Biomedical subjects

Kayoko Kato

Publications and source records attributed to Kayoko Kato.

14 recordsLinked to original sources

PubMed2006

Quantifying phthalate metabolites in human meconium and semen using automated off-line solid-phase extraction coupled with on-line SPE and isotope-dilution high-performance liquid chromatography--tandem mass spectrometry.

We developed an analytical method using off-line solid-phase extraction (SPE) coupled with on-line SPE and isotope-dilution high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) to determine the concentrations of phthalate metabolites in human meconium and in semen. First, we used off-line SPE to remove interfering proteins and other biomolecules from the samples. Then, we preconcentrated the phthalate metabolites in the extract using on-line SPE before measuring them by HPLC-MS/MS. For most of the analytes, the limits of detection ranged between 0.2 and 0.7 ng/g for meconium and between 0.3 and 0.7 ng/mL for semen. The recovery after off-line SPE varied for most analytes between 65 and 99% at concentrations ranging from 3.0 to 30.0 ng/mL in semen and between 67 and 103% at concentrations ranging from 2.0 to 10.0 ng/mL in meconium. Precision measured by the relative standard deviation ranged from 3.2 to 19.1% for intraday and from 3.9 to 18.6% for interday. We validated this novel approach--which is applicable to other biological matrixes, including serum and breast milk--on spiked samples and on five meconium samples and one pooled semen sample from people with no known occupational exposure to phthalates.

Animals↗

PubMed2006

Urinary biomarkers of di-isononyl phthalate in rats.

Commercial di-isononyl phthalate (DiNP) is a mixture of various branched-chain dialkyl phthalates mainly containing nine-carbon alkyl isomers. At high doses in rodents, DiNP is a carcinogen, and a developmental toxicant. After exposure, the diester isomers are de-esterified to form hydrolytic monoesters, monoisononyl phthalates (MiNP), which subsequently metabolize to form oxidative metabolites. These metabolites can be excreted in urine or feces. The urinary excretion of DiNP metabolites was monitored in adult female Sprague-Dawley rats after oral administration of a single dose (300 mg/kg) of commercial DiNP. The metabolites were extracted from urine, resolved with high performance liquid chromatography, analyzed by mass spectrometry, and tentatively identified based on their chromatographic separation and mass spectrometric fragmentation pattern. Because DiNP is an isomeric mixture, its metabolites were also isomeric mixtures that eluted from the HPLC column with close retention times. Mono(carboxy-isooctyl)phthalate (MCiOP) was identified as the major metabolite of DiNP; in addition, mono(hydroxy-isononyl)phthalate (MHiNP) and mono(oxo-isononyl)phthalate (MOiNP) were present. Furthermore, metabolites of di-isooctyl phthalate (DiOP) and di-isodecyl phthalate (DiDP) were also detected. Excretion toxicokinetics of the DiNP metabolites in urine followed a biphasic pattern with initial rapid decay in concentration. Despite potential differences in the metabolism of DiNP among species, MCiOP, MHiNP and MOiNP were detected in humans with no known exposure to DiNP at levels significantly higher than MiNP suggesting that these oxidative metabolites may be better urinary biomarkers of human exposure to DiNP than is MiNP.

Administration, Oral↗

PubMed2005

Urinary metabolites of di-n-octyl phthalate in rats.

Di-n-octyl phthalate (DnOP) is a plasticizer used in polyvinyl chloride plastics, cellulose esters, and polystyrene resins. The metabolism of DnOP results in the hydrolysis of one ester linkage to produce mono-n-octyl phthalate (MnOP), which subsequently metabolizes to form oxidative metabolites. We investigated the toxicokinetics of DnOP in adult female Sprague-Dawley rats by monitoring the excretion of DnOP metabolites in urine after oral administration of DnOP (300 mg/kg). By using authentic standards, the presence of urinary phthalic acid (PA), MnOP, and the major DnOP metabolite, mono-(3-carboxypropyl) phthalate (MCPP) was clearly established. Furthermore, we identified five additional urinary DnOP oxidative metabolites based on their chromatographic behavior and mass spectrometric fragmentation pattern. These DnOP oxidative metabolites, are postulated to be mono-carboxymethyl phthalate (MCMP), mono-(5-carboxy-n-pentyl) phthalate (MCPeP), mono-(7-carboxy-n-heptyl) phthalate (MCHpP), and isomers of mono-hydroxy-n-octyl phthalate (MHOP) (e.g., mono-(7-hydroxy-n-octyl) phthalate) and of mono-oxo-n-octyl phthalate (MOOP) (e.g., mono-(7-oxo-n-octyl) phthalate). The urinary excretion of DnOP metabolites followed a biphasic excretion pattern. The metabolite levels decreased significantly after the first day of DnOP administration although MCPP, MCHpP, MHOP, and MOOP were detectable after 4 days. We also studied the in vitro metabolism of DnOP and MnOP by rat liver microsomes. DnOP produced MnOP, MHOP, and PA in vitro whereas, MnOP produced MHOP and PA in vitro at detectable levels.

