The presence of multiple heavy metal contaminants is of grave concern and has received considerable attention in Taiwan.1 Heavy metals such as mercury (Hg) are toxic contaminants, which can cross the placenta and affect fetal growth.2,3 Essential minerals such as iron (Fe), copper (Cu), and manganese (Mn) are both nutrients and potential toxicants, depending on the amount of exposure.4 These essential minerals are metabolized similarly to the heavy metals,5 and they are also important metallic cofactors in catalyzing redox reactions.4 The developing brain is highly sensitive to oxidative damage, so the concentrations of essential minerals play a crucial role in fetal brain development.6 Few data are available on the concentrations of both essential minerals and heavy metals in maternal/fetal medicine.7–10 Butler Walker et al7 reported levels of total Hg and methyl Hg were significantly higher in cord blood than in maternal blood (p<0.0001), whereas maternal Cu levels were significantly higher than those in cord blood (p<0.0001). The confounding factors were not analyzed except for ethnicity and smoking habits; furthermore, they did not evaluate the association of these metals with birth outcome. Rudge et al8 showed that Hg levels in cord blood were almost twice those of the mothers (n = 62), suggesting that the fetus may act as a filter for maternal Hg levels during pregnancy. Mn and Cu levels did not show statistically significant correlations between the two compartments. However, they also did not evaluate the confounding factors or association of these metals with birth outcome. A Taiwanese study (n = 308) by Lin et al9 demonstrated that cord blood lead was lower where the mother had a higher blood concentration of Mn (p = 0.02). Kopp et al10 studied the association of multiple heavy metals and trace elements between maternal and cord blood (n = 50). Hg accumulated in the fetus resulting in more than a three-fold increase in fetal exposure compared with maternal exposure. Their results also showed no association between internal exposure to any metals and maternal use of nutritional supplement during pregnancy.10
These studies7–10 demonstrated that the levels of essential minerals in healthy pregnant women were significantly different from those of the general population. There is a need to evaluate more pregnant women, especially from different races, for the purpose of establishing specific normative levels of essential minerals. Although Taiwan is a high-fish-consuming island country, data on the accumulation of Hg in pregnant woman remain limited.3 Furthermore, there is no Taiwanese information regarding Hg and essential minerals in both mother and fetus. The purposes of this study in south Taiwan were to (1) assess the correlation of the Hg, Mn, Fe, and Cu levels in paired maternal/fetal blood samples, and (2) study potential confounding factors such as socioeconomic factors, smoking, vitamin intake, and seafood consumption during pregnancy.
2.1. Studying individuals and sampling
This study was a prospective cross-sectional study. Women were considered eligible if they were 20–45 years old and pregnant with a term singleton fetus. Women with chronic medical conditions or infectious diseases, or those who reported illicit drug use were excluded. A total of 150 women consented to participate and initially met eligibility requirements. Two women withdrew because of twin fetuses, one withdrew because of stillbirth, and two others withdrew consent prior to being discharged from the hospital.
A total of 145 healthy pregnant women with a mean age of 28.06 years were recruited at the Department of Obstetrics and Gynecology of Fooyin University Hospital in Tong Gang, Taiwan, from September 30, 2010 to May 25, 2011. The city of Tong Gang is a major seaside fishing area in southern Taiwan. The participants received a detailed explanation of the study procedures before consenting to participate. The research protocol was approved by the Institutional Review Board of Fooyin University (FYH-IRB-099-04-02-A), and written informed consent was obtained from all participants. Information on the gestational age, prenatal examination, and characteristics of the birth and newborn, were obtained from the medical records. We used interviewer-administered questionnaires to collect information on demographic characteristics, smoking habits, alcohol drinking habits, betel nut chewing habits, use of Chinese medicine, seafood consumption, nutritional supplement (vitamin), and degree of education. The degree of seafood consumption was defined as the sum scores of seafood consumption items in the questionnaire. A total of nine items were about the amount of fresh fish and seafood consumption of each participant during pregnancy. High sum scores revealed more seafood consumption. The cutoff score of high and low seafood consumption was defined as the median value of the sum scores in the questionnaire. The participants were asked to record their nutritional supplement of vitamin consumption during pregnancy in the questionnaire. Anthropometric measurement of newborns was made by delivery room staff, using standard anthropometric procedures.
