The ameliorative and toxic effects of selenite on Caenorhabditiselegans

Selenium is an essential trace nutrient that has a narrow exposure window between its beneficial and detrimental effects. We investigated how selenium affected the development, fertility, and cholinergic signaling of the nematode, Caenorhabditiselegans. Our results showed that selenite supplementation at 0.01 and 0.05 μM accelerated development and increased the brood size, while the addition of 20 μM selenite retarded the developmental rate and decreased the brood size. We also showed that the 0.01 μM selenite-pretreated nematodes were more resistant to paralysis induced by an acetylcholinesterase inhibitor, aldicarb, and a nicotinic acetylcholine receptor agonist, levamisole, compared to untreated worms. In contrast, 20 μM selenite-pretreated animals were more sensitive to aldicarb- and levamisole-induced paralysis compared to untreated worms. We measured the internal selenium in supplemented worms using inductively coupled plasma atomic emission spectroscopy, and the data obtained suggested that selenite added to growth medium was taken up by the worms. Taken together, these results suggest that selenite exerts both ameliorative and toxic effects on C.elegans, depending on the amount. Our investigations here thus reinforce our understanding of the ameliorative and toxic effects of selenium on development, reproduction, and cholinergic signaling.     

Differential accumulation of paralytic shellfish toxins from Alexandrium minutum in the pearl oyster, Pinctada imbricata

To investigate the potential for differential accumulation of paralytic shellfish toxins (PSTs) in various tissues of the akoya pearl oyster, Pinctada imbricata, two feeding trials were carried out using the PST-producing dinoflagellate, Alexandrium minutum. When fed with A. minutum at concentrations between 100 and 1300 cells ml⁻¹, the maximum clearance by P. imbricata was shown to occur at a density of 300 cells ml⁻¹. When fed twice daily at this rate for up 12 days, P. imbricata accumulated analogues of gonyautoxins (GTXs): GTXs 1,4 and 2,3. The levels of GTXs in the viscera increased progressively on days 4, 8 and 12 to peak at 17.9 ± 4.47 μg STX-equivalent 100 g⁻¹ biomass. Following 12 days of depuration, in the absence of A. minutum, GTX levels fell by approximately 65% to 6.0 ± 2.20 μg STX-equivalent 100 g⁻¹ biomass. No GTX was found in the oysters at the start of the trial or in untreated controls. The accumulation of GTX was found to be tissue specific. No GTX was detected in the muscle tissue of P. imbricata during the feeding trial.     

Concentration- and time-dependent behavioral changes in Caenorhabditis elegans after exposure to nicotine

Nicotine induces profound behavioral responses in the model organism Caenorhabditis elegans. We tested the effect of a broad range of concentrations of nicotine (from 0.001 mM to 30 mM in nematode growth medium) on C. elegans locomotor behavior. We also followed the time-course influence on the sensitivity of C. elegans to nicotine (from 0 min to 300 min). A low concentration (0.001 mM) of this alkaloid causes a reduction of the speed of movement. By contrast, moderate concentrations (0.01 and 0.1 mM) induced acceleration of the mean speed of locomotion of C. elegans. High doses of nicotine (above 1 mM) induced slowing down of the movements and, finally, paralysis. Time-dependent analysis revealed that the stimulating effect of nicotine abolished the slowing down of C. elegans in control experiments after 30 min in the presence of 0.001, 0.1 and 10 mM nicotine. In the presence of 0.1 mM nicotine, the stimulation phase lasted up to 70 min. The evidence indicates that nicotine can have dual effects on the speed of locomotion, which is dependent on differences in its dosage and treatment time.     

