Association of perfluorinated chemical exposure in utero with maternal and infant thyroid hormone levels in the Sapporo cohort of Hokkaido Study on the Environment and Children's Health.

OBJECTIVES: Perfluorooctane sulfonate (PFOS) and perfluorooctanoate (PFOA) have been widely used as industrial products, and are persistent organic pollutants due to their chemical stability. Previous studies suggested that PFOS and PFOA might disrupt thyroid hormone (TH) status. Although TH plays an important role in fetal growth during pregnancy, little attention has been paid to the relationships between maternal exposure to perfluorocarbons and TH statuses of mothers and fetuses. We investigated the effects of low levels of environmental PFOS and PFOA on thyroid function of mothers and infants. METHODS: Of the eligible subjects in a prospective cohort, 392 mother-infant pairs were selected. Concentration of maternal serum PFOS and PFOA was measured in samples taken during the second and third trimesters or within 1 week of delivery. Blood samples for measuring thyroid stimulating hormone (TSH) and free thyroxine (FT4) levels were obtained from mothers at early gestational stage (median 11.1 weeks), and from infants between 4 and 7 days of age, respectively. RESULTS: Median concentrations of PFOS and PFOA were 5.2 [95 % confidence interval (CI) 1.6-12.3] and 1.2 (95 % CI limitation of detection-3.4) ng/mL, respectively. Maternal PFOS levels were inversely correlated with maternal serum TSH and positively associated with infant serum TSH, whereas maternal PFOA showed no significant relationship with TSH or FT4 among mothers and infants. CONCLUSIONS: These findings suggest that PFOS may independently affect the secretion and balances of maternal and infant TSH even at low levels of environmental exposure.

Molecular orbital imaging of the acetone S2 excited state using time-resolved (e, 2e) electron momentum spectroscopy.

We report a time-resolved (e, 2e) experiment on the deuterated acetone molecule in the S2 Rydberg state with a lifetime of 13.5 ps. The acetone S2 state was prepared by a 195 nm pump laser and probed with electron momentum spectroscopy using a 1.2 keV incident electron beam of 1 ps temporal width. In spite of the low data statistics as well as of the limited time resolution (±35 ps) due to velocity mismatch, the experimental results clearly demonstrate that electron momentum spectroscopy measurements of short-lived transient species are feasible, opening the door to time-resolved orbital imaging in momentum space.

Development of a closed drug preparation method for oral anticancer drugs.

The objective of this article is to reduce the preparation time for oral anticancer drugs, reduce the exposure to drug preparations, and develop drug preparation equipment without external drug leaks in a closed state. In the newly developed closed oral drug preparation device, a 10 mL disposable syringe that was replaced with one projection for crushing tablets and a no-processing 30 mL disposable syringe were connected to a three-way stopcock. Using this instrument, Endoxan(®) tablets (principal components: cyclophosphamide) were crushed and suspended in water in a closed state. The drug was prepared to suspension and flowed out via a feeding tube by switching the handle of the three-way stopcock. To assess human exposure to cyclophosphamide, a high-performance volatile organic compound-solvent desorption passive sampler was attached to the preparer's mouth to collect air drifting in the vicinity, and cyclophosphamide levels were subsequently measured by liquid chromatography with tandem mass spectrometry. Using the developed drug preparation equipment, Endoxan(®) tablets were suspended in a closed state. According to liquid chromatography with tandem mass spectrometry analysis, the exposure of the preparer to cyclophosphamide was greatly reduced when using the developed device; cyclophosphamide was detected in only two of the five samples, though only at trace levels. The closed oral drug preparation device may permit the preparation and administration of toxic drugs to patients while greatly reducing the risk of occupational exposure among health-care workers and caregivers.

Determination of exposure of dispensary drug preparers to cyclophosphamide by passive sampling and liquid chromatography with tandem mass spectrometry.

