Brasiliensic acid: in vitro cytotoxic and genotoxic, in vivo acute toxicity and in silico pharmacological prediction of a new promising molecule.

Brasiliensic acid (Bras) is a chromanone isolated from Calophyllum brasiliense Cambèss. bark extracts with confirmed potential activity on gastric ulcer and Helicobacter pylori infection. This study aimed to investigate the in vitro and in vivo toxicity of Bras and molecular docking studies on its interactions with the H. pylori virulence factors and selected gastric cancer-related proteins. Cytotoxicity was evaluated by alamarBlue© assay, genotoxicity by micronucleus and comet assays, and on cell cycle by flow cytometry, using Chinese hamster epithelial ovary cells. Bras was not cytotoxic to CHO-K1 cells, and caused no chromosomal aberrations, nor altered DNA integrity. Furthermore, Bras inhibited damages to DNA by H2O2 at 1.16 µM. No cell cycle arrest was observed, but apoptosis accounted for 31.2% of the cell death observed in the CHO-K1 at 24 h incubation of the IC50. Oral acute toxicity by Hippocratic screening test in mice showed no relevant behavioral change/mortality seen up to 1,000 mg/kg. The molecular docking approach indicated potential interactions between Bras and the various targets for peptic ulcer and gastric cancer, notably CagA virulence factor of H. pylori and VEGFR-2. In conclusion, Bras is apparently safe and an optimization for Bras can be considered for gastric ulcer and cancer.Communicated by Ramaswamy H. Sarma.

Evaluation of the gastroprotective and ulcer healing properties by Fridericia chica (Bonpl.) L.G. Lohmann hydroethanolic extract of leaves.

ETHNOPHARMACOLOGICAL RELEVANCE: Fridericia chica (Bonpl.) L.G. Lohmann (Bignoniaceae), is a climber native to Brazil, found in all Brazilian biomes. It is mostly known in Brazil as "carajiru," and home medicines made from the leaves have been used to cure disorders including stomach ulcers and other gastrointestinal disorders. AIM OF THE STUDY: The objective of the study was to investigate the F. chica hydroethanolic extract of leaves (HEFc) preventative and curative antiulcer gastrointestinal efficacy as well as the mechanisms of action using in vivo rodent models. MATERIALS AND METHODS: F. chica was collected in the municipality of Juína, Mato Grosso, and its leaves were used to prepare the extract by maceration technique (70% hydroethanol in the 1:10 ratio, w/v) to obtain the HEFc. The chromatographic analysis of HEFc was carried out by High Performance Liquid Chromatography-Photo Diode Array-Electrospray Ionization-Mass Spectrometry (HPLC-PDA-ESI-MS)- LCQ Fleet™ system. To determine the potential antiulcer potential of HEFc (1, 5 and 20 mg/kg, p.o.), the gastroprotective activity was assessed in various animal models of stomach ulcers caused by acidified ethanol, water constraint stress, indomethacin, (acute), and acid acetic (chronic). Additionally, the prokinetic properties of the HEFC were assessed in mice. The gastroprotective underlying mechanisms were evaluated by the histopathological analysis and determination of gastric secretion (volume, free and total acidity), gastric barrier mucus, activation of PGs, NO, K +ATP channels, α2-adrenoceptor, antioxidant activity (GSH, MPO and MDA), NO and mucosal cytokines (TNF-α, IL-1ß, and IL-10) levels. RESULTS: The chemical composition of HEFc was analyzed and apigenin, scutellarin, and carajurone were identified. HEFc (1, 5 and 20 mg/kg) showed effect against acute ulcers induced by HCl/EtOH with a reduction in the ulcerated area of 64.41% (p < 0.001), 54.23% (p < 0.01), 38.71% (p < 0.01), respectively. In the indomethacin experiment, there was no change in the doses tested, whereas in the water immersion restraint stress ulcer there was a reduction of lesions at doses of 1, 5, and 20 mg/kg by 80.34% (p < 0.001), 68.46% (p < 0.01) and 52.04% (p < 0.01). HEFc increased the mucus production at doses of 1 and 20 mg/kg in 28.14% (p < 0.05) and 38.36% (p < 0.01), respectively. In the pyloric ligation-induced model of gastric ulceration, the HEFc decreased the total acidity in all doses by 54.23%, 65.08%, and 44.40% (p < 0.05) and gastric secretory volume in 38.47% at dose of 1 mg/kg (p < 0,05) and increased the free acidity at the dose of 5 mg/kg by 11.86% (p < 0.05). The administration of EHFc (1 mg/kg) showed a gastroprotective effect possibly by stimulating the release of prostaglandins and activating K+ATP channels and α2-adrenoreceptors. Also, the gastroprotective effect of HEFc involved an increase in CAT and GSH activities, and a reduction in MPO activity and MDA levels. In the chronic gastric ulcer model, the HEFc (1, 5 and 20 mg/kg) decreased the ulcerated area significantly (p < 0.001) at all doses by 71.37%, 91.00%, and 93.46%, respectively. In the histological analysis, HEFc promoted the healing of gastric lesions by stimulating the formation of granulation tissue and consequently epithelialization. On the other hand, regarding the effect of HEFc on gastric emptying and intestinal transit, it was observed that the extract did not alter gastric emptying, but there was an increase in intestinal transit at the dose of 1 mg/kg (p < 0.01). CONCLUSION: These outcomes confirmed the advantages of Fridericia chica leaves for the treatment of stomach ulcers, which are well-known. HEFc was discovered to have antiulcer characteristics through multitarget pathways, which might be related to an increase in stomach defense mechanisms and a decrease in defensive factor. HEFc can be regarded as a potential new antiulcer herbal remedy because of its antiulcer properties, which may be attributed to the mixture of flavonoids, apigenin, scutellarin and carajurone.

Evidence for the Involvement of Cytokines Modulation and Prokinetic Properties in Gastric Ulcer Healing Effects of Helicteres sacarolha A. St.-Hil. A. Juss.

