Re-examination of the anion derivatives of isoflavones by radical fragmentation in negative electrospray ionization tandem mass spectrometry: experimental and computational studies

This paper reports theoretical and experimental studies of gas-phase fragmentation reactions of four naturally occurring isoflavones. The samples were analyzed in negative ion mode by direct infusion in ESI-QqQ, ESI-QqTOF and ESI-Orbitrap systems. The MS/MS and MS(n) spectra are in agreement with the fragmentation proposals and high-resolution analyses have confirmed the formulae for each ion observed. As expected, compounds with methoxyl aromatic substitution have showed a radical elimination of center dot CH(3) as the main fragmentation pathway. A second radical loss (center dot H) occurs as previously observed for compounds which exhibit a previous homolytic center dot CH(3) cleavage (radical anion) and involves radical resonance to stabilize the anion formed. However, in this study we suggest another mechanism for the formation of the main ions, on the basis of the enthalpies for each species. Compounds without methoxy substituent dissociate at the highest energies and exhibit the deprotonated molecule as the most intense ion. Finally, energy-resolved experiments were carried out to give more details about the gas-phase dissociation reaction of the isoflavones and the results are in agreement with the theoretical approaches. Copyright (C) 2011 John Wiley & Sons, Ltd.

Fragmentation studies and electrospray ionization mass spectrometry of lapachol: protonated, deprotonated and cationized species

Electrospray ionization mass spectrometric analysis of lapachol (2-hydroxy-3-(3-methy1-2-butenyl)-1,4-naphthoquinone) was accomplished in order to elucidate the gas-phase dissociation reactions of this important biologically active natural product. The occurrence of protonated and cationized species in the positive mode and of deprotonated species in the negative mode was explored by means of collision-induced dissociation (CID) experiments. For the protonated molecule, the H(2)O and C(4)H(8) losses occur by two competitive channels. For the deprotonated molecule, the even-electron rule is not conserved, and the radicalar species are eliminated by formation of distonic anions. The fragmentation mechanism for each ion was suggested on the basis of computational thermochemistry. Atomic charges, relative energies, and frontier orbitals were employed aiming at a better understanding of the gas-phase reactivity of lapachol. Potential energy surfaces for fragmentation reactions were obtained by the B3LYP/6-31+G(d,p) model. Copyright (C) 2010 John Wiley & Sons, Ltd.

Metal embryotoxicity from urban particles in Sao Paulo city: An experimental study in chicken embryos

Chicken eggs were inoculated with suspensions of ambient air particles (<= 10 mu m, PM(10)) from Sao Paulo city in 3, 0.3 or 0.03 mu g doses on one of the four early days of embryo development. On the eleventh day of development alterations were observed on embryos inoculated with PM(10) 3 mu g on the third day. Particles analysis showed high content of metals. Hence, embryos were also inoculated with PM(10) (3 mu g) combined with metal chelating EDTA. PM(10) (3 mu g) embryos presented underdevelopment (stage 29.44 +/- 11.4) compared to vehicle and positive controls (stage 36.44 +/- 0.51 Saline and stage 31.20 +/- 9.7 Cyclophosphamide, p <= 0.05); higher (47%) mortality rate (23% Saline and 42% Cyclophosphamide) and low (68%) viability (100% Saline and 70% Cyclophosphamide, p = 0.04). Effects were attenuated when embryos received PM(10) + EDTA (stage 33.63 +/- 0.94, 18.9% mortality rate and 82% viability). PM(10) from Sao Paulo city is embryotoxic and metal may be implicated in the toxic mechanism. (C) 2010 Elsevier Inc. All rights reserved.

Predictors of Lethality in Severe Leptospirosis in Urban Brazil

To ascertain prognostic factors associated with fatal outcomes in severe leptospirosis, a retrospective case-control study was done using population-based surveillance data. Centralized death certificate reporting of leptospirosis mortality was combined with details of patients` hospitalizations, which were obtained from hospitals representing all sectors of Sao Paulo city. Among identified leptospirosis cases, 89 lethal cases and 281 survivor cases were analyzed. Predictors of death included age > 40 years, development of oliguria, platelet count < 70,000/mu L, creatinine > 3 mg/dL. and pulmonary involvement. The latter was the strongest risk Factor with all estimated odds ratio of 6.0 (95% confidence interval: 3.0-12.0). Serologic findings with highest titer against Leptospira interrogans serovar Copenhageni did not show significant differences between survivors and non-survivors. Lung involvement was an important predictor of death in leptospirosis in Sao Paulo, of relevance in leptospirosis-endemic regions where this complication is common.