OBJECTIVES: To determine cyclophosphamide exposure to preparers during tablet crushing and subsequent handling by analyzing indoor air collected using a high-performance volatile organic compounds-solvent desorption (VOC-SD) passive air sampler. METHODS: The passive sampler was taped to the mask over the mouth of the preparer and indoor air was collected during crushing and preparation of cyclophosphamide tablets (Endoxan®). After collection, the carbon molecular sieve adsorbent of the passive sampler was placed in a centrifuge tube, and 1 mL of carbon disulfide was used to elute cyclophosphamide from the adsorbent. Liquid-liquid extraction with 1 mL of water was performed, and the aqueous phase was used as the test solution. Cyclophosphamide concentration was determined by liquid chromatography with ultraviolet and tandem mass spectrometry detection. RESULTS: Cyclophosphamide concentration was detected in the range of 7.6-157.7 ng/sampler. Our results showed that low-level exposure occurred near the mouth of the preparer, which could present risks for long-term exposure, especially if combined with multiple toxic drug exposures. CONCLUSION: The anticancer drug monitoring methodology described here is a simple exposure assessment that can be used to ensure the safety of hospital pharmacy tablet preparers. Furthermore, since the anticancer drug exposure risk is very high for preparers, preparation should be in hood or with face mask.

Prenatal exposure to perfluorinated chemicals and relationship with allergies and infectious diseases in infants.

BACKGROUND: Recent studies have shown effects of prenatal exposure to perfluorooctane sulfonate (PFOS) and perfluorooctanoate (PFOA) on infants in the general environmental levels. Laboratory animal studies have shown that exposure to PFOS and PFOA is associated with immunotoxic effects. OBJECTIVES: To investigate the relationship between maternal PFOS and PFOA levels and infant allergies and infectious diseases during the first 18 months of life. Cord blood immunoglobulin (Ig) E levels were also evaluated. METHODS: We conducted a prospective cohort study of pregnant women from 2002 to 2005 in Sapporo, Japan. Maternal PFOS and PFOA levels were measured in relation to cord blood IgE concentrations (n=231) and infant allergies and infectious diseases (n=343). Characteristics of mothers and their infants were obtained from self-administered questionnaires and medical records. Development of infant allergies and infectious diseases was determined from self-administered questionnaires at 18 months of age. Concentrations of PFOS and PFOA in maternal serum and concentrations of IgE in umbilical cord serum at birth were measured. RESULTS: Cord blood IgE levels decreased significantly with high maternal PFOA concentration among female infants. However, there were no significant associations among maternal PFOS and PFOA levels and food allergy, eczema, wheezing, or otitis media in the 18 month-old infants (adjusted for confounders). CONCLUSIONS: Although cord blood IgE level decreased significantly with high maternal PFOA levels among female infants, no relationship was found between maternal PFOS and PFOA levels and infant allergies and infectious diseases at age in 18 months.

Comparison of fluorescence reagents for simultaneous determination of hydroxylated phenylalanine and nitrated tyrosine by high-performance liquid chromatography with fluorescence detection.

Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are well-known and important contributors to oxidative and nitrosative stress in several diseases. Hydroxylated phenylalanine and nitrated tyrosine products appear to be particularly susceptible targets of oxidative and nitrosative stress. We compared fluorescence reagents for their potential use in the analysis of hydroxylated phenylalanine and nitrated tyrosine products with a high-sensitivity and high-specificity HPLC-UV-FL technique. The analytes were extracted from serum via solid-phase extraction on Waters Oasis MCX cartridges. Chromatographic separation was achieved on an ODS column (Capcell Pak MG II; 150 × 2.0 mm) using a gradient mobile phase consisting of 20 mm sodium phosphate buffer (adjusted to pH 3.0) and acetonitrile. The method quantification limit for 4-nitrophenylalanine, m-tyrosine, and 3-nitrotyrosine was 0.1 µm. The relative standard deviation of the precision and accuracy was acceptable at the spiked concentration of 0.1 µm for 4-nitrophenylalanine, m-tyrosine and 3-nitrotyrosine. The method could be used for the in vitro analysis of serum samples.

