Ayotte et al. to inhibit enzymes, in vitro experiments have been conducted to investigate the inhibitory effects of toxic metals on PON1 activity. Gonzalvo et al. (1997) first reported that cop- per and mercurials were potent inhibitors of PON1 activity in human liver microsomes. Similar results were obtained in other in vitro experiments conducted with pooled human serum of subjects with the PON1Q192 geno- type; manganese (Mn2+), cobalt (Co2+), cad- mium (Cd2+), and nickel (Ni2+) also inhibited PON1 activity in these experiments (Debord et al. 2003). Subsequent studies have reported that metals were more effective inhibi- tors of the PON1R192 isozyme than of the PON1Q192 isozyme (Cole et al. 2002; Costa et al. 2005; Gençer and Arslan 2009). Despite these compelling results from in vitro experi- ments, treatment of mice with Cd, MeHg, or dietary iron, leading to metal serum concen- trations > 1 Mμ, did not alter serum or liver PON1 activity (Cole et al. 2002). To our knowledge, the association between MeHg exposure and PON1 activity has yet to be tested in humans and may prove to be of pub- lic health relevance in Inuit who are highly exposed to MeHg through fish and marine mammal consumption (Fontaine et al. 2008). This population is also exposed to lead, a metal whose blood levels were significantly associated with decreased serum PON1 activ- ity in lead workers (Li et al. 2006). We conducted a comprehensive health survey in the Inuit population of Nunavik (Québec, Canada) during the fall of 2004. In the course of this study, we investigated the relation between blood mercury concen- trations and plasma PON1 activities in 896 Inuit adults living in Nunavik, while taking into account the potential protective role of selenium, which has been shown to coun- teract the toxicity of mercurials (Khan and Wang 2009). We also investigated the pos- sible confounding or modifying role of several factors, including PON1 gene variants. Materials and Methods Study population and data collection. This health survey was conducted in Nunavik, a northern region of Québec where approxi- mately 9,500 Inuit live in 14 communities along the coasts of Hudson Bay, Hudson Strait, and Ungava Bay. Informed consent was obtained from all participants before enroll- ing them in the study, which was approved by the Comité d’éthique de la recherche de l’Université Laval and the Comité d’éthique de santé publique du Québec. The target popula- tion of this study was permanent Inuit residents of Nunavik from 18 to 74 years of age. To obtain a standard representation of the target population, the study used a stratified random sampling of private Inuit households, with the community being the stratification variable. 1078 Several self-administered and interviewer- completed questionnaires were used to obtain information regarding demographics, lifestyle habits, nutrition, and health indicators. In addition, individuals were asked to participate in a clinical session where blood samples were taken and anthropometric measurements were performed. Data on body composition (body fat percentage, lean body mass) were obtained by bioelectrical impedance using the Tanita TBF-300A analyzer (Tanita Corporation of America, Inc., Arlington Heights, IL, USA). Medical history and related drug consump- tion of each participant were documented by medical chart review. Among the 677 contacted households, 521 agreed to participate (household response rate of 77.8%), 1,056 individuals signed a consent form, and 917 people agreed to the collection of blood samples for clinical and toxicologi- cal analyses (final participation rate of 67%). Because of various problems (insufficient plasma volume, broken vials), we measured PON1 activity for 899 individuals. Three participants were excluded from the database because of extreme values of blood mercury (> 800 nmol/L) and blood selenium (> 30 mμol/L). Data on the consumption of traditional foods were obtained from a food frequency questionnaire, which was designed to measure season-specific consumption of food items derived from fishing and hunting during the year before the survey. Season-specific intakes were computed by multiplying the