Mundkinajeddu Deepak

Antiinflammatory activity and chemical composition of extracts of Verbena officinalis

In an attempt to locate the biologically active fraction(s) of the plant Verbena officinalis Linn. (Verbenaceae), a preliminary screening of successive petroleum ether, chloroform and methanol extracts of aerial parts for antiinflammatory activity using carrageenan paw oedema model was carried out. All three extracts were found to exhibit antiinflammatory activity with the chloroform extract being the most active. Chemical investigations of petroleum ether and chloroform extracts led to the isolation of β‐sitosterol, ursolic acid, oleanolic acid, 3‐epiursolic acid, 3‐epioleanolic acid, and minor triterpenoids of derivatives of ursolic acid and oleanolic acids. Chromatographic purification of the methanol extract yielded two iridoid glucosides, verbenalin and hastatoside, a phenylpropanoid glycoside, verbascoside and β‐sitosterol‐D‐glucoside. Copyright

Evaluation of the genotoxic potential and acute oral toxicity of standardized extract of Andrographis paniculata (KalmCold).

Andrographis paniculata is used in the traditional medicine for cold and influenza remedy. The main endeavor in this study was to assess the genotoxicity of the standardized extract of A. paniculata (KalmCold) through three different in vitro tests: Ames, chromosome aberration (CA), and micronucleus (MN). Ames test was performed at 5000 microg/ml, 1581 microg/ml, 500 microg/ml, 158 microg/ml, 50 microg/ml, 16 microg/ml, while the clastogenicity tests were performed at 80 microg/ml, 26.6 microg/ml, 8.8 microg/ml for short-term treatment without S9; 345 microg/ml, 115 microg/ml, 38.3 microg/ml for short-term treatment with S9; and 46 microg/ml, 15.3 microg/ml, 5.1 microg/ml for long-term without S9 using DMSO as a vehicle control. Results of Ames test confirmed that KalmCold did not induce mutations both in the presence and absence of S9 in Salmonella typhimurium mutant strains TA98 and TAMix. In CA and MN, KalmCold did not induce clastogenicity in CHO-K1 cells in vitro. Based on our results, it is evident that KalmCold is genotoxically safe. Additionally in acute oral toxicity study, female rats were treated at 5000 mg/kg of KalmCold and observed for signs of toxicity for 14 days. KalmCold did not produce any treatment-related toxic effects in rats.

Tribulosin and β-sitosterol-D-glucoside, the anthelmintic principles of Tribulus terrestris

Successive extracts of Tribulus terrestris prepared using petroleum ether, chloroform, 50% methanol and water were tested for anthelmintic activity in-vitro using the nematode Caenorhabditis elegans. The activity could be detected only in 50% methanol extract which on further bioactivity guided fractionation and chromatographic separation yielded a spirostanol type saponin, tribulosin and beta-sitosterol-D-glucoside. Both the compounds exhibited anthelmintic activity with ED50 of 76.25 and 82.50 microg/ml respectively.

Dual inhibitory effect of Glycyrrhiza glabra (GutGard™) on COX and LOX products.

Glycyrrhiza glabra and its phytoconstituents have been known to possess widespread pharmacological properties as an anti-inflammatory, anti-viral, antitumour and hepatoprotective drug. In this study, we examined the inhibitory potential of extract of G. glabra (GutGard™) root and its phytoconstituents (glabridin, glycyrrhizin, and isoliquiritigenin) on both cyclooxygenase (COX) and lipoxygenase (LOX) products in order to understand the mechanism of its anti-inflammatory action. Inhibitory effect of GutGard™ and its phytoconstituents on lipopolysaccharide (LPS) induced prostaglandin E(2) (PGE(2)), calcimycin (A23187) induced thromboxane (TXB(2)), and leukotriene (LTB(4)) release was studied using murine macrophages (J774A.1) and human neutrophil (HL-60) cells. Results revealed that, G. glabra and glabridin significantly inhibited PGE(2), TXB(2) (COX) and LTB(4) (LOX), while, isoliquiritigenin exerted inhibitory effect only against COX products but failed to suppress LOX product. However, glycyrrhizin at the tested concentrations failed to exhibit inhibitory effect on both COX and LOX products. Here, we report for the first time that G. glabra (almost devoid of glycyrrhizin) exhibits anti-inflammatory property likely through the inhibition of PGE(2), TXB(2) and LTB(4) in mammalian cell assay system, which could be influenced in part by glabridin and isoliquiritigenin.