Characterization of nitrated phenolic compounds for their anti-oxidant, pro-oxidant, and nitration activities.

Coffee is one of the most widely consumed beverages worldwide. Evidence of the health benefits and the important contribution of coffee brew to the intake of anti-oxidants in the diet has increased coffee consumption. Chlorogenic acid (ChA) and caffeic acid (CaA) are the major phenolic compounds in coffee. However, phenolic compounds, which are generally effective anti-oxidants, can become pro-oxidants in the presence of Cu(2+) to induce DNA damage under certain conditions. On the other hand, sodium nitrite (NaNO(2)) is widely used as a food additive to preserve and tinge color on cured meat and fish. It is possible that phenolic compounds react with NaNO(2) under acidic conditions, such as gastric juice. In this study, we identified compounds produced by the reaction between ChA or CaA in coffee and NaNO(2) in artificial gastric juice. The identified phenolic compounds and nitrated phenolic compounds were assessed for their anti-oxidant, pro-oxidant, and nitration activities by performing an in vitro assay. The nitrated phenolic compounds seemed to show increased anti-oxidant activity and decreased pro-oxidant activity. However, one nitrated CaA compound that has a furoxan ring showed the ability to release NO(2)(-) in the neutral condition.

Identification and detection method for genetically modified papaya resistant to papaya ringspot virus YK strain.

Unauthorized genetically modified (GM) papaya (Carica papaya LINNAEUS) was detected in a commercially processed product, which included papaya as a major ingredient, in Japan. We identified the transgenic vector construct generated based on resistance to infection with the papaya ringspot virus (PRSV) YK strain. A specific detection method to qualitatively monitor papaya products for contamination with the GM papaya was developed using the real-time polymerase chain reaction.

Epigenetic assessment of environmental chemicals detected in maternal peripheral and cord blood samples.

Epigenetic alteration is an emerging paradigm underlying the long-term effects of chemicals on gene functions. Various chemicals, including organophosphate insecticides and heavy metals, have been detected in the human fetal environment. Epigenetics by DNA methylation and histone modifications, through dynamic chromatin remodeling, is a mechanism for genome stability and gene functions. To investigate whether such environmental chemicals may cause epigenetic alterations, we studied the effects of selected chemicals on morphological changes in heterochromatin and DNA methylation status in mouse ES cells (ESCs). Twenty-five chemicals, including organophosphate insecticides, heavy metals and their metabolites, were assessed for their effect on the epigenetic status of mouse ESCs by monitoring heterochromatin stained with 4¢,6-diamino-2-phenylindole (DAPI). The cells were surveyed after 48 or 96 h of exposure to the chemicals at the serum concentrations of cord blood. The candidates for epigenetic mutagens were examined for the effect on DNA methylation at genic regions. Of the 25 chemicals, five chemicals (diethyl phosphate (DEP), mercury (Hg), cotinine, selenium (Se) and octachlorodipropyl ether (S-421)) caused alterations in nuclear staining, suggesting that they affected heterochromatin conditions. Hg and Se caused aberrant DNA methylation at gene loci. Furthermore, DEP at 0.1 ppb caused irreversible heterochromatin changes in ESCs, and DEP-, Hg- and S-421-exposed cells also exhibited impaired formation of the embryoid body (EB), which is an in vitro model for early embryos. We established a system for assessment of epigenetic mutagens. We identified environmental chemicals that could have effects on the human fetus epigenetic status.

Development and validation of a hydrophilic interaction chromatography-tandem mass spectrometry for quantification of nicotine and its metabolites in human maternal and cord sera.

