Antioxidant activity evaluation of dried herbal extracts: an electroanalytical approach

ABSTRACT The prevention of chronic and degenerative diseases, is a health concern deeply associated with oxidative stress. Such progressive phenomena can be avoided through exogenous antioxidant intake, which set up a reductant cascade, mopping up damaging free radicals. Medicinal herbs are commonly associated with high antioxidant potential, and hence their health benefits. The commerce of dried herbal extracts movements a big portion of developing countries economy. The determination of medicinal herbs the antioxidant activity capacity is of utmost importance. The assessment of antioxidant activity in phytotherapics is mostly achieved by spectrophotometric assays, however colored substances can produce interferences that do not occur in electroanalytical methods. Therefore, the aim of this paper is to compare spectrophotometric and voltammetric techniques to evaluate antioxidant activity in herbal drugs such as: Ginkgo biloba L., Camellia sinensis (L.) Kuntze, Theaceae; Hypericum perforatum L., Hypericaceae; Aesculus hippocastanum L., Sapindaceae; Rosmarinus officinalis L., Lamiaceae; Morinda citrifolia L., Rubiaceae; Centella asiatica (L.) Urb., Apiaceae; Trifolium pratense L., Fabaceae; Crataegus oxyacantha L., Rosaceae; and Vaccinium macrocarpon Aiton, Ericaceae. The spectrophotometric methods employed were DPPH (2,2-diphenyl-1-picrylhydrazyl), ABTS (2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) and the Folin-Ciocalteu assays. The electroanalytical method used was voltammetry and it was developed a phenoloxidase based biosensor. The redox behavior observed for each herbal sample resulted in distinguishable voltammetric profiles. The highest electrochemical indexes were found to G. biloba and H. perforatum, corroborating to traditional spectrophotometric methods. Thus, the electroanalysis of herbal drugs, may be a promising tool for antioxidant potential assessment.

Protective Effects of Ellagic Acid on Cardiovascular Injuries Caused by Hypertension in Rats.

Ellagic acid is described as having antioxidant and antiproliferative properties. Hence, it was hypothesized that ellagic acid could improve cardiovascular damage caused by hypertension. In this work, hypertension was induced in rats with Nω-Nitro-L-arginine methyl ester hydrochloride (60 mg/kg/day in drinking water) for 6 weeks. Ellagic acid was coadministered (10 or 30 mg/kg/day by gavage) between the second and sixth week. Blood pressure was recorded every week by tail-cuff plethysmography. After 6 weeks, the rats were sacrificed, the hearts and kidneys were weighed, and blood was collected. Aortas were isolated and set up to isometric recordings in an organ bath for histological assay and measuring of calcium content. Hypertension (233.6 ± 9.5 mmHg) was reduced (p < 0.01) by treatment with ellagic acid 10 or 30 mg/kg. The blood levels of nitrate/nitrite were reduced in hypertensive rats and the ellagic acid restored these levels. While the vascular relaxations to acetylcholine and sodium nitoprusside and the contraction to phenylephrine were impaired in the hypertensive group, they were improved after ellagic acid treatment. The alkaline phosphatase activity was increased by hypertension and returned to control levels after ellagic acid treatment. In the aorta, the administration of ellagic acid resulted in less aortic wall thickening and less calcification. In conclusion, ellagic acid attenuates hypertension, possibly improving nitric oxide bioavailability. The vascular response to acetylcholine, sodium nitroprusside, and phenylephrine was impaired by hypertension and improved after treatment with ellagic acid. Moreover, plasmatic alkaline phosphatase activity, calcium content, and hypertrophy in vascular tissues during hypertension were attenuated by treatment with ellagic acid.

Radical Scavenger Capacity of Jabuticaba Fruit (Myrciaria cauliflora) and Its Biological Effects in Hypertensive Rats.

Jabuticaba is an exotic fruit native to Brazil that has been arousing medicinal interest. Using chemical (HPLC-PDA, resonance mass spectra, and NMR), electroanalytical (differential pulse voltammetry, radical scavenging assay), and pharmacological (in vivo and in vitro) approaches, we have identified its bioactive compounds and hypotensive effects on hypertensive rats. The hydroalcoholic extract of jabuticaba (HEJ) presents a great quantity of phenolic compounds, and several molecules with hydroxyl groups present high efficiency as an antioxidant. The treatment with HEJ (100 and 300 mg/kg/day, for four weeks) presented hypotensive effects on L-NAME-induced hypertensive rats, possibly improving the nitric oxide bioavailability because of its high antioxidant potential. Furthermore, renal and cardiac hypertrophies were also attenuated after the HEJ treatment. Moreover, the vascular responses to contractile and dilating agonists were improved with the HEJ treatment, which is also able to induce nitric oxide production in endothelial cells.

