Exploring a new method for the assessment of metal exposure by analysis of exhaled breath of welders.

PURPOSE: Air monitoring has been the accepted exposure assessment of toxic metals from, e.g., welding, but a method characterizing the actual dose delivered to the lungs would be preferable. Sampling of particles in exhaled breath can be used for the biomonitoring of both endogenous biomarkers and markers of exposure. We have explored a new method for the sampling of metals in exhaled breath from the small airways in a study on welders. METHODS: Our method for particle sampling, Particles in Exhaled Air (PExA®), is based on particle counting and inertial impaction. We applied it on 19 stainless steel welders before and after a workday. In parallel, air monitoring of chromium, manganese and nickel was performed as well as blood sampling after work. RESULTS: Despite substantial exposure to welding fumes, we were unable to show any significant change in the metal content of exhaled particles after, compared with before, exposure. However, the significance might be obscured by a substantial analytical background noise, due to metal background in the sampling media and possible contamination during sampling, as an increase in the median metal contents were indicated. CONCLUSIONS: If efforts to reduce background and contamination are successful, the PExA® method could be an important tool in the investigations of metals in exhaled breath, as the method collects particles from the small airways in contrast to other methods. In this paper, we discuss the discrepancy between our findings and results from studies, using the exhaled breath condensate (EBC) methodology.

Benzene Exposure and Biomarkers in Alveolar Air and Urine Among Deck Crews on Tankers Transporting Gasoline.

INTRODUCTION: Increased rates of leukaemia have been found among tanker crews. Occupational exposures to the leukomogen benzene during loading, unloading, and tank cleaning are possible causes. Studies on older types of tankers carrying gasoline with most handling being done manually have revealed important exposures to benzene. Our study explores benzene exposures on tankers with both automatic and manual systems. Correlations between benzene exposure and benzene in alveolar air (AlvBe), benzene in urine (UBe), and trans,trans-muconic acid (ttMA) in urine were investigated. METHODS: Forty-three male seafarers (22 deck crewmembers and 21 not on deck) on five Swedish different product and chemical tankers transporting 95- or 98-octane gasoline were investigated between 1995 and 1998. The tankers used closed systems for the loading and unloading of gasoline but stripping and tank cleaning were done manually. Benzene in respiratory air was measured using personal passive dosimeters during a 4-h work shift. Samples for biomarker analyses were collected pre- and post-shift. Smoking did occur and crewmembers did not use any respiratory protection during work. RESULTS: The average 4-h benzene exposure level for exposed was 0.45 mg m-3 and for non-exposed 0.02 mg m-3. Benzene exposure varied with type of work (range 0.02-143 mg m-3). AlvBe, UBe, and ttMA were significantly higher in post-shift samples among exposed and correlated with exposure level (r = 0.89, 0.74, and 0.57, respectively). Smoking did not change the level of significance among exposed. DISCUSSION: Benzene in alveolar air, unmetabolized benzene, and ttMA in urine are potential biomarkers for occupational benzene exposure. Biomarkers were detectable in non-exposed, suggesting benzene exposure even for other work categories on board tankers. Work on tankers carrying gasoline with more or less closed handling of the cargo may still lead to significant benzene exposure for deck crewmembers, and even exceed the Swedish Occupational Exposure Limit (OEL; 8-h time-weighted average [TWA]) of 1.5 mg m-3.

Two techniques to sample non-volatiles in breath-exemplified by methadone.

