RESUMO
The aim of this work was to investigate the in vitro cytotoxic effect of previously developed nanocapsules, nanoemulsion, and microemulsion based on bullfrog oil (BFO) against human melanoma cells (A2058). The nanosystems were produced as described in previous studies and characterized according to droplet/particle distribution and zeta potential. The biocompatibility was evaluated by the determination of the hemolytic potential against human erythrocytes. The cytotoxicity assessment was based on MTT and cell death assays, determination of Reactive Oxygen Species (ROS) levels, and cell uptake. The nanosystems were successfully reproduced and showed hemolytic potential smaller than 10% at all oil concentrations (50 and 100 µg.mL-1) (p < 0.05). The MTT assay revealed that the nanosystems decreased the mitochondrial activity up to 92 ± 2% (p < 0.05). The study showed that the free BFO induced cell apoptosis, while all the nanostructured systems caused cell death by necrosis associated with a ROS overproduction. This can be related to the increased ability of the nanostructured systems to deliver the BFO across all cellular compartments (membrane, cytoplasm, and nucleus). Finally, these results elucidate the in vitro BFO nanosystems cytotoxic effect against human melanoma cells (A2058), revealing the emulsified ones as the most cytotoxic systems. Overall, the findings suggest that the safety and antineoplastic activity of these systems can be further investigated by in vivo studies.
Assuntos
Antineoplásicos/farmacologia , Melanoma/tratamento farmacológico , Nanoestruturas , Neoplasias Cutâneas/tratamento farmacológico , Animais , Antineoplásicos/isolamento & purificação , Antineoplásicos/toxicidade , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Emulsões , Eritrócitos/efeitos dos fármacos , Eritrócitos/metabolismo , Hemólise/efeitos dos fármacos , Humanos , Melanoma/patologia , Mitocôndrias/metabolismo , Nanocápsulas , Óleos/isolamento & purificação , Óleos/farmacologia , Óleos/toxicidade , Tamanho da Partícula , Rana catesbeiana/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Neoplasias Cutâneas/patologiaRESUMO
Melanoma is the most lethal form of skin cancer, with a worldwide increase in incidence. Despite the increased overall survival of metastatic melanoma patients given recent advances in targeted and immunotherapy, it still has a poor prognosis and available treatment options carry diverse severe side effects. Polysaccharides from seaweed have been shown to exert antitumor activities. Here we show in vitro and in vivo antitumor activities of a sulfated homogalactan (named 3G4S) from Codium isthmocladum seaweed in the B16-F10 murine melanoma cell line. 3G4S did not induce cytotoxicity or proliferation changes; however, it was able to reduce solid tumor growth and metastasis, while not inducing side effects in mice. B16-F10 cells traits related to the metastatic cascade were also impaired by 3G4S, reducing cell invasion, colony-forming capacity and membrane glycoconjugates. Therefore, 3G4S shows promising antitumor activities without the commonly associated drawbacks of cancer treatments and can be further explored.
Assuntos
Galactanos/farmacologia , Química Verde , Melanoma Experimental/prevenção & controle , Alga Marinha/química , Sulfatos/química , Animais , Apoptose , Proliferação de Células , Feminino , Humanos , Masculino , Melanoma Experimental/secundário , Camundongos , Camundongos Endogâmicos C57BL , Células Tumorais CultivadasRESUMO
Melon (Cucumis melo L.) has high economic value and in recent years, its production has increased; however, part of the fruit is wasted. Usually, inedible parts such as peel and seeds are discarded during processing and consumption. Extracts of melon residues were prepared and their phenolic compounds, antioxidants and antiproliferative activities were evaluated. Total phenolic compounds were found in hydroethanolic, hydromethanolic, and aqueous extracts, especially for melon peel (1.016 mg gallic acid equivalent/100 g). Flavonoids total content found for melon peel aqueous extract was 262 µg of catechin equivalent (CA)/100 g. In all extracts of melon peel significant amounts of gallic acid, catechin, and eugenol were found. For total antioxidant capacity, reported as ascorbic acid equivalent, the hydroethanolic and hydromethanolic extracts in peels and hydromethanolic in seeds were 89, 74, and 83 mg/g, respectively. Different extracts of melon showed iron and copper ions chelating activity at different concentrations, especially melon peel aqueous extract, reaching values of 61% for iron and 84% for copper. The hydroethanolic extract of melon peel presented a significant ability for hydroxyl radicals scavenging (68%). To assess the antiproliferative potential in human cancer cell lines, such as kidney carcinoma, colorectal carcinoma, cervical adenocarcinoma and cervical carcinoma, MTT assay was performed. The proliferation was inhibited by 20-85% at extracts concentrations of 0.1-1.0 mg/mL in all cancer cell lines. The results suggest that melon residues extracts display a high antioxidant activity in in vitro assays and have effective biological activity against the growth of human tumor cells.