Xiaoliu Zhou

Automated on-line column-switching high performance liquid chromatography isotope dilution tandem mass spectrometry method for the quantification of bisphenol A, bisphenol F, bisphenol S, and 11 other phenols in urine.

Human exposure to bisphenol A (BPA) is widespread. However, in recent years, bisphenol analogs such as bisphenol S (BPS) and bisphenol F (BPF) are replacing BPA in the production of some consumer products. Because human exposure to these alternative bisphenols may occur, biomonitoring of these bisphenol analogs is warranted. In the present study, we developed and validated a sensitive and selective method that uses on-line solid phase extraction coupled to high performance liquid chromatography-isotope dilution tandem mass spectrometry with peak focusing to measure BPA, BPF, BPS, and 11 other environmental phenols in urine. The method required a small amount of sample (100μL) and minimal sample pretreatment. The limits of detection were 0.03ng/mL (BPS), 0.06ng/mL (BPF), 0.10ng/mL (BPA), and ranged from 0.1ng/mL to 1.0ng/mL for the other 11 phenols. In 100 urine samples collected in 2009-2012 from a convenience group of anonymous adults in the United States, of the three bisphenols, we detected BPA at the highest frequency and median concentrations (95%, 0.72ng/mL), followed by BPS (78%, 0.13ng/mL) and BPF (55%, 0.08ng/mL). This sensitive, rugged, and labor and cost-effective method could be used for the analysis of large number of samples for epidemiologic studies.

Canned Soup Consumption and Urinary Bisphenol A: A Randomized Crossover Trial

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Potential External Contamination with Bisphenol A and Other Ubiquitous Organic Environmental Chemicals during Biomonitoring Analysis: An Elusive Laboratory Challenge

Background: Biomonitoring studies are conducted to assess internal dose (i.e., body burden) to environmental chemicals. However, because of the ubiquitous presence in the environment of some of these chemicals, such as bisphenol A (BPA), external contamination during handling and analysis of the biospecimens collected for biomonitoring evaluations could compromise the reported concentrations of such chemicals. Objectives: We examined the contamination with the target analytes during analysis of biological specimens in biomonitoring laboratories equipped with state-of-the-art analytical instrumentation. Discussions: We present several case studies using the quantitative determination of BPA and other organic chemicals (i.e., benzophenone-3, triclosan, parabens) in human urine, milk, and serum to identify potential contamination sources when the biomarkers measured are ubiquitous environmental contaminants. Conclusions: Contamination with target analytes during biomonitoring analysis could result from solvents and reagents, the experimental apparatus used, the laboratory environment, and/or even the analyst. For biomonotoring data to be valid—even when obtained from high-quality analytical methods and good laboratory practices—the following practices must be followed to identify and track unintended contamination with the target analytes during analysis of the biological specimens: strict quality control measures including use of laboratory blanks; replicate analyses; engineering controls (e.g., clean rooms, biosafety cabinets) as needed; and homogeneous matrix-based quality control materials within the expected concentration ranges of the study samples.

Urinary Concentrations of Bisphenol A and Three Other Bisphenols in Convenience Samples of U.S. Adults during 2000–2014

Because of regulatory actions and public concerns, the use of bisphenol A (BPA) may decrease, while the use of BPA alternatives may increase. Although BPA alternatives are considered safer than BPA, their effects on health are still largely unknown. For risk assessment, understanding exposure to these chemicals is necessary. We measured the urinary concentrations of BPA and three bisphenol analogs, bisphenol S (BPS), bisphenol F (BPF), and bisphenol AF (BPAF), in 616 archived samples collected from convenience samplings of U.S. adults at eight time points between 2000 and 2014. We detected BPA at the highest frequency and geometric mean (GM) concentrations (74-99%, 0.36-2.07 μg/L), followed by BPF (42-88%, 0.15-0.54 μg/L) and BPS (19-74%, < 0.1-0.25 μg/L); BPAF was rarely detected (<3% of all samples). Although concentrations of BPF were generally lower than for other bisphenols, the 95th percentile concentration of BPF was often comparable or higher than that of BPA. We did not observe obvious exposure trends for BPF. However, the significant changes in GM concentrations of BPA and BPS suggest that exposures may be declining (BPA) or on the rise (BPS). Nationally representative data will be useful to confirm these findings and to allow monitoring future exposure trends to BPA and some of its bisphenol alternatives.