Jabuticaba-Induced Endothelium-Independent Vasodilating Effect on Isolated Arteries / Efeito Vasodilatador Independente do Endotélio Induzido pela Jabuticaba (M. cauliflora) em Artérias Isoladas

Abstract Background: Despite the important biological effects of jabuticaba, its actions on the cardiovascular system have not been clarified. Objectives: To determine the effects of jabuticaba hydroalcoholic extract (JHE) on vascular smooth muscle (VSM) of isolated arteries. Methods: Endothelium-denuded aortic rings of rats were mounted in isolated organ bath to record isometric tension. The relaxant effect of JHE and the influence of K+ channels and Ca2+ intra- and extracellular sources on JHE-stimulated response were assessed. Results: Arteries pre-contracted with phenylephrine showed concentration-dependent relaxation (0.380 to 1.92 mg/mL). Treatment with K+ channel blockers (tetraethyl-ammonium, glibenclamide, 4-aminopyridine) hindered relaxation due to JHE. In addition, phenylephrine-stimulated contraction was hindered by previous treatment with JHE. Inhibition of sarcoplasmic reticulum Ca2+ ATPase did not change relaxation due to JHE. In addition, JHE inhibited the contraction caused by Ca2+ influx stimulated by phenylephrine and KCl (75 mM). Conclusion: JHE induces endothelium-independent vasodilation. Activation of K+ channels and inhibition of Ca2+ influx through the membrane are involved in the JHE relaxant effect.

Resumo Fundamentos: Embora a jabuticaba apresente importantes efeitos biológicos, suas ações sobre o sistema cardiovascular ainda não foram esclarecidas. Objetivos: Determinar os efeitos do extrato de jabuticaba (EHJ) sobre o músculo liso vascular (MLV) em artérias isoladas. Métodos: Aortas (sem endotélio) de ratos foram montadas em banho de órgãos isolados para registro de tensão isométrica. Foram verificados o efeito relaxante, a influência dos canais de K+ e das fontes de Ca2+ intra- e extracelular sob a resposta estimulada pelo EHJ. Resultados: Artérias pré-contraídas com fenilefrina apresentaram relaxamento concentração-dependente (0,380 a 1,92 mg/mL). O tratamento com bloqueadores de canais de K+ (tetraetilamônio, glibenclamida, 4-aminopiridina) prejudicaram o relaxamento pelo EHJ. A contração estimulada com fenilefrina também foi prejudicada pelo tratamento prévio com EHJ. A inibição da Ca2+ATPase do reticulo sarcoplasmático não alterou o relaxamento pelo EHJ. Além disso, o EHJ inibiu a contração causada pelo influxo de Ca2+ estimulado por fenilefrina e KCl (75 mM). Conclusão: O EHJ induz vasodilatação independente do endotélio. Ativação dos canais de K+ e inibição do influxo de Ca2+ através da membrana estão envolvidas no efeito relaxante do EHJ.

Jabuticaba-Induced Endothelium-Independent Vasodilating Effect on Isolated Arteries.