Effect of phosphatidylinositol-3 kinase inhibition on ovotoxicity caused by 4-vinylcyclohexene diepoxide and 7, 12-dimethylbenz[a]anthracene in neonatal rat ovaries

4-vinylcyclohexene diepoxide (VCD) is an ovotoxicant that specifically destroys primordial and small primary follicles in the ovaries of mice and rats. In contrast, 7,12-dimethylbenz[a]anthracene (DMBA) is ovotoxic to all ovarian follicle classes. This study investigated phosphatidylinositol-3 kinase signaling involvement in VCD- and DMBA-induced ovotoxicity. Postnatal day (PND) 4 Fischer 344 (F344) rat whole ovaries were cultured for 2-12 days in vehicle control, VCD (30 μM), or DMBA (1 μM), ± PI3 kinase inhibitor LY294002 (20 μM) or its inactive analog LY303511 (20 μM). Following culture, ovaries were histologically evaluated, and healthy follicles were classified and counted. PI3 kinase inhibition had no effect on primordial follicle number, but reduced (P < 0.05) small primary and larger follicles beginning on day 4. VCD caused primordial and small primary follicle loss (P < 0.05) beginning on day 6. With PI3 kinase inhibition, VCD did not affect primordial follicles (P > 0.05) at any time, but did cause loss (P < 0.05) of small primary follicles. DMBA exposure caused primordial and small primary follicle loss (P < 0.05) on day 6. Further, DMBA-induced primordial and small primary follicle loss was greater with PI3 kinase inhibition (P < 0.05) than with DMBA alone. These results support that (1) PI3 kinase mediates primordial to small primary follicle recruitment, (2) VCD, but not DMBA, enhances ovotoxicity by increasing primordial to small primary follicle recruitment, and (3) in addition to xenobiotic-induced ovotoxicity, VCD is also a useful model chemical with which to elucidate signaling mechanisms involved in primordial follicle recruitment.     

Antibacterial effects of enniatins J1 and J3 on pathogenic and lactic acid bacteria

Enniatins (ENs) are N-methylated cyclohexadepsipeptides, secondary metabolites produced by various species of the genus Fusarium. They are known to act as antifungal, antiyeast and antibacterial and to possess antiinsecticidal and phytotoxic properties. In this study we evaluated for the first time the antibiotic effect of pure fractions of EN J₁ and J₃ on several pathogenic strains and lactic acid bacteria.The ENs J₁ and J₃ were purified from the fermentation extract of Fusarium solani growth on solid medium of wheat kamut, using the technique of the low pressure liquid chromatography (LPLC) followed by a semipreparative liquid chromatography (LC). The purity and the structure of the isolated compound were confirmed by electrospray ionization-mass spectrometry study-linear ion trap (ESI-MS-LIT).The use of both chromatographic techniques have permitted to produce and purify 47 mg of the En J₁ and 50 mg of the EN J₃ with a mean purity of 98% completely characterized with the technique of the ESI-MS-LIT.Microbial bioassay analyses were carried out by incubation in MRSA and TSA for acid lactic and pathogenic bacteria, respectively during 24 h at 37 °C. None of the tested strains were inhibited by a 1 ng dose of EN J₁ and J₃. These compounds were only not effective against Listeria monocytogenes, Pseudomonas aeruginosa and Salmonella enteric. This study highlight ENs J₁ and J₃ could be potentially effective antibacterial agents against several pathogenic and lactic acid bacteria.     

Infection by mycotoxigenic fungal species and mycotoxin contamination of maize grain in Umbria, central Italy

