In vitro relaxant and spasmolytic effects of essential oil of Pistacia integerrima Stewart ex Brandis Galls.

ETHNOPHARMACOLOGICAL RELEVANCE: Pistacia integerrima J.L. Stewart ex Brandis (Family: Anacardiaceae) galls are used in Indian ethnomedicine for its anti-asthmatic, sedative and spasmolytic properties, however, there are no scientific studies demonstrating its spasmolytic activity. The present investigation deals with the evaluation of relaxant and spasmolytic activities of the essential oil isolated from the galls of Pistacia integerrima J.L. Stewart ex Brandis (EOPI). MATERIALS AND METHODS: In vitro pharmacological assays were carried out on rabbit jejunum spontaneous contractions, guinea pig ileum. The present investigation studied the relaxation of basal tone of isolated guinea pig ileum by possible involvement of NO, prostaglandins, membrane Na(+) channels, potassium channel, enteric nervous system, adrenoceptors, Ca(2+) channels. Additional studies were conducted for comparison of the relaxant effects of EOPI on CaCl2 induced contraction in calcium free tyrode solution, effect on nifedipine insensitive component of ACh-induced contraction and on the contractile machinery to intracellular [Ca(2+)] on isolated guinea pig ileum. RESULTS: EOPI at non-relaxing dose potentiated the isoprenaline induced relaxation of rabbit jejunum. EOPI (50 µg/mL) exhibited 28% relaxation of basal tone of 60 mM K(+) induced contraction which is unaltered by preincubation with 0.5 mM hexamethonium, 0.5 µM Tetrodotoxin, 1 µM indomethacin, and 100 µM L-NG-Nitroarginine Methyl Ester (L-NAME). EOPI inhibited Ca(2+) induced contraction of isolated guinea pig ileum in Ca(2+) free medium. EOPI (10 µg/ml) potentiated the reversal of a KCl-induced tonic contraction has been observed in Ca(2+) free medium. CONCLUSION: The present investigation reinforces the use of Pistacia integerrima Stewart ex Brandis as antispasmodic in folk medicine. Moreover, it is demonstrated the involvement of ß- adrenoceptors and calcium channels in this activity, but not the participation of nicotinic receptors, Na(+) channels, prostaglandins or nitric oxide.

Embelia ribes ameliorates lipopolysaccharide-induced acute respiratory distress syndrome.

ETHNOPHARMACOLOGICAL RELEVANCE: Embelia ribes Burm. f. (Fam. Myrsinaceae) locally known as Vidanga have been used for treating tumors, ascites, bronchitis, jaundice, diseases of the heart and brain in traditional Indian medicine. However, no scientific studies providing new insights in its pharmacological properties with respect to acute respiratory distress syndrome have been investigated. AIM: The present investigation aimed to elucidate the effectiveness of Embelin isolated from Embelia ribes seeds on attenuation of LPS-induced acute respiratory distress syndrome in murine models. METHODS: Embelin (5, 10 and 20 mg/kg/day, i.p.) and Roflumilast (1 mg/kg/day, p.o.) were administered for four days and prior to LPS in rats (i.t.). Four hour after LPS challenge animals were anesthesized and bronchoalveolar lavage was done with ice-cold phosphate buffer. Assessment of BAL fluid was done for albumin, total protein, total cell and neutrophil count, TNF-α levels, nitrosoative stress. Superior lobe of right lung was used for histopathologic evaluation. Inferior lobe of right lung was used to obtain lung edema. Left lung was used for myeloperoxidase estimation. Arterial blood was collected immediately and analyzed for pH, pO2 and pCO2 were estimated. RESULTS: Pretreatment with embelin (5, 10 and 20 mg/kg, i.p.) decreased lung edema, mononucleated cellular infiltration, nitrate/nitrite, total protein, albumin concentrations, TNF-α in the bronchoalveolar lavage fluid and myeloperoxidase activity in lung homogenate. Embelin markedly prevented pO2 down-regulation and pCO2 augmentation. Additionally, it attenuated lung histopathological changes in acute respiratory distress syndrome model. CONCLUSION: The study demonstrates the effectiveness of Embelia ribes Burm. f. (Fam. Myrsinaceae) seeds in acute respiratory distress syndrome possibly related to its anti-inflammatory and protective effect against LPS induced airway inflammation by reducing nitrosative stress, reducing physiological parameters of blood gas change, TNF-α and mononucleated cellular infiltration indicating it as a potential therapeutic agent for acute respiratory distress syndrome.

Protective effect of Dillenia indica L. on acetic acid induced colitis in mice.

