Comprehensive LC-MS/MS method combined with tandem hybrid hydrolysis for multiple exposure assessment of multiclass environmental pollutants.

Environmental pollutants (EPOLs), such as phthalates, volatile organic compounds, phenols, parabens, polycyclic aromatic hydrocarbons, pyrethroids, and environmental tobacco smoke, are highly heterogeneous compounds. Recently, attention has been drawn to the assessment of the combinatory effects of multiple EPs. To correlate multiple exposures with potential health implications, advanced comprehensive analytical methods covering multiclass EPOLs are essential. However, because of several technical problems associated with enzyme hydrolysis, simultaneous extraction, and multiresidue liquid chromatography-tandem mass spectrometry analysis, it is difficult to establish a comprehensive method covering a number of EPOLs in a single sample preparation and analytical run. We developed tandem hybrid hydrolysis, modified direct injection, and a comprehensive mobile phase to overcome these technical problems and established a comprehensive analytical method for simultaneous biomonitoring of multiclass EPOLs. Tandem hybrid hydrolysis using ß-glucuronidase and consecutive acid hydrolysis allowed selective hydrolysis of glucuronide- and sulfate-conjugated metabolites without phthalate degradation. The comprehensive mobile phase composed of 0.01% acetic acid and acetonitrile enabled us to simultaneously analyze 86 EPOLs, with good chromatographic behavior and ionization efficiency. Modified direct injection allowed a small amount of sample and simultaneous urinary extraction. The method was validated and applied to 39 urine samples from 19 mother-newborn pairs for multiple exposure assessment. Results showed that BP-3, a general component in sunblock products, and monoethyl phthalate, a metabolite of diethyl phthalate, exhibit a clear positive correlation between mothers and newborns. Therefore, the developed method has potential as a novel analytical tool for long-term, large-scale, and data-rich human biomonitoring of EPOLs.

Multiresidue analysis of 85 persistent organic pollutants in small human serum samples by modified QuEChERS preparation with different ionization sources in mass spectrometry.

In this study, a multiresidue analytical method was developed, validated, and applied for quantifying 85 persistent organic pollutants (POPs), including 38 polychlorinated biphenyls (PCBs), 23 polybrominated diphenyl ethers (PBDEs), and 24 organochlorine pesticides (OCPs) from 200 µL of human serum. A modified QuEChERS (Quick, Easy, Cheap, Effective, Rugged, and Safe) method was applied to minimize the required sample amount and optimize various conditions including the extraction solvent and the number of extractions. The extraction efficiency was optimized using double extraction with an ethyl acetate/hexane/acetone mixture. Gas chromatography coupled with triple-quadrupole mass spectrometry was used for analysis, and two different ionization sources, electron impact ionization (EI) and atmospheric pressure chemical ionization (APCI), were used to compare their sensitivity. The APCI source employed soft ionization at atmospheric pressure, producing abundant molecular ion formation with minimal fragmentation, in contrast to extensive fragmentation caused by EI. Of the 85 POPs analyzed, 59 target compounds (69.4%) showed lower limits of detection that were two- to fifty-fold lower in APCI than those determined using EI. The developed method was validated for its detection limit (0.5-10 pg/mL for PCBs, 2-20 pg/mL for PBDEs, and 2-40 pg/mL for OCPs), precision (0.8%-34.3% of coefficient of variation), recovery (49.6%-77.1%), matrix effect (46.7%-156.9%), and accuracy (81.2%-113.1% for PCBs, 85.8%-112.2% for PBDEs, and 55.2%-113.9% for OCPs). Its linearity was R2 > 0.99 for 84 compounds, and 96% average accuracy (for APCI) was obtained using the National Institute of Standards and Technology (NIST) standard reference materials (NIST 1957 and 1958). These ionization methods were compared by analyzing 25 real human serum samples. The observed species were 1.1-24.6 pg/mL of 28 PCBs, 2.5 pg/mL of BDE-47, and 6.5-195.1 pg/mL of 6 organochlorine pesticides (median concentration for each species), and only 11 compounds were detected with APCI owing to its enhanced sensitivity.

Analysis of Nicotine Metabolites in Hair and Nails Using QuEChERS Method Followed by Liquid Chromatography-Tandem Mass Spectrometry.

