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To study how traditional (t,t-muconic acid--t,t-MA and S-phenylmercapturic--SPMA) and new (urinary benzene) urinary biomarkers of internal dose can contribute to exclude an occupational source of exposure to extremely low concentrations of benzene, also analyzing the influence that non-occupational sources of exposure, such as cigarette smoking and urban pollution, can have on the levels of these biomarkers.Assessment was made of 6 workers employed at a groundwater purification plant polluted by benzene (exposed) and 6 administrative clerks employed at the same plant (controls); both groups included smokers and non-smokers. Environmental monitoring (fixed and personal samplings lasting 8 hours) and biological monitoring (determinations of t,t-MA, SPMA, urinary benzene, and urinary creatinine so as to apply suitable adjustments) were performed in exposed workers on 10 successive days, including also rest days (background exposure), and in controls only once.Airborne benzene always resulted lower than the limit of detection of the analytical method in both fixed and personal samplings done on exposed workers and controls during working days, while personal samplings done on exposed workers during rest days showed benzene concentrations even higher than 5 microg/m3, that is the limit value for ambient air quality. Concentrations of t,t-MA, SPMA and urinary benzene did not show differences between exposed workers, regardless of whether they were studied on working or rest days, and controls and appeared to be largely within the reference value range for the Italian population. All biomarkers of internal dose examined in the study showed significantly higher values in smokers than non-smokers. In the latter, SPMA was always below the limit of detection, while urinary benzene resulted higher than the limit of detection in 60.0% and 87.5% of the determinations done on working and on rest days, respectively.In situations of occupational exposure to extremely low doses of benzene or of absence of exposure, the application of an integrated environmental--biological monitoring approach, involving the determination of SPMA and/or urinary benzene, together with a careful evaluation of those factors determining non-occupational exposure to the toxicant, seems indispensable in order to be able to exclude the presence of occupational exposure. In these particular situations of occupational exposure to benzene, the interpretation of the results of environmental and biological monitoring should not only consider the TLV or BEI, but also the limit value for ambient air quality and the reference value for the general population, since benzene is able to determine genotoxic carcinogenic effects even at exposure to extremely low concentrations of the toxicant.

To verify which of the various biomarkers of internal dose of benzene can be considered reliable for biological monitoring of exposure to the low concentrations present nowadays in working and living environments.The specific literature was analyzed to assess the reliability of the different biomarkers of internal dose.T,t-muconic acid is a non specific biomarker for benzene, valid for exposure to concentrations up to one order of magnitude less than the threshold limit of 3250 microg/m3. S-phenylmercapturic acid (SPMA) is a reliable marker even for exposure to concentrations up to two orders below the threshold value of 3250 microg/m3, and can be considered the biomarker of choice for biological monitoring of workers exposed to benzene. Urinary benzene does not seem to have any real advantages over SPMA for monitoring occupational exposure to benzene, but it does seem to be more reliable than SPMA to assess exposure to concentrations like those present in living environments. A smoking habit influences the urinary excretion of all the described biomarkers, and for the current low levels of occupational and environmental exposure to benzene, must be taken into account when interpreting the results of biological monitoring.

This study was aimed to identify useful biomarkers of exposure and effect in workers exposed to low levels of benzene, and to evaluate any correlations existing between these parameters. Benzene exposure was measured in 33 petrochemical industry operators (PIO), 28 service station attendants (SSA), 21 gasoline pump maintenance workers (GPMW) and 51 non-exposed controls by GC-FID analysis. Samples were collected with personal passive samplers (Radiello ® ). End-shift urine samples were collected for t,t-muconic acid (t,t-MA) determination by HPLC and for S-phenylmercapturic acid (S-PMA) measurement by HPLC-MS/MS. The alkaline version of the comet assay and, in a subgroup of 19 SSA and 16 control subjects, chromosomal aberrations (CA) and glutathione (GSH) levels were measured in peripheral blood lymphocytes. Personal benzene exposure was significantly higher in PIO, SSA and GPMW as compared to controls. The urinary excretion of the two metabolites showed a significant increase in SSA (p = 0.0258 and p = 0.0001, for t,t-MA and S-PMA, respectively) and in PIO (p = 0.0013 and p = 0.0001, for t,t-MA and S-PMA, respectively) as compared with the control group, while no such increase was observed for GPMW, for whom occupational exposure was not continuous and occurred on specific working days only. Significant increases of DNA damage were found by the comet assay for tail moment (TM) and tail length (TL) in SSA (p < 0.0001 and p = 0.008, for TM and TL, respectively) and PIO (p < 0.0001 and p < 0.0001, for TM and TL, respectively) when compared with controls. The PIO group also displayed a significant increase in the number of cells with comet (p < 0.0001). Smoking habits did not appear to interfere with these results in any of the groups. No difference was found in percentage of CA between exposed workers and controls. Significant correlations were found, in all groups, between benzene exposure and the more representative comet parameter TM (r = 0.509, p = 0.007; r = 0.525, p = 0.017 and r = 0.420, p = 0.046 in SSA, GPMW, and PIO, respectively). A trend of negative correlation was observed between DNA damage and either GSH or urine S-PMA for exposed workers. In summary, in present study urinary S-PMA and DNA damage by the comet assay were both sensitive to exposure to low levels of benzene, and GSH seems to play an important defence role against benzene-dependent DNA damage.

