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.     

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.     

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.     

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.     

Antispasmodic effect of Mentha piperita essential oil on tracheal smooth muscle of rats

Aim of the study

Mentha piperita is a plant popularly known in Brazil as “hortelã-pimenta” whose essential oil is used in folk medicine for its anti-inflammatory, antispasmodic, expectorant actions and anti-congestive. Here, it was investigated the effect of Mentha piperita essential oil (peppermint oil) in rat tracheal rings along with its mechanism of action.

Materials and methods

Tracheal tissue from Male Wistar rats (250–300 g) were used. Peppermint oil was added in cumulative concentrations [1–300 μg/ml] to the tissue basal tonus or pre-contracted by carbachol [10 μM] at 10 min intervals, incubated or not with indomethacin [10 μM], l-N-metyl-nitro-arginine [100 μM], hexamethonium [500 μM], or tetraethylammonium [5mM].

Results

Peppermint oil [100 and 300 μg/ml] inhibited the contractions induced by carbachol, which was reversed by indomethacin, l-N-metyl-nitro-arginine and hexamethonium, but not by tetraethylammonium. These data suggest the participation of prostaglandin E₂, nitric oxide and autonomic ganglions in the peppermint oil relaxant effect and may be correlated with its popular use in respiratory diseases.

Conclusions

Peppermint oil exhibited antispasmodic activity on rat trachea involving prostaglandins and nitric oxide synthase.     

Comparative antioxidant and anti-inflammatory effects of [6]-gingerol, [8]-gingerol, [10]-gingerol and [6]-shogaol

Ethnopharmacological relevance

Zingiber officinale Rosc. (Zingiberaceae) has been traditionally used in Ayurvedic, Chinese and Tibb-Unani herbal medicines for the treatment of various illnesses that involve inflammation and which are caused by oxidative stress. Although gingerols and shogaols are the major bioactive compounds present in Zingiber officinale, their molecular mechanisms of actions and the relationship between their structural features and the activity have not been well studied.

Aim of the study

The aim of the present study was to examine and compare the antioxidant and anti-inflammatory activities of gingerols and their natural analogues to determine their structure–activity relationship and molecular mechanisms.

Materials and methods

The in vitro activities of the compounds [6]-gingerol, [8]-gingerol, [10]-gingerol and [6]-shogaol were evaluated for scavenging of 1,1-diphenyl-2-picyrlhydrazyl (DPPH), superoxide and hydroxyl radicals, inhibition of N-formyl-methionyl-leucyl-phenylalanine (f-MLP) induced reactive oxygen species (ROS) production in human polymorphonuclear neutrophils (PMN), inhibition of lipopolysaccharide induced nitrite and prostaglandin E₂ production in RAW 264.7 cells.

Results

In the antioxidant activity assay, [6]-gingerol, [8]-gingerol, [10]-gingerol and [6]-shogaol exhibited substantial scavenging activities with IC₅₀ values of 26.3, 19.47, 10.47 and 8.05 μM against DPPH radical, IC₅₀ values of 4.05, 2.5, 1.68 and 0.85 μM against superoxide radical and IC₅₀ values of 4.62, 1.97, 1.35 and 0.72 μM against hydroxyl radical, respectively. The free radical scavenging activity of these compounds also enhanced with increasing concentration (P < 0.05). On the other hand, all the compounds at a concentration of 6 μM have significantly inhibited (P < 0.05) f-MLP-stimulated oxidative burst in PMN. In addition, production of inflammatory mediators (NO and PGE₂) has been inhibited significantly (P < 0.05) and dose-dependently.

Conclusions

6-Shogaol has exhibited the most potent antioxidant and anti-inflammatory properties which can be attributed to the presence of α,β-unsaturated ketone moiety. The carbon chain length has also played a significant role in making 10-gingerol as the most potent among all the gingerols. This study justifies the use of dry ginger in traditional systems of medicine.     

Sesquiterpenoids from antidiabetic Psacalium decompositum block ATP sensitive potassium channels

Ethnopharmacological relevance

The hypoglycemic effect of root and rhizome aqueous decoction of Psacalium decompositum (Asteraceae), a medicinal herb from Mexico, has been experimentally demonstrated, leading to the identification of several hypoglycemic sesquiterpenoids, such as cacalol, and the mixture of 3-hydroxycacalolide, and epi-3-hydroxycacalolide; however, the mechanism of action of these compounds is unknown.

