Assessment of acute, sub-acute, chronic and genotoxicity of polyherbal formulation DCD-684 in mice.

Digas colic drops (DCD-684) a polyherbal formulation containing Carum carvi, Foeniculum vulgare, Mentha arvensis, Mentha piperita and Zingiber officinale is widely used in Pakistan against gastrointestinal ailments including infantile colic. The DCD-684 (0.03-3ml/kg.bw) administered orally in acute (7-days) and sub-acute toxicity (14-days) tests, displayed neither mortality nor toxicological changes in physical, behavioral, biochemical and histopathological parameters. In chronic study (90-days), DCD-684 (0.3-12ml/kg.bw) also revealed no changes. However, at 18 and 36 ml/kg.bw, liver demonstrated mild inflammation correlating with raised aspartate transaminase (AST), alkaline phosphatase (ALP) and alpha fetoprotein (AFP) levels. Increased levels of urea and inflamed renal parenchyma indicated mild nephro-toxicity with high alanine aminotransferase (ALT) at 36ml/kg.bw. The LD50 of DCD-684 in mice was 27.5 ml/kg.bw. In hepatocytes at 36ml/kg.bw, elevated mRNA expression of pro-inflammatory chemokines and cytokines were evident. DCD-684 neither damaged DNA nor induced cytotoxicity in micronucleus assay. In conclusion, polyherbal DCD-684 caused neither hepatic, renal, genotoxicity nor any undesirable effect in mice. Higher doses administered for 90 days showed mild toxic effects with no sign of necrosis, fibrosis or genotoxicity. Thus, in mice DCD-684 demonstrated a wide margin of safety to be used for the relief of infantile colic.

Antispasmodic activity and mechanism of action of polyherbal formulation DCD-684 on rabbit jejunum.

Digas colic drops (DCD-684) is a polyherbal formulation containing decoctions of five medicinal plants namely Carum carvi L., Foeniculum vulgare Mill, Mentha arvensis L., Mentha piperita L. and Zingiber officinale Roscoe. These plants have been extensively used in traditional medicine for the treatment of various gastrointestinal diseases including abdominal colic. This study was conducted to determine the spasmolytic effect of DCD-684 (100% v/v) and its individual plant components on isolated rabbit jejunum (in vitro) and their possible mechanism of action. The effects were evaluated on spontaneous and pre-contracted tissues using KCl (80mM) and other contractile agonists including acetylcholine (0.3µM), carbamylcholine (0.3µM), serotonin (10 µM) and histamine (100µM) in the presence and absence of DCD-684. The various concentrations of DCD-684 (0.1-3% v/v) demonstrated spasmolytic effects on both spontaneous (IC50=0.75%) and KCl-induced contractions (IC50=1.6%), respectively. It also inhibited the contractions induced by acetylcholine (IC50=0.45%), carbamylcholine (IC50=0.95%), serotonin (IC50=0.95%) and histamine (IC50=0.87%). The DCD-684 exhibited synergistic effect due to its five plant components suggesting that spasmolytic cascade is probably governed by muscarinic and/or nicotinic receptors, serotonergic histaminergic, as well as calcium channel blocking mechanisms. Thereby, providing the pharmacological basis of its therapeutic use in the gastrointestinal motility disorders and related inflammatory ailments.

Gamma-linolenic acid ameliorated glycation-induced memory impairment in rats.

CONTEXT: γ-Linolenic acid (GLA) is an important constituent of anti-ageing supplements. OBJECTIVE: The current study investigates the anti-ageing effect of GLA in Sprague-Dawley rats. MATERIALS AND METHODS: GLA (0.1, 0.2, 0.4, 2, 10, 20 and 24 µM) was initially evaluated for its effect on the formation of advanced glycation end products (AGEs) in vitro. For in vivo assessment (1, 5 or 15 mg/kg), the rat model of accelerated ageing was developed using d-fructose (1000 mg/kg (i.p.) plus 10% in drinking water for 40 days). Morris water maze was used to evaluate impairment in learning and memory. The blood of treated animals was used to measure glycosylated haemoglobin (HbA1c) levels. The interaction of GLA with active residues of receptor of AGE (RAGE) was analyzed using AutoDock Vina. RESULTS: Our data showed that GLA inhibited the production of AGEs (IC50 = 1.12 ± 0.05 µM). However, this effect was more significant at lower tested doses. A similar pattern was also observed in in vivo experiments, where the effect of fructose was reversed by GLA only at lowest tested dose of 1 mg/kg. The HbA1c levels also revealed significant reduction at lower doses (1 and 5 mg/kg). The in silico data exhibited promising interaction of GLA with active residues (Try72, Arg77 and Gln67) of RAGE. CONCLUSION: The GLA, at lower doses, possesses therapeutic potential against glycation-induced memory decline.

