Vasorelaxant and antihypertensive effects of methanolic extracts from Hymenocardia acida Tul

Ethnopharmacological relevance

In Congolese traditional medicine, decoctions of Hymenocardia acida root bark (HaRB) and trunk bark (HaTrB) are used for the treatment of conditions assumed to be hypertension. In this work, we propose to study the vasorelaxant effect of HaRB and HaTrB methanolic extracts on isolated rat thoracic aorta, to characterize the group of molecules responsible for the observed vasorelaxant activity, to evaluate the in vitro antioxidant activity of these extracts and to determine the antihypertensive activity of the HaRB extract on spontaneously hypertensive rats (SHR).

Materials and methods

The vasorelaxant effect of the HaRB and HaTrB methanolic extracts was studied on endothelium-intact aortic rings pre-contracted with phenylephrine (PE, 1 μM). The mechanism of this vasorelaxant effect was investigated on endothelium-denuded vessels and on endothelium-intact aortic rings in the presence of three inhibitors: l-N^G-nitroarginine methyl ester (100 μM), indomethacin (10 μM) and 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (10 μM). To determine the nature of the compounds responsible for the vasorelaxant activity, we carried out a fractionation of the extracts and a thiolysis of the most active fraction followed by a liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) analysis. The extracts antioxidant activity was determined by the 2,2-diphenyl-1-picrylhydrazyl (DPPH) colorimetric assay. In vivo anti-hypertensive activity of the HaRB extract was conducted on SHR.

Results

HaRB and HaTrB methanolic extracts produced a concentration-dependent vasorelaxation on intact aortic rings pre-contracted with PE (1 μM). The vasorelaxant responses obtained were 95.3±1.5% (5 μg/ml) and 100.6±3.0% (1 μg/ml), respectively. The effect was markedly attenuated by removal of endothelium or pretreatment of aortic rings with all inhibitors except indomethacin. The LC/ESI-MS analysis of the thiolysis products indicated that the fraction which caused the most important vasorelaxation (97.9±2.5% at 3 μg/ml) was a mixture of procyanidins and prodelphinidins, with a predominance of procyanidins. Both extracts and all fractions from HaRB extract showed a DPPH scavenging activity, ranging from 0.4 to 0.8 quercetin-equivalents. The HaRB methanolic extract reduced the systolic blood pressure in SHR (from 214±3 mmHg to 194±4 mmHg) after a 5-week treatment.

Conclusions

The methanolic extracts of Hymenocardia acida root and trunk bark have vasorelaxant activity. The vasorelaxant effect observed is endothelium-dependent and seems mainly mediated through the NO-cGMP pathway. The COX pathway is not involved. The vasorelaxant activity appears to be due to polymeric procyanidins and prodelphinidins. These extracts also have an antioxidant effect. The extract of Hymenocardia acida root bark shows a significant but weak antihypertensive activity in SHR.     

Vascular effects and antioxidant activity of two Combretum species from Democratic Republic of Congo

Ethnopharmacological relevance

Combretum racemosum P. Beauv (Combretaceae) leaves (CrLv) and root bark (CrRB) and Combretum celastroides subsp. laxiflorum Welw (Combretaceae) leaves (ClLv) are used in Congolese traditional medicine for several therapeutic purposes, notably for the treatment of conditions consistent with hypertension. The present study aims to investigate the vasorelaxant and in vitro antioxidant activities of these plants polar extracts and to examine the in vivo antihypertensive effect of the extract which displays the most potent vasorelaxant effect.

Material and methods

The vasorelaxant effect of CrLv, CrRB and ClLv methanolic extracts was studied on rat aorta rings pre-contracted with phenylephrine (PE, 1 μM) in the presence or absence of the endothelium. In some experiments, prior to the addition of the extract, rings were incubated for 30 min with either L-N^G-nitroarginine methyl ester (L-NAME; 100 μM), a nitric oxide synthase (NOS) inhibitor, indomethacin (10 μM), a cyclooxygenase inhibitor, or 1 H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ; 10 μM), a guanylate cyclase inhibitor. The antioxidant activity was determined by the measurement of the scavenging ability of extracts towards the stable free radical 2,2-diphenyl-1-picrylhydrazyl (DPPH). Blood pressure was measured on normotensive Wistar rats and spontaneously hypertensive rats (SHR) treated orally with a daily dose (40 mg/kg) of the CILv extract for 5 weeks. Tested extracts have been characterised by TLC profiles targeted at flavonoids.

Results

All tested extracts showed an important DPPH scavenging activity, ranging from 0.6 to 1.1 quercetin-equivalents. They caused a concentration-dependent vasorelaxation on intact aortic rings pre-contracted with PE (1 μM). The responses to CrRB and CrLv methanolic extracts reached 74.0±5.1% and 62.2±8.6% at a cumulative concentration of 50 μg/ml, respectively. The ClLv (10 μg/ml) extract was more active and, in the same conditions, relaxed aortic rings by 90.3±5.8%. The vasorelaxant activity of all extracts disappeared or was significantly attenuated by removal of the endothelium or after pretreatment with L-NAME or ODQ. Indomethacin only inhibited the activity of CrLv and CrRB extracts. The ClLv extract was able to lower the systolic blood pressure in SHR rats by 7% after a 5-week treatment.