In vitro anti-Helicobacter pylori activity of a flavonoid rich extract of Glycyrrhiza glabra and its probable mechanisms of action

ETHNOPHARMACOLOGICAL RELEVANCE Glycyrrhiza glabra Linn. is regarded useful for peptic ulcer in traditional systems of medicine in India and Helicobacter pylori has been considered as one of the causative factors for peptic ulcer. Aim of the present study is to evaluate the anti-Helicobacter pylori action of GutGard(®), a flavonoid rich extract of Glycyrrhiza glabra and further to elucidate the possible mechanisms of its anti-Helicobacter pylori action. MATERIALS AND METHODS Agar dilution and microbroth dilution methods were used to determine the minimum inhibitory concentration of GutGard(®) against Helicobacter pylori. Protein synthesis, DNA gyrase, dihydrofolate reductase assays and anti-adhesion assay in human gastric mucosal cell line were performed to understand the mechanisms of anti-Helicobacter pylori activity of GutGard(®). RESULTS GutGard(®) exhibited anti-Helicobacter pylori activity in both agar dilution and microbroth dilution methods. Glabridin, the major flavonoid present in GutGard(®) exhibited superior activity against Helicobacter pylori while glycyrrhizin did not show activity even at 250 μg/ml concentration. In protein synthesis assay, GutGard(®) showed a significant time dependent inhibition as witnessed by the reduction in (35)S methionine incorporation into Helicobacter pylori ATCC 700392 strain. Additionally, GutGard(®) showed a potent inhibitory effect on DNA gyrase and dihydrofolate reductase with IC(50) value of 4.40 μg/ml and 3.33 μg/ml respectively. However, the extract did not show significant inhibition on the adhesion of Helicobacter pylori to human gastric mucosal cell line at the tested concentrations. CONCLUSION The present study shows that, GutGard(®) acts against Helicobacter pylori possibly by inhibiting protein synthesis, DNA gyrase and dihydrofolate reductase.

In vitro Safety Evaluation and Anticlastogenic Effect of BacoMind™ on Human Lymphocytes

OBJECTIVE BacoMind (BM) is a standardized extract of Bacopa monnieri, which belongs to the family Scrophulariaceae and is a creeping annual plant found throughout the Indian subcontinent. It has been used by Ayurvedic medicinal practitioners in India for almost 3000 years and is classified as a medharasayana, a substance which improves memory and intellect. With the widespread traditional use as well as scientific validation of Bacopa monnieri for nootropic activity, a bioactive-rich unique phytochemical composition-BacoMind was developed from B. monnieri for use as a cognition and memory enhancing agent. The present study aimed to investigate the in vitro toxicity of this formulation of BacoMind on human lymphocytes and to rule out its possible contribution to mutagenicity. METHODS In the present investigation the active ingredients present in BM were identified and quantified by high performance liquid chromatography (HPLC) and high performance thin-layer chromatography (HPTLC). Antioxidant and anticlastogenic properties of BM were studied in vitro with and without metabolic activation. Doses of BM were chosen on the basis of mitotic index (MI) and cytokinesis-block proliferation index (CBPI). Clastogenicity assays were performed at 31.2 microg/mL, 62.5 microg/mL, and 125 microg/mL, while the Salmonella reverse mutation assay (Ames test) was performed at doses of 61.72, 185.18, 555.55, 1666.67, and 5000.00 microg/plate. RESULTS HPLC and HPTLC analysis of BM revealed the presence of bacoside A3, bacopaside I, bacopaside II, jujubogenin isomer of bacopasaponin C, bacosine, luteolin, apigenin, bacosine, and beta-sitosterol D glucoside. BM demonstrated significant antioxidant activity. The number of chromosomal aberrations and the frequency of micronuclei induced by BM were not statistically significant up to a dose of 62.5 microg/mL. A subsequent dose of 125 microg/mL prior to metabolic activation induced mild clastogenicity, but it was found to be biologically insignificant as this effect was not seen post metabolic activation. BM also demonstrated a dose-dependent protection against the clastogens used in this study using the above tests for clastogenicity. Maximum protection was observed in presence of metabolic activation. Moreover, BM demonstrated no mutagenic effect on the tested strains, as observed in the Ames test. CONCLUSION BM protected human lymphocytes against various clastogens. BM also exhibited high antioxidant activity which might be responsible for the observed protective effects against the clastogens since the used clastogens are known to induce their clastogenic effects via production of oxidative radicals.

