Environ. Sci. Technol. 2010, 44, 7123–7129
Pre- and Postnatal Exposure to Perfluorinated Compounds (PFCs) H E R M A N N F R O M M E , * ,† CHRISTINE MOSCH,† MARIA MOROVITZ,‡ IRENE ALBA-ALEJANDRE,‡ SIGRUN BOEHMER,† MANDY KIRANOGLU,† FABIENNE FABER,§ IRIS HANNIBAL,§ ´ NY,§ ORSOLYA GENZEL-BOROVICZE BERTHOLD KOLETZKO,§ AND ¨ LKEL† WOLFGANG VO Department of Chemical Safety and Toxicology/ Biomonitoring, Bavarian Health and Food Safety Authority, Pfarrstrasse 3, D-80538 Munich, Germany, Department of Obstetrics and Gynecology, Ludwig-Maximilians University, Maistrasse 11, D-80337 Munich, Germany, and Dr. von Hauner Children’s Hospital, Ludwig-Maximilians University, Lindwurmstrasse 4, D-80337 Munich, Germany
Received April 14, 2010. Revised manuscript received July 24, 2010. Accepted July 27, 2010.
Perfluorinated compounds (PFCs) are a group of chemicals widely used for many applications. In this study PFCs were investigated in maternal blood during pregnancy (at two time points) (n ) 40 and 38) and 6 months after delivery (n ) 47), in cord blood (n ) 33) and in blood of infants six (n ) 40) and nineteen months (n ) 24) after birth, and monthly in breast milk samples in Germany. Concentrations in maternal serum ranged from 0.5 to 9.4 µg/L for perfluorooctane sulfonate (PFOS) and 0.7 to 8.7 µg/L for perfluorooctanoic acid (PFOA). In cord serum, the values ranged from 0.3 to 2.8 µg/L and from 0.5 to 4.2 µg/L for PFOS and PFOA, respectively. The median results from serum at six and nineteen months of age were 3.0 and 1.9 µg/L for PFOS and 6.9 and 4.6 µg/L for PFOA, respectively. In breast milk samples, PFOS ranged from 60 years of age were analyzed (29). The authors concluded that PFC concentrations increased with age, but this was significant for PFOS and PFOA only in the sampling year 2007, not in 1976, 1987, or 1998. In another study from Australia, using pooled blood samples from subjects 0 to >60 years old, a constant increase was reported for PFOS according to age group (35). For PFOA the lowest mean levels were observed VOL. 44, NO. 18, 2010 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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in subjects 0-0.5 years old, whereas in the other age groups the concentrations appeared to be relatively stable. Animal studies suggest that PFCs can cross the placenta and enter the fetus (31). Investigations from Japan, Germany, and Denmark, in which PFCs were detected simultaneously in the umbilical cord and maternal blood, indicated that the human fetus is also exposed to PFCs (14, 32-36). The mechanism by which perfluorinated substances are transferred from the mother’s blood to breast milk is not fully understood. Two animal studies showed that PFOS and PFOA enter breast milk, and therefore, suckling pups will be exposed to PFCs (31, 37). PFOS and PFOA concentrations found in milk of experimental animals were between 3% and 49% (37) and 10% (31), respectively, of the concentrations in maternal serum samples. Results for PFCs in human milk have also been reported in various studies from Asia (38-40), the U.S. (37, 41, 42), and Europe (43-46). To our best knowledge, longitudinal data on the body burden of mothers (at different time points), fetuses, and infants in the first months of their life have not been reported so far. Thus, the main objective of the study was to investigate maternal, fetal, and infant body burden of PFCs and improve our knowledge of the exposure in utero and during critical periods of infant development. For this reason, we analyzed PFCs in maternal blood twice during pregnancy and once after delivery, in cord blood, and in the blood of infants six and nineteen months old. Moreover, the same target substances were repeatedly measured in breast milk samples up to five months after delivery to characterize the exposure of infants during lactation. Finally, the concentrations of PFCs were quantified in a limited number of infant formulas. This study is a pilot study of the Mother and Infant Biomonitoring Survey (MIBS), which aims to characterize the body burden in a considerably larger randomly selected population and compare it with health outcomes.