Preparations of Helicteres sacarolha (Malvaceae) leaves and roots are used in the form of decoction, infusion or maceration, to treat gastrointestinal disturbances, among others. Studies supporting some of its ethnomedicinal uses are still incipient. The present study aimed to investigate it potential effect on chronic ulcer, ulcerative colitis and possible prokinetic activities as part of its mechanism of action. The powdered leaves of Helicteres sacarolha (HEHs) was prepared by maceration in 70 % hydroethanolic solution. Its qualitative phytochemical constituents were investigated by direct flow injection analysis coupled to atmospheric pressure chemical ionization ion trap tandem mass spectrometry (FIA-APCI-IT-MSn ). The gastric ulcer healing effect was evaluated in acetic acid induced chronic ulcer in mice and the lesions were evaluated, including analysis of blood plasma cytokine levels. The prokinetic properties (gastric emptying and intestinal transit) were carried out in mice. Potential anti-ulcerative colitis activity was evaluated in rats using 2,4,6-trinitrobenzenesulfonic acid (5 % TNBS) -induced colitis. All animal experiments were carried out at the doses of 20, 50 and 250 mg/kg (p.o.). Eight compounds were putatively identified, specifically lariciresinol, and its derivatives, kaempferol derivatives and Tricin-O-Glc. The extract promoted increased gastric ulcer healing at all doses tested. Modulation of the cytokines involved inhibition of some key pro-inflammatory cytokines with maximum effect on IL-1ß (70 %, 50 mg/kg, p<0.05), TNF-α (79 %, 20 mg/kg, p<0.01), and in the anti-inflammatory cytokines, namely IL-10 (57 %, 50 mg/kg, p<0.05) and IL-17 (79 %, only at 50 mg/kg, p<0.05). Histological findings demonstrated a mitigated inflammatory activity, and tissues undergoing regeneration. HEHs treatment caused delayed gastric emptying, and increased intestinal transit, but had no effect in the experimentally induced ulcerative colitis. We report for the first time putatively the presence of Lariciresinol and tricin derivatives from the hydroethanolic leaves extract of H. sacarolha. Its possible mechanism of actions of gastric ulcer healing involves cytokines modulation, mitigation of inflammatory response and tissue regeneration and provoked opposing effect in the gastrointestinal system. The present study demonstrates the therapeutic potential of H. sacarolha leaves used in Brazilian ethnomedicine in the treatment of chronic gastric ulcer.

Chemical Characterization, Antioxidant and Cytotoxic Activities of the Edible Fruits of Brosimun gaudichaudii Trécul, a Native Plant of the Cerrado Biome.

In Brazil, there is a large diversity of species of small edible fruits that are considered sources of nutrients and functional properties. They present a high innovation domain for the pharmaceutical, cosmetic and food industries due to their health-promoting properties. Edible fruits from Brosimum gaudichaudii (Moraceae) are widely consumed and used in folk medicine and in feed by the population of the Brazilian Cerrado. Nevertheless, detailed information on the chemical fingerprint, antiradical activity and safety aspects of these fruits is still unknown. Thus, the aim of this work was to investigate the bioactive compounds of hydroethanolic extracts of fruits from Brosimum gaudichaudii using high-performance liquid chromatography combined with mass spectrometry using electrospray ionization (HPLC ESI-MS). Eighteen different compounds, including flavonoids, coumarins, arylbenzofurans, terpenoids, stilbenes, xanthones and esters, were detected. Moreover, the study indicated that the hydroethanolic extract of fruits from B. gaudichaudii presented low scavenging activity against 2,2-diphenyl-1-picrylhydrazyl radicals (IC50 >800 µg mL-1 ) and was cytotoxic (IC50 <30 µg mL-1 ) in Chinese hamster ovary cells (CHO-K1) by an in vitro assay. This is the first report of the chemical profile, antioxidant activity and cytotoxic properties of the hydroethanolic extract of fruits from B. gaudichaudii.

Canthin-6-one ameliorates TNBS-induced colitis in rats by modulating inflammation and oxidative stress. An in vivo and in silico approach.

Canthin-6-one (Cant) is an indole alkaloid found in several botanical drugs used as medicines, reported to be gastroprotective, anti-inflammatory, anti-microbial, anti-diarrheal and anti-proliferative. We aimed to explore Cant in the management of colitis using a trinitrobenzenesulfonic acid (TNBS)-induced rat model. Cant (1, 5 and 25 mg/kg) was administered by oral gavage to Wistar rats followed by induction of colitis with TNBS. Macroscopic and histopathological scores, myeloperoxidase (MPO), malondialdehyde (MDA) and reduced glutathione (GSH) were assessed in colon tissues. Pro- (TNF-α, IL-1ß and IL-12p70) and anti-inflammatory (IL-10) cytokines, and vascular endothelial growth factor (VEGF) were also quantified. Mitogen-activated protein kinase 14 (MAPK14) and Toll-like receptor-8 (TLR8), as putative targets, were considered through in silico analysis. Cant (5 and 25 mg/kg) reduced macroscopic and histological colon damage scores in TNBS-treated rats. MPO and MDA were reduced by up to 61.69% and 92.45%, respectively, compared to TNBS-treated rats alone. Glutathione concentration was reduced in rats administered with TNBS alone (50.00% of sham group) but restored to 72.73% (of sham group) with Cant treatment. TNF-α, IL-1ß, IL-12p70 and VEGF were reduced, and anti-inflammatory IL-10 was increased following Cant administration compared to rats administered TNBS alone. Docking ligation results for MAPK14 (p38α) and TLR8 with Cant, confirmed that these proteins are feasible putative targets. Cant has an anti-inflammatory effect in the intestine by down-regulating molecular immune mediators and decreasing oxidative stress. Therefore, Cant could have therapeutic potential for the treatment of inflammatory bowel disease and related syndromes.

Copaifera malmei Harms leaves infusion attenuates TNBS-ulcerative colitis through modulation of cytokines, oxidative stress and mucus in experimental rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Ethnobotanical studies show that the infusion of the leaves from Copaifera malmei Harms (Fabaceae) has been utilized in the Brazilian traditional medicine to treat provocative and gastrointestinal diseases, among others. Recently, our research team has shown that an infusion extract of the leaves of C. malmei has a strong antiulcer activity and its oral use gives no indications of toxicity. AIM OF THE STUDY: The aim of the study is to evaluate the anti-inflammatory intestinal effect of an infusion extract from the leaves of Copaifera malmei (IECm) in an animal model of ulcerative colitis induced by 2,4,6-trinitrobenzenesulfonic acid (TNBS). MATERIALS AND METHODS: Acute intestinal inflammation was induced in male Wistar rats by TNBS in 20% EtOH (0.25 mL). IECm was administered by oral gavage (for 72, 48, 24 and 2 h) preceding the induction of ulcerative colitis. The colon damage and degree of inflammation were evaluated by morphological observation scores and colon weight. The improved colonic mucosal injury, oxidative stress and inflammatory response were assessed by histopathological investigation and by estimating myeloperoxidase (MPO) activity, malondialdehyde (MDA) and glutathione (GSH) levels and tumor necrosis factor (TNF), interleukin 1ß (IL1-ß), IL-17 and IL-10 colon tissue concentrations. The histopathological changes were done on the colon tissues by hematoxylin and eosin and Periodic Acid-Schiff staining were utilized to measure the mucus. RESULTS: Pre-treatment (25, 100 and 400 mg/kg) with IECm altogether diminished the intestinal inflammation prompted by TNBS in rats by diminishing colonic score by 69.12% (p < 0.01), 19.87% (p < 0.05) and 67.60% (p < 0.01), individually. Improvement of colonic mucosal injury by treatment with IECm was shown by a decline in MPO activity at dosages 25 and 400 mg/kg by 67.98% and 59.68% (p < 0.001), MDA levels 64.80% and 80.00% (p < 0.01) and an expansion in GSH content at all portions (62.53%, 53.38% and 81.20% p < 0.05) compared with vehicle control group. IECm additionally prevention of intestinal inflammation as confirm by decreased cytokine levels, for example, TNF (31.26%, p < 0.05, 50.68% and 45.95%, p < 0.01), IL1-ß (56.41%, 58.83% and 56.65%, p < 0.001), IL-17 (51.66%, p < 0.001, 22.23%, p < 0.05 and 49.67%, p < 0.001) and increased the IL-10 levels at 25 and 400 mg/kg (57.13%, p < 0.01 and 35.83%, p < 0.05) respectively. Histopathological examination of the colon tissue displayed recovery of ulcerative colitis of IECm treated animals by reducing leukocyte infiltrate, epithelial, submucosal and muscular layer damages and maintaining mucus production. CONCLUSION: These findings revealed that IECm was effective and possess anti-colitic activities in a rodent model of UC and can be useful in inflammatory bowel diseases (IBD). The pre-treatment with IECm decreased intestinal inflammation by reducing macroscopical and microscopical colon injury. In addition, the present study demonstrated that IECm ameliorates TNBS-colitis by promoting antioxidant effect, modulation of cytokines release and restauration of mucus production. The study reinforces the traditional use of the Copaifera malmei leaves infusion to inflammatory gastrointestinal disorders and makes IECm a potential herbal medicine for the treatment of IBD.