Conclusions

The present study shows that methanolic extracts from ClLv, CrRB and CrLv have an antioxidant activity and an endothelium-dependent vasorelaxant effect. ClLv induces the vasorelaxant effect through the NO-cGMP pathway while CrLv and CrRB extracts also act via a prostanoid pathway. ClLv extract demonstrated a modest but significant antihypertensive activity in SHR rats.     

In vitro xanthine oxidase inhibitory activity of the fractions of Erythrina stricta Roxb.

Aim of the study

To assay the in vitro xanthine oxidase inhibitory activity of the various fractions of the hydromethanolic extract of the leaves of Erythrina stricta and to determine its enzyme inhibition mechanism.

Materials and methods

Xanthine oxidase inhibitory activity was assayed spectrophotometrically under aerobic conditions and the degree of enzyme inhibition was determined by measuring the increase in absorbance at 295 nm associated with uric acid formation. Enzyme kinetics was carried out using Lineweaver-Burk plots using xanthine as the substrate.

Results

Among the fractions tested, the chloroform fraction exhibited highest potency (IC₅₀ 21.2 ± 1.6 μg/ml) followed by the pet–ether (IC₅₀ 30.2 ± 2.2 μg/ml), ethyl acetate (IC₅₀ 44.9 ± 1.4 μg/ml) and residual (IC₅₀ 100 ± 3.3 μg/ml) fractions. The IC₅₀ value of allopurinol used, as the standard was 6.1 ± 0.3 μg/ml. Enzyme inhibition mechanism indicated that the mode of inhibition was of a mixed type.

Conclusion

These results suggest that the use of Erythrina stricta for the treatment of gout could be attributed to its xanthine oxidase inhibitory activity.     

Vasorelaxant effects of extracts of the stem bark of Terminalia superba Engler & Diels (Combretaceae)

Aim of the study

The stem bark of Terminalia superba (Combretaceae) (TS) is used in traditional Cameroonian medicine as antihypertensive remedy. In the present study, we investigated the vasorelaxant properties of different extracts of TS and their underlying mechanisms.

Materials and methods

Activities of aqueous (AQU), methanolic (MET), methylene chloride (MC), and methylene chloride–methanol (MCM) extracts of TS were evaluated on isolated rat aortic rings precontracted with phenylephrine (PE) or high KCl.

Results

All extracts induced a vasodilating effect both on KCl- and PE-induced contractions. The effects of MC and MCM extracts were greater than those of AQU or MET extracts (P < 0.05). MC had an endothelium-independent effect and reduced Ca⁺⁺-induced contraction following PE or KCl challenge (P < 0.05). After incubation with verapamil, MC induced a relaxation in rings precontracted by PE (P < 0.001). By contrast, the effect of MCM was endothelium-dependent and decreased by the nitric oxide synthase inhibitor N_W-nitro-l-arginine methyl ester (P < 0.05).

Conclusions

These data demonstrate that the MC extract exhibits vasorelaxant effects that are partly due to inhibition of extracellular Ca⁺⁺ influx and/or inhibition of intracellular Ca⁺⁺ release in vascular smooth muscle cells. By contrast, the effect of the MCM extract was found to be endothelium- and nitric oxide dependent.     

Vasorelaxant effects and mechanisms of action of Heracleum sphondylium L. (Apiaceae) in rat thoracic aorta

Ethnopharmacological relevance

Aerial parts of Heracleum sphondylium L. (HS) are used in traditional medicine to treat hypertension. To provide pharmacological basis for this use, we investigated the vasorelaxant effects of a dichloromethane extract of HS (HSDE) and the mechanisms involved.

Materials and methods

Activity of HSDE was evaluated on rat isolated thoracic aortic rings.

Results

HSDE induced vasorelaxation in phenylephrine (PE, 10⁻⁶ mol/L) and high KCl—(6×10⁻² mol/L) pre-contracted aortic rings that was independent on the presence of endothelium. HSDE markedly decreased extracellular Ca²⁺-induced contraction in high-KCl and PE pre-challenged rings. It also inhibited the intracellular Ca²⁺ release sensitive to PE (10⁻⁶ M). The relaxant effect of HSDE were blunted by 4-amino-pyridine (4-AP, 10⁻³ mol/L), an inhibitor of voltage-dependent K⁺ channels.

Conclusion

Our results provide the first evidence that a dichloromethane extract of Heracleum sphondylium L. exhibits vasorelaxant properties through endothelium-independent mechanisms involving the inhibition of Ca²⁺ mobilization and changes in Kv channel conductances. These data argue for its use as antihypertensive therapy in traditional medicine.     