2.2. Blood sampling and sample analysis
Umbilical cord blood and maternal venous whole blood samples were collected into 9 mL standard laboratory issued EDTA tubes, separately. All samples were processed within 2 hours of delivery and stored at –80°C. Samples were sent to the Department of Biomedical Engineering and Environmental Sciences, Ultra Trace Micro-Analysis Laboratory at National Tsing Hua University, and were analyzed by a inductively coupled plasma-mass spectrometer (7500ce; Agilent, Tokyo, Japan). The limit of detection was 0.353 μg/L (ppb) for Hg, 0.125 μg/L (ppb) for Mn, 0.061 μg/mL (ppm) for Fe, and 0.00066 μg/mL (ppm) for Cu.11
2.3. Statistical analysis
We used Pearson's correlation coefficient to reveal the association between pairs of the following variables: heavy metal concentrations (Hg, Mn, Fe, and Cu) in maternal blood, heavy metal concentrations (Hg, Mn, Fe, and Cu) in cord blood, vitamin, and seafood consumption. Log-transformed data of heavy metals were used to reveal the association between maternal and fetal pairs.
Multivariate logistic regression was used to reveal the association of heavy metal (Hg, Mn, Fe, and Cu) concentrations in maternal and cord blood by controlling maternal age, race, smoking, vitamin use, and seafood consumption. Unadjusted odds ratios and fully adjusted odds ratios were reported. The significance of p value was set at 0.05. All analyses were performed using SPSS software (version 20; SPSS Institute Inc., Chicago, IL, USA).
A total of 150 women consented to participate and initially met eligibility requirements. Two women withdrew because of twin fetuses, one withdrew because of stillbirth, and two others withdrew consent prior to being discharged from the hospital. A total of 145 healthy pregnant women with a mean age of 28.06 years were recruited at the Department of Obstetrics and Gynecology of Fooyin University Hospital in Tong Gang, Taiwan, from September 30, 2010 to May 25, 2011.
Specific characteristics of the paired pregnant women and their neonates are shown in Table 1. The mean age of pregnant women in this study was 28.06±5.18 years. Of the study participants, 72.41% were Taiwanese and approximately 48.97% were high school graduates. Prepregnant and perinatal body mass indexes were 22.04±3.33% and 27.82±3.86%, respectively. The percentages of smoking, alcohol drinking, and areca nut chewing during pregnancy were 8.97%, 7.59%, and 6.9%, respectively. A minority of the participants (9.66%) used Chinese medicine prenatally. Forty pregnant women (27.6%) used prenatal vitamin >3 times/wk. There were 71 female and 74 male neonates. Mean birth weight, length, and head circumference were 3081.59±387.31 g, 49.61±2.49 cm, and 34.3±1.4 cm respectively. A large majority of the neonates (74.48%) were full term, and only 10 neonates (6.9%) had a low birth weight (<2500 g).
Table 2 shows data on metals in paired maternal and umbilical cord blood. Median maternal and fetal Hg levels were 2.24 μg/L and 2.3 μg/L, respectively. There was a significant association between maternal and fetal Hg (Fig. 1A, r = 0.78, p<0.001). Median maternal and fetal Mn levels were 44.96 μg/L and 61.68 μg/L, respectively, and there was a significant association between maternal and fetal Mn (Fig. 1B, r = 0.31, p<0.001). Median maternal and fetal Fe levels were 288.2 mg/L and 449.4 mg/L, respectively; there was a significant association between maternal and fetal Fe (Fig. 1C, r = 0.17, p = 0.038). Median maternal and fetal Cu levels were 1.47 mg/L and 0.72 mg/L, respectively, and there was a significant association between maternal and fetal Cu (Fig. 1D, r = 0.21, p = 0.01).
