Higher PBDE serum concentrations may be associated with feline hyperthyroidism in Swedish cats.

Serum from 82 individual cats was analyzed for decabromobiphenyl (BB-209), polybrominated diphenyl ethers (PBDEs), hydroxylated PBDEs (OH-PBDEs), and 2,4,6-TBP in order to study differences in body burden between healthy and sick cats diagnosed with Feline Hyperthyroidism (FH). Within the study group, 60 of these cats had a euthyroid (n = 23) or hyperthyroid (n = 37) status, all of which were used in the comparison. This study shows that hyperthyroid compared to euthyroid cats have higher serum concentrations for some of the investigated PBDEs (BDE-99, BDE-153, and BDE-183) and CB-153 on a fat weight basis. Further, it is intriguing, and beyond explanation, why the flame retardant BB-209 (discontinued in 2000) is present in all of the cat serum samples in concentrations similar to BDE-209. Median BDE-47/-99 ratios are 0.47 and 0.32 for healthy and euthyroid cats, respectively, which differs significantly from Swedes, where the ratio is 3.5. Another important finding is the occurrence of very low levels or the absence of hydroxylated PBDE metabolites in the cats. In addition, the major OH-PBDE, 6-OH-BDE47, is likely of natural origin, probably ingested via cat food. The statistics indicate an association between elevated PBDE concentrations in the cats and FH.

Recovery discrepancies of OH-PBDEs and polybromophenols in human plasma and cat serum versus herring and long-tailed duck plasma.

Hydroxylated polybrominated diphenyl ethers (OH-PBDEs) have been identified as metabolites of polybrominated diphenyl ethers (PBDEs) and/or as natural products. The OH-PBDEs and polybromophenols have come into focus over the last decade due to their abundance in biota and their potential adverse health effects. The present recovery study aims to validate a commonly used method (published by Hovander et al. 2000) for OH-PBDE analysis in human plasma. Further, the authors intended to determine the method's applicability to serum/plasma matrices from other species than humans. The investigated matrices were human plasma, cat serum, herring- and long-tailed duck plasma. The recovery study included nine OH-PBDEs, four polybromophenols and three methoxylated PBDEs (MeO-PBDEs). Five replicates of each matrix were spiked with these compounds at two dose levels; a low dose (0.5 ng) and a high dose (5 ng) and were cleaned up according to the Hovander method. The recovery of OH-PBDEs and polybromophenols in human plasma and cat serum were high and reproducible at both dose levels whereas the recovery for herring and long-tailed duck plasma were low and insufficient with great variability amongst OH-PBDE congeners at both dose levels. Our data show that the method can be fully applied to matrices like human plasma and cat serum but not for herring and long-tailed duck plasma without further method development. Hence care needs to be taken when applying the method onto other blood matrices without validation since the present study have demonstrated that the recoveries may differ amongst OH-PBDE congeners and specie.

Decabromobiphenyl, polybrominated diphenyl ethers, and brominated phenolic compounds in serum of cats diagnosed with the endocrine disease feline hyperthyroidism.

The incidence of cats being diagnosed with feline hyperthyroidism (FH) has increased greatly since it was first described in 1979. The cause of FH has not been established. Hypothetically, there is a link between increasing FH and exposure to brominated flame retardants. Much greater polybrominated diphenyl ethers (PBDE) concentrations have been reported in cat serum compared with human serum, likely due to cat licking behaviour. This study aimed to extend the present identification of brominated compounds in cat serum, with a focus on hydroxylated metabolites of PBDE, to improve the understanding of feline metabolism of PBDEs. A pooled serum sample from 30 Swedish pet cats with FH was analysed, and brominated species were identified. The results showed exposure to the discontinued flame retardant decabromobiphenyl (BB-209) and technical penta- and octa-BDEs. Altogether 12 PBDE congeners were identified along with 2'-MeO-BDE68. Furthermore, 2,4-dibromophenol, 2,4,6-, 2,4,5- and 2,3,4-tribromophenol plus 2'-OH-BDE68, 6-OH-BDE47, 5-OH-BDE47, 4'-OH-BDE49 were identified. 2,4,6-tribromophenol and 6-OH-BDE47 were the most prominent species in cat serum. Considering that these are natural products, it can be concluded that metabolism of PBDEs to OH-PBDEs is not a major route of PBDE elimination in cats. It is notable that BB-209, 6-OH-BDE47, and 2,4,6-tribromophenol all suggested that endocrine-disrupting chemicals were present in high concentrations in cat serum.

Optimization of digestion parameters for protein quantification.