The inflammatory bowel disease (IBD) is an idiopathic, immune mediated and chronic inflammation of the intestine. The study aimed to elucidate the ameliorative effect of methanolic extract of Dillenia indica (DIME), hexane fraction (HFDI) and chloroform fraction (CFDI) of Dillenia indica in acetic acid induced experimental colitis in mice. Macroscopic score, colon weight, colonic catalase (CAT), superoxide dismutase (SOD), glutathione (GSH), myeloperoxidase (MPO), malondialdehyde (MDA), tumor necrosis factor (TNF-alpha), and histological changes were recorded after the treatment regimen of 7 days. Intra-rectal instillation of acetic acid caused enhanced macroscopic score, colon weight, colonic MPO, MDA, and TNF-alpha level. It caused significant decreased level of CAT, SOD and GSH. DIME (800 mg/kg), HFDI (200 mg/kg) and CFDI (200 mg/kg) treatment exhibited significant effect in lowering macroscopic score, colon weight, MPO, MDA, TNF-alpha levels and elevation of CAT, GSH and SOD levels. The results suggest that D. indica has ameliorating effects on experimental colitis by inhibiting the proinflammatory mediators like TNF-alpha production.

Estimation of the Long-term Cardiovascular Events Using UKPDS Risk Engine in Metabolic Syndrome Patients.

Long-term cardiovascular complications in metabolic syndrome are a major cause of mortality and morbidity in India and forecasted estimates in this domain of research are scarcely reported in the literature. The aim of present investigation is to estimate the cardiovascular events associated with a representative Indian population of patients suffering from metabolic syndrome using United Kingdom Prospective Diabetes Study risk engine. Patient level data was collated from 567 patients suffering from metabolic syndrome through structured interviews and physician records regarding the input variables, which were entered into the United Kingdom Prospective Diabetes Study risk engine. The patients of metabolic syndrome were selected according to guidelines of National Cholesterol Education Program - Adult Treatment Panel III, modified National Cholesterol Education Program - Adult Treatment Panel III and International Diabetes Federation criteria. A projection for 10 simulated years was run on the engine and output was determined. The data for each patient was processed using the United Kingdom Prospective Diabetes Study risk engine to calculate an estimate of the forecasted value for the cardiovascular complications after a period of 10 years. The absolute risk (95% confidence interval) for coronary heart disease, fatal coronary heart disease, stroke and fatal stroke for 10 years was 3.79 (1.5-3.2), 9.6 (6.8-10.7), 7.91 (6.5-9.9) and 3.57 (2.3-4.5), respectively. The relative risk (95% confidence interval) for coronary heart disease, fatal coronary heart disease, stroke and fatal stroke was 17.8 (12.98-19.99), 7 (6.7-7.2), 5.9 (4.0-6.6) and 4.7 (3.2-5.7), respectively. Simulated projections of metabolic syndrome patients predict serious life-threatening cardiovascular consequences in the representative cohort of patients in western India.

Investigation into the mechanism of action of essential oil of Pistacia integerrima for its antiasthmatic activity.

ETHNOPHARMACOLOGICAL RELEVANCE: Pistacia integerrima J.L. Stewart ex Brandis locally known as Karkatashringi is an important medicinal plant whose galls are valued in traditional medicine used in India for the treatment of asthma, chronic bronchitis, phthisis, diarrhea, fever, other ailments for the respiratory tract, and as antispasmodic, carminative, antiamoebic and anthelmintic. However, in vitro and in vivo investigations providing new insights into its pharmacological properties have not been thoroughly investigated yet. The present investigation aimed to elucidate the probable mechanism of antiasthmatic action of essential oil of Pistacia integerrima J.L. Stewart ex Brandis galls (EOPI). METHODS: EOPI was tested using in vitro studies such as antioxidant activity, mast cell degranulation, angiogenesis, isolated guinea pig ileum preparation and soyabean lipoxidase enzyme activity. In vivo studies included lipopolysaccharide-induced bronchial inflammation in rats and airway hyperresponsiveness in ovalbumin in sensitized guinea pigs using spirometry. RESULTS: EOPI (5-30 µg/ml) inhibits 5-lipoxidase enzyme activity with IC50 of 19.71 µg/ml and DPPH scavenging activity up to 100 µg/ml with maximum inhibition of 44.93 ± 2.53% at 100 µg/ml. Pre-treatment with EOPI inhibited erythropoietin-induced angiogenesis. It showed dose dependent (10, 30 and 100 µg/ml) anti-allergic activity by inhibiting compound 48/80 induced mast cell degranulation to an extent 19.08 ± 0.47%. The finding that essential oil induced inhibition of transient contraction of acetylcholine in calcium free medium, and relaxation of S-(-)-Bay 8644-precontracted isolated guinea pig ileum jointly suggests that the L-subtype Cav channel is involved in spasmolytic action of EOPI. Treatment with EOPI dose dependently (7.5, 15 and 30mg/kg i.p.) inhibited lipopolysaccharide-induced increase in total cell count, neutrophil count, nitrate-nitrite, total protein, albumin levels in bronchoalveolar fluid and myeloperoxidase levels in lung homogenates. Roflumilast was used as a standard. EOPI reduced the respiratory flow due to gasping in ovalbumin sensitized guinea pigs. CONCLUSION: The study demonstrates the effectiveness of essential oil of Pistacia integerrima J.L. Stewart ex Brandis galls in bronchial asthma possibly related to its ability to inhibit L-subtype Cav channel, mast cell stabilization, antioxidant, angiostatic and through inhibition of 5-lipoxygenase enzyme.