However, metal levels observed in this series differed from those observed in previous studies of mother–infant cohorts (Table 3).3,7–10,12–17
Tables 4–7 show multiple logistic analysis of metals with the potential confounding factors. Maternal and fetal metal levels were not associated with maternal age except for maternal Cu level. There was a borderline significant association between maternal age and maternal Cu level [adjusted odds ratio (AOR) 0.936, p = 0.057]. Maternal and fetal metal levels were not associated with race and smoking. High seafood consumption is associated with lower maternal Fe (AOR 0.404, p = 0.017) and Cu (AOR 0.434, p = 0.034). Prenatal vitamin use (>3 times/wk) was significantly associated with lower maternal Hg (AOR 0.272, p = 0.005) and lower maternal Cu (AOR 0.267, p = 0.004) levels.
In this series, we found a positive correlation of Hg, Mn, Fe, and Cu in maternal and umbilical cord blood of paired mother/child samples. These data may contribute to establishing reference levels in pregnant women, and studying the role and mode of action of environmental metals in both mother and fetus. Further analysis of confounding factors showed that prenatal vitamin use decreased the maternal levels of Hg and Cu. To the best of our knowledge, this is the first report about the effect of prenatal vitamin use on maternal metals.
The Hg concentrations found in Taiwanese studies, including our investigation, are higher than those found in foreign studies.7,10,12 The dietary habit during pregnancy may partially account for the difference between our series and foreign studies. Fish consumption during pregnancy is generally higher in Taiwan than in other countries because of the traditional idea of eating fish to provide improved nutrition for the fetus.3 Fish consumption can be a major source of Hg during pregnancy. A Taiwanese study by Chien et al18 showed that 21.6–24.3% and 45.6–57.4% of the daily Hg dose estimates exceeded the reference doses for typical and high seafood consumers, respectively. Their analysis suggested that the acceptable ingestion rate of fish for women during childbearing is 90.8±15.7 g/d. Although there was no significant association of Hg level and birth outcome in this series, Hg has been reported to be associated with developmental delay in children whose mothers were exposed to it during pregnancy.19,20 Pregnant woman should be educated about the risk of high Hg level associated with overingestion of specific types and quantities of fish to help protect their children's health.
There was a positive correlation between umbilical cord blood levels and maternal concentrations in terms of Fe, Cu, and Mn. High seafood consumption was associated with lower maternal Fe and Cu levels in multiple logistic regression. This relationship requires detailed ingredients of seafood to elucidate the mechanism. However, the distribution of three essential minerals between maternal and umbilical cord blood was different. Median fetal Mn level (61.68 μg/L) was 40% higher than maternal Mn level (44.96 μg/L), whereas median fetal Fe level (449.40 μg/L) was 60% higher than maternal Fe level (288.20 μg/L). In the study by Kopp et al,10 median fetal Fe level (635.8 mg/L) was 20% higher than maternal Fe level (530.5 mg/L), but median fetal Mn level (28.8 μg/L) was 70% higher than maternal Mn level (17.0 μg/L). Median maternal Fe level was much lower, and median maternal Mn level was far higher in this series than in Kopp et al.'s10 study. Women with low Fe stores absorbed about 5% of dietary Mn, but women with normal Fe stores absorbed only about 1% of dietary Mn.21 The effect of Fe deficiency on Mn absorption is apparently due to the ability of the divalent metal transporter 1.22 Therefore, Fe deficiency, particularly among women of reproductive age, is a potential risk factor for Mn toxicity when intestinal Mn exposure is high. The low Fe status may partially account for the higher maternal Mn level in this series than in other studies in America and Europe (Table 3). Overall, the interaction of Fe and Mn in fetus is somewhat complicated. The fetus requires increased amounts of Fe for high oxygen and energy consumption. However, Fe is also capable of generating harmful reactive oxygen species via Fenton chemistry.23 Mn superoxide dismutase is a mitochondrial enzyme, which can selectively decrease oxidative damage without affecting Fe-mediated oxygen transport and energy production.24 In addition, Chen et al25 observed a dramatic decrease of reactive oxygen species as a consequence of upregulation of Mn-dependent superoxide dismutase and catalase during osteogenic differentiation of human mesenchymal stem cells. The biological roles of fetal Fe and Mn can be further elucidated by the abovementioned mechanism. The simultaneous increase of fetal Mn and Fe levels observed in this series is beneficial rather than detrimental. Further studies are required to study the potentially high Mn exposure and its relationship with Fe absorption in Taiwan.
