Trends in occupational exposure to styrene in the European glass fibre-reinforced plastics industry.

AIM: This study presents temporal trends of styrene exposure for workers in the European glass fibre-reinforced plastics (GRP) industry during the period 1966-2002. METHODS: Data of personal styrene exposure measurements were retrieved from reports, databases and peer-reviewed papers. Only sources with descriptive statistics of personal measurements were accepted. The styrene exposure data cover personal air samples and biological monitoring data, that is, urinary styrene metabolites (mandelic acid and/or phenylglyoxylic acid) and styrene in blood. Means of series of measurements were categorized by year, country, production process, job and sampling strategy. Linear mixed models were used to identify temporal trends and factors affecting exposure levels. RESULTS: Personal exposure measurements were available from 60 reports providing data on 24145 1-8-h time-weighted average shift personal air samples. Available data of biological exposure indicators included measurements of mandelic acid in post-shift urine (6361 urine samples being analysed). Trend analyses of the available styrene exposure data showed that the average styrene concentration in the breathing zone of open-mould workers in the European GRP industry has decreased on average by 5.3% per year during the period 1966-1990 and by only 0.4% annually in the period after 1990. The highest exposures were measured in Southern Europe and the lowest exposures in Northern Europe with Central Europe in between. Biological indicators of styrene (mandelic acid in post-shift urine) showed a somewhat steeper decline (8.9%), most likely because urine samples were collected in companies that showed a stronger decrease of styrene exposure in air than GRP companies where no biological measurements were carried out.

Polycyclic aromatic hydrocarbon-DNA adducts in white blood cell DNA and 1-hydroxypyrene in the urine from aluminum workers: relation with job category and synergistic effect of smoking.

We examined a group of 105 workers from a primary aluminum plant for the presence of polycyclic aromatic hydrocarbon (PAH)-DNA adducts in their WBC and 1-hydroxypyrene in their urine. Workers were recruited from five job categories with different PAH exposure: the anode factory; the bake oven; and the electrolysis and the pot-relining departments. Unexposed workers from the foundry department served as the control group. The exposure to PAH was measured by personal monitoring, and the average PAH concentrations in the work atmosphere ranged from 0.4 micrograms/m3 in the foundry to 150 micrograms/m3 in the pot-relining department. The average exposure to benzo(a)pyrene was under the Swedish exposure limit of 5 micrograms/m3. The internal dose of pyrene was measured utilizing the 1-hydroxypyrene concentration in pre- and postshift urine samples. Higher exposure to PAH in the work atmosphere was associated with increased concentrations of 1-hydroxypyrene in the urine. The average increase in concentration of 1-hydroxypyrene ranged from 0.2 mumol/mol creatinine in the control group to 5.9 mumol/mol creatinine in the pot-relining department; an accumulation of 1-hydroxypyrene over a 5-day working period was observed. A good correlation was found between PAH exposure and the concentration of 1-hydroxypyrene in the urine on a group level (rs = 0.90; P = 0.02). PAH-DNA adducts were determined by 32P-postlabeling analysis (nuclease P1 enrichment procedure).(ABSTRACT TRUNCATED AT 250 WORDS)

Urinary excretion of 3-hydroxy-benzo[a]pyrene after percutaneous penetration and oral absorption of benzo[a]pyrene in rats.

A method for the monitoring of exposure to benzo[a]pyrene was applied to rat urine samples. Benzo[a]pyrene was administered percutaneously and orally with repeated doses of 10, 20 and 50 mumol/kg body wt. At low dose levels, the urinary excretion of 3-OH-benzo[a]pyrene was higher after percutaneous treatment than after oral administration. The excretion of 4 benzo[a]pyrene metabolites, including 3-OH-benzo[a]pyrene, is faster after oral administration. The percutaneous absorption of small doses of benzo[a]pyrene appears to be greater than the oral absorption.

Microsomal lauric acid hydroxylase activities after treatment of rats with three classical cytochrome P450 inducers and peroxisome proliferating compounds.