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Antioxidantes/farmacologia , Proliferação de Células/efeitos dos fármacos , Cucurbitaceae/química , Extratos Vegetais/farmacologia , Antineoplásicos Fitogênicos/isolamento & purificação , Antioxidantes/isolamento & purificação , Linhagem Celular Tumoral , Cromatografia Líquida de Alta Pressão , Cromatografia em Camada Fina , Flavonoides/isolamento & purificação , Flavonoides/farmacologia , Humanos , Fenóis/isolamento & purificação , Fenóis/farmacologia , Sementes/química , Taninos/isolamento & purificação , Taninos/farmacologiaRESUMO
Sulfated polysaccharides from 11 species of tropical marine algae (one edible specie of Rhodophyta, six species of Phaeophyta and four species of Chlorophyta) collected from Natal city coast (Northeast of Brazil) were evaluated for their anticoagulant, antioxidant and antiproliverative in vitro activities. In the activated partial thromboplastin time (APTT) test, which evaluates the intrinsic coagulation pathway, seven seaweeds presented anticoagulant activity. Dictyota cervicornis showed the highest activity, prolonging the coagulation time to double the baseline value in the APTT with only 0.01 mg/100 microl of plasma, 1.4-fold lesser than Clexane, a low molecular weight heparin. In the protrombin time (PT) test, which evaluates the extrinsic coagulation pathway, only Caulerpa cupresoides showed anticoagulant activity. All species collected showed antioxidant activities. This screening emphasized the great antioxidant potential (total capacity antioxidant, power reducing and ferrous chelating) of four species: C. sertularioide; Dictyota cervicornis; Sargassum filipendula and Dictyopteris delicatula. After 72 h incubation, HeLa cell proliferation was inhibited (p<0.05) between 33.0 and 67.5% by S. filipendula; 31.4 and 65.7% by D. delicatula; 36.3 and 58.4% by Caulerpa prolifera and 40.2 and 61.0% by Dictyota menstrualis at 0.01-2mg/mL algal polysaccharides. The antiproliferative efficacy of these algal polysaccharides were positively correlated with the sulfate content (r=0.934). Several polysaccharides demonstrated promising antioxidant, antiproliferative an/or anticoagulant potential and have been selected for further studies on bioguided fractionation, isolation and characterization of pure polysaccharides from these species as well as in vivo experiments are needed and are already in progress.
Assuntos
Clorófitas/química , Phaeophyceae/química , Polissacarídeos/farmacologia , Rodófitas/química , Anticoagulantes/isolamento & purificação , Anticoagulantes/farmacologia , Antioxidantes/isolamento & purificação , Antioxidantes/farmacologia , Proliferação de Células/efeitos dos fármacos , Enoxaparina/farmacologia , Células HeLa , Humanos , Tempo de Tromboplastina Parcial , Polissacarídeos/isolamento & purificação , Tempo de ProtrombinaRESUMO
Sulfated polysaccharides (fucans and fucoidans) from brown algae show several biological activities, including anticoagulant and anti-inflammatory activities. We have extracted a sulfated heterofucan from the brown seaweed Lobophora variegata by proteolytic digestion, followed by acetone fractionation, molecular sieving, and ion-exchange chromatography. Chemical analyses and 13C-NMR and IR spectroscopy showed that this fucoidan is composed of fucose, galactose, and sulfate at molar ratios of 1 : 3 : 2. We compared the anticoagulant activity of L. variegata fucoidan with those of a commercial sulfated polysaccharide (also named fucoidan) from Fucus vesiculosus and heparin. The experimental inflammation models utilized in this work revealed that fucoidan from L. variegata inhibits leukocyte migration to the inflammation site. Ear swelling caused by croton oil was also inhibited when sulfated polysaccharides from F. vesiculosus and L. variegata were used. The precise mechanism of different action between homo- and heterofucans is not clear; nevertheless, the polysaccharides studied here may have therapeutic potential in inflammatory disorders.
