Structure-related blockage of calcium channels by vasodilator alkamides in mice mesenteric artery.

The development of new calcium channel blockers is still relevant for the understanding of their physiological role and pharmacological and therapeutic purposes. For this task, natural products represent a relevant source of new drugs. The present work investigated the mechanism and the structural relationship of the vasodilator effect of riparins I, II and III in mouse small mesenteric artery. Riparins I, II and III induced an endothelium-independent and concentration-dependent vasodilator effect in mesenteric arteries. Riparins II and III were more potent than riparin I, suggesting a structural relationship of the effect of these drugs. All riparins inhibited the contractile effect of KCl, similarly to nifedipine. However, the inhibitory profile was different for the contractile responses to phenylephrine and caffeine, passing from similar to nifedipine with riparin I, for similar to SKF-96365 with riparin III. A comparable effect was observed for the increase in the intracellular calcium concentration induced by caffeine and phenylephrine. These results suggest that the higher hydroxylation provides the alkamides the ability to inhibit non-selective cation channels in addition to the inhibition of L-type calcium channels in mouse mesenteric arteries. These observations may give support to the development of new selective inhibitors of non-selective cation channels using alkamides as leading compounds.

Hancornia speciosa Gomes (Apocynaceae) as a potential anti-diabetic drug.

ETHNOPHARMACOLOGICAL RELEVANCE: The leaves of Hancornia speciosa Gomes are traditionally used to treat diabetes in Brazil. The aim of the study is to evaluate the potential anti-diabetic effect of Hancornia speciosa extract and derived fractions. MATERIALS AND METHODS: The ethanolic extract from Hancornia speciosa leaves and chromatographic fractions thereof were evaluated on α-glucosidase assay, on hyperglycemic effect and glucose uptake. The chemical composition of the extract and its most active fraction was investigated by ESI-LC-MS. RESULTS: The ethanolic extract and derived fractions inhibited α-glucosidase in vitro. However, only the crude extract and the dichloromethane fraction inhibited the hyperglycemic effect induced by starch or glucose. Both the extract and dichloromethane fraction were also able to increase glucose uptake in adipocytes. Bornesitol, quinic acid, and chorogenic acid were identified in the extract, along with flavonoid glycosides, whereas the dichloromethane fraction is majorly composed by esters of lupeol and/or α/ß-amirin. CONCLUSIONS: Hancornia speciosa has a potential anti-diabetic effect through a mechanism dependent on inhibition of α-glucosidase and increase on glucose uptake. These results give support to the use on traditional medicine of this medicinal plant.

Impairment of α1-adrenoceptor-mediated calcium influx in contralateral carotids following balloon injury: beneficial effect of superoxide anions.

There are many evidences indicating a compensatory mechanism in contralateral carotids following balloon injury. Previously it was observed α1-adrenoceptor-mediated hyper-reactivity and impairment of calcium influx in contralateral carotids 4 days after injury. At a later stage, α1-adrenoceptor-mediated contraction is similar to the control and we hypothesized that downstream signaling was normal. In the present study, we aimed to evaluate α1-adrenoceptor-mediated calcium influx in contralateral carotids 15 days after balloon injury. Concentration-response curves for CaCl2 in presence of the α1-adrenoceptor agonist (phenylephrine), measurement of the intracellular calcium transient and the levels of reactive oxygen species using fluorescent dyes were performed in control and contralateral carotids. Phenylephrine-induced intracellular calcium mobilization in contralateral carotids was not altered, while phenylephrine-induced calcium influx was reduced in the contralateral artery. Nitric oxide synthase inhibitors, L-NAME or L-NNA, restored this response, but nitrite and nitrate levels were decreased in contralateral carotids. Additionally, a rise in oxygen free radicals was observed in contralateral carotids. Furthermore, Tiron, a superoxide anion scavenger, restored α1-adrenoceptor-mediated calcium influx in contralateral carotids to the control level. Similar results were observed with the selective potassium channels blockers 4-aminopyridine and charybdotoxin. In conclusion, data showed that balloon catheter injury resulted in increased superoxide anions levels, activation of potassium channels (Kv and BKCa), inhibition of calcium channels (Cav) and preservation of α1-adrenoceptor-mediated contraction at a later stage after injury.

Mechanism of the vasodilator effect of mono-oxygenated xanthones: a structure-activity relationship study.

