Dragana Mitić-Ćulafić

Protective effect of linalool, myrcene and eucalyptol against t-butyl hydroperoxide induced genotoxicity in bacteria and cultured human cells

We studied the protective effect of monoterpenes myrcene, eucalyptol and linalool against t-butyl hydroperoxide (t-BOOH) induced genotoxicity in reverse mutation assay with Escherichia coli WP2 IC185 strain and its oxyR mutant IC202, and with the comet assay in human hepatoma HepG2 and human B lymphoid NC-NC cells. The monoterpenes were tested in concentration ranges 0.05-1.5 mg/plate and 0.01-1.0 microg/ml in bacteria and mammalian cells, respectively. Suppression of t-BOOH induced mutagenesis was detected only in IC202 strain, and correlated with the observed inhibition of lipid peroxidation by the three monoterpenes. Linalool and myrcene strongly suppressed t-BOOH induced mutagenesis. Eucalyptol, in addition to moderate suppression of t-BOOH induced mutagenesis, suppressed also spontaneous mutagenesis. In NC-NC cells linalool and myrcene reduced t-BOOH induced DNA damage by about 50% at 0.01 microg/ml, while eucalyptol was less efficient (about 50% reduction at 1.0 microg/ml). In HepG2 cells linalool and eucalyptol reduced DNA damage by 30% and 40%, respectively, while myrcene was ineffective. The repair of t-BOOH induced DNA damage, studied in HepG2 cells, was not affected by monoterpenes. The results indicate that linalool, eucalyptol and myrcene have substantial protective effect against oxidant induced genotoxicity, which is predominately mediated by their radical scavenging activity.

Essential Oil of Myrtus communis L. as a Potential Antioxidant and Antimutagenic Agents

The present study describes DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging activity and antimutagenic properties of the essential oil of myrtle (Myrtus communis L.). Plant samples were collected from the two distant localities (southernmost and northern point) of the Montenegro coastline. Chemical profiles of the two samples were evaluated by GC-MS. In both of the samples monoterpenes were found to be the predominant compounds. Among them α-pinene, linalool, 1,8-cineole, and myrtenyl acetate were the major compounds. Significant differences between the samples were found in the ranges of α-pinene (14.7%–35.9%) and myrtenyl acetate (5.4%–21.6%). Both oils exhibited moderate DPPH scavenging activity, with IC50 values of 6.24 mg/mL and 5.99 mg/mL. The antimutagenic properties were assayed against spontaneous and t-BOOH-induced mutagenesis in Escherichia coli oxyR mutant IC202, a bacterial strain deficient in removing ROS. Reduction of the spontaneous mutagenesis in presence of myrtle EO was only slight, up to 13% at the highest concentration tested. When the oxidative mutagen was used, EO expressed higher reduction of mutagenesis, in a concentration dependent manner, with statistical significance for effect at the highest concentration tested (28%). Suppression of t-BOOH induced mutagenesis was correlated with the observed scavenging activity.

Modulation of genotoxicity and DNA repair by plant monoterpenes camphor, eucalyptol and thujone in Escherichia coli and mammalian cells

The aim of this work was to examine the antigenotoxic potential of plant monoterpenes: camphor, eucalyptol and thujone in prokaryotic and eukaryotic cells and to elucidate their effect on DNA repair. We compared the effect of monoterpenes on spontaneous, UV- and 4NQO-induced mutagenesis in Escherichia coli K12 repair proficient, and MMR and NER deficient strains. Positive controls tannic acid and vanillin were included in bacterial tests. We also examined protective effect of monoterpenes against 4NQO-induced genotoxicity in Vero cell line by alkaline comet assay. The results obtained in repair proficient strain indicated antimutagenic potential of monoterpenes against UV- and 4NQO-induced mutagenesis, which was diminished with NER deficiency. Camphor and eucalyptol maintained UV-induced SOS response longer than in controls, while thujone decreased SOS response and reduced general protein synthesis and the growth rate. The three monoterpenes increased spontaneous and UV-induced recombination in recA730 and camphor additionally in recA(+) cells. Incubation of 4NQO-pretreated Vero cells with monoterpenes resulted in significant reduction of tail moment. However, higher concentrations of monoterpenes induced DNA strand breaks. Obtained results indicate that by making a small amount of DNA lesions camphor, eucalyptol and thujone can stimulate error-free DNA repair processes and act as bioantimutagens.