Metabolic products of soluble epoxide hydrolase are essential for monocyte chemotaxis to MCP-1 in vitro and in vivo.

Monocyte chemoattractant protein-1 (MCP-1)-induced monocyte chemotaxis is a major event in inflammatory disease. Our prior studies have demonstrated that MCP-1-dependent chemotaxis requires release of arachidonic acid (AA) by activated cytosolic phospholipase A(2) (cPLA(2)). Here we investigated the involvement of AA metabolites in chemotaxis. Neither cyclooxygenase nor lipoxygenase pathways were required, whereas pharmacologic inhibitors of both the cytochrome-P450 (CYP) and the soluble epoxide hydrolase (sEH) pathways blocked monocyte chemotaxis to MCP-1. To verify specificity, we demonstrated that the CYP and sEH products epoxyeiscosatrienoic acids (EETs) and dihydroxyeicosatrienoic acids (DHETs), respectively, restored chemotaxis in the presence of the inhibitors, indicating that sEH-derived products are essential for MCP-1-driven chemotaxis. Importantly, DHETs also rescued chemotaxis in cPLA(2)-deficient monocytes and monocytes with blocked Erk1/2 activity, because Erk controls cPLA(2) activation. The in vitro findings regarding the involvement of CYP/sEH pathways were further validated in vivo using two complementary approaches measuring MCP-1-dependent chemotaxis in mice. These observations reveal the importance of sEH in MCP-1-regulated monocyte chemotaxis and may explain the observed therapeutic value of sEH inhibitors in treatment of inflammatory diseases, cardiovascular diseases, pain, and even carcinogenesis. Their effectiveness, often attributed to increasing EET levels, is probably influenced by the impairment of DHET formation and inhibition of chemotaxis.

Evaluation of antinociceptive effect of Aegiceras corniculatum stems extracts and its possible mechanism of action in rodents.

ETHNOPHARMACOLOGICAL RELEVANCE: Aegiceras corniculatum (Linn.) Blanco is used in various traditional medicinal system(s) for the treatment of rheumatism, painful arthritis and inflammation. Therefore, the pharmacological studies of its antinociceptive effect was undertaken to validate its traditional use. MATERIALS AND METHODS: n-Hexane, ethyl acetate and methanol extract(s) derived from Aegiceras corniculatum (stems) were studied using various nociceptive model(s) induced chemically or thermally in mice including acetic acid-induced writhing, formalin-induced paw licking and hot plate test. RESULTS: In acetic acid-induced writhing test, plant extracts dose dependently decreased the writhing numbers. The methanolic extract (1-10mg/kg, i.p. in mice) of the plant was more potent than acetaminophen and acetyl salicylic acid, with an IC(50) of 4.2 ± 0.99 mg/kg. Moreover, the time of nociceptive behaviors induced by intraplantar formalin injection was also suppressed during 1st and 2nd phases in the presence of ethyl acetate extract whereas, n-hexane and methanolic extracts inhibited the paw licking in mice during the 1st (IC(50) 12 ± 0.76 mg/kg) and 2nd phases (IC(50) 3.8 ± 0.55 mg/kg). Naloxone, ß-funaltrexamine, and naltrindole antagonized the n-hexane extract-induced antinociception in the first phase of formalin test indicating its non-selective analgesic response via opioid receptor(s). However, ethyl acetate extract was devoid of any opioid action. Additionally, these extracts significantly inhibited the pain stimulation in hot plate test. Withdrawal syndrome of morphine dependence was also diminished in the presence of plant extracts via potentiation of GABAergic system. CONCLUSION: These results suggested that Aegiceras corniculatum extract(s) possesses analgesic properties and acts on the central nervous system, thereby suppressing the inflammatory pain justifying its folklore use.

Aegiceras corniculatum extract suppresses initial and late phases of inflammation in rat paw and attenuates the production of eicosanoids in rat neutrophils and human platelets.