The present study characterized the mechanisms involved in the vasodilator effect of two mono-oxygenated xanthones, 4-hydroxyxanthone and 4-methoxyxanthone. 9-Xanthenone, the base structure of xanthones, was used for comparison. 4-Hydroxyxanthone and 9-xanthenone induced a concentration-dependent and endothelium-independent vasodilator effect in arteries precontracted with phenylephrine (0.1 µmol ·â€ŠL-1) or KCl (50 mmol ·â€ŠL-1). 4-Methoxyxanthone induced a concentration-dependent vasodilator effect in arteries precontracted with phenylephrine, which was partially endothelium-dependent, and involved production of nitric oxide. In endothelium-denuded arteries precontracted with KCl, the vasodilator effect of 4-methoxyxanthone was abolished. The vasodilator effect of 4-hydroxyxanthone (96.22 ± 2.10 %) and 4-methoxyxanthone (96.57 ± 12.40 %) was significantly higher than observed with 9-xanthenone (53.63 ± 8.31 %). The presence of an oxygenated radical in position 4 made 4-hydroxyxanthone (pIC50 = 4.45 ± 0.07) and 4-methoxyxanthone (pIC50 = 5.04 ± 0.09) more potent as a vasodilator than 9-xanthenone (pIC50 = 3.92 ± 0.16). In addition, 4-methoxyxanthone was more potent than the other two xanthones. Ca2+ transients in vascular smooth muscle cells elicited by high K+ were abolished by 4-hydroxyxanthone and 9-xanthenone. The endothelium-independent effect of 4-methoxyxanthone was abolished by inhibition of K+ channels by tetraethylammonium. The current work shows that an oxygenated group in position 4 is essential to achieve Emax and to increase the potency of xanthones as vasodilators. Substitution of an OH by OCH3 in position 4 increases the potency of the vasodilator effect and changes the underling mechanism of action from the blockade of L-type calcium channels to an increase in NO production and activation of K+ channels.

Mechanism of the antihypertensive and vasorelaxant effects of the flavonoid tiliroside in resistance arteries.

Hypertension is a leading cause of death and disability globally, and its prevalence continues to accelerate. The cardiovascular effects of the flavonoid tiliroside have never been reported. In this work, using complementary in vivo and in vitro approaches, we describe the antihypertensive effect of tiliroside and the underlying mechanisms involved in the reduction of blood pressure. Tiliroside (1, 5 or 10 mg/kg) induced a dose-dependent long-lasting decrease in blood pressure in conscious DOCA-salt hypertensive rats that was accompanied by an increased heart rate. Tiliroside also induced a concentration-dependent vasodilation of mesenteric resistance arteries precontracted with phenylephrine. Removal of the endothelium or pretreatment of the preparation with L-NAME or indomethacin did not modify the vasodilator response for tiliroside. When vessels were precontracted with a high K⁺ (50 mM) solution, tiliroside exhibited a vasodilator effect similar to that observed in vessels precontracted with phenylephrine. Experiments carried out in nominally Ca²âº-free solution showed that tiliroside antagonized CaCl2-induced contractions. Moreover, tiliroside reduced the rise in intracellular Ca²âº concentration induced by membrane depolarization in vascular smooth muscle cells. Finally, tiliroside decreased the voltage-activated peak amplitude of the L-type Ca²âº channel current in freshly dissociated vascular smooth muscle cells from mesenteric arteries. Altogether, our results point to an antihypertensive effect of tiliroside due to a reduction in peripheral resistance through blockage of voltage-activated peak amplitude of the L-type Ca²âº channel in smooth muscle cells.

Inhibition of α-glucosidase and hypoglycemic effect of stilbenes from the Amazonian plant Deguelia rufescens var. urucu (Ducke) A. M. G. Azevedo (Leguminosae).

The control of blood glucose levels is critical in the treatment of diabetes mellitus. α-Glucosidase inhibitors are of great importance in reducing hyperglycemia, and plants have provided many of these agents. The present study aimed at investigating the effect of two stilbenes, lonchocarpene and 3,5-dimethoxy-4'-O-prenyl-trans-stilbene (DPS), isolated from the Amazonian plant Deguelia rufescens var. urucu, on α-glucosidase activity and on mice postprandial hyperglycemia. Lonchocarpene and DPS inhibited α-glucosidase in vitro, with pIC(50) values of 5.68 ± 0.12 and 5.73 ± 0.08, respectively. In addition, when given orally, DPS produced a significant reduction of hyperglycemia induced by an oral tolerance test, while lonchocarpene did not. Data suggest that DPS may have a potential use as an antidiabetic drug.