Vasorelaxant and antihypertensive effect of Cocos nucifera Linn. endocarp on isolated rat thoracic aorta and DOCA salt-induced hypertensive rats

Ethnopharmacological relevance

The fruits of Cocos nucifera Linn. (Arecaceae) have long been used in traditional medicine for the treatment of cardio-metabolic disorders.

Aim of the study

To evaluate the ethanolic extract of Cocos nucifera Linn. endocarp (CNE) for its vasorelaxant activity on isolated rat aortic rings and antihypertensive effects in deoxycorticosterone acetate (DOCA) salt-induced hypertensive rats.

Materials and methods

Cocos nucifera Linn. endocarp was extracted with ethanol and characterized by HPLC. CNE was examined for its in vitro vascular relaxant effects in isolated norepinephrine, phenylephrine or potassium chloride pre-contracted aortic rings (both intact endothelium and denuded). In vivo anti-hypertensive studies were conducted in DOCA salt-induced uninephrectomized male Wistar rats.

Results

Removal of endothelium or pretreatment of aortic rings (intact endothelium) with l-NNA (10 μM) or ODQ (10 μM) followed by addition of contractile agonists prior to CNE significantly blocked the CNE-induced relaxation. Indomethacin (10 μM) and atropine (1 μM) partially blocked the relaxation, whereas glibenclamide (10 μM) did not alter it. CNE significantly reduced the mean systolic blood pressure in DOCA salt-induced hypertensive rats (from 185.3 ± 4.7 mmHg to 145.6 ± 6.1 mmHg). The activities observed were supported by the polyphenols, viz. chlorogenic acid, vanillic acid and ferulic acid identified in the extract.

Conclusions

These findings reveal that the vasorelaxant and antihypertensive effects of CNE, through nitric oxide production in a concentration and endothelium-dependent manner, is due to direct activation of nitric oxide/guanylate cyclase pathway, stimulation of muscarinic receptors and/or via cyclooxygenase pathway.     

Artemisia copa aqueous extract as vasorelaxant and hypotensive agent

Ethnopharmacological relevance

Artemisia copa Phil. (Asteraceae) is a medicinal plant commonly used in traditional medicine in Argentina.

Aim of the study

The vasorelaxant and hypotensive activities of the aqueous extract of Artemisia copa have been investigated.

Materials and methods

The in vitro effect of the extract and isolated compounds from Artemisia copa was investigated using isolated rat aortic rings. The acute effect caused by the intravenous (i.v.) infusion (0.1–300 mg/kg) on blood pressure and heart rate was evaluated in spontaneous hypertensive rats. In addition, a phytochemical analysis of the extract was performed by HPLC.

Results

Artemisia copa had a relaxant effect in endothelium-intact aortic rings that had been pre-contracted with 10⁻⁷ M phenylephrine (E_max=96.7±1.3%, EC₅₀=1.1 mg/ml), 10⁻⁵ M 5-hydroxytriptamine (E_max=96.7±3.5%, EC₅₀=1.5 mg/ml) and 80 mM KCl (E_max=97.9± 4.4%, EC₅₀=1.6 mg/ml). In denuded aortic rings contracted by phenylephrine, a similar pattern was observed (E_max=92.7±6.5%, EC₅₀=1.8 mg/ml). l-NAME, indomethacin, tetraethylammonium and glibenclamide were not able to block the relaxation induced by the extract. Nevertheless, the pre-treatment with Artemisia copa attenuated the CaCl₂-induced contraction in a concentration-dependent manner (E_max: 86% of inhibition for 3 mg/ml and 52% de-inhibition for 1 mg/ml). This pre-treatment also induced a significant attenuation of the norepinephrine-induced contraction in a concentration-dependent manner (E_max: 72.7% of inhibition for 3 mg/ml and 27% de inhibition for 1 mg/ml) in a Ca²⁺ free medium. Upon analyzing the composition of the extract, the presence of p-coumaric acid, isovitexin, luteolin and chrysoeriol were found. Luteolin (CE₅₀: 1.5 μg/ml), chrysoeriol (CE₅₀: 13.2 μg/ml) and p-coumaric acid (CE₅₀: 95.2 μg/ml), isolated from the aqueous extract, caused dilatation of thoracic aortic rings pre-contracted with phenylephrine. Artemisia copa administered i.v. also induced a decrease in the mean arterial pressure but did not affect the heart rate in hypertensive rats.

Conclusions

The aqueous extract of Artemisia copa proved to have vasorelaxing and hypotensive effects through the inhibition of Ca²⁺ influx via membranous calcium channels and intracellular stores. The presence of luteolin, chrysoeriol and p-coumaric acid found in this plant could be involved in this effect.     