Many studies have analyzed nicotine metabolites in blood and urine to determine the toxicity caused by smoking, and assess exposure to cigarettes. Recently, hair and nails have been used as alternative samples for the evaluation of smoking, as not only do they reflect long-term exposure but they are also stable and easy to collect. Liquid-liquid or solid-phase extraction has mainly been used to detect nicotine metabolites in biological samples; however, these have disadvantages, such as the use of toxic organic solvents and complex pretreatments. In this study, a modified QuEChERS method was proposed for the first time to prepare samples for the detection of nicotine metabolite cotinine (COT) and trans-3'-hydroxycotinine (3-HCOT) in hair and nails. High-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to analyze traces of nicotine metabolites. The established method was validated for selectivity, linearity, lower limit of quantitation, accuracy, precision and recovery. In comparison with conventional liquid-liquid extraction (LLE), the proposed method was more robust, and resulted in higher recoveries with favorable analytical sensitivity. Using this method, clinical samples from 26 Korean infants were successfully analyzed. This method is expected to be applicable in the routine analysis of nicotine metabolites for environmental and biological exposure monitoring.

Effect of sulfidation and dissolved organic matters on toxicity of silver nanoparticles in sediment dwelling organism, Chironomus riparius.

The properties, fate, and toxicity of silver nanoparticles (AgNPs) are readily modified in the environment. Thus, in order to predict the environmental impact of AgNPs, the toxicity test should be conducted to assess the interactions of AgNPs with environmental matrices. Dissolved organic matter (DOM) is known to mitigate AgNPs toxicity in natural systems, and it is also known that silver binds strongly to sulfur. Little is known, however, about the effect of sulfidation and to what extent it could compete with DOM in the sediment. We therefore investigated the effect of sulfide on a sediment dwelling organism, Chironomus riparius using ecotoxicity endpoints. We then investigated how sulfide and a combination of sulfide and DOM affect the toxicity of AgNPs in C. riparius. We also monitored the concentrations of silver in the water and sediment compartments, as well as in C. riparius tissue, in the presence and absence of sulfide. Finally, in order to investigate how sulfide and DOM affect the release of ions from AgNPs, we also monitored released Ag(+) in each treatment. In the presence of sulfide, AgNPs were found to be less toxic to C. riparius in acute and chronic endpoints than AgNPs alone, whereas DOM treatment did not modulate the toxicity of AgNPs. Sulfide treatment reduced the release of Ag(+) from AgNPs. Water-spiked AgNPs with sulfide were found to be more slowly incorporated into both sediment and larvae as compared to the AgNP alone. Overall, the results suggest that the presence of sulfide in sediment mitigates the ecotoxicity of AgNPs in C. riparius.

Possible Role of Phthalate in the Pathogenesis of Endometriosis: In Vitro, Animal, and Human Data.

CONTEXT: Although phthalates were shown to have several negative effects on reproductive function in animals, its role in the pathogenesis of endometriosis remains to be elucidated. OBJECTIVE: We aimed to investigate the in vitro and in vivo effects of di-(2-ethylhexyl)-phthalate (DEHP) and to compare the urinary levels of several phthalate metabolites between women with and without endometriosis. DESIGN: For experimental studies, we used endometrial cell culture and nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mouse models. We also performed a prospective case-control study for human sample analyses. SETTING: The study was conducted at an academic center. MAIN OUTCOME MEASURES: The activities of matrix metalloproteinase (MMP)-2 and 9, cellular invasiveness, phosphorylation of extracellular signal-regulated kinase (Erk), and expression of p21-activated kinase 4 were analyzed in endometrial cells treated with DEHP. The implant size was compared between NOD/SCID mice fed with and without DEHP. Urinary concentrations of several phthalate metabolites were compared between women with and without endometriosis. RESULTS: In vitro treatment of endometrial cells with DEHP led to significant increases of MMP-2 and 9 activities, cellular invasiveness, Erk phosphorylation, and p21-activated kinase 4 expression. The size of the endometrial implant was significantly larger in the NOD/SCID mice fed with DEHP compared with those fed with vehicle. The urinary concentration of mono (2-ethyl-5-hydroxyhexyl) phthalate, mono (2-ethyl-5-oxohexyl) phthalate, and mono (2-ethyl-5-carboxyphentyl) phthalate were significantly higher in women with endometriosis compared with controls. CONCLUSION: These findings strongly suggest that exposure to phthalate may lead to establishment of endometriosis by enhancing invasive and proliferative activities of endometrial cells.