BACKGROUND:: Despite the important biological effects of jabuticaba, its actions on the cardiovascular system have not been clarified. OBJECTIVES:: To determine the effects of jabuticaba hydroalcoholic extract (JHE) on vascular smooth muscle (VSM) of isolated arteries. METHODS:: Endothelium-denuded aortic rings of rats were mounted in isolated organ bath to record isometric tension. The relaxant effect of JHE and the influence of K+ channels and Ca2+ intra- and extracellular sources on JHE-stimulated response were assessed. RESULTS:: Arteries pre-contracted with phenylephrine showed concentration-dependent relaxation (0.380 to 1.92 mg/mL). Treatment with K+ channel blockers (tetraethyl-ammonium, glibenclamide, 4-aminopyridine) hindered relaxation due to JHE. In addition, phenylephrine-stimulated contraction was hindered by previous treatment with JHE. Inhibition of sarcoplasmic reticulum Ca2+ ATPase did not change relaxation due to JHE. In addition, JHE inhibited the contraction caused by Ca2+ influx stimulated by phenylephrine and KCl (75 mM). CONCLUSION:: JHE induces endothelium-independent vasodilation. Activation of K+ channels and inhibition of Ca2+ influx through the membrane are involved in the JHE relaxant effect. FUNDAMENTOS:: Embora a jabuticaba apresente importantes efeitos biológicos, suas ações sobre o sistema cardiovascular ainda não foram esclarecidas. OBJETIVOS:: Determinar os efeitos do extrato de jabuticaba (EHJ) sobre o músculo liso vascular (MLV) em artérias isoladas. MÉTODOS:: Aortas (sem endotélio) de ratos foram montadas em banho de órgãos isolados para registro de tensão isométrica. Foram verificados o efeito relaxante, a influência dos canais de K+ e das fontes de Ca2+ intra- e extracelular sob a resposta estimulada pelo EHJ. RESULTADOS:: Artérias pré-contraídas com fenilefrina apresentaram relaxamento concentração-dependente (0,380 a 1,92 mg/mL). O tratamento com bloqueadores de canais de K+ (tetraetilamônio, glibenclamida, 4-aminopiridina) prejudicaram o relaxamento pelo EHJ. A contração estimulada com fenilefrina também foi prejudicada pelo tratamento prévio com EHJ. A inibição da Ca2+ATPase do reticulo sarcoplasmático não alterou o relaxamento pelo EHJ. Além disso, o EHJ inibiu a contração causada pelo influxo de Ca2+ estimulado por fenilefrina e KCl (75 mM). CONCLUSÃO:: O EHJ induz vasodilatação independente do endotélio. Ativação dos canais de K+ e inibição do influxo de Ca2+ através da membrana estão envolvidas no efeito relaxante do EHJ.

Electrochemical behavior and determination of major phenolic antioxidants in selected coffee samples.

The redox behavior of commercial roasted coffee products were evaluated by electroanalysis, whereas high performance liquid chromatography (HPLC) was used for determination of cinnamic acid markers, the total phenolic content (TPC) was achieved by Folin-Ciocalteu assay, and the traditional DPPH (1-diphenyl-2-picrylhydrazyl) method for antioxidant power determination. In turn, an optimized electrochemical index was proposed to estimate the antioxidant power of different samples and it was found a great correlation between all methods. The voltammetric profile of all coffee samples was quite similar, presenting the same number of peaks at the same potential values. Minor differences on current levels were in agreement with the total phenolic and major markers content, as well as, to the antioxidant power. Therefore, the electrochemical methods showed to be practical, low cost and very useful to evaluate the antioxidant properties of coffee samples, which is a relevant quality parameter of this widely consumed beverage.

Blocking the L-type Ca2+ channel (Cav 1.2) is the key mechanism for the vascular relaxing effect of Pterodon spp. and its isolated diterpene methyl-6α-acetoxy-7ß-hydroxyvouacapan-17ß-oate.

Pterodon spp. Vogel (Fabaceae), popularly known as "sucupira", has ethnopharmacological application which is described as having antispasmodic and relaxant effects. Hence, it was hypothesized that sucupira oil-resin (SOR) could induce smooth muscle relaxation. So, this study investigated the mechanisms involved in the vasorelaxant effect of SOR and its isolated diterpene (methyl-6α-acetoxy-7ß-hydroxyvouacapan-17ß-oate). Vascular reactivity experiments were performed using rat aortic rings (n=5-8) with (E+) or without endothelium (E-) in an isolated bath organ. The SOR (0-56 µg/mL) relaxed phenylephrine (E+: 86.7±7.1%; E-: 92.3±4.7%) and KCl contracted rings (E-: 97.1±2.8%). In the same way, diterpene (0-48 µg/mL) also relaxed phenylephrine (E+: 94.5±3.6%; E-: 92.2±3.4%) and KCl contracted rings (E-: 99.7±0.2%). The pre-incubation of arterial rings with cyclopiazonic acid (reticular Ca2+-ATPase inhibitor), tetraethylammonium (K+ channels blocker) or MDL-12,330A (adenylyl cyclesinhibitor) did not modify either SOR- or diterpeneinduced vasorelaxation. However, ODQ (guanylyl cyclase inhibitor) impaired only diterpene-induced vasorelaxation. SOR and diterpene significantly reduced CaCl2-induced contraction stimulated by Bay K8644 (1 µM), phenylephrine (0.1 µM) or KCl solution (40 mM). Computational molecular docking studies demonstrated that the vasodilator effect of diterpene relies on blocking the Cav 1.2 channel, and patch clamp results showed that diterpene substantially decreased the ionic current through Cav 1.2 in freshly dissociated vascular smooth muscle cells. These findings suggest that SOR and its isolated diterpene induce endothelium-independent vascular relaxation by blocking the L-type Ca2+ channel (Cav 1.2).