We present a rapid and efficient in-solution enzymatic digestion protocol suitable for mass spectrometry-based absolute protein quantification techniques. The digestion method employs RapiGest SF (an acid-labile surfactant), an excess amount of modified trypsin (enzyme-to-substrate ratio of 2.5:1), and an incubation time of 2 h. No reduction/alkylation reagents are used. Digestion parameters were varied systematically to monitor their effect on rate and completeness of digestion. To demonstrate the general applicability of the method, the optimization was done using a viral hemagglutinin (HA) as a model protein and then applied to ricin, a potent protein toxin extracted from the castor bean (Ricinus communis). The parameters that were optimized included incubation time, concentration of RapiGest SF, enzyme-to-substrate ratio, and incubation temperature. The optimization was done by comparing the yields from two protein-specific peptides originating from two different sites of the HA protein. The analysis was performed by liquid chromatography-tandem mass spectrometry in multiple reaction monitoring mode using isotopically labeled peptide standards for quantification.

Identification of human urinary metabolites of acetochlor in exposed herbicide applicators by high-performance liquid chromatography-tandem mass spectrometry.

Acetochlor is a preemergent chloroacetanilide herbicide used to control annual grasses and small-seeded broadleaf weeds. It is the second most abundantly applied herbicide on corn crops in the United States; however, human metabolites associated with known exposure to acetochlor have not been positively identified and confirmed. We positively identified acetochlor mercapturate (ACM) as a metabolite of acetochlor in urine samples collected during a 24-h period from custom (commercial) applicators who had applied acetochlor on either the day of or the day before urine collection. Concentrations in applicator urine samples ranged from 0.5 to 449 microg/l (0.3-121 microg/g creatinine). We found that ACM accounted for as much as 42% of the total acetochlor-derived metabolites; however, as the exposure level decreased (based on total acetochlor metabolite level), ACM became a less abundant metabolite of acetochlor (<17%). Unmetabolized acetochlor was also measured in the urine samples analyzed. At high exposures (classified as >100 microg/l), acetochlor accounted for about 0.8% of the total excreted acetochlor metabolites (approximately 2% of the ACM concentrations). At lower exposures (classified as ACM<10 microg/l), ACM and acetochlor concentrations were similar. Additionally, we tentatively identified another acetochlor metabolite that appeared to be important at low levels of exposure.

Quantification of six herbicide metabolites in human urine.

We developed a sensitive, selective and precise method for measuring herbicide metabolites in human urine. Our method uses automated liquid delivery of internal standards and acetate buffer and a mixed polarity polymeric phase solid phase extraction of a 2 mL urine sample. The concentrated eluate is analyzed using high-performance liquid chromatography-tandem mass spectrometry. Isotope dilution calibration is used for quantification of all analytes. The limits of detection of our method range from 0.036 to 0.075 ng/mL. The within- and between-day variation in pooled quality control samples range from 2.5 to 9.0% and from 3.2 to 16%, respectively, for all analytes at concentrations ranging from 0.6 to 12 ng/mL. Precision was similar with samples fortified with 0.1 and 0.25 ng/mL that were analyzed in each run. We validated our selective method against a less selective method used previously in our laboratory by analyzing human specimens using both methods. The methods produced results that were in agreement, with no significant bias observed.

Hydrolysis of environmental contaminants as an experimental tool for indication of their persistency.

To predict the persistency of a chemical in the environment, the chemical's physical-chemical properties and its reactivity in the environment need to be known or at least estimated. The partitioning of a chemical can be described on the basis of its water solubility, its octanol/water partitioning coefficient, and its vapor pressure. The mechanisms by which a chemical can be transformed may be categorized as being hydrolysis, oxidation, reduction, and photolysis. This study establishes a method for estimating the relative susceptibility of some potential environmental pollutants to undergo hydrolysis reactions. The method used the second-order rate constant for the reaction with sodium methoxide in methanol/N,N-dimethylformamide (DMF) as an indicator of relative susceptibility toward hydrolysis. The decabromodiphenyl ether is rapidly hydrolyzed, that is, undergoes nucleophilic aromatic substitution, while the rate of reaction of less brominated diphenyl ethers decreased by roughly a factor of 10 for each decrease in the level of bromination. Hexachlorobenzene was found to have a similar rate to a nonabromodiphenyl ether. 2,2-Bis(4-chlorophenyl)-1,1,1-trichloroethane (DDT) was transformed to 2,2-bis(4-chlorophenyl)-1,1-dichloroethene (DDE) immediately under these conditions, while DDE showed no apparent reaction. The results show that chemicals that can undergo elimination reactions are rapidly transformed, as are perhalogenated chemicals that can undergo substitution reactions. These chemicals are not likely to persist in the environment, while those that did not show any observable reactivity under similar hydrolytic conditions may persist for a very long time.