Decline in non-smoking workers' urine cotinine levels after increased smoking regulation in Korea.

OBJECTIVES: To identify any association between implementing smoking regulation policies and workers' urine cotinine concentration levels in Korea. METHODS: From the first stage of the Korean National Environmental Health Survey conducted by the National Institute of Environmental Research from 2009 to 2011, 2,475 non-smoking workers selected. We analyzed the trend in the changes of cotinine concentration in urine using the general linear model and linear regression, in various jobs as categorized by the National Center for Health Statistics (NCHS) and Korea Standard Classification of Occupations (KSCO). RESULTS: The urine cotinine concentration tended to decrease every year (2.91 ng/ml in 2009, 2.12 ng/ml in 2010, and 1.31 ng/ml in 2011), showing a decreasing trend (P < 0.001). The total subjects' decreased cotinine concentration in urine between 2009 and 2011 was 2.72 ng/ml (54.1 % relative decrease). The changes in each subgroup's urine cotinine concentration ranged from 1.59 to 6.03 ng/ml (33.2 to 77.5 %). All groups except for the managerial group (n = 49), which had a small sample size, had statistically significant negative regression coefficients (p < 0.05). The ranges of the decrease in urine cotinine were 2.75 ng/ml (53.6 %) for males and 2.72 ng/ml (54.9 %) for females. The negative slope in urine cotinine level was statistically significantly greater in men than women. The changes in urine cotinine by occupation as classified by the NCHS occupational categories ranged from 2.43 to 3.36 ng/ml (46.6 to 61.5 % relative decrease). The negative slopes in urine cotinine levels of the white-collar and farm workers were statistically significantly greater than those of the service workers and blue-collar workers. The change by occupation as classified by the KSCO ranged from 1.59 to 6.03 ng/ml (a 33.2 to 77.5 % relative decrease). The negative slopes in urine cotinine levels of the professionals and related workers and clerks were statistically significantly greater than those of the service workers and plant and machine operators and assemblers. CONCLUSIONS: The cotinine concentration in urine among non-smoking worker groups tended to decline from 2009 to 2011. Such a result may be an indirect indicator of the effectiveness of smoking regulation policies including the revision of the National Health Promotion Act.

Association between urine phthalate levels and poor attentional performance in children with attention-deficit hyperactivity disorder with evidence of dopamine gene-phthalate interaction.

Although there is some evidence supporting the existence of an association between prenatal maternal or postnatal child's urine phthalate metabolite concentrations and poor attentional performances, the interaction between urine phthalate metabolite levels and genetic variation for neuropsychological deficit of attention-deficit hyperactivity disorder (ADHD) has not been examined. The aim of this study was to determine whether phthalate metabolites in urine are associated with poor neuropsychological performance in children with ADHD, and whether such association is affected by genotype-phthalate interaction. A cross-sectional examination of urine phthalate metabolite concentrations and the continuous performance test (CPT) were performed in 179 Korean children with ADHD recruited from department of psychiatry of university hospital. Correlations between urine phthalate metabolite concentrations and the CPT scores were investigated, and the interaction of phthalate metabolite levels with the selected polymorphisms at major candidate genes for ADHD, namely dopamine receptor D4 (DRD4), dopamine transporter, α-2A-adrenergic receptor, and norepinephrine transporter genes. For the subjects with the DRD4 4/4 genotype, there were significant associations of the urine phthalate metabolite concentrations with the number of omission errors, the number of commission errors, and the response time variability scores on the CPT. However, for the subjects without the DRD4 4/4 genotype, no significant associations were found. The results of this study suggest a possible association between phthalate metabolite concentrations and poor attentional performances of ADHD as well as a genetic influence on this association. Further prospective and epigenetic studies are needed to investigate causality and pathophysiological mechanisms.

