A. Alt

Fast determination of urinary S-phenylmercapturic acid (S-PMA) and S-benzylmercapturic acid (S-BMA) by column-switching liquid chromatography-tandem mass spectrometry

Benzene and toluene are important industrial chemicals and ubiquitous environmental pollutants. The urinary mercapturic acids of benzene and toluene, S-phenylmercapturic acid (S-PMA) and S-benzylmercapturic acids (S-BMA) are specific biomarkers for the determination of low-level exposures. We have developed and validated a fast, specific and very sensitive method for the simultaneous determination of S-PMA and S-BMA in human urine using an automated multidimensional LC-MS-MS-method that requires no additional sample preparation. Analytes are stripped from urinary matrix by online extraction on a restricted access material, transferred to the analytical column and subsequently determined by tandem mass spectrometry using isotopically labelled S-PMA as internal standard. The lower limit of quantification (LLOQ) for both analytes was 0.05 microg/L urine and sufficient to quantify the background exposure of the general population. Precision within series and between series for S-PMA and S-BMA ranged from 1.0% to 12.2%, accuracy was 108% and 100%, respectively. We applied the method on spot urine samples of 30 subjects of the general population with no known exposure to benzene or toluene. Median levels (range) for S-PMA and S-BMA in non-smokers (n=15) were 0.14 microg/L (<0.05-0.26 microg/L) and 8.2 (1.6-77.4 microg/L), respectively. In smokers (n=15), median levels for S-PMA and S-BMA were 1.22 microg/L (0.17-5.75 microg/L) and 11.5 microg/L (0.9-51.2 microg/L), respectively. Due to its automation, our method is well suited for application in large environmental studies.

Plasma Polychlorinated Biphenyls (PCB) Levels of Workers in a Transformer Recycling Company, their Family Members, and Employees of Surrounding Companies

In spring 2010, high internal exposures (up to 236 μg/L plasma) for the sum of indicator polychlorinated biphenyls (PCB) were discovered in workers in a transformer recycling company in Germany, where PCB-contaminated material was not handled according to proper occupational hygiene. The release of PCB from this company raised growing concerns regarding possible adverse human health effects correlated with this exposure. This provided a basis for a large biological monitoring study in order to examine the internal exposure to PCB in individuals working in that recycling company, their family members, and relatives, as well as subjects working or living in the surroundings of this company. Blood samples from 116 individuals (formerly) employed in the transformer recycling company and 45 direct relatives of these persons were obtained. Further, blood samples of 190 subjects working in close vicinity of the recycling plant, 277 persons working in the larger area, and 41 residents of the area were investigated. Plasma samples were analyzed for the 6 indicator PCB (PCB 28, 52, 101, 138, 153, 180) and 12 dioxin-like PCB using gas chromatography/mass spectroscopy (GC/MS; limit of detection [LOD] at 0.01 μg/L). Median concentrations (maximum) for the sum of the 6 indicator PCB in blood of the employees, their relatives, individuals working in close vicinity, persons working in the larger area, and the residents were 3.68 (236.3), 1.86 (22.8), 1.34 (22.9), 1.19 (6.42), and 0.85 (7.22) μg/L plasma, respectively. The (former) employees of the transformer recycling plant partly showed the highest plasma PCB levels determined thus far in Germany. Even family members displayed highly elevated levels of PCB in blood due to contaminations of their homes by laundering of contaminated clothes. Vicinity to the recycling plant including reported contact with possibly contaminated scrap was the main contributor to the PCB levels of the workers of the surrounding companies. Residents of the area did not show significantly elevated blood PCB levels compared to the general population. Our biomonitoring results served as a basis for individual risk communication and successful risk management.

Biological monitoring of indoor-exposure to dioxin-like and non-dioxin-like polychlorinated biphenyls (PCB) in a public building

The release of PCBs from sealant material in public buildings and the resulting indoor air levels have raised growing concerns about possible human health effects connected with this exposure. Ambient monitoring of PCBs in a public building has revealed a contamination with the more volatile lower chlorinated PCB 28, PCB 52 and PCB 101. This gave reason for a large biological monitoring study in order to examine the internal exposure to PCBs in persons working in that building. Blood samples from 209 persons employed in the PCB-contaminated building were drawn. 98 persons matched for age and gender working in non-contaminated buildings served as control group. Plasma samples were analysed for the six indicator PCBs (PCB 28, 52, 101, 138, 153, 180) and 12 dioxin-like PCBs using GC/MS (LOD: 0.01 μg/L). Significant differences between both collectives were only found for the plasma levels of the lower chlorinated PCB 28, PCB 52 and PCB 101 and for the dioxin-like congeners PCB 105 and PCB 118, which are due to inhalative exposure to these congeners via indoor air. Median plasma levels of PCB 28, PCB 52 and PCB 101 in the employees of the contaminated building were 0.087 μg/L, 0.024 μg/L and 0.012 μg/L, respectively. The concentrations of the higher chlorinated PCBs and all other dioxin-like congeners investigated were within the normal range of the general population. There was no relationship between indoor air measurements and internal exposure of the employees in the corresponding office, but estimated lifetime exposure of the employees turned out to be a significant factor for plasma levels of PCB 28. Our biomonitoring results served as a basis for individual risk communication and successful risk management.