Comparative study of genotoxic, antigenotoxic and cytotoxic activities of monoterpenes camphor, eucalyptol and thujone in bacteria and mammalian cells

Genotoxic/antigenotoxic, mutagenic/antimutagenic and cytotoxic effects of monoterpenes camphor, eucalyptol and thujone were determined in bacteria and mammalian cells using alkaline comet assay, Escherichia coli K12 reversion test and MTT assay, respectively. When applied in low doses (up to 200 μM in bacterial assay and 50 μM in comet assay) monoterpenes protected repair proficient E. coli and Vero cells against UV-induced mutagenesis and 4NQO-induced DNA strand breaks, respectively. Antimutagenic response was not detected in nucleotide excision repair (NER) deficient bacteria. When monoterpenes were applied in higher doses, a weak mutagenic effect was found in mismatch repair (MMR) and NER deficient E. coli strains, while induction of DNA strand breaks was evident in human fetal lung fibroblasts MRC-5, colorectal carcinoma HT-29 and HCT 116 cells, as well as in Vero cells. Moreover, the involvement of NER, MMR and RecBCD pathways in repair of DNA lesions induced by monoterpenes was demonstrated in E. coli. Camphor, eucalyptol and thujone were cytotoxic to MRC-5, HT-29 and HCT 116 cells. The most susceptible cell line was HCT 116, with IC50 values of 4.5 mM for camphor, 4 mM for eucalyptol and 1 mM for thujone. Observed effects of monoterpenes are consistent with hormesis response, characterized by a low dose beneficial effect and a high dose adverse effect of a stressor agent, and provide a basis for further study of both chemopreventive and chemotherapeutic potential of camphor, eucalyptol and thujone.

Essential oil of Eucalyptus gunnii hook. As a novel source of antioxidant, antimutagenic and antibacterial agents.

The present study describes radical scavenging capacity (RSC), antimutagenic and antibacterial properties of the essential oil (EO) of the leaves of Eucalyptus gunnii Hook. (Southern Montenegro). Chemical composition was evaluated by gas chromatography-mass spectrometry (GC-MS). In oil, 1,8-cineole (67.8%) and α-pinene (14.12%) were the major compounds comprising almost 82% of total EO. EO exhibited moderate DPPH (2,2-diphenyl-1-picrylhydrazyl) scavenging activity, with IC50 value of 7.19 µL/mL. The antimutagenic properties were assayed against the spontaneous and t-BOOH-induced mutagenesis in Escherichia coli IC202 oxyR mutant strain, deficient in removing radical oxygen species (ROS). Reduction of the spontaneous mutagenesis in the presence of E. gunnii EO was only slight, up to 12% at the highest concentration tested. However, when the oxidative mutagen was used, EO displayed more significant reduction of mutagenesis (maximum 23%) in a concentration dependent manner. Antibacterial activity was tested against the selected strains from ATTC and NCIB collections: Staphylococcus aureus, Staphylococcus epidermidis, Pseudomonas aeruginosa, Escherichia coli, Bacillus subtilis, Micrococcus flavus, Klebsiella pneumoniae, and the two Escherichia coli strains from our laboratory collection (SY252 and IB112) using both the disk-diffusion and MIC assays. The greatest sensitivity was shown by M. flavus, K. pneumoniae and E. coli lpcA (MIC = 0.83 mg/mL), while the highest resistance was shown by E. coli (ATTC 25922) and S. epidermidis. This study represents the first report on chemical composition and biological activity of the Eucalyptus gunnii in the South Balkan region and beyond.

The Protective Effects of Probiotic Bacteria on Cadmium Toxicity in Rats

One of the useful properties of probiotic bacteria is their capacity to bind different targets, thus eliminating them through feces. It is supposed that one of these targets could be cadmium, a widespread environmental toxicant that causes various disturbances in biological systems. This study examined the protective effects of probiotic supplementation against cadmium-induced toxicity in the rat. The experiment was conducted in the course of 5 weeks. Animals were divided into four groups: (1) controls, (2) probiotics treated, (3) cadmium treated, and (4) probiotics + cadmium treated. The cadmium concentration was measured in the blood, liver, kidney, and feces, as well as the blood alanine aminotransferase (ALT) and aspartate aminotransferase (AST) as biomarkers of the liver function. Histomorphological changes in the liver and kidney were also determined. Our results revealed that probiotics combined with cadmium increase this metal concentration in feces. As a result, blood, liver, and kidney Cd levels, as well as blood ALT and AST activities were lessened compared to the rat group treated with cadmium only. Besides, probiotics consumed simultaneously with cadmium attenuated histomorphological changes in the liver and kidney caused by cadmium. The rise in lactobacilli number in feces of rats treated simultaneously with cadmium and probiotics results in strong correlation with the increase of Cd concentration in their feces and the decrease of Cd concentration in their blood. We speculate that probiotics actively contribute to cadmium excretion through feces, probably, by its binding to their bacterial cell wall.