The time course of vasoconstriction and endothelin receptor A expression in pulmonary arterioles of mice continuously exposed to ambient urban levels of air pollution

The present study aimed to verify the time course of the effects of environmental levels of urban air pollution toxicity on lung arterioles. BALB/c mice (n = 56) were continuously exposed to selective chambers equipped with (filtered, F) or without (non-filtered, NF) filter devices for particles and toxic gases for 24 h/day, over 14, 21, 30 or 45 days. After exposure, we evaluated the lumen-wall relationship (an estimator of arteriolar narrowing), endothelial nitric oxide synthase (eNOS) and endothelin type A receptor (ETAr) expression in the vascular wall and inflammatory influx of the peribronchiolar area. Concentrations of fine particulate matter (PM <= 2.5 mu g/m(3)), nitrogen dioxide (NO(2)), black smoke (BS), humidity and temperature in both the environment and inside the chambers were measured daily. Filters cleared 100% of BS and 97% of PM inside the F chamber. The arteriole wall of the lungs of mice from NF chamber had an increased ETAr expression (p <= 0.042) concomitant to a decrease in the lumen/wall ratio (p = 0.02) on the early days of exposure, compared to controls. They also presented a progressive increment of inflammatory influx in the peribronchiolar area during the study (p = 0.04) and decrement of the eNOS expression on the 45th day of exposure in both vascular layers (p <= 0.03). We found that after 14 days of exposure, the ambient levels of air pollutants in Sao Paulo induced vasoconstriction that was associated with an increase in ETAr expression. These vascular results do not appear to be coupled to the progressive inflammatory influx in lung tissue, suggesting a down-regulation of vasoconstrictive mechanisms through an imbalance in the cytokines network. It is likely that these responses are protective measures that decrease tissue damage brought about by continuous exposure to air pollutants. (C) 2010 Elsevier Inc. All rights reserved.

Derivation and external validation of a simple prediction model for the diagnosis of type 2 Diabetes Mellitus in the Brazilian urban population

A risk score model was developed based in a population of 1,224 individuals from the general population without known diabetes aging 35 years or more from an urban Brazilian population sample in order to select individuals who should be screened in subsequent testing and improve the efficacy of public health assurance. External validation was performed in a second, independent, population from a different city ascertained through a similar epidemiological protocol. The risk score was developed by multiple logistic regression and model performance and cutoff values were derived from a receiver operating characteristic curve. Model`s capacity of predicting fasting blood glucose levels was tested analyzing data from a 5-year follow-up protocol conducted in the general population. Items independently and significantly associated with diabetes were age, BMI and known hypertension. Sensitivity, specificity and proportion of further testing necessary for the best cutoff value were 75.9, 66.9 and 37.2%, respectively. External validation confirmed the model`s adequacy (AUC equal to 0.72). Finally, model score was also capable of predicting fasting blood glucose progression in non-diabetic individuals in a 5-year follow-up period. In conclusion, this simple diabetes risk score was able to identify individuals with an increased likelihood of having diabetes and it can be used to stratify subpopulations in which performing of subsequent tests is necessary and probably cost-effective.

Metabolic syndrome determinants in an urban population from Brazil: Social class and gender-specific interaction