Secondhand smoke exposure and urine cotinine concentrations by occupation among Korean workers: results from the 2008 Korea National Survey for Environmental Pollutants in the Human Body.

This study aimed to estimate the status of secondhand smoke (SHS) exposure through urine cotinine analysis among nonsmoking workers in Korea and to analyze factors affecting urine cotinine concentrations. Data were based on "The 2008 Korea National Survey for Environmental Pollutants in the Human Body," a cross-sectional study of the National Institute of Environmental Research of Korea. We selected 1448 nonsmoking adult workers from 200 localities to participate in this survey. Urine cotinine concentrations were analyzed using a gas chromatograph-mass selective detector. We calculated separate covariate-adjusted geometric means for socio-demographic variables for males, females, and total subjects by analysis of covariance (ANCOVA). Statistical analyses were performed using SPSS version 18.0 (SPSS Inc., Chicago, Ill.). The prevalence of self-reported exposure to SHS was 36.9%. The geometric mean (95% confidence interval) of urine cotinine concentrations among all participants was 16.50 (14.48-18.80) µg/L. Gender, living area, education, and SHS exposure showed significant differences in urine cotinine concentrations. The urine cotinine concentrations of farmworkers and blue-collar workers such as skilled agricultural, forestry, and fishery workers, and elementary occupations were higher than those of white-collar workers such as clerical support workers, technicians, and associate professionals. Such a high proportion of the population having high urine cotinine levels indicates widespread exposure to SHS among nonsmoking workers in Korea. Furthermore, the urine cotinine levels among nonsmoking workers exposed to SHS varied by occupation. The measured urine cotinine concentration is suggested to be a valuable indication of SHS exposure in Korea.

Assessment of arsenic exposure by measurement of urinary speciated inorganic arsenic metabolites in workers in a semiconductor manufacturing plant.

OBJECTIVES: The purpose of this study was to evaluate the exposure to arsenic in preventive maintenance (PM) engineers in a semiconductor industry by detecting speciated inorganic arsenic metabolites in the urine. METHODS: The exposed group included 8 PM engineers from the clean process area and 13 PM engineers from the ion implantation process area; the non-exposed group consisted of 14 office workers from another company who were not occupationally exposed to arsenic. A spot urine specimen was collected from each participant for the detection and measurement of speciated inorganic arsenic metabolites. Metabolites were separated by high performance liquid chromatography-inductively coupled plasma spectrometry-mass spectrometry. RESULTS: Urinary arsenic metabolite concentrations were 1.73 g/L, 0.76 g/L, 3.45 g/L, 43.65 g/L, and 51.32 g/L for trivalent arsenic (As3+), pentavalent arsenic (As5+), monomethylarsonic acid (MMA), dimethylarsinic acid (DMA), and total inorganic arsenic metabolites (As3+ + As5+ + MMA + DMA), respectively, in clean process PM engineers. In ion implantation process PM engineers, the concentrations were 1.74 g/L, 0.39 g/L, 3.08 g/L, 23.17 g/L, 28.92 g/L for As3+, As5+, MMA, DMA, and total inorganic arsenic metabolites, respectively. Levels of urinary As3+, As5+, MMA, and total inorganic arsenic metabolites in clean process PM engineers were significantly higher than that in the non-exposed group. Urinary As3+ and As5+ levels in ion implantation process PM engineers were significantly higher than that in non-exposed group. CONCLUSION: Levels of urinary arsenic metabolites in PM engineers from the clean process and ion implantation process areas were higher than that in office workers. For a complete assessment of arsenic exposure in the semiconductor industry, further studies are needed.

Korea National Survey for Environmental Pollutants in the human body 2008: 1-hydroxypyrene, 2-naphthol, and cotinine in urine of the Korean population.

