enzyme adsorption/crosslinking (EAC), respectively. EAPC retained 29% of its initial activity after incubation at 40 °C for 128 h, while EA and EAC showed no residual activities, respectively. In biosensing a model phenolic compound of catechol, the sensitivities of EA, EAC and EAPC electrodes on glassy carbon electrode (GCE) were 34, 281 and 675 µA/mM/cm2, respectively. When 90 w/w% CNTs were added to the enzyme electrodes, the sensitivities of EA, EAC, and EAPC electrodes were 146, 427, and 1160 µA/mM/cm2, respectively, and the EAPC electrode showed a 2.3-fold increase in sensitivity upon CNT addition. Catechol and phenol could also be detected by EAPC on the screen-printed electrode (SPE), with sensitivities of 1340 and 1170 µA/mM/cm2, respectively. The sensitivity of EAPC-SPE for phenol detection in the effluent from real municipal wastewater treatment plant was 1100 µA/mM/cm2. The sensitivity of EAPC-SPE retained 74% of its initial sensitivity after incubation at 40 °C for 12 h. The combination of EAPC immobilization and CNT addition has great potential for application in the development of sensitive enzyme biosensors for various analytes and phenols in water environments.


Assuntos
Agaricales/enzimologia , Técnicas Biossensoriais/métodos , Enzimas Imobilizadas/química , Monofenol Mono-Oxigenase/química , Nanotubos de Carbono/química , Fenóis/análise , Poluentes Químicos da Água/análise , Catecóis/análise , Reagentes para Ligações Cruzadas/química , Eletrodos , Limite de Detecção , Modelos Moleculares , Nanotubos de Carbono/ultraestrutura , Fenol/análise , Águas Residuárias/análise
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