Jangwoo Lee

Concentrations of phthalate metabolites in breast milk in Korea: Estimating exposure to phthalates and potential risks among breast-fed infants

Phthalates have been associated with endocrine disruption and developmental effects in many experimental and epidemiological studies. Developing infants are among the most susceptible populations to endocrine disruption. However, limited information is available on phthalate exposure and its associated risks among breast-fed newborn infants. In the present study, breast milk samples were collected from 62 lactating mothers at 1 month post-partum from four cities of Korea in 2012 and were evaluated for six phthalate metabolites (mono-isobutyl phthalate (MiBP), mono-n-butyl phthalate (MnBP), mono(2-ethyl-hexyl) phthalate (MEHP), mono-(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP), mono-(2-ethyl-5-oxohexyl) phthalate (MEOHP) and monoethyl phthalate (MEP)). MEP was detected in all breast milk samples, with a median concentration of 0.37 μg/L, and MiBP, MnBP and MEHP were detected in 79-89% of samples, with median concentrations of 1.10, 1.70, and 2.08 μg/L, respectively. However, MEHHP and MEOHP, the oxidized forms of di-ethyl-hexyl phthalate (DEHP), were detected in only one sample. For exposure assessment, the levels of phthalate diesters were estimated based on the parent:metabolite ratios in the breast milk that are reported elsewhere. For risk assessment, the endocrine-related toxicity of the monoester was assumed to be the same as that of its diester form. Median daily intake estimates of phthalates, including both monoester and diester forms, through breast milk consumption ranged between 0.91 and 6.52 μg/kg body weight (bw) for DEHP and between 0.38 and 1.43 μg/kg bw for di-n-butyl phthalate (DnBP). Based on the estimated daily intake, up to 8% of infants exceeded the reference dose of anti-androgenicity (RfD AA) for DEHP, and 6% of infants exceeded the tolerable daily intake (TDI) for DnBP. Breast milk MiBP and MnBP concentrations showed significant positive associations with maternal consumption of whipped cream or purified water. Considering vulnerability of young infants, efforts to mitigate phthalate exposure among lactating women are warranted.

Association of diethylhexyl phthalate with obesity-related markers and body mass change from birth to 3 months of age

Background Several studies have suggested potential links of phthalates to obesity in children and adults. Limited evidence, however, has been available for the relations between diethylhexyl phthalate (DEHP) and obesity-related markers or body mass change in early life. Methods 128 healthy pregnant women were recruited and, after delivery, their newborns’ first urine and umbilical cord blood samples were collected. We measured urinary levels of two DEHP metabolites, mono-(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP) and mono-(2-ethyl-5-oxohexyl) phthalate (MEOHP). We also measured the levels of leptin, total cholesterol and triglyceride (TG) in cord serum, and used them along with weight, length, head circumference and ponderal index (PI, 100u2005g/cm3) at birth, as obesity-related markers, and estimated the relations between DEHP metabolites and obesity-related markers using generalised linear models. For the evaluation of body mass increase by early life DEHP exposure, body mass index (BMI) z-score change during 3u2005months after birth by DEHP metabolites in the first urine samples of the newborns were evaluated using logistic regression. Results DEHP exposure was associated with decrease of PI and increase of TG (PI, β=−0.11, p=0.070 and TG, β=0.14, p=0.027), especially for boys (PI, β=−0.13, p=0.021; and TG, β=0.19, p=0.025). Moreover, DEHP exposure was positively associated with body mass increase during 3u2005months after birth (change of BMI z-scores, OR=4.35, p=0.025). Conclusions Our findings suggest that DEHP exposure may affect body mass change in early life through changes of obesity-related markers.

Bisphenol A distribution in serum, urine, placenta, breast milk, and umbilical cord serum in a birth panel of mother-neonate pairs

Bisphenol A (BPA) exposure during the perinatal and postnatal periods increases the susceptibility to disease over the life cycle. However, information on the BPA delivered to fetuses or infants via the placenta and breastfeeding is limited. We determined the BPA exposure levels in various bodily fluids and tissues of pregnant women and described fetus and infant exposures to BPA based on associations and BPA ratios in mother-neonate paired samples. Maternal serum, urine, placenta, breast milk, cord serum, and neonatal urine samples were collected from 318 mother-neonate pairs at six university hospitals in Korea. BPA levels were detected using liquid chromatography tandem mass spectrometry. The ratios of the BPA levels in the other sample types to the levels in maternal serum were calculated. BPA was detected in 79.5-100% of the maternal and fetal samples. The median BPA concentration in the samples decreased in the order of neonatal urine (4.75ng/mL), maternal urine (2.86ng/mL), cord serum (1.71ng/mL), maternal serum (1.56ng/mL), breast milk (0.74ng/mL), and the placenta (0.53ng/g). We estimated the ratios of BPA levels in the other sample types to those in maternal serum. The median (95th percentile) cord serum-to-maternal serum ratio was 1.12 (15.2) for 160 mother-fetal pairs, in which BPA was detected in both samples. The placenta-, maternal urine-, neonatal urine-, and breast milk-to-maternal serum ratios were 0.28 (5.31), 1.79 (29.9), 1.98 (28.2), and 0.51 (10.5), respectively. In addition, the median (95th percentile) cord serum-to-placenta ratio was 4.03 (45.8), and the neonatal urine-to-cord serum ratio was 1.95 (25.6). The 95th percentile values were 14-20-fold greater than the medians. Urine contained the highest BPA concentrations, followed by serum, breast milk, and the placenta. The variations of BPA ratio show individual differences in the amounts of BPA delivered from mother to fetus.

