Jeongae Lee

Mass spectrometry based metabolomic approaches in urinary biomarker study of women's cancers.

BACKGROUND The metabolomic approaches for mining biomarkers of womens cancers based on gas chromatography-mass spectrometry and liquid chromatography-mass spectrometry combined with partial least squares-discriminant analysis are described. METHODS To identify urinary potential biomarkers, the qualitative and quantitative analyses were introduced with 10 breast, 9 ovarian and 12 cervical cancer patients as well as 22 normal controls, which were considered with their ages and menopausal state. RESULTS For comprehensive metabolomic approaches, the non-targeted qualitative profiling was first achieved to get metabolic patterns of collected samples and the targeted quantitative analysis focused on hormonal metabolism was also conducted. Two known biomarkers, i.e., 5-hydroxymethyl-2-deoxyuridine and 8-hydroxy-2-deoxyguanosine, in breast cancer were also confirmed using the present methods. In addition, 3 potential biomarkers for ovarian cancer i.e. 1-methyladenosine, 3-methyluridine, and 4-androstene-3,17-dione, which were categorized in significantly increased level using one way of variance analysis (p<0.05), were identified as quantitatively targeted metabolites with pattern analysis. The cancer markers identified in this study are highly related to metabolites which are responsible for oxidative DNA damage and DNA methylation process. CONCLUSION The present metabolomic approaches are not only useful for diagnostic tools and patient stratification, but may be mapped on metabolic network to reflect disease states.

Urinary metabolites before and after cleanup and subjective symptoms in volunteer participants in cleanup of the Hebei Spirit oil spill.

BACKGROUND On December 7th, 2007, the Hong Kong tanker Hebei Spirit (HS) (146,848 tons) was crushed by a crane ship near the shore of Taean, Korea. More than 12,547 kl of crude oil spilled into the sea and contaminated the western coastline of the Korean peninsula. For a period of six months after the accident, approximately 1,000,000 volunteers participated in the cleanup. Our goal in this study was to examine the exposure status and acute health effects on volunteers that participated in the oil spill cleanup. METHODS A survey questionnaire was filled out by 565 volunteers, requesting information regarding physical symptoms. Out of the total number of participants, urine samples from 105 university student volunteers were collected before and after the cleanup work, and metabolite levels of volatile organic compounds and polycyclic aromatic hydrocarbons were analyzed. RESULTS Volunteers that participated for longer cleanup work reported an increase in physical symptoms including visual disturbance, nasal and bronchus irritation, headaches, heart palpitations, fatigue and fever, memory and cognitive disturbance, and abdominal pain. The levels of t,t-muconic acid, mandelic acid, and 1-hydroxypyrene were significantly higher in samples after cleanup than those measured before participation (p<0.05). Other than the associated risk of dermal irritation with the difference in the t,t-muconic acid level between the post- to pre-cleanup levels, no other physical symptoms demonstrated a significant association with changes observed in the levels of urinary metabolites. CONCLUSIONS Based on the significant increase of subjective symptoms in volunteers participating in the study, monitoring of the long term health effects, focusing on those with longer exposure, is warranted.

Association between urinary levels of bisphenol-A and estrogen metabolism in Korean adults

Bisphenol-A (BPA) possesses estrogenic properties both in vitro and in vivo as an endocrine disrupting chemical. Humans experience a long-term and cumulative exposure to BPA. BPA was detectable in 97.3% of 1904 urine specimens from Korean adults. We investigated urinary estrogen concentrations in subjects with low and high BPA concentrations and its possible association with estrogen metabolism. Urine samples were collected from a high BPA concentration group (BPA-H; n=100, 11.05 ± 20.47 μg/g creatinine) and a low BPA concentration group (BPA-L; n=100, 0.70 ± 0.22 μg/g creatinine) from Korea Biomonitoring Program of Hazardous Materials Survey 2009-2010. Urinary estrogens were enzymatically hydrolyzed, extracted, and then derivatized for quantitative analysis by gas chromatography-mass spectrometry. Estrogen levels were higher in the BPA-H group than in the BPA-L group. Concentrations of estrone, 17β-estradiol, and their hydroxylated metabolites in both men and women were significantly higher in the BPA-H group than in the BPA-L group (p<0.04). Furthermore, in the BPA-H group, estrogen metabolism to 4-hydroxy-estrone and 4-hydroxy-17β-estradiol was more active than that to 2-hydroxy-estrone and 2-hydroxy-17β-estradiol. Although single measurement and/or single spot urine samples limit the measurement of long-term exposure to BPA, we found significant differences of estrogen metabolism in the BPA-H and the BPA-L groups. The increase of hydroxyestrogens, especially 4-hydroxyestrogens, can be an important factor resulting negative effects of prolonged exposure to BPA.