Diuretic effects and urinary electrolyte excretion induced by Aspidosperma subincanum Mart. and the involvement of prostaglandins in such effects.

ETHNOPHARMACOLOGICAL RELEVANCE: Aspidosperma subincanum Mart. is a medicinal herb known for its diuretic properties and used for the treatment of cardiovascular-related illnesses. Although our earlier study has shown that the ethanol extract of Aspidosperma subincanum (EEAS) induces hypotension and vasodilation, no scientific data have been recorded to evaluate the diuretic effects of this Brazilian medicinal plant. The aim of this study was to evaluate the diuretic activity of EEAS, and possible mechanism of action, using Wistar rats. MATERIAL AND METHODS: EEAS (60 and 120mg/kg), furosemide (20mg/kg) or saline (control) were orally administered to rats individually held in metabolic cages for urine collection 1, 2, 4, 6, 8, 12 and 24h after treatment. In order to evaluate the involvement of prostaglandins in the diuretic action of EEAS, the animals received piroxicam (5mg/kgi.p.), a nonselective inhibitor of cyclooxygenase, before treatment with EEAS at 120mg/kg. The control groups received only saline (NaCl, 0.9%), or saline and piroxicam. Urinary volume, electrolyte excretion and pH were measured. RESULTS: Oral administration of EEAS 60 and 120mg/kg significantly increased diuresis and electrolyte excretion of Na(+) and K(+) on a continuous basis throughout the study period. Both EEAS 60 and 120mg/kg caused a relative increase of around 77% and 142%, respectively, in cumulative diuresis compared with the control group. From 4th hour until the end of the experiment, the group treated with EEAS 120mg/kg provided a greater excretion of Na(+) than the furosemide group. The diuretic effects of EEAS were neutralized by piroxicam between 4 and 8h after treatment. CONCLUSION: The results suggest that EEAS could present compound(s) responsible for diuretic activities, and the mechanism could involve the prostaglandin system.

Hypotensive effect of Aspidosperma subincanum Mart. in rats and its mechanism of vasorelaxation in isolated arteries.

ETHNOPHARMACOLOGICAL RELEVANCE: Aspidosperma subincanum is a medicinal herb that is known to be useful for the treatment of cardiovascular-related illnesses. However, its effects and pharmacological mechanisms of action have not been studied. The aim of the present study was to determine the effect of an ethanol extract of Aspidosperma subincanum (EEAS) on blood pressure (in vivo) and vascular tension (in vitro) in the rat thoracic aorta. MATERIALS AND METHODS: Catheters were inserted into the right femoral vein and artery of anesthetized rats for EEAS infusion and the measurement of blood pressure, heart rate and aortic blood flow (flow probes were placed around the aorta). Moreover, the vasodilator effect of EEAS in isolated pre-contracted rat aortas was examined. RESULTS: Intravenous infusion of EEAS resulted in significant and dose-dependent hypotension, bradycardia and increased aortic blood flow. In isolated arteries, EEAS (0-27 µg/mL) induced a concentration-dependent relaxation of pre-contracted aortic rings; endothelial denudation potentiated this effect. Pre-treatment of the aortic rings with ODQ, an inhibitor of soluble guanylyl cyclase (sGC); MDL-12,330A, an inhibitor of adenylyl cyclase (AC); or CPA, a SERCA inhibitor, reduced EEAS-induced vasorelaxation. Treatment with an EEAS impaired contractions induced by phenylephrine (an adrenergic agonist) and Bay K 8644 (an L-type Ca(2+) channel activator). The blockade of K(+) channels with tetraethylammonium, clotrimazole, glibenclamide or 4-aminopyridine reduced the relaxation stimulated by EEAS. CONCLUSIONS: These findings suggest that EEAS induces hypotension associated with bradycardia. EEAS induces endothelium-independent vascular relaxation. The sGC/cGMP and AC/cAMP pathways, SERCA activation and Ca(2+) and K(+) flux across the sarcolemma, are likely involved in this relaxation.