Glenn Wilson

Determination of parent and substituted polycyclic aromatic hydrocarbons in high-fat salmon using a modified QuEChERS extraction, dispersive SPE and GC-MS

A fast and easy modified QuEChERS (quick, easy, cheap, rugged and safe) extraction method has been developed and validated for determination of 33 parent and substituted polycyclic aromatic hydrocarbons (PAHs) in high-fat smoked salmon that greatly enhances analyte recovery compared to traditional QuEChERS procedures. Sample processing includes extraction of PAHs into a solution of ethyl acetate, acetone and isooctane followed by cleanup with dispersive SPE and analysis by GC-MS in SIM mode. Method performance was assessed in spike recovery experiments (500 μg/g wet weight) in three commercially available smoked salmon with 3-11% fat. Recoveries of some 2-, 3- and 5-ring PAHs were improved 50-200% over traditional methods, while average recovery across all PAHs was improved 67%. Method precision was good with replicate extractions typically yielding relative standard deviations <10%, and detection limits were in the low ng/g range. With this method, a single analyst could extract and clean up ≥60 samples for PAH analysis in an 8 h work day.

Ketone and quinone-substituted polycyclic aromatic hydrocarbons in mussel tissue, sediment, urban dust, and diesel particulate matrices.

Polycyclic aromatic hydrocarbons (PAHs) substituted with a ketone or quinone functionality (OPAHs) may be important environmental contaminants. The OPAHs from environmental samples have demonstrated toxicity and may be more harmful than PAHs. Knowledge gaps concerning the occurrence of OPAHs in the total environment arise from analytical difficulties, as well as limited standards and methodologies. An optimized method was developed to quantify five ketone and four quinone OPAHs from matrices ranging from biological tissue to diesel particulates. Five National Institute of Standards and Technology Standard Reference Materials (SRMs) were analyzed. This is the first report of OPAH quantitation in SRM 2977 (mussel tissue), SRM 1944 (New York/New Jersey, USA waterway sediment), SRM 1975 (diesel extract), and SRM 1650b (diesel particulate matter) and among the few to report concentrations from SRM 1649 (urban dust). Furthermore, this is one of the first reports of OPAHs in biological tissue. Σ₉OPAHs were 374 ± 59 mg/kg (mussel tissue), 5.4 ± 0.5 mg/kg (sediment), 16.9 ± 1.6 mg/kg (urban dust), 33.4 ± 0.4 mg/kg (diesel extract), and 150 ± 43 mg/kg (diesel particulate matter). In all SRMs, the levels of OPAHs were similar to or exceeded levels of PAHs. Of the OPAHs tested, the most frequently occurring in the environmental matrices were 9-fluorenone, 9,10-anthraquinone, benzofluorenone, and 7,12-benz[a]anthracenequinone.

Structurally distinct polycyclic aromatic hydrocarbons induce differential transcriptional responses in developing zebrafish.

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous in the environment as components of fossil fuels and by-products of combustion. These multi-ring chemicals differentially activate the aryl hydrocarbon receptor (AHR) in a structurally dependent manner, and induce toxicity via both AHR-dependent and -independent mechanisms. PAH exposure is known to induce developmental malformations in zebrafish embryos, and recent studies have shown cardiac toxicity induced by compounds with low AHR affinity. Unraveling the potentially diverse molecular mechanisms of PAH toxicity is essential for understanding the hazard posed by complex PAH mixtures present in the environment. We analyzed transcriptional responses to PAH exposure in zebrafish embryos exposed to benz(a)anthracene (BAA), dibenzothiophene (DBT) and pyrene (PYR) at concentrations that induced developmental malformations by 120 h post-fertilization (hpf). Whole genome microarray analysis of mRNA expression at 24 and 48 hpf identified genes that were differentially regulated over time and in response to the three PAH structures. PAH body burdens were analyzed at both time points using GC-MS, and demonstrated differences in PAH uptake into the embryos. This was important for discerning dose-related differences from those that represented unique molecular mechanisms. While BAA misregulated the least number of transcripts, it caused strong induction of cyp1a and other genes known to be downstream of the AHR, which were not induced by the other two PAHs. Analysis of functional roles of misregulated genes and their predicted regulatory transcription factors also distinguished the BAA response from regulatory networks disrupted by DBT and PYR exposure. These results indicate that systems approaches can be used to classify the toxicity of PAHs based on the networks perturbed following exposure, and may provide a path for unraveling the toxicity of complex PAH mixtures.

Toxicology of brominated fatty acids: metabolite concentration and heart and liver changes.

Rats were fed for 35 days diets containing 2% of either brominated corn oil (BCO), monoglyceride of dibromostearate (DBS), monoglyceride of tetrabromostearate (TBS) or a mixture of the two monoglycerides (BMG) which provided proportions of brominated acids comparable to that of the BCO. Hearts from all animals fed BCO were yellow colored and firm in texture. Myocardial cellular degeneration, mild to moderate edema and occasional small necrotic foci were observed. Hearts from animals fed DBS showed moderate edema and some slight necrosis. All diets produced an increase in lipid content of heart. Animals fed the experimental diets developed enlarged livers and showed elevated liver lipid content. The tetrabromostearate appeared to be the more active in producing these changes, in particular a severe intracellular fatty degeneration. Shorter-chain (C-16, C-14) metabolites of di- and tetrabromostearate were identified and the concentration of brominated fatty acids in heart, liver and adipose tissue determined and found to account for 80% of the bromine detected in these tissues by neutron activation analysis. TBS accumulated in liver while the highest concentration of DBS was observed in heart lipids. Although the concentrations of brominated acids in heart and liver lipids were comparable in rats fed BCO or BMG, BCO produced the more pronounced effects. This differential could be due to additional active components in BCO or to a variation in response associated with changes in the location of the fatty acid on the glycerol molecule.

