Yusuke Iwasaki

Correlations between Prenatal Exposure to Perfluorinated Chemicals and Reduced Fetal Growth

Background Perfluorooctane sulfonate (PFOS) and perfluorooctanoate (PFOA) are man-made, ubiquitous, and persistent contaminants in the environment, wildlife, and humans. Although recent studies have shown that these chemicals interfere with fetal growth in humans, the results are inconsistent. Objectives Our goal was to investigate the correlation between relatively low levels of PFOS and PFOA in maternal serum and birth weight and birth size. Methods We conducted a hospital-based prospective cohort study between July 2002 and October 2005 in Sapporo, Japan. A total of 428 women and their infants were involved in the study. We obtained characteristics of the mothers and infants from self-administered questionnaire surveys and from medical records. We analyzed maternal serum samples for PFOS and PFOA by liquid chromatography–tandem mass spectrometry (LC/MS/MS). Results After adjusting for confounding factors, PFOS levels negatively correlated with birth weight [per log10 unit: β = −148.8 g; 95% confidence interval (CI), −297.0 to −0.5 g]. In addition, analyses stratified by sex revealed that PFOS levels negatively correlated with birth weight only in female infants (per log10 unit: β = −269.4 g; 95% CI, −465.7 to −73.0 g). However, we observed no correlation between PFOA levels and birth weight. Conclusion Our results indicate that in utero exposure to relatively low levels of PFOS was negatively correlated with birth weight.

A new strategy for ionization enhancement by derivatization for mass spectrometry

Liquid chromatography-mass spectrometry (LC-MS) using atmospheric pressure ionization is drastically different from hitherto available analytical methods used to detect polar analytes. The electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) sources of MS have contributed to the advancement of LC-MS and LC-MS/MS techniques for the analysis of biological samples. However, one major obstacle is the weak ionization of some analytes in the ESI and APCI techniques. In this review, we introduce high-sensitivity methods using several derivatization reagents for ionization enhancement. We also present an overview of chemical derivatization methods that have been applied to small molecules, such as amino acids and steroids, in biological samples.

Chromatographic and mass spectrometric analysis of glutathione in biological samples.

Biological thiol compounds are classified into high-molecular-mass protein thiols and low-molecular-mass free thiols. Endogenous low-molecular-mass thiol compounds, namely, reduced glutathione (GSH) and its corresponding disulfide, glutathione disulfide (GSSG), are very important molecules that participate in a variety of physiological and pathological processes. GSH plays an essential role in protecting cells from oxidative and nitrosative stress and GSSG can be converted into the reduced form by action of glutathione reductase. Measurement of GSH and GSSG is a useful indicator of oxidative stress and disease risk. Many publications have reported successful determination of GSH and GSSG in biological samples. In this article, we review newly developed techniques, such as liquid chromatography coupled with mass spectrometry and tandem mass spectrometry, for identifying GSH bound to proteins, or for localizing GSH in bound or free forms at specific sites in organs and in cellular locations.

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.

Determination of uric acid in human saliva by high-performance liquid chromatography with amperometric electrochemical detection.

The aim of the present study is to establish a highly sensitive method for the determination of uric acid (UA) in human saliva. The monitoring of UA levels in less invasive biological samples such as saliva is suggested for the diagnosis and therapy of gout, hyperuricemia, and the Lesch-Nyhan syndrome, and for detecting such conditions as alcohol dependence, obesity, diabetes, high cholesterol, high blood pressure, kidney disease, and heart disease. Reversed-phase high-performance liquid chromatography with electrochemical detection (HPLC-ED) was employed for the determination of UA obtained by solid-phase extraction from saliva. To quantify UA, we compared the ED efficiencies of an amperometric ED (Ampero-ED) with a single electrode and a coulometric ED (Coulo-ED) with a multiple electrode array. The results showed that the detection limits (S/N=3) were 3 nM for Ampero-ED and 6 nM for Coulo-ED, and the linearity of the calibration curves of 60-6000 nM had correlation coefficients exceeding 0.999. In addition, the total analytical time was 10 min. In the sample preparation of UA in saliva, an Oasis MAX solid-phase cartridge was used. The recoveries of UA spiked at 0.6 and 3 microM in saliva were above 95% with a relative standard deviation (RSD) of less than 15%. Therefore, the present method may be used in the routine and diagnostic determination of UA in human saliva.