Impaired calcium influx despite hyper-reactivity in contralateral carotid following balloon injury: eNOS involvement.

Balloon catheter injury results in hyper-reactivity to phenylephrine in contralateral carotids. Decreased nitric oxide (NO) modulation and/or increased intracellular calcium concentration triggers vascular smooth muscle contraction. Therefore, this study explores the participation of NO signaling pathway and calcium mobilization on hyper-reactivity to phenylephrine in contralateral carotids. Concentration-response curves for calcium (CaCl(2)) and phenylephrine were obtained in control and contralateral carotids four days after balloon injury, in the presence and absence of the inhibitors (L-NAME, L-NNA, 1400W, 7-NI, Oxyhemoglobin, ODQ or Tiron). Confocal microscopy using Fluo-3AM or DHE was performed to detect the intracellular levels of calcium and reactive oxygen species, respectively. The modulation of NO on phenylephrine-induced contraction was absent in the contralateral carotid. Phenylephrine-induced intracellular calcium mobilization was not altered in contralateral carotids. However, extracellular calcium mobilization by phenylephrine was reduced in the contralateral carotid compared to control arteries, and this result was confirmed by confocal microscopy. L-NAME increased phenylephrine-induced extracellular calcium mobilization in the contralateral carotid to the control levels. Results obtained with L-NNA, 1400W, 7-NI, OxyHb, ODQ or Tiron showed that this response was mediated by products from endothelial NOS (eNOS) different from NO and without soluble guanylate cyclase activation, but it involved superoxide anions. Furthermore, Tiron or L-NNA reduced the levels of reactive oxygen species in contralateral carotids. Data suggest that balloon catheter injury promoted eNOS uncoupling in contralateral carotids, which generates superoxide rather than NO, and reduces phenylephrine-induced extracellular calcium mobilization, despite the hyper-reactivity to phenylephrine in contralateral carotids.

An apparent paradox: attenuation of phenylephrine-mediated calcium mobilization and hyperreactivity to phenylephrine in contralateral carotid after balloon injury.

Balloon catheter injury promotes hyperreactivity to phenylephrine (Phe) in the contralateral carotid. Phe-induced contraction involves calcium mobilization, a process that may be sensitive to reactive oxygen species. In this study, we investigated whether increased reactivity to Phe in the contralateral carotid is due to alterations in calcium mobilization by Phe and reactive oxygen species signaling. Concentration-response curves to Phe were obtained in control and contralateral arteries 4 days after balloon injury. Tiron did not modify Emax to Phe in control arteries but reduced this parameter in the contralateral carotid to control levels. Moreover, immunofluorescence to dihydroethydine showed increased basal oxidative stress in the contralateral artery compared with control artery. Intracellular calcium mobilization by Phe in the contralateral artery was not different from control, but Phe-induced extracellular calcium mobilization was reduced in the contralateral artery compared with that in the control. These data were confirmed by confocal microscopy using Fluo 3-AM. Tiron and SC-236 increased Phe-induced calcium influx in the contralateral artery, which was similar to controls in the same conditions. However, catalase did not modify this response. Taken together, our results suggest that superoxide anions and prostanoids from cyclooxygenase-2 alter pathways downstream of alpha1-adrenoceptor activation in the contralateral carotid in response to injury. This results in reduced Phe-induced calcium influx, despite hyperreactivity to Phe.

Identification of the antimicrobial substances produced by Solanum palinacanthum (Solanaceae).

To find out natural antimicrobial agents as alternative in therapeutics and to preserve food, the methanol extract of Solanum palinacanthum aerial parts was submitted to purification steps guided by antibacterial and antifungal assays. As a consequence, the flavonoid rutin and 3,5-dicaffeoylquinic acid were isolated by column chromatography and high performance liquid chromatography, and identified by mass and nuclear magnetic resonance spectrometry. Minimal inhibitory concentrations (MIC) of the quinic acid derivative against Aeromonas hydrophila, Bacillus subtilis, Staphylococcus aureus and the fungus Aspergillus ochraceus were 250, 1000, 1000 and > 568 microg/mL, respectively. Against the same microorganisms, MIC for rutin were 1000, > 1000, > 1000 and 35 microg/mL, respectively. Rutin was very promising for A. ochraceus control, since its MIC against such fungus was close to the one observed for benzalkonium chloride, which is used as a fungicide in Brazil.