The Korea National Survey for Environmental Pollutants in the human body conducts representative Korean population studies, which were first initiated in 2005 in Korea. This study was conducted from 2008 to 2009 to determine the exposure levels of polycyclic aromatic hydrocarbons and nicotine in the Korean general population. The study population consisted of 4702 adult subjects from 196 sampling locations including coastal, rural, and urban areas. The urinary levels of 1-hydroxypyrene, 2-naphthol, and cotinine were measured for exposure of polycyclic aromatic hydrocarbons and nicotine. The geometric means of the urinary 1-hydroxypyrene, 2-naphthol and cotinine concentrations in the Korean general population were 0.15 µg/L (95% confidence interval (CI): 0.13-0.17), 3.84 µg/L (95% CI: 3.57-4.11) and 47.42 µg/L (95% CI: 40.52-54.32) respectively. When these values were compared with reference ranges for the United States and Germany, the levels of 1-hydroxypyrene, 2-naphthol, and cotinine were very similar for Korea and Germany, however, these levels were slightly lower in the United States. This study is the first nationwide survey of exposure to polycyclic aromatic hydrocarbons and nicotine in Korea and provides a background reference range for exposure to polycyclic aromatic hydrocarbons and nicotine in the Korean general population.

Korea National Survey for Environmental Pollutants in the Human Body 2008: heavy metals in the blood or urine of the Korean population.

BACKGROUND: Recently, there have been several nationwide episodes involving imported toys contaminated with toxic metals and environmental hormones. In addition, cadmium intoxication has occurred due to soil contamination with cadmium from abandoned metal mines. OBJECTIVES: To investigate the distribution, extent and factors influencing the levels of toxic metals in the blood or urine of the Korean general population over twenty years of age, we studied the blood or urine concentrations of heavy metals in a representative sample of 5087 Koreans in 2008. METHODS: Multiple biological substrates were collected from each participant to determine the most suitable samples for an environmental health survey system. Information regarding exposure conditions of all subjects was collected by questionnaire-based interviews. RESULTS: The geometric means of the blood lead, mercury and manganese levels were 19.1, 3.23 and 10.8 µg/L, respectively. The geometric means of urinary arsenic and cadmium concentrations were 43.5 and 0.65 µg/L, respectively. Blood mercury and urinary arsenic levels in the Korean general population were significantly higher than in European and American populations. CONCLUSIONS: The higher levels of blood mercury and urinary arsenic could be explained by the greater seafood consumption among the Korean population. This biomonitoring study of blood or urine heavy metals in the Korean general population provides important reference data stratified by demographic and lifestyle factors that will be useful for the ongoing surveillance of environmental exposure of Koreans to toxic metals.

Hebei spirit oil spill exposure and subjective symptoms in residents participating in clean-up activities.

OBJECTIVES: This study was conducted to examine the relationship between crude oil exposure and physical symptoms among residents participating in clean-up work associated with the Hebei Spirit oil spill, 2007 in Korea. METHODS: A total of 288 residents responded to a questionnaire regarding subjective physical symptoms, sociodemographic characteristics and clean-up activities that occurred between two and eight weeks after the accident. Additionally, the urine of 154 of the respondents was analyzed for metabolites of volatile organic compounds (VOCs) and polycyclic aromatic hydrocarbons (PAHs) and heavy metals. To compare the urinary levels of exposure biomarkers, the urine of 39 inland residents who were not directly exposed to the oil spill were analyzed. RESULTS: Residents exposed to oil remnants through clean-up work showed associations between physical symptoms and the exposure levels defined in various ways, including days of work, degree of skin contamination, and levels of some urinary exposure biomarkers of VOCs, metabolites and metals, although no major abnormalities in urinary exposure biomarkers were observed. CONCLUSIONS: This study provides evidence of a relationship between crude oil exposure and acute human health effects and suggests the need for follow-up to evaluate the exposure status and long-term health effects of clean-up participants.

[The effects of wearing protective devices among residents and volunteers participating in the cleanup of the Hebei Spirit oil spill].