Administration, Oral↗

PubMed2005

Determination of 16 phthalate metabolites in urine using automated sample preparation and on-line preconcentration/high-performance liquid chromatography/tandem mass spectrometry.

We developed an on-line solid-phase extraction (SPE) method, coupled with isotope dilution high-performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS) and with automated sample preparation, to simultaneously quantify 16 phthalate metabolites in human urine. The method requires a silica-based monolithic column for the initial preconcentration of the phthalate metabolites from the urine and a silica-based conventional analytical column for the chromatographic separation of the analytes of interest. It uses small amounts of urine (100 microL), is sensitive (limits of detection range from 0.11 to 0.90 ng/mL), accurate (spiked recoveries are approximately 100%), and precise (the inter- and intraday coefficients of variation are <10%). The method is not labor intensive, and, because pretreatment of the urine samples was performed automatically using an HPLC autosampler, involves minimal sample handling, thus minimizing exposure to hazardous chemicals. The method was validated on spiked, pooled urine samples and on urine samples from 43 adults with no known exposure to phthalates. The high sensitivity and high throughput (HPLC run time, including the preconcentration step, is 27 min) of this analytical method combined with the ease of use and effective automated sample preparation procedure make it suitable for large epidemiological studies to evaluate the prevalence of human exposure to phthalates.

Adult↗

PubMed2005

Determination of total phthalates in urine by isotope-dilution liquid chromatography-tandem mass spectrometry.

Diesters of 1,2-benzenedicarboxylic acid are a family of industrial compounds called "phthalates." The physical and chemical properties of these diesters, and therefore their potential uses, depend on the structure of the dialkyl or alkyl/aryl side chain. The urinary concentrations of phthalate monoesters, which are metabolites, have been used as biomarkers of human exposure to specific phthalates. However, several phthalates, particularly those with side chains of eight or more carbon atoms, are complex mixtures of isomers. For these, the phthalate metabolites to be used as biomarkers of exposure have not been unequivocally identified. We developed a method for assessing total exposure to phthalates, including the isomeric mixtures of high molecular weight phthalates, by measuring the concentration of phthalic acid (PA) in human urine after acid hydrolysis of the phthalate metabolites to PA. The present method accurately assesses total exposure to phthalates without noticeable contamination from the ubiquitous phthalates in the environment, but it gives no information about the parent phthalate.

Adult↗

PubMed2004

Mono(2-ethyl-5-hydroxyhexyl) phthalate and mono-(2-ethyl-5-oxohexyl) phthalate as biomarkers for human exposure assessment to di-(2-ethylhexyl) phthalate.

Exposure to di-(2-ethylhexyl) phthalate (DEHP) is prevalent based on the measurement of its hydrolytic metabolite mono-(2-ethylhexyl) phthalate (MEHP) in the urine of 78% of the general U.S. population studied in the 1999-2000 National Health and Nutrition Examination Survey (NHANES). However, despite the high level of production and use of DEHP, the urinary MEHP levels in the NHANES samples were lower than the monoester metabolites of phthalates less commonly used than DEHP, suggesting metabolic differences between phthalates. We measured MEHP and two oxidative DEHP metabolites, mono-(2-ethyl-5-oxohexyl) phthalate (MEOHP) and mono (2-ethyl-5-hydroxyhexyl) phthalate (MEHHP) to verify whether these other metabolites account for a greater proportion of DEHP metabolic products in 127 paired human urine and serum samples. We found that the urinary levels of MEHHP and MEOHP were 10-fold higher than levels of MEHP; concentrations of urinary MEOHP and MEHHP were strongly correlated (r = 0.928). We also found that the serum levels of MEOHP and MEHHP were comparatively lower than those in urine. Furthermore, the glucuronide-bound conjugates of the oxidative metabolites were the predominant form in both urine and serum. MEOHP and MEHHP cannot be formed by serum enzymes from the hydrolysis of any contamination from DEHP potentially introduced during blood collection and storage. Therefore, concentrations of MEHHP and MEOHP in serum may be a more selective measure of DEHP exposure than is MEHP. However, additional data on the absorption, distribution, metabolism, and elimination of these oxidative metabolites are needed to completely understand the extent of DEHP exposure from the serum concentrations of oxidative DEHP metabolites.

Biomarkers↗

PubMed2003

Glucuronidation patterns of common urinary and serum monoester phthalate metabolites.