daily con- sumption frequency by the portion size for each food item and season. Season-specific intakes were then averaged over the four sea- sons to yield the mean annual consumption (grams per day). The following three compos- ite variables were created: a) wild fish com- prised clam, mussel, oyster, scallop, seaweed, urchin, arctic char, cod, whitefish, trout, salmon, dried fish, and other fish such as pike, cisco, and walleye; b) marine mammal fat comprised beluga and seal fat; and c) marine mammal meat comprised beluga meat, dried beluga, beluga muktuk (skin), beluga liver, other parts of beluga, seal meat, walrus meat, igunak (aged meat), seal kidney, seal liver, other parts of seal, and walrus parts. Plasma PON1 activity. Blood was collected from a cubital vein in a 6-mL plastic Vacutainer containing sodium heparin as the anticoagu- lant (BD Medical, Mississauga, ON, Canada). Blood was centrifuged within 2 hr of collection, and the plasma was stored frozen at –80°C until time of analysis. Although in most previ- ous studies PON1 activity has been measured in serum samples, the suitability of heparin- plasma samples was demonstrated by Ferré et al. (2005). PON1 activity was measured in 5 Lμ plasma, diluted to a final concentration of 2%, using the EnzChek Paraoxonase Assay Kit (Molecular Probes, Invitrogen, Carlsbad, CA, USA). This highly sensitive, homogeneous fluorometric assay for the organophosphatase activity of PON1 is based on the hydrolysis of a fluorogenic organophosphate analog (7-diethyl- phospho-6,8-difluor-methylumbelliferyl). The intensity of emitted light was measured with a Victor2 plate reader (PerkinElmer–Cetus Life Sciences, Boston, MA, USA) using 355- and 460-nm filters for excitation and emission, respectively. Enzymatic activity was calcu- lated by converting fluorescence intensity into activity using a standard curve. Activity was expressed as kilounits per liter, where one unit of PON1 is defined as the amount of enzyme that produces 1 nmol fluorescent product per minute at 37°C. Aliquots of a plasma sample obtained from a volunteer were stored frozen at –80°C along with the study samples; this “control” sample was analyzed in each analyti- cal batch. The mean value was 13.3 kU/L, and the between-day coefficient of variation was 7.6% (n = 23). Metal analyses. Mercury, selenium, and lead concentrations were determined in whole- blood samples collected from a cubital vein in a 6-mL plastic Vacutainer containing potassium EDTA as the anticoagulant (BD Medical). Blood samples were stored frozen at –80°C until time of analysis. Determination was per- formed by inductively coupled plasma mass spectrometry on an ELAN DRC II instrument for mercury and selenium and an Elan 6000 instrument for lead (Perkin-Elmer SCIEX, Concord, ON, Canada). Blood samples were diluted 20-fold in an ammonia solution before analysis. Limits of detection for mercury, lead, and selenium were 0.5 nmol/L, 1 nmol/L and 0.1 μmol/L, respectively, and each run of samples included a standard. Between-day coefficients of variation for mercury, lead, and selenium measurements were 2.1%, 2.8%, and 6.1%, respectively. Analyses were per- formed by the Laboratoire de Toxicologie of the Institut National de Santé Publique du Québec, which has ISO 17025 accredita- tion and participates in the quality assurance/ quality control programs of the Canadian Northern Contaminants Program and the Arctic Monitoring Assessment Program. Plasma lipids. Concentrations of total cho- lesterol and triacylglycerol in plasma samples were determined by enzymatic methods using a Hitachi 917 autoanalyzer and reagents, both from Roche Diagnostics (Laval, QC, Canada). HDL cholesterol (HDL-C) was measured directly by selectively inhibiting reaction with other lipoproteins. LDL cholesterol (LDL-C) was calculated using the Friedewald formula [LDL-C (grams per liter) = total cholesterol (grams per liter) – HDL-C (grams per liter) – triacylglycerol (grams per liter)/5] (Tremblay et al. 2004). Lipid analyses were performed at the Centre de Recherche sur les Maladies Lipidiques (Centre de Recherche du Centre ovlume 119 | number 8 | August 2011 • Environmental Health Perspectives