Introduction: The metabolic syndrome (MS) is characterized by multiple cardiovascular risk factors such as central obesity, arterial hypertension, dislipidemia and hyperinsulinemia and is associated with a higher incidence of cardiovascular events and mortality. The aim of the present work is to describe the prevalence of MS in an urban population from a highly admixed developing country and to characterize the different correlations between this diagnosis, cardiovascular risk factors and demographic variables distributed in this population. Materials and methods: A cross-sectional study of risk factors for cardiovascular diseases was performed in the urban population of Vitoria, Brazil (n= 1507). Major cardiovascular risk factors such as smoking habits, alcohol intake, amount of physical activity, diabetes and hypertension were inquired. Blood biochemical assays were performed by standard techniques in 12 h fasting blood sample and Metabolic Syndrome (MS) was characterizes following the ATP III criteria. Results: The analysis of 1507 individuals showed a 25.43% general prevalence of MS without any significant difference between sexes, but a clear relation of the prevalence with progressing age (p=<0.0001). Even though both sexes showed similar prevalence rates, distribution of risk factors that defined MS was different between men and women, with the prevalence of hypertension, fasting hyperglycemia and hypertriglyceridemia being higher in men. Race was not an important risk factor for MS in this population as opposed to social economic class that was highly associated with the risk of MS in women as their social class was lower, but not in men. Conclusion: This cross-sectional study from a large urban population in Brazil showed a high general prevalence of MS (25.4%), which is increased as the population becomes older (especially in women) and poorer. Although prevalence was very similar in both genders, the frequency of components defining the syndrome varied greatly amongst them. In particular, a significant interaction between gender and social class was observed and may shed light in our understanding of the complex interplay between demographic and biological risk factors for metabolic syndrome. (C) 2007 Elsevier Ireland Ltd. All rights reserved.

International study of temperature, heat and urban mortality: the `ISOTHURM` project

Background This study describes heat- and cold-related mortality in 12 urban populations in low- and middle-income countries, thereby extending knowledge of how diverse populations, in non-OECD countries, respond to temperature extremes. Methods The cities were: Delhi, Monterrey, Mexico City, Chiang Mai, Bangkok, Salvador, So Paulo, Santiago, Cape Town, Ljubljana, Bucharest and Sofia. For each city, daily mortality was examined in relation to ambient temperature using autoregressive Poisson models (2- to 5-year series) adjusted for season, relative humidity, air pollution, day of week and public holidays. Results Most cities showed a U-shaped temperature-mortality relationship, with clear evidence of increasing death rates at colder temperatures in all cities except Ljubljana, Salvador and Delhi and with increasing heat in all cities except Chiang Mai and Cape Town. Estimates of the temperature threshold below which cold-related mortality began to increase ranged from 15 degrees C to 29 degrees C; the threshold for heat-related deaths ranged from 16 degrees C to 31C. Heat thresholds were generally higher in cities with warmer climates, while cold thresholds were unrelated to climate. Conclusions Urban populations, in diverse geographic settings, experience increases in mortality due to both high and low temperatures. The effects of heat and cold vary depending on climate and non-climate factors such as the population disease profile and age structure. Although such populations will undergo some adaptation to increasing temperatures, many are likely to have substantial vulnerability to climate change. Additional research is needed to elucidate vulnerability within populations.

Evaluation of the genotoxicity of treated urban sludge in the Tradescantia micronucleus assay

The sludge produced in sewage treatment plants can contain toxic substances. Among these, the genotoxic substances are of great concern. The present paper aimed at evaluating the genotoxicity of treated sludge samples collected at four different sewage treatment plants (STP) located in the State of Sao Paulo Brazil, using the Trad-MN assay. Another objective of the study was to compare the responses of the Clone #4430 with the Tradescantia pallida. Sludge samples mixed with reference soil in concentrations of 10, 25 and 50% (v/v) were tested in experiments with 3 months exposure in the field. Negative and positive controls (arsenic trioxide) were also tested with both plants. In Clone #4430 two sludge samples induced genotoxicity while in T pallida three were positive, although no clear dose-response were observed for both plants. Results with the negative and positive controls suggest that T pallida presented similar results when compared to the Clone #4430. The protocol using plants chronically exposed to sludge mixed with soil seems to be a promising tool to assess the genotoxicity of sludge although time consuming. (C) 2008 Elsevier B.V. All rights reserved.

Biomonitoring genotoxic risks under the urban weather conditions and polluted atmosphere in Santo Andre, SP, Brazil, through Trad-MCN bioassay

The present study was made to check if the Trad-MCN bioassay, developed with inflorescences of Tradescantia pallida cv. Purpurea, might discriminate genotoxic risks in areas of the city of Santo Andre (SE Brazil) contaminated by different air pollutants, and periods of the year when risks are higher, and to determine if the variations in the frequency of micronuclei (MCN) can be explained by environmental factors that characterize the stressful situation in each site. Potted plants were exposed in sites highly contaminated by ozone (Capuava and School) and in sites reached by high vehicular emissions (downtown and Celso Daniel Park). Pedroso Park, far from the polluted areas, was taken as reference. From September 2003 to September 2004, 20 young inflorescences were collected twice a week from each place and the frequencies of MCN were estimated. The environmental conditions observed in the polluted sites were stressful enough to promote an increase of MCN, mainly in sites reached by high vehicular emissions. But MCN rates in Capuava and at Celso Daniel Park could not be predicted only by pollutants which characterized the air contamination in these sites. More severe weather conditions, mainly low temperature, relative humidity and rainfall, caused an increase of MCN. Improvement of the biomonitoring system is recommended to minimize this negative influence of weather factors. (C) 2008 Elsevier Inc. All rights reserved.