Dilodendron bipinnatum Radlk. extract alleviates ulcerative colitis induced by TNBS in rats by reducing inflammatory cell infiltration, TNF-α and IL-1ß concentrations, IL-17 and COX-2 expressions, supporting mucus production and promotes an antioxidant effect.

ETHNOPHARMACOLOGICAL RELEVANCE: Dilodendron bipinnatum (Sapindaceae) stem bark decoction and macerate were used to treat uterine inflammation, pain in general, dermatitis and bone fractures. These homemade preparations also have diuretic, stimulant, expectorants and sedative effects and are effective in treating worm infections in the Brazilian Pantanal population. Our previous research confirmed the anti-inflammatory activity of the hydroethanolic extract of inner stem bark of D. bipinnatum (HEDb). AIM: This work aimed to investigate the efficacy of HEDb in ameliorating experimental colitis in rats and to elucidate the possible mechanisms involved in the anti-ulcerative colitis properties of HEDb in rats and Caco-2 cell line. MATERIALS AND METHODS: The effects on cell viability, IL-8 and TNF-α in human colon adenocarcinoma (Caco-2) were determined by flow cytometer and ELISA. Wistar rats (n = 6-7) were orally gavaged with, vehicle (0.9% saline), HEDb at doses of 20, 100 or 500 mg/kg, or mesalazine at a dose of 500 mg/kg, at 48, 24 and 1 h prior to the administration of trinitrobenzene sulfonic acid via rectal administration to induce colitis. The anti-inflammatory effects of HEDb were assessed macroscopically, by myeloperoxidase (MPO) activity and for glutathione (GSH) concentration in the colon. Additionally, colonic histopathological analyses of UC severity were conducted by different staining methods (H&E, PAS and toluidine blue). Pro-inflammatory cytokines TNF-α and IL-1ß were quantified in colonic tissue by ELISA and colonic expressions of COX-2 and IL-17 were analyzed by western blotting. RESULTS: HEDb was shown to be non-cytotoxic with mean viability of 80% in Caco-2 cells. HEDb pre-treatments of 1, 5 or 20 µg/mL significantly reduced TNF-α production in Caco-2 cells by 21.8% (p < 0.05), 60.5 and 82.1% (p < 0.001) respectively following LPS treatment compared to LPS alone. However, no change in IL-8 production was observed. HEDb pre-treatment of rats subjected to TNBS significantly (p < 0.001) reduced colonic lesion score. Higher doses (100 and 500 mg/kg) caused a sharp downregulation of haemorrhagic damage, leukocyte infiltration, edema and restoration of mucus production. Moreover, mast cell degranulation was inhibited. Colonic MPO activity was reduced following all doses of HEDb, reaching 51.1% ± 1.51 (p < 0.05) with the highest dose. GSH concentration was restored by 58% and 70% following 100 and 500 mg/kg of HEDb, respectively. The oral treatment of HEDb at doses 20, 100 and 500 mg/kg decreased the concentrations of TNF-α and IL-1ß at all doses in comparison to vehicle treated control. In addition, HEDb inhibited the COX-2 and IL-17 expressions with maximal effect at 500 mg/kg (60.3% and 65% respectively; p < 0.001). In all trials, the effect of HEDb at all doses being 20, 100 and 500 mg/kg was statistically comparable to mesalazine (500 mg/kg). CONCLUSIONS: HEDb reduces colonic damage in the TNBS colitis model and relieves oxidative and inflammatory events, at least in part, by increasing mucus production, reducing leukocyte migration and reducing TNF-α (in vivo and in vitro), IL-1ß, IL-17 and COX-2 expression. Therefore, HEDb requires further investigation as a candidate for treating IBD.

Piper umbellatum L. (Piperaceae): Phytochemical profiles of the hydroethanolic leaf extract and intestinal anti-inflammatory mechanisms on 2,4,6 trinitrobenzene sulfonic acid induced ulcerative colitis in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Piper umbellatum L. leaves, commonly found in the Amazon, Cerrado and Atlantic rain forest regions of Brazil, are widely used as a traditional medicine to treat gastrointestinal disorders and inflammation, among others diseases. Also, previous scientific studies demonstrated that P. umbellatum has gastroprotective and anti-inflammatory activity. AIM: To investigate the phytochemical profiles and the intestinal anti-inflammatory effect of the hydroethanolic extract of P. umbellatum (HEPu) leaf on ulcerative colitis in rats. MATERIALS AND METHODS: In this study, the chemical composition of HEPu was analyzed by gas chromatography-mass spectrometry (GC-MS) and liquid chromatography coupled to mass spectrometry (LC/MS). Also, this work studied the effects of HEPu on ulcerative colitis induced by 2,4,6-trinitrobenzene sulfonic acid (TNBS, 30 mg/mL in 20% ethanol) by intrarectal administration in rats. Simultaneously, animals were pre-treated orally with HEPu (30, 100 and 300 mg/kg), mesalazine (500 mg/kg), or vehicle. At the end of the experimental period, clinical signs of ulcerative colitis were evaluated by determination of weight loss, gross appearance, ulcer area and histological changes. Reduced glutathione (GSH), lipoperoxides (MDA) and nitric oxide (NO) levels, and superoxide dismutase (SOD), myeloperoxidase (MPO) and catalase (CAT) activities were determined in colon tissues. Also, pro-inflammatory mediators such as tumor necrosis factor alpha (TNF-α) and interleukin 1 beta (IL- 1ß) were quantified by immunoassay on the surface of fluorescent-coded magnetic beads (Luminex MagPix System). RESULTS: GC-MS analysis showed the presence of 17 different phytochemical compounds in the HEPu. LC/MS analyses revealed the presence of compounds in HEPu as protocatechuic acid, ferulic acid, kaempferol, rosmarinic acid, apigenin and ursolic acid. Treatment with HEPu significantly ameliorated weight loss, macroscopic damage, ulcerated area and histopathological changes such as sub-mucosal edema, cell infiltration, ulceration and necrosis (p < 0.001). Furthermore, HEPu (30, 100, and 300 mg/kg, p.o) inhibited the levels of oxidative parameters, such as MPO (49%, 53%, and 62%, p < 0.001), NO (20%, 19%, 22%, p < 0.01), and MDA (75%, 83%, 70%, p < 0.001), whereas increased the antioxidant activities such as SOD (208%, 192%, 64%, p < 0.001), GSH (94%, 75%, 49%, p < 0.01), and CAT (92%, 69%, 108%, p < 0.01). The extract also inhibited the pro-inflammatory cytokines TNF-α (81%, 85%, 85%, p < 0.001) and IL-1ß (95%, 79%, 89%, p < 0.001) levels. CONCLUSION: Together, these results revealed that P. umbellatum L. is a promising source of metabolites to be used in the treatment of inflammatory bowel disease.