Surveys were carried out in 2006 and 2007 in Umbria (central Italy) to evaluate the presence of mycotoxigenic fungi and mycotoxins in maize grain sampled at harvest. Fusarium spp., were the most abundant species detected in maize kernels, followed by Aspergillus species of sections Flavi and Nigri and by Penicillium spp. Among Fusarium species, F. verticillioides was the most prevalent species, as detected by PCR directly on the kernels and on the fungi isolated from the kernels, followed by F. proliferatum and F. subglutinans. Fumonisins were the predominant mycotoxins with values, on average, of 4.3 and 5.7 mg kg⁻¹, in 2006 and 2007, respectively, with a maximum of 76.3 mg kg⁻¹ in the second year. Deoxynivalenol ranged from 0.2 to 3.98 mg kg⁻¹ in 2006 (average 1.04 mg kg⁻¹) and from undetectable levels to 14 mg kg⁻¹ in 2007 (average 0.86 mg kg⁻¹). Aflatoxins, analyzed only in 2007, averaged 26.3 μg kg⁻¹, with a maximum of 820 μg kg⁻¹. Zearalenone content was always very low. Results indicate that EU legal limits for these mycotoxins were rarely exceeded with low levels across most of the examined area, suggesting that this region could be considered suitable for the production of healthy maize.     

Separation and determination of four ganoderic acids from dried fermentation mycelia powder of Ganoderma lucidum by capillary zone electrophoresis

Ganoderic acids (GAs) were bioactive secondary metabolites produced by a traditional mushroom Ganoderma lucidum. We describe a simple and efficient method for the separation and quantitative determination of four GAs, namely Ganoderic acid T (GA-T), Ganoderic acid Mk (GA-Mk), Ganoderic acid Me (GA-Me) and Ganoderic acid S (GA-S) from dried triterpene-enriched extracts of G. lucidum mycelia powder by capillary zone electrophoresis (CZE). Under the optimum conditions, the four GAs reached the baseline separation in 9 min with Glycyrrhetinic acid (GTA) as internal standard. The four GAs and internal standard (GTA) were detected at a wavelength 245 nm. All calibration curves showed good linearity (r² > 0.9958) within test ranges. Limit of detection (LOD) and limit of quantification (LOQ) were less than 0.6 and 1.8 μg/mL, respectively. The relative standard deviation (R.S.D.) values of precision and recoveries were less than 5% and recoveries ranged from 91.4% to 103.6%. This was the first report on simultaneous determination of the four GAs and the results provided a firm basis for the trace analysis of GAs in dried fermentation mycelia powder of G. lucidum with high accuracy.     

Chemical composition,antifungal and antiaflatoxigenic activities of Ocimum sanctum L. essential oil and its safety assessment as plant based antimicrobial

The study deals with the efficacy of Ocimum sanctum essential oil (EO) and its major component, eugenol against the fungi causing biodeterioration of food stuffs during storage. O. sanctum EO and eugenol were found efficacious in checking growth of Aspergillus flavus NKDHV8; and, their minimum inhibitory concentrations (MICs) were recorded as 0.3 and 0.2 μl ml⁻¹, respectively. The O. sanctum EO and eugenol also inhibited the aflatoxin B₁ production completely at 0.2 and 0.1 μl ml⁻¹, respectively. Both of these were found superior over some prevalent synthetic antifungals and exhibited broad fungitoxic spectrum against 12 commonly occurring fungi. The LD₅₀ value of O. sanctum EO on mice was found to be 4571.43 μl kg⁻¹ suggesting its non-mammalian toxic nature. The findings of present study reveals the possible exploitation of O. sanctum EO and eugenol as plant based safe preservatives against fungal spoilage of food stuffs during storage.     

Protective effect of Acorus calamus L. in rat model of vincristine induced painful neuropathy: An evidence of anti-inflammatory and anti-oxidative activity