Exacerbation of alcohol-induced oxidative stress in rats by polyunsaturated Fatty acids and iron load.

The hypothesis that excessive intake of vegetable oil containing polyunsaturated fatty acids and iron load precipitate alcohol-induced liver damage was investigated in a rat model. In order to elucidate the mechanism underlying this synergism, the serum levels of iron, total protein, serum glutamate pyruvate transaminase, liver thiobarbituric acid reactive substances, and activities of antioxidant enzymes superoxide dismutase, catalase in liver of rats treated with alcohol, polyunsaturated fatty acids and iron per se and in combination were examined. Alcohol was fed to the rats at a level of 10-30% (blood alcohol was maintained between 150-350 mg/dl by using head space gas chromatography), polyunsaturated fatty acids at a level of 15% of diet and carbonyl iron 1.5-2% of diet per se and in combination to different groups for 30 days. Hepatotoxicity was assessed by measuring serum glutamate pyruvate transaminase, which was elevated and serum total protein, which was decreased significantly in rats fed with a combination of alcohol, polyunsaturated fatty acids and iron. It was also associated with increased lipid peroxidation and disruption of antioxidant defense in combination fed rats as compared to rats fed with alcohol or polyunsaturated fatty acids or iron. The present study revealed significant exacerbation of the alcohol-induced oxidative stress in presence of polyunsaturated fatty acids and iron.

Determination of actarit from human plasma for bioequivalence studies.

An analytical method based on high-performance liquid chromatography with ultraviolet detection (245 nm) was developed for the determination of actarit in human plasma. Coumarin was used as an internal standard. Chromatographic separation was achieved with a C8 column using a mobile phase of methanol: 1% acetic acid (50-50, v/v) with a flow rate of 1.0 ml/min. The calibration curve was linear over the range of 0.1-4.0 µg/ml (r(2) > 0.99) and the lower limit of quantification was 0.1 µg/ml. The method was validated for sensitivity, accuracy, precision, recovery and stability. The method was used to determine the concentration-time profiles of actarit in the plasma following oral administration of 100 mg actarit tablets.

Mechanism of vasorelaxant activity of a fraction of root extract of Sesamum indicum Linn.

The petroleum ether soluble fraction (SIPE) of the root extract of S. indicum was evaluated for the vasorelaxant activity using isolated rat aorta. SIPE up to 180 microg/ml concentration significantly inhibited phenylephrine- and KCl-induced contraction to the extent of 98.13 +/- 6.37 and 70.19 +/- 3.43% respectively in isolated rat aorta in a concentration dependent manner. The vasorelaxant activity was not blocked by propranolol (10 microM), atropine (1 microM) indomethacin (10 microM) and glibenclamide (10 microM). Influence of SIPE on phenylephrine-induced contractions in aortic preparations in absence of functional endothelium and on pre-incubating the tissue with L-NAME (300 microM) or methylene blue (10 microM) was also studied. SIPE at 180 microg/ml concentration could elicit partial relaxation in presence of L-NAME or methylene blue to the extent of 34.26 +/- 6.13 and 25.66 +/- 10.95% respectively. However, in absence of functional endothelium, SIPE exhibited little relaxation to the extent of 6.70 +/- 4.87%. These studies revealed that the vasorelaxant activity of SIPE was chiefly mediated through endothelium-dependent pathway.

A simplified HPLC method for quantification of torsemide from human plasma and its application to a bioequivalence study.