Sorocea guilleminiana Gaudich.: Wound healing activity, action mechanisms, and chemical characterization of the leaf infusion.

ETHNOPHARMACOLOGICAL RELEVANCE: Sorocea guilleminina Gaudich. is a tree or shrub endemic to Brazil. Its leaves are used in Brazilian folk medicine for the healing of wounds, stomach problems, inflammation and as diuretic. The present study evaluates the activity and action mechanisms of the healing properties of the aqueous extract of S. guilleminiana leaves (AESg), in experimental models in vivo and in vitro, as well as performs a phytochemical analysis of the extract. MATERIALS AND METHODS: The AESg was prepared by infusion: Ten g of dry leaves powder in 1 L hot water, soaked for 15 min, filtered, lyophilized, and stored at -30 °C. Phytochemical analyses were realized by colorimetry and HPLC/ESI/MS. Its' in vitro cytotoxicity was evaluated on fibroblastic N3T3 cells. The potential of the wound healing activity in vivo was evaluated using excision and incision wound rat models, by histopathology of the injured skin along with the determination of nitric oxide, cytokines (IL-1ß, IL-10, and TNF-α), and antioxidant parameters (GSH, MPO and CAT). In vitro wound healing activity was also demonstrated in scratched N3T3 cells, by measuring the proliferation/migration rate. RESULTS: The phytochemical analysis of the AESg revealed a strong presence of polar compounds, especially flavonoids (4 majoritarian), as well as terpenes and/or sterols (2 majoritarian). The AESg showed no toxicity in the N3T3 cell line (IC50 > 800 µg/mL). Topical treatment with the AESg showed an increase (p < 0.05) in wound contraction with 2 mg/g cream on days 5 and 9 (43.56% and 6.70% increase, respectively), and with 50 mg/g on days 7 and 9 (10.88% and 7.91%, respectively), compared to the vehicle (non-ionic neutral cream). Topical application of AESg (2 or 50 mg/g non-ionic cream) in incised wounds caused an increase in the force necessary for the rupture of the wound when compared to the vehicle group. No changes in cytokines (IL-1ß, IL-10, or TNF-α) or NO accumulation was found with up to 50 mg/g AESg treatment. For antioxidant activity on the incision wound, an increase in GSH levels was denoted with the AESg use, at the lowest and highest dose (2 and 50 mg/g) by 75.86% and 61.20% respectively, when compared to the vehicle. Also, the CAT activity was accentuated by AESg at the highest dose (50 mg/g) by 85.87%. Finally, the AESg at all doses attenuated MPO activity significantly in the incision wound by 71.35%, 73.21%, 78.08%, respectively. In the scratch test on N3T3 cells, the treatment with AESg resulted also in an increase in fibroblast proliferation/migration rate, compared to the vehicle. CONCLUSION: AESg is not cytotoxic. The results confirm the popular use of the leaf infusion of S. guilleminiana for the treatment of cutaneous wounds, possibly by stimulating the proliferation of fibroblasts with a consequent deposition of collagen, fastening rearrangement of collagen fibers, and greater transformation into myofibroblasts, essential in the healing process. Preliminary chemical analyzes of AESg revealed the presence mainly of phenolic compounds, being salicylic acid, gallic acid, pinocembrin and isoquercitrin the majoritarian ones.

Cochlospermum regium (Mart. ex Schrank) Pilg.: Evaluation of chemical profile, gastroprotective activity and mechanism of action of hydroethanolic extract of its xylopodium in acute and chronic experimental models.