In order to investigate a proposed relationship between induction of hepatic microsomal lauric acid hydroxylase activity and peroxisome proliferation in the liver, male Wistar rats were treated with peroxisome proliferating compounds, and the lauric acid hydroxylase activity, the immunochemical detectable levels of cytochrome P450 4A1 and the activities of peroxisomal enzymes were determined. In addition, the levels of cytochrome P450 4A1 and lauric acid hydroxylase activities were studied after treatment of rats with three cytochrome P450 inducers. After treatment with aroclor-1254, phenobarbital or 3-methylcholanthrene total cytochrome P450 was 1.7-2.7 times induced. However, no induction of lauric acid omega-hydroxylase activities or P450 4A1 levels were found. After treatment of rats with di(2-ethylhexyl)phthalate (DEHP) a dose-dependent induction of lauric acid omega-hydroxylase activities, levels of cytochrome P450 4A1 and peroxisomal fatty acid beta-oxidation was found. Even at a dose-level of 100 mg DEPH/kg body weight per day a significant induction of these activities was observed. The main metabolites of DEHP, mono(2-ethylhexyl)phthalate and 2-ethyl-1-hexanol, also caused an induction of levels of P450 4A1, lauric acid omega-hydroxylase activities and the activity of peroxisomal palmitoyl-CoA oxidase. 2-Ethyl-1-hexanoic acid did not influence lauric acid omega-hydroxylase activities, but did induce levels of P450 4A1 and palmitoyl-CoA oxidase activities. Three other compounds (perfluoro-octanoic acid, valproate and nafenopin) induced both lauric acid omega-hydroxylase activity and peroxisomal palmitoyl-CoA oxidase activity. The plasticizer, di(2-ethylhexyl)adipate, did not induce levels of P450 4A1, lauric acid omega-hydroxylase activities or palmitoyl-CoA oxidase activities. With the compounds tested a close association between the induction of lauric acid omega-hydroxylase activities and peroxisomal palmitoyl-CoA oxidase activity was found. These data support the theory that peroxisome proliferating compounds do induce lauric acid omega-hydroxylase activities and that there might be a mechanistic inter-relationship between peroxisome proliferation and induction of lauric acid omega-hydroxylase activities.

Effects of the peroxisome proliferator mono(2-ethylhexyl)phthalate in primary hepatocyte cultures derived from rat, guinea pig, rabbit and monkey. Relationship between interspecies differences in biotransformation and peroxisome proliferating potencies.

Primary hepatocyte cultures derived from rat, rabbit, guinea pig and monkey have been treated in vitro with metabolites of di(2-ethylhexyl)phthalate, i.e. mono(2-ethylhexyl)phthalate (MEHP), mono(5-carboxy-2-ethylpentyl)phthalate (metabolite V) and mono(2-ethyl-5-oxohexyl)phthalate (metabolite VI). In rat hepatocyte cultures MEHP and metabolite VI were equally potent in inducing peroxisome proliferation, while metabolite V was much less potent. In rat hepatocytes a 50% increase in both peroxisomal palmitoyl-CoA oxidase activity and microsomal lauric acid omega-hydroxylation activity was found after treatment with 5-15 microM MEHP. In guinea pig, rabbit and monkey hepatocyte cultures, a 50% increase in peroxisomal palmitoyl-CoA oxidase activity was found after treatment with 408-485 microM MEHP. No induction of lauric acid omega-hydroxylation activity was found. These results indicate that peroxisome proliferation can be induced by MEHP in rabbit, guinea pig and monkey hepatocytes, but that these species are at least 30-fold less sensitive to peroxisome proliferation induction than rats. The proposed mechanistic inter-relationship between induction of lauric acid omega-hydroxylation activity and peroxisome proliferation is found in rat hepatocytes, but not in hepatocytes of the other three species. Treatment of guinea pig hepatocyte cultures with MEHP resulted in an increase in triglyceride concentrations in the hepatocytes. In rat and rabbit hepatocyte cultures, triglyceride concentrations were much less altered by MEHP. In monkey hepatocytes a decrease in hepatic triglyceride concentration was found after treatment with MEHP. These effects are in agreement with in vivo effects observed before. After treatment of primary hepatocyte cultures with MEHP, high concentrations of omega- and (omega-1)-hydroxylated metabolites of MEHP were found in media from rat, rabbit and guinea pig cultures. The formation of these metabolites did not decline in time. During treatment the metabolite profile in media from rat hepatocyte cultures moved towards omega-hydroxy metabolites of MEHP. In media from monkey hepatocyte cultures the lowest concentrations of hydroxylated metabolites were determined. No major species differences were found in the potency to form oxidized MEHP metabolites, and thus no unique metabolite differences were found, which could explain the species differences in sensitivity for peroxisome proliferation.

Effect of di(2-ethylhexyl)phthalate on enzyme activity levels in liver and serum of rats.

