K. Pandima Devi

Eugenol (an essential oil of clove) acts as an antibacterial agent against Salmonella typhi by disrupting the cellular membrane.

AIM OF THE STUDY To evaluate the antibacterial activity of eugenol and its mechanism of bactericidal action against Salmonella typhi. MATERIALS AND METHODS The antibacterial activity was checked by disc-diffusion method, MIC, MBC, time course assay and pH sensitivity assay. The chemo-attractant property of eugenol was verified by chemotaxis assay. The mode of action of eugenol was determined by crystal violet assay, measurement of release of 260 nm absorbing material, SDS-PAGE, FT-IR spectroscopy, AFM and SEM. RESULTS Treatment with eugenol at their MIC (0.0125%) and MBC (0.025%) reduced the viability and resulted in complete inhibition of the organism. Eugenol inactivated Salmonella typhi within 60 min exposure. The chemo-attractant property of eugenol combined with the observed high antibacterial activity at alkaline pH favors the fact that the compound can work more efficiently when given in vivo. Eugenol increased the permeability of the membrane, as evidenced by crystal violet assay. The measurement of release of 260 nm absorbing intracellular materials, SDS-PAGE, SEM and AFM analysis confirmed the disruptive action of eugenol on cytoplasmic membrane. The deformation of macromolecules in the membrane, upon treatment with eugenol was verified by FT-IR spectroscopy. CONCLUSION The results suggest that the antibacterial activity of eugenol against Salmonella typhi is due to the interaction of eugenol on bacterial cell membrane.

Silymarin protects PBMC against B(a)P induced toxicity by replenishing redox status and modulating glutathione metabolizing enzymes - an in vitro study.

PAHs are a ubiquitous class of environmental contaminants that have a large number of hazardous consequences on human health. An important prototype of PAHs, B(a)P, is notable for being the first chemical carcinogen to be discovered and the one classified by EPA as a probable human carcinogen. It undergoes metabolic activation to QD, which generate ROS by redox cycling system in the body and oxidatively damage the macromolecules. Hence, a variety of antioxidants have been tested as possible protectors against B(a)P toxicity. Silymarin is one such compound, which has high human acceptance, used clinically and consumed as dietary supplement around the world for its strong anti-oxidant efficacy. Silymarin was employed as an alternative approach for treating B(a)P induced damage and oxidative stress in PBMC, with an emphasis to provide the molecular basis for the effect of silymarin against B(a)P induced toxicity. PBMC cells exposed to either benzopyrene (1 microM) or silymarin (2.4 mg/ml) or both was monitored for toxicity by assessing LPO, PO, redox status (GSH/GSSG ratio), glutathione metabolizing enzymes GR and GPx and antioxidant enzymes CAT and SOD. This study also investigated the protective effect of silymarin against B(a)P induced biochemical alteration at the molecular level by FT-IR spectroscopy. Our findings were quite striking that silymarin possesses substantial protective effect against B(a)P induced oxidative stress and biochemical changes by restoring redox status, modulating glutathione metabolizing enzymes, hindering the formation of protein oxidation products, inhibiting LPO and further reducing ROS mediated damages by changing the level of antioxidant enzymes. The results suggest that silymarin exhibits multiple protections and it should be considered as a potential protective agent for environmental contaminant induced immunotoxicity.