ETHNOPHARMACOLOGICAL RELEVANCE: Cochlospermum regium (Bixaceae) is a native shrub of Brazil and its xylopodium (infusion/decoction) is being used for the treatment of gastritis, ulcers, arthritis, intestinal infections, gynaecological infections, skin diseases, among others. The aim of the present study was to evaluate the gastroprotective/antiulcer activity and the mechanism of action of hydroethanolic extract of C. regium xylopodium (HECr), using in vitro and in vivo models. Additionally, phytochemical constituents were identified by high-performance liquid chromatography (HPLC). MATERIALS AND METHODS: C. regium xylopodium was macerated with ethanol/water to obtain the HECr. The phytochemical characterisation was carried out by HPLC. The antiulcer efficacy of HECr (25, 100 and 400 mg/kg, p.o.) was evaluated using acute acidified ethanol (HCl/EtOH), piroxicam and water immersion-induced experimental ulcer models. Chronic gastric ulcer healing activity of HECr was evaluated through acetic acid (99.8%) - induced model. Histological analysis and myeloperoxidase (MPO), glutathione (GSH), catalase (CAT) activities were also evaluated in chronic ulcer induced gastric tissues. The plausible mode of action of the HECr was assessed by estimation of gastric wall mucus production and the role of gastric secretion in pylorus ligature. The animals were also pre-treated with various inhibitors which includes indomethacin (10 mg/kg, p.o.) a selective inhibitor of cyclooxygenase, L-NAME (10 mg/kg, i.p.), an inhibitor of nitric oxide synthase, glibenclamide, a ATP-sensitive potassium channels (K+ATP) blocker (5 mg/kg, p.o.) or yohimbine (2 mg/kg, i.p.), an α2-adrenergic receptor antagonist. In vitro, Helicobacter pylori action was done by broth microdilution method. RESULTS: The HPLC analysis data revealed the presence of gallic acid, rutin, myricetin, morin and kaempferol. HECr promoted protective effect against acute ulcers induced by HCl/EtOH with inhibitions of 47.52% (p < 0.01) and 62.69% (p < 0.001) at 100 and 400 mg/kg, and in piroxicam by 34.11% (p < 0.05), 49.14% (p < 0.01) and 61.34% (p < 0.001), at 25, 100 or 400 mg/kg, respectively, and in water restraint stress by 78.26% inhibition, p < 0.001, at the dose of 400 mg/kg when compared to the vehicle control group respectively. In the chronic gastric ulcer model, HECr (25, 100 and 400 mg/kg p.o.) significantly (p < 0.001) decreased the injured area by 58.80%, 77.87% and 71.10% respectively. Histological examination indicated that oral treatment of HECr promoted healing of gastric lesions by regenerating gastric mucosa layer with less inflammatory cells. HECr augmented the GSH, CAT activities and reduced MPO level. The pre-treatment with HECr increased the gastric wall mucus production. It also significantly altered the gastric secretion parameters by causing the reduction in the gastric juice volume, elevated the pH level and reduced the total acidity at all doses tested when compared with the vehicle group. HECr at the most active dose (100 mg/kg) reversed completely the reduction of PGs, NO production, closure of K+ATP- channels and α2-adrenoreceptor blockage - induced damages. In microdilution assay, the HECr showed good anti-Helicobacter pylori effect with MIC = 100 µg/mL. CONCLUSION: The HECr presented preventive and curative effects in the experimental gastric ulcer models, besides good anti-Helicobacter pylori activity, which supports the traditional medicinal use of the xylopodium of this plant for gastrointestinal diseases. The underlying mechanisms of this antiulcerogenic/antiulcer action involve, at least, augmentation of mucus production, inhibition of gastric secretion, stimulation of PGs and NO synthesis. And that it involves activation of K+ATP channels and α-2-adrenergic receptors, in addition to an antioxidant activity, probably due to the presence of gallic acid and flavonoids in HECr.

Chemical characterization and evaluation of gastric antiulcer properties of the hydroethanolic extract of the stem bark of Virola elongata (Benth.) Warb.

ETHNOPHARMACOLOGICAL RELEVANCE: Virola elongata is a tree species belonging to the Myristicaceae family, distributed in the North and Midwest regions of Brazil, in the phytogeographic domain of the Amazon. The aqueous infusion or the hydroethanolic macerate of the stem bark of V. elongata are used in Brazilian and Ecuadorian indigenous folk medicine for several ethnopharmacological purposes, principally, in the treatment of stomach pain, indigestions, and gastric ulcers. This study was aimed to investigate the gastroprotective activity of this plant in order to support its popular use with scientific evidence. MATERIALS AND METHODS: The stem bark hydroethanolic extract of the plant (HEVe) was prepared by maceration. Its qualitative and quantitative phytochemical constituents were investigated by classical colorimetric techniques, HPLC, and electrospray ionization-multiple stage fragmentation (ESI-MSn). The gastroprotective and antiulcer activity of HEVe at doses of 100, 300 and 900 mg/kg p.o. were tested using three acute (acidified ethanol, piroxicam, and in-water-restrain stress), and one chronic (acetic acid) animal ulcer models. The probable mode of action of the HEVe was evaluated by analyzing gastric acid secretion, mucus content, nitric oxide effect, and its antioxidant properties (on catalase, myeloperoxidase, and GSH content) in experimental rodents. The direct extract's activity on the growth of Helicobacter pylori was also investigated. RESULTS: Total phenolic content in the HEVe was of 146.20 ±â€¯1.07 mg, being flavonoids about 50% (71.79 ±â€¯0.70 mg) of it. Comparative HPLC fingerprint analysis revealed the presence of known phenolic antiulcer compounds, such as gallic acid, catechin, and rutin. Also, methanol/water fractionation and ESI-MSn analysis of the HEVe reveals the presence of quinic acid, 3,3',4-trihydroxystilbene, juruenolid D, one catechin dimer, one C-glycosyl flavonoid, one polyketide and two neolignans as the major components of the extract. The HEVe attenuated gastric ulceration in all the different models of acute gastric ulcer, by enhancing gastroprotection through its antioxidant properties in vivo, and reducing also considerably the gastric secretion and total acidity. The HEVe also presented healing properties against the induced chronic ulceration process. On the other hand, the HEVe did not exhibit direct activity against H. pylori. CONCLUSION: The HEVe exhibited significant gastroprotective/antiulcer effects and contain a relative high proportion of phenolic compounds, especially flavonoids, that could likely account, at least in part, for its pharmacological properties. The results justify its traditional usage and provided scientific evidence for its potential as a new herbal medicine to treat gastric ulcers.

Chemical characterisation and toxicity assessment in vitro and in vivo of the hydroethanolic extract of Terminalia argentea Mart. leaves.