Aim of the study

To establish whether cacalol, cacalone epimer mixture and cacalol acetate may block adenosine triphosphate-sensitive potassium channels (K_ATP channels) in a similar way to the antidiabetic drug glibenclamide.

Materials and methods

Cacalol, cacalone epimer mixture, and cacalol acetate were tested on the diazoxide-induced relaxation of male rat aortic rings precontracted with phenylephrine (3.2 × 10⁻⁶ M).

Results

Cacalol (10⁻⁵ M), cacalol acetate and the cacalone epimer mixture (10⁻⁴ M) inhibited the diazoxide effect, in a similar manner and concentration as glibenclamide (10⁻⁵ M). Cacalone epimer mixture exerted this effect in a concentration-dependent manner (P < 0.01). Cacalol (10⁻⁴ M), irreversibly inhibited the diazoxide-induced relaxation, and displayed activity at a lower concentration (10⁻⁵ M) than cacalone epimer mixture and cacalol acetate.

Conclusions

These results suggest that the studied compounds block K_ATP channels in a similar way to glibenclamide in rat aorta. However, controversial data indicate that Psacalium decompositum sesquiterpenoids are less effective than glibenclamide in lowering plasma glucose levels, suggesting that cacalol and cacalone epimer mixture, as well as cacalol acetate, may display a higher affinity to SUR2 subunit of K_ATP channels in aortic smooth muscle than to SUR1 subunit in pancreatic β-cells.     

Synergistic vasorelaxant and antihypertensive effects of Ligusticum wallichii and Angelica gigas

Aim of the study

The synergistic vasorelaxant and antihypertensive effects of Ligusticum wallichii and Angelica gigas were examined in isolated rat aorta rings and spontaneously hypertensive rats (SHRs).

Materials and methods

The ethanol extract of Ligusticum wallichii (LwEx) or Angelica gigas (AgEx) or their combinations at ratios Ligusticum wallichii:Angelica gigas = 1:1 (MxEx11), 1:3 (MxEx13), and 3:1 (and MxEx31), and their successive water soluble (LwDw, AgDw, MxDw11, MxDw13 and MxDw31) or n-butanol soluble fractions (LwBt, AgBt, MxBt11, MxBt13, and MxBt31) were examined for their vasorelaxant effects. In an antihypertensive study, LwEx, AgEx, or MxEx11 (100 mg/kg) was orally administered to SHRs, and the systolic, diastolic, and mean blood pressure were measured using the tail-cuff method before and 1, 3, 5, 7, and 24 h after oral administration.

Results

Each of the ethanol extracts caused long-term relaxation in endothelium-intact or endothelium-denuded rat aorta preconstricted with norepinephrine (NE, 300 nM). All of the water phases of the ethanol extracts elicited an endothelium-dependent acute relaxation, and the water phase of MxDw11 (EC₅₀ values: 1.08 mg/mL, P < 0.05) had the highest activity. MxDw11-induced acute relaxation was abolished by pretreatment with N^G-nitro-l-arginine (10 μM), methylene blue (1.0 μM), or atropine (0.1 μM), indicating that the response to MxDw involves the enhancement of the nitric oxide-cGMP system. On the other hand, all of the butanol phases showed an endothelium-independent long-term relaxation, and MxBt11 (85 ± 7% relaxation of NE-preconstricted active tone at 20 min after the addition, P < 0.05) displayed the highest activity. MxBt11-induced gradual relaxation was significantly attenuated by an inward rectifier potassium-channel inhibitor, but not by an ATP-sensitive or a large conductance Ca²⁺-activated potassium-channel blocker. Calcium concentration-dependent contraction curves in high-potassium, depolarizing medium were shifted significantly to the right and downward after incubation with MxBt11 (0.03, 0.1, and 0.3 mg/mL), implying that MxBt11 is also involved in the inhibition of extracellular calcium influx to vascular smooth muscle. MxEx11 (100 mg/kg) significantly reduced systolic blood pressure of SHRs at 3, 5, and 7 h after oral administration, but this effect was not induced by Ligusticum wallichii or Angelica gigas alone.