In order to identify non-invasive, biochemical indicators of di(2-ethylhexyl)phthalate (DEHP) exposure, we have compared the effects in blood serum with biochemical effects in liver in rats fed a diet containing 0, 0.25, 0.75 and 2% DEHP for 2 weeks. After 3 days of treatment serum arylesterase activity levels and serum triglycerides were decreased to 60% and 20% of control values, respectively. After a 2-week treatment with DEHP the effects were generally stronger. Compared to a control group, serum arylesterase activity levels, serum triglycerides and serum cholesterol were decreased to 40%, 20% and 50%, respectively. Serum cholinesterase activity levels and serum albumin concentrations were increased by the DEHP treatment to 290% and 135% of control values, respectively. In the livers a hepatomegaly, an induction of cytochrome P-450 IVA1 and induction of the activity of palmitoyl-CoA oxidase and carnitine acetyl-CoA transferase was found to be 180%, 1080%, 1300% and 1700% of control values, respectively. The liver is a more sensitive target for DEHP exposure compared to the biochemical effects in serum, but determination of the serum parameters can be used to determine early biological effects of exposure to DEHP.

Biological monitoring of environmental exposure to polycyclic aromatic hydrocarbons; 1-hydroxypyrene in urine of people.

A biomarker of human exposure to chemical agents provides a valuable parameter for assessing the extent and significance of the uptake by giving a measurement that is direct and integrated over time and exposure routes. Urinary 1-hydroxypyrene is currently tested as a biomarker for the assessment of low level environmental exposure of people to polycyclic aromatic hydrocarbons (PAH). Five examples of the application of urinary 1-hydroxypyrene methodology in the assessment of environmental exposure to PAH are presented: inhalation of tobacco smoke; inhalation of urban outdoor air; windsurfers sailing on polluted water; absorption of contaminated food; exposure in an urban area with many heavy industries. The examples illustrate that the urinary 1-hydroxypyrene test is sufficiently sensitive. Urinary 1-hydroxypyrene is an effective biomarker for the assessment of human environmental exposure to PAH.

Mutagenicity of bi-, tri- and tetra-cyclic aromatic hydrocarbons in the "taped-plate assay" and in the conventional salmonella mutagenicity assay.

Aromatic hydrocarbons in the range of 1-4 nuclear rings were examined for mutagenicity in the so-called "taped-plate assay". This modification of the Ames assay is particularly equipped for the detection of volatile mutagens. Of the many compounds tested only phenanthrene, pyrene, benzo[c]phenanthrene and benzoacenaphthylene were positive in this assay. The present data underline the exceptional behaviour of fluoranthene by being a rather potent bacterial mutagen with a volatile nature (as found in a previous study).

Detection of volatile mutagens in creosote and coal tar.

Creosote and coal tar were examined for the presence of volatile mutagens by use of the so-called "taped plate assay". Application of this method, recently published by Distlerath et al., reveals that vapour escaping from creosote and coal tar at 37 degrees C was able to revert the Salmonella typhimurium strains TA98 and TA100 in the presence of S9 mix. The simplicity of this method makes it useful for routine screening of industrial fluids or solid products for the presence of volatile mutagens.

Non-mutagenicity of 4 metabolites of di(2-ethylhexyl)phthalate (DEHP) and 3 structurally related derivatives of di(2-ethylhexyl)adipate (DEHA) in the Salmonella mutagenicity assay.

Four metabolites of the rat liver carcinogen di(2-ethylhexyl)phthalate (DEHP) (mono-(2-ethylhexyl)phthalate, mono-(2-ethyl-5-hydroxyhexyl)phthalate, mono-(2-ethyl-5-oxohexyl)phthalate, and mono-(5-carboxy-2-ethylpentyl)phthalate) and 3 structurally related derivatives of di(2-ethylhexyl)adipate (DEHA) (mono-(2-ethylhexyl)adipate, mono-(2-ethyl-5-hydroxyhexyl)adipate, and mono-(2-ethyl-5-oxohexyl)adipate) were tested for mutagenicity in the Ames assay using Salmonella typhimurium strains TA97, TA98, TA100, and TA102, with and without a metabolic activation preparation. Aroclor 1254-induced rat liver S9 and DEHP-induced rat liver S9 were used. Concentrations of these compounds up to 1000 micrograms/plate were negative with all tester strains in the presence or absence of metabolic activation.

1-Hydroxypyrene as an indicator of the mutagenicity of coal tar after activation with human liver preparations.

