1. 9th International Symposium on Biological Monitoring in Occupational and Environmental Health (ISBM 2013)GENOTOXIC EFFECTS OF OCCUPATIONAL AND ENVIRONMENTAL EXPOSURE TO LOW CONCENTRATIONS OF BENZENE Lovreglio P. 1, Maffei F.2, Carrieri M.3, DErrico M.N.1, Drago I.1, Hrelia P.2, Bartolucci G.B.3, Soleo L.1 1InterdisciplinaryDepartment of Medicine, Section of Occupational Medicine E.C. Vigliani, University of Bari, Italy 2Department of Pharmacology, University of Bologna, Italy 3Department of Molecular Medicine, Section of Occupational Medicine, University of Padova, Italy8-11 September 2013, Manchester, UK

2. INTRODUCTION BENZENEOccupational toxicantEnvironmental toxicantMain sources: - automobile exhaust - cigarette smokeExposure to benzene urban pollution (always at low or very low concentrations) can affect: - not only the general population , - but also workers in urban traffic, such as traffic wardens, bus drivers, taxi drivers, etc.Because of its effects on human health (also carcinogenic), the use of benzene in industrial applications has been strictly limited by specific regulations in western nations.Nevertheless, occupational exposure to low or very low concentrations is still possible in workers in: - upstream/downstream petrochemical industry; - chemical synthesis industry; - coke ovens; - fuel distribution. 3. INTRODUCTION Benzene has a well known clastogenic effect in humans CHROMOSOME DAMAGE BIOMARKERS Frequency of chromosome aberrations (CA) in peripheral blood lymphocytesFrequency of micronuclei (MN) in peripheral blood lymphocytesWorkers exposed to concentrations of benzene exceeding or near to the TLV-TWA of the ACGIH, i.e. 1600 g/m3 Workers exposed to lower concentrations, up to 2 orders of magnitude below the TLV-TWABoth workers exposed to benzene concentrations exceeding or near to the TLV-TWA and workers exposed to lower concentrations than the TLV-TWA .Evident increase in the frequency of CA, reported in numerous studies. Few studies, with limited information... studies have shown conflicting results and poor agreement.For the genotoxic effects of low and very low concentrations of benzene, like those currently present in the workplace and living environment, the exposure-response relationship is controversial yet. 4. OBJECTIVE OF THE RESEARCH To study possible genotoxic effects of benzene, by determining the frequency of CA and MN in the peripheral blood lymphocytes of workers occupationally exposed to low and very low concentrations of the toxicant. 5. METHODS 19 male fuel tanker drivers EXPOSED 24 male filling station attendantsCONTROLS31 male workers with no occupational exposure to benzene, resident in the same geographic areaAll subjects were administered a questionnaire probing: - general characteristics - current and previous work experience - smoking history and alcohol consumption - clinical history - use of medicinal drugs and exposure to ionizing radiation in the last 12 months - possible exposure to non occupational sources of benzene. Exclusion criteria for subjects enrolled in the study: - previous treatment with ionizing radiation or chemotherapeutic drugs; - radiography in the 12 months before the sampling; - a previous diagnosis of cancer; - for exposed workers, less than 1 year's work in the current job. 6. METHODS EXTERNAL DOSE BIOMARKER: Airborne benzene- passive personal sampling (Radiello) for 8 hours; - blinded analyses performed by GC-FID; limit of detection (LOD)=3.0 g/m3.INTERNAL DOSE BIOMARKERS: blinded analyses performed on urine samples collected at the end of the environmental sampling.- t,t-muconic acid (HPLCUV method at 264 nm, after solid phase extraction (SPE); LOD=10 g/L) - SPMA (HPLCESIMS/MS; LOD=0.20 g/L) - Urinary benzene (GC/MS analysis; LOD=0.02 g/L) - Urinary creatinine (colorimetric method; LOD=0.01 mg/dL).EFFECT BIOMARKERS: peripheral blood collection before the start of the environmental sampling; heparinized blood samples were code labeled, transported at room temperature and cultured for cytogenetic tests within 24 hours of collection; all the phases following blood collection were performed blinded.- CA by standard technique; at least 100 metaphases were counted and scored for each subject; assessment was made of total CA and separately, of chromosome and chromatid breaks; gaps were excluded from the analyses.- MN by standard cytokinesis block technique; for each subject, 2000 binucleated cells (BN) were examined; the frequency was expressed as total MN per 1000 cells analyzed.- Nuclear Division Index (NDI): calculated by scoring at least 1000 cells per series of cultures,adopting the formula NDI = (M1+2M2+3M3+4M4)/n, [M1M4 =number of cells with 1-4 nuclei; n = total number of cells scored]. 7. RESULTS Table 1: General characteristics and lifestyle of the fuel tanker drivers, filling station attendants and controls included in the study. FUEL TANKER DRIVERS N.MeanSD/% Median RangeAge (years)1942.17.542.0Body mass index (Kg/m2)1927.84.026.8Duration of employment (years)1916.69.317.02 4 1311 8Alcohol consumption - teetotal - 10 g/day29-57FILLING STATION ATTENDANTS N. MeanSD/% Median RangeN. MeanSD/% Median243141.79.13126.74.8 25.8 20.9-44.420.8-35.9 24 5-3740.79.6 40.019-6026.13.9 26.5 20.3-37.2 13.5CONTROLS2414.28.73-3610.5% 21.1% 68.4%7 7 1029.2% 29.2% 41.6%10 9 1212 1250.0% 50.0%16 1551.6% 48.4%1616.610.622-6232.3% 29.0% 38.7%57.9% 42.1%43.0Range-Smoking habit Smoker Non smokerCigarettes/day 11 (smokers)21.510.620.06-501217.711.120.0Packs/year (smokers)20.411.221.53.3-401227.017.625.0112-40 1-621623.419.411.05-4012.77-61 8. Table 2: Airborne benzene concentrations in the fuel tanker drivers, filling station attendants and controls. FUEL TANKER DRIVERS (N. 19) MeanSD Median Range Airborne benzene 306.7266.7 246.6 7.4-1017.1 (g/m3)aFILLING STATION ATTENDANTS (N. 24) MeanSD Median Range 23.317.019.94.5-66.3CONTROLS (N. 31) MeanSD Median Range 4.62.64.3