Mecanismos de ação relacionados à atividade antiúlcera de Kalanchoe pinnata (Lam.) Pers. (Crassulaceae) / Mechanisms of action underlying antiulcer activity of Kalanchoe pinnata (Lam.) Pers. (Crassulaceae).

Kalanchoe pinnata (Lam.) Pers. (Crassulaceae) é uma espécie muito empregada na medicina tradicional no Brasil e em outras partes do mundo, especialmente Índia, países da África e China. É indicada popularmente para diversos fins incluindo o tratamento de úlceras gástricas. A análise fitoquímica revelou a presença de vários constituintes, em especial os flavonoides. O tratamento de úlcera gástrica convencional apresenta diversos efeitos colaterais e, na maioria das vezes, não evita a recidiva da lesão. Dessa maneira, é interessante encontrar uma terapêutica mais segura e efetiva. Com o objetivo de avaliar a segurança, foi realizado ensaio de citotoxicidade do extrato bruto, in vitro, com valor de IC50 igual a 0,926 mg/mL, sendo possível predizer um valor de LD50 (1341,46 mg/kg). Já em relação ao ensaio de citotoxicidade, in vitro, da fração acetato de etila não foi encontrado um valor de IC50. Resultados de fototoxicidade, in vitro, mostraram que o extrato bruto e fração acetato de etila de K. pinnata não possuem potencial fototóxico. A contagem microbiana na droga vegetal para bactérias aeróbias/mesófilas foi de 6,9 x 104 UFC/g e a contagem de bolores e leveduras foi de 2,4 x 103 UFC/g, ambos valores dentro do limite estabelecido pela OMS. Análise de endotoxinas também foi realizada para o extrato bruto (<4,0.105 UE/kg) e fração acetato de etila (<2,7.105 EU/kg) de K. pinnata. Referente à fitoquímica, diversos flavonoides foram identificados no extrato bruto e fração acetato de etila de K. pinnata. Paralelamente ao estudo fitoquímico foi verificado que a atividade gastroprotetora do extrato bruto envolve a ação das prostaglandinas e grupamentos sulfidrila. Já o mecanismo de gastroproteção da fração acetato de etila é dependente de prostaglandinas e óxido nítrico. A atividade cicatrizante do extrato bruto de K. pinnata também foi avaliada. De acordo com os resultados macroscópicos, as doses de 200mg/kg e 400 mg/kg reduziram a área de lesão, com uma taxa de 33% e 39%, respectivamente, após 7 dias de tratamento (p<0,05). Análise histológica dos grupos tratados com o extrato bruto (200 e 400 mg/kg) indicou melhor recuperação da lesão, verificada pela regeneração da mucosa gástrica e pelo restabelecimento da arquitetura glandular. As enzimas antioxidantes (catalase, superóxido dismutase e glutationa peroxidase) e a expressão de VEGF foram avaliadas no mecanismo de cicatrização de úlceras gástricas. Os resultados mostraram que a atividade antiulcerogênica foi mediada pela ação antioxidante da enzima SOD. Não foi evidenciado in vivo o aumento da expressão de VEGF e nem o sequestro do radical peroxil nos animais tratados com o extrato bruto. Os resultados dos ensaios in vitro (ORAC) mostraram uma maior capacidade de sequestro de radicais peroxil da fração acetato de etila (1192,35 ± 112,61 µmol equivalente de Trolox/g de amostra seca) quando comparado com o extrato bruto (431,32 ± 7,17 µmol equivalente de Trolox/g de amostra seca). A atividade anti Helicobacter pylori também foi avaliada, no entanto, o extrato bruto não apresentou atividade anti H.pylori. Ademais, o extrato bruto demonstrou um potencial anti-inflamatório, pois foi observada uma redução nos níveis de TNF-α e L-selectina, após o tratamento em neutrófilos estimulados com LPS. Analisando os resultados sugere-se que K. pinnata possui um potencial terapêutico no combate de úlceras gástricas e possivelmente, anti-inflamatório, sendo que os flavonoides podem estar relacionados com o efeito biológico observado.