Improvement and validation the method using dispersive liquid-liquid microextraction with in situ derivatization followed by gas chromatography-mass spectrometry for determination of tricyclic antidepressants in human urine samples.

A simple, rapid and sensitive method termed dispersive liquid-liquid microextraction (DLLME) combined with gas chromatography-mass spectrometry (GC/MS) was developed for the determination of tricyclic antidepressants (TCAs) in human urine sample. An appropriate mixture of methanol (disperser solvent), carbon tetrachloride (extraction solvent), and acetic anhydride (derivatization reagent) was injected rapidly into human urine sample. After extraction, the sedimented phase was analyzed by GC/MS. The calibration curves obtained with human urine were linear with a correlation coefficient of over 0.99 in the range of 2.0/5.0-100 ng mL(-1). Under the optimum conditions (carbon tetrachloride: 10 μL, methanol: 150 μL), the detection limits and the quantification limits of the tricyclic antidepressants were 0.5-2.0 ng mL(-1) and 2.0-5.0 ng mL(-1), respectively. The average recoveries of TCAs were 88.2-104.3%. Moreover, the inter- and intra-day precision and accuracy was acceptable at all concentrations. The results showed that DLLME is applicable to the determination of trace amounts of TCAs in human urine sample.

The Association of Prenatal Exposure to Perfluorinated Chemicals with Maternal Essential and Long-Chain Polyunsaturated Fatty Acids during Pregnancy and the Birth Weight of Their Offspring: The Hokkaido Study

Background Fatty acids (FAs) are essential for fetal growth. Exposure to perfluorinated chemicals (PFCs) may disrupt FA homeostasis, but there are no epidemiological data regarding associations of PFCs and FA concentrations. Objectives We estimated associations between perfluorooctane sulfonate (PFOS)/perfluorooctanoate (PFOA) concentrations and maternal levels of FAs and triglyceride (TG) and birth size of the offspring. Methods We analyzed 306 mother–child pairs in this birth cohort between 2002 and 2005 in Japan. The prenatal PFOS and PFOA levels were measured in maternal serum samples by liquid chromatography–tandem mass spectrometry. Maternal blood levels of nine FAs and TG were measured by gas chromatography–mass spectrometry and TG E-Test Wako kits, respectively. Information on infants’ birth size was obtained from participant medical records. Results The median PFOS and PFOA levels were 5.6 and 1.4 ng/mL, respectively. In the fully adjusted model, including maternal age, parity, annual household income, blood sampling period, alcohol consumption, and smoking during pregnancy, PFOS but not PFOA had a negative association with the levels of palmitic, palmitoleic, oleic, linoleic, α-linolenic, and arachidonic acids (p < 0.005) and TG (p-value = 0.016). Female infants weighed 186.6 g less with mothers whose PFOS levels were in the fourth quartile compared with the first quartile (95% CI: –363.4, –9.8). We observed no significant association between maternal levels of PFOS and birth weight of male infants. Conclusions Our data suggest an inverse association between PFOS exposure and polyunsaturated FA levels in pregnant women. We also found a negative association between maternal PFOS levels and female birth weight. Citation Kishi R, Nakajima T, Goudarzi H, Kobayashi S, Sasaki S, Okada E, Miyashita C, Itoh S, Araki A, Ikeno T, Iwasaki Y, Nakazawa H. 2015. The association of prenatal exposure to perfluorinated chemicals with maternal essential and long-chain polyunsaturated fatty acids during pregnancy and the birth weight of their offspring: the Hokkaido Study. Environ Health Perspect 123:1038–1045; http://dx.doi.org/10.1289/ehp.1408834