The particles in exhaled breath provide a promising matrix for the monitoring of pathological processes in the airways, and also allow exposure to exogenous compounds to be to assessed. The collection is easy to perform and is non-invasive. The aim of the present study is to assess if an exogenous compound-methadone-is distributed in the lining fluid of small airways, and to compare two methods for collecting methadone in particles in exhaled breath. Exhaled particles were collected from 13 subjects receiving methadone maintenance treatment. Two different sampling methods were applied: one based on electret filtration, potentially collecting exhaled particles of all sizes, and one based on impaction, collecting particles in the size range of 0.5-7 µm, known to reflect the respiratory tract lining fluid from the small airways. The collected samples were analyzed by liquid chromatography mass spectrometry, and the impact of different breathing patterns was also investigated. The potential contribution from the oral cavity was investigated by rinsing the mouth with a codeine solution, followed by codeine analysis of the collected exhaled particles by both sampling methods. The results showed that methadone was present in all samples using both methods, but when using the method based on impaction, the concentration of methadone in exhaled breath was less than 1% of the concentration collected by the method based on filtration. Optimizing the breathing pattern to retrieve particles from small airways did not increase the amount of exhaled methadone collected by the filtration method. The contamination from codeine present in the oral cavity was only detected in samples collected by the impaction method. We conclude that methadone is distributed in the respiratory tract lining fluid of small airways. The samples collected by the filtration method most likely contained a contribution from the upper airways/oral fluid in contrast to the impaction method.

Characterization of exhaled breath particles collected by an electret filter technique.

Aerosol particles that are present in exhaled breath carry nonvolatile components and have gained interest as a specimen for potential biomarkers. Nonvolatile compounds detected in exhaled breath include both endogenous and exogenous compounds. The aim of this study was to study particles collected with a new, simple and convenient filter technique. Samples of breath were collected from healthy volunteers from approximately 30 l of exhaled air. Particles were counted with an optical particle counter and two phosphatidylcholines were measured by liquid chromatography-tandem mass spectrometry. In addition, phosphatidylcholines and methadone was analysed in breath from patients in treatment with methadone and oral fluid was collected with the Quantisal device. The results demonstrated that the majority of particles are <1 µm in size and that the fraction of larger particle contributes most to the total mass. The phosphatidylcholine PC(16 : 0/16 : 0) dominated over PC(16 : 0/18 : 1) and represented a major constituent of the particles. The concentration of the PC(16 : 0/16 : 0) homolog was significantly correlated (p < 0.001) with total mass. From the low concentration of the two phosphatidylcholines and their relative abundance in oral fluid a major contribution from the oral cavity could be ruled out. The concentration of PC(16 : 0/16 : 0) in breath was positively correlated with age (p < 0.01). An attempt to use PC(16 : 0/16 : 0) as a sample size indicator for methadone was not successful, as the large intra-individual variability between samplings even increased after normalization. In conclusion, it was demonstrated that exhaled breath sampled with the filter device represents a specimen corresponding to surfactant. The possible use of PC(16 : 0/16 : 0) as a sample size indicator was supported and deserves further investigations. We propose that the direct and selective collection of the breath aerosol particles is a promising strategy for measurement of nonvolatiles in breath.

Supercritical fluid extraction (SFE) for determination of metalworking fluid aerosols.

A common methodology for analyzing metalworking fluid (MWF) aerosols in workplace air is based on gravimetry before and after organic solvent extraction of the MWFs from a suitable collection filter. Because MWFs have different chemical and physical properties, various mixtures of organic solvents have been used to extract the MWFs from their collection device. An alternative to organic solvents, used in the work presented in this article, is the use of a supercritical fluid. The efficiency of supercritical fluid extraction (SFE) was investigated by weighing conditioned filters before and after extraction of samples spiked with MWFs at different concentrations using the American Society for Testing and Materials method. For three common straight oil MWFs spiked on filters, supercritical carbon dioxide gave recoveries of 92-101% with a low standard deviation (0.2-1.9%). For semisynthetic MWFs, carbon dioxide had to be mixed with methanol to obtain recoveries above 80%. With the optimized method using 7% methanol in carbon dioxide, the 10 investigated MWFs could be extracted in 30 min with a recovery of 90-98%. The amount of MWFs spiked on the filters varied between 0.10-1.65 mg. In Sweden, the limit value for MWFs is 1 mg/m3. Thus, our spike level is in the range of 10-200% of the limit value if sampling for 8 hours with 2 L/min. The use of SFE methodology results in small volume extracts (3 mL) with concentrations at such high levels that analysis of chemical components in the MWF can be carried out without further volume reduction.

Exposure to volatile methacrylates in dental personnel.

