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Selenium is an element present in trace amounts and different chemical forms. It may exert both beneficial and adverse effects on cellular redox status and on the generation of reactive oxygen species. 8-oxo-7,8-dihydro-2'deoxyguanosine (8-oxodG) is an oxidized derivative of deoxyguanosine, and a sensitive biomarker of oxidative stress and genotoxicity. The present study assessed the extent to which selenium status was associated with urinary 8-oxodG concentrations in a Northern Italian population. We recruited healthy, non-smoking blood donors living in the Reggio Emilia province during 2017-2019. We measured urinary 8-oxodG concentrations and used restricted cubic spline regression analyses to investigate the association between selenium status (estimated using food frequency questionnaires, urinary concentrations, and serum concentrations of selenium and selenium species) and 8-oxodG/g creatinine. Among 137 participants aged 30-60 years, median urinary selenium and 8-oxodG concentrations were 22.02 μg/L and 3.21 μg/g creatinine, respectively. Serum samples and selenium speciation analyses were available for 104 participants. Median total serum selenium levels and dietary intake were 116.5 μg/L and 78.7 μg/day, respectively. In spline regression analysis, there was little association between dietary, serum, or urinary selenium with 8-oxodG concentrations. In sex-specific analyses, urinary selenium showed a positive association with the endpoint among males. For single selenium species, we observed positive associations with urinary 8-oxodG for serum organic selenium species, and negative associations for inorganic selenium forms. In the most adjusted analysis, urinary 8-oxodG concentrations showed a strong positive association with selenomethione-bound selenium (Se-Met) and a negative association with inorganic tetravalent selenium, selenite. In sex-specific analyses, these associations were considerably stronger in males than in females. Overall, study findings indicate that selenium species exhibited very different patterns of associations with the biomarker of oxidative stress, and that these associations also depended on sex. Background exposure to Se-Met appears to be strongly and positively associated with oxidative stress., Competing Interests: Declaration of competing interest Dr. Lauren A. Wise is supported by grants from the National Institutes of Health. She also serves as a fibroid consultant on for AbbVie, Inc. She also receives in-kind donations for primary data collection in Pregnancy Study Online (PRESTO) from Swiss Precision Diagnostics (home pregnancy tests) and Kindara.com (fertility apps). All of these relationships are for work unrelated to this manuscript. The remaining authors declare no conflict of interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

An ever-growing burden of scientific evidence links air pollution to different aspects of human health even at very low concentrations; the impact increases for those living in urban environments, especially the youngest and the elderly. This study investigated the exposure to air pollution of urban school children of Milan, Italy, by personal and biological monitoring, in the frame of the MAPS-MI project. A total of 128 primary school children (7-11 years) were involved in a two-season monitoring campaign during spring 2018 and winter 2019. Personal exposure to airborne VOCs and eBC, and biological monitoring of urinary benzene (BEN-U) and methyl-tert-butyl ether (MTBE-U) were performed. Time-activity patterns, environmental tobacco smoke (ETS), spatial, and meteorological information were evaluated as determinants in mixed effects regression analysis. Children personal exposure was mostly quantifiable with median (5th-95th percentile) levels 1.9 (0.8-7.5) μg/m 3 for eBC, and 1.1 (<0.6-3.4) and 0.8 (0.3-1.8) μg/m 3 for benzene and MTBE, respectively; with values 2-3-fold higher in winter than in spring. In urine, median (5th-95th) BEN-U and MTBE-U levels were 44.9 (25.7-98.6) and 11.5 (5.0-35.5) ng/L, respectively. Mixed effect regression models explained from 72 to 93 % of the total variability for air pollutants, and from 58 to 61 % for biomarkers. Major contributors of personal exposure were season, wind speed, mobility- or traffic-related variables; biomarkers were mostly predicted by airborne exposure and ETS. Our results suggest that traffic-mitigation actions, together with parents' educational interventions on ETS and commuting mode, should be undertaken to lower children exposure to air pollution., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Objectives: Slowness, generally assessed by walking speed (WS), is an estimator of frailty and its outcomes. Because of potential difficulties in assessing WS, the Moberg picking-up test (MPUT) might be an alternative. This study investigated the capacity of slowness measurements (WS and MPUT) to predict nonfatal adverse consequences of frailty: primarily, decline in basic activities of daily living (BADL); and secondarily, decline in instrumental activities of daily living (IADL), fall, hospitalization, and incident disease., Design: Observational (prospective longitudinal study)., Setting and Participants: This study used data from the population-based Lausanne cohort 65+. At baseline, 1887 individuals (aged 72-77 years) completed both WS (time to walk 20 m at usual pace) and MPUT (time to pick up 12 objects) assessments., Methods: All outcomes, assessed at 1- and 4-year follow-ups, were entered in separate logistic regression models with adjustment for age, sex, and respective values at baseline. The prediction of all outcomes by either WS or MPUT was assessed using area under the receiver operating characteristic curve and compared by χ 2 tests., Results: There were positive associations between slowness either assessed by WS [relative risk (RR) = 2.48; P < .001] or MPUT (RR = 1.91; P < .001) and decline in BADL at 1-year follow-up. These associations remained significant at 4-year follow-up for both WS (RR = 2.28; P < .001) and MPUT (RR = 1.95; P < .001). There was no significant difference between predictive values of slow WS and MPUT for decline in BADL at 1-year (P = .328) and 4-year follow-ups (P = .413). The prediction was not significantly different for secondary outcomes, except for decline in IADL for which the prediction was slightly better for WS., Conclusions and Implications: MPUT may be an alternative measurement of slowness with predictive value of functional decline. No significant difference in predictive capabilities of MPUT and WS for specific adverse consequences of frailty is promising in favor of using MPUT for measuring slowness., (Copyright © 2022 AMDA – The Society for Post-Acute and Long-Term Care Medicine. Published by Elsevier Inc. All rights reserved.)