Sedative and anticonvulsant activities of methanol extract of Dorstenia arifolia in mice

Ethnopharmacological relevance

Dorstenia arifolia is a plant that has been used in the folk medicine to produce hypnotic, sedative and ansiolitic effects but the pharmacological properties have not yet been studied. In addition, the smoke of its rhizome is reputed to induce lethargic sensation.

Aims of the study

The present study investigated possible activities of the methanol extract (ME) of Dorstenia arifolia rhizome on the central nervous system (CNS).

Materials and methods

ME was tested for sedative, hypnotic and anticonsulsant effects using locomotor activity evaluation, pentobarbital-induced sleeping time and pentylenetetrazol (PTZ)-induced convulsion, respectively.

Results

Intraperitoneal administration of ME (10 and 50 mg/kg) significantly decreased locomotor activity from 205.2 ± 25.6 movements/min (DMSO) to 112.1 ± 18.4 (P < 0.05) and 114.9 ± 16.9 (P < 0.05), respectively. Flumazenil (10 mg/kg), an antagonist of GABA_A receptor, prevented the ME-induced sedation. Treatment with ME (50 mg/kg) significantly increased the duration of pentobarbital-induced sleeping time from 41.0 ± 2.3 to 57.9 ± 2.9 min (P < 0.05). The latencies to seizures after intraperitoneal injection of PTZ was recorded and compared between groups. ME promoted a significant protection of PTZ-induced seizures and mortality in a dose-dependent manner.

Conclusions

Our findings indicate that ME of Dorstenia arifolia rizhome has pronounced central effects, and that the sedative and anticonvulsant activities may be related to a facilitation of the GABAergic transmission.     

Hypotensive and endothelium-independent vasorelaxant effects of methanolic extract from Curcuma longa L. in rats

Ethnopharmacological relevance

Curcuma longa L. (CL) is a yellow rhizome that is used in African traditional medicine to treat palpitation, hypertension or other related blood circulation disorders.

Aim of the study

To justify the use of CL in ethnomedicine, we investigated the vasorelaxant effect of methanolic extract of CL (CLME) and its underlying mechanisms in isolated rat mesenteric artery.

Materials and methods

The effect of CLME on the mean arterial pressure (MAP) and heart rate (HR) (pulse interval) were determined in vivo in non-anaesthetized rats. Superior mesenteric rings were isolated, suspended in organ baths containing Tyrode solution at 37 °C and gassed with 95% O₂ + 5% CO₂, under a resting tension of 0.75 g. The vasorelaxant effects of CLME were studied by means of isometric tension recording experiments.

Results

In normotensive rats, CLME (10, 20 and 30 mg/kg, i.v.) induced dose-dependent hypotension (2.0 ± 0.5%; 27.1 ± 5.0% and 26.7 ± 4.6%, respectively), and pronounced bradycardia (5.8 ± 1.2%, 19.3 ± 3.2% and 22.9 ± 4.6%, respectively). CLME (1–1000 μg/mL) induced concentration-dependent relaxation of tonic contractions evoked by phenylephrine (Phe) (10 μM) and KCl (80 mM) in rings with intact-endothelium (E_max = 82.3 ± 3.2% and 97.7 ± 0.7%) or denuded-endothelium (E_max = 91.4 ± 1.0% and 97.8 ± 1.1%). Also, in a depolarized, Ca²⁺ free medium, CLME inhibited CaCl₂ (1 μM–30 mM)-induced contractions and caused a concentration-dependent rightward shift of the response curves, indicating that CLME inhibited the contractile mechanisms involving extracellular Ca²⁺ influx. In addition, in Ca²⁺ free media containing EGTA (1 mM), CLME inhibited the transient contraction of denuded rings constricted with Phe, but not those evoked by caffeine (20 mM). In contrast, neither glibenclamide, BaCl₂, tetraethylammonium nor 4-aminopyridine affected CLME-induced relaxation.

Conclusions

These results demonstrate the hypotensive and bradycardic effects of CLME, as well as its potent vasodilation of rat mesenteric arteries. These effects, may in part, be due to the inhibition of extracellular Ca²⁺ influx and/or inhibition of intracellular Ca²⁺ mobilization from Phe-sensitive stores.     

Vasoactive effects of different fractions from two Panamanians plants used in Amerindian traditional medicine

Ethnopharmacological relevance

Cecropia obtusifolia (Cecropiaceae) and Psychotria poeppigiana (Synonym: Cephaelis elata, Rubiaceae) are two Latin American plants broadly used in traditional Amerindian medicine. The former, together with many other species of the genus Cecropia, share the folk reputation of curing heart failure, cough, asthma and bronchitis. The latter is used in Panama by Kuna and Ngäbe Buglé (Guaymies) native Indians for the treatment of dyspnea.

Aim of the study

Based on screening of selected medicinal Panamanian plants by radioligand-binding techniques by Caballero-George et al. (2001), the present study was carried out in order to investigate the vasoactive effects of different fractions from both P. poeppigiana and C. obtusifolia on rat thoracic aorta and identify active fractions and their chemical constituents.