Kalanchoe pinnata (Lam.) Pers. (Crassulaceae) is a commonly used species in traditional medicine in Brazil and in other parts of the world, especially India, Africa and China, for the treatment of various diseases, including gastric ulcers. Phytochemical analysis revealed the presence of several constituents in this plant, especially flavonoids. The available pharmaceutical products to treat peptic ulcer have several side effects and, in most cases, do not prevent recurrence of the gastric lesions. Therefore, it is important to find a safer and more effective therapy. In order to evaluate safety, the in vitro cytotoxicity assay of crude extract from K. pinnata was performed. The IC50 value was 0,926 mg/mL corresponding to LD50 value (1341, 46 mg/kg). It was not determined IC50 value in vitro cytotoxicity assay for ethyl acetate fraction from K. pinnata. Neither the crude extract nor ethyl acetate fraction from K. pinnata showed phototoxicity. Microbial counting was performed on the K. pinnata-based drug in order to investigate microbiological contamination. The microbial count for aerobic / mesophilic bacteria was 6.9 x 104 CFU/g, and yeast count was 2.4 x 103 CFU/g, both values in agreement with the limits established by WHO. Endotoxin analysis was also performed for the crude extract (<4,0.105 UE/kg) and for ethyl acetate fraction (<2,7.105 UE/kg) from K. pinnata. In the phytochemical analysis several flavonoids were identified in the crude extract and ethyl acetate fraction of K. pinnata. In parallel to the phytochemical study, it was verified that the gastroprotective activity of the crude extract of K. pinnata involved prostaglandins and sulfhydril compounds. On the other hand, the mechanism of gastroprotection of the ethyl acetate fraction of K. pinnata is dependent on prostaglandins and nitric oxide. The healing activity of the crude extract of K. pinnata was also evaluated. According to the macroscopic results the dose of 200 mg/kg and 400mg/kg reduced the injury area, with a rate of 33% and 39%, respectively, after 7 days of treatment (p <0.05). Histological analysis showed regeneration of the gastric mucosa and re-establishment of the glandular architecture in groups treated with the crude extract (200 and 400 mg/kg). Antioxidant enzymes (catalase, superoxide dismutase and glutathione peroxidase) and VEGF expression were evaluated in the mechanism of gastric ulcer healing. The results showed that the antiulcerogenic activity was mediated by SOD. It was not demonstrated an increase in VEGF expression and nor in the in vivo sequestration of the peroxyl radical in the animals treated with crude extract. The results of in vitro assay (ORAC) showed a greater sequestering of peroxyl radical to the ethyl acetate fraction (1192,35 ± 112,61 µmol equivalent of Trolox/g of ethyl acetate fraction) when compared to the crude extract (431,32 ± 7,17 µmol equivalent of Trolox/g of crude extract) of K. pinnata. The anti Helicobacter pylori activity was also evaluated; however, the crude extract did not show anti H. pylori activity. However, the crude extract of K. pinnata demonstrated an anti-inflammatory potential, because TNF-α and L-selectin levels were reduced after treatment in LPS-stimulated neutrophils. The analysis of the results suggests that K. pinnata has a therapeutic potential against gastric ulcers and possible anti-inflammatory properties, and the flavonoids may be linked to the biological effect.

Gastroprotective activity of the hydroethanolic extract and ethyl acetate fraction from Kalanchoe pinnata (Lam. ) Pers

ABSTRACT Peptic ulcers are an important pathology, and the search for safer and more effective treatment methods is of paramount importance. In this study, we assess the gastroprotective effects of the hydroethanolic extract (HE) and ethyl acetate fraction (EAF) from Kalanchoe pinnata leaves against an ethanol/HCl-induced ulcer model in rats. The HE reduced gastric lesions by approximately 47% (400 mg/kg). A significant inhibition of the gastric lesions by 50% was observed after pretreatment with the EAF (200 mg/kg). Quercetrin and quercetin 3-O-α-L-arabinopyranosyl-(1→2)-α-L-rhamnopyranoside were isolated and identified in the flavonoid fraction (EAF) by HPLC and NMR analyses because this fraction showed the highest gastroprotective effect. This fraction demonstrated high antioxidant activities (CE50=41.91 µg/mL) by DPPH in comparison with Trolox(r) and 11.33 mmol Trolox(r) equivalent by ORAC. In conclusion, the HE and FAE from K. pinnata displayed gastroprotective activity in rats, most likely due to the presence of flavonoids.