Conclusions

The combination of Ligusticum wallichii and Angelica gigas elicits a synergistic effect on vasorelaxation in isolated rat aortas and antihypertension in SHRs. The ratio of Ligusticum wallichii:Angelica gigas = 1:1 was the most effective of all combinations tested.     

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.     

Endothelium-dependent and independent vasorelaxation induced by an n-butanolic fraction of bark of Scutia buxifolia Reiss (Rhamanaceae)

Ethnopharmacological relevance

Scutia buxifolia has been widely used in Brazilian folk medicine as an anti-hypertensive agent.We evaluated the vascular effects and mechanism involved in the relaxation of aorta induced by an n-butanolic fraction (BuOH) from Scutia buxifolia.

Materials and Methods

Rat aortic rings precontracted by phenylephrine (1 μM) were exposed to cumulative concentrations (3–3000 μg/ml) of crude extracts or fractions obtained from bark or leaves of Scutia buxifolia. Classical receptor antagonists, channel and enzymatic inhibitors were used to check the mechanisms involved.

Results

The crude extracts of both leaves and bark of Scutia buxifolia, as well as several fractions, were able to induce partial or total relaxation of rat aortic rings. The BuOH fraction of bark of Scutia buxifolia was the most potent in endothelium-intact (E+) preparations, and also induced a partial, but very significant relaxation in endothelium-denuded (E−) vessels. The non-selective nitric oxide synthase inhibitor L-NAME, as well as the soluble guanylate cyclase inhibitor ODQ, vanished the relaxation in E+. In E− preparations, K⁺ channel blockers, such as tetraethylammonium, glibenclamide, 4-aminopyridine, and the large-conductance calcium-activated K⁺ channel blocker iberiotoxin, were able to significantly reduce the maximum relaxation elicited by BuOH fraction.

Conclusion

Our results demonstrated that BuOH fraction obtained from barks of Scutia buxifolia induced both endothelium-dependent and -independent relaxation in rat aortic rings. The endothelium-dependent relaxation is fully dependent on NO/cGMP system, while direct activation of K⁺ channels may explain, at least in part, the endothelium-independent relaxation induced by BuOH fraction of Scutia buxifolia.     

Antinociceptive and anti-inflammatory kaempferol glycosides from Sedum dendroideum

Aim of the study

To identify the compounds responsible for the antinociceptive and anti-inflammatory effects previously described for Sedum dendroideum, through bioassay-guided fractionation procedures.

Materials and methods

Antinociceptive activity was evaluated through mouse acetic acid-induced writhing model. The anti-inflammatory activity was assessed through croton oil-induced mouse ear oedema and carrageenan-induced peritonitis.

Results

The Sedum dendroideum juice afforded seven known flavonoids identified with basis on NMR data. The oral administration of the major kaempferol glycosides kaempferitrin [1] (17.29 μmol/kg), kaempferol 3-O-β-glucopyranoside-7-O-α-rhamnopyranoside [2] (16.82 μmol/kg), kaempferol 3-O-neohesperidoside-7-O-α-rhamnopyranoside [3] (13.50 μmol/kg) or α-rhamnoisorobin [5] (23.13 μmol/kg) inhibited by 47.3%, 25.7%, 60.2% and 58.0%, respectively, the acetic acid-induced nociception (indomethacin: 27.95 μmol/kg, p.o.; 68.9%). Flavonoids 1, 2, 3 or 5, at the same doses, reduced by 39.5%, 46.5%, 35.6% and 33.3%, respectively, the croton oil-induced oedema (dexamethasone: 5.09 μmol/kg, s.c.; 83.7%) and impaired leukocyte migration by 42.9%, 46.3%, 50.4% and 49.6%, respectively (dexamethasone: 5.09 μmol/kg, s.c.; 66.1%).

Conclusions

Our findings show that the major kaempferol glycosides may account for the renowned medicinal use of Sedum dendroideum against pain and inflammatory troubles.     

Comparative acute toxicities and immunomodulatory potentials of five Eastern Nigeria mistletoes

Ethnopharmacological relevance

Traditionally, mistletoes of Eastern Nigeria origin, Loranthus micranthus Linn. have been used as immunostimulant for the management of certain diseases with high profile immune depleting potentials. This practice has remained till date without scientific validation.