In contrast, median Cu level (0.73 mg/L) in our study was approximately 50% lower than that in maternal blood (1.47 mg/L). Our result is consistent with several previously published studies,8,10,26 which showed a 50–60% decrease in Cu level in the fetus. The observed decrease in fetal Cu, a major metallic cofactor in a variety of oxidoreductases, may reduce the potential of cellular oxidative damage in the developing fetus.27 However, Cu is an essential mineral, and its deficiency can result in many nutritional and vascular disorders.28 Maintaining an adequate amount of Cu in the human body is important, especially for the newborns who are dependent on stored Cu.
Prenatal vitamin use significantly decreased the maternal levels of Hg and Cu in this series. From animal studies, some data on the effect of vitamin E on heavy metals are available.29–31 Al-attar's29 study suggested that the administration of vitamin E protects against heavy metal-induced renal and testicular oxidative stress and injuries in male mice. Another Al-attar's30 study showed that vitamin E protects against the heavy metal-induced liver injury in albino mice, and the attenuating effect of vitamin E may be due to its antioxidant activity. Abd El-Aziz et al31 reported that vitamin E may ameliorate some aspects of methyl Hg developmental toxicity in rat fetuses. Kim et al32 also found a negative association between serum folate and blood Hg concentrations in pregnant Korean women. Their findings suggest that folate is associated with the blood Hg level by participating in the Hg detoxification process.32 It is still unclear by which mechanism prenatal vitamin use can reduce maternal levels of Hg and Cu. The effect of vitamin use on heavy metals in pregnant woman still requires further investigation.
Several limitations in this study need to be specified. This study was a single-center investigation of pregnant woman with a modest sample size. In addition, this is a cross-sectional study without level change of Hg and essential minerals during the entire pregnancy. Therefore, it was a limitation of the representative in terms of metal levels based on the collection time by delivery. Additionally, the data regarding potential exposure sources for heavy metals and essential minerals were not complete. The collection of maternal vitamin use is not detailed in the ingredients. In addition, assessment of seafood consumption was not precise, and Fe deficiency of the participants was not evaluated. Therefore, multicenter studies with a large sample size and precise assessment of seafood consumption are suggested for future investigations.
In conclusion, there was a positive correlation of Hg, Fe, Cu, and Mn in maternal and umbilical cord blood of paired mother/child samples in this series. However, the distribution of Hg and three essential minerals between maternal and umbilical cord blood was different. Median Hg, Mn, and Fe levels were higher in cord blood than in maternal blood, while median Cu level was lower in cord blood than in maternal blood. The low Fe status in pregnant women may partially account for the higher maternal Mn level. Our findings raise the possibility of reducing maternal Hg and Cu levels via prenatal vitamin supplementation, although the effect of vitamin use on heavy metals during pregnancy requires further study.
Part of this research was supported by the Kaohsiung Veterans General Hospital (VGHKS103-81, VGHKS103-82, VGHUST104-G7-7-3, VGHKS105-106, and VGHUST105-G3-1-3) and Kaohsiung Medical University (MOST104-2314-B-037-012-MY2, and MOST105-2632-B-037-002). We thank Huei-Han Liou in Kaohsiung Veterans General Hospital for the statistical analysis.
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