The study investigates the protective effect of Acorus calamus L. (AC) in vincristine-induced painful neuropathy. Vincristine (75 μg/kg, i.p. for 10 consecutive days) was administered to induce painful neuropathy in rats. Various tests were performed to assess the degree of painful neuropathy at different days i.e., 0, 1, 7, 14, and 21st day. Sciatic nerve TNF-α, superoxide anion generation, total calcium, and myeloperoxidase activity level were also estimated after 21st day of study. Hydro-alcoholic extract of AC (HAE-AC, 100 and 200 mg/kg, p.o.) and pregabalin (10 mg/kg, p.o.) were administered for 14 consecutive days. Vincristine significantly induced neuropathic pain manifested in the terms of thermal hyperalgesia and allodynia (increase in hind paw licking, lifting or jumping from hot plate); mechanical hyperalgesia (increase in left hind paw lifting duration in pin-prick test) and allodynia (left hind paw withdrawal reflexes to non-noxious stimuli in Von Frey test); and sciatic functional index (analysis of footprints of the feet) along with rise in the levels of various biochemicals. HAE-AC attenuated vincristine induced behavioral, and biochemical changes comparable to that of pregabalin (positive control). HAE-AC attenuated vincristine induced painful neuropathy, which may be attributed to its multiple effects including anti-oxidative, anti-inflammatory, and calcium inhibitory actions.     

Evaluation of the immunogenicity and in vivo toxicity of the antimicrobial peptide P34

Immunogenicity and toxicity of antimicrobial peptide P34 were evaluated in vivo. BALB/c mice were inoculated intraperitoneally with peptide P34 alone and associated with Freund's adjuvant. For acute toxicity testing, different concentrations of the peptide P34 (82.5, 165.0, 247.5 and 330.0 mg/kg) were orally administered. To evaluate the sub-chronic toxicity the tested dose of 0.825 mg/kg/day of the peptide P34 or nisin were administered for 21 days. There were no hypersensitivity reactions or significant increase in antibody titer during the immunogenicity experiment or death of animals during the acute or sub-chronic toxicity tests. The LD₅₀ was higher than 332.3 ± 0.76 mg/kg. No significant changes in serum biochemical parameters were observed in the animals treated with the peptide P34 unlike nisin-treated group showed a significant increase in alanine transaminase levels in comparison to controls. The group treated with 0.825 mg/kg/day of nisin showed histological changes in the spleen, skin and liver. In the group treated with peptide P34 histological changes in the spleen were observed, with the presence of megakaryocytes. Few studies report the use of animal models to evaluate the in vivo toxicity of antimicrobial peptides and such investigation is an essential step to ensure it safe use in foods.     

Literature review on the safety of Toyocerin,a non-toxigenic and non-pathogenic Bacillus cereus var. toyoi preparation

Bacillus cereus var. toyoi is a naturally occurring, non-toxigenic and non-pathogenic strain of B. cereus. Safety studies were conducted on a B. toyoi preparation (Toyocerin^®), including but not limited to enterotoxicity, eye irritation, genotoxicity, acute, subchronic and chronic toxicity studies and human clinical trials. In rabbits, Toyocerin^® did not exhibit enterotoxicity and was only slightly irritating to the eyes. It was non-mutagenic in an Ames assay at up to 10,000 μg/plate and did not exhibit clastogenic activity in a chromosomal aberration test at up to 450 mg/ml. It was non-toxic in acute and repeated-dose (30 and 60 days and 1 year) toxicity studies in rats and mice at up to 3 × 10¹¹ spores/kg bw/day. In an eight-day human clinical trial, Toyocerin^® did not cause any adverse effects in healthy male and female subjects at 1 × 10⁹ and 1 × 10¹⁰ spores/kg bw/day. In feeding trials, Toyocerin^® did not cause any adverse effects in rabbits, pigs, chickens, turkeys and cattle at doses ranging from 8.5 × 10⁷ to 4 × 10⁹ spores/kg bw/day for durations of 2 weeks to 18 months. Taken together, these studies demonstrate that Toyocerin^® is safe at the doses tested.     