Metabolism of most diesters of phthalic acid in humans occurs by an initial phase I biotransformation in which phthalate monoesters are formed, followed by a phase II biotransformation in which phthalate monoesters react with glucuronic acid to form their respective glucuronide conjugates. The phase II conjugation increases water solubility and facilitates urinary excretion of phthalate, and reduces the potential biological activity because the putative biologically active species is the monoester metabolite. In this study, we report percentages of glucuronidation of four common phthalate monoesters, monoethyl (mEP), monobutyl (mBP), monobenzyl (mBzP), and mono-2-ethylhexyl phthalate (mEHP) in a subset of urine (mEP n=262, mBP n=283, mBzP n=328, mEHP n=119) and serum (mEP n=93, mBP n=149, mEHP n=141) samples from the general US population. The percentages of free and conjugated monoester excreted in urine differed for the various phthalates. For the more lipophilic monoesters (i.e., mBP, mBzP, and mEHP), the geometric mean of free monoester excretion ranged from 6 to 16%. The contrary was true for the most hydrophilic monoester, mEP, for which about 71% was excreted in urine as its free monoester. Furthermore, percentages of free and conjugated monoesters were similar for mEP, mBP and mEHP among serum and urine samples. Serum mBzP was largely below the method limit of detection. Interestingly, the serum mEP and mBP levels were less than 3% and 47%, respectively, of their urinary levels, whereas the level of mEHP was similar both in urine and serum.

Environmental Pollutants↗

PubMed2003

Improved quantitative detection of 11 urinary phthalate metabolites in humans using liquid chromatography-atmospheric pressure chemical ionization tandem mass spectrometry.

Phthalates are widely used as industrial solvents and plasticizers, with global use exceeding four million tons per year. We improved our previously developed high-performance liquid chromatography-atmospheric pressure chemical ionization-tandem mass spectrometric (HPLC-APCI-MS/MS) method to measure urinary phthalate metabolites by increasing the selectivity and the sensitivity by better resolving them from the solvent front, adding three more phthalate metabolites, monomethyl phthalate (mMP), mono-(2-ethyl-5-oxohexyl)phthalate (mEOHP) and mono-(2-ethyl-5-hydroxyhexyl)phthalate (mEHHP); increasing the sample throughput; and reducing the solvent usage. Furthermore, this improved method enabled us to analyze free un-conjugated mono-2-ethylhexyl phthalate (mEHP) by eliminating interferences derived from coelution of the glucuronide-bound, or conjugated form, of the mEHP on measurements of the free mEHP. This method for measuring phthalate metabolites in urine involves solid-phase extraction followed by reversed-phase HPLC-APCI-MS/MS using isotope dilution with (13)C(4) internal standards. We further evaluated the ruggedness and the reliability of the method by comparing measurements made by multiple analysts at different extraction settings on multiple instruments. We observed mMP, monoethyl phthalate (mEP), mono-n-butyl phthalate (mBP), monobenzyl phthalate (mBzP), mEHP, mEHHP and mEOHP in the majority of urine specimens analyzed with DEHP-metabolites mEHHP and mEOHP present in significantly higher amounts than mEHP.

Atmospheric Pressure↗

PubMed2003

Determination of three phthalate metabolites in human urine using on-line solid-phase extraction-liquid chromatography-tandem mass spectrometry.

An on-line solid-phase extraction-liquid chromatography-tandem mass spectrometry (on-line SPE-HPLC-MS/MS) method was developed for the analysis of metabolites of three phthalate esters in human urine at the low nanogram per milliliter level. The recoveries were above 84.3% and relative standard deviations varied from 0.8 to 4.8%. The compounds along with their deuterated internal standards were detected in the negative ion mode by selective reaction monitoring and the accuracy of the method was improved by isotope dilution. Monobutyl phthalate was detected with median level of 22.5 ng/ml. The median levels for monobenzyl phthalate and monoethylhexyl phthalate were less than the limit of quantitation (LOQ). The on-line SPE-HPLC-MS/MS method allowed the possibility of determining these metabolites within a short time, with increased sensitivity and by using decreased amounts of sample and solvent.

Calibration↗

PubMed2003

Different subtype distributions in two cities in Myanmar: evidence for independent clusters of HIV-1 transmission.

A molecular epidemiological investigation was conducted in two major cities in Myanmar (Yangon and Mandalay). The study revealed a unique predominance of HIV-1 subtype B' (Thailand variant of subtype B) among injecting drug users in Yangon, indicating the strong founder effect of this variant. In contrast, multiple lineages of HIV-1 strains were found in Mandalay, leading to the evolution of various forms of intersubtype recombinants. The results showed independent clusters of HIV-1 transmission in Myanmar.

Female↗

PubMed2003

Assessing human exposure to phthalates using monoesters and their oxidized metabolites as biomarkers.