Improvements in pollutant monitoring: optimizing silicone for co-deployment with polyethylene passive sampling devices.

Sequestering semi-polar compounds can be difficult with low-density polyethylene (LDPE), but those pollutants may be more efficiently absorbed using silicone. In this work, optimized methods for cleaning, infusing reference standards, and polymer extraction are reported along with field comparisons of several silicone materials for polycyclic aromatic hydrocarbons (PAHs) and pesticides. In a final field demonstration, the most optimal silicone material is coupled with LDPE in a large-scale study to examine PAHs in addition to oxygenated-PAHs (OPAHs) at a Superfund site. OPAHs exemplify a sensitive range of chemical properties to compare polymers (log Kow 0.2-5.3), and transformation products of commonly studied parent PAHs. On average, while polymer concentrations differed nearly 7-fold, water-calculated values were more similar (about 3.5-fold or less) for both PAHs (17) and OPAHs (7). Individual water concentrations of OPAHs differed dramatically between silicone and LDPE, highlighting the advantages of choosing appropriate polymers and optimized methods for pollutant monitoring.

Modified ion source triple quadrupole mass spectrometer gas chromatograph for polycyclic aromatic hydrocarbon analyses.

We describe modified gas chromatography electron-impact/triple-quadrupole mass spectrometry (GC-EI/MS/MS) utilizing a newly developed hydrogen-injected self-cleaning ion source and modified 9mm extractor lens. This instrument, with optimized parameters, achieves quantitative separation of 62 polycyclic aromatic hydrocarbons (PAHs). Existing methods historically limited rigorous identification and quantification to a small subset, such as the 16 PAHs the US EPA has defined as priority pollutants. Without the critical source and extractor lens modifications, the off-the-shelf GC-EI/MS/MS system was unsuitable for complex PAH analysis. Separations were enhanced by increased gas flow, a complex GC temperature profile incorporating multiple isothermal periods, specific ramp rates, and a PAH-optimized column. Typical determinations with our refined GC-EI/MS/MS have a large linear range of 1-10,000pgμl(-1) and detection limits of <2pgμl(-1). Included in the 62 PAHs, multiple-reaction-monitoring (MRM) mode enabled GC-EI/MS/MS identification and quantitation of several constituents of the MW 302 PAH isomers. Using calibration standards, values determined were within 5% of true values over many months. Standard curve r(2) values were typically >0.998, exceptional for compounds which are archetypally difficult. With this method benzo[a]fluorene, benzo[b]fluorene, benzo[c]fluorene were fully separated as was benzo[b]fluoranthene, benzo[k]fluoranthene, and benzo[j]fluoranthene. Chrysene and triphenylene, were sufficiently separated to allow accurate quantitation. Mean limits of detection (LODs) across all PAHs were 1.02±0.84pgμl(-1) with indeno[1,2,3-c,d] pyrene having the lowest LOD at 0.26pgμl(-1) and only two analytes above 2.0pgμl(-1); acenaphthalene (2.33pgμl(-1)) and dibenzo[a,e]pyrene (6.44pgμl(-1)).

Tissue fatty acid changes and tumor incidence in C3H mice ingesting cottonseed oil

The incidence of spontaneous mammary tumors in C3H mice at 35 wk was higher in mice fed rations containing cottonseed oil than in mice fed fats of comparable fatty acid composition. The time to 50% (T50) incidence was also shorter in the first group. The fatty acid composition of tissue lipids from mice fed the cottonseed oil showed changes indicating the presence of cyclopropene fatty acids-higher levels of saturates and lower oleate/stearate and palmitoleate/palmitate ratios. A possible association between the development of a spontaneous mammary tumor in the C3H mouse and the presence of cyclopropene fatty acids in the cottonseed oil is indicated.

Preparation and performance features of wristband samplers and considerations for chemical exposure assessment

Wristbands are increasingly used for assessing personal chemical exposures. Unlike some exposure assessment tools, guidelines for wristbands, such as preparation, applicable chemicals, and transport and storage logistics, are lacking. We tested the wristband’s capacity to capture and retain 148 chemicals including polychlorinated biphenyls (PCBs), pesticides, flame retardants, polycyclic aromatic hydrocarbons (PAHs), and volatile organic chemicals (VOCs). The chemicals span a wide range of physical–chemical properties, with log octanol–air partitioning coefficients from 2.1 to 13.7. All chemicals were quantitatively and precisely recovered from initial exposures, averaging 102% recovery with relative SD ≤21%. In simulated transport conditions at +30 °C, SVOCs were stable up to 1 month (average: 104%) and VOC levels were unchanged (average: 99%) for 7 days. During long-term storage at −20 °C up to 3 (VOCs) or 6 months (SVOCs), all chemical levels were stable from chemical degradation or diffusional losses, averaging 110%. Applying a paired wristband/active sampler study with human participants, the first estimates of wristband–air partitioning coefficients for PAHs are presented to aid in environmental air concentration estimates. Extrapolation of these stability results to other chemicals within the same physical–chemical parameters is expected to yield similar results. As we better define wristband characteristics, wristbands can be better integrated in exposure science and epidemiological studies.

Fatty acid composition of liver lipids in rats fed brominated fatty acids

Feeding rats diets containing brominated corn oil or di- or tetrabromostearate as the monoglyceride produced changes in fatty acid composition of liver lipids. Those changes associated with the feeding of brominated corn oil or tetrabromosterates could be explained by the accumulation of triglyceride, and the changes associated with the feeding of dibromostearate could result from the proliferation of a membrane system. A unique response to the feeding of diets containing brominated corn oil is an increase in the level of γ-linolenic acid.