A selective and sensitive HILIC-MS/MS method for the simultaneous quantification of nicotine and its metabolites in human maternal and cord sera was developed and validated. After solid-phase extraction, LC separation was achieved on a hydrophilic interaction chromatography. The validated method is capable of selective identification as well as accurate and sensitive quantification. Analyte recovery ranged from 86.2 to 107.7% and intra- and inter-day assay precision were less than 15% relative standard deviation. This sensitive HILIC-MS/MS method can be used to determine nicotine and its metabolic profile in smokers. This validated method is useful for the determination of nicotine and its metabolites in human serum in future studies of the effects of nicotine exposure on neonatal outcome.

Development of quality assessment method for optically active food flavor chemicals.

A quality assessment method for commercially available, optically active flavor compounds, namely, menthol, menthyl acetate, borneol, perillaldehyde, and 1,8-cineol, was developed. A gas chromatograph equipped with a flame ionization detector and a DB-5ms capillary column was used for the chemical purity test. A GC/MS with a beta-DEX cyclodextrin column was used for the optical purity test, by which the enantiomeric separation of each flavor compound was achieved. Enantiomeric excess was calculated as an expression of optical purity. Of the 25 standard samples subjected to the chemical purity test, six were found to have lower purity than the data provided by the manufacturers. When the same samples were subjected to the optical purity test, 11 were found to have lower purity than that indicated on the reagent labels. These results suggest that there is a need to conduct an optical purity test, in addition to a chemical purity test, for the quality assessment of flavor standards.

Determination of proline enantiomers in honey and royal jelly by LC-UV.

The chiral separation and quantification of D-proline and L-proline in honey and royal jelly were examined by LC with UV detection. Most of the endogenous compounds existing in honey, such as sugars, were removed by using SPE cartridges containing C18 and strong cation-exchange sorbent. Other components, such as primary amino acids, were also removed by two-step derivatization with o-phthalaldehyde (OPA) and 9-fluorenylmethyl chloroformate (FMOC-CI). The components that were derivatized with OPA were separated from proline with a C18 cartridge. Proline was then converted into an FMOC derivative that could be subsequently measured by LC-UV. Sufficient chiral separation of D-proline and L-proline was achieved with an LC chiral column made of a beta-cyclodextrin phase in the polar organic-phase mode. The average recoveries of D-proline and L-proline from honey and royal jelly were in the range of 81.3-98.6% (RSD of < 1.8%). When this method was applied to commercial honey and royal jelly samples, L-proline was detected at concentrations of 369-1930 microg/g, whereas D-proline was not detected.

Determination of phthalates in diet and bedding for experimental animals using gas chromatography-mass spectrometry.

We have developed a gas chromatography-mass spectrometry method to measure five phthalates (dibutyl phthalate, butylbenzyl phthalate, di-2-ethylhexyl phthalate, diisooctyl phthalate, and diisononyl phthalate) in diets and beddings for experimental animals. The recoveries from diets and beddings spiked with five phthalates were 98.8%-148% with coefficients of variation of 0.4%-7.8% for diets and 94.7%-146% with coefficients of variation of 1.0%-5.0% for beddings. We analyzed commercial animal diets and beddings, and found that the levels of phthalates varied from sample to sample; the concentrations of five phthalates were 141-1,410 ng/g for diets and 20.5-7,560 ng/g for beddings.

Determination of five phthalate monoesters in human urine using gas chromatography-mass spectrometry.