A biomarker approach to estimate the daily intake of benzene in non-smoking and smoking individuals in Germany.

Owing to its carcinogenic properties, benzene is one of the most important environmental air pollutants. We have applied a simple pharmacokinetic model to estimate the individual daily exposure of persons of the general population to benzene using their urinary excretion of S-phenylmercapturic acid as biomarker of exposure. On the basis of a non-representative convenience sample of the general population, spontaneous urine samples of 43 non-smoking persons, 13 persons with exposure to environmental tobacco smoke (ETS) (as determined by urinary cotinine) and 72 smokers were analyzed for S-phenylmercapturic acid, and benzene exposure was back calculated on the basis of the results. The pharmacokinetic model was based either on estimated daily urinary volume or creatinine excretion. Median daily exposure of non-smokers was calculated to be 47 μg/day (volume-based model) and 63 μg benzene/day (creatinine-based model). ETS-exposed persons had a slightly higher median daily exposure of 65 μg/day (volume-based model) and 72 μg benzene/day (creatinine-based model). The daily exposure of smokers was significantly higher with median values of 491 μg benzene/day (volume-based model) and 693 μg benzene/day (creatinine-based model). Our biomarker-based model gave plausible results for daily benzene exposure that were in good agreement with exposure estimations published earlier. As it is purely based on the determination of individual internal dose, our model provides a powerful tool for the risk assessment of environmental benzene.

Simple and sensitive GC/MS-method for the quantification of urinary phenol, o- and m-cresol and ethylphenols as biomarkers of exposure to industrial solvents.

We have developed and validated a simple and sensitive method for the determination of urinary phenol as well as the urinary metabolites of toluene and ethylbenzene in one analytical run. After enzymatic hydrolysis for the cleavage of conjugates overnight, the analytes are extracted from the matrix with a liquid-liquid extraction procedure using toluene as solvent under acidic conditions. The analytes are then derivatised to volatile ethers using N,O-bis(trimethylsilyl) trifluoroacetamid (BSTFA) for cresols and ethylphenols as well as N-tert-butyldimethylsilyl-N-methyltrifluoroacetamid (MTBSTFA) for the determination of phenol. Separation and detection was carried out using capillary gas chromatography with mass-selective detection (GC-MS). Deuterium-labeled o-cresol served as internal standard for the quantification of all metabolites and guaranteed good accuracy of the results. No matrix effects were observed in the quantification of the analytes. The limit of detection for o- and m-cresol and 2- and 4-ethylphenol was 10 and 20μg/l urine and linearity ranged up to 3 and 12mg/L urine, respectively. The limit of detection for urinary phenol was 0.5mg/L with a linear range up to 200mg/L. The relative standard deviation of the within-series imprecision ranged between 3.0 and 7.2% at two spiked concentrations of 60 and 400μg/l and the relative recovery was between 84 and 104%, depending on the analyte. The method was successfully applied in proficiency testing for urinary o-cresol and phenol. This method was used for the analysis of urine samples of 17 non-smoking and 13 smoking persons from the general population without known exposure to solvents. Smokers showed a significantly higher excretion of o-cresol (median: 23 vs. 33μg/l), m-cresol (median: 43 vs. 129μg/l) as well as 4-ethylphenol (median: 25 vs. 124μg/l). Especially excretion of 4-ethylphenol was significantly correlated to smoking habits. The method seems to be suitable for biological monitoring of low-level solvent exposures and allows determination of background values in the general population.

Human biomonitoring of polychlorinated biphenyls (PCBs) in plasma of former underground miners in Germany – A case-control study

Polychlorinated biphenyls (PCBs) are very persistent organic pollutants of severe environmental concern due to their toxic properties. Former underground miners might have been exposed to this substance group due to the widespread use of PCBs in hydraulic oils from the late 1960s to the mid 1980s. We have conducted a blinded case-control study in order to evaluate the possibility of retrospective exposure assessment of PCBs using human biomonitoring in former underground miners decades after the last possible exposure. We have identified n = 34 male former underground miners and n = 136 age-matched male control persons from the database of patients of our occupational outpatient clinic aged between 47.9 and 83.7 years  at the time of sampling (June 2006-June 2016). These archived plasma samples have been blinded and analysed for 21 different PCB-congeners using a validated and quality controlled procedure using GC/MS (LOQ: 0.01 μg/L). Highly significant differences between cases and age-matched controls were only found for the PCB-congeners PCB 74 and PCB 114. The median (95th percentile) levels of PCB 74 in cases and controls were 0.126 μg/L plasma (0.899 μg/L plasma) vs. 0.058 μg/L plasma (0.368 μg/L plasma) and the 95th percentile levels for PCB 114 were 0.039 μg/L plasma vs. 0.017 μg/L plasma. Linear regression models revealed that this difference in plasma levels was unequivocally attributed to the underground mining activity. Thus, retrospective exposure assessment for underground miners by use of human biomonitoring seems feasible and further studies with a particular focus on this special group of workers should be performed.