3α,24-dihydroxy-urs-12-en-28-oic acid from verbena officinalis

A new triterpenoid, 3α,24-dihydroxy-urs-12-en-28-oic acid, has been isolated from the bioactive chloroform extract of aerial parts of Verbena officinalis along with 3α,24-dihydroxy-olean-12-en-28-oic acid and ursolic acid.

Reproductive and fertility effects of an extract of Andrographis paniculata in male Wistar rats.

The possible effect of extract of Andrographis paniculata Nees (A paniculata) standardized to ≥10% andrographolide, the main bioactive component, on male fertility in albino Wistar rats was evaluated, by orally administering 0, 20, 200, and 1000 mg/kg of body weight per day, for 65 days prior to mating and 21 days during mating. The treated groups showed no signs of dose-dependent toxicity. The body weight gain and feed consumption were not affected at any of the dose levels. The testosterone levels and fertility indices in treatment groups were found to be comparable with that of the control indicating no effect on fertility. Total sperm count and sperm motility were not affected. The testes and epididymides did not show any gross and histopathological changes. Based on these findings, it can be concluded that the no-observed adverse effect level of extract of A paniculata (≥10% andrographolide) was found to be more than 1000 mg/kg per day.

Bioactive caffeic acid esters from Glycyrrhiza glabra

Thin layer chromatography bioautography (using DPPH spray reagent) guided fractionation of Glycyrrhiza glabra led to the isolation of two caffeic acid derivative esters, viz. eicosanyl caffeate (1) and docosyl caffeate (2). The two compounds exhibited potent elastase inhibitory activity, with IC50 values of 0.99 µg mL−1 and 1.4 µg mL−1 for 1 and 2, respectively. The compounds also showed moderate antioxidant activity in DPPH and ABTS scavenging assays. The results indicate a possible role of caffeic acid derivatives, in addition to flavonoids in the anti-ulcer properties of G. glabra.

Quantitative determination of the major constituents of Verbena officinalis using high performance thin layer chromatography and high pressure liquid chromatography

A pentacyclic triterpenoid, ursolic acid (1), two iridoid glucosides, verbenalin (2) and hastatoside (3), and a phenylpropanoid glycoside, verbascoside (4), were isolated from Verbena officinalis Linn. (Verbenaceae), a plant listed in the Chinese Pharmacopoeia and the British Herbal Pharmacopoeia. A procedure for the optimised extraction of these constituents for quantitative estimations has been established. An HPTLC method was employed for the determination of 1 using Liebermann Burchard reagent. A reversed-phase HPLC system with photodiode array detector was used to resolve compounds 2, 3 and 4 in the methanol extracts of different parts of the plant. Tender parts of the plant were rich in all of these constituents (0.24–0.34%, w/w) while the roots, which are not official in the Pharmacopoeias, contained a maximum amount (0.32%, w/w) of the bioactive verbascoside (4). The assay methods described are simple, rapid and accurate, and may form part of future drug authentication protocols. Copyright