In vitro activities of ceftobiprole combined with amikacin or levofloxacin against Pseudomonas aeruginosa: evidence of a synergistic effect using time-kill methodology

Ceftobiprole is an investigational intravenous broad-spectrum cephalosporin with in vitro activity against Gram-positive and Gram-negative pathogens, including meticillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa. Pseudomonas aeruginosa is a frequent nosocomial pathogen, increasingly associated with complicated skin and skin-structure infections. Combination antimicrobial therapy is recommended as empirical therapy for serious infections where P. aeruginosa is suspected. Therefore, in this study the interaction of ceftobiprole with two other antipseudomonal agents (amikacin and levofloxacin) was investigated. Time-kill studies were performed for each single agent and for the combination of ceftobiprole 4 mg/L with either amikacin or levofloxacin at 0.5×, 1× and 2× the minimum inhibitory concentration. Five clinical isolates of P. aeruginosa as well as the P. aeruginosa ATCC 27853 reference strain were tested at initial inocula of 5 × 10⁵ colony-forming units (CFU)/mL (low inoculum) or 5 × 10⁷ CFU/mL (high inoculum). Synergy was defined as a decrease of ≥2 log₁₀ CFU/mL with the combination compared with the most active single drug at 6 h and 24 h. At low inoculum with ceftobiprole as a single agent, viable counts were decreased by 1.5-2 log₁₀ at 6 h. Addition of either amikacin or levofloxacin resulted in synergistic bactericidal activity at 24 h. At high inoculum the combination of ceftobiprole with amikacin or levofloxacin demonstrated synergism in one of three and three of five strains, respectively. This study demonstrated that the combination of ceftobiprole at a clinically achievable concentration of 4 mg/L with amikacin or levofloxacin exhibited synergistic activity against P. aeruginosa. There was no evidence of antagonism for either combination.     

In vitro activity of S-(3,4-dichlorobenzyl)isothiourea hydrochloride and novel structurally related compounds against multidrug-resistant bacteria, including Pseudomonas aeruginosa and Burkholderia cepacia complex

The aim of this study was to establish the antimicrobial activities of S-(3,4-dichlorobenzyl)isothiourea hydrochloride (A22) and a series of structurally related compounds against multidrug-resistant (MDR) bacteria. The minimum inhibitory concentrations (MICs) of 21 compounds were determined against 18 strains of pathogenic bacteria in addition to Pseudomonas aeruginosa (n = 19) and Burkholderia cepacia complex (BCC) (n = 20) isolated from the sputa of cystic fibrosis patients. Selected compounds were tested against further isolates, including P. aeruginosa (n = 100), BCC (n = 12) and Stenotrophomonas maltophilia (n = 19). The interaction of S-(4-chlorobenzyl)isothiourea hydrochloride (C2) in combination with conventional antimicrobials was examined against 10 P. aeruginosa strains. Selected compounds were also tested against Enterobacteriaceae producing NDM-1 carbapenemase (n = 64) and meticillin-resistant Staphylococcus aureus (MRSA) (n = 37). Of the 21 compounds, 14 showed antimicrobial activity that was generally more pronounced against Gram-negative bacteria. Against P. aeruginosa, the most active compound was C2 [MIC for 50% of the organisms (MIC₅₀) = 32 μg/mL]. This compound was also the most active against BCC, with all isolates inhibited by 64 μg/mL. For all ten strains of P. aeruginosa subjected to combination testing with C2 and conventional antimicrobials, a bactericidal effect was achieved with at least one combination. C2 and A22 both showed strong activity [MIC for 90% of the organisms (MIC₉₀) = 4 μg/mL] against Enterobacteriaceae that produced NDM-1 carbapenemase. Finally, S-(4-chlorobenzyl)-N-(2,4-dichlorophenyl)isothiourea hydrochloride showed good activity (MIC₉₀ = 8 μg/mL) against MRSA. This work establishes the activity of isothiourea derivatives against a broad range of clinically important MDR bacteria.     