We have developed a gas chromatography-mass spectrometry (GC-MS) method to determine five phthalate monoesters (monoethyl phthalate (MEP), mono-n-butyl phthalate (MBP), mono-(2-ethylhexyl) phthalate (MEHP), monoisononyl phthalate (MINP) and monobenzyl phthalate (MBz)) in human urine. Human urine samples were subjected to enzymatic deconjugation of the glucuronides followed by extraction with hexane. The extracted phthalate monoesters were methylated with diazomethane, purified on a Florisil column and then subjected to GC-MS analysis. The recoveries from urine spiked with five phthalate monoesters were 86.3%-119% with coefficients of variation of 0.6%-6.1%. We measured phthalate monoester levels in human urine by analyzing 36 samples from volunteers. MBP and MEP were detected in all samples, and their median concentrations were 60.0 and 10.7 ng/mL, respectively. MBzP and MEHP were found in 75% and 56% of samples, and their median concentrations were 10.9 and 5.75 ng/mL, respectively. MINPs were not detected in most samples (6% detectable). Women had significantly (p < 0.05) higher mean concentrations of MBP and MEP than men. The estimated daily exposure levels for the four parent phthalates excluding diisononyl phthalate ranged from 0.27 to 5.69 mug/kg/day (median).

Miniaturized hollow fiber assisted liquid-phase microextraction and gas chromatography-mass spectrometry for determination of benzophenone and derivates in human urine sample.

The determination of benzophenones (BPs) in human urine sample by miniaturized hollow fiber assisted liquid-phase microextraction (HF-LPME) and gas chromatography-mass spectrometry (GC-MS) is described. As analytes, BP, its metabolites benzhydrol (BP-OH) and 2-hydroxybenzophenone (2OH-BP), and its derivatives 2-hydroxy-4-methoxybenzophenone (BP-3) and 2-hydroxy-4-methoxy-4'-methylbenzophenone (BP-10) were selected. The detection limit and the quantification limit of BPs in human urine sample are 5-10 and 20-50 pg mL(-1), respectively. The calibration curve for BPs is linear with correlation coefficient higher than 0.99 in the range of 0.02-10 or 0.05-10 ng mL(-1). The average recoveries of BPs in human urine samples spiked with 0.5 and 5 ng mL(-1) BPs are 89.8-100.2% (RSD: 2.5-9.3%) and 89.3-99.9% (RSD: 2.9-3.7%), respectively. Ten human urine samples were analyzed using the present method. BP-OH and BP-3 were detected in all the samples within the range of 0.24-5.91 and 0.43-5.17 ng mL(-1), respectively. This simple, sensitive, and selective analytical method was successfully applied to the determination of trace amounts of BPs in human urine samples.

Stir bar sorptive extraction and thermal desorption-gas chromatography-mass spectrometry for trace analysis of triclosan in water sample.

A simple and highly sensitive method called stir bar sorptive extraction (SBSE) and thermal desorption (TD)-gas chromatography-mass spectrometry (GC-MS), which is used for the determination of trace amounts of 5-chloro-2-(2,4-dichlorophenoxy)phenol (triclosan) in river water samples, is described. A stir bar coated with polydimethylsiloxane (PDMS) is added to a 10 mL water sample and stirring is carried out for 120 min at room temperature (25 degrees C) in a vial. Then, the PDMS stir bar is subjected to TD-GC-MS. The detection limit of triclosan is 5 ng L(-1) (ppt). The method shows linearity over the calibration range (0.02-20 microg L(-1)) and the correlation coefficient is higher than 0.997 for triclosan standard solution. The recovery of triclosan in river water samples ranges from 91.9 to 108.3% (RSD: 4.0-7.0%). This simple, accurate, sensitive, and selective analytical method may be used in the determination of trace amounts of triclosan in river water samples.

Mercury speciation and analysis in drinking water by stir bar sorptive extraction with in situ propyl derivatization and thermal desorption-gas chromatography-mass spectrometry.