Cadmium is a heavy metal with established adverse effects on human health, namely on bone, liver and kidney function and the cardiovascular system. We assessed cadmium exposure and its correlation with biomarkers of toxicity. We recruited 137 non-smoking blood donors without a history of chronic disease or cancer who resided in the Northern Italy province of Reggio Emilia (mean age 47 years, range 30-60 years) in the 2017-2019 period. We used a semi-quantitative food frequency questionnaire to estimate dietary cadmium intake and urine samples to assess concentrations of urinary cadmium and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG). Median urinary cadmium and 8-oxodG concentrations were 0.21 μg/L (interquartile range (IQR): 0.11-0.34 μg/L) and 3.21 μg/g creatinine (IQR: 2.21-4.80 μg/g creatinine), respectively, while median dietary cadmium intake was 6.16 μg/day (IQR: 5.22-7.93 μg/day). We used multivariable linear and spline regression models to estimate mean differences exposure concentrations. Dietary and urinary cadmium were positively correlated, and both were positively and linearly correlated with 8-oxodG. We found a positive association of urinary cadmium with blood alanine aminotransferase (ALT), total cholesterol, low-density lipoprotein (LDL)-cholesterol and thyroid-stimulating hormone (TSH) concentrations. We also observed a positive association with triglycerides, in both linear (beta regression coefficient = 77.03, 95% confidence interval 32.27-121.78) and non-linear spline regression analyses. Despite the positive correlation between dietary and urinary cadmium estimates, dietary cadmium intake showed inconsistent results with the study endpoints and generally weaker associations, suggesting a decreased capacity to reflect actual cadmium exposure. Overall, these findings suggest that even low levels of cadmium exposure may adversely alter hematological and biochemical variables and induce oxidative stress., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Introduction: Children with smoking parents are potentially exposed to Environmental Tobacco Smoke (ETS). The aims of this study were: 1) to assess ETS exposure in Milan schoolchildren, by measuring urinary cotinine (COT-U), 2) to compare the parents' perception of children ETS exposure, with the actual ETS exposure measured by COT-U, 3) to explore the factors influencing COT-U, including smoking bans at home, the season, and children characteristics., Methods: One-hundred school children (7-11 years) and their parents were recruited for the study in Spring 2018 (n = 81) and in Winter 2019 (n = 94), 75 children participated to both campaigns, for a sum of 175 observations. A questionnaire was submitted to parents to collect information about smoking habits in the house. COT-U was measured by LC-MS/MS in spot urine sample collected in the morning from children., Results: Detectable COT-U levels were found in 42% and 57% of children, in spring and winter, in contrast with 17% and 13% of parents acknowledging ETS exposure. Children living with smokers or e-cigarette users (vapers) (30% of the participants) had higher COT-U levels than children not living with smokers or vapers (median 0.67, 0.46, and <0.1 μg/L in spring, and 0.98, 0.85, and 0.11 μg/L in winter, respectively). Increasingly higher COT-U levels were observed in children living in homes where smoking was completely banned, allowed in the external parts of the home, or allowed in some rooms. The multiple regression analysis confirmed the positive significant effect of living with smokers, a partial smoking ban and absence of smoking ban at home, the winter season, and BMI as determinants of COT-U., Conclusion: ETS exposure resulted in measurable urinary cotinine in children. Smoking parents underestimate exposure to ETS of their children. Living with smokers is a determinant of COT-U, only slightly mitigated by adopting partial smoking ban., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Objectives: Slow walking speed (WS) is predictive of mortality but may be difficult to measure, which compromises the assessment of frailty, based on Fried et al's phenotype. The timed Moberg picking-up test (MPUT), developed to evaluate hand's function, was found moderately but significantly correlated with WS. We compared the relationship between slowness, assessed by MPUT and WS tests, and mortality., Design: Observational (prospective cohort study)., Setting and Participants: 4731 community-dwelling adults included in 2004, 2009, or 2014 in the ongoing Lausanne cohort 65+ (Lc65+) were assessed at the age of 66-71 years., Method: Mortality was compared for individuals above and below percentile 80 of MPUT, and respectively WS performance time, according to the Fried criterion. Multivariable analysis using Cox's regression models were adjusted for age, sex, height and grip strength. The predictive capability of MPUT and WS was assessed in adjusted models using Harrell C., Results: Slowness in MPUT and in WS test was associated with mortality at 4, 9, and 14 years (P < .001). Survival curves showed lower survival rates in the highest percentile for both tests (P < .001), regardless of the follow-up period. Cox models indicated a higher risk of death at 4 years [adjusted hazard ratio (95% confidence interval): MPUT, 2.1 (1.5-3.0); WS, 2.2 (1.5-3.1)], 9 years [MPUT 1.7 (1.3-2.3); WS 2.0 (1.5-2.6)] and 14 years [MPUT 1.8 (1.4-2.3); WS 1.8 (1.4-2.4)] for participants above the 80th percentile (all P < .001). The 2 tests had similar predictive capability (Harrell C: MPUT, between 61% and 68%; WS, between 62% and 69%)., Conclusions and Implications: Poor performance in MPUT is associated with increased mortality at the short and long term among community-dwelling older adults. This alternative to WS in the assessment of slowness has similar predictive capability for mortality and avoids biased estimates because of nonrandom exclusion of individuals unable to complete WS., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Human exposure to air pollutants, and specifically to particulate matter (PM) and volatile organic compounds (VOCs), may pose a relevant risk on human health., Aim: To evaluate the personal exposure of adults living and working in Milan (Italy) by environmental and biological monitoring., Methods: Personal exposure of 51 volunteer adults to PM 2.5 , PM 2.5 - 10 and selected VOCs, including benzene, toluene, ethylbenzene, o-xylene, m + p-xylene, methyl tert-butyl ether, naphthalene, hexane, cyclohexane, heptane, and limonene was assessed along a 24-h period via personal cascade impactors and radial diffusive samplers. Urine spot samples were collected to investigate the corresponding urinary biomarkers. Time-activity patterns were filled in by participants to explore the performed activities. Multiple regression models were applied to investigate the association between personal exposure, biomarker levels, and tobacco smoke, traffic exposure, commuting mode, cooking activities, and personal characteristics., Results: Median personal exposure to PM 2.5 , PM 2.5 - 10 , benzene, toluene, ethylbenzene o-xylene, m + p-xylene, methyl tert-butyl ether, naphthalene, hexane, cyclohexane, heptane, and limonene were 36.1, 7.8, 2.3, 7.8, 2.1, 1.8, 4.7, 0.8, 0.3, 1.4, 2.5, 1.6, and 59.9 μg/m 3 , respectively. Median levels of urinary benzene, toluene, ethylbenzene o-xylene, m + p-xylene, naphthalene, hexane, and heptane were 78.0, 88.1, 21.5, 15.2, 43.9, 21.0, 11.0, and 22.5 ng/L, respectively. For personal exposure, multiple regression models explained up to 67% (PM 2.5 ) and 61% (benzene) of variability, with major contribution from commuting mode and environmental exposure. For biological monitoring, multiple regression analysis explained up to 74% of urinary benzene, with a major contribution given by creatinine, and secondary contributions by commuting mode, personal exposure to airborne benzene and smoking., Conclusions: Personal exposure to air pollutants was lower than that measured in the past in Milan. Personal exposure was mainly driven by traffic variables, while internal dose was mainly driven by personal characteristics and smoking habit., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Objectives: Fear of falling (FOF) is common in older people and is related to negative outcomes. This study aimed to investigate whether 2 different instruments, the Falls Efficacy Scale-International (FES-I) and the single question on FOF and activity restriction (SQ-FAR), were associated with incident disability at 3 years., Design: Prospective observational study., Setting and Participants: Participants (n = 1219, 57.4% women) were disability-free community-dwelling persons enrolled in the Lausanne cohort 65+, aged 66 to 71 years, in 2005., Measures: Baseline covariates included demographic, cognitive, affective, and health status. Basic activities of daily living (BADL) assessment was recorded annually from a self-administered questionnaire. Disability outcome was defined as reporting difficulty or help needed in ≥1 of 5 BADL in ≥2 consecutive years, or being institutionalized during follow-up., Results: At 3 years, disability was reported by 77 participants (6.3%). Reporting the highest level of fear at FES-I [adjusted odds ratio (aOR) 5.14, 95% confidence interval (CI) 1.82-14.55, P = .002] or "FOF with activity restriction" with SQ-FAR (aOR 3.23, 95% CI 1.29-8.08, P = .012) were both associated with increased odds of disability even after adjusting for covariates. The FES-I model explained incident disability slightly better than the SQ-FAR one [Bayesian information criterion (BIC) values of 466.70 and 469.43, respectively]., Conclusions and Implications: High FOF and related activity restriction, assessed with FES-I and SQ-FAR, are associated with incident disability in young-old community-dwelling people. The SQ-FAR is suitable as a screening tool to proactively detect a potentially reversible risk factor for disability. Using the FES-I may serve additional clinical purposes, such as FOF characterization and management., (Copyright © 2020 AMDA – The Society for Post-Acute and Long-Term Care Medicine. Published by Elsevier Inc. All rights reserved.)

Pesticides are chemicals widely applied in agriculture and proven environmental contaminants; their hazards include harmful effects on human health, therefore the evaluation of exposure is relevant to risk assessment. Hair is a non-invasive specimen that incorporates pollutants allowing an extended exposure window to be surveyed. Aim of this work was to develop and validate an assay for measuring 41 pesticide active principles in human hair. Under optimised conditions, analytes were extracted by soaking hair in acetonitrile, in the presence of internal standards, under stirring and heating condition. Chemical separation was achieved using liquid chromatography with silica-based bonded phase chromatographic column. Detection and quantification were performed, with both positive and negative electrospray ionization, by a hybrid triple quadrupole/linear ion trap mass spectrometer operating in the scheduled selected reaction monitoring mode. The validated assay showed a linear dynamic range up to 10000 ng/L or 400 pg/mg hair, inter- and intra-run precisions <7%, and accuracies within 10% of theoretical concentrations. Limits of quantification were 1 ng/L or 0.04 pg/mg hair for most of the investigated pesticides. Matrix effect experiments showed that the use of internal standards allowed for the control of biases. The method was applied to the determination of pesticides in hair samples form occupationally and non-occupationally exposed individuals. The results of this study indicate that the developed assay is useful to assess pesticides in human hair following different exposure scenarios., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Objectives: Slowness is a marker of frailty captured by the Fried phenotype by a walking speed test which, for health or logistical reasons, is sometimes difficult to perform. The Moberg picking-up test (MPUT) is another timed functional test. It measures hand motor activity and might represent an alternative to assess slowness when the walking speed cannot be evaluated. This study aimed to evaluate the relationship between MPUT and walking speed., Design: Cross-sectional., Setting and Participants: In total, 2748 individuals aged 66 to 83 years who participated in the latest examination (2015-2017) of the population-based Lausanne cohort 65+ and completed both tests., Methods: Walking speed (time to walk 20 meters at usual pace) and MPUT (time to pick up 12 objects) were compared using scatter graphs. Multivariate regression models further investigated the relationship between MPUT and walking times with adjustment for height, grip strength, body mass index, and Mini-Mental State Examination. All analyses were stratified by sex., Results: MPUT and walking times were moderately, positively correlated in men (r = 0.38, P < .001) and in women (r = 0.38, P < .001). Higher grip strength and Mini-Mental State Examination performances were correlated to shorter MPUT and walking times. Men and women slower at the MPUT were also significantly slower at the walking speed test when adjusting for height (P < .001) as well as in fully adjusted models (P < .001)., Conclusions and Implications: These preliminary results point to a positive association between MPUT and walking speed independent of muscle strength and cognition. Further research is needed to investigate the capacity of MPUT to predict adverse health outcomes before considering this test as an alternative measure of slowness in the assessment of frailty., (Copyright © 2020. Published by Elsevier Inc.)