Phthalates are a group of industrial chemicals with many commercial uses, such as solvents, additives, and plasticizers. For example, di-(2-ethylhexyl) phthalate (DEHP) is added in varying amounts to certain plastics, such as polyvinyl chloride, to increase their flexibility. In humans, phthalates are metabolized to their respective monoesters, conjugated, and eliminated. However, despite the high production and use of DEHP, we have recently found that the urinary levels of the DEHP metabolite mono-(2-ethylhexyl) phthalate (MEHP) in 2,541 persons in the United States were lower than we anticipated, especially when compared with urinary metabolite levels of other commonly used phthalates. This finding raised questions about the sensitivity of this biomarker for assessing DEHP exposure. We explored the utility of two other DEHP metabolites, mono-(2-ethyl-5-oxohexyl) phthalate (MEOHP) and mono-(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP), as additional DEHP biomarkers. These metabolites are formed by oxidative metabolism of MEHP. In urine from 62 people, both the range and the mean urinary levels of MEOHP and MEHHP were on average 4-fold higher than those of MEHP; the mean of the individual ratios of MEHHP/MEOHP, MEHHP/MEHP, and MEOHP/MEHP were 1.4, 8.2, and 5.9, respectively. These data suggest that MEOHP and MEHHP are more sensitive biomarkers of exposure to DEHP than is MEHP. These findings also suggest a predominant human metabolic route for DEHP hydrolysis to MEHP followed by oxidation of MEHP; they also imply that a similar mechanism may be relevant for other high-molecular-weight phthalates, such as di-n-octyl, di-isononyl, and di-isodecyl phthalates.

Biomarkers↗

PubMed2002

Isolation and characterization of replication-competent molecular DNA clones of HIV type 1 CRF01_AE with different coreceptor usages.

We have isolated replication-competent molecular clones of HIV-1 circulating recombinant form CRF01_AE with different coreceptor usages. After lambda phage cloning of unintegrated circular proviral DNAs derived from a CRF01_AE strain (HIV-1NH1), isolated in Japan, the infectious molecular clone, designated p93JP-NH1, was reconstituted. 93JP-NH1 showed an X4 and R5 phenotype in NP2 cell-based coreceptor utilization assays and exerted robust replication in human T cell lines, including MT2, M8166, and PM1 cells, whereas it propagated modestly in peripheral blood mononuclear cells. The CRF01_AE molecular clone with R5 phenotype (p93JP-NH2env) was then constructed by replacing the env gene of p93JP-NH1 with that of a nearly isogenic CRF01_AE R5 strain isolated from an epidemiologically linked case. The phylogeny and recombination break-point analysis confirmed that these clones shared an A/E recombinant structure similar to that of the prototype CRF01_AE strain, CM240. These replication-competent CRF01_AE molecular clones with different coreceptor usages would be useful tools for the study of CRF01_AE, one of the most prevalent strains in Asia.

Base Sequence↗

PubMedDate not supplied

Quantitative detection of nine phthalate metabolites in human serum using reversed-phase high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry.

We developed a highly sensitive method for the quantitative detection of nine phthalate ester metabolites in human serum. This method requires denaturation of the serum enzymes immediately after blood collection to avoid the hydrolysis of the contaminant diester parent compounds introduced during blood collection and storage. Before analysis, the samples were subjected to an enzymatic deconjugation to hydrolyze the glucuronidated phthalate monoesters and a solid-phase extraction to isolate the monoesters from other serum components. The extracts were analyzed using reversed-phase high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry. The limits of detection of all nine phthalate monoesters in serum were in the low nanogram-per-milliliter range (0.6-1.3 ng/mL). Stable isotope-labeled internal standards for all analytes were used to improve precision and for recovery corrections. This highly selective method permits the analysis of phthalate monoesters without interferences resulting from the hydrolysis of the ubiquitous contaminant phthalate diesters by serum enzymes. In addition, it allows the direct measurement of the active phthalate monoester metabolites reportedly responsible for the reproductive and developmental toxicity of certain phthalates.

Chromatography, High Pressure Liquid↗

PubMedDate not supplied

Preparation of samples for gas chromatography/mass spectrometry analysis of phthalate and adipate esters in plasma and beverages by steam distillation and extraction.

Phthalate and adipate esters are present in relatively large amounts in the environment, resulting in their large blank values at analysis and making precise analysis difficult. We developed a highly sensitive analytical method for phthalate and adipate esters in plasma and beverages by lowering the blank values that interfere with analysis. The method uses a closed distillation cleanup system in which steam distillation and extraction are performed simultaneously. The recoveries from beverages and plasma were both satisfactory, ranging from 90.2 to 118.3%, relative standard deviation (RSD) = 2.8-5.3%, and 96.2-134.4%, RSD = 2.2-6.5%, respectively. The detection limits of dibutyl phthalate and di-2-ethyl hexyl phthalate were 5 ng/mL, and those of diethyl phthalate, butyl benzyl phthalate, and di-2-ethyl hexyl adipate were 10 ng/mL in rabbit plasma and beverages.

Adipates↗