A method for mercury analysis and speciation in drinking water was developed, which involved stir bar sorptive extraction (SBSE) with in situ propyl derivatization and thermal desorption (TD)-GC-MS. Ten millilitre of tap water or bottled water was used. After a stir bar, pH adjustment agent and derivatization reagent were added, SBSE was performed. Then, the stir bar was subjected to TD-GC-MS. The detection limits were 0.01 ng mL(-1) (ethylmercury; EtHg), 0.02 ng mL(-1) (methylmercury; MeHg), and 0.2 ng mL(-1) (Hg(II) and diethylmercury (DiEtHg)). The method showed good linearity and correlation coefficients. The average recoveries of mercury species (n=5) in water samples spiked with 0.5, 2.0, and 6.0 ng mL(-1) mercury species were 93.1-131.1% (RSD<11.5%), 90.1-106.4% (RSD<7.8%), and 94.2-109.6% (RSD<8.8%), respectively. The method enables the precise determination of standards and can be applied to the determination of mercury species in water samples.

Simultaneous analysis of benzophenone sunscreen compounds in water sample by stir bar sorptive extraction with in situ derivatization and thermal desorption-gas chromatography-mass spectrometry.

A method for the simultaneous measurement of benzophenone (BP) sunscreen compounds, its derivatives 2,4-dihydroxybenzophenone (BP-1), 2-hydroxy-4-methoxybenzophenone (BP-3), 2-hydroxy-4-methoxy-4'-methylbenzophenone (BP-10), 2-hydroxybenzophenone (2OH-BP), 3-hydroxybenzophenone (3OH-BP) and 4-hydroxybenzophenone (4OH-BP), in water samples was developed using stir bar sorptive extraction (SBSE) with in situ derivatization followed by thermal desorption (TD)-gas chromatography-mass spectrometry (GC-MS). The detection limit is 0.5-2 ng L(-1) (ppt) for the seven BPs. The method shows good linearity and the correlation coefficients are equal to or higher than 0.990 for all the analyte. The average recoveries of BPs range from 102.0 to 128.1% (RSD<15.4%, n=6). Trace amounts of BPs in river water samples were determined by the present method.

Miniaturized hollow fiber assisted liquid-phase microextraction with in situ derivatization and gas chromatography-mass spectrometry for analysis of bisphenol A in human urine sample.

A new method that involves miniaturized hollow fiber assisted liquid-phase microextraction (HF-LPME) with in situ derivatization and gas chromatography-mass spectrometry (GC-MS) is described for the determination of trace amounts of bisphenol A (BPA) in human urine samples. The detection limit and the quantification limit of BPA in human urine sample are 0.02 and 0.1 ng ml(-1) (ppb), respectively. The calibration curve for BPA is linear with a correlation coefficient of >0.999 in the range of 0.1-50 ng ml(-1). The average recoveries of BPA in human urine samples spiked with 1 and 5 ng ml(-1) BPA are 101.0 (R.S.D.: 6.7%) and 98.8 (R.S.D.: 1.8%), respectively, with correction using the added surrogate standard, bisphenol A-(13)C12. This simple, accurate, sensitive and selective analytical method can be applicable to the determination of trace amounts of BPA in human urine samples.

Determination of urinary triclosan by stir bar sorptive extraction and thermal desorption-gas chromatography-mass spectrometry.

We have developed an analytical method for the determination of urinary 5-chloro-2-(2,4-dichlorophenoxy)phenol (triclosan), which utilizes stir bar sorptive extraction (SBSE) and thermal desorption (TD)-gas chromatography-mass spectrometry (GC-MS). Human urine sample is de-conjugated by treatment with beta-glucuronidase and sulfatase. A stir bar coated with polydimethylsiloxane (PDMS) is added to the urine sample in a vial and the sample is stirred for 60 min at room temperature (25 degrees C). Then, the PDMS stir bar is subjected to TD-GC-MS. The detection limit of triclosan is 0.05 ng mL(-1). The method shows linearity over the calibration range (0.1-10 ng mL(-1)) and the correlation coefficient (r) is higher than 0.993 for triclosan standard solution. The average recoveries of triclosan in human urine sample are 102.8-113.1% (RSD: 2.4-6.7%). This simple, sensitive, and selective analytical method may be used in the determination of trace amounts of triclosan in human urine samples.