Cytotoxicity and genotoxicity of a low-shrinkage monomer and monoacylphosphine oxide photoinitiator: Comparative analyses of individual toxicity and combination effects in mixtures

OBJECTIVE To compare cytotoxicity and genotoxicity of novel urethane-based monomer FIT-852 and monoacylphosphine oxide photoinitiator (Lucirin TPO) with conventional Bisphenol A-glycidyl-methacrylate (BisGMA) and triethylene glycol dimethacrylate (TEGDMA) monomers and camphorquinone (CQ)/amine photoinitiator system, respectively. Moreover, we quantified and analyzed the combinatorial effects of individual substances in resin-based mixtures concerning the nature of the combinatorial effects. METHODS Cytotoxic and genotoxic effects of BisGMA, FIT, TEGDMA, CQ, DMAEMA and TPO and their combined toxicity in four clinically relevant mixtures (FIT/TPO, FIT/CQ, BisGMA/TPO, BisGMA/CQ) were tested on human fetal lung fibroblasts MRC-5 using MTT and Comet assays. We assessed combination effects of monomers and photoinitiators on overall toxicity from the measured concentration-effect relationships. Combination index (CI) was calculated on the basis of the median-effect equation derived from the mass-action law principle. RESULTS Individual substances showed decreasing cytotoxic effects in the following order: BisGMA>TPO>FIT>CQ>DMAEMA>TEGDMA. Experimental mixtures showed decreasing cytotoxic effects in the order BisGMA/TPO>BisGMA/CQ>FIT/CQ>FIT/TPO. FIT-based mixtures exhibited antagonistic cytotoxic effects between components while BisGMA-based mixtures demonstrated synergistic effects at ED50. TPO amplified both antagonistic and synergistic cytotoxic effects in mixtures. Pure substances showed genotoxicity in the following order: TPO>BisGMA>FIT>CQ>TEGDMA. We did not detect the genotoxic potential of DMAEMA. The rank of genotoxic concentrations of the mixtures was: BisGMA/TPO>BisGMA/CQ>FIT/CQ>FIT/TPO. SIGNIFICANCE Lower cytotoxicity and genotoxicity of FIT than BisGMA suggests its greater biocompatibility. Conversely, photoinitiator TPO was significantly more cytotoxic and genotoxic than both CQ and DMAEMA. CI values showed that components of FIT-based mixtures exhibit an antagonistic cytotoxic effect, while compontents of BisGMA-based mixtures show synergism.

Characterization of some potentially probiotic Lactobacillus strains of human origin

A novel preparation for human use was investigated for probiotic properties of new lactobacilli isolates from oral and fecal samples of children. Identified strains were Lactobacillus plantarum (Lac1, Lac2, Lac6, and Lac7), Lactobacillus casei (Lac3), and Lactobacillus paracasei (Lac4). Isolates were non-hemolytic, produced organic acids, were tolerant to wide ranges of temperature, NaCl, and pH, and were highly resistant to lysozyme, acidity, and bile salts. High survival rates in artificial gastric and intestinal fluids indicated abilities to survive passage through the gastrointestinal tract. Antimicrobial activities were restricted to bacteria, attributed to low pH values. Isolated strains possessed good aggregation abilities, high hydrophobicity values, and moderate abilities to adhere to HCT 116 cells. Substantial probiotic features were identified for all isolates. Lac2, Lac6, and Lac7 were identified as the most advantageous candidates for further study of other probiotic and technological properties.

Molecular Mechanisms of Action of Antimutagens from Sage (Salvia officinalis) and Basil (Ocimum basilicum)

© 2012 Nikolić et al., licensee InTech. This is an open access chapter distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Molecular Mechanisms of Action of Antimutagens from Sage (Salvia officinalis) and Basil (Ocimum basilicum)

Chemical characterization, antioxidant, genotoxic and in vitro cytotoxic activity assessment of Juniperus communis var. saxatilis

Chemical composition and antioxidative, genotoxic and cytotoxic potential of essential oil (EO) and post-distillation waste (PDW) of Serbian Juniperus communis L. var. saxatilis Pall. was studied in human lung carcinoma (A549) and normal lung fibroblast (MRC-5) cells. GC-MS analysis identified 93.95% of total EO content and determined α-pinen as a dominant component (23.61%). LC-MS/MS analysis of PDW pointed at rutin (12.2 mg g-1) and quinic acid (11.1 mg g-1) as the most abundant. Antioxidativity of PDW was strong in DPPH (IC50 was 5.27 μg mL-1), and moderate in TBA and FRAP assays. Both substances were more cytotoxic to A549 than to MRC-5 cells. Obtained IC50 values were 69.4 μg mL-1 and 120 μg mL-1 for EO, and 1.27 mg mL-1 and 2.86 mg mL-1 for PDW, respectively. PDW was genotoxic (0.3 mg mL-1 and 1 mg mL-1 in A549 and MRC-5 cells, respectively) and induced apoptosis and arrested cell cycle in G2/M phase in A549 cells (0.3 mg mL-1). In mixtures with doxorubicin cytotoxicity of EO and PDW increased, and combination index values (0.12-0.18) revealed clear synergistic effect, stronger in cancer cells. This indicates that J. communis var. saxatilis could decrease the chemotherapeutic doses of doxorubicin, potentially reducing its side effects.