Aim of study

To obtain and validate evidence for or against its continued use as immunostimulant and afford data for further studies on this specie of mistletoe. The present work is an in vivo proof of ethnopharmacological concept of the age long immunomodulatory use of our local mistletoe.

Materials and methods

Aqueous-methanol extracts of the plant leaves from five different host trees were evaluated for immunomodulatory activity using four in vivo models in mice or rats, namely; total and differential leukocyte count (TLC and DLC), the cellular mediated delayed-type hypersensitivity reaction (DTHR) test, the humoral mediated antibody titration (AT) test and the cyclophosphamide-induced myelosuppression (CIM) test at different dose levels (100, 200 and 400 or 50, 100 and 250 mg kg⁻¹; depending on model) against standard controls. Phytochemical and acute toxicity tests were equally carried out on all the extracts.

Results

Results obtained indicate that all the mistletoes contained the same phytochemical constituents, although in varying amounts. The mistletoes exhibited statistically significantly different (p < 0.05 or p < 0.001, ANOVA) immunomodulatory (up-regulatory) activities in the overall order of that from Kola acuminata > Citrus spp > Persia americana > Parkia biglobosa > Pentaclatra macrophylla. LD₅₀ values were generally greater than 5000 mg/kg.

Conclusion

The present study confirms the Eastern Nigeria mistletoe as a potent and safe alternative or complementary medicine for the management of immunodeficiency diseases.     

A fraction of stem bark extract of Entada africana suppresses lipopolysaccharide-induced inflammation in RAW 264.7 cells

Ethnopharmacological relevance

Entada africana is a plant used in African traditional medicine for the treatment of stomachache, fever, liver related diseases, wound healing, cataract and dysentery.

Aims of the study

This study aimed at evaluating the anti-inflammatory activity of fractions of the stem bark extract of the plant using lipopolysaccharide (LPS)-induced inflammation in RAW 264.7 macrophages model.

Materials and methods

The crude extract was prepared using the mixture CH₂Cl₂/MeOH (1:1, v/v) and fractionated by flash chromatography using solvents of increasing polarity to obtain five different fractions. The effects of the fractions on the cells viability were studied by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and their inhibitory activity against LPS-induced nitric oxide (NO) production screened by Griess test. The most active fraction was further investigated for its effects on reactive oxygen species (ROS) production using flux cytometry, the expression of inducible nitric oxide synthase (iNOS), pro-and anti-inflammatory cytokines (IL1β, TNFα, IL6, IL10 and IL13) by RT-PCR, and the activity of the enzyme p38 MAPK kinase by enzyme-linked immunosorbent assay (ELISA).

Results

The fractions presented no significant effect on the viability of macrophages at 100 μg/ml after 24 h incubation. The CH₂Cl₂/MeOH 5% (Ea5) fraction was found to be the most potent in inhibiting NO production with a half inhibition concentration (IC₅₀)=18.36 μg/ml, and showed the highest inhibition percentage (89.068%) in comparison with Baicalin (63.34%), an external standard at 50 μg/ml. Ea5, as well as Baicalin significantly (P<0.05) inhibited the expression of TNFα, IL6 and IL1β mRNA, attenuated mRNA expression of inducible NO synthase in a concentration-dependent manner, stimulated the expression of anti-inflammatory cytokines (IL10 and IL13), and showed a 30% inhibition of the activity of p38 MAPK kinase.

Conclusion

The results of the present study indicate that the fraction Ea5 of Entada africana possesses most potent in vitro anti-inflammatory activity and may contain compounds useful as a therapeutic agent in the treatment of inflammatory related diseases cause by over-activation of macrophages.     

Antisecretory activity from the flowers of Chiranthodendron pentadactylon and its flavonoids on intestinal fluid accumulation induced by Vibrio cholerae toxin in rats

Ethnopharmacological relevance

The flowers of Chiranthodendron pentadactylon Larreat. (Sterculiaceae) has been traditionally used as folk medicine in Mexico for the treatment of gastrointestinal disorders such as diarrhea and dysentery.

Aim of the study

This study aimed to assess the antisecretory activity which supports the therapeutic use of Chiranthodendron pentadactylon and its flavonoids to treat diarrhea.