This study reports the development of a method based on headspace (HS)-solid-phase microextraction (SPME)-gas chromatography (GC)-triple quadrupole tandem mass spectrometry (MS/MS) for the quantification of 2- to 4-ring polycyclic aromatic hydrocarbons (PAHs) in saliva samples. Eight unmetabolized compounds (naphthalene, acenaphthylene, acenaphthene, fluorene, phenanthrene, anthracene, fluoranthene and pyrene) were quantified using six deuterated PAHs as surrogate internal standards. The absence of matrix effect allowed saliva samples to be quantified by external calibration method. The optimized method resulted easy, with minimal sample pre-treatment (homogenization of the sample), and it achieved the highest sensitivity up to date: limits of quantification (LOQ) were in the 0.8-26.4 ng L -1 range, with a significant improvement in comparison with the few existing methods. Intra- and inter-run precisions provided CV values <18.1%, and accuracies within 20% of the spiked concentration. The application of the method to the analysis of fresh saliva samples collected by spitting from smokers (n = 10) and non-smokers (n = 10) showed that PAHs were quantifiable in all samples and that smokers had higher levels of all compounds than non-smokers. These results show that the method is suitable for quantifying low-boiling PAHs in saliva samples from individuals exposed at different PAH levels., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: Although thousands of different chemicals have been identified in cigarette smoke, the characterization of their urinary metabolites still requires significant research. The aim of this work was to perform an untargeted metabolomic approach to a pilot cross-sectional study conducted on subjects with different smoking habits and to compare the results with those of the targeted measurement of mercapturic acids., Methods: Urine samples from 67 adults, including 38 non-smokers, 7 electronic cigarette users, and 22 traditional tobacco smokers were collected. Samples were analysed by liquid chromatography/time-of flight mass spectrometry. Data were processed using the R-packages IPO and XCMS to perform feature detection, retention time correction and alignment. One-way ANOVA test was used to identify different features among groups. Quantitative determination of 17 mercapturic acids was available from a previous study., Results: One hundred and seventeen features, out of 3613, were different among groups. They corresponded to 91 potential metabolites, 5 of which were identified vs authentic standards, 43 were putatively annotated and 13 were attributed to chemical classes. Among identified compounds there were the mercapturic acids of acrolein, 1,3-butadiene, and crotonaldehyde; among putatively annotated compounds there were the glucuronide conjugated of 3-hydroxycotinine and the sulfate conjugate of methoxyphenol; with the lowest degree of confidence several sulfate conjugates of small molecules were annotated. Considering mercapturic acids, the coherence between the targeted and untargeted approach was found for a limited number of chemicals, typically the most abundant., Conclusions: Differences in the urinary levels of several compounds were associated to the different smoking habits, suggesting that the proposed approach is useful for the investigation of the metabolite patterns related to the exposure to toxicants. However, limitations were highlighted, in particular regarding the identification of low concentration compounds., Competing Interests: Declaration of Competing Interest The authors report no declarations of interest., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: While tobacco smoke contains thousands of chemicals, some of which are carcinogenic to humans, the content of electronic cigarette smoke is less known. This work aimed to assess and compare the exposure associated with different smoking habits by profiling urinary mercapturic acids as biomarkers of toxic compounds., Methods: In this pilot study, sixty-seven healthy adults with different smoking habits were investigated: 38 non-smokers (NS), 7 electronic cigarette users (ECU), and 22 traditional tobacco smokers (TTS). Seventeen urinary mercapturic acids, metabolites of 1,3-butadiene (DHBMA, MHBMA), 4-chloronitrobenze (NANPC), acrolein (3-HPMA), acrylamide (AAMA, GAMA), acrylonitrile (CEMA), benzene (SPMA), crotonaldehyde (CMEMA, HMPMA), ethylating agents (EMA), methylating agents (MMA), ethylene oxide (HEMA), N,N-dimethylformamide (AMCC), propylene oxide (2-HPMA), styrene (PHEMA), and toluene (SBMA), were quantified, along with urinary nicotine and cotinine., Results: Median urinary cotinine was 0.4, 1530 and 1772 μg/L in NS, ECU and TTS, respectively. Most mercapturic acids were 2-165 fold-higher in TTS compared to NS, with CEMA, MHBMA, 3-HPMA and SPMA showing the most relevant increases. Furthermore, some mercapturic acids were higher in ECU than NS; CEMA and 3-HPMA, in particular, showed significant increases and were 1.8 and 4.9 fold-higher, respectively., Conclusions: This study confirms that tobacco smoking is a major source of carcinogenic chemicals such as benzene and 1,3-butadiene; electronic cigarette use is a minor source, mostly associated with exposure to chemicals with less carcinogenic potential such as acrylonitrile and acrolein., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Introduction: Vineyard is a crop where a large number of pesticides are applied; exposure to pesticides may occur in farmers and the general population living close to the treated area. This work aimed to investigate hair as a matrix for the assessment of cumulative and aggregate exposure to pesticides in potentially exposed individuals., Methods: Twenty agricultural workers (AW), 4 agricultural worker relatives (AR), and 5 research staff members (RS) were involved in the study. Hair samples were collected before and after the application season (PRE- and POST-EXP samples) to obtain 18 paired samples. Records with the name and the quantity of applied pesticides were obtained; twenty-seven pesticides were measured in hair by solvent extraction and LC-MS/MS., Results: During the study season, AW applied 14 different pesticides with median amount ranging from 12 to 7200 g. The most popular pesticides were dimethomorph, penconazole, cyazofamid, fenamidone and quinoxyfen, applied from 94 to 69% of AW. In AW, in PRE-EXP samples the majority of used pesticides was detectable (with detection rates from 6 to 88%), with median concentrations of few pg/mg hair; in the POST-EXP samples the frequency of detected values increased (from 25 to 100%), with median concentrations up to two orders of magnitude higher. In AR, most pesticides were quantifiable only in POST-EXP samples and with lower concentration in comparison with AW; in RS, in both PRE- and POST-EXP samples only a few pesticides were quantifiable with very low levels. In AW, a linear correlation (r = 0.682 on log-transformed data, p < 0.01) was found between the total amounts of applied pesticides during the season and their concentration in hair., Conclusion: The study shows that the majority of assessed pesticides was incorporated into hair of AW and AR. The increased frequency of detection and level at the end of the season and the correlation between pesticide in hair and the amount of applied pesticides, reinforce the use of hair for quantitative biomonitoring of cumulative exposure to pesticides., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Introduction: The European Environment Agency has identified Northern Italy as one of the most polluted areas in Europe. Among air contaminants, black carbon (BC) has been identified as a sensitive marker of traffic related air pollution. This study aims to investigate the spatial distribution of BC in the catchment area of an elementary school of Milan, the biggest city in Northern Italy, using Land Use Regression (LUR) models and focusing especially on Morning Rush Hour (MRH)., Methods: Two recruitment campaigns were performed asking schoolchildren's parents and residents of the study area to host a monitoring site in their own dwellings. Finally, 34 monitoring sites and 1 reference site were sampled. BC was measured in two seasonal campaigns using eight micro-aethalometers. Six seasonal and annual LUR models were developed, 3 focused on MRH., Results: Overall, median BC was 3247 and 1309 ng/m 3 in the cold and warm season, respectively. In both seasons, there was a significant spatial variation between the monitoring sites. MRH values were higher than the daily values with median concentrations of 4227 and 2331 ng/m 3 , respectively. Developed LUR models showed that BC variability is well explained only by traffic variables; R 2 ranged from 0.52 to 0.79 and from 0.65 to 0.81, for seasonal/annual and MRH LUR models respectively., Discussion: LUR models based on traffic variables explain most of the measured BC distribution variability for both warm and cold season. MRH represents a critical moment for BC during all the year, with an increase of 1000 ng/m 3 respective to the daily median value and differences in magnitude according to location. Our results highlight that the mobility issue is one of the most important challenges to reduce air pollution in the city of Milan and this is of particular concern for elementary schoolchildren that commute to school during MRH., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Introduction: Mercapturic acids are urinary metabolites of occupational and environmental toxicants. The aim of this work was to develop and validate an analytical assay for the determination of several urinary mercapturic acids to be used as biomarkers of exposure., Method: An isotope dilution tandem mass spectrometric method, coupled with reversed-phase liquid chromatography, was developed for the analysis of mercapturic acids derived from several compounds, including those of benzene, toluene, 1,3-butadiene, styrene, acrylonitrile, 4-chloronitrobenzene, acrylamide, acrolein, propylene oxide, N,N-dimethylformamide, crotonaldehyde, ethylene oxide, and methylating and ethylating agents. Samples were prepared by simple filtration after dilution. A validation was carried out, including the assessment of calibration curves, sensitivity, accuracy, precision, process efficiency, and stability, along with external verification. The assay was applied to the analysis of 14 end-of-shift urine samples from unexposed workers and gasoline station attendants., Results: The chromatographic run lasted 18 min. Limits of quantitation ranged from 0.01 to 3.2 μg/L; precision, expressed as relative standard deviation, ranged from 0.6 to 20.9%; and accuracy ranged from 93.4 to 114.9% of theoretical values. The use of deuterated internal standards was suitable for control of the matrix effect. The assay allowed the simultaneous quantitation of urinary mercapturic acids at different concentration ranges. The external verification exercise produced good results. The application of the assay to urine samples from workers revealed differences in mercapturic acid profiles in agreement with the expected patterns of exposure., Conclusion: This high-throughput method is valid and useful for the quantitation of urinary mercapturic acids, and is suitable for human biomonitoring of occupational and environmental exposure., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Penconazole (PEN) is a fungicide used in agriculture. The aim of this work was to evaluate the exposure to PEN in vineyard workers focusing on urinary biomarkers. Twenty-two agricultural workers were involved in the study; they were investigated during PEN applications and re-entry work, performed for 1-4 consecutive working days, for a total of 42 mixing and applications and 12 re-entries. Potential and actual dermal exposure, including hand exposure, were measured using pads and hand washes. Urine samples were collected starting before the first application, continuing during the work shift, and ending 48 h after the last shift. The determination of PEN in dermal samples and PEN metabolites in urine was performed by liquid chromatography tandem mass spectrometry. Dermal potential body exposure and actual total exposure showed median levels ranging from 18 to 3356µg and from 21 to 111 µg, respectively. Urinary monohydroxyl-derivative PEN-OH was the most abundant metabolite; its excretion rate peaked within 24 h after the work shift. In this period, median concentrations of PEN-OH and the carboxyl-derivative PEN-COOH ranged from 15.6 to 27.6 µg/L and from 2.5 to 10.2 µg/L, respectively. The concentration of PEN-OH during the work shift, in the 24 h after and in the 25-48 h after the work shift were correlated with actual body and total dermal exposure (Pearson's r from 0.279 to 0.562). Our results suggest that PEN-OH in the 24 h post-exposure urine is a promising candidate for biomonitoring PEN exposure in agricultural workers., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Penconazole (PEN) and tebuconazole (TEB) are fungicides widely used in vineyards. The aim of this the study was to assess the suitability of hair to assess long-term exposure to PEN and TEB. Hair samples of agricultural workers exposed to PEN (AW-PEN, 18 subjects) or TEB (AW-TEB, 2 subjects) during the application of fungicides, agricultural workers relatives (AR, 4 subjects), and research staff technicians (RS, 5 subjects) were collected before (PRE-EXP) and after (POST-EXP) the application season. PEN in PRE-EXP samples was quantifiable in all AW and AR (medians from 1.4 to 7.9 pg/mg hair) and in one RS (1.4 pg/mg hair); PEN in POST-EXP samples was always quantifiable (medians from 2.6 to 23.7 pg/mg hair), with higher levels in AW. Comparing PRE- vs. POST-EXP samples, an increase in PEN level in AW and RS was found. TEB in PRE-EXP samples was quantifiable in most AW and AR (median from 2.1 to 15.5 pg/mg hair), but not in RS; TEB in POST-EXP samples was similarly quantifiable in AW and AR, and was quantifiable also in RS (from 1.4 to median of 141.3 pg/mg hair). Comparing PRE- vs. POST-EXP samples, an increase in TEB level in AW and RS was found. In AW, a positive correlation between the number of PEN treatments during the season and the POST-EXP level of PEN in hair was found (N = 8, Spearman rho = 0.794, p = 0.019). Our results suggest that PEN and TEB accumulate in hair during the agricultural season and that hair is a promising matrix for biomonitoring long-term exposure., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The aim of this study was to measure lead (Pb) levels in blood (Pb-blood) in consumers of game meat, taking into account other possible sources of lead exposure. A spot blood sample was obtained from 95 subjects, and a questionnaire was used to collect general information and data on game meat consumption, hunting, wine drinking and other possible sources of lead exposure. Pb-blood was measured by inductively coupled plasma-mass spectrometry. Pb-blood was not influenced by age, sex, residence in an urban or rural area, consumption of game meat, tobacco smoking or hobbies associated with potential exposure to lead, and median Pb-blood was 1.7 (5th-95th percentile 1.0-5.3)µg/dL and 3.4 (0.9-6.1)µg/dL for game meat non-eaters and eater, respectively. A multiple linear regression analysis (containing the covariates sex, age, hunting, wine drinking, game meat consumption, tobacco smoking, shooting range, and occupational exposure) found an association with hunting (Pb-blood almost double in hunters) and wine drinking (40% higher in drinkers) but not with consumption of game meat or other parameters. Whether the higher Pb-blood level was due to inhalation of lead fumes while shooting with lead ammunition, to handling lead ammunition or both could not be ascertained., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The aims of this study were (1) to explore the behavioral and sociodemographic factors influencing urinary cotinine (COT-U) levels in active smokers and in environmental tobacco smoke (ETS)-exposed individuals, (2) to assess the specificity and sensitivity of the questionnaire for identifying active smokers and nonsmokers, and (3) to derive the upper reference value of COT-U in non-ETS exposed individuals. The COT-U levels of 495 adults (age range 18-69 years) who classified themselves as active smokers (29%) or as nonsmokers with (17%) or without (83%) ETS exposure were quantified by LC-MS-MS (quantification limit: 0.1µg/L, range of linearity: 0.1-4000µg/L). Median COT-U levels in these groups were 883, 1.38, and 0.39µg/L, respectively. Significant determinants of COT-U levels in active smokers were the number of cigarettes per day, type of smoking product, smoking environment, as well as time between the last cigarette and urine collection. Among ETS-exposed nonsmokers, significant determinants were living with smokers, being exposed to smoke at home, ETS exposure duration, as well as time between the last exposure and urine collection. When a 30-µg/L COT-U cut-off value was used to identify active daily smoking, the sensitivity and specificity of the questionnaire were 94% and 98%, respectively. For ETS exposure, the COT-U value of 1.78 (0.90 confidence interval 1.75-1.78) µg/L, corresponding to the 95th percentiles of the COT-U distribution in non-ETS-exposed participants, is proposed as upper reference value to identify environmental exposure., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Hydroquinone (HQ) is an important benzene-derived metabolite associated with acute myelogenous leukemia risk. Although altered DNA methylation has been reported in both benzene-exposed human subjects and HQ-exposed cultured cells, the inventory of benzene metabolite effects on the epigenome is only starting to be established. In this study, we used a monocytic leukemia cell line (THP-1) and hematopoietic stem cells (HSCs) from cord blood to investigate the effects of HQ treatment on the expression of the three most important families of retrotransposons in the human genome: LINE-1, Alu and Endogenous retroviruses (HERVs), that are normally subjected to tight epigenetic silencing. We found a clear tendency towards increased retrotransposon expression in response to HQ exposure, more pronounced in the case of LINE-1 and HERV. Such a partial loss of silencing, however, was generally not associated with HQ-induced DNA hypomethylation. On the other hand, retroelement derepression was also observed in the same cells in response to the hypomethylating agent decitabine. These observations suggest the existence of different types of epigenetic switches operating at human retroelements, and point to retroelement activation in response to benzene-derived metabolites as a novel factor deserving attention in benzene carcinogenesis studies., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Terbuthylazine (TBA) is an herbicide widely used in corn cultivation. Herein we evaluate the measurement of hair TBA as biomarkers of exposure. Five Sprague Dawley rats were gavaged with TBA for 3 days, and then the back hair was shaved and analyzed for TBA. In addition, head hair samples from 10 corn farmers, 9 rural residents, and 6 urban residents were collected at the end of the application season. Hair TBA was detected by liquid chromatography triple quadrupole mass spectrometry after solvent extraction. TBA was quantifiable in all rat samples with a mean concentration of 0.92 (±0.26)ng/mg, which corresponds to a 0.12% incorporation rate. TBA was quantifiable in all farmer samples (median: 0.67ng/mg), in 75% of rural resident samples (0.01ng/mg) and in none of the urban resident samples (<0.01ng/mg), with a statistical difference among groups (P<0.01). Our results suggest that TBA is incorporated in hair and prompt further investigation on the use of hair TBA as a potential biomarker of cumulative exposure., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Biological monitoring of low level exposure to pollutants is a very challenging analytical activity, and the quality of results is difficult to assess, especially when a certified reference material is unavailable. The aim of this work was to evaluate the reliability of the assays used to measure urinary benzene (Benz-U) and S-phenylmercapturic acid (SPMA), by applying an internal quality control protocol. Urine spot samples from 705 subjects who were either members of the general urban population, gasoline station attendants, or refinery plant workers were assayed for Benz-U and SPMA, using GC/MS and LC/MS/MS, with quantification limits of 15 ng/L and 0.10 μg/L. The median Benz-U concentration was 263 ng/L (60-2789 ng/L, 5th-95th percentile), and the median SPMA concentration was 0.19 μg/L (<0.1-2.5 μg/L, 5th-95th percentile). Linearity of both assays was good, but a less-than-proportional response was found for SPMA concentrations below 1 μg/L. Between-run precision and accuracy for Benz-U concentration determination were assessed using quality controls at 120 ng/L and 1000 ng/L and were 10.3% and 4.8%, and 104.8% and 98.9%, respectively; while the precision and accuracy for SPMA concentration determination at 0.3 μg/L, 2.5 μg/L, and 20 μg/L were 40.3%, 6.2%, and 6.2%, and 48.3%, 96.3%, and 98.8%, respectively. Precision, estimated using duplicates of unknown samples, was 13.4% for Benz-U and 26.5% for SPMA analyses. Control charts for the means of the slope of the linear calibration curve of Benz-U showed good stability of the means over a five-year period. For SPMA, a two-laboratory comparison revealed acceptable agreement between ln-transformed data pairs, with a slope of the linear regression of 0.863 (confidence interval 0.774-0.952), null intercept, and a Pearson's r value of 0.844. Reliable results were obtained for Benz-U analyses over the entire concentration range, and for high and medium SPMA levels. However, the determination of SPMA concentrations at levels close to the limit of quantification was less reliable., (Copyright © 2009 Elsevier B.V. All rights reserved.)