In vitro antioxidant and anti-cholinesterase activities of Rhizophora mucronata

Abstract Context: Rhizophora mucronata Lam. (Rhizophoraceae), commonly known as Asiatic mangrove, has been used traditionally among Asian countries as folk medicine. Objective: This study investigates the cholinesterase inhibitory potential and antioxidant activities of R. mucronata. Materials and method: Rhizophora mucronata leaves were successively extracted using solvents of varying polarity and a dosage of 100–500 µg/ml were used for each assay. Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory activities were assessed according to the method of Ellman. In vitro antioxidant activity was assessed using free radical scavenging, reducing power, and metal-chelating activity (duration – 3 months). Total phenolic and flavonoid content were quantified spectrophotometrically. Compound characterization was done using column chromatography, NMR, FTIR, and LC-MS analysis. Results: Methanolic leaf extract (500 µg/ml) exhibited the highest inhibitory activity against AChE (92.73 ± 0.54%) and BuChE (98.98 ± 0.17%), with an IC50 value of 59.31 ± 0.35 and 51.72 ± 0.33 µg/ml, respectively. Among the different solvent extracts, methanolic extract exhibited the highest antioxidant activity with an IC50 value of 47.39 ± 0.43, 401.45 ± 18.52, 80.23 ± 0.70, and 316.47 ± 3.56 µg/ml for DPPH, hydroxyl, nitric oxide radical, and hydrogen peroxide, respectively. Total polyphenolic and flavonoid contents in methanolic extract were observed to be 598.13 ± 1.85 µg of gallic acid equivalent and 48.85 ± 0.70 μg of rutin equivalent/mg of extract. Compound characterization illustrated (+)-catechin as the bioactive compound responsible for cholinesterase inhibitory and antioxidant activities. Conclusion: The presence of rich source of flavonoids, in particular catechin, might be responsible for its cholinesterase inhibitory and antioxidant activities.

Silymarin prevents benzo(a)pyrene-induced toxicity in Wistar rats by modulating xenobiotic-metabolizing enzymes:

Benzo(a)pyrene (B(a)P), which is commonly used as an indicator species for polycyclic aromatic hydrocarbon (PAH) contamination, has a large number of hazardous consequences on human health. In the presence of the enzyme cytochrome-P-450 1A1 (CYP1A1), it undergoes metabolic activation to form reactive intermediates that are capable of inducing mutagenic, cytotoxic, teratogenic and carcinogenic effects in various species and tissues. Research within the last few years has shown that flavonoids exhibit chemopreventive effect against these toxins. In the present study, the protective effect of silymarin (a flavonoid) against B(a)P-induced toxicity was monitored in Wistar rats by evaluating the levels of hepatic phase I (CYP1A1), phase II enzymes (glutathione-S-transferase, epoxide hydroxylases, uridinediphosphate glucuronosyltransferases, NAD(P)H: quinone oxidoreductase 1, sulfotransferases), cellular antioxidant enzyme heme oxygenase and total glutathione. The results reveal that silymarin possesses substantial protective effect against B(a)P-induced damages by inhibiting phase I detoxification enzyme CYP1A1 and modulating phase II conjugating enzymes, which were confirmed by histopathological analysis. Overall, the inhibition of CYP1A1 and the modulation of phase II enzymes may provide, in part, the molecular basis for the effect of silymarin against B(a)P.

Synergistic effect of hydroxypropyl-β-cyclodextrin encapsulated soluble ferrocene and the gold nanocomposite modified glassy carbon electrode for the estimation of NO in biological systems

An electrochemical assay for sensing NO in biological systems is described in this paper. The ferrocene mediated reduction of NO, facilitated by the gold nanocomposite modified glassy carbon electrode is followed by an amperometric procedure. The analytical protocol involves the modification of a glassy carbon electrode by an overlayer of Au nanocomposites prepared through galvanic reduction. Additional overlayers can be built on the surface by repetition of the procedure. The modification leads to the decrease of the over-potential required for the analysis and results in a non-biofouling surface. Since the procedure is based on the electrochemical reduction of NO, the potential interferences from species like dopamine, ascorbic acid, etc., are overcome. The sensitivity, detection limit and response time achieved through this protocol for the modified electrode containing three Au overlayers are 0.03 nA/nM, 25.75 nM and <5 s. Analysis of NO has been carried out in real samples like liver extract, peripheral blood mononuclear cells (PBMCs) and miconazole nitrate ointment and the values obtained are comparable with that obtained by Griess analysis.