Liver S9 fractions were prepared from male Wistar rats, either non-induced or induced with Aroclor 1254 and from 5 human kidney transplant donors. The preparations were compared for their ability to metabolize the premutagens present in coal tar to mutagenic metabolites in the Salmonella mutagenicity assay towards strain TA98. Low levels of mutagenicity of coal tar were seen with human S9 preparations. The differences between the S9 mix of the 5 donors in capacity to activate premutagens were approximately 6-fold. The activation of coal tar by rat liver S9 preparations was higher than by the human S9 preparations. The metabolic conversion of pyrene in coal tar to 1-hydroxypyrene by the same human S9 preparations was determined in a parallel assay. 3 human preparations showed a high correlation between the formation of 1-hydroxypyrene and bioactivation of coal tar to mutagenic metabolites. The slope values of the individual regression lines were equal, suggesting that 1-hydroxypyrene is a good indicator for the activation of premutagens present in coal tar.

Fluoranthene, a volatile mutagenic compound, present in creosote and coal tar.

Creosote, a coal-tar distillation product, contains mutagens which are volatile at 37 degrees C. After distillation of creosote we found that these volatile mutagens were present in the distillation fraction with the highest boiling range (greater than 360 degrees C). The "volatile mutagenic activity" was connected with the presence of fluoranthene, a polycyclic aromatic hydrocarbon. Commercially available fluoranthene was positive in the so-called "taped-plate assay" (the test system used for the detection of volatile mutagens) towards the strains TA98 and TA100 in the presence of S9 mix. The tested creosote and coal tar contained fluoranthene in concentrations of 5.2 and 2.2%, respectively.

Urinary 1-hydroxypyrene as a biomarker of polycyclic aromatic hydrocarbons exposure of workers on a contaminated site: influence of exposure conditions.

The aim of the study was to determine the exposure levels of workers to polycyclic aromatic hydrocarbons on gasworks sites by the measurement of urinary 1-hydroxypyrene. Start-shift and end-shift urine samples were taken every day during an entire week (Monday to Friday), once in November and a second time in June. Four groups of workers were selected according to their activity. Increased exposure was only found among volunteers involved in the remediation of a site, 0.16 to 2.31 mumol/mol creatinine in non-smokers. The median of the non-smoker referent group was 0.02 mumol/mol creatinine (95% confidence interval, 0.01 to 0.04). Smokers had greater exposure levels than non-smokers in every group. Within and between variability was around 200%. Assessment of the exposure of persons on contaminated soil is possible, with the condition that the exposed subjects come in direct contact with the soil.

Effect of the reduction of skin contamination on the internal dose of creosote workers exposed to polycyclic aromatic hydrocarbons.

Ten creosote-exposed workers of a wood impregnation plant participated in this study, which took place in two consecutive weeks on a Monday, after a weekend off. On one of the two days each worker wore Tyvek coveralls underneath his normal workclothes. Dermal contamination measurements (pyrene on exposure pads) and biological monitoring (urinary 1-OH-pyrene) were performed to measure the reduction of both the skin contamination and the internal dose. The total pyrene skin contamination of workers not wearing coveralls ranged between 47 and 1510 micrograms.d-1 (0.2-7.5 mumol.d-1). On the average, the coveralls reduced the pyrene contamination on the workers' skin by about 35 (SD 63)%. The excreted amount of 1-OH-pyrene in urine decreased significantly from 6.6 to 3.2 micrograms (30.2 to 14.7 nmol). Multiple regression analysis showed that skin contamination by polycyclic aromatic hydrocarbons is the main determinant of the internal exposure dose of creosote workers.

Biological monitoring of polycyclic aromatic hydrocarbons. Metabolites in urine.

Assays of urinary mutagenicity, urinary 3-hydroxy-benzo(a)pyrene, and urinary 1-hydroxypyrene were used to study their suitability in estimating exposure to polycyclic aromatic hydrocarbons (PAH) in coal tar products. Rats exposed to coal tar solutions applied on the dorsal skin excreted mutagens, 3-hydroxy-benzo(a)pyrene, and 1-hydroxypyrene dose dependently in their urine. The correlation between the three parameters was high. Five dermatologic patients undergoing topical coal tar treatment excreted low concentrations of 3-hydroxy-benzo(a)pyrene and high concentrations of 1-hydroxypyrene. A significant correlation between the excretion of the two metabolites was found. The smoking workers of a coal tar distillation plant showed a significantly enhanced urinary mutagenicity compared with their nonsmoking colleagues, but an increase due to occupational exposure was not found. However, the concentration of 1-hydroxypyrene in the urine of these workers highly exceeded the upper 95 percentile of a reference population. The urinary excretion of 1-hydroxypyrene of smoking referents was not significantly increased compared with that of nonsmoking referents. The data suggest that urinary 1-hydroxypyrene is a sensitive and specific marker for the assessment of occupational exposure to PAH.

Methods for routine biological monitoring of carcinogenic PAH-mixtures.