AIM OF THE STUDY: The present study is designed to explore the anti-inflammatory potential of Aegiceras corniculatum Linn. Blanco stems extracts and their mechanism of action against various pro-inflammatory mediators and to validate its traditional use against inflammatory diseases. MATERIALS AND METHODS: Rat paw edema and peritonitis models were employed for in vivo studies. For in vitro studies human platelets and rat neutrophils were stimulated with Ca(2+)-ionophore A23187 leading to the production of various pro-inflammatory metabolites, i.e., 12-HTT, 12-HETE and LTB(4) and 5-HETE which were quantified by HPLC. RESULTS: The highly polar methanol extract (100mg/kg) caused approximately 90% reduction in the carrageenan- and prostaglandin E2-induced paw edema in rats. It also caused the inhibition of cycloxygenase-1 metabolite, 12-HHT (IC(50) 41.1+/-1.5microg/ml) with a concomitant rise in 12-lipoxygenase metabolite, 12-HETE in A23187 stimulated human platelets. Conversely, the non-polar hexane extract attenuated (IC(50) 0.36+/-0.12microg/ml) 12-HETE formation with a parallel rise in 12-HHT, thereby displaying a selectivity towards 12-lipoxygenase. Non-polar hexane extract also antagonized the production of 5-lipoxygenase metabolites, i.e., leukotriene B(4) and 5-HETE in the rat neutrophils. Furthermore, ethyl acetate extract inhibited both COX and 5-LOX with a marked decline in the production of 12-HHT (IC(50) 0.08+/-0.002microg/ml) and LTB(4) (IC(50) 0.86+/-0.03microg/ml), respectively. The anti-inflammatory effect of hexane and ethyl acetate extracts was also reflected by the diminution of carrageenan-induced cell infiltration in rat peritoneum. Additionally, plant extracts caused approximately 60% suppression in dextran-induced paw edema implying that they also ameliorate histamine and serotonin release. CONCLUSION: Hexane, ethyl acetate and methanol extracts derived from Aegiceras corniculatum possess significant anti-inflammatory activity via multiple mechanisms and validate their traditional use against inflammation-related diseases.

A study on antioxidant, free radical scavenging, anti-inflammatory and hepatoprotective actions of Aegiceras corniculatum (stem) extracts.

AIM OF THE STUDY: The present study was conducted to evaluate the antioxidant, anti-inflammatory and hepatoprotective potential of Aegiceras corniculatum Linn. Blanco (Aegicerataceae). METHODS AND RESULTS: The n-hexane, ethyl acetate and methanol extracts, derived from Aegiceras corniculatum stems, scavenged superoxide anions (O2*) and hydroxyl radicals (*OH) in nitro blue tetrazolium reduction and deoxyribose degradation assays, respectively. All the extracts inhibited the process of lipid peroxidation at its initiation step. Additionally, in rat liver microsomes n-hexane and ethyl acetate extracts also caused termination of radical chain reaction supporting their scavenging action towards lipid peroxy radicals (LOO*). Moreover, increased production of O2* in human neutrophils, stimulated by phorbol-12-myristate-13-acetate (PMA) and/or opsonized zymosan were also suppressed (IC50 approximately 3-20 microg/mL). Thereby, revealing the ability of plant extracts to antagonize the oxidative stress via interference with NADPH oxidase metabolic pathway. These in vitro results coincide with the reduction in the glucose oxidase-induced paw edema in mice in the presence of ethyl acetate and methanol extracts (10, 50, and 100mg/kg, i.p.). Plant extracts (250, 500 and 1000 mg/kg, p.o.) also significantly protected the carbon tetrachloride (CCl4)-induced oxidative tissue injury in rat liver. This was reflected by a approximately 60% decline in the levels of serum aminotransferase enzymes. CONCLUSION: Aegiceras corniculatum extracts found to possess pronounced antioxidant effect that may be at least in part related to its anti-inflammatory and hepatoprotective activities. This study provides a scientific basis for the ethnomedical claims that Aegiceras corniculatum is effective against inflammation and liver injury.

Analgesic and antioxidant activity of mangiferin and its derivatives: the structure activity relationship.

Mangiferin, 2-beta-D-glucopyranosyl-1,3,6,7-tetrahydroxy-9H-xanthen-9-one, obtained directly from methanolic extracts of Bombax ceiba leaves in substantial amounts demonstrated strong antioxidant activity (EC(50) 5.8+/-0.96 mug/ml or 13.74 muM) using DPPH assay comparable to rutin, commonly used as antioxidant for medical purposes. The acetyl and cinnamoyl derivatives were found to be less active than mangiferin whereas, methyl and 3,6,7-trimethylether tetraacetate derivatives were inactive implying that for antioxidant activity, free hydroxyl groups and catechol moiety are essential. Moreover, mangiferin showed hepatoprotective activity against carbon tetrachloride induced liver injury further supporting the free radical scavenging property in the in vivo system. Additionally, plant extracts and mangiferin failed to exhibit acute anti-inflammatory activity whereas, it displayed significant analgesic effect in acetic acid-induced writhing and hot plate tests in mice. Using naloxone, it was revealed that plant extracts induced analgesia was independent of opioid receptor, whereas, mangiferin demonstrated significant interaction with it at peripheral site with a slight contribution at the neuronal level.