Materials and methods

Both acid and neutral methanol fractions (P-AMeOH and P-NMeOH) and acid and neutral dichlorometane fractions (P-ADCM and P-NDCM) were obtained from P. poeppigiana crude methanolic and dichlorometane extracts, respectively. Identical fractionation was carried out for C. obtusifolia (C-AMeOH, C-NMeOH, C-ADCM and C-NDCM. Vasorelaxant effect of all fractions, and their inhibition of contractile responses to angiotensin II were evaluated in isolated aortic rings.

Results

P-AMeOH, P-NMeOH and P-ADCM fractions induced a concentration-dependent relaxation (43.9 ± 1.8%, 35.3 ± 4.7% and 52.9 ± 3.5%, respectively) in the endothelium-intact aorta precontracted by phenylephrine (PE, 10⁻⁶ M). The relaxation produced by C-AMeOH and C-NMeOH (57.3 ± 2.5% and 53.3 ± 3.3%, respectively) was greater than the effect produced by C-ADCM and C-NDCM (42.2 ± 3.4% and 21.8 ± 0.8%, respectively). Only the incubation of the aortic rings with P-AMeOH reduced the maximum contraction induced by angiotensin II at 20.08 ± 0.55%.

Conclusions

The direct vasorelaxation effect observed could explain in part the ethnomedical use of these plants in Amerindian traditional medicine. The most active fractions contain phenolic and aromatic acid compounds. Furthermore, P-AMeOH, the only fraction that showed both vasorelaxant effect and inhibition of contractile responses to angiotensin II, is the most rich in aromatic acids compounds and the only one that contains scopoletin.     

Characterization of the antinociceptive and anti-inflammatory activities of fractions obtained from Copaifera multijuga Hayne

Ethnopharmacological relevance

Copaifera multijuga Hayne (Leguminosae) is a tree that produces an oleoresin, which is extensively commercialized in Brazil as capsules or crude oil for the treatment of several disorders. Ethnopharmacological studies show a diversity of indications such as anti-inflammatory and epidermal wound cicatrization.

Aim of the study

In the present work three fractions obtained from Copaifera multijuga oleoresin (hexane (HF), chloroform (CF), and methanol (MF) from a KOH impregnated silica gel column chromatography, representing the three main classes of compounds in the Copaifera genus (hydrocarbon sesquiterpenes, oxygenated sesquiterpenes and acidic diterpenes), were evaluated using antinociceptive and anti-inflammatory models.

Materials and methods

HF, CF, and MF (doses ranging between 1 and 150 mg/kg, depending on the model used), Copaifera multijuga oleoresin (CMO, 100 mg/kg, p.o.) and the reference drug morphine (5 mg/kg, p.o.) were evaluated using models for analgesia (acetic acid-induced contortions and tail flick) or inflammation (rat paw oedema and increase in vascular permeability). To elucidate the mechanism of action from the fractions, animals were pre-treated with naloxone (opioid receptor antagonist, 5 mg/kg, i.p.).

Results

Fractions significantly inhibited (in a concentration-dependant way) the number of contortions induced by acetic acid and the second phase of formalin-induced licking response. Similar results were observed in the tail flick model. The central antinociceptive effect for HF and CF at the doses of 50 and 100 mg/kg was higher than the one observed for morphine (1 mg/kg). Administration of naloxone inhibited the antinociceptive effect of fractions indicating that HF, CF, and MF may be acting on opioid receptors. All three fractions also inhibited rat paw oedema and the increase in vascular permeability induced by several phlogistic agents (carrageenan, histamine, and serotonin).

Conclusions

Our results indicate that fractions obtained from Copaifera multijuga Hayne demonstrate an antinociceptive effect probably mediated by opioid receptors, and anti-inflammatory activity through inhibition of histaminergic and serotoninergic pathways.     

MAO-A inhibitory activity of quercetin from Calluna vulgaris (L.) Hull.

Aim of the study

This study investigated MAO-A inhibitory activity of methanol extract of Calluna vulgaris (L.) Hull., which traditionally has been used as a nerve calming remedy.

Materials and methods

A methanolic extract of Calluna vulgaris was partitioned against heptane, ethyl acetate and water. The three fractions were tested in a photometric peroxidase linked MAO-A bioassay. The ethyl acetate phase showed the highest MAO-A inhibitory activity. Quercetin was isolated by VLC through bioassay-guided fractionation and purified by re-crystallisation. The structure was elucidated by LC–MS and ¹H NMR.

Results

The IC₅₀-value for MAO-A inhibition by quercetin was 18 ± 0.2 μM in an assay where the IC₅₀-value for MAO-A inhibition by clorgylin was 0.2 ± 0.02 μM.

Conclusion

The content of quercetin in Calluna vulgaris might explain the reported nerve calming effect of the plant.     