ETHNOPHARMACOLOGICAL RELEVANCE: Terminalia argentea Mart. (Combretaceae), known mainly as "capitão", is a native tree, not endemic, that occurs in the Amazon, Caatinga, Cerrado and Atlantic Forest in Brazil. Leaf infusion is popularly mentioned by riverine communities that inhabit the microregion of Northern Araguaia (Mato Grosso, Brazil) for the treatment of gastric ulcer, bronchitis and haemorrhage. Considering the wide medicinal use, lack of studies that evaluate the safety of use and the scarcity of phytochemical studies of T. argentea leaves, this work was carried out with the objective of evaluating the toxicity of the hydroethanolic extract of the leaves of T. argentea Mart. (HETa) in experimental models in vivo and in vitro, as well as to advance the phytochemical analysis of HETa. MATERIALS AND METHODS: HETa was prepared by macerating the leaf powder in hydroethanolic solution. Phytochemical characterisation was carried out by thin-layer chromatography (TLC), high-performance liquid chromatography (HPLC) and mass spectrometry through direct flow infusion coupled with electrospray ionization and ion-trap analyzer (DFI-ESI-IT-MS analyses) The contents of phenols, flavonoids and phytosterols were analysed by colorimetric methods. Cytotoxicity was assessed by the Alamar blue assay on Chinese hamster ovary epithelial cells (CHO-K1) and human gastric adenocarcinoma cells (AGS). In vitro genotoxicity of HETa (10, 30 or 100 µg/mL) was assessed by micronucleus (MN) and comet tests using CHO-K1 cells. The acute toxicity assessment was performed by oral administration of HETa in single dose Swiss mice (males and females) up to 2000 mg/kg and sub-chronic toxicity by daily oral administration of HETa (50, 200 and 800 mg/kg) in Wistar rats for 30 days. The parameters related to the clinical and toxicological observations were determined every 6 days and at the end of the treatment the blood was collected for biochemical and haematological analysis, and some organs were removed for macroscopic and histopathological analysis. RESULTS: Preliminary phytochemistry and TLC analysis of HETa revealed the presence of phenolic compounds (18.8%), flavonoids (10.8%), saponins, tannins and phytosterols (19%). The HPLC data revealed the presence of gallic acid, rutin, ellagic acid, catechin, quercetin and kaempferol. In the analysis by DFI-ESI-IT-MS, the presence of gallic acid, rutin, ellagic acid and quercetin was confirmed and identified caffeic acid, quinic acid, galloylmucic acid, quercetin xyloside, quercetin rhamnoside, quercetin glucoside, caffeoyl ellagic acid, quercetin galloyl xyloside, terminalin, quercetin galloyl glucose, corilagin, quercetin digalloyl xyloside, quercetin digalloyl glucoside, punicalin and punicalagin. HETa showed no cytotoxic effect on CHO-K1 and AGS cells. In the MN assay, HETa increased the number of MNs and nuclear buds (NBUDs) in binucleate cells at the three concentrations tested and the nucleoplasmic bridges (NPBs) number at 30 µg/mL. In the comet test, HETa (10 and 100 µg/mL) alone showed a genotoxic effect on CHO-K1 cells. In pre-treatment, HETa at all concentrations tested prevented DNA damage induced by H2O2. In co-treatment with H2O2, HETa showed genotoxic effects at the three concentrations, and post-treatment DNA damage in exposed CHO-K1 cells to H2O2 was repaired in 22.5% with 10 µg/mL HETa. In the acute toxicity test, the HETa did not cause death in the mice, being verified only by piloerection and reversible in 2 h in males and in 4 days in females. No macroscopic changes were observed in the analysed organs. In the sub-chronic toxicity test, the HETa did not cause death in the rats after 30 days and the few changes were: absolute (103/mm3) and relative (%) values of basophils increased by 477.8% and 423% (p < 0.001), respectively, with 50 mg/kg; reduction in feed intake (23.6%, p < 0.01) only on day 18; total cholesterol concentration (13.1%, p < 0.05) and relative heart weight (13.2% %, p < 0.05) at a dose of 800 mg/kg. These effects were not dose-dependent nor followed by clinical signs and symptoms of intoxication, nor of macroscopic and histopathological changes in the organs of animals treated with HETa. CONCLUSIONS: The results demonstrated that HETa had no cytotoxic in vitro effects for CHO-K1 and AGS cells. In in vitro genotoxicity assays, the HETa induced different responses, according to concentration and experimental condition. In the MN test the HETa presented genotoxic potential by increasing the number of MNs, NBUDs and NPBs. In the comet assay, HETa was genotoxic by itself and in the co-treatment protocol with H2O2. In pre-treatment or post-treatment protocols with H2O2, HETa presented an antigenotoxic effect by preventing or repairing, respectively, the genotoxicity induced by H2O2. In the in vivo models, HETa was shown to be relatively safe after acute administration in mice [no-observed-adverse effect level (NOAEL) of 2000 mg/kg] and sub-chronic in rats (NOAEL of 800 mg/kg), confirming the riverine information that it is non-toxic in the dosage used. Phytochemical analysis of HETa revealed the presence of phenolic compounds, flavonoids, saponins, tannins and phytosterols. Among the flavonoids and tannins, we highlight gallic acid, rutin, ellagic acid, quercetin, caffeic acid, quinic acid, corilagin, punicalin and punicalagin. Thus, it can be stated that HETa has a good safety margin for therapeutic use.

Sphenodesme involucrata var. paniculata (C.B. Clarke) Munir.: Chemical characterization, anti-nociceptive and anti-inflammatory activities of methanol extract of leaves.

ETHNOPHARMACOLOGICAL RELEVANCE: Sphenodesme involucrata var. paniculata (C. B. Clarke) Munir is native as well as endemic to South India. Its leaves are used in folklore medicine to treat pain and rheumatism. OBJECTIVE: This study was aimed to investigate the chemical characterization, anti-nociceptive and mode of action underlying the anti-inflammatory effects of methanol extract of S. involucrata leaves (MESi). METHODS: Phytoconstituents of MESi was analyzed using colorimetric and liquid chromatography-mass spectrometry (LC-MS) methods, and the oral acute toxicity was evaluated in mice up to 2000 mg/kg. The anti-nociceptive effect was evaluated in hot plate and writhing tests; whereas the anti-inflammatory effect was investigated using carrageenan, cotton pellet and lipopolysaccharide (LPS)-induced peritonitis models at doses of 100, 200 and 400 mg/kg. Additionally nitric oxide (NO) and inflammatory cytokines levels were also evaluated. RESULTS: MESi exhibited the high content of phenolics and flavonoids as well as compounds like austricine, benzylglucosinolate, gossypin, justicidin B and cirsimarin were detected in LC-MS. In the acute toxicity study, oral administration of MESi did not cause any toxic effect and mortality up to 2000 mg/kg body weight in mice. In the anti-nociceptive tests, MESi augmented the latency period at higher dose (400 mg/kg), on the other hand attenuated writhings at the dose of 400 mg/kg by 87.87% (p < 0.001). In the carrageenan induced paw oedema MESi significantly inhibited the oedema formation at dose 400 mg/kg by 32.1%; besides, anti-inflammatory effect was registered in the cotton pellets-induced inflammation model at doses 200 and 400 mg/kg by 27.09% (p < 0.001) and 35.47% (p < 0.001) respectively. On the other hand, MESi appreciably reduced leukocyte, neutrophils infiltration, nitric oxide, TNF-α and IL-1ß levels and increased the IL-10 level in the (LPS)-induced peritonitis model. CONCLUSION: The results conclude that MESi has no acute toxic effect and it demonstrated potent anti-nociceptive and anti-inflammatory activities. Its anti-nociceptive activities are probably mediated through peripheral and central mechanisms. The anti-inflammatory effect of MESi involved the inhibition of neutrophils migration and the modulation of Th1 and Th2 cytokines, besides the attenuation of production of PGE2 and NO. LC-MS analysis revealed the predominant presence of the austricine, benzylglucosinolate, gossypin, justicidin B and cirsimarin compounds, which are possibly involved in the anti-nociceptive and anti-inflammatory effects of MESi. The current study provided supportive evidence for the folklore use of S. involucrata in the treatment of pain and inflammatory conditions.

Evaluation of genotoxicity and subchronic toxicity of the standardized leaves infusion extract of Copaifera malmei Harms in experimental models.