In vitro and ex vivo angiogenic effects of roxarsone on rat endothelial cells

Roxarsone, a feed additive, is being used worldwide to promote animal growth. However, the potential effect of roxarsone on angiogenesis has not been extensively characterized. We examined the ability of roxarsone to promote angiogenesis of rat endothelial cells in vitro and from rat aorta rings ex vivo. Endothelial cells from rats were exposed to 0.01–10.00 μM roxarsone, 5 ng/mL vascular endothelial growth factor (VEGF) as a positive control or phosphate buffer saline (PBS) as a negative control. Cell proliferation was measured by MTT assay, and the content of VEGF in supernatants was measured by enzyme-linked immunosorbent assay and Western blotting. A Matrigel-induced tube formation assay was used to evaluate the effects of roxarsone on endothelial cells. Additionally, the total number and length of microvessels sprouted from rat aortic rings were measured for ex vivo investigation of angiogenesis. Results showed that the cell viability and total number and length of capillary-like tube formations after roxarsone treatment was significantly higher than that of negative (P < 0.05), with a maximum effect at 1.00 μM exposure. Furthermore, the number of microvessels sprouted from aortic rings treated for 4 h with 0.1–10.0 μM roxarsone was significantly higher than that of PBS treatment, with a peak value of 1.0 μM. These results further demonstrate the potential of roxarsone to promote angiogenesis in vitro and ex vivo.     

Genetic polymorphisms in hMTH1, hOGG1 and hMYH and risk of chronic benzene poisoning in a Chinese occupational population

Oxidative damage to DNA induced by benzene is an important mechanism of its genotoxicity, which leads to chronic benzene poisoning (CBP). Therefore, genetic variation in DNA repair genes may contribute to susceptibility to CBP in the exposed population. We hypothesized that single nucleotide polymorphisms (SNPs) in hMTH1, hOGG1 and hMYH genes are associated with risk of CBP. We genotyped SNPs at codon 83 of hMTH1, codon 326 of hOGG1, and codon 324 of hMYH in 152 CBP patients and 152 healthy workers occupationally exposed to benzene without poisoning manifestations. The genotypes were determined by polymerase chain reaction-restrained fragment length polymorphism (PCR-RFLP) technique. There were 2.51-fold [adjusted odds ratio (OR_adj), 2.51; 95% CI, 1.14-5.49; P = 0.02] and 2.49-fold (OR_adj, 2.49; 95% CI: 1.52-4.07; P < 0.01) increased risk of CBP for individuals carrying genotypes of hMTH1 83Val/Met + Met/Met and hOGG1 326Cys/Cys, respectively. Compared with individuals carrying genotypes of hOGG1 326Cys/Cys and hMYH 324His/His at the same time, there was a 0.33-fold (OR_adj, 0.33; 95% CI: 0.15-0.72; P < 0.05) decreased risk of CBP for those with genotypes of hOGG1 326Ser/Cys + Ser/Ser and hMYH 324His/Gln + Gln/Gln. In the smoking group, there was a 0.15-fold (OR_adj, 0.15; 95% CI, 0.03-0.68; P = 0.01) decreased risk of CBP for subjects carrying genotypes of hMYH 324His/Gln + Gln/Gln compared with those of genotype of hMYH 324His/His. Therefore, our results suggested that polymorphisms at codons 83 of hMTH1 and codon 326 of hOGG1 might contribute to CBP in a Chinese occupational population.     

Induction of hsp70, hsp60, hsp83 and hsp26 and oxidative stress markers in benzene, toluene and xylene exposed Drosophila melanogaster: Role of ROS generation