DEHP, DEP and DBP Exposure Analysis using Urinary Metabolites of Gyonggi Province University Students

Objectives: Phthalates are used as plasticizers in polyvinyl chloride (PVC) plastics. As phthalate plasticizers are not chemically bound to the PVC, they can leach, migrate or evaporate into indoor air and atmosphere, foodstuffs, other materials, etc. Therefore, humans are exposed through ingestion, inhalation, and dermal exposure over their entire lifetime, including during intrauterine development. In particular, university students have a great number of opportunities to contact products including phthalates during campus life (food packaging, body care products, cosmetic, lotions, aftershave, perfume etc.). The purpose of this study was to examine levels of phthalate exposure as undergraduate students begin to use pharmaceuticals and personal care products including phthalates. Methods: Phthalate metabolites, mono-ethyl phthalate (MEP), mono-n-butyl phthalate (MnBP), mono-isobutyl phthalate (MiBP), mono-2- ethylhexyl phthalate (MEHP), {(mono-(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP}, and mono-(2-ethlyl-5-oxohexyl) phthalate (MEOHP} were examined. 80 urine samples collected from university students were analyzed using LC/MS/MS(API 4000, Applied Bioscience) with on-line enrichment and columnswitching techniques. This study was carried out at Y university located in Gyonggi Province from 2008 to 2011. Results: The detection limit of phthalate metabolites were 0.03 ng/mL for MEP, 0.11 ng/mL for MnBP, 0.08 ng/mL for MiBP, 0.93 ng/mL for MEHP, 0.19 ng/mL for MEOHP and 0.16ng/mL for MEHHP. MnBP showed the highest urinary levels (median: 31.6 ㎍/L, 24.8 ug/g creatinine (cr)). Concentrations were also high for MEHHP (median: 24.1 ㎍/L, 19.0 ㎍/g cr), followed by MEOHP (median: 22.8 ㎍/L, 17.9 ㎍/g cr). In individual cases, the maximum level reached up to 348 ㎍/L, and 291 ㎍/g cr, respectively. The urinary and creatinine adjusted levels of MEP were lower than those for DBP and DEHP metabolites, but were higher in 95th percentiles. As a result, the mean daily DEP intake value was 2.3 ㎍/㎏ bw/day, 3.5 ㎍/㎏ bw/day for DEHP and 4.9 ㎍/㎏ bw/day for DBP. Conclusion: These students’ phthalate exposure levels were below the international safe level set by the EU, but higher than the 2012 KFDA survey of the age group from 3 to 18.

Urinary parabens and triclosan concentrations and associated exposure characteristics in a Korean population—A comparison between night-time and first-morning urine

Parabens and triclosan have been widely used in many personal care products and cosmetics. The endocrine disrupting potential of these compounds is of increasing public health concern. The aim of this study is to understand the current exposure profile of these chemicals in last void before bedtime (night-time) and first-morning void (first-morning) urines among a Korean population and to characterize their exposure sources and pathways. A total of 261 people, including infants (0-2 years), toddlers (3-6 years), children (7-12 years), adolescents (13-18 years), and adults (≥19 years), were recruited, and sampled for night-time urine and first-morning urine of the following day. Methyl (MeP), ethyl (EtP), propyl (PrP) and butyl paraben (BuP), and triclosan were measured in urine. The demographic characteristics, use of personal care products, and food consumption were obtained through a questionnaire. Among the target compounds, EtP and MeP were most frequently detected at the highest concentrations. The median concentration of EtP in night-time urine was 32.4u202fμg/L (interquartile range: 8.37-82.8u202fμg/L), which is higher than previously reported worldwide. Unlike other test compounds, compared to those measured from first-morning urine, the EtP concentrations were significantly higher in night-time urine, suggesting the presence of different exposure sources. Among adults, the MeP and PrP concentrations in night-time urine were associated with frequent use of skin care products, colored cosmetics, bath products, toothpaste, vinyl food packaging, or consumption of canned food. The MeP and PrP concentrations were higher in females than in males, especially in night-time urine. The results of this study also show that multiple urine samples are necessary to capture the diurnal variation of non-occupational exposure to environmental chemicals, such as parabens.