Materials and methods

The methanol extract of Chiranthodendron pentadactylon, subsequent fractions, and flavonoids were evaluated on cholera toxin-induced intestinal secretion in rat jejunal loops model.

Results

Three antisecretory flavonoids were isolated by bioassay-guided purification, namely, isoquercitrin 3, (+)-catechin 4 and (−)-epicatechin 5. Among them, epicatechin exhibited the most potent antisecretory activity with ID₅₀ of 8.3 μM/kg. Its potency was close that of to loperamide (ID₅₀ 6.1 μM/kg), drug used as control. Isoquercitrin (ID₅₀ 19.2 μM/kg) and catechin (ID₅₀ 51.7 μM/kg) showed moderate and weak activity, respectively.

Conclusion

The results of the present study lend some support to the anecdotal report for the traditional use of the flowers of Chiranthodendron pentadactylon in the control of dysentery.     

Vasorelaxant properties of acid and neutral fractions of Dimerocostus strobilaceus Kuntze used by Kuna Indians of Panama

Ethnopharmacological relevance

Dimerocostus strobilaceus is used by the Kuna Indians of Panama for the treatment of hypertension and other cardiovascular diseases.

Aim of the study

We investigated the vascular effects of acid and neutral fractions obtained from methanol and dichloromethane extracts of Dimerocostus strobilaceus.

Materials and methods

The acid and neutral methanol fractions (A-MeOH and N-MeOH) or acid and neutral dichlorometanic fractions (A-DCM and N-DCM) were tested using isolated rat aortic rings with or without endothelium pre-contracted by phenylephrine. We examined the ability of these different fractions at different concentrations to modify vascular responses induced by angiotensin II using endothelium-denuded aortic rings from Spontaneously Hypertensive Rats (SHR).

Results

In aortic rings with intact endothelium A-MeOH, N-MeOH and A-DCM fractions produced a concentration-dependent vasorelaxation (62.4 ± 5.2, 64.5 ± 5.0 and 63.7 ± 5.0%, respectively), whereas the N-DCM fraction did not produce any vasorelaxant effect. Maximal relaxation evocated by vasoactive fractions was substantially inhibited on aortic rings without endothelium.Our study demonstrates that A-MeOH, N-MeOH, A-DCM and N-DCM significantly reduce contractile responses induced by angiotensin-II on aortic rings.

Conclusions

Our findings may contribute to a better understanding of the potential link between vascular properties observed with Dimerocostus strobilaceus and their ethnobotanical use.     

In vitro biological activities of niloticane,a new bioactive cassane diterpene from the bark of Acacia nilotica subsp. kraussiana

Ethnopharmacological relevance

Acacia nilotica subsp. kraussiana was reported in African traditional medicine for the treatment of various ailments. Isolation of an active compound in this study from the bark extract may lead to the validation of its efficiency as a traditional crude drug.

Aims of the study

This study aimed to isolate active compound(s) from an ethyl acetate bark extract of Acacia nilotica subsp. kraussiana and to investigate some of its biological activity.

Materials and methods

The isolation process was carried out using bioassay-guided fractionation. The isolated compound was tested for antibacterial activity using the micro-dilution assay; anti-inflammatory activity using the COX-1 and COX-2 assays and investigated for inhibitory effect against acetylcholinesterase using the microplate assay.

Results

A new bioactive compound was isolated and identified as a cassane diterpene, niloticane. Niloticane showed antibacterial activity against Gram-positive bacteria Bacillus subtilis and Staphylococcus aureus with MIC values of 4 and 8 μg/mL, respectively. With Gram-negative bacteria, niloticane showed weak activity. MIC values obtained were 16 and 33 μg/mL against Klebsiella pneumonia and Escherichia coli, respectively. In the cyclooxygenase test, niloticane possessed activity with IC₅₀ values of 28 and 210 μM against COX-1 and COX-2, respectively. IC₅₀ values observed with indomethacin (positive control) were 3.6 μM for COX-1 and 189 μM for COX-2. In the acetylcholinesterase test, niloticane showed anti-cholinesterase activity with an IC₅₀ value of 4 μM. IC₅₀ values obtained by the galanthamine (positive control) was 2.0 μM.