OBJECTIVES: To assess the protective effects of wearing protective devices among the residents and volunteers who participated in the cleanup of the Hebei Spirit oil spill. METHODS: A total of 288 residents and 724 volunteers were surveyed about symptoms, whether they were wearing protective devices and potential confounding variables. The questionnaires were administered from the second to the sixth week following the accident. Spot urine samples were collected and analyzed for metabolites of 4 volatile organic compounds (VOCs), 2 polycyclic aromatic hydrocarbons (PAHs), and 6 heavy metals. The association between the wearing of protective devices and various symptoms was assessed using a multiple logistic regression adjusted for confounding variables. A multiple generalized linear regression model adjusted for the covariates was used to test for a difference in least-square mean concentration of urinary biomarkers between residents who wore protective devices and those who did not. RESULTS: Thirty nine to 98% of the residents and 62-98% of volunteers wore protective devices. Levels of fatigue and fever were higher among residents not wearing masks than among those who did wear masks (odds ratio 4.5; 95% confidence interval 1.23-19.86). Urinary mercury levels were found to be significantly higher among residents not wearing work clothes or boots (p<0.05). CONCLUSIONS: Because the survey was not performed during the initial high-exposure period, no significant difference was found in metabolite levels between people who wore protective devices and those who did not, except for mercury, whose biological half-life is more than 6 weeks.

Serum selenium level in healthy Koreans.

Of trace elements in the serum of living organisms, selenium (Se) is an essential mineral and plays the role of an antioxidant as selenoproteins protecting the organism against oxidative damage induced by hydrogen peroxide, other lipid hydroperoxides, and their derivatives. The aim of this study was to determine the mean serum Se levels in healthy Korean volunteers (50 males and 50 females) by using an inductively coupled plasma-mass spectrometry method. The samples were collected at the Health Promotion Centre of Kangnam St. Mary's Hospital, College of Medicine, the Catholic University of Korea, Kangnam District, Seoul in accordance with procedures approved by the Institutional Review Board of the Catholic University of Korea. The mean serum Se level in healthy subjects was 112.05 +/- 30.42 microg/l. For gender, it was 120.81 +/- 27.37 microg/l for females and 103.29 +/- 31.05 microg/l for males. From the study result, there was a significant difference between the mean Se concentrations of gender groups (p = 0.0035). Also, the study indicated no effect of age on Se levels (p > 0.05) in the healthy individuals.

Evaluation of toxicological monitoring markers using proteomic analysis in rats exposed to formaldehyde.

Formaldehyde (FA) is known as a low molecule weight organic compound and one of major components that causes sick building syndrome (SBS), and it has been reported that FA has cytotoxic, hemotoxic, immunotoxic, and genotoxic properties. The International Agency for Research on Cancer (IARC) has characterized FA as a carcinogen. In this study, we investigated the effects of FA on rat plasma proteins by using proteomic approach. Rats were exposed to three different concentrations of FA (0, 5, 10 ppm) for 2 weeks at 6 hours/day and 5 days/week in an inhalation chamber. Malondialdehyde (MDA) assay and carbonyl spectrometric assay were conducted to determine lipid peroxidation and protein oxidation levels and Comet assays were used for genotoxicity evaluation. Level of MDA, carbonyl insertion and DNA damage in plasma, livers, and in the lymphocytes of rats exposed to FA were found to be dose dependently increased. Proteomic analysis using three different pI ranges (3.5-5.6, 5.3-6.9, 6-9) and large size two-dimensional gel electrophoresis (2-DE) showed the presence of 3491 protein spots. A total of 32 (19 up- and 13 down-regulated) proteins were identified as biomarkers of FA, all showed dose dependent expressions in the plasma of rats exposed to FA and of these, 27 protein spots were identified by MALDI-TOF/MS. Several differentiated protein groups were found. Proteins involved in apoptosis, transportation, signaling, energy metabolism, and cell structure and motility were found to be up- or down-regulated. Among these, the identities of SNAP 23, apolipoprotein A-1 and E, clusterin, kinesin, and fibrinogen gamma were confirmed by Western blot assay, and apo E was further analyzed by using 2-DE immunoblot assays to determine isoform patterns. Two cytokine including IL4 and INF-gamma were measured in plasma with respect to fibrinogen gamma changes. In summary, cytotoxicity, and genotoxicity assays, namely MDA lipid peroxidation assay, the carbonyl protein oxidation assay, and Comet genotoxic assay showed that these effects increased on increasing FA levels. Proteomic analysis with three different pI ranges and long size 2-DE gel electrophoresis showed that 32 protein spots were up-or down-regulated. Of these 32 proteins, 7 proteins were confirmed by western blot assay. They could be potential biomarkers for human diseases associated with FA exposure.