Exposure to benzene, toluene and xylene in the human population may pose a health risk. We tested a working hypothesis that these test chemicals cause cellular toxicity to a non-target organism, Drosophila melanogaster. Third instar larvae of D. melanogaster transgenic for hsp70, hsp83 and hsp26 and Oregon R⁺ strain were exposed to 1.0-100.0 mM benzene, toluene and xylene for 2-48 h to examine the heat shock proteins (hsps), ROS generation, anti-oxidant stress markers and developmental end points. The test chemicals elicited a concentration- and time-dependent significant (p < 0.01) induction of the hsps in the exposed organism in the order of hsp70 > hsp83 ≥ hsp26 as evident by β-galactosidase activity after 24 h. RT-PCR amplification studies in Oregon R⁺ larvae revealed a similar induction pattern of these genes along with hsp60 in the order of hsp70 > hsp60 > hsp26 ≥ hsp83. Under similar experimental conditions, a significant induction of ROS generation and oxidative stress markers viz. superoxide dismutase, catalase, glutathione S-transferase, thioredoxin reductase, glutathione, malondialdehyde and protein carbonyl content was observed. Sub-organismal response was propagated towards organismal response i.e., a delay in the emergence of flies and their reproductive performance. While hsp70 was predominantly induced in the organism till 24 h of treatment with the test chemicals, a significant or insignificant regression of Hsp70 after 48 h was concurrent with a significant induction (p < 0.01) of hsp60 > hsp83 ≥ hsp26 in comparison to the former. A significant positive correlation was observed between ROS generation and these hsps in the exposed organism till 24 h and a negative correlation between ROS generation and hsp70 in them after 48 h indicating a modulatory role of ROS in the induction of hsps. The study suggests that among the tested hsps, hsp70 may be used as an early bioindicator of cellular toxicity against benzene, toluene and xylene and D. melanogaster as an alternative animal model for screening the risk posed by environmental chemicals.     

Gastroprotective and toxicological evaluation of the Lithothamnion calcareum algae

Lithothamnion calcareum is a red alga of the Corallinacea family whose main feature is the formation of calcium carbonate precipitate in its cell walls. L. calcareum is marketed as a nutritional supplement for calcium and other minerals in Brazil and other countries under the pharmaceutical name of Vitality 50+®. In this study, gastroprotective and pre-clinical toxicity assays were performed on this product. Doses of 30, 120 and 480 mg/kg were used in the gastroprotective study on Wistar rats. A dose of 2000 mg/kg was used in the preclinical acute toxicity study and oral doses of 1000 and 2000 mg/kg were used in the subchronic toxicity evaluation. L. calcareum played no significant role in the protection of the rats’ gastric mucosa, nor did it cause increase in gastric irritation. No impact on the acute toxicity test was identified. In the subchronic toxicity test, serum levels of albumin, total protein and calcium decreased, and creatinine levels increased, suggesting hypercalcemia and possible kidney damage associated with liver damage, given that the majority of these parameters were irreversible. Thus, this work aims to discuss the relationship of the high concentration of calcium in the product with the observed effects.     

Toxic essential oils: Anxiolytic,antinociceptive and antimicrobial properties of the yarrow Achillea umbellata Sibth. et Sm. (Asteraceae) volatiles

Many plant species are used for medicinal purposes without the knowledge of their possible toxic effect. The ethnopharmacologically renowned genus Achillea L. (Asteraceae) is even more troublesome in this respect since different taxa are believed to have the same beneficial properties as A. millefolium. According to the median lethal i.p. dose (LD₅₀ = 853 mg/kg, mice), the volatiles of Achillea umbellata Sibth. et Sm. are more toxic than the thujone-containing essential oils (LD₅₀ > 960 mg/kg). A GC–MS analysis of A. umbellata oil revealed the presence of a series of fragranyl esters (six new natural products). The major constituents of this oil, the rare monoterpene alcohol fragranol and fragranyl acetate, and one more ester (benzoate), as well as the oil itself, showed antianxiety, analgesic and, in some instances, paralyzing properties at 50–150 mg/kg but these are very likely sign of intoxication and not of possible beneficial effects of the plant volatiles. Testing of antimicrobial activity demonstrated that the oil possesses moderate activity against pathogenic microorganisms, but the effect of the oil differs in pro- and eukaryotic cells. According to the results obtained, fragranol may be considered as the main active principle responsible for the observed activity/toxicity.     