The “Wonder Plant” Kalanchoe pinnata (Linn.) Pers.: A Review.

Kalanchoe pinnata (Linn.) Pers. is a plant found mostly in the temperate and tropical regions of the world. It is traditionally known to exhibit a wide range of pharmacological activities that involves treatment for the most serious disorders related to mankind. In this review, the therapeutic and medicinal values of the plant which includes its wound-healing, antioxidant, anticancerous, antiproliferative, antimicrobial, antiviral, antiprotozoal, antileishmanial, anthelmentic, insecticidal, anti-allergic, analgesic, antinociceptive, anti-oedematogenic, anti-inflammatory, muscle-relaxant, antipyretic, anticonvulsant, antidepressant, sedative, antilithiatic, hepatoprotective, gastroprotective, antidiabetic, nephroprotective, haemoprotective, antihistamine, antihypertensive and immunosuppressant activities have been comprehensively discussed. In ethnomedicine, it is known for its anthroposophical and tocolytic effects in pregnant women. Also, it is used to facilitate dropping of placenta during child birth. Scientists have explored the different parts of the plant and have established the clinical potentials of the plant as a whole and its parts successfully. Few scientific validations have even lead to the isolation and determination of the applications of the bioactive compounds from various solvent extracts of the plant. Further research and clinical trials have to be carried out in order to commercialise the potential pharmaceutical uses of the plant for which one should thoroughly know about the pharmacognostic properties of the plant.

Hypoglycemic and hypocholesterolemic activities of the aqueous preparation of Kalanchoe Pinnata leaves in streptozotocin-induced diabetic rats

Objective: To evaluate the hypoglycemic and hypocholesterolemic activities of the aqueous preparation of Kalanchoe pinnata (K. pinnata) leaves in streptozotocin-induced diabetic rats. Methods: Diabetes mellitus was induced in rats by a single administration of streptozotocin (60 mg/Kg). Diabetic rats were then treated with aqueous K. pinnata for 30 d. Serum glucose, proteins, lipid composition, liver and kidney function indices, inflammatory markers, and key enzymes of hepatic carbohydrate and lipid metabolism were determined. Results: The untreated and treated diabetic groups lost weight and consumed less food compared to the normal group. We noted 37.9% decrease in fasting blood glucose in the treated diabetic group compared to 13.2% and 17.0% increases in normal and untreated diabetic groups respectively. Serum cholesterol and triglyceride levels were significantly (P<0.05) reduced in the treated diabetic group compared to the untreated diabetic group. Blood urea nitrogen was significantly (P<0.05) elevated in the untreated and treated diabetic groups compared to the normal group. Serum alkaline phosphatase and hepatic pyruvate kinase activities were significantly (P<0.05) elevated in the treated diabetic group. Serum albumin level was significantly (P<0.05) reduced in the untreated diabetic group. Serum IL-6 was significantly (P<0.05) depressed in the treated diabetic group. Conclusions: The observed decrease in body weight, blood glucose and cholesterol level suggests that the aqueous K. pinnata preparation consumption may be beneficial in the management of diabetes mellitus. The observed adverse effect on alkaline phosphatase activity may be due to the combined effect of streptozotocin-induced diabetes and K. pinnata preparation administration.

Comparative pharmacognosy of Pashanbhed.

