Vasorelaxant and hypotensive effects of the extract and the isolated flavonoid rutin obtained from Polygala paniculata L.

OBJECTIVES: This study aimed to investigate the in-vitro and in-vivo cardiovascular effects of the crude hydroalcoholic extract from Polygala paniculata (HEPP) in rats. METHODS: The procedures were performed on aortic rings and on normotensive anaesthetized rats. KEY FINDINGS: When tested in endothelium-intact aorta rings, HEPP (30-1000 µg/ml) produced a significant non-concentration-dependent relaxing effect (∼40%), which was completely prevented by incubation with L-NAME (nitric oxide synthase inhibitor), ODQ (soluble guanylate cyclase inhibitor) and partially inhibited by tetraethylammonium (TEA; a non-selective potassium channel blocker) and charybdotoxin (a large- and intermediate-conductance calcium-activated potassium channel blocker). In contrast, atropine (a muscarinic receptor antagonist) or pyrilamine(a histamine H1 receptor antagonist) had no effect. Furthermore, oral administration of HEPP (30-300 mg/kg) in anaesthetized rats caused a dose-dependent and sustained hypotensive action. This effect was unchanged by atropine or TEA, but was strongly reduced in rats continuously infused with L-NAME or methylene blue. Moreover, rutin (1-3 mg/kg) administered by an intravenous route also caused a dose-dependent hypotensive effect in rats. CONCLUSIONS: Our results demonstrated that the extract obtained from P. paniculata induces potent hypotensive and vasorelaxant effects that are dependent on the nitric oxide/guanylate cyclase pathway. These effects could be related, at least in part, to the rutin contents in this extract.

Myricitrin as a substrate and inhibitor of myeloperoxidase: implications for the pharmacological effects of flavonoids.

Flavonoids are increasingly being ingested by the general population as chemotherapeutic and anti-inflammatory agents. They are potentially toxic because of their conversion to free radicals and reactive quinones by peroxidases. Little detailed information is available on how flavonoids interact with myeloperoxidase, which is the predominant peroxidase present at sites of inflammation. This enzyme uses hydrogen peroxide to oxidize chloride to hypochlorous acid, as well as to produce an array of reactive free radicals from organic substrates. We investigated how the flavonoid myricitrin is oxidized by myeloperoxidase and how it affects the activities of this enzyme. Myricitrin was readily oxidized by myeloperoxidase in the presence of hydrogen peroxide. Its main oxidation product was a dimer that underwent further oxidation. In the presence of glutathione, myricitrin was oxidized to a hydroquinone that was conjugated to glutathione. When myeloperoxidase oxidized myricitrin and related flavonoids it became irreversibly inactivated. The number of hydroxyl groups in the B ring of the flavonoids and the presence of a free hydroxyl m-phenol group in the A ring were important for the inhibitory effects. Less enzyme inactivation occurred in the presence of chloride. Neutrophils also oxidized myricitrin to dimers in a reaction that was partially dependent on myeloperoxidase. Myricitrin did not affect the production of hypochlorous acid by neutrophils. We conclude that myricitrin will be oxidized by neutrophils at sites of inflammation to produce reactive free radicals and quinones. It is unlikely to affect hypochlorous acid production by neutrophils.

Gastroprotective activity of the hydroalcoholic extract obtained from Polygala paniculata L. in rats.

The possible gastroprotective effects of the hydroalcoholic extract of Polygala paniculata in rats have been evaluated. We have investigated the effects of this hydroalcoholic extract on acute lesions induced by ethanol (70%, p.o.) and indomethacin (20 mg kg(-1), s.c.). Its influence on mucus secretion was investigated, measured as the amount of Alcian blue dye estimated by colorimetry, and antisecretory effects were assessed in the pylorus ligature model. The treatment of rats with a crude hydroalcoholic extract of P. paniculata (HEPP; 30, 100, 300 mg kg(-1), p.o., or 3, 10 and 30 mg kg(-1), i.p.) decreased the ulcer index, and maintained the gastric mucus production in acute gastric lesions caused by ethanol 70%. In addition, the extract partially protected the mucosa against indomethacin-induced lesions. The extract did not change the volume and acidity of gastric secretion in the pylorus-ligated rat. An additional antioxidant activity of the extract and its isolated flavonoid compound rutin, in the DPPH free radical scavenging assay, was observed. In conclusion, HEPP exhibited marked gastroprotection; these effects may have involved prostaglandins and be related to cytoprotective factors, such as antioxidant activity and maintenance of mucus production.

Involvement of p38MAPK on the antinociceptive action of myricitrin in mice.

Previous studies from our group investigated the analgesic and anti-inflammatory properties of the flavonoid myricitrin. Here, we demonstrated the role of interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha) and mitogen-activated protein kinases (MAPKs) on the antinociceptive action of myricitrin. The nociceptive response was evaluated by monitoring biting behaviour following intratecal (i.t.) administration of IL-1beta and TNF-alpha in mice. Western blot analyses of total and phosphorylated MAPKs: p38(MAPK), extracellular-signal regulated kinase (ERK1/2) and c-Jun amino-terminal kinases (JNK1/2) from the spinal cord of mice injected with cytokines were measured. Myricitrin (0.03-30mg/kg) or vehicle (control) was administered 30 min beforehand by intraperitoneal (i.p.) injection. Myricitrin pre-treatment prevented cytokine-induced biting behaviour. The calculated ID(50) of myricitrin were 6.8 (4.6-9.0) and 2.6 (0.3-4.9) mg/kg and maximal inhibition of 83+/-9 and 100+/-0% for IL-1beta and TNF-alpha, respectively. Intrathecal injection of IL-1beta and TNF-alpha significantly increased p38(MAPK) phosphorylation and this was inhibited by myricitrin treatment. Cytokines administration did not alter ERK1/2 and JNK1/2 phosphorylation. Myricitrin prevented cytokine-induced biting behaviour and inhibited p38(MAPK) phosphorylation in response to cytokines stimulation. Taken together, it suggests that the mechanism for antinociceptive action of myricitrin in response to cytokines may involve a blockage on p38(MAPK) pathway. This finding could explain, at least in part, the antinociceptive action of this flavonoid in process like neuropathic and inflammatory chronic pain.