The aim of this work was to evaluate urinary benzene, toluene, ethylbenzene, m+p-xylene, o-xylene (BTEX), methyl tert-butyl ether (MTBE), ethyl tert-butyl ether (ETBE), and naphthalene (NAP) as biomarkers of exposure to environmental pollutants. Personal air and urine samples from 108 subjects belonging to the Italian general population were compared. Urinary profiles were obtained by headspace gas chromatography-mass spectrometry. BTEX, MTBE, ETBE and NAP median airborne exposures during a 5-h sampling were 4.0, 25.3, 3.8, 9.3, 3.4, 3.4, <0.8, and 3.4 microg/m(3), respectively. Meanwhile, median urinary levels, as geometric means of three determinations were: 122, 397, 74, 127, 43, 49, <15, and 46 ng/L, respectively. Urinary benzene and toluene concentrations were 4.6- and 1.2-fold higher in smokers than in non-smokers. For most chemicals, significant positive correlations between airborne exposure (log-transformed) and the corresponding biological marker (log-transformed) were found, with Pearson's r values for correlation, ranging from 0.228 to 0.396. Multiple linear regression analysis showed that the urinary level of these chemicals was influenced by personal airborne exposure, urinary creatinine, and urinary cotinine, with R(2) 0.733 for benzene. Urinary chemicals are useful biomarkers of environmental exposure. Given the ease of rapidly obtaining urine samples, they represent a non-invasive alternative to blood chemical analysis. The possibility of obtaining urinary exposure profiles makes this method an appealing tool for environmental epidemiology., (Copyright 2010 Elsevier B.V. All rights reserved.)