Silymarin prevents the toxicity induced by benzo(a)pyrene in human erythrocytes by preserving its membrane integrity: An in vitro study

Silymarin, the purified extract from milk thistle Silybum marianum (L.) Gaertn, consists mainly of four isomeric flavonolignans: silibinin, isosilibinin, silidianin, and silichristin. The present study was carried out to evaluate the protective potential of silymarin in human erythrocytes against in vitro exposure to the carcinogen benzo(a)pyrene (B(a)P). Erythrocytes isolated from human blood were divided into four groups and treated with Vehicle [Group I], B(a)P (300 μM) [Group II], Silymarin (500 μM) + B(a)P (300 μM) [Group III], and Silymarin alone (500 μM)] [Group IV]. Silymarin treatment maintains the integrity of erythrocytes by preventing hemolysis, protein thiol oxidation and by decreasing the activity of AChE. SEM observations indicate that B(a)P induced significant alteration in the morphology of erythrocytes to echinocytes, which may be due to the interaction of B(a)P with the membranes outer phopholipid monolayer. The light microscopic and SEM images show that silymarin treatment maintains the normal discocytic morphology of erythrocytes. The protective effect of silymarin might be attributed to its chemical structure and membranotrophic nature. The components silibinin, silydianin, and silychristin have OH in the 3rd, 5th, and 7th carbon atoms that may account for its increased antioxidant activity and removal of ROS formed during B(a)P metabolism.

Study of p53 codon 72 polymorphism and codon 249 mutations in Southern India in relation to age, alcohol drinking and smoking habits

Germline polymorphisms of genes involved in different steps of tumorigenesis like p53, the tumor suppressor gene, are reported to determine the individual susceptibility to cancer. Lung cancer is one of the most common and lethal cancers and tobacco smoking remains its most important etiologic factors. The most frequently p53 mutated codons of lung cancer are 72 (exon 4) and 249 (exon 7). Since mutations in the p53 gene are present in ∼40% of all human lung cancers and are more common in smokers than in nonsmokers, we aimed to detect the status of p53 at codon 72 for Arg/Arg or Arg/Pro or Pro/Pro allele polymorphism and p53 codon 249 mutation in smokers and nonsmokers of South India. Allele frequencies in the nonsmokers were 0.16 for the Arg/Pro allele and 0.84 for the Pro/Pro allele in our study population. Among the smokers, the frequencies of the Arg/Pro, Arg/Arg, and Pro/Pro alleles were 0.88, 0.04, and 0.08, respectively. No mutation was detected in both smokers and nonsmokers in p53 codon 249. From the worldwide scenario, it can be speculated that the smokers, with Arg/Pro genotype are more prone for lung cancer or to other types of cancer.

Antioxidant compounds in the seaweed Gelidiella acerosa protects human Peripheral Blood Mononuclear Cells against TCDD induced toxicity

Abstract 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a persistent environmental toxin formed as an unintentional by-product of incomplete combustion. Several therapeutic approaches have evolved to combat its toxicity since it elicits immunotoxicity, neurotoxicity, hepatotoxicity, carcinogenicity and lethality. Search for drugs from natural resources especially from seaweeds has become intense due to their enormous pharmacological potential. Hence, the present study aims at revealing the protective effect of methanolic extract of G. acerosa (MEGA) in Peripheral Blood Mononuclear Cells (PBMC) against TCDD induced toxicity, by assessing the antioxidant, anti-apoptotic and cytoprotective activities. The results of antioxidant assays suggests that MEGA reverted TCDD induced toxicity by causing an alteration in the levels of antioxidant enzymes (Catalase [CAT], Superoxide dismutase [SOD], Glutathione peroxidase [GPx], Glutathione-S-transferase [GST]) and Glutathione [GSH]. The results of lipid peroxidation assay and protein carbonyl content reveal that MEGA protects PBMC from TCDD induced macromolecular damage. MEGA was found to exhibit significant (p < 0.05) anti-apoptotic activity as verified by evaluation of mitochondrial membrane potential and AO-EtBr dual staining. In addition, PBMC co-treated with MEGA prevented TCDD induced oxidative DNA damage. Levels of phase-I detoxification enzymes determined by EROD assay and semi-quantitative RT-PCR showed that TCDD up-regulates the expression of CYP1A1 and upon co-treatment with MEGA, the expression got slightly decreased suggesting its protective role. Preliminary phytochemical analysis demonstrates that the extract is rich in cardiac glycosides and terpenoids. LC-MS analysis revealed the presence of antioxidants including caffeic acid, phytol and mannoheptulose in MEGA, which could be attributed for the observed protective effect against TCDD induced toxicity.