Broberg et al. water they consumed (tap water/bottled water and source of the tap water), their dietary habits (what do you eat for breakfast, lunch, and dinner?), diseases, and time of residence in the area, including that of their parents and grandparents. These questions revealed that all women almost exclusively drank tap water, their diets (mainly corn, beans, chicken, and pork) were very similar, and only 3 of 202 women reported ongoing use of any medica- tion: One was being treated for gastritis and two for high blood pressure. Their time of residence in the area and other characteristics are presented in Table 1. Height and weight were measured using standardized protocols, and the women were asked to donate a urine sample (~ 20 mL) and blood samples (4 mL for separation of plasma). All the samples were collected at the hospital or the local health clinics during the daytime, and study participants were allowed to eat and drink before sampling. All the volunteers gave oral and written consent. The study was approved by the Ministry of Health in Salta, Argentina, and the ethical committee at Karolinska Institute (Sweden). Sample collection. Spot-urine samples were collected in disposable paper cups and immediately transferred to 20-mL acid-washed polyethylene bottles. After testing for pH and presence of protein and glucose by urine test strips (Siemens Multistix, Tarrytown, NY, USA), the urine samples were frozen at –20°C. Markers of thyroid function were measured in plasma, which was obtained from collected venous blood samples (heparin tubes) that were centrifuged within 10 min after blood withdrawal. Plasma and urine samples were kept at –20°C until they were transported (with cooling blocks) to Sweden for analysis. Analysis of elements and markers of thy- roid function. We evaluated arsenic, boron, and cesium in urine because our previous study had demonstrated that these elements were elevated in drinking water in the same regions as lithium (Concha et al. 2010). Of note, cesium is known to accumulate in the thyroid (Bandazhevsky 2003). We also evalu- ated selenium and iodine because these ele- ments are important for thyroid function [Lu and Holmgren 2009; World Health Organization (WHO) et al. 2007]. The uri- nary samples were analyzed 3 months after sampling using inductively coupled plasma mass spectrometry with collision/reaction cell (Agilent 7500ce; Agilent Technologies, Waldbronn, Germany) as described previously (Concha et al. 2010). Iodine was measured after diluting the urine with 0.1% NH4OH (Suprapur; Merck, Darmstadt, Germany). Analytical accuracy was ascertained by analyz- ing commercially available urine reference materials (Concha et al. 2010). For iodine, we analyzed Seronorm Urine No. 2525 (refer- ence value, 282 gμ/L) and OK4636 (analyti - cal value, 139 μg/L; SERO AS, Billingstad, Norway) and obtained measured values of 294 ± 7.5 and 133 ± 4.4 μg/L ( n = 11), respectively. To compensate for dilution, we Table 1. Characteristics, measured concentrations of elements and hormones in urine and plasma, and correlations between the variables and T4 and TSH (n = 202).a aAll values were based on 202 observations with the exception of years living in the area (n = 198), parity (n = 197), plasma T4 (n = 194), and TSH (n = 196), because of either missing information (years living in the area and parity) or insufficient amount of sample left for analysis (plasma T4 and TSH). *p < 0.05, **p < 0.01, and #p < 0.001, for the Spearman’s correla- tions between the variables and T4 and TSH. Table 2. Association between a 1,000-μg/L increase in urine lithium concentration and markers of thyroid function and BMI by linear regression Mean 37.2 24.7 25.6 4.3 4,404 15,000 244 473 28 147 17 2.2 Median 34 25 24.7 3 3,914 14,578 230 470 25 125 17 1.9 Variable Age (years) BMI (kg/m2) Parity (n) Lithium Boron Arsenic Cesium Selenium Iodine Years living in the area Urinary concentration (μg/L) Plasma concentration T4 (pmol/L) TSH (mIU/L) analysis.a Outcome T 4 TSH BMI Unadjusted Adjustedb Unadjusted Adjustedc 828 β-Coefficient –0.19 –0.17 0.096 0.089 95% CI –0.31 to –0.068 –0.32 to –0.015 0.033 to 0.16 0.024 to 0.15 p-Value 0.002 0.032 0.003 0.007 Unadjusted 0.18 –0.039 to 0.40 0.11 Adjustedd 0.14 –0.059 to 0.35 0.16 aAdjusted models include covariates with p < 0.25 in a univariate analysis, apart from boron, which was highly correlated to lithium (rs = 0.85; p = 0.001). bAdjusted for urinary arsenic, cesium, and selenium; age; parity; and BMI. cAdjusted for selenium and BMI. dAdjusted for urinary iodine, age, and parity. 5th, 95th percentiles 18, 64 3, 51 18.8, 35 0, 11 268, 10,392 2,602, 27,907 21, 545 26, 880 11, 51 37, 354 13, 21 0.68, 4.9 ) L / lo 25 m p ( 4 T a sm 20 a l P 35 30 15 10 ovlume Minimum–maximum 12–80 0.3–71 16–40 0–14 117–14,343 1,292–45,313 10–1,250 8.0–2,243 4.0–74 16–857 11–32 0–8.8 Spearman rank (rs) correlation T TSH 4 –0.19** –0.048 –0.096 –0.012 –0.24** 0.063 –0.28# –0.064 –0.25** –0.024** –0.20** –0.27# 0.23** 0.035 — –0.11 0.10 0.025 0.002 –0.005 –0.18* –0.063 –0.11 — 0 2,000 4,000 6,000 8,000 10,000 12,000 14,000 Lithium (µg/L) Figure 1. Association between urinary lithium concentration and plasma free T4. 119 | number 6 | June 2011 • Environmental Health Perspectives