Assuntos
Anti-Inflamatórios/isolamento & purificação , Anticoagulantes/isolamento & purificação , Phaeophyceae/química , Polissacarídeos/isolamento & purificação , Animais , Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Anticoagulantes/química , Anticoagulantes/farmacologia , Galactanos/química , Galactanos/isolamento & purificação , Galactanos/farmacologia , Polissacarídeos/química , Polissacarídeos/farmacologia , RatosRESUMO
Brown algae have two kinds of acid polysaccharides present in the extracellular matrix: sulfated fucan and alginic acid. We have previously isolated and characterized fucans from several species of brown seaweed. The characterized fucans from Dictyotaceae are heterofucans containing mainly fucose, galactose, glucose, xylose, and/or uronic acid. The fucan from Fucus vesiculosus is a homofucan containing only sulfated fucose. We assessed the activity of these fucans as inhibitors of HIV from reverse transcriptase (RT). Using activated DNA and template primers poly(rA)-oligo(dT), we found that fucans at a concentration of 0.5-1.0 microg/mL had a pronounced inhibitory effect in vitro on the avian reverse transcriptase, with the exception of xylogalactofucan isolated from Spatoglossum schröederi, which had no inhibitory activity. The alginic acid (1.0 microg/mL) inhibited the reverse transcriptase activity by 51.1% using activated DNA. The inhibitory effect of fucans was eliminated by their desulfation. Furthermore, only xylofucoglucuronan from S. schröederi lost its activity after carboxyreduction. We suggest that fucan activity is not only dependent on the ionic changes but also on the sugar rings that act to spatially orientate the charges in a configuration that recognizes the enzyme, thus determining the specificity of the binding.
Assuntos
Fármacos Anti-HIV/química , Transcriptase Reversa do HIV/antagonistas & inibidores , Phaeophyceae/química , Polissacarídeos/química , Inibidores da Transcriptase Reversa/química , Transcriptase Reversa do HIV/química , Relação Estrutura-AtividadeRESUMO
The present study shows the cytotoxic effect of three different classes of algal polysaccharides on HL60 cells. Three galactofucans, fucoidan, and glucan were the polysaccharides utilized in this analysis. The parameters used for evaluating cell viability were [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] (MTT) reduction, protein content, and phosphatase activity. We demonstrated stimulation of phosphatase activity, MTT reduction, and protein content in relation to three types of galactofucans (1, 2, and 3) with different molecular weights (1600, 1200, and 360 kD). However, when HL60 cells were treated with galactofucan type 3, the total protein remained unchanged. Under the same experimental conditions, an expressed increase in the phosphatase activity was detected when galactofucan 3 was utilized. In relation to the mitochondrial function, the stimulation was higher in cells treated with galactofucan type 1. Fucoidan did not have a significant effect on MTT reduction, but protein content was decreased (IC50 around 30 microg/ml). Glucan also activated all the parameters that were analyzed, and this effect was more expressed in the phosphatase activity and in the protein content. This study provides new insights into the cytotoxic action of polysaccharides on HL60 cells and suggests for the first time the possible involvement of phosphatases in this process.
Assuntos
Citotoxinas/farmacologia , Fucus/química , Galactanos/farmacologia , Glucanos/farmacologia , Monoéster Fosfórico Hidrolases/metabolismo , Polissacarídeos Bacterianos/farmacologia , Polissacarídeos/farmacologia , Alga Marinha/química , Citotoxinas/química , Galactanos/química , Glucanos/química , Células HL-60 , Humanos , Polissacarídeos/química , Polissacarídeos Bacterianos/químicaRESUMO
The brown algae Padina gymnospora contain different fucans. Powdered algae were submitted to proteolysis with the proteolytic enzyme maxataze. The first extract of the algae was constituted of polysaccharides contaminated with lipids, phenols, etc. Fractionation of the fucans with increasing concentrations of acetone produced fractions with different proportions of fucose, xylose, uronic acid, galactose, and sulfate. One of the fractions, precipitated with 50% acetone (v/v), contained an 18-kDa heterofucan (PF1), which was further purified by gel-permeation chromatography on Sephadex G-75 using 0.2 M acetic acid as eluent and characterized by agarose gel electrophoresis in 0.05 M 1,3 diaminopropane/acetate buffer at pH 9.0, methylation and nuclear magnetic resonance spectroscopy. Structural analysis indicates that this fucan has a central core consisting mainly of 3-beta-D-glucuronic acid 1-> or 4-beta-D-glucuronic acid 1 ->, substituted at C-2 with alpha-L-fucose or beta-D-xylose. Sulfate groups were only detected at C-3 of 4-alpha-L-fucose 1-> units. The anticoagulant activity of the PF1 (only 2.5-fold lesser than low molecular weight heparin) estimated by activated partial thromboplastin time was completely abolished upon desulfation by solvolysis in dimethyl sulfoxide, indicating that 3-O-sulfation at C-3 of 4-alpha-L-fucose 1-> units is responsible for the anticoagulant activity of the polymer.