Changes in steroid metabolism among girls with precocious puberty may not be associated with urinary levels of bisphenol A

Precocious puberty (PP) refers to the appearance of physical and hormonal signs of pubertal development at an abnormally early age. Urinary steroid signatures obtained from 42 patients with central PP and 40 patients with peripheral PP were assessed to compare metabolic changes. Levels of androgens such as testosterone, androstenedione, androstenediol, 16α-hydroxy-dehydroepiandrosterone, and 5α-androstenedione tended to be high in both PP groups, and the level of 17β-estradiol was higher in the central-PP group (P<0.01) than in the peripheral-PP and 32 age-matched healthy girls. Altered steroid metabolism was also associated with urinary BPA levels, and levels of testosterone, 17β-estradiol, and pregnenolone were significantly increased among individuals with high BPA levels. In particular, a correlation was observed between estrogen metabolism and BPA levels irrespective of the type of PP. These findings suggest that in girls, BPA exposure causes metabolic changes in steroidogenesis, but not the early onset of PP.

Biomonitoring of urinary di(2-ethylhexyl) phthalate metabolites of mother and child pairs in South Korea

Di(2-ethylhexyl) phthalate (DEHP) is one of the common phthalate plasticizers used primarily in soft polyvinyl chloride, which is a plastic polymer that makes up the total weight of goods from 1% up to 40% in many consumer products. The aims of this study were to examine the urinary DEHP metabolites in South Korean children and to investigate the correlation between mother and child DEHP urine excretion. Three kinds of urinary DEHP metabolites: mono(2-ethylhexyl) phthalate (MEHP), mono(2-ethyl-5-hydroxyhexyl) phthalate (5-OH-MEHP) and mono(2-ethyl-5-oxohexyl) phthalate (5-oxo-MEHP), were analyzed. The total of 954 samples (nChildren=392, nMothers=265, nAadults=297), including 258 mother and child pairs, were analyzed using isotope dilution gas chromatography-mass spectrometry. Many studies present higher concentration of DEHP metabolites detected from adults in reproductive age than adults in other ages. Therefore, adults who are age-matched to mothers were evaluated to serve as a standard of comparison against mothers. All statistical analysis was made by adjusting detected volume concentrations (μg/L) with respect to creatinine concentrations (mg/dL) since urinary DEHP metabolites were studied using human reference. The difference in median levels of sum of urinary DEHP metabolites was only significant when children were analyzed in relation to region (p-value≤0.005). Among the three DEHP metabolites, only MEHP of children was significantly correlated to that of paired mothers (p-value≤0.01). The present paper defines the relative metabolic rate (RMR) of DEHP metabolism for the first time in study on phthalates. Children had faster RMR than mothers and adults, specifically in the first step of DEHP metabolism (RMR1: MEHP hydroxylation to 5-OH-MEHP), and RMR1 of children between 1 and 24 months was the fastest. The above results may be used to study and assess human health risk from DEHP exposures, especially among mothers and children in Korea.

The Effects of Wearing Protective Devices among Residents and Volunteers Participating in the Cleanup of the Hebei Spirit Oil Spill

OBJECTIVES To assess the protective effects of wearing protective devices among the residents and volunteers who participated in the cleanup of the Hebei Spirit oil spill. METHODS A total of 288 residents and 724 volunteers were surveyed about symptoms, whether they were wearing protective devices and potential confounding variables. The questionnaires were administered from the second to the sixth week following the accident. Spot urine samples were collected and analyzed for metabolites of 4 volatile organic compounds (VOCs), 2 polycyclic aromatic hydrocarbons (PAHs), and 6 heavy metals. The association between the wearing of protective devices and various symptoms was assessed using a multiple logistic regression adjusted for confounding variables. A multiple generalized linear regression model adjusted for the covariates was used to test for a difference in least-square mean concentration of urinary biomarkers between residents who wore protective devices and those who did not. RESULTS Thirty nine to 98% of the residents and 62-98% of volunteers wore protective devices. Levels of fatigue and fever were higher among residents not wearing masks than among those who did wear masks (odds ratio 4.5; 95% confidence interval 1.23-19.86). Urinary mercury levels were found to be significantly higher among residents not wearing work clothes or boots (p<0.05). CONCLUSIONS Because the survey was not performed during the initial high-exposure period, no significant difference was found in metabolite levels between people who wore protective devices and those who did not, except for mercury, whose biological half-life is more than 6 weeks.