Concentrations of bisphenol A and seven other phenols in pooled sera from 3-11 year old children: 2001-2002 National Health and Nutrition Examination Survey.

Concerns exist regarding childrens exposure to bisphenol A (BPA) and other phenols because of the higher sensitivity, compared to adults, of childrens developing organs to endocrine disruptors. Several studies reported the urinary concentrations of these phenols in children, but data on levels of these compounds in childrens serum are limited. We present here the total (free plus conjugated) and free concentrations of BPA and seven other phenols in 24 pooled serum samples prepared from individual specimens collected from 936 children 3-11 years old who participated in the 2001-2002 National Health and Nutrition Examination Survey. We detected benzophenone-3, triclosan, 2,4-dichlorophenol, 2,5- dichlorophenol, and three parabens in at least 60% of the pools suggesting childrens exposure to these compounds or their precursors. Conjugated phenols were the major species. However, although many previous studies have shown widespread detection of BPA in childrens urine, we only detected total or free BPA in 3 and 2 pooled serum samples, respectively, at concentrations of 0.1-0.2 μg/L. The nonpersistent nature of BPA and the phenols examined and the likely episodic nature of the exposures to these compounds (or their precursors) suggest that for general population biomonitoring of these nonpersistent phenols, urine, not serum or plasma, is the preferred matrix.

In-vitro oxidation of bisphenol A: Is bisphenol A catechol a suitable biomarker for human exposure to bisphenol A?

The extensive use of bisphenol A (BPA) in the manufacture of consumer products results in widespread human exposure to the chemical. In the body, BPA undergoes first-pass metabolism to form BPA glucuronide, considered to be a major BPA byproduct. Concentrations of total (free plus conjugated) urinary species of BPA are used to assess human exposure to BPA. However, because BPA can be present in numerous consumer and household products, potential contamination with parent BPA during collection and handling may pose a challenge when measuring BPA in such biological samples as blood or urine. In this study we investigated the in-vitro phase I metabolism of BPA in rat and human liver microsomes by using on-line solid-phase extraction–high-performance liquid chromatography–tandem mass spectrometry to identify phase I metabolites (e.g., BPA oxidation products) that could be used as potential alternative biomarkers of BPA exposure. We unambiguously identified 5-hydroxy BPA (BPA catechol) as an in-vitro oxidative metabolite of BPA, but human microsomes oxidized only about 10% of BPA to BPA catechol. We evaluated the usefulness of BPA catechol as a potential biomarker of human exposure to BPA by measuring total concentrations of BPA catechol and BPA in 20 urine samples. We detected BPA catechol at much lower concentrations and frequency than those of BPA. Furthermore, we found that free BPA catechol was rather unstable in urine, which highlights the importance of sampling techniques to adequate interpretation of biomonitoring data. Together, these findings suggest that BPA catechol may not be a suitable biomarker of environmental exposure to BPA, but could be used to confirm BPA exposure in special populations or in situations when urine specimens were potentially contaminated with BPA.

Urinary Concentrations of 2,4-Dichlorophenol and 2,5-Dichlorophenol in the U.S. Population (National Health and Nutrition Examination Survey, 2003–2010): Trends and Predictors