Material and Methods Study Population and Sampling. Samples of maternal blood, cord blood, blood from infants, and breast milk were collected during December 2007 and October 2009 in Munich, Germany. The study population consisted of randomly selected females who were participating in a birthing class at the Department of Obstetrics and Gynecology at the Ludwig-Maximilians University. At 32 weeks of gestation, women were asked to participate and were informed about the aim and study protocol during an interview. A total of 53 women agreed to participate. All of them consumed a normal mixed diet, were healthy subjects, and had normal pregnancies. All participants received a questionnaire about age, weight, smoking habits, socioeconomic status, family and housing conditions, and other potential influencing factors. Blood samples (20-30 mL) were collected by venipuncture from women at the 34th to 37th week of pregnancy, again when they arrived at the hospital for delivery, and around 6 months after birth. Umbilical cord blood (10 mL) was obtained immediately after birth. Around 6 months (5-7 months) and 19 months (16-20 months) after birth, 1-2 mL of blood was collected from the infants by venipuncture. In all samples, plasma was separated by centrifugation. All samples were stored at -20 °C until analysis. Over a 5-month period after delivery, monthly breast milk samples (50-150 mL) were obtained by mothers using milk pumps. For quality measures, the pumps were tested for contamination of the target analytes. Overall, five different pumps were ternary rinsed with 100 mL of ethanol. After solid-phase extraction PFCs were quantified with LC-MS/ MS (TSQ Quantum Ultra AM) equipped with an electrospray ionization source. In none of the samples PFCs could be detected. 7124
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Furthermore, four infant formulas were purchased from retail stores and prepared just before analysis with tap water as described by the producers. We could not measure PFCs in the tap water used, but according to information available in our institution no PFCs could be measured above the quantification limit of 4 ng/L in the water distribution system of the study region. The study protocol was approved by the ethics committee of the Bavarian Chamber of Physicians. All participants provided written informed consent and both parents gave consent for child participation. Sample Preparation and Analysis. Methanol (LiChrosolv) and acetonitrile (LC-MS grade) were purchased from Merck KGaA (Darmstadt, Germany), water (Rotisolv HPLC Gradient grade) was purchased from Roth (Karlsruhe, Germany), and ammonium acetate p.a. was from Riedel-de-Hae¨n (Hannover, Germany). Perfluorobutanesulfonic acid tetrabutylammonium salt (PFBS), potassium perfluorooctanesulfonate (PFOS), perfluorononanoic acid (PFNA), perfluorodecanoic acid (PFDA), and perfluorododecanoic acid (PFDoA) were from SigmaAldrich (Taufkirchen, Germany). Perfluorooctanoic acid (PFOA), sodium perfluorohexanesulfonate (PFHxS), and the corresponding isotope-labeled internal standards sodium perfluoro-1-hexane [18O2] sulfonate (MPFHxS), sodium perfluoro-1-[1,2,3,4-13C4] octanesulfonate (MPFOS), perfluoron-[1,2,3,4-13C4] octanoic acid (MPFOA), perfluoro-n-[1,2,3,4,513 C5] nonanoic acid (MPFNA), perfluoro-n-[1,2-13C2] decanoic acid (MPFDA), and perfluoro-n-[1,2-13C2] dodecanoic acid (MPFDoA) were from Wellington Laboratories (Ontario, Canada). Lipase Type VII (Enzyme Commission number: 3.1.1.3; EC: 232-619-9) and Protease Type XIV (Mix of different enzymes; EC: 232-909-5) were purchased from Sigma-Aldrich (Taufkirchen, Germany). Sample preparation, analysis, and quality criteria were previously described in detail by Mosch et al. (47). For analysis, an online extraction LC-MS/MS system was used and PFCs were quantified with a triple-stage quadrupole mass spectrometer (API 3200 QTRAP Applied Biosystems, Darmstadt, Germany) equipped with TurboIonSpray interface. Limits of quantification (LOQ) in serum samples were 0.4 µg/L for PFOA, PFNA, PFDoA, PFDA, and PFBS, 0.1 µg/L for PFOS, and 0.2 µg/L for PFHxS. LOQ in breast milk samples and infant formula based on a 10-fold peak-to-noise ratio for PFOS, PFOA, and PFHxS were 0.03, 0.15, and 0.02 µg/L, respectively. Statistical Analysis. We analyzed our data using SPSS software, version 13.0. Unless stated otherwise, samples below the LOQ were assigned a value of half of the LOQ. All correlations were evaluated using the Spearman rank correlation coefficient. Differences among the measurement periods were assessed using the Wilcoxon rank sum test. The mean change in blood concentrations in infants 6 months to 19 months after birth and the change from cord blood to blood of infants 6 months after birth were evaluated using the t test for paired observations. Stepwise linear multivariate regression analysis was used to evaluate the effect of concentrations in cord blood and breast milk on infant blood (dependent variable).
Results The maternal age at inclusion in the study ranged from 21 to 43 years (mean: 33 years). Before pregnancy, the subjects had body weights between 49 and 91 kg, with an average of 62 kg. Overall, 62% of mothers were primiparae, and 34% gave birth to their second or third child; 83% of the babies were delivered vaginally and 17% were delivered by caesarean section. All newborns were healthy, full-term babies, born between March and June 2008, with a mean birth weight of
TABLE 1. Overall Descriptive Statistics of PFC in Blood Samples of the Study Subjects mother pregnancy
at delivery
N (% > LOQa) mean median 95th percentile
44 (100) 3.5 3.2 6.8
38 (100) 3.5 3.2 6.1
N (% > LOQ) mean median 95th percentile
44 (100) 2.6 2.4 5.5
N (% > LOQ) mean median 95th percentile
fetus/infant cord blood
6 months after birth
19 months after birth
PFOS 47 (100) 3.2 2.9 6.3
33 (100) 1.1 1.0 2.2
40 (100) 3.3 3.0 8.1
24 (100) 2.2 1.9 4.6
38 (100) 2.3 1.9 5.2
PFOA 47 (100) 1.7 1.5 3.9
33 (100) 1.7 1.4 3.7
40 (100) 8.0 6.9 19.5
24 (100) 5.1 4.6 11.4
44 (98) 0.6 0.5 1.3
38 (97) 0.6 0.5 1.5
PFHxS 47 (96) 0.4 0.3 0.9
33 (73) 0.3 0.2 0.9
40 (98) 0.7 0.6 1.6
24 (100) 0.7 0.6 1.2
N (% > LOQ) mean median 95th percentile
44 (86) 0.8 0.6 2.8
38 (83) 0.8 0.6 3.0
47 (83) 0.7 0.5 2.0
33 (30) 0.4 LOQ) mean median 95th percentile
44 (32)