Oxidative damages in the DNA, lipids, and proteins of rats exposed to isofluranes and alcohols.

DNA damage, lipid peroxidation and protein oxidation were evaluated in rats exposed to a 1% isoflurane atmosphere with or without alcohol administration (administrated by gastric intubation at 4 g/kg body weight as a 50% solution). Single cell gel electrophoresis assays were performed in order to evaluate DNA damage occurring in the lymphocytes, spleen, bone marrow, brain, livers and lung of rats exposed to 1% isoflurane for 30 or 60 min with/without ethanol. Levels of malondialdehydes (MDA), a metabolite of lipid peroxidation, were determined in plasma and tissues. Carbonyl contents were also analyzed to determine levels of protein oxidation in plasma and tissues. Levels of DNA damage in lymphocytes, bone marrow, and the organ tissues of rats exposed to isoflurane were found to increase time dependently, and alcohol increased DNA damage. Lipid peroxidation and protein oxidation results showed patterns that differed from those of DNA damage. Levels of MDA in plasma, bone marrow, spleen, and the livers of rats exposed to isoflurane with/without ethanol were found to be time dependently increased, but this was not observed in the brain or lung. However, protein oxidation levels were significantly increased in the plasma, brains, and lungs of rats exposed to isoflurane, and exposure to isoflurane and alcohol, significantly increased these levels in plasma and brain. The present study demonstrates that isoflurane exposure results in significant DNA damage in rat lymphocytes, bone marrow, spleen, brain, livers, and lung. Moreover, alcohol was found to be as a strong inducer of DNA damage, lipid peroxidation and protein oxidation. However, no evidence in association between DNA damage, lipid peroxidation and protein oxidation was found. Regarding the effects of isoflurane and alcohol on oxidative damages, single strand DNA damages may be a useful biomarkers and blood cells and plasma appear to be more sensitive targets to oxidative damage than other tissues.

Comparison of immunnological and genotoxicological parameters in automobile emission inspectors exposed to polycyclic aromatic hydrocarbons.

In this study, we investigated the immunotoxicities of polycyclic aromatic hydrocarbons in 54 automobile emission inspectors and in 84 control subjects, and evaluated associations between immunological and genotoxicological parameters. Specific surface antigens of peripheral lymphocytes, namely, CD3, CD4, CD8, CD19, and CD69 were subjected to measure immune status in automobile emission inspectors and control subjects. T-and B-cells showed no significant differences between automobile emission inspectors and control subjects (p=0.740 and 0.395). In addition, the ratio of T helper cells to T cytotoxic cells was not deferent (p=0.144). However, T-cell activation was found to be significantly higher in automobile emission inspectors (p=0.041), but not B-cell activation. The levels of two cytokines (IL-4 an INF-γ) and four immunoglobulins (IgA, IgE, IgG, and IgM) were also determined in automobile emission inspectors and control subjects. All immunoglobulin types were lower in automobile emission inspectors, but this was significant only for IgG (0.047). In addition, the levels of two cytokines, IL-4 and INF-γ, were also higher in automobile emission inspectors, though this was not significant. DNA damage in mononuclear and polynuclear lymphocytes and in the level of urinary metabolites, 1-OHP and 2-naphthol, were evaluated in automobile emission inspectors and in control subjects and significant differences were found between the two groups. Examinations of urinary metabolites, DNA damage, and immunological parameters, including leukocyte subpopulations, immunoglobulins, and cytokines, showed that the cytokines levels were associated with the levels of two urinary metabolites, 1-OHP and 2-naphthol.

DNA damage in T and B lymphocytes, bone marrow, spleens, and livers of rats exposed to benzene.