Aim of the study was the assessment of exposure of coke-oven workers to polycyclic aromatic hydrocarbons (PAHs) by determination of urinary profiles of hydroxylated and unmetabolized PAHs. Fifty-five Polish coke-oven workers were investigated by measurement of 12 hydroxylated metabolites of PAHs (OHPAHs) (1-, 2-hydroxynaphthalene; 2-, 9-hydroxyfluorene; 1-, 2-, 3-, 4-, 9-hydroxyphenanthrene; 1-hydroxyypyrene, 6-hydroxychrysene and 3-hydroxybenzo[a]pyrene) and 13 unmetabolized PAHs (U-PAHs) (from naphthalene to benzo[a]pyrene), in spot urine samples collected at the end of the workshift. U-PAHs with four or less rings were detected in all samples. In particular, median levels for urinary naphthalene, phenanthrene, pyrene, chrysene and benz[a]anthracene were 0.806, 0.721, 0.020, 0.032 and 0.035 microg/L. OHPAHs up to 1-hydroxypyrene were found in all samples, while high molecular-weight OHPAHs were always below quantification limit. Median level of 1-hydroxyypyrene was 15.4 microg/L. In all subjects significant correlations between OHPAHs and U-PAHs were observed (0.27 < r < 0.70, p < 0.01). Our results suggest that both hydroxylated metabolites and unmetabolized PAHs in urine are useful biomarkers of exposure to PAHs. Moreover, the simultaneous determination of several biomarkers permits to obtain specific excretion profiles that might help in exposure characterization and in better defining the excretion patterns.

The aim of this work is to compare several short-term biomarkers of styrene exposure, namely urinary styrene (StyU), mercapturic acids (M1+M2), mandelic acid (MA), phenylglyoxylic acid (PGA), phenylglycine (PHG), and 4-vinylphenol conjugates (VP), for use as biomarkers of exposure in epidemiologic studies. A repeated measurements protocol (typically 4 measurements per worker over 6 weeks) was applied to measure airborne styrene (StyA) and urinary biomarkers in 10 varnish and 8 fiberglass reinforced plastic workers. Estimated geometric mean personal exposures to StyA were 2.96mg/m(3) in varnish workers and 15.7mg/m(3) in plastic workers. The corresponding levels of StyU, M1+M2, MA, PGA, MA+PGA, PHG and VP were 5.13microg/L, 0.111, 38.2, 22.7, 62.6, 0.978, and 3.97mg/g creatinine in varnish workers and 8.38microg/L, 0.303, 146, 83.4, 232, 2.85 and 3.97mg/g creatinine in plastic workers. Within-worker (sigma(wY)(2)) and between-worker (sigma(bY)(2)) variance components were estimated from the log-transformed data as were the corresponding fold ranges containing 95% of the respective lognormal distributions of daily levels ((w)R(0.95)) and subject-specific mean levels ((b)R(0.95)). Estimates of (w)R(0.95) (range: 4-26) were generally smaller than those of (b)R(0.95) (range: 5-790) for both environmental and biological markers; this indicates that exposures varied much more between workers than within workers in these groups. Since attenuation bias in an estimated exposure-response relationship increases with the variance ratio lambda=sigma(wY)(2)/sigma(bY)(2), we estimated values of lambda for all exposure measures in our study. Values of lambda were typically much less than one (median=0.220) and ranged from 0.089 for M1+M2 in plastic workers to 1.38 for PHG in varnish workers. Since values of lambda were 0.147 and 0.271 for StyA in varnish workers and plastic workers, respectively, compared to 0.178 and 0.210 for MA in the same groups, our results suggest that either air measurements or conventional biomarker measurements (urinary MA) would be comparable surrogates for individual exposures in epidemiologic studies., (Copyright 2009 Elsevier Ireland Ltd. All rights reserved.)