Dental personnel are exposed to acrylates due to the acrylic resin-based composites and bonding agents used in fillings. It is well known that these compounds can cause contact allergy in dental personnel. However, in the 1990s, reports emerged on asthma also caused by methacrylates. The main volatile acrylates in dentistry are 2-hydroxyethyl methacrylate and methyl methacrylate. The aim of this study was to quantify the exposure to these acrylates in Swedish dental personnel. We studied the exposure to 2-hydroxyethyl methacrylate and methyl methacrylate in five randomly selected public dental clinics and at the Faculty of Odontology at Göteborg University. In total, 21 whole-day and 46 task-specific short-term (1-18 min) measurements were performed. The median 8-hour time-weighted averages were 2.5 microg/m3 (dentists) and 2.9 microg/m3 (dental nurses) for 2-hydroxyethyl methacrylate, and 0.8 microg/m3 (dentists) and 0.3 microg/m3 (dental nurses) for methyl methacrylate. The maximum short-term exposure levels were 79 microg/m3 for 2-hydroxyethyl methacrylate and 15 microg/m3 for methyl methacrylate, similar in dentists and dental nurses. The observed levels are much lower than in complete denture fabrication. We found only one previous study in dentistry and it showed similar results (though it reported short-term measurements only). Irritant effects would not be expected in healthy people at these levels. Nevertheless, occupational respiratory diseases due to methacrylates may occur in dental personnel, and improvements in the handling of these chemicals in dentistry are warranted. This includes better vials for the bonding agents and avoiding evaporation from discarded materials.

Optimization and validation of an ion chromatographic method for the simultaneous determination of sodium, ammonium and potassium in exhaled breath condensate.

An ion chromatographic method with conductivity detection for the simultaneous quantification of sodium, ammonium and potassium in exhaled breath condensate (EBC) was developed and validated. A factorial design was used to optimize the chromatographic conditions, which resulted in baseline separations of the cations within 6 min. The method requires no pre-treatment of EBC samples. The optimized method was used for the intra-individual screening of cations in EBC of 10 healthy volunteers. The LOQs were low (0.3, 0.1 and 0.2 microM for sodium, ammonium and potassium, respectively), compared with levels detected in healthy volunteers. The responses were linear with good precision, and samples could be stored for at least 10 weeks at refrigerating conditions.

Determination of hydrogen peroxide in exhaled breath condensate by flow injection analysis with fluorescence detection.

A method for the determination of hydrogen peroxide in exhaled breath condensate (EBC) by automated flow injection analysis (FIA) with fluorescence detection was developed and validated. In the enzymatic assay a fluorescent dimer of para-hydroxyphenyl acetic acid (HPAA) was formed by the redox coupling reaction between hydrogen peroxide and horseradish peroxidase (HRP). The calibration curve of hydrogen peroxide was linear over a range of 40-5000 nM. The coefficient of variation (CV) for within-day precision was 1-3%; for between-day precision, it was 2-5% over the validated range. The assay requires a small sample aliquot (150 microl) and no incubation time, and has an analytical runtime of < 2 min. It is therefore suitable for larger studies. The method was used to detect hydrogen peroxide in EBC of asthmatic patients and healthy volunteers. A statistically significant difference was found between patients with asthma (n = 19) and control subjects without asthma (n = 19), 780 nM versus 480 nM (P = 0.03).

A combination of micro-porous membrane liquid-gas extraction and solid-phase trapping for ultra trace determination of benzene in urine.

A method was developed for the determination of benzene in urine. The sample was pumped through the donor channel of a membrane extraction unit with a micro-porous membrane, separating the donor channel from an identical acceptor channel purged with nitrogen. The analyte reached the acceptor channel by diffusion through the membrane and was then swept by the carrier to a solid sorbent tube, where it was trapped. The analyte was subsequently thermally desorbed and analyzed by gas chromatography (GC) with mass selective detection (MS). After optimization, the recovery was close to quantitative, or 95%. Purging the membrane unit with pure water in between the samples eliminated any memory effects. The linearity was good in the concentration range examined (20-4000 ng l(-1)), with a correlation coefficient of 0.9996. The repeatability at 50 ng l(-1) and 400 ng l(-1) was 1.4% and 1.2%, respectively. The limit of detection was 12 ng l(-1) and the limit of quantification 35 ng l(-1). This enables assessment of benzene exposures of occupationally exposed subjects, of smokers and the majority of the general population. The developed method can be easily automated.