A simple, rapid and selective method was developed. The method was validated and found to be linear in the range of 100-4000 ng/ml. Chromatographic peaks were separated by means of a 5 mum, C18 silica column using acetonitrile and phosphate buffer (0.05 M) in proportion of 40:60 (pH 4.0) as a mobile phase. The retention time of torsemide was 5.00+/-0.20 min. The chromatograms showed good resolution and no interference from plasma. The mean recovery from human plasma was found to be above 82%. Both inter-day and intra-day accuracy and precision data showed good reproducibility. This method was applied to a single dose bioequivalence study. Log transformed values were compared by ANOVA followed by classical 90% confidence interval. Confidence limits for C(max), AUC(0-t) and AUC(0-inf) ranged from 98.6 to 102.8, 101.8 to 105.3 and 102.4 to 105.5 respectively. These results suggested that the analytical method was linear, precise and accurate. Test and reference product were found to be bioequivalent.

Mechanism of spasmolytic activity of a fraction of Sarcostemma brevistigma Wight.

The effect of chloroform soluble fraction (F-A) of twigs of Sarcostemma brevistigma on contractions induced by KCl, histamine, and acetylcholine in the isolated guinea pig ileum and taenia coli smooth muscles has been evaluated. F-A (19.5 microg/ml) significantly inhibited the contraction induced by 40 mM KCl to the extent of 87.6% in the isolated guinea pig ileum. In the isolated guinea pig ileum, F-A (64.3 and 59.2 microg/ml) significantly inhibited the contractions induced by acetylcholine and histamine to the extent of 85 and 83% respectively. In the isolated guinea pig taenia coli, F-A (65.2 microg/ml) significantly inhibited the contraction induced by 40 mM KCl to the extent of 96.0%. The inhibitory effect of F-A (40 microg/ml) on the isolated guinea pig taenia coli was reduced by Bay K 8644 (10(-6) M) to the extent of 61.6 from 73.6%. These results suggest that the F-A may exhibit smooth muscle relaxant activity by blocking the Ca2+ channels.

Evaluation of sesquiterpene lactone fraction of Saussurea lappa on transudative, exudative and proliferative phases of inflammation.

The sesquiterpene lactone fraction of Saussurea lappa roots was evaluated for its effect on the transudative, exudative and proliferative phases of inflammation using the cotton pellet granuloma assay in rats. The fraction (25-100 mg/kg, p.o.) showed significant dose-dependent inhibition of the increase in wet weight of the cotton pellet at 3 h (transudative phase), leakage of dye from the bloodstream around granuloma at 24 h (exudative phase) and increase in dry weight of the cotton pellet on day 6 (proliferative phase). It significantly lowered the elevated biochemical parameters such as alkaline phosphatase, acid phosphatase, gamma-glutamyltranspeptidase and significantly elevated the lowered albumin concentration in serum. The studies suggest that the antiinflammatory activity of the sesquiterpene lactone fraction of S. lappa may, in part, be due to stabilization of lysosomal membranes and an antiproliferative effect.

Studies on the immunomodulatory activity of flavonoidal fraction of Tephrosia purpurea.

The flavonoid fraction of Tephrosia purpurea (FFTP) was studied for its effect on cellular and humoral functions and on macrophage phagocytosis in mice. Oral administration of FFTP (10-40 mg/kg) significantly inhibited sheep red blood cells (SRBC)-induced delayed-type hypersensitivity reactions. It also produced a significant, dose-related decrease in sheep erythrocyte-specific haemagglutination antibody titre. However, the fraction failed to show a significant change in the macrophage phagocytic activity. The results obtained indicate the ability of the flavonoidal fraction of T. purpurea to modulate both the cell-mediated and the humoral components of the immune system.

Investigations into the immunomodulatory activity of Argyreia speciosa.

The objective of the present study was to investigate the immunomodulatory activity of Argyreia speciosa on cellular and humoral immunity. Oral administration of the ethanolic extract of A. speciosa root (ASEE), at the doses of 50, 100 and 200 mg/kg in mice, dose-dependently potentiated the delayed-type hypersensitivity reaction induced both by sheep red blood cells (SRBC) and oxazolone. It significantly enhanced the production of circulating antibody titre in mice in response to SRBC. ASEE failed to show any effect on macrophage phagocytosis. Chronic administration of ASEE significantly ameliorated the total white blood cell count and also restored the myelosuppressive effects induced by cyclophosphamide. The present investigation reveals that ASEE possesses immunomodulatory activity.

Preliminary evaluation of anti-inflammatory and anti-arthritic activity of S. lappa, A. speciosa and A. aspera.