Objectives This study analyzes the validity of new, more sensitive and speciWc urinary biomarkers of internal dose, namely, urinary benzene for benzene and urinary toluene and S-benzylmercapturic acid (SBMA) for toluene, to assess their eYcacy when compared to traditional biomarkers for biological monitoring of occupational exposure to low concentrations of these two toxic substances. Methods Assessment was made of 41 workers occupationally exposed to benzene and toluene, 18 fuel tanker drivers and 23 Wlling-station attendants, as well as 31 subjects with no occupational exposure to these toxic substances (controls). Exposure to airborne benzene and toluene was measured using passive Radiello® personal samplers worn throughout the work shift. In urine samples collected from all subjects at the end of the workday, both the traditional and the new internal dose biomarkers of benzene and toluene were assessed, as well as creatinine so as to apply suitable adjustments. Results Occupational exposure to benzene and toluene resulted signiWcantly higher in the fuel tanker drivers than the Wlling-station attendants, and higher in the latter than in controls. SigniWcantly higher concentrations of t,t-muconic acid (t,t-MA), S-phenylmercapturic acid (SPMA), urinary benzene, SBMA and urinary toluene were found in the drivers than the Wlling-station attendants or the controls. Instead, urinary phenol and hippuric acid were not diVerent in the three groups. In the entire sample, airborne benzene and toluene values were signiWcantly correlated, as were the respective urinary biomarkers, showing coeYcients ranging from 0.36 to 0.98. Subdividing the subjects by smoking habit, higher coeYcients were evident in nonsmokers than in smokers; at multiple regression analysis t,t-MA, SPMA and urinary benzene and toluene were dependent on the number of cigarettes smoked daily and on airborne benzene and toluene, respectively. Instead, SBMA was dependent only on airborne toluene. Conclusions Our research conWrmed the validity of t,t-MA and SPMA for use in the biological monitoring of exposure to low concentrations of benzene. Urinary benzene showed comparable validity to SPMA; both parameters are aVected by smoking cigarettes in the hours before urine collection, so it is best to ask subjects to refrain from smoking for 2 h before urine collection. Urinary toluene was found to be a more speciWc biomarker than SBMA.

OBJECTIVE: To verify whether urinary benzene is an applicable biomarker of occupational exposure to very low concentrations of benzene, considering the influence of cigarette smoke and benzene-toluene co-exposure. MATERIALS AND METHODS: 23 filling station attendants with occupational exposure to benzene and 31 controls were analyzed. Occupational and environmental exposure was monitored and t,t-muconic acid (t,t-MA), S-phenylmercapturic acid (SPMA), urinary benzene and creatinine in the urine samples were tested. RESULTS: Occupational exposure to benzene and toluene was significantly higher in the filling station attendants than in the controls, whereas t,t-MA, SPMA and urinary benzene were not different in the two groups. Instead, the smoker group showed significantly higher values for the above biomarkers than the non-smoker group, each of which included both exposed workers and controls. SPMA was dependent on airborne benzene and cigarette smoking, and urinary benzene only on cigarette smoking, while t,t-MA was not dependent on either of these variables. CONCLUSIONS: At very low concentrations of occupational exposure to benzene, urinary benzene is less valid than SPMA as a biomarker, even if both are strongly influenced by smoking habit. Abstention from smoking should therefore be recommended for at least two hours before urine collection.