Comparison of sampling efficiency and storage stability on different sorbents for determination of solvents in occupational air.

This study describes the sampling efficiency and storage stability of compounds typically present in occupational atmospheres on the sorbents Anasorb CSC, Anasorb 747, and Chromosorb 106. The selection of compounds included in the study contained aliphatic and aromatic hydrocarbons, alcohols, esters, glycol ethers, ketones, and halogenated compounds, thus representing a wide range of chemical and physical properties. The different sorbent tubes were simultaneously exposed to the selected compounds as three different mixtures of solvent vapours in air, and storage both at room temperature and at -22 degrees C was investigated. The sorbent tubes were stored and analyzed at two different laboratories. The sampling efficiencies of all the investigated compounds were excellent on Anasorb CSC and Anasorb 747, while Chromosorb 106 did not give such good results for the most volatile compounds under study. The room temperature storage stability on Chromosorb 106, however, was good for all compounds, although formation of artefacts was observed during storage, a disadvantage that was substantially reduced by storage at -22 degrees C. The room temperature storage stability on Anasorb CSC was good for all compounds except some of the ketones. The room temperature storage stability of these ketones, especially cyclohexanone and 2-butanone, was much better on Anasorb 747, which still showed the same excellent storage stability for the remaining compounds. When stored in a freezer, the storage stability of all compounds, including the ketones, was very good on all sorbents. Among the sorbents under study, Anasorb 747 appears to be the most suitable all-round sorbent for monitoring volatile compounds in occupational air, with satisfactory capabilities regarding both sampling efficiency and storage stability.

Determination of ethane, pentane and isoprene in exhaled air using a multi-bed adsorbent and end-cut gas-solid chromatography.

A method for the determination of exhaled ethane, pentane and isoprene was developed and validated. The method was based on pre-concentration of the analytes on a multi-bed solid adsorbent tube containing Tenax TA, Carboxen 569 and Carboxen 1000, thermal desorption and gas chromatography (GC) with flame ionisation detection (FID). A pre-column in an end-cut GC system was used to avoid problems with water and strongly retained substances. The detection limits were 5, 2 and 6 pmol per sample for ethane, pentane and isoprene, respectively, using a sample volume of 500 ml. The linearity was good for all analytes with correlation coefficients exceeding 0.999. The repeatability for exhaled air samples was 7, 10 and 12% for ethane, pentane and isoprene, respectively. Analysis of a certified reference material of ethane and pentane did not differ significantly from the certified values. Ethane and pentane levels were stable up to six days of storage in sample tubes. Isoprene levels were not stable during storage in the sample tubes used here, but using Carbopack X instead of Carboxen 569, levels were stable up to two days. The levels of exhaled ethane, pentane and isoprene in healthy subjects (n = 4) were 8.1+/-5.8 pmol l(-1), 11+/-5.8 pmol l(-1) and 2.4+/-0.90 mnol l(-1), respectively. The method could, with minor modifications, be used to determine other low-molecular hydrocarbons in exhaled air as well.

Determination of malondialdehyde in breath condensate by high-performance liquid chromatography with fluorescence detection.

An automated and rapid method for quantifying malondialdehyde (MDA) in breath condensate was developed and validated. The method is based on derivatisation with thiobarbituric acid, HPLC separation and fluorescence detection and is optimised for determination of MDA in breath condensate. Sample collection is non-invasive and simple. The detection limit (4.1 nM) is low, precision is good and the analysis time is short. The response is linear in the concentration range of 0.020 to 1.0 microM. Samples could be stored for 1 month at -20 degrees C and for 3 months at -80 degrees C without losses. Using this method, there was no statistically significant difference between patients with asthma and patients without asthma. However, among females, subjects with asthma had higher MDA levels as compared to females without asthma (0.17 vs. 0.12 pmol/s, p=0.04). The use of the method when studying airway inflammation has to be further evaluated.