Single-cell gel electrophoresis assays were performed in order to evaluate DNA damage occurring in the T and B lymphocytes, spleens, bone marrow, and livers of rats exposed to benzene at a concentration of 100, 200, or 400 ppm for 2 or 4 wk. The level of t,t-muconic acid (t,t-MA), which is a urinary benzene metabolite, was determined. In the control rats, mean Olive tail moments in the T and B lymphocytes were 1.507 +/- 0.398 and 1.579 +/- 0.206, respectively. DNA damage in the T and B lymphocytes exposed to 400 ppm benzene for 4 wk caused those rats to exhibit the highest Olive tail moments, with their values measured as 4.351 +/- 0.510 and 3.140 +/- 0.631, respectively. Also, the t,t-MA levels increased directly with increasing benzene exposure time and dose during the 4 wk. After 4 wk, the levels of t,t-MA in urine from rats exposed to 100, 200, and 400 ppm were 19.30 +/- 5.62, 30.36 +/- 4.46, and 46.93 +/- 9.10 mg/g creatinine. In conclusion, the present study demonstrates that benzene exposure results in significant DNA damage in the T and B lymphocytes, bone marrow, spleens, and livers of rats. DNA damage in the blood cells and organs was also discovered to vary directly with benzene exposure, in both a dose-dependent and time-dependent manner. In addition, a similar trend regarding DNA damage was found in the blood cells and organs, and evidenced a good association with the level of t,t-MA in the urine.

Evaluation of immuno- and reproductive toxicities and association between immunotoxicological and genotoxicological parameters in waste incineration workers.

In this study, we investigated immunotoxicity levels of 2,3,7,8-tetrachlorodibenzo-p-dioxin in 31 waste incineration workers and in 84 control subjects, and evaluated the association between immunological and genotoxicological parameters. DNA damage in mononuclear and polynuclear lymphocytes, and the level of the urinary metabolites, 1-OHP and 2-naphthol, were evaluated in both waste incineration workers and control subjects. Significant differences were detected in these values between exposed and control groups. Number of sperms was lower in the waste incineration workers than in the control subjects, as was the percentage of motile sperms, but a significant difference existed only in the number of sperms (p=0.05). DNA damage in the spermatozoa of waste incineration workers and control subjects measured 1.40+/-0.08 and 1.26+/-0.03, respectively (p=0.001). Specific surface antigens of peripheral lymphocytes, namely, CD3, CD4, CD8, CD19, and CD69 were used to measure immune status in waste incineration workers and control subjects. There was no significant difference in T- and B-cell profiles showed between waste incineration workers and control subjects (p=0.684 and 0.157). In addition, the ratio of T helper cells to T cytotoxic cells was also not remarkably different between groups (p=0.174). However, T-cell activation was found to be significantly higher in the waste incineration workers than in the control subjects (p=0.001), although B-cell activation did not exhibit this trend. The levels of two cytokines (IL-4 an INF-gamma) and four immunoglobulins (IgA, IgE, IgG, and IgM) were also measured in the experimental population. All immunoglobulin types were found in lower amounts in the waste incineration workers, but this diaparity was not significant one. In addition, the levels of two cytokines, IL-4 and INF-gamma, were also found to be lower in the waste incineration workers than in the control subjects, and only in IL-4 was a significant difference determined to exist.

DNA damage in lymphocytes of benzene exposed workers correlates with trans,trans-muconic acids and breath benzene levels.

Benzene causes many kinds of blood disorders in workers employed in many different environments. These diseases include myelodisplastic syndrome and acute and chronic myelocytic leukemia. In the present study, five occupational work places, including six industrial process types, namely, printing, shoe-making, methylene di-aniline (MDA), nitrobenzene, carbomer, and benzene production were selected, and the levels of breath benzene, and trans,trans-muconic acids (t,t-MA) and phenol in urine were evaluated, as well as hematological changes and lymphocyte DNA damage. The concentration of benzene in breath was less than 3 ppm in the workplaces, and benzene exposure was found to be higher in work places where benzene is used, than in those where benzene is produced. At low levels of benzene exposure, urinary t,t-MA correlated strongly with benzene in air. Highest Olive tail moments were found in workers producing carbomer. Levels of breathzone benzene were found to be strongly correlated with Olive tail moment values in the lymphocytes of workers, but not with hematological data in the six workplaces types. In conclusion, the highest benzene exposures found occurred in workers at a company, which utilized benzene in the production of carbomer. In terms of low levels of exposure to benzene, urinary t,t-MA and DNA damage exhibited a strong correlation with breath benzene, but not with hematological data. We conclude that breath benzene, t,t-MA and lymphocytic DNA damage are satisfactory biomonitoring markers with respect to benzene exposure in the workplace.