Identification of the antimicrobial substances produced by Solanum palinacanthum (Solanaceae)

To find out natural antimicrobial agents as alternative in therapeutics and to preserve food, the methanol extract of Solanum palinacanthum aerial parts was submitted to purification steps guided by antibacterial and antifungal assays. As a consequence, the flavonoid rutin and 3,5-dicaffeoylquinic acid were isolated by column chromatographyand high performance liquid chromatography, and identified by mass and nuclear magnetic resonance spectrometry. Minimal inhibitory concentrations (MIC) of the quinic acid derivative against Aeromonas hydrophila, Bacillus subtilis, Staphylococcus aureus and the fungus Aspergillus ochraceus were 250, 1000, 1000 and > 568µg/mL, respectively. Against the same microorganisms, MIC for rutin were 1000, > 1000, > 1000 and 35µg/mL, respectively. Rutinwas very promising for A. ochraceus control, since its MIC against such fungus was close to the one observed for benzalkonium chloride, which is used as a fungicide in Brazil.

Com vistas a descobrir antimicrobianos de origem natural para uso terapêutico ou para a preservação de alimentos, o extrato metanólico das partes aéreas de Solanum palinacanthum foi submetido a fracionamentos direcionados por testes para avaliar a atividade antibacteriana e antifúngica. Em decorrência, o flavonóide rutina e o ácido 3,5-dicafeoilquínico foram isolados por cromatografia em coluna e por cromatografia líquida de alta eficiência, para serem identificados por espectrometria de massas e de ressonância magnética nuclear. As concentrações inibitórias mínimas (CIM) do derivado do ácido cafeico contra Aeromonas hydrophila, Bacillus subtilis, Staphylococcus aureus e o fungo Aspergillus ochraceus foram 250, 1000, 1000 e > 568µg/mL, respectivamente. Contra os mesmos organismos, os valores de CIM para a rutina foram 1000, > 1000, > 1000 e 35µg/mL, respectivamente. A rutina mostrou-se muito promissora para o controle de A. ochraceus, pois seu valor de CIM contra tal fungo foi bem próximo ao observado para o cloreto de benzalcônio, que é empregado como fungicida no Brasil.

Purification of an antibacterial compound from Lantana lilacina / Purificação de um composto antibacteriano de Lantana lilacina

Observou-se, em estudo preliminar, que o extrato metanólico das folhas de L. lilacina, coletadas no município de Lavras (MG, Brasil), apresentava atividade antibacteriana. Em decorrência, buscou-se purificar e identificar a substância responsável por tal efeito, através de fracionamento do referido extrato direcionado por testes de difusão em agar com Aeromonas hydrophila, Bacillus subtilis, Pseudomonas aeruginosa e Staphylococcus aureus. Após partições com solventes e vários processos cromatográficos, isolou-se o [β-3,4-diidroxifenil)etil]-(3'-O-α-L-ramnopiranosil)-(4'- O-cafeoil)-β-D-glicopiranosídeo, que é conhecido como acteosídeo. A concentração inibitória mínima e a concentração bactericida mínima desta substância para A. hydrophila, B. subtilis, P. aeruginosa e S. aureus foram de 0,12, 1,00, 1,00 e 0,25 mg/mL, respectivamente.

Since the methanol extract of Lantana lilacina leaves collected in the city of Lavras (MG, Brazil) showed antibacterial properties in a preliminary study, a fractionation process guided by agar diffusion assays with Aeromonas hydrophila, Bacillus subtilis, Pseudomonas aeruginosa and Staphylococcus aureus was carried out to purify and identify the active compounds. After solvent partition and several chromatographic steps, [ß-3,4-dihydroxyphenyl)-ethyl]-(3'-O-α-L-rhamnopyranosyl)-(4'- O-cafeoyl)-β-D-glycopyranoside, known as acteoside, was isolated. The minimal inhibition concentration and the minimal bactericidal concentration of such substance against A. hydrophila, B. subtilis, P. aeruginosa and S. aureus were 0.12, 1.00, 1.00 and 0.25 mg/mL, respectively.

Antibacterial activity of plant extracts from Brazilian southeast region.

A screening was conducted with 26 plants collected in the Brazilian southeast region, to identify plant extracts with antibacterial properties against Aeromonas hydrophila, Bacillus subtilis, Pseudomonas aeruginosa and Staphylococcus aureus. Initially, the agar diffusion method was employed. Then, those extracts presenting activity were submitted to a broth microdilution assay to determine the minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC). It was observed that 13 of the tested extracts showed antibacterial activity. The best results were obtained with those from Lantana lilacina and Phyllanthus tenellus.