Background: Pashanbhed is a commercially available diuretic and lithotropic drug, used to treat renal problems. It is a controversial name as it is assigned to various plants such as Bergenia ligulata, Kalanchoe pinnata, Coleus aromaticus and Rotula aquatica. Objective: To perform the comparative preliminary phytochemical screening, diuretic activity, and thin layer chromatography (TLC) fi nger printing profi le of three plants (B. ligulata, C. aromaticus, and K. pinnata), most commonly used as Pashanbhed. Materials and Methods: Diuretic potential of methanolic extract (ME) of three plants were evaluated at two dose levels (500 and 1,000 mg/kg p.o.), using normal Wistar rats (Lipschitz method). Furosemide (20 mg/kg p.o.) was used as a standard drug. The effect on urine output and electrolyte changes were measured for 24 h and compared. All MEs were screened preliminarily for their constituents and their TLC fi nger printing profi les were prepared. One-way analysis of variance (ANOVA) followed by Bonferroni’s multiple comparison test. P < 0.05 was considered statistically signifi cant. Results: The MEs of all three plants have shown diuresis in normal rats. However, in intercomparison of the ME C. aromaticus (1,000 mg/kg p.o.) produced more signifi cant diuresis (P < 0.05) and electrolyte excretion compared to other test groups, the effect was at par with furosemide. The ME of these plants showed presence of alkaloids, glycosides, steroids, terpenoids, saponins, fl avonoids, etc. Conclusion: The ME of C. aromaticus (1,000 mg/kg p.o.) has showed highest diuretic action (4.2) among the tested extracts. This suggests the use of C. aromaticus leaves as “Pashanbhed”; the most effective diuretic drug.

Antilithiatic activity of poly-herbal formulation tablets by in-vitro method.

Hydroalcoholic extracts of Kalanchoe pinnata and Rotula aquatica and its combination were formulated herbal tablets, evaluated for antilithiatic in vitro method. The homogenous precipitation method was used. The study was carried out in glass tubes. The buffer system used was TRIS buffer pH 7.4. The experiment consists of the following tubes for control and test, 25 ml each of 25 mM CaCl2. 2H2O, 25 mM Na2HPO4. 2H2O or 25mM Na2C2O4. To the tubes of each set, tablet formulation or an equal amount of vehicle was added. The tubes were incubated at 37°C for 4 h. The precipitate of calcium phosphate was generated by mixing 1 ml of solution from the tubes having calcium chloride dihydrate and disodium hydrogen phosphate monohydrate and Calcium oxalate precipitate was generated by mixing 1 ml of solutions from the tubes having calcium chloride dihydrate and sodium oxalate solutions. Calcium was estimated using titrimetry and phosphorus was estimated using colorimetric analysis. Appropriate standard curves were done with each set of experiments. The amounts of precipitate of calcium and phosphate were determined in each of the respectively. The percent inhibition of the test was calculated in comparison with the control samples. Herbal tablet formulation showed antilithiatic activity to the marketed formulation in terms of inhibiting the formation of phosphate precipitate but showed a significantly better potential in preventing the formation of the calcium precipitate. The herbal tablet formulation of Kalanchoe pinnata and Rotula aquatica have inhibitory effect on calcium oxalate crystallization thus may be beneficial in the treatment of renal lithiasis.

3’,4’-Dimethoxy quercetin, a flavonol compound isolated from Kalanchoe Pinnata.

From methanol extract of Kalanchoe pinnata (Lam.) Pers. leaves, a flavonol compound has been isolated. Fractionation of the methanol extract with ethyl acetate, followed by ethyl acetate fraction purification using column chromatography method with ethyl acetate : n-hexane as mobile phase gave a yellow crystal compound (A). Further analysis using 1D- and 2D-nuclear magnetic resonance (NMR), confirmed with mass spectrometer (LC-MS), compound A identified as 3’,4’-dimethoxy quercetin.

Wound healing potential of ethanolic extract of Kalanchoe pinnata Lam. leaf—A preliminary study.

The extract of K. pinnata was evaluated for its wound healing activity by using excision wound model in rats. On day 11, animals treated with the ethanolic leaf extract exhibited 86.33 % reduction in the wound area, compared to petroleum jelly treated control (69.36%) and the mupirocin treated standard (85.49%). The hydroxyproline content of extract treated animals was higher, as compared to control and the standard groups. Histological analysis was also consistent with the proposal that K. pinnata leaf extract exhibits significant wound healing potential. The increased rate of wound contraction and hydroxyproline content in the extract treated animals supports the claims made by traditional healers of the benefits obtained from the medicinal use of K. pinnata.