New Approaches for Analysis of Metabolism Compounds in Hydrophilic Interaction Chromatography

Abstract Hydrophilic interaction chromatography (HILIC) was developed by Andrew Alpert for separation of highly polar compounds. HILIC is characterized by a high organic mobile phase concentration and the hydrophilic stationary phase. Thus, the retention times of highly polar compounds are increased with the increase of the hydrophilicity of the solutes. Recently, it became necessary to determine highly polar compounds, such as pharmaceutical metabolism products. Therefore, HILIC has been promoted by the demand to analyze polar compounds in a complex matrix. Another reason for the increase in popularity is the widespread use of mass spectrometry (MS) coupled to liquid chromatography (LC). It is easy to promote ionization of several compounds, because there is low concentration of aqueous mobile phase. This review attempts to summarize the ongoing discussion of the separation mechanism and gives an overview of the stationary phases used and the applications addressed with this separation mode in LC.

Determination of tris(2-ethylhexyl)trimellitate released from PVC tube by LC-MS/MS.

Tris(2-ethylhexyl)trimellitate (TOTM) is used as an alternative plasticizer of polyvinyl chloride (PVC) medical devices. A method for the determination of TOTM released from PVC medical devices into intravenous preparations was developed, which uses liquid chromatography-tandem mass spectrometry (LC-MS/MS). A PVC tube was filled with an intravenous preparation and extraction was carried out by shaking for 1h at room temperature. LC was performed with an Inertsil-C8 (50 mm x 2.1 mm, 5 microm) column. The isocratic mobile phase was acetonitrile:purified water (90:10, v/v) at a flow rate of 0.2 ml/min. MS detection was accomplished with an MS/MS detector equipped with a turbo ionspray ionization source in the positive ion mode. The limit of detection and the limit of quantification for the standard solution of TOTM was 0.5 ng/ml (S/N=3) and 1.0 ng/ml (S/N > or =10), respectively. When Prograf (tacrolimus) was used, the average recovery of TOTM was 101.1% (R.S.D.=4.72%; n=3). When our method was applied to the determination of TOTM released from unsterilized and gamma-ray-sterilized PVC tubes, we found that a higher concentration of TOTM was released from the unsterilized PVC tube than from the gamma-ray-sterilized one.

Demographic, behavioral, dietary, and socioeconomic characteristics related to persistent organic pollutants and mercury levels in pregnant women in Japan

Persistent organic pollutants and mercury are known environmental chemicals that have been found to be ubiquitous in not only the environment but also in humans, including women of reproductive age. The purpose of this study was to evaluate the association between personal lifestyle characteristics and environmental chemical levels during the perinatal period in the general Japanese population. This study targeted 322 pregnant women enrolled in the Hokkaido Study on Environment and Childrens Health. Each participant completed a self-administered questionnaire and a food-frequency questionnaire to obtain relevant information on parental demographic, behavioral, dietary, and socioeconomic characteristics. In total, 58 non-dioxin-like polychlorinated biphenyls, 17 dibenzo-p-dioxins and -dibenzofuran, and 12 dioxin-like polychlorinated biphenyls congeners, perfluorooctane sulfonate, perfluorooctanoic acid, and mercury were measured in maternal samples taken during the perinatal period. Linear regression models were constructed against potential related factors for each chemical concentration. Most concentrations of environmental chemicals were correlated with the presence of other environmental chemicals, especially in the case of non-dioxin-like polychlorinated biphenyls and, polychlorinated dibenzo-p-dioxins and -dibezofurans and dioxin-like polychlorinated biphenyls which had similar exposure sources and persistence in the body. Maternal smoking and alcohol habits, fish and beef intake and household income were significantly associated with concentrations of environmental chemicals. These results suggest that different lifestyle patterns relate to varying exposure to environmental chemicals.