Urinary phthalate metabolites over the first 15 months of life and risk assessment – CHECK cohort study

Phthalates are important group of endocrine disruptors. Infants and young children are susceptible to phthalate exposure. However, information on the phthalate exposure during the early stages of life is very limited. This study was conducted to understand the temporal trend of exposure to major phthalates among infants of Korea during the first 15months after birth, and to estimate associated risks. A total of 286 urine samples were collected from 171 children at 3, 9, 12, or 15months of age, with 77 children sampled for two or more times. Four phthalates, i.e., di(2-ethylhexyl) phthalate (DEHP), di-isobutyl phthalate (DiBP), di-n-butyl phthalate (DnBP), and diethyl phthalate (DEP) were chosen, and their major metabolites were analyzed in the urine. The DEHP metabolites were detected in 100% of the urine samples at relatively higher levels compared to those reported in other countries. The levels of mono-ethyl phthalate (MEP) were generally lower. Urinary concentrations of most phthalate metabolites, especially DEHP metabolites, increased as children grew older. Intra-class correlation coefficients (ICCs) calculated for DEHP metabolites over time were high (0.7-0.8), suggesting persistence of consistent exposure sources during this sensitive period of life. Hazard quotient (HQ) and hazard index (HI) were calculated from daily intake estimates divided by recommended toxicity thresholds. Among the study population, 4, 16, and 26% of the children showed HI >1 at 9, 12, and 15months of age, respectively. DEHP exposure explained most of the risk estimates. Considering vulnerability of young children to endocrine disruption, efforts to identify sources of exposure and to develop appropriate mitigation options are warranted.

Development and Validation of On-line Column Switching HPLC-MS/MS Method for 10 Phthalate Metabolites in Human Urine

Phthalates, such as di (2-ethylhexyl) phthalate (DEHP), dibutyl phthalate (DBP) have been proved to be teratogenics and endocrine disruptors, metabolized rapidly and excreted in the urine. In this study, a simultaneous analytical method for 10 phthalate metabolites, MnBP, MiBP, MBzP, MCHP, MEHP, MEHHP, MEOHP, MnOP, MiNP and MiDP, in human urines, based on switching system with on-line pretreatment column using HPLC-MS/MS has been developed. This method was validated according to the guideline of bioanalytical method validation of National Institute of Toxicological Research. Limits of detection range between 0.2 and 0.9 ng/ml for 10 phthalate metabolites. The calibration curves showed linearity in the range 0.997~0.999, and the results of the intra- and inter-day validations were in the range from 0.4 to 14.7% RSD and from 0.3 to 9.4% RSD, respectively. Recoveries of phthalate metabolites varied from 87.0 to 116.1%. This analytical method showed high accuracy and stable precision for all metabolites, and seems to be suitable for biomonitoring of phthalates in human urine.

Association of phthalate exposures with urinary free cortisol and 8-hydroxy-2′-deoxyguanosine in early childhood

Several studies suggested potential links of phthalates to stress-related outcomes. However, limited evidence has been available for the relationships between phthalate metabolites and free cortisol and 8-hydroxy-2-deoxyguanosine (8-OHdG) in perinatal and postnatal environments. Therefore, we evaluated the relationships between phthalate metabolites and free cortisol and 8-OHdG in mother-child pairs. We repeatedly collected urine samples of 287 mother-child pairs from just before delivery to 15u202fmonths of age to measure the levels of four phthalate metabolites - mono-(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP), mono-(2-ethyl-5-oxohexyl) phthalate (MEOHP), mono-isobutyl phthalate (MiBP), and mono-n-butyl phthalate (MnBP) - and free cortisol and 8-OHdG. We used linear mixed effect models and generalized additive mixed models to estimate the relationship between the phthalate metabolites and free cortisol and 8-OHdG after adjusting for the childs gender, urine collection time, and maternal smoking status. The four phthalate metabolite levels were strongly correlated each other (all, pu202f<u202f.0001), and intra-class correlation for each metabolite in children ranged from 0.18 to 0.96. All four phthalate metabolites were positively associated with both free cortisol (MEHHP, βu202f=u202f0.18 and pu202f<u202f.0001; MEOHP, βu202f=u202f0.17 and pu202f<u202f.0001; MiBP, βu202f=u202f0.13 and pu202f=u202f.0001; MnBP, βu202f=u202f0.21 and pu202f<u202f.0001; and molar sum of metabolites, βu202f=u202f0.21 and pu202f<u202f.0001) and 8-OHdG (MEHHP, βu202f=u202f0.20 and pu202f<u202f.0001; MEOHP, βu202f=u202f0.18 and pu202f<u202f.0001; MiBP, βu202f=u202f0.23 and pu202f<u202f.0001; MnBP, βu202f=u202f0.28 and pu202f<u202f.0001; and molar sum of metabolites, βu202f=u202f0.29 and pu202f<u202f.0001) in childhood. Our findings suggest that phthalate exposures increase free cortisol and 8-OHdG levels in early childhood.

Analysis of dutasteride in human serum by LC-MS/MS

The determination and confirmation of dutasteride in human serum was performed by a liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI/MS/MS). Beclomethasone as an internal standard (I.S.) was added to the serum and the mixed sample was pretreated by liquid-liquid extraction (LLE) with methyl tert-butyl ether (MTBE). The mass transitions of dutasteride and I.S. monitored in multiple reaction monitoring (MRM) were m/z 529.6461.5 and m/z 409.3391.2, respectively, and the retention times were 6.45 and 5.46 min, respectively. The calibration curve was linear in the concentration range of 0.5~30.0 ng/mL (