Background: 2,4-Dichlorophenol (2,4-DCP), 2,5-dichlorophenol (2,5-DCP), and their precursors are widely used in industry and in consumer products. Urinary concentrations of these dichlorophenols (DCPs) have been measured as part of four National Health and Nutrition Examination Survey (NHANES) cycles in order to assess the exposure to these compounds or their precursors among the general U.S. population. Objectives: We identified predictors and evaluated trends in DCP concentrations according to race/ethnicity, age, sex, family income, and housing type. Methods: We used analysis of covariance to examine associations of various demographic parameters and survey cycle with urinary concentrations of DCPs during NHANES 2003–2010. We also conducted weighted logistic regressions to estimate associations of DCP concentrations above the 95th percentile with housing type, race/ethnicity, and income. Results: We detected DCPs in at least 81% of participants. Geometric mean (GM) urinary concentrations were higher for 2,5-DCP (6.1–12.9 μg/L) than 2,4-DCP (0.8–1.0 μg/L) throughout 2003–2010. Adjusted GM concentrations of the DCPs among children (6–11 years of age) and adults > 60 years of age were higher than among adolescents and other adults. Adjusted GM concentrations among non-Hispanic whites were lower than among non-Hispanic blacks and Mexican Americans, although differences according to race/ethnicity were less pronounced among participants in high-income households. Among non-Hispanic blacks and Mexican Americans, adjusted GM concentrations were lowest among high-income participants relative to other income groups, with a monotonic decrease with increasing income among Mexican Americans. Type of housing and race/ethnicity were significant predictors of DCP urinary concentrations above the 95th percentile. Furthermore, urinary DCP concentrations have showed a downward trend since 2003. Conclusions: Exposure to DCPs and their precursors was prevalent in the general U.S. population in 2003–2010. We identified age and race/ethnicity, family income, and housing type as predictors of exposure to these compounds. Citation: Ye X, Wong LY, Zhou X, Calafat AM. 2014. Urinary concentrations of 2,4-dichlorophenol and 2,5-dichlorophenol in the U.S. population (National Health and Nutrition Examination Survey, 2003–2010): trends and predictors. Environ Health Perspect 122:351–355; http://dx.doi.org/10.1289/ehp.1306816

Does the composition of urine change when collected from disposable diapers and other absorbent materials

The free and conjugated urinary species of non-persistent environmental chemicals or their breakdown products are valid human exposure biomarkers. For convenience, disposable diapers and other absorbent materials are widely used to collect urine specimens from infants and young toddlers. However, the extent to which the different urinary species of the target analytes and other components are recovered after the urine is extracted from these absorbent materials is unknown. In this proof-of-concept study, we investigated the extraction recovery from disposable diapers, cotton pads, and gauzes of the free versus glucuronidated urinary species of three example chemicals: bisphenol A, triclosan, and 4-methylumbelliferone. Although the glucuronides were almost fully recovered, the free species were not. Our results suggest that, in addition to other sampling considerations, the binding affinity and extraction recovery of the target biomarkers to the material used to collect the urine should be considered. Alternative collection approaches that do not require such an extraction (e.g., urine bags routinely used in hospitals) may be worth exploring. Despite its shortcomings, having urinary concentrations for biomonitoring considerably strengthens the exposure assessment, particularly for infants and young toddlers, and the benefits of including biomonitoring data outweigh their potential limitations.

Impact of enzymatic hydrolysis on the quantification of total urinary concentrations of chemical biomarkers

Human exposure to consumer and personal care products chemicals such as phenols, including parabens and other antimicrobial agents, can be assessed through biomonitoring by quantifying urinary concentrations of the parent chemical or its metabolites, often after hydrolysis of phase II conjugates. Developing suitable analytical methods for the concurrent quantification of multiple exposure biomarkers is challenging because optimal conditions for the hydrolysis of such conjugates (e.g., O-glucuronides, N-glucuronides, sulfates) may differ depending on the biomarker. We evaluated the effectiveness of seven commercial hydrolytic enzymes to simultaneously hydrolyze N-glucuronides (using the antibacterial triclocarban as example compound) and other conjugates (using select phenols and parabens as examples) by using on-line solid phase extraction-high performance liquid chromatography-isotope dilution-tandem mass spectrometry. Incubation (30 min, 55 °C) with a genetically engineered β-glucuronidase (IMCS, ≥15 units/μL urine) hydrolyzed N-glucuronide triclocarban, but did not fully hydrolyze the conjugates of phenols and parabens. By contrast, incubation (4 h, 37 °C) with solid β-glucuronidase (Helix pomatia, Type H-1, ≥30 units/μL urine) or liquid β-glucuronidase/arylsulfatase (Helix pomatia, 30 units/μL urine [i.e., 30 μL/100 μL urine]) in the presence of 100 μL methanol for 100 μL urine completely hydrolyzed N-glucuronide triclocarban and the conjugates of several phenols and parabens, without cleaving the ester bond of the parabens to form p-hydroxybenzoic acid. These results highlight the relevance of method validation procedures that include optimizing the hydrolysis of phase II urinary conjugates (e.g., enzyme type and amount used, reaction time, temperature) to quantify accurately and concurrently multiple exposure biomarkers for biomonitoring purposes.