Antinociceptive action of myricitrin: involvement of the K+ and Ca2+ channels.

The present study was designed to investigate the mechanisms involved in the antinociception afforded by myricitrin in chemical models of nociception in mice. Myricitrin given by intrathecal (i.t.) or intracerebroventricular (i.c.v.) route produced dose-related antinociception when evaluated against acetic acid-induced visceral pain in mice. In addition, the intraperitoneal administration of myricitrin caused significant inhibition of biting behaviour induced by i.t. injection of glutamate, substance P, capsaicin, interleukin 1 beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha). The antinociception caused by myricitrin in the acetic acid test was fully prevented by i.t. pre-treatment with pertussis toxin, a Gi/o protein inactivator, and by i.c.v. injection of calcium chloride (CaCl(2)). In addition, the i.t. pre-treatment of mice with apamin, a blocker of small (or low)-conductance calcium-gated K(+) channels and tetraethylammonium, a blocker of voltage-gated K(+) channels significantly reversed the antinociception induced by myricitrin. The charybdotoxin, a blocker of large (or fast)-conductance calcium-gated K(+) channels and glibenclamide, a blocker of the ATP-gated K(+) channels had no effect on myricitrin-induced antinociception. Calcium uptake analysis revealed that myricitrin inhibited (45)Ca(2+) influx under a K(+)-induced depolarization condition. However, calcium movement was modified in a non-depolarizing condition only when the highest concentration of myricitrin was used. In summary, our findings indicate that myricitrin produces consistent antinociception in chemical models of nociception in mice. These results clearly demonstrate an involvement of the Gi/o protein dependent mechanism on antinociception caused by myricitrin. The opening of voltage- and small-conductance calcium-gated K(+) channels and the reduction of calcium influx led to the antinociceptive of myricitrin.

Anti-allodynic property of flavonoid myricitrin in models of persistent inflammatory and neuropathic pain in mice.

The aim of the present study was to investigate the effects of myricitrin, a flavonoid with anti-inflammatory and antinociceptive action, upon persistent neuropathic and inflammatory pain. The neuropathic pain was caused by a partial ligation (2/3) of the sciatic nerve and the inflammatory pain was induced by an intraplantar (i.pl.) injection of 20 microL of complete Freund's adjuvant (CFA) in adult Swiss mice (25-35 g). Seven days after sciatic nerve constriction and 24 h after CFA i.pl. injection, mouse pain threshold was evaluated through tactile allodynia, using Von Frey Hair (VFH) filaments. Further analyses performed in CFA-injected mice were paw edema measurement, leukocytes infiltration, morphological changes and myeloperoxidase (MPO) enzyme activity. The intraperitoneal (i.p.) treatment with myricitrin (30 mg/kg) significantly decreased the paw withdrawal response in persistent neuropathic and inflammatory pain and decreased mouse paw edema. CFA injection increased 4-fold MPO activity and 27-fold the number of neutrophils in the mouse paw after 24 h. Myricitrin strongly reduced MPO activity, returning to basal levels; however, it did not reduce neutrophils migration. In addition, myricitrin treatment decreased morphological alterations to the epidermis and dermis papilar of mouse paw. Together these results indicate that myricitrin produces pronounced anti-allodynic and anti-edematogenic effects in two models of chronic pain in mice. Considering that few drugs are currently available for the treatment of chronic pain, the present results indicate that myricitrin might be potentially interesting in the development of new clinically relevant drugs for the management of this disorder.

Protective effects of Polygala paniculata extract against methylmercury-induced neurotoxicity in mice.

We have examined the possible protective effects of Polygala paniculata extract against methylmercury (MeHg)-induced neurotoxicity in adult mice. MeHg was diluted in drinking water (40 mg L(-1), freely available) and the hydroalcoholic Polygala extract was diluted in a 150 mM NaCl solution and administered by gavage (100 mg kg(-1) b.w., twice a day). After a two-week treatment, MeHg exposure significantly inhibited glutathione peroxidase and increased glutathione reductase activity, while the levels of thiobarbituric acid reactive substances were increased in the cerebral cortex and cerebellum. These alterations were prevented by administration of Polygala extract, except for glutathione reductase activity, which remained elevated in the cerebral cortex. Behavioural interference in the MeHg-exposed animals was evident through a marked deficit in the motor performance in the rotarod task, which was completely recovered to control levels by Polygala extract co-administration. This study has shown, for the first time, the in-vivo protective effects of Polygala extract against MeHg-induced neurotoxicity. In addition, our findings encourage studies concerning the beneficial effects of P. paniculata on neurological conditions related to excitotoxicity and oxidative stress.