Conclusion

The results obtained support the traditional uses of the bark of Acacia nilotica subsp. kraussiana in African traditional medicine for the treatment of some ailments that relate to microbial diseases, inflammation and central nervous system disorders.     

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.     

The Canadian medicinal plant Heracleum maximum contains antimycobacterial diynes and furanocoumarins

Ethnopharmacological relevance

Heracleum maximum is amongst the most commonly used plants by the indigenous peoples of North America. The First Nations of the eastern Canada use infusions of Heracleum maximum roots for the treatment of respiratory ailments including tuberculosis. Previous investigations of extracts derived from the roots of Heracleum maximum have shown it to possess antimycobacterial activity. Aim of the study: To isolate and identify antimycobacterial constituents from the roots of Heracleum maximum.

Materials and methods

A methanolic extract of Heracleum maximum roots was subjected to bioassay guided fractionation using the microplate resazurin assay (MRA) to assess inhibitory activity against Mycobacterium tuberculosis strain H37Ra. The antimycobacterial constituents were identified by NMR, MS and polarimetry.

Results

The polyacetylene (3R,8S)-falcarindiol and the furanocoumarins bergapten, isobergapten, angelicin, sphondin, pimpinellin, isopimpinellin and 6-isopentenyloxyisobergapten were isolated from the Heracleum maximum root extract. (3R,8S)-Falcarindiol and 6-isopentenyloxyisobergapten exhibited MICs of 24 μM and 167 μM and IC₅₀s of 6 μM and 27 μM against Mycobacterium tuberculosis H37Ra respectively. The remaining furanocoumarins bergapten, isobergapten, angelicin, sphondin, pimpinellin, and isopimpinellin were less active, with MICs of 925, 1850, 2149, 1859, 812 and 1625 μM and IC₅₀s of 125, 344, 350, 351, 389 and 406 μM.

Conclusions

(3R,8S)-Falcarindiol, bergapten, isobergapten, angelicin, sphondin, pimpinellin, isopimpinellin and 6-isopentenyloxyisobergapten were identified as the principal constituents responsible for the antimycobacterial activity of the roots of Heracleum maximum. This work supports the ethnopharmacological use of Heracleum maximum by Canadian First Nations and Native American communities as a treatment for infectious diseases, specifically tuberculosis.     

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.     

Antiarrhythmic effects of dehydroevodiamine in isolated human myocardium and cardiomyocytes

Ethnopharmacological relevance

Dehydroevodiamine alkaloid (DeHE), a bioactive component of the Chinese herbal medicine Wu-Chu-Yu (Evodiae frutus), exerted antiarrhythmic effect in guinea-pig ventricular myocytes. We further characterize the electromechanical effects of DeHE in the human atrial and ventricular tissues obtained from hearts of patients undergoing corrective cardiac surgery or heart transplantation.

Materials and methods

The transmembrane potentials of human myocardia were recorded with a traditional microelectrode technique while sarcolemmal Na⁺ and Ca²⁺ currents in single human cardiomyocytes were measured by a whole-cell patch-clamp technique. The intracellular pH (pH_i) and Na⁺–H⁺ exchanger (NHE) activity were determined using BCECF-fluorescence in human atria.

Results

In human atria, DeHE (0.1–0.3 μM) depressed upstroke velocity, amplitude of action potential, and contractile force, both in slow and fast response action potential. Moreover, the similar depressant effects of DeHE were found in human ventricular myocardium. Both in isolated human atrial and ventricular myocytes, DeHE (0.1–1 μM) reversibly, concentration-dependently decreased the Na⁺ and Ca²⁺currents. Moreover, DeHE (0.1 and 0.3 μM) suppressed delayed afterdepolarizations and aftercontractions, induced by epinephrine and high [Ca²⁺]_o in atria. In human ventricular myocardium, the strophanthidin-induced triggered activities were attenuated by pretreating DeHE (0.3 μM). The resting pH_i and NHE activity were also significantly increased by DeHE (0.1–0.3 μM).

Conclusions

We concluded for the first time that, in the human hearts, DeHE could antagonize triggered arrhythmias induced by cardiotonic agents through a general reduction of the Na⁺ and Ca²⁺ inward currents, while increase of resting pH_i and NHE activity.