Assuntos
Anticoagulantes/química , Phaeophyceae/química , Polissacarídeos/química , Alga Marinha/química , Anticoagulantes/isolamento & purificação , Anticoagulantes/farmacologia , Coagulação Sanguínea/efeitos dos fármacos , Eletroforese em Gel de Ágar , Humanos , Espectroscopia de Ressonância Magnética , Tempo de Tromboplastina Parcial , Polissacarídeos/isolamento & purificação , Polissacarídeos/farmacologia , Tempo de TrombinaRESUMO
The brown algae Padina gymnospora contain different fucans. Powdered algae were submitted to proteolysis with the proteolytic enzyme maxataze. The first extract of the algae was constituted of polysaccharides contaminated with lipids, phenols, etc. Fractionation of the fucans with increasing concentrations of acetone produced fractions with different proportions of fucose, xylose, uronic acid, galactose, and sulfate. One of the fractions, precipitated with 50 percent acetone (v/v), contained an 18-kDa heterofucan (PF1), which was further purified by gel-permeation chromatography on Sephadex G-75 using 0.2 M acetic acid as eluent and characterized by agarose gel electrophoresis in 0.05 M 1,3 diaminopropane/acetate buffer at pH 9.0, methylation and nuclear magnetic resonance spectroscopy. Structural analysis indicates that this fucan has a central core consisting mainly of 3-ß-D-glucuronic acid 1-> or 4-ß-D-glucuronic acid 1 ->, substituted at C-2 with alpha-L-fucose or ß-D-xylose. Sulfate groups were only detected at C-3 of 4-alpha-L-fucose 1-> units. The anticoagulant activity of the PF1 (only 2.5-fold lesser than low molecular weight heparin) estimated by activated partial thromboplastin time was completely abolished upon desulfation by solvolysis in dimethyl sulfoxide, indicating that 3-O-sulfation at C-3 of 4-alpha-L-fucose 1-> units is responsible for the anticoagulant activity of the polymer.
Assuntos
Humanos , Phaeophyceae/química , Anticoagulantes/química , Polissacarídeos/química , Alga Marinha/química , Anticoagulantes/isolamento & purificação , Anticoagulantes/farmacologia , Coagulação Sanguínea/efeitos dos fármacos , Eletroforese em Gel de Ágar , Espectroscopia de Ressonância Magnética , Tempo de Tromboplastina Parcial , Polissacarídeos/isolamento & purificação , Polissacarídeos/farmacologia , Tempo de TrombinaRESUMO
Fucan is a term used to denote a family of sulfated L-fucose-rich polysaccharides which are present in the extracellular matrix of brown seaweed and in the egg jelly coat of sea urchins. Plant fucans have several biological activities, including anticoagulant and antithrombotic, related to the structural and chemical composition of polysaccharides. We have extracted sulfated polysaccharides from the brown seaweed Dictyota menstrualis by proteolytic digestion, followed by separation into 5 fractions by sequential acetone precipitation. Gel electrophoresis using 0.05 M 1,3-diaminopropane-acetate buffer, pH 9.0, stained with 0.1% toluidine blue, showed the presence of sulfated polysaccharides in all fractions. The chemical analyses demonstrated that all fractions are composed mainly of fucose, xylose, galactose, uronic acid, and sulfate. The anticoagulant activity of these heterofucans was determined by activated partial thromboplastin time (APTT) using citrate normal human plasma. Only the fucans F1.0v and F1.5v showed anticoagulant activity. To prolong the coagulation time to double the baseline value in the APTT, the required concentration of fucan F1.0v (20 g/ml) was only 4.88-fold higher than that of the low molecular weight heparin Clexane (4.1 g/ml), whereas 80 g/ml fucan 1.5 was needed to obtain the same effect. For both fucans this effect was abolished by desulfation. These polymers are composed of fucose, xylose, uronic acid, galactose, and sulfate at molar ratios of 1.0:0.8:0.7:0.8:0.4 and 1.0:0.3:0.4:1.5:1.3, respectively. This is the fist report indicating the presence of a heterofucan with higher anticoagulant activity from brown seaweed.