Antidiabetic potential of unripe Carissa carandas Linn. fruit extract

Ethnopharmacological relevance

Carissa carandas commonly known as Karanda have a long history of use in traditional system of medicine. It is used by tribal healers of Western Ghat region of Karnataka as hepatoprotective and antihyperglycemic. However, no scientific data is available to validate the folklore claim. The present study has been designed to evaluate its unripe fruit for the antidiabetic activity.

Aim

In the present study, methanol extract of unripe fruits and its fractions were studied for its antidiabetic potential.

Materials and methods

The methanol extract and its fractions were screened for antidiabetic activity in alloxan induced diabetic rats. The polyphenolic, flavonoid and flavanone contents of methanolic extract and its fractions were also determined and correlated with its antidiabetic activity.

Results

The experimental data indicated that the methanol extract and its ethyl acetate soluble fraction has significantly lowered the elevated blood glucose levels by 48% (p < 0.001) and 64.5% (p < 0.001) respectively at dose level of 400 mg/kg per oral after 24 h as compared to diabetic control. In order to assess the role of polyphenolic components in the relevant activity, polyphenolic and flavonoid contents were determined. The polyphenolic and flavonoid content of methanol extract and its ethyl acetate soluble fraction were found to be 15.8 ± 1.2 mg and 18.55 ± 0.34 mg (gallic acid equivalent/g extract) and flavonoid content 2.92 ± 0.03 mg and 1.534 ± 0.30 mg (rutin equivalent/g extract) respectively.

Conclusion

The increased antidiabetic potential of ethyl acetate fraction over methanol extract is due to its partial purification achieved by fractionation which resulted in increase in degree of polymerization and segregation of secondary metabolites.     

Antihypertensive activity of Salvia elegans Vahl. (Lamiaceae): ACE inhibition and angiotensin II antagonism

Ethnopharmacological relevance

Salvia elegans Vahl. (Lamiaceae), recognized with the popular name of “mirto” is widely used in Mexico for healing purposes, and also them as antihypertensive treatment.

Aim of the study

The high prevalence of this illness and the side effects of antihypertensive drugs conducted us to the evaluation of the Salvia elegans extract on angiotensin II action.

Materials and methods

The acute response of blood pressure to angiotensin II administration was measured in mice. We also tested in vitro the inhibitory effect on angiotensin convertase enzyme. Additionally, characterization of the pharmacological effect of the extract fraction was obtained.

Results

We obtained dose–response curve for the administration of complete extract and extract fractions. Due to the hydroalcoholic extract (SeHA) treatment blood pressure decreased significantly from systolic dose of 0.75 mg kg⁻¹ (p < 0.05) and even had an antihypertensive effect that was greater than that treatment with losartan. SeHA extract decreased the E_max of the AG II hypertensive effect by about 20% in both systolic and diastolic pressures, treatment with losartan also decreased the same parameter between 6% and 8% for systolic and diastolic pressures, respectively. Fractions SeF8 and SeF8-8 showed similar levels of AG II ED₅₀ for both pressures compared with losartan, these fractions showed major compounds with maximum absorbance peaks at 221, 289 and 330 nm typical of flavonoids. In the inhibition assay the activity of angiotensin converting enzyme (ACE), the extract SeHA showed percentage inhibition (%IACE) of 50.27 ± 5.09% (n = 5). SeBuOH fraction is found to have greater inhibitory capacity of achieving a IACE 78.40 ± 2.24% (n = 5), which was similar to the values obtained in the presence of the SeF8-22 fraction (82.61 ± 1.74%) and lisinopril (87.18 ± 1.16%). The changes in the value of K_M suggest that components of the extracts and fractions were recognized by the enzyme's active site. The main compounds of the fractions SeBuOH, SeF8-22 were by flavonoid and phenyl propanoid types, according to UV absorption spectra of the fractions.

Conclusions

The experimental results demonstrated the antihypertensive effect of Salvia elegans and it was due to the AG II antagonism and inhibition of angiotensin converting enzyme.     

Hypoglycemic effect of quassinoids from Brucea javanica (L.) Merr (Simaroubaceae) seeds

Ethnopharmacological relevance

The seeds of Brucea javanica (L.) Merr (Simaroubaceae) are recommended by traditional practitioners for the treatment of diabetes mellitus.

Aim of the study

To identify the compounds responsible for blood glucose lowering effect and evaluate the acute toxicity of the compounds.

Materials and methods

Extracts, fractions and subfractions were administered to normoglycemic mice and the blood glucose concentration was monitored for 8 h. Bioactive compounds isolated through column chromatography were administered to normoglycemic mice and streptozotocin (STZ) rats with monitoring of blood glucose concentration at 0–8 h. The acute toxicity was evaluated in mice.