An innovative and simple headspace solid-phase microextraction method using a novel diethoxydiphenylsilane fibre based on in situ derivatization with acetic anhydride was optimized and validated for the gas chromatography-mass spectrometry determination of some monohydroxy metabolites of polycyclic aromatic hydrocarbons, namely 1-hydroxynaphthalene, 2-hydroxynaphtalene, 9-hydroxyfluorene, 2-hydroxyfluorene and 1-hydroxypyrene at trace levels in human urine. Enzymatic hydrolysis was applied before derivatization, whereas extraction conditions, i.e. the effects of time and temperature of extraction and salt addition were investigated by experimental design. Regression models and desirability functions were applied to find the experimental conditions providing the highest global extraction response. These conditions were found in correspondence of an extraction temperature of 90 degrees C, an extraction time of 90 min and 25% NaCl added to urine samples. The capabilities of the developed method were proved obtaining limit of quantitations in the 0.1-2 microg/l range, thus allowing the bio-monitoring of these compounds in human urine. A good precision was observed both in terms of intra-batch and inter-batch repeatability with RSD always lower than 14%. Recoveries ranging from 98(+/-3) to 121(+/-1)% and extraction yields higher than 72% were also obtained. Finally, the analysis of urine specimens of coke-oven workers revealed analytes' concentrations in the 2.2-164 microg/l range, proving the exposure to PAHs of the involved workers.

Polycyclic aromatic hydrocarbons (PAHs) are common environmental pollutants in both living and working environments. The aim of this study was the development of a headspace solid-phase microextraction gas chromatography-isotope dilution mass spectrometry (HS-SPME/GC-IDMS) method for the simultaneous quantification of 13 PAHs in urine samples. Different parameters affecting PAHs extraction by HS-SPME were considered and optimized: type/thickness of fiber coatings, extraction temperature/time, desorption temperature/time, ionic strength and sample agitation. The stability of spiked PAHs solutions and of real urine samples stored up to 90 days in containers of different materials was evaluated. In the optimized method, analytes were absorbed for 60min at 80 degrees C in the sample headspace with a 100mum polydimethylsiloxane fiber. The method is very specific, with linear range from the limit of quantification to 8.67 x 10(3)ngL(-1), a within-run precision of <20% and a between-run precision of <20% for 2-, 3- and 4-ring compounds and of <30% for 5-ring compounds, trueness within 20% of the spiked concentration, and limit of quantification in the 2.28-2.28 x 10(1)ngL(-1) range. An application of the proposed method using 15 urine samples from subjects exposed to PAHs at different environmental levels is shown.

The aim of this study was the development of a method for the determination of 12 urinary monohydroxy metabolites of PAHs, namely 1-hydroxynaphthalene, 2-hydroxynaphthalene, 2-hydroxyfluorene, 9-hydroxyfluorene, 1-hydroxyphenanthrene, 2-hydroxyphenanthrene, 3-hydroxyphenanthrene, 4-hydroxyphenanthrene, 9-hydroxyphenanthrene, 1-hydroxypyrene, 6-hydroxychrysene, and 3-hydroxybenzo[a]pyrene. Analytes were determined in the presence of deuterated analogues as internal standards, by GC/MS operating in the electron impact mode. Sample preparation was performed by enzymatic hydrolysis of glucoronate and sulphate conjugates of hydroxy metabolites of PAHs, liquid-liquid extraction with n-hexane, and derivatization with a silylating reagent. The method is very specific, limits of quantification are in the range 0.1-1.4 microg/l, and range of linearity is from limit of detection to 208 microg/l. Within- and between-run precision, expressed as coefficient of variation, are <20%; accuracy for most analytes is within 20% of the theoretical value. An application of the proposed method to the analysis of 10 urine samples from coke-oven workers shows that 1-hydroxynaphthalene and 2-hydroxyfluorene were the most abundant compounds (median 61.4 and 69.7 microg/l, respectively), while 6-hydroxychrysene, and 3-hydroxybenzo[a]pyrene were always below the quantification limit.

Methyl tert-butyl ether (MTBE), ethyl tert-butyl ether (ETBE) and tert-amyl methyl ether (TAME) are oxygenated compounds added to gasoline to enhance octane rating and to improve combustion. They may be found as pollutants of living and working environments. In this work a robotized method for the quantification of low level MTBE, ETBE and TAME in human urine was developed and validated. The analytes were sampled in the headspace of urine by SPME in the presence of MTBE-d12 as internal standard. Different fibers were compared for their linearity and extraction efficiency: carboxen/polydimethylsiloxane, polydimethylsiloxane/divinylbenzene, and polydimethylsiloxane. The first, although highly efficient, was discarded due to deviation of linearity for competitive displacement, and the polydimethylsiloxane/divinylbenzene fiber was chosen instead. The analysis was performed by GC/MS operating in the electron impact mode. The method is very specific, with range of linearity 30-4600 ng L(-1), within- and between-run precision, as coefficient of variation, <22 and <16%, accuracy within 20% the theoretical level, and limit of detection of 6 ng L(-1) for all the analytes. The influence of the matrix on the quantification of these ethers was evaluated analysing the specimens of seven traffic policemen exposed to autovehicular emissions: using the calibration curve and the method of standard additions comparable levels of MTBE (68-528 ng L(-1)), ETBE (<6 ng L(-1)), and TAME (<6 ng L(-1)) were obtained.

This study was carried out to define reference values for urinary ethylenethiourea (ETU) in the Northern Italy population and to identify the sources of exposure. Ninety-five healthy subjects were selected. A spot urine sample was collected in the morning, and analyzed using GC/MS in the EI/SIM mode. Thirty-nine subjects showed urinary ETU concentrations lower than the limit of detection (LOD, 0.4 microg/g creatinine), and the remainders ETU concentrations ranging from 0.5 to 11.6 microg/g creatinine. No correlation was shown between smoke or alcohol intake and urinary ETU concentrations. Based on data on ethylene-bis-dithiocarbamate (EBDC) concentrations in food, we estimated a total EBDCs intake of 31.7-50.1 microg/day. These values are largely below the ADIs, but explain the presence of small amounts of ETU in the urine samples we have analyzed. Finally, it was estimated that the mean ETU in urine in the Italian general population is 0.6-0.8 microg/g creatinine, with a 95th percentile of 4.5-5.0 microg/g creatinine. These values can be used as reference, to compare the results of biological monitoring activities carried out on EBDCs occupationally and environmentally exposed populations.

In this paper we evaluated the possibility to assess occupational exposure to polycyclic aromatic hydrocarbons (PAHs) measuring unmetabolized PAHs in urine. With this aim, 24 road paving (RP) workers, exposed to bitumen fumes, and 6 road construction workers (CW), exposed to diesel exhausts, were investigated. Median personal exposure to low boiling PAHs (from naphthalene to pyrene) during the work shift ranged from 0.5 to 369 ng/m(3), with naphthalene as the most abundant compound. Three urine samples were collected for each worker: baseline (after 2 days of vacation), before- and end-shift samples (in the second part of the work week). The following urinary compounds were measured by headspace-solid phase microextraction GC/MS: naphthalene (U-NAP), acenaphthylene (U-ACY), acenaphthene (U-ACE), fluorene (U-FLE), phenanthrene (U-PHE), anthracene (U-ANT), fluoranthene (U-FLU), pyrene (U-PYR). Urinary PAHs were detected in almost all samples. Median levels for U-NAP, U-PHE, U-PYR and U-FLE in end-shift samples were 82, 48, 54 and 21 ng/L in RP and 69, 14, 24 and 15 ng/L in CW, respectively. Significant differences in the levels of U-PHE, U-FLU and U-PYR were found between RP and CW (p<0.05). Moreover in RP samples the urinary excretion of most analytes increased during the work shift (p<0.05). These results suggest that urinary PAHs may be useful biomarkers of occupational exposure.