Particulate urban air pollution affects the functional morphology of mouse placenta

in humans, adverse pregnancy outcomes (low birth weight, prematurity, and intrauterine growth retardation) are associated with exposure to urban air pollution. Experimental data have also shown that such exposure elicits adverse reproductive outcomes. We hypothesized that the effects of urban air pollution on pregnancy outcomes could be related to changes in functional morphology of the placenta. To test this, future dams were exposed during pregestational and gestational periods to filtered or nonfiltered air in exposure chambers. Placentas were collected from near-term pregnancies and prepared for microscopical examination. Fields of view on vertical uniform random tissue slices were analyzed using stereological methods. Volumes of placental compartments were estimated, and the labyrinth was analyzed further in terms of its maternal vascular spaces, fetal capillaries, trophoblast, and exchange surface areas. From these primary data, secondary quantities were derived: vessel calibers (expressed as diameters), trophoblast thickness (arithmetic mean), and total and mass-specific morphometric diffusive conductances for oxygen of the intervascular barrier. Two-way analysis of variance showed that both periods of exposure led to significantly smaller fetal weights. Pregestational exposure to nonfiltered air led to significant increases in fetal capillary surface area and in total and mass-specific conductances. However, the calibers of maternal blood spaces were reduced. Gestational exposure to nonfiltered air was associated with reduced volumes, calibers, and surface areas of maternal blood spaces and with greater fetal capillary surfaces and diffusive conductances. The findings indicate that urban air pollution affects placental functional morphology. Fetal weights are compromised despite attempts to improve diffusive transport across the placenta.

Effect of pre- and postnatal exposure to urban air pollution on myocardial lipid peroxidation levels in adult mice

Exposure to air pollution can elicit cardiovascular health effects. Children and unborn fetuses appear to be particularly vulnerable. However, the mechanisms involved in cardiovascular damage are poorly understood. It has been suggested that the oxidative stress generated by air pollution exposure triggers tissue injury. To investigate whether prenatal exposure can enhance oxidative stress in myocardium of adult animals, mice were placed in a clean chamber (CC, filtered urban air) and in a polluted chamber (PC, Sao Paulo city) during the gestational period and/or for 3 mo after birth, according to 4 protocols: control group-prenatal and postnatal life in CC; prenatal group-prenatal in PC and postnatal life in CC; postnatal group-prenatal in CC and postnatal life in PC; and pre-post group-prenatal and postnatal life in PC. As an indicator of oxidative stress, levels of lipid peroxidation in hearts were measured by malondialdehyde (MDA) quantification and by quantification of the myocardial immunoreactivity for 15-F2t-isoprostane. Ultrastructural studies were performed to detect cellular alterations related to oxidative stress. Concentration of MDA was significantly increased in postnatal (2.45 +/- 0.84 nmol/mg) and pre-post groups (3.84 +/- 1.39 nmol/mg) compared to the control group (0.31 +/- 0.10 nmol/mg) (p < .01). MDA values in the pre-post group were significantly increased compared to the prenatal group (0.71 +/- 0.15 nmol/mg) (p = .017). Myocardial isoprostane area fraction in the pre-post group was increased compared to other groups (p <= .01). Results show that ambient levels of air pollution elicit cardiac oxidative stress in adult mice, and that gestational exposure may enhance this effect.