ETHNOPHARMACOLOGICAL RELEVANCE: Copaifera malmei Harms (Fabaceae), known mainly as óleo-mirim, is a native and endemic plant found in the states of Mato Grosso and Goiás of Brazil. The plant's leaves infusion is popularly used by riverine communities of the northern Araguaia microregion, Mato Grosso, Brazil, for the treatment of gastric ulcers and inflammatory diseases of the respiratory tract. The gastric antiulcer activity of the standardized leaves infusion extract of Copaifera malmei (SIECm) in rodents has been reported. The objective of this study was to advance the investigation of the safety profile of SIECm by evaluating the genotoxicity and subchronic toxicity using in vitro and in vivo experimental models. MATERIALS AND METHODS: SIECm was prepared by infusion, by incubating the powdered dried leaves material in boiled water for 15min. In vitro genotoxicity of SIECm (10, 30 or 100µg/mL) was assessed by micronucleus and comet tests using Chinese hamster ovary (CHO-k1) epithelial cells. The evaluation of subchronic toxicity profile was performed by daily oral administration of SIECm (100, 400 or 1000mg/kg) to Wistar rats for 30 days. Clinical observations of toxicological related parameters were done every 6 days. After the treatment period, blood was collected for hematological and biochemical analysis, and some organs were removed for macroscopic and histopathological analysis. RESULTS: In the micronucleus assay, SIECm demonstrated anti-mutagenic activity. In the comet assay, SIECm presented anti-genotoxic effect preventing DNA damage at all the three concentrations tested with pre-treatment, while the same effect was only observed in the co-treatment at the lowest concentration. Post-treatment with SIECm increased the genetic damage induced by hydrogen peroxide (H2O2) at the highest concentration. In the subchronic toxicity test, few changes were observed, such as increase in feed consumption in the group of animals treated with 100mg/kg of the SIECm, which reversed after 6 days. There were no macroscopic, histological and relative weights changes in the organs of animals treated with SIECm. No toxicologically relevant changes were observed in the hematological analysis. Subchronic administration of SIECm reduced levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in animals treated with 100mg/kg and serum triglyceride levels at 400 and 1000mg/kg. However, the hematological and biochemical changes observed are within the physiological ranges for this animal species. CONCLUSION: The results demonstrate that SIECm is not genotoxic, and does not present toxicity when used orally for up to 30 days. In addition, it showed protection to the genetic damage induced by H2O2. The SIECm therefore has a high safety margin for therapeutic use.

Vitexin inhibits inflammation in murine ovalbumin-induced allergic asthma.

Vitexin is an important component of various medicinal plants frequently used to treat asthma, such as Crataegus spp., Vitex spp., Passiflora spp., and Echinodorus spp. However, there is no information about the vitexin potential as anti-asthmatic. The aim of the present work was to evaluate the anti-hypersensitive activity of vitexin in a murine ovalbumin (OVA)-induced allergic asthma model. Mice were sensitized to OVA by i.p. injection on days 1st and 10th, followed by a daily challenge with OVA using a nebulizer, from days 19th to 24th. Vitexin or dexamethasone were orally administered 1h before each OVA challenge. Vitexin attenuates migration induced by OVA-hypersensitivity of eosinophil, neutrophil, and mononuclear cells in bronchoalveolar lavage fluid (BALF). Histological analysis of the lungs shows that vitexin suppressed leukocyte infiltration, mucus production and pulmonary edema. Increases in Th2 cytokines in BALF in OVA-induced asthma is also attenuated by vitexin, as well as plasma levels of IgE. Overall, these results suggest that vitexin can suppress OVA-induced allergic inflammation in mice and provide a strong rationale for further developing vitexin as a candidate treatment for allergic hypersensitivity. These data corroborate the popular use of vitexin-rich plants for asthma treatment.

Heteropterys tomentosa A. Juss: Toxicological and adaptogenic effects in experimental models.

BACKGROUND: The constant pursuit of improved athletic performance characterizes high-performance sport and the use of medicinal plants as dietary supplements is becoming widespread among athletes to enhance long-term endurance performance. AIM: The present study evaluated the toxicity of Heteropterys tomentosa (HEHt) and its acute adaptogenic effects. METHODS: The in vitro safety profile was evaluated on CHO-k1 cells using the alamar Blue assay, at concentrations ranging from 3.125 to 200 µg/mL. In vivo acute oral toxicity was conducted in male and female mice with oral administration of graded doses of HEHt from 400 to 2000 mg/kg. A subchronic oral toxicity study was completed by oral administration of HEHt (50, 200 or 1000 mg/kg) and vehicle for 30 days in male Wistar rats. Clinical observations and toxicological related parameters were determined. Blood was collected for biochemical and hematological analyses, while histological examinations were performed on selected organs. Thereafter, an adaptogenic test consisting of progressive loads until exhaustion was conducted in rats ( n = 5/group) orally pre-treated with the vehicle and HEHt (25, 100 or 400 mg/kg). RESULTS: HEHt exhibited no cytotoxic effects on the CHO-k1 cells and, apparently, no acute toxicity in mice and no subchronic toxicity in rats. An ergogenic effect was observed only at the dose of 25 mg/kg compared with the vehicle in relation to time to exhaustion and exercise load ( p = .011 and .019, respectively). HEHt is safe at up to 400 mg/kg, contains astilbin and taxifolin as the major phytochemical compounds, and exhibited a potential adaptogenic effect. CONCLUSIONS: These results justify its anecdotal usage as a tonic, show that the hydroethanolic maceration of the root does not cause toxicity, and provide scientific evidence of its potential as a source of new adaptogenic substance(s).

Chemical characterization, toxicology and mechanism of gastric antiulcer action of essential oil from Gallesia integrifolia (Spreng.) Harms in the in vitro and in vivo experimental models.

Gallesia integrifolia is a Brazilian Amazon tree whose bark decoction is popularly used to treat peptic ulcer. The essential oil from the inner stem bark of G. integrifolia (EOGi) was chemically characterized by GC/MS. The in vitro cytotoxicity and genotoxicity were evaluated in CHO-K1 cells, while the in vivo oral acute toxicity was performed in mice. The gastroprotective effect of EOGi was assessed in acidified ethanol and piroxicam and ulcer healing on acetic acid -induced ulcer models in rodents. Anti-secretory, mucus, K+-ATP channels, prostaglandins (PGs), nitric oxide (NO), TNF-α, IL-1ß, IL-10, catalase (CAT) and myeloperoxidase (MPO) activities and in vitro Helicobacter pylori action by EOGi were evaluated. EOGi exhibited cytotoxic effects only at 72h and no acute toxicity. EOGi showed gastroprotective and ulcer healing effects. EOGi gastroprotection was attenuated by indomethacin pre-treatment. Gastric volume and total acidity were reduced, while gastric pH was elevated. EOGi increased mucus and NO productions and CAT activity, and inhibited MPO activity, TNF-α and IL-1ß concentrations and augmented IL-10. EOGi was not active against H. pylori. These results indicated that EOGi is safe and exerts preventive and curative gastric ulcer effects by multitarget actions. Twenty compounds were identified and (-)-alpha-santalene was the main compound.