S-Phenylmercapturic acid (S-PMA), is a urinary metabolite of benzene, thought to be derived from the condensation product of benzene oxide with glutathione. S-PMA may be determined by GC, HPLC (UV or fluorescence detection), GC-MS, LC-MS/MS or immunoassays. The limit of sensitivities of most of these techniques is 1 microg/l urine or below. It has been suggested that S-PMA may have value as a biomarker for low level human exposure to benzene, in view of the facts that urinary excretion of S-PMA has been found to be related to airborne benzene in occupationally exposed workers, and that only low background levels of S-PMA have been found in control subjects. We have evaluated the use of S-PMA as a biomarker, using a commercially available analytical service, in a multicentre European study of populations exposed to varying levels of benzene, in Italy (Milan, Genoa) and in Bulgaria (Sofia). These were filling station attendants, urban policemen, bus drivers, petrochemical workers and referents (a total of 623 subjects). S-PMA was measured at the end of the work shift by an immunoassay procedure. Urinary benzene (in Milan only) and the benzene metabolite trans,trans-muconic acid (t,t-MA) were measured before and after the work shift. Air-borne benzene was measured as a monitor of exposure. Urinary benzene was the most discriminatory biomarker and showed a relationship with airborne benzene at all levels of exposure studied (including groups exposed to <0.1 ppm benzene), whereas t,t-MA and S-PMA, as determined by immunoassay, were suitable only in the highest exposed workers (petrochemical industry, geometric mean 1765 microg/m3 (0.55 ppm) benzene). All three biomarkers were positively correlated with smoking as measured by urinary cotinine).

This research compared the capability of urinary trans,trans-muconic acid (t,t-MA), S-phenylmercapturic acid (S-PMA) and benzene excreted in urine (U-benzene) to monitor low benzene exposure and evaluated the influence of smoking habit on these indices. Gasoline attendants, urban policemen, bus drivers and two groups of referents working in two large Italian cities (415 people) were studied. Median benzene exposure was 61, 22, 21, 9 and 6 microg/m3, respectively, with higher levels in workers than in referents. U-benzene, but not t,t-MA and S-PMA, showed an exposure-related increase. All the biomarkers were strongly influenced by cigarette smoking, with values up to five-fold higher in smokers compared to non-smokers. In conclusion, in the range of investigated benzene exposure (<478 microg/m3 or <0.15 ppm), the smoking habit may be regarded as a major source of benzene intake; among the study indices, U-benzene is the marker of choice for the biological monitoring of occupational and environmental exposure.

ortho-Cresol (o-C) and meta-cresol (m-C) are minor urinary metabolites of toluene, a widely used chemical with neurotoxicological properties. A new assay for their determination in human urine is here proposed. Urinary cresol sulphates and glucuronates are submitted to acid hydrolysis, urine is neutralized, added with o-cresols-d8, and analytes are sampled in the headspace of urine by SPME using a polydimethylsiloxane fiber. Analysis is performed by GC/MS using, for separation, either a SupelcoWax10 (for o-C) or a chiral CP Cresol (for o-C and m-C) column. The method is very specific, with a range of linearity 0-5.0 mg/l, within- and between-run precision, as coefficient of variation, <15% and <19%, limit of detection of 0.006 mg/l for o-C and 0.007 mg/l for m-C. The procedure is applied to the quantification of cresols in urine from workers exposed to toluene and from subjects belonging to the general population.

Ethylenethiourea (ETU) is a major metabolite of ethylenebisdithiocarbamate pesticides: a sensitive and specific assay for its determination in human urine is proposed below. ETU is extracted on a diatomaceous earth column using dichloromethane and derivatized with the mixture of N-(tert-butyldimethylsilyl)-N-methyltrifluoroacetamide and tert-butyldimethyilsilyl chloride. The derivative is analyzed using GC/MS in the EI/SIM mode. The whole procedure is carried out in the presence of ethylenethiourea-d(4) as internal standard. The analytical features of the method are: high specificity, >90% recovery, range of linearity 0-200 microg/L, within- and between-run precision as coefficient of variation, <17 and <20%, respectively, limit of quantification 2 microg/L. In specimens stored in the dark at -20 degrees C ETU is stable for at least 6 months. The procedure was successfully applied to the biological monitoring of vineyard workers exposed to EBDTC and of a matched group of subjects from the general population.

Exposure to 1,3-butadiene (BD), a probable carcinogen to humans, was investigated in two groups of subjects working in a petrochemical plant where BD is produced and used to prepare polymers: 42 occupationally exposed workers and 43 internal non-occupationally exposed controls. BD personal exposure was very low but significantly different in the two groups (median airborne BD 1.5 and 0.4 microg/m(3) in exposed and controls, respectively). Similarly, BD in blood and urine, but not in exhaled air, was higher in the exposed workers than in controls (blood BD 3.7 ng/l versus <1.8 ng/l, urinary BD 2.4 ng/l versus <1.0 ng/l). These three biomarkers correlated significantly with personal exposure ( 0.283 < or = Pearson's r < or = 0.383) and between them (0.780 < or = r < or = 0.896). Excretion of urinary mercapturic acids N-acetyl-S-(3,4-hydroxybutyl)-l-cysteine (MI), N-acetyl-S-(1-hydroxymethyl-2-propenyl)-l-cysteine and N-acetyl-S-(2-hydroxy-3-butenyl)-l-cysteine (MII), chromosomal aberrations (CA), and sister chromatid exchanges (SCE) in peripheral blood lymphocytes were not influenced by occupational exposure. Our results show that unmetabolised BD in biological fluids, and particularly urinary BD, represents the biomarker of choice for assessing occupational exposure to low airborne concentrations of BD.

In the present study, the personal exposure to mancozeb and/or ethilenethiourea (ETU) in 13 Italian vineyard workers and in 13 subjects without occupational exposure to pesticides was investigated. With this aim, the level of ETU in urine and the dermal exposure to mancozeb were determined. Baseline urinary ETU results were lower than the analytical limit of detection for all controls (<0.5 microg/g creatinine) and for ten workers (median <0.5, range <0.5-3.4 microg/g creatinine). In workers, urinary ETU was significantly increased at the end of shift (2.5, <0.5-95.2 microg/g creatinine) compared with baseline levels. End-shift urinary ETU was higher in operators using open tractors (n=7) than in those using closed tractors (n=5) (16.2 vs. 2.4 microg/g creatinine), but the difference was not significant (P=0.073). End-shift urinary ETU was positively correlated with dermal exposure to mancozeb determined both over the clothes and on the skin (Spearman's rho=0.770 and 0.702, P=0.009 and 0.024, respectively). Wine consumption positively influenced the excretion of ETU.

Hemoglobin adducts were determined as biomarkers of 1,3-butadiene (BD) in 30 workers and 10 controls from an Italian BD plant and in 14 diesel-exposed miners. N-(2,3,4-trihydroxybutyl)valine (THBVal), an N-terminal valine globin adduct of reactive butadiene metabolites, was analyzed by gas chromatography/high resolution mass spectrometry after a modified Edman degradation and further acetylation. The BD exposure for the plant workers was 31 microg/m(3) (personal sampling). Whereas there was no detectable difference in hemoglobin adduct levels (range 17.7-61.4 pmol/g globin) between the total group of exposed and controls, slight but significant differences could be found between two subgroups of workers from different production units as well as one subgroup and controls (P<0.05), between smoking (n=13) and non-smoking exposed workers (n=17; P=0.066) as well as between smoking exposed workers and controls (P=0.055). Adduct levels of the miners (all non-smokers) were in the same range as those of the Italian BD-workers and controls. The internal exposure and strain measured by THBVal levels resulting from a very low occupational BD exposure was in the range of the contribution of moderate smoking.