Effects of semen storage and separation techniques on sperm DNA fragmentation

Objective: To determine the effect of semen storage and separation techniques on sperm DNA fragmentation. Design: Controlled clinical study. Setting: An assisted reproductive technology laboratory. Patient(s): Thirty normoozospermic semen samples obtained from patients undergoing infertility evaluation. Intervention(s): One aliquot from each sample was immediately prepared (control) for the sperm chromatin dispersion assay (SCD). Aliquots used to assess storage techniques were treated in the following ways: snap frozen by liquid nitrogen immersion, slow frozen with Tris-yolk buffer and glycerol, kept on ice for 24 hours or maintained at room temperature for 4 and 24 hours. Aliquots used to assess separation techniques were processed by the following methods: washed and centrifuged in media, swim-up from washed sperm pellet, density gradient separation, density gradient followed by swim-up. DNA integrity was then measured by SCD. Main Outcome Measure(s): DNA fragmentation as measured by SCD. Result(s): There was no significant difference in fragmentation among the snap frozen, slow frozen, and wet-ice groups. Compared to other storage methods short-term storage at room temperature did not impact DNA fragmentation yet 24 hours storage significantly increased fragmentation. Swim-up, density gradient and density gradient/swim-up had significantly reduced DNA fragmentation levels compared with washed semen. Postincubation, density gradient/swim-up showed the lowest fragmentation levels. Conclusion(s): The effect of sperm processing methods on DNA fragmentation should be considered when selecting storage or separation techniques for clinical use. (Fertil Steril (R) 2010;94:2626-30. (C) 2010 by American Society for Reproductive Medicine.)

Chronic exposure to ambient levels of urban particles affects mouse lung development

Rationale- Chronic exposure to air pollution has been associated with adverse effects on children`s lung growth. Objectives: We analyzed the effects of chronic exposure to urban levels of particulate matter (PM) on selected phases of mouse lung development. Methods: The exposure occurred in two open-top chambers (filtered and nonfiltered) placed 20 m from a street with heavy traffic in Sao Paulo, 24 hours/day for 8 months. There was a significant reduction of the levels of PM(2.5) inside the filtered chamber (filtered = 2.9 +/- 3.0 mu g/m(3), nonfiltered = 16.8 +/- 8.3 mu g/m(3); P = 0.001). At this exposure site, vehicular sources are the major components of PM(2.5) (PM <= 2.5 mu m). Exposure of the parental generation in the two chambers occurred from the 10th to the 120th days of life. After mating and birth of offspring, a crossover of mothers and pups occurred within the chambers, resulting in four groups of pups: nonexposed, prenatal, postnatal, and pre+postnatal. Offspring were killed at the age of 15 (n = 42) and 90 (n = 35) days; lungs were analyzed by morphometry for surface to volume ratio (as an estimator of alveolization). Pressure-volume curves were performed in the older groups, using a 20-ml plethysmograph. Measurements and Main Results: Mice exposed to PM(2.5) pre+postnatally presented a smaller surface to volume ratio when compared with nonexposed animals (P = 0.036). The pre+postnatal group presented reduced inspiratory and expiratory volumes at higher levels of transpulmonary pressure (P = 0.001). There were no differences among prenatal and postnatal exposure and nonexposed animals. Conclusions: Our data provide anatomical and functional support to the concept that chronic exposure to urban PM affects lung growth.

Candidate-Gene Approach in Posttraumatic Stress Disorder After Urban Violence: Association Analysis of the Genes Encoding Serotonin Transporter, Dopamine Transporter, and BDNF

Posttraumatic stress disorder (PTSD) is a prevalent, disabling anxiety disorder marked by behavioral and physiologic alterations which commonly follows a chronic course. Exposure to a traumatic event constitutes a necessary, but not sufficient, factor. There is evidence from twin studies supporting a significant genetic predisposition to PTSD. However, the precise genetic loci still remain unclear. The objective of the present study was to identify, in a case-control study, whether the brain-derived neurotrophic factor (BDNF) val66met polymorphism (rs6265), the dopamine transporter (DAT1) three prime untranslated region (3`UTR) variable number of tandem repeats (VNTR), and the serotonin transporter (5-HTTPRL) short/long variants are associated with the development of PTSD in a group of victims of urban violence. All polymorphisms were genotyped in 65 PTSD patients as well as in 34 victims of violence without PTSD and in a community control group (n = 335). We did not find a statistical significant difference between the BDNF val66met and 5-HTTPRL polymorphism and the traumatic phenotype. However, a statistical association was found between DAT1 3`UTR VNTR nine repeats and PTSD (OR = 1.82; 95% CI, 1.20-2.76). This preliminary result confirms previous reports supporting a susceptibility role for allele 9 and PTSD.