The ability of a biomarker to provide an assessment of the integrated individual dose following uptake through multiple routes is especially valuable for mixtures of polycyclic aromatic hydrocarbons (PAH), due to methodological and practical difficulties of collecting and analysing samples from the various environmental compartments like air, water and soil and various media such as diet, cigarette smoke and workroom air. Since 1980, a large variety of novel approaches and techniques have been suggested and tested, e.g. urinary thioethers, mutagenicity in urine, levels of PAH or PAH-metabolites in blood and urine and methods for determination of adducts in DNA and proteins. Two approaches are more frequently reported: PAH-DNA-adduct monitoring in blood cells and urinary 1-hydroxypyrene monitoring. A large research effort has been made to use the extent of binding of PAH to DNA as a biomarker of exposure. The 32P-post-labeling assay detects the total of aromatic DNA-adducts and the adduct level in white blood cells is claimed to be an indicator of the biological effect of the PAH-mixture. However, the levels of aromatic DNA-adducts may be subject to appreciable analytical and biological variation. The present technical complexity of the method makes it more convenient for research applications than for routine application in occupational health practice. Pyrene is a dominant compound in the PAH mixture and is mainly metabolised to the intermediary 1-hydroxypyrene to form 1-hydroxypyrene-glucuronide, which is excreted in urine. Since the introduction of the determination of 1-hydroxypyrene in urine as a biomarker for human exposure assessment in 1985, many reports from different countries from Europe, Asia and America confirmed the potential of this novel approach. The conclusion of the first international workshop on 1-hydroxypyrene in 1993 was that urinary 1-hydroxypyrene is a solid biological exposure indicator of PAH. Studies with a comparison of several biomarkers confirmed that 1-hydroxypyrene in urine is a valid and sensitive indicator of exposure. Periodical monitoring of 1-hydroxypyrene appears to be a powerful method in controlling occupational PAH-exposure in industries. The reference level and the biological exposure limit of 1-hydroxypyrene in urine are discussed.

S-phenyl-N-acetylcysteine in urine of rats and workers after exposure to benzene.

An HPLC method for the determination of S-phenyl-N-acetylcysteine in urine is described. The sensitivity is 6 mumol/L (CV = 9%) urine. Exposure of rats to six different concentrations of benzene, ranging from 0-30 ppm, was highly associated with urinary excretion of S-phenyl-N-acetylcysteine (r = 0.86) and with total phenol (r = 0.81). A background level of phenol was found in urine of both non-exposed rats and of non-exposed referents. However, no background excretion of S-phenyl-N-acetylcysteine was found, either in rats or in humans. In urine of exposed rats, the level of S-phenyl-N-acetylcysteine was approximately five times lower than the phenol level. Workers occupationally exposed to benzene, showing high levels of urinary phenol, revealed low concentrations of urinary S-phenyl-N-acetylcysteine. The biological monitoring of industrial exposure to benzene by determination of S-phenyl-N-acetylcysteine in urine is not better than the determination of phenol in urine.

Dermal exposure to polycyclic aromatic hydrocarbons among primary aluminium workers.

Large amounts of PAH's are released in the electrode production departments of pre-bake cell aluminium reduction plants. Emission sources are mixing, shaping and baking of the anode (paste plant and bake oven) and pot relining operations. A study was performed to quantify the importance of dermal uptake of PAH's among exposed workers. Twenty workers in the anode production departments (paste plant (N = 8) and bake oven (N = 5)) and the pot relining department (N = 7) volunteered for the study. Monitoring was performed over a period of 5 consecutive days using personal air sampling, dermal contamination sampling and biological monitoring. Pyrene concentrations measured in the respirable air samples, ranged up to 320 micrograms/m3. Dermal contamination of pyrene was monitored at three skin sites (wrist, jaw/neck and groin) using exposure pads as pseudo-skin. The skin contamination with pyrene ranged up to 375 ng/cm2. Contamination of the groin skin site, although covered by work clothes ranged up to 106 ng/cm2. The concentration of 1-hydroxypyrene in pre and post-shift urine ranged up to 27 mumol/mol creatinine and showed an increase during the day and a decrease during the night. Pyrene in air and pyrene on the skin were tested for significance of correlation with urinary 1-hydroxypyrene in samples taken at several moments: end-of-shift, pre-shift next morning and weekly increase. The correlation coefficients between dermal contamination and urinary 1-hydroxypyrene were equal or higher than the correlation coefficient between pyrene air concentration and urinary 1-hydroxypyrene. The total skin contamination in exposed workers is estimated to be more than three times higher than the intake via the respiratory tract. The contribution of dermal exposure to the total PAH body burden of exposed workers therefore appears to be significant.