Saussurea lappa, Argyreia speciosa and Achyranthes aspera are well known Indian medicinal plants used in the indigenous systems of medicine for the treatment of inflammatory conditions. The ethanolic extracts of the plants at the doses of 50, 100 and 200 mg/kg, p.o. were screened for their effect on acute and chronic inflammation induced in mice and rats. S. lappa and A. speciosa were found to significantly inhibit paw edema induced by carrageenan and Freund's complete adjuvant and to prevent accumulation of inflammatory cells in carrageenan-induced peritonitis at doses of 50-200 mg/kg. A. aspera inhibited these inflammatory responses at doses of 100-200 mg/kg. The studies reveal that the ethanolic extracts of S. lappa, A. speciosa and A. aspera possess anti-inflammatory and anti-arthritic activity and support the rationale behind the traditional use of these plants in inflammatory conditions.

Influence of ethanolic extract of Tephrosia purpurea Linn. on mast cells and erythrocytes membrane integrity.

The ethanolic extract of T. purpurea Linn. was studied for its in vitro effect on rat mast cell degranulation and erythrocyte membrane integrity in vitro. The extract in concentration of 25-200 microg/ml showed a dose-dependant inhibition of rat mast cell degranulation induded by compound 48/80 and egg albumin. T. purpurea extract was found to inhibit haemolysis of erythrocytes induced by hypotonic solution but accelerated haemolysis induced by heat at a concentration of 100 microg/ml. The studies reveal that the ethanolic extract of T. purpurea may inhibit degranulation of mast cells by a mechanism other than membrane stabilization.

Studies on the immunomodulatory effects of Boerhaavia diffusa alkaloidal fraction.

The alkaloidal fraction of Boerhaavia diffusa was studied for its effect on cellular and humoral functions in mice. Oral administration of the fraction (25-100 mg/kg) significantly inhibited SRBC-induced delayed hypersensitivity reactions in mice. However, the inhibition was observed only during post-immunisation drug treatment, while no effect during pre-immunisation drug treatment was observed. A significant dose-related increase in antibody titre was observed during pre- and post-immunisation treatment. The alkaloidal fraction failed to show any blastogenic responsiveness of murine splenocytes to Concanvalin A (Con A) and lipopolysaccharide (LPS). Similarly, it did not display any mitogenic activity. Thus, the present study has shown the in vivo immunostimulatory activity of B. diffusa alkaloidal fraction without an in vitro effect.

Studies on the anti-inflammatory and analgesic activity of Cedrus deodara (Roxb.) Loud. wood oil.

The volatile oil extracted by steam distillation of the wood of Cedrus deodara was examined for its oral anti-inflammatory and analgesic activity at the doses of 50 and 100 mg/kg body weight. It produced significant inhibition of carrageenan-induced rat paw edema and of both exudative-proliferative and chronic phases of inflammation in adjuvant arthritic rats at doses of 50 and 100 mg/kg body weight. The oil at both tested doses was found to possess analgesic activity against acetic acid-induced writhing and hot plate reaction in mice.

Mast cell stabilizing and lipoxygenase inhibitory activity of Cedrus deodara (Roxb.) Loud. wood oil.

Volatile oil of C. deodara, administered orally at the doses of 50, 100 and 200 mg/kg body weight, significantly inhibited the pedal edema induced by compound 48/80 in rats. The oil significantly inhibited compound 48/80 induced degranulation of isolated rat peritoneal mast cells at concentrations ranging from 25-200 micrograms/ml. C. deodara wood oil also significantly inhibited the enzyme lipoxygenase at a concentration of 200 micrograms/ml. Thus, the anti-inflammatory activity of C. deodara wood oil could be attributed to its mast cell stabilizing activity and the inhibition of leukotriene synthesis.

Antiallergic potential of furosemide.

Recent reports have indicated the effectiveness of furosemide in inhibiting responses to inhaled allergen and in treating allergic conjunctivitis. In the present study furosemide was tested for its antiallergic potential using compound 48/80 induced paw edema and in vitro mast cell degranulation. Furosemide was found to significantly inhibit compound 48/80 paw edema and compound 48/80 induced histamine release. Furosemide was also found to inhibit histamine release during passive peritoneal anaphylaxis in rats. The results suggest that furosemide may be inhibiting the release of mediators of anaphylaxis from the mast cells.

Studies on the mechanism of action of Semecarpus anacardium in rheumatoid arthritis.

Semecarpus anacardium nuts are used for variety of disorders in Ayurveda. A chloroform extract of the nut significantly reduced acute carrageenan-induced paw oedema in rats and was active against the secondary lesions of adjuvant-induced arthritis. Delayed hypersensitivity induced in mice by sheep red blood cells as an antigen was potentiated by the extract.