A method was developed for the determination in human urine of S-phenylmercapturic (PMA) and S-benzylmercapturic (BMA) acids, metabolites respectively of benzene and toluene. PMA and BMA were determined, after alkaline hydrolysis, to give respectively thiophenol and benzylmercaptan, and coupling of the thiol-containing compounds with monobromobimane (MB), by reversed-phase HPLC on a diphenyl-silica bonded cartridge (100 x 4.6 mm I.D., 5 microm particle size) with fluorimetric detection. Wavelengths for excitation and emission were 375 and 480 nm, respectively. The recovery of PMA and BMA from spiked urines was >90% in the 10-500 microg/l range; the quantification limits were respectively 1 and 0.5 microg/l; day-to-day precision at 42 microg/l was C.V. <7%. The suitability of the proposed procedure for the biological monitoring of exposure to low-level airborne concentrations of benzene and toluene, was evaluated by analyzing the urinary excretion of PMA and BMA in subjects exposed to different sources of aromatic hydrocarbons, namely occupationally-unexposed referents (non-smokers, n=15; moderate smokers, n=8; mean number of cigarettes smoked per-day=17 cig/day) and non-smoker workers occupationally exposed to toluene in maintenance operations of rotogravure machines (non-smokers, n=17). Among referents, non-smokers showed values of PMA ranging from <1 to 4.6 microg/l and BMA from 1.0 to 10.4 microg/l; in smokers, PMA values ranging from 1.2 to 6.7 microg/l and BMA from 9.3 to 39.9 microg/l, were observed. In occupationally exposed non-smoker subjects, BMA median excretion value (23.6 microg/l) was higher than in non-smoker referents (3.5 microg/l) (P<0.001) and individual BMA values (y, microg/l) were associated and increased with airborne toluene concentration (x, mg/m3) according to the equation y=6.5+0.65x (r=0.69, P<0.01, n=17). The proposed analytical method appears to be a sensitive and specific tool for biological monitoring of low-level exposure to benzene and toluene mixtures in occupational and environmental toxicology laboratory.

The analysis of benzene in urine of the general population or of exposed workers can be performed with different methods using the 'purge and trap' or 'solid-phase microextraction' techniques in combination with gas chromatographic analysis and photoionisation or mass spectrometric detection. The published results, however, are deeply conflicting. Differences in sample preparation by different research groups and our own preliminary observations prompted us to investigate pre-analytical and analytical factors potentially capable of modifying the urinary benzene quantification results. Benzene concentrations were measured in 20 urine samples in relation to different conditioning conditions (at 24, 40 and 80 degrees C) and at basic or acid pH. Urinary protein concentrations were measured in the same samples. Urine heating at 80 degrees C yields benzene concentrations on average five times higher than at 24 degrees C. On acidification of urine, the benzene released increases up to 28-fold in comparison to that obtained at uncorrected 'physiological' pH. Despite a widely scattered data distribution, a statistically significant linear correlation was found between 'heat-released' and 'acid-labile' benzene values. There was no correlation between total urinary proteins present in 'physiological' concentrations (between 12 and 110 mg/l) and the different kinds of benzene in urine. Our results could perhaps be explained if it is supposed that part of the benzene in urine is absorbed onto sediment, or bound to specific proteins, or derived from parent molecules and is released with pH modification or heat administration. Our observations may also help to explain why the urinary benzene concentrations reported by different investigators vary considerably even when environmental levels are comparable.

A method for the determination of benzene, toluene, ethylbenzene and xylenes (BTEX) in urine of people exposed to these airborne pollutants present in the living environment, has been described. Solid-phase microextraction has been used for sampling BTEX from the headspace of urine and gas chromatography-mass spectrometry has been applied for the selective analysis of chemicals. The method has the following features: small volume of urine (2 ml) needed, linearity in the range of interest (from the limit of detection up to 5000 ng/l) with coefficient of correlation > or =0.998, limit of detection in the range 12-34 ng/l, good repeatability (coefficient of variation 2-7%), high specificity. The stability of the urine sample during storage (-20 degrees C) was evaluated: BTEX remained stable for up to 2 months. The assay has been successfully applied to the biological monitoring of two subjects environmentally exposed to airborne BTEX in an urban area.

Methylhippuric acid isomers (MHAs), urinary metabolites of xylenes, were determined, after clean-up by C18-SPE and esterification with hexafluoroisopropanol and diisopropylcarbodiimide, by GC with ECD detection, on an SPB-35 capillary column (30 m, 0.32 mm I.D., 0.25 microm film thickness, beta = 320). S-benzyl-mercapturic acid was used for internal standardization. Chromatographic conditions were: oven temperature 162 degrees C, for 14.2 min; ramp by 30 degrees C/min to 190 degrees C, for 3.5 min; ramp by 30 degrees C/min to 250 degrees C, for 4 min; helium flow rate: 1.7 ml/min; detector and injector temperature: 300 degrees C. The sample (1 microl) was injected with a split injection technique (split ratio 5:1). MHA recovery was >95% in the 0.5-20 micromol/l range; the limit of detection was <0.25 micromol/l; day-to-day precision, at 2 micromol/l, was Cv<10%. Urinary MHAs were determined in subjects exposed to different low-level sources of xylenes: (a) tobacco smoking habit and (b) BTX urban air pollution (airborne xylene ranging from 0.1 to 3.7 micromol/m3). Study (a) showed a significant difference between urinary MHA median excretion values of nonsmokers and smokers (4.6 micromol/l vs. 8.1 micromol/l, p<0.001). Study (b) revealed a significant difference between indoor workers and outdoor workers (4.3 micromol/l vs. 6.9 micromol/l, p<0.001), and evidenced a relationship between MHAs (y, micromol/mmol creatinine) and airborne xylene (x, micromol/m3) (y = 0.085+0.34x; r = 0.82, p<0.001, n = 56). Proposed biomarkers could represent reliable tools to study very low-level exposure to aromatic hydrocarbons such as those observed in the urban pollution due to vehicular traffic or in indoor air quality evaluation.

trans, trans-Muconic acid (1,3-butadiene-1, 4-dicarboxylic acid, MA), a minor urinary metabolite of benzene exposure, was determined, after clean-up by solid-phase anion-exchange chromatography, by reversed-phase HPLC on a C18 column (5 x 0.46 cm I.D., 3 microns particle size), using formic acid-tetrahydrofuran-water (14:17:969) as mobile phase and UV detection at 263 nm. The recovery of MA from spiked urine was > 95% in the 50-500 microgram/l range; the quantification limit was 6 micrograms/l; day-to-day precision, at 300 micrograms/l, was C.V. = 9.2%; the run time was less than 10 min. Urinary MA excretion was measured in two spot urine samples of 131 benzene environmentally exposed subjects: midday values obtained in non-smokers (mean +/- S.D. = 77 +/- 54 micrograms/l, n = 82) were statistically different from those of smokers (169 +/- 85 micrograms/l, n = 30) (P < 0.0001); each group showed a statistically significant increase between MA excretion in midday over morning samples. Moreover, in subjects grouped according to tobacco-smoke exposure level, median values of MA were positively associated with and increased with daily smoking habits.

Environmental and biological monitoring of airborne aromatic hydrocarbons has been performed in 20 policemen working as traffic wardens exposed to motor vehicle exhausts and in 19 peers employed as clerks. Airborne benzene, toluene, ethylbenzene and xylene concentrations, measured during the workshift, resulted in significantly higher outdoor than indoor concentrations (benzene and related aromatic hydrocarbons mean values, respectively of 53 and 350 micrograms/m3 vs. 29 and 180 micrograms/m3). Blood benzene, toluene, ethylbenzene and xylene concentrations did not differ significantly between indoor and outdoor workers; no differences were found between values obtained at the beginning (07:30 h) and the end of shift (00:30) in either group. Blood hydrocarbon concentrations seem to reflect airborne pollution, whilst the blood benzene concentration determined after the workshift poorly reflects airborne benzene morning peaks. Endshift blood benzene mean concentration in smokers (462 ng/l, n = 9) differs significantly from non-smokers (292 ng/l, n = 39).

Protein tyrosine kinases (PTK) are important signal transducing enzymes involved in the modulation of normal cellular growth and differentiation and have been associated with the etiology of various human cancers. The development of properly designed inhibitors, which block their function by interfering with the substrate binding, may therefore offer an unique target for selective anticancer chemotherapy. Here we describe synthesis and biochemical testing of a novel series of non-peptide PTK inhibitors which have as characteristic active pharmacophore the cinnamamide moiety. For testing we used an exogenous substrate kinase assay based on the phosphorylation of (Val5)-angiotensin II with radiolabelled ATP by the catalytic domain of the PTK encoded by the v-abl oncogene (p45 v-abl). The most potent compounds were found in the class of 3-arylidene-2-oxindoles (II) with IC50 values in the 1 microM range. Among these the 2-tetralylmethylene-, 4-quinolylmethylene-, 5-quinolylmethylene- and 3-indolylmethylene-2-oxindole compounds of formulae 16, 20, 21 and 24 respectively were selected for further investigation.