Hydroethanolic extract from Echinodorus scaber Rataj leaves inhibits inflammation in ovalbumin-induced allergic asthma.

ETHNOPHARMACOLOGICAL RELEVANCE: Echinodorus scaber, Alismataceae, is popularly known in Brazil as "chapéu-de-couro". The plant leaves are used by the population as decoction, infusion, or maceration in bottled spirits, to treat inflammatory respiratory diseases. AIM OF THE STUDY: To investigate the anti-inflammatory mechanism of the hydroethanolic extract of leaves of Echinodorus scaber (HEEs) in allergic asthma. A phytochemical analysis of the extract was performed as well. MATERIALS AND METHODS: The leaves of Echinodorus scaber were prepared by maceration in 75% ethanol. Preliminary phytochemical analysis was carried out using basic classical methods, and the secondary metabolites detected in HEEs were analyzed and confirmed by high-performance liquid chromatography (HPLC). The in vivo anti-inflammatory activity of HEEs was evaluated in Swiss male albino mice sensitized and challenged by OVA. The HEEs (1, 5 and 30mg/kg, p.o.) was administered to mice twice a day, 1h before the challenge, from days 19 through 24. The mechanism of action of HEEs was studied by evaluating the levels of TH2 cytokines (IL-4, IL-5 and IL-13) in bronchoalveolar lavage fluid (BALF) and IgE production in blood plasma. Histopathological changes triggered by OVA-sensitization/challenge in the lung tissue were also investigated. RESULTS: HEEs reduced total leukocyte, eosinophil, neutrophil, and mononuclear cell counts at all doses tested, with maximum effect at 30mg/kg (73.9%, 75.9%, 75.5%, and 65.2% reduction, p<0.001, respectively). Increases in TH2 cytokine secretion (IL-4, IL-5 and IL-13) and in IgE levels were also attenuated by HEEs. Preliminary phytochemical screening seems to indicated the presence of phenolic compounds, flavonoids and alkaloids. HPLC analyses evidenced the presence of phenolic compounds, such as gallic acid, rutin and vitexin. CONCLUSION: Our findings provided pharmacological preclinical evidence for the popular use of the leaves of Echinodorus scaber in allergic inflammation. Its anti-inflammatory effect was dependent on the decrease in migratory inflammatory cells, and both TH2 cytokines and IgE levels. It is suggested that vitexin, gallic acid and rutin, known anti-inflammatory compounds, may participate in the anti-asthamtic effect of the HEEs, by acting jointly along with other components present in the extract.

Mandevilla longiflora (Desf.) Pichon improves airway inflammation in a murine model of allergic asthma.

ETHNOPHARMACOLOGICAL RELEVANCE: Mandevilla longiflora, popularly known as "velame" in central Brazil, is a subshrub widely distributed in South America. Its xylopodium is used in the form of a decoction or infusion to treat inflammation and other ailments. AIM OF THE STUDY: This study aimed to evaluate the anti-inflammatory potential of M. longiflora in an in vivo model of ovalbumin-induced immediate hypersensitivity, identifying its effects on leukocyte infiltration, IgE and LTB4 levels, and Th2 cytokine production. In addition, HPLC fingerprint of the extract was performed. MATERIAL AND METHODS: The hydroethanolic extract 70% of M. longiflora (HEMI) was obtained by maceration of the plant xylopodium. Swiss mice were sensitized by i.p. injection OVA-aluminium hydroxide on days 1 and 10. Nine days after the last sensitisation animals were challenged for 6 consecutive days with OVA solution for 20min daily in a closed chamber under continuous flow of aerosol. The animals were treated with HEMl (20, 50 and 200mg/kg p.o.), 2 times per day, and euthanized 24h later. Animals treated with vehicle (2% Tween-20) or dexamethasone were used as negative and positive controls, respectively. The recruitment of inflammatory cells into the pulmonary cavity was evaluated by counting cells present in broncho-alveolar lavage fluid (BALF). Lung tissue was also collected for histopathology and infiltration analysis. Quantification of IL-4, IL-5 and IL-13 from the BALF, and IgE, and LTB4 from plasma, were conducted by immunoassay. RESULTS AND CONCLUSIONS: The HEMl attenuated leukocyte migration into the airways, which was evidenced by a decrease in eosinophils, neutrophils and mononuclear cells, both in BALF quantification and by histopathological analysis, as well as decreasing the concentrations of IL-4, IL-5, IL-13, IgE and LTB4. All of these events are typical of air-mucosa inflammatory disease. These findings scientifically evidence for the first time the ethnopharmacological use of M. longiflora to treat chronic inflammatory events, such as asthma, and suggest a potential therapeutic use or complementary therapy for this plant extract.

Evaluation of the safety, gastroprotective activity and mechanism of action of standardised leaves infusion extract of Copaifera malmei Harms.

ETHNOPHARMACOLOGICAL RELEVANCE: Copaifera malmei Harms (Fabaceae) is a plant that occurs in the central region of Brazil, where the plant's leaves infusion is popularly used to treat gastric ulcer and inflammatory diseases. This study was aimed to investigate the gastroprotective activity and mode of action of the plants' leaves infusion in order to establish the scientific basis for such usage, and to assess its potential as a source of an anti-ulcer agent. MATERIALS AND METHODS: Leaves infusion extract of the plant (SIECm) was prepared, freeze dried and lyophilised. Its qualitative and quantitative phytochemical constituents were investigated using TLC and HPLC techniques. The safety profile was evaluated on CHO-k1 epithelial cells viability using the Alamar blue assay, and by acute toxicity test in mice. The gastroprotection and anti-ulcer efficacy of the SIECm (25, 100 and 400mg/kg, p.o.) were tested using acute (acidified ethanol, piroxicam and water restrain stress), and chronic (acetic acid) experimental ulcer models. The plausible mode of action of the SIECm was assessed using gastric secretion, gastric barrier mucus, nitric oxide, and its antioxidant (myeloperoxidase and catalase) effects in mice and rats. The histopathological analyses of the ulcerated tissues as well as the extract's activity on Helicobacter pylori were also investigated. RESULTS: Phytochemical tests indicated the presence of mainly phytosterols, phenolics and flavonoids. The SIECm exhibited no cytotoxic effects on the CHO-k1 cells, and no oral acute toxicity in mice. It prevented against the acute induced ulcerations by enhancing gastroprotection through gastric mucus production, NO modulation, antioxidant, reduced gastric secretion and enhanced chronic ulcers healing process, as shown by reduction/prevention of epithelial and vascular damage, in addition to reduction in leucocyte infiltration. The SIECm however did not exhibit activity against H. pylori. CONCLUSION: The SIECm is safe, contain useful phytochemicals and exhibited significant gastroprotective/anti-ulcer effects. The results justify its folkloric usage, and provided scientific evidence of its potential as a source of new phytodrug to treat gastric ulcers.