Results

Bioactivity-guided fractionation led to the isolation of bruceines E (1) and D (2). Normoglycemic mice administered with 1 mg/kg of 1 and 2 exhibited significant blood glucose concentration reduction of 40.07 ± 11.45% and 48.82 ± 13.34%, respectively. STZ induced diabetic rats administered with 1 and 2 exhibited significant blood glucose concentration reduction of 73.57 ± 13.64% and 87.99 ± 2.91%, respectively.

Conclusion

The reduction of blood glucose concentration by both bruceines was comparable to glibenclamide and they might act as an insulin secretagogue. The presence of a hydroxyl moiety at C₂ in 1 reduced the toxic effect by 9-fold compared to 2.     

Antiplasmodial activity of root extract and fractions of Croton zambesicus

Aim of the study

Antiplasmodial activity of root extract and fractions of Croton zambesicus were evaluated to ascertain the folkloric claim of its antimalarial activity and elucidate its antiplasmodial mechanism of action.

Material and method

The crude ethanolic root extract (27–81 mg/kg) and gradient fractions ( n- hexane, chloroform, ethyl acetate and methanol; 54 mg/kg) of Croton zambesicus were investigated for antiplasmodial activity against chloroquine - sensitive Plasmodium berghei infections in mice. The antiplasmodial activity during early and established infections as well as the prophylactic activity were investigated. Chloroquine (5 mg/kg) and pyrimethamine (1.2 mg/kg) were used as positive controls. Thin films made from tail blood of each mouse were used to assess the level of parasitaemia of the mice. Oxidant generation potentials of the crude extract and fractions was also evaluated to elucidate their mechanism of action.

Results

The crude root extract (27 – 81 mg/kg) demonstrated significant (P < 0.01–0.001) schizonticidal activity during early and established infections and also had prophylactic activity. The activity was comparable to that of the standard drug used (chloroquine 5 mg/kg, pyrimethamine 1.2 mg/kg). Methanol, ethyl acetate and chloroform fractions had comparative in vivo antiplasmodial activity and oxidant generation potentials.

Conclusion

The antiplasmodial activity of this root extract and fractions which is likely to be through peroxidation confirms the folkloric use of this plant.     

Pharmacological mechanisms underlying the vascular activities of Loranthus ferrugineus Roxb. in rat thoracic aorta

Aim of the study

The present study was aimed to investigate the pharmacological basis for the use of Loranthus ferrugineus in hypertension.

Materials and methods

Loranthus ferrugineus methanol extract (LFME) was obtained using Soxhelt extractor and then successively fractionated using chloroform, ethyl acetate and n-butanol. The n-butanol fraction of LFME (NBF-LFME) was studied using isolated rat thoracic aorta.

Results

NBF-LFME (1.0 × 10⁻⁵ to 3.0 mg/ml) was found to be the most potent to concentration-dependently relax the endothelium-intact phenyephrine (PE, 1 μM)- and high K⁺ (80 mM)-precontracted rat aortic rings. Removal of the endothelium completely abolished the vascular relaxing properties of NBF-LFME. Pretreatment with atropine (1 μM), l-NAME (10 μM), indomethacin (10 μM) and methylene blue (10 μM) significantly blocked NBF-LFME-mediated relaxation. Endothelium-dependent and -independent relaxations induced by acetylcholine (ACh) and sodium nitroprusside (SNP), respectively, were significantly enhanced in aortic rings pretreated with NBF-LFME when compared to those observed in control aortic rings. On the contrary, glibenclamide (10 μM), propranolol (1 μM) and prazosin (0.01 μM) did not alter NBF-LFME-induced relaxation.

Conclusions

The results suggest that NBF-LFME induced vascular relaxation by stimulating muscarinic receptors, activating the endothelium-derived nitric oxide-cGMP-relaxant pathway, promoting prostacyclin release and/or possibly through its ability to lengthen the released nitric oxide half-life. The present data further supports previous in vivo findings and explain the traditional use of Loranthus ferrugineus as an anti-hypertensive agent.     

Isolation of seselin from Clausena anisata (Rutaceae) leaves and its effects on the feeding and development of Lucilia cuprina larvae may explain its use in ethnoveterinary medicine

Ethnopharmacological relevance

The leaves of Clausena anisata are used traditionally to expel maggots from wounds of animals in Zimbabwe. We have previously proved in the laboratory that the plant certainly affects the behaviour and growth of blowfly larvae. The objective of this study was to isolate and identify the active compounds responsible for this activity.

Materials and methods

The acetone extract of Clausena anisata leaf powder was separated by solvent–solvent partition into five fractions. The n-hexane fraction was the most active in the larvicidal assay and therefore subjected to open column chromatography on silica gel.

Results

The isolated compound was identified by nuclear magnetic resonance (NMR) and mass spectroscopy (MS) as the pyranocoumarin, seselin, chemically known as 2′,2′-dimethylpyranocoumarin. It inhibited feed intake in the first and second instars of blowfly larvae at the minimum concentration tested of 1 ppm resulting in significant lower mass pupae (13.5±0.5 mg and 22.4±0.4 mg for the first and second instar larvae respectively) compared to the solvent control group (26.19±0.8 mg) (p<0.05).