Antigenic, microbicidal and antiparasitic properties of an l-amino acid oxidase isolated from Bothrops jararaca snake venom

Venoms from the bee Apis mellifera, the caterpillar Lonomia achelous, the spiders Lycosa sp. and Phoneutria nigriventer, the scorpions Tityus bahiensis and Tityus serrulatus, and the snakes Bothrops alternatus, Bothrops jararaca, Bothrops jararacussu, Bothrops moojeni, Bothrops neuwiedi, Crotalus durissus terrificus, and Lachesis muta were assayed (800 μg/mL) for activity against Staphylococcus aureus. Venoms from B. jararaca and B. jararacussu showed the highest S. aureus growth inhibition and also against other Gram-positive and Gram-negative bacteria. To characterize the microbicidal component(s) produced by B. jararaca, venom was fractionated through gel exclusion chromatography. The high molecular weight, anti-S. aureus P1 fraction was further resolved by anion exchange chromatography through Mono Q columns using a 0-0.5 M NaCl gradient. Bactericidal Mono Q fractions P5 and P6 showed significant LAAO activity using l-leucine as substrate. These fractions were pooled and subjected to Heparin affinity chromatography, which rendered a single LAAO activity peak. The anti-S. aureus activity was abolished by catalase, suggesting that the effect is dependent on H₂O₂ production. SDS-PAGE of isolated LAAO indicated the presence of three isoforms since deglycosylation with a recombinant N-glycanase rendered a single 38.2 kDa component. B. jararaca LAAO specific activity was 142.7 U/mg, based on the oxidation of l-leucine. The correlation between in vivo neutralization of lethal toxicity (ED₅₀) and levels of horse therapeutic antibodies anti-LAAO measured by ELISA was investigated to predict the potency of Brazilian antibothropic antivenoms. Six horses were hyperimmunized with Bothrops venoms (50% from B. jararaca and 12.5% each from B. alternatus, B. jararacussu, B. neuwiedii and B. moojeni). To set up an indirect ELISA, B. jararaca LAAO and crude venom were used as antigens. Correlation coefficients (r) between ED₅₀ and ELISA antibody titers against B. jararaca venom and LAAO were 0.846 (p < 0.001) and 0.747 (p < 0.001), respectively. The hemolytic and leishmanicidal (anti-Leishmania amazonensis) activity of LAAO was also determined.     

Influence of GSTs, CYP2E1 and mEH polymorphisms on 1, 3-butadiene-induced micronucleus frequency in Chinese workers

1,3-butadiene (BD) has been classified as a human carcinogen, however, the relationship between chromosomal damage and its metabolic polymorphisms is not clear. The present study used the CBMN assay to detect chromosomal damage in the peripheral lymphocytes of 166 exposed workers and 41 non-exposed healthy individuals. PCR and PCR-RFLP were applied to detect GSTT1, GSTM1, CYP2E1 c1c2 and mEH Tyr113His, His139Arg polymorphisms. The results demonstrated that the micronucleus (MN) frequency of the exposed workers was significantly higher than controls (P < 0.01). Among the exposed workers, the individuals with high BD exposures are more susceptible to chromosomal damage than those with low exposures (FR = 1.30, 95% CI 1.14-1.53; P < 0.05). Gender-difference was also found in our study: males got lower micronucleus frequency than females. Workers who carried the genotypes of GSTM1 (+), CYP2E1 (c1c2/c2c2) and mEH intermediate (I) group had significantly higher MN frequency than those carrying the genotypes of GSTM1 (−) (FR = 1.29, 95% CI 1.05-1.59; P < 0.05), CYP2E1 (c1c1) (FR = 1.55, 95% CI 1.24-1.93; P < 0.01) or mEH high (H) group (FR = 1.57, 95% CI 1.08-2.34; P < 0.05), respectively. Our data indicated that the current BD exposure level could cause significantly higher MN frequency in workers than controls. Polymorphisms of GSTM1, CYP2E1 and mEH are susceptible to altered chromosome damage.