Simultaneous analysis of urinary phthalate metabolites of residents in Korea using isotope dilution gas chromatography–mass spectrometry

Phthalates are used in industry products, household items, and medical tools as plasticizers. Human exposure to phthalates has raised concern about its toxicity. In the present study, optimization was conducted for the simultaneous analysis of eight kinds of phthalate metabolites using gas chromatography-mass spectrometry (GC-MS): MEP, MiBP, MnBP, MBzP, MiNP, MEHP, MEOHP, and MEHHP. In order to minimize the matrix effect and to do quantitative analysis, isotope dilution and LLE-GC-MS methods were performed. Urine samples were enzymatically hydrolyzed, extracted with a mixture of n-hexane and ethyl ether (8:2; v:v), and subsequently derivatized with trimethylsilylation. All eight kinds of analytes showed clear resolution and high reproducibility in GC-MS results. The method detection limit ranged from 0.05 ng/mL to 0.2 ng/mL. Calibration curves were found to be linear from 0.2 to 100 ng/mL with -(2)>0.992. The relative standard deviation of the intraday precision using water and urine ranged from 2.1% to 16.3%. The analysis was performed with urine samples that were collected from adults residing in the Republic of Korea. The analyzed concentration results were compared according to gender and region. As a result, DEHP metabolites showed the highest detected concentration (75.92 μg/g creatinine, 100%), and MiNP, a metabolite of DiNP, showed the lowest detected concentration (0.42 μg/g creatinine, 22.5%). On average, female urine (200.76 μg/g creatinine) had a higher detected concentration of ∑8 phthalate metabolites than male urine. Samples from rural regions (211.96 μg/g creatinine) had higher levels than samples from urban regions.

Spectroscopic measurement of the acid dissociation constant of 2-naphthol and the second dissociation constant of carbonic acid at elevated temperatures

2-Naphthol can be used to measure the pH of aqueous solutions if the acid dissociation constant for 2-naphthol is known at a given temperature. The temperature dependence of the acid dissociation constant for 2-naphthol was spectroscopically determined in borate buffer solutions under vapor-saturated pressure up to 200°C. The result was where T is in Kelvin. The pH of carbonate buffer solutions was measured by obtaining the UV–visible spectra of 2-naphthol and utilized to determine the second dissociation constant of carbonic acid up to 175°C under vapor-saturated pressure. The result was where T is in Kelvin. By using the temperature dependence of the dissociation constant, the isocoulombic reaction was studied. The plot of -log Kisoc against 1/T was approximately linear in the temperature range studied. The linearity can be used to predict the dissociation constant by the extrapolation of -log Kisoc to higher temperatures.

Urinary metabolic profiling of volatile organic compounds in acute exposed volunteers after an oil spill in Republic of Korea

The Herbei Spirit oil spill occurred in western Korea. A large number of people who participated in the cleanup tasks of the contaminated area were exposed to crude oil component. We developed a method to monitor volatile organic compound (VOC) metabolites in urine, and evaluate the acute exposure caused by the oil spill in exposed volunteer workers (n = 100, 20.7 +/- 2.1 years, mean +/- SD). Acidified urine samples were extracted by SPE, derivatized with trimethylsilyl, and analyzed using gas chromatography-mass spectrometry. Calibration curves were found to be linear from 3 to 1000 ng/mL (r(2) > 0.993). Accuracy was over 82.4%, and precision was less than 24.8%. Using this method, the VOC metabolites, except hippuric acid, were present at higher levels in the urine samples of volunteers after cleanup work. The levels of mandelic acid (MA) and trans,trans-muconic acid (t,t-MU) were increased significantly (p < 0.001). The exposure effect was greater in women than in men. The effect of smoking was analyzed in all exposed and non-exposed groups, with non-smokers showing increased MA and t,t-MU levels related to exposure. The present method was reliable to determine VOC metabolites in urine and could be useful for biomonitoring of acute exposure effects of VOCs.

Identification of a pyrovalerone metabolite in the rat by gas chromatography-mass spectrometry and determination of pyrovalerone by gas chromatography-nitrogen-phosphorus detection.

Pyrovalerone and its hydroxylated metabolite have been identified by gas chromatography-mass spectrometry in rat urine and plasma. A sensitive gas chromatographic method for the quantitative analysis of pyrovalerone in rat urine and plasma is described. The method also permits the quantitative monitoring of the urinary excretion of the drug and its metabolite. Pyrovalerone and its hydroxylated metabolite are detected up to 18 h after a single oral administration to the rat at a dose of 20 mg/kg.