Conclusions

This is the first report of the isolation of seselin from the leaves of Clausena anisata and the first report of the compound having an effect against blowfly larvae.     

Vasorelaxant effects of Cerebralcare Granule are mediated by NO/cGMP pathway,potassium channel opening and calcium channel blockade in isolated rat thoracic aorta

Ethnopharmacological relevance

Cerebralcare Granule (CG), one of the famous classical recipes in traditional Chinese medicine, is developed from the “Decoction of Four Drugs”. It has been used for treatment of cerebrovascular related diseases, such as hypertension. It is well known that vasodilatation plays a very important role in hypertensive. Despite the popular medicinal use of CG, little data was available to its activity and mechanism involved in vasodilatation. Therefore, we aimed to investigate the vasorelaxant effects of CG on isolated rat thoracic aorta so as to assess some of the possible mechanisms. The present study was performed to examine the vasodilative activity of CG and its mechanisms in isolated rat thoracic aorta.

Materials and methods

CG was studied on isolated rat thoracic aorta in vitro, including endothelium-intact and endothelium-denuded aortic rings. In present study, specific inhibitors including NO synthase inhibitor N^G-nitro-l-arginine methyl ester (l-NAME), cyclooxygenase (COX) inhibitor indomethacin (INDO), non-selective K⁺ channel inhibitor tetraethylammonium chloride (TEA), K_ir channel inhibitor BaCl₂, K_ATP channel inhibitor Glibenclamide (Gli) and cholinergic receptor antagonist atropine were used, they were added 20 min before NE contraction and then added CG-induced vasodilation.

Results

Removal of endothelium or pretreatment of aortic rings (intact endothelium) with l-NAME (0.1 mM) or INDO (0.01 mM) significantly blocked the CG induced relaxation. Pretreatment with the non-selective K⁺ channel inhibitor TEA (1 mM), or the K_ir channel inhibitor BaCl₂ (0.1 mM), neither of them had no influence on the CG-induced response (p>0.05). However, pretreatment with the K_ATP channel inhibitor Gli (0.01 mM) produced significant inhibition on the CG-induced response (p<0.01). Besides, CG also inhibited the contraction triggered by NE in endothelium-denuded rings in Ca²⁺-free medium. CG (0.4, 0.8 and 3.2 mg/mL) produced rightward parallel displacement of CaCl₂ curves and reduced the maximum contraction induced by 30 mM CaCl₂ to 31.1±9.3%, 18.8±6.9% and 9.4±4.5%, respectively. The relaxation, induced by CG on endothelium-intact rat aortic rings pre-contracted with NE, was significantly attenuated in the presence of atropine (EC₅₀=3.7 mg/mL, p<0.01).

Conclusions

Our results suggest that CG induces relaxation in rat aortic rings through an endothelium-dependent pathway mediated by NO/cGMP pathway and an endothelium-independent pathway involving blockade of Ca²⁺ channels, inhibition of Ca²⁺ mobilization from intracellular stores, opening of K_ATP channel. In addition, the muscarinic receptor stimulation is also one of the vasorelaxant mechanisms.     

Cochlospermum vitifolium induces vasorelaxant and antihypertensive effects mainly by activation of NO/cGMP signaling pathway

Aim of the study

Cochlospermum vitifolium is a medicinal plant used for the treatment of diabetes, hepatobilary and cardiovascular illnesses. The aim of current study was to determine the in vivo antihypertensive and in vitro functional vasorelaxant mechanism of methanol extract of Cochlospermum vitifolium (MECv) and naringenin (NG).

Materials and methods

Test material was assayed on rat isolated aorta rings test with- and without-endothelium to determine their vasorelaxant mechanism. Also, the in vivo antihypertensive effect was evaluated on spontaneously hypertensive rat (SHR) model. In addition, presence of NG into the extract was confirmed by reverse phase high performance liquid chromatography (RP-HPLC) analysis.

Results

MECv (120 mg/kg) and NG (50 and 160 mg/kg) showed acute antihypertensive effects on SHR when systolic and diastolic pressure were decreased at 1 h and 24 h after administration, respectively. Vasorelaxant effect of MECv and NG was shifted to the right when endothelium-intact aortic rings were pre-incubated with L-NAME (10 μM) and ODQ (1 μM). Also, NG relaxant curves were displaced to the right in the presence of tetraethylammonium (TEA, 1 mM) and 2-aminopyridine (2-AP, 100 μM) on endothelium-denuded aortic rings.

Conclusion

Experiments described above showed that MECv play an important role in hypertension regulation through NO synthesis and may be PGI₂ production and potassium channel activation on excessive endothelial dysfunction conditions. Unfortunately, presence of NG into the extract is not significant on bioactivity of the extract; however, this compound could be tested and evaluated as structural scaffold for future drug design for development of antihypertensive agents.