After obtaining information on the study and signing an informed consent, participants completed a detailed and validated Danish questionnaire with the assistance of a qualified interviewer during a face-to-face interview. The questionnaire was adapted from the Mediterranean Osteoporosis Study Questionnaire (MEDOS) [27 (link)] and allowed to document the following risk factors for osteoporosis: smoking habits, physical activity, daily milk products and calcium supplement consumption, current use of HRT and former contraceptive use. Questions were also asked to document secondary causes of osteoporosis such as Cushing's disease, renal and liver deficiency, hyper- and hypothyroidism, bone cancer and rheumatoid arthritis.
Weight, height and waist, abdominal and hip girth were measured using standardized techniques. Women were considered postmenopausal if they had no menses for at least one year before the recruitment, if they underwent bilateral oophorectomy more than 6 months ago and if analysis had revealed a follicle stimulating hormone concentration greater than 40 IU/L.
Blood samples (10 ml) were collected in vials containing EDTA as the anticoagulant, centrifuged (10 min, 3000 rpm) and the plasma poured into glass vials pre-rinsed with hexane. First morning urine samples were collected in plastic vials. Plasma and urine samples were stored frozen at -80°C until time of analysis for OCs and cadmium respectively, at the Centre de Toxicologie (CTQ) of the Institut National de Santé Publique du Québec (Québec, Canada). This laboratory is accredited by the Canadian Association for Environmental Analytical Laboratories.
Fourteen PCB congeners [International Union for Pure and Applied Chemistry (IUPAC) no. 28, 52, 99, 101, 105, 118, 128, 138, 153, 156, 170, 180, 183, 187], p, p'-DDT (dichlorodiphenyltrichloroethane) and its major metabolite p, p'-DDE (p, p'-dichlorodiphenyl-dichloroethylene), hexachlorobenzene (HCB) and 8 other chlorinated pesticides [α-chlordane, γ-chlordane, aldrin, β-hexachlorocyclohexane (β-HCH), mirex, oxychlordane, cis-nonachlor, and trans-nonachlor] were analysed by high-resolution gas chromatography with electron capture detection. A 1:1:3 mixture of ammonium sulfate:ethanol:hexane was first added to the plasma to extract OCs. The extracts were then concentrated and purified on two Florosil columns (60–100 mesh; Fisher Scientific, Nepean, Ontario, Canada). The OCs were measured in the purified extracts with an HP 5890 high-resolution gas chromatograph equipped with dual-capillary columns (HP Ultra I and Ultra II) and dual Ni-63 electron capture detectors (Hewlett-Packard, Palo Alto, CA, USA). The limit of detection was based on 3 times the average standard deviation of noise and was 0.08 μg/L for p, p'-DDE, p, p'-DDT and β-HCH, and 0.04 μg/L for all other compounds. The average percent recoveries were greater than 95%. Coefficients of variations (CVs) based on repeated analyses of a standard reference material (SRM 1589; N = 15) ranged from 3.9% to 18.5%, except for PCB 105 and p, p'-DDT, for which CVs were 31.6% and 26.2%, respectively.
Plasma OC concentrations were expressed on a lipid basis. We calculated the total plasma lipid concentration from the concentrations of cholesterol esters, free cholesterol, triglycerides and phospholipids [28 (link)], which were measured using standard enzymatic procedures.
Urine samples were analysed for cadmium by graphite furnace atomic absorption spectrometry and concentrations were corrected for urinary creatinine content to take into account differences in urinary output between participants. Duplicates were run every 10 samples. The CV of the method based on repeated analyses of a standard reference material from CTQ interlaboratory comparison program (N = 34) was 2.0%, and the limit of detection was 0.2 μg/L.
Weight, height and waist, abdominal and hip girth were measured using standardized techniques. Women were considered postmenopausal if they had no menses for at least one year before the recruitment, if they underwent bilateral oophorectomy more than 6 months ago and if analysis had revealed a follicle stimulating hormone concentration greater than 40 IU/L.
Blood samples (10 ml) were collected in vials containing EDTA as the anticoagulant, centrifuged (10 min, 3000 rpm) and the plasma poured into glass vials pre-rinsed with hexane. First morning urine samples were collected in plastic vials. Plasma and urine samples were stored frozen at -80°C until time of analysis for OCs and cadmium respectively, at the Centre de Toxicologie (CTQ) of the Institut National de Santé Publique du Québec (Québec, Canada). This laboratory is accredited by the Canadian Association for Environmental Analytical Laboratories.
Fourteen PCB congeners [International Union for Pure and Applied Chemistry (IUPAC) no. 28, 52, 99, 101, 105, 118, 128, 138, 153, 156, 170, 180, 183, 187], p, p'-DDT (dichlorodiphenyltrichloroethane) and its major metabolite p, p'-DDE (p, p'-dichlorodiphenyl-dichloroethylene), hexachlorobenzene (HCB) and 8 other chlorinated pesticides [α-chlordane, γ-chlordane, aldrin, β-hexachlorocyclohexane (β-HCH), mirex, oxychlordane, cis-nonachlor, and trans-nonachlor] were analysed by high-resolution gas chromatography with electron capture detection. A 1:1:3 mixture of ammonium sulfate:ethanol:hexane was first added to the plasma to extract OCs. The extracts were then concentrated and purified on two Florosil columns (60–100 mesh; Fisher Scientific, Nepean, Ontario, Canada). The OCs were measured in the purified extracts with an HP 5890 high-resolution gas chromatograph equipped with dual-capillary columns (HP Ultra I and Ultra II) and dual Ni-63 electron capture detectors (Hewlett-Packard, Palo Alto, CA, USA). The limit of detection was based on 3 times the average standard deviation of noise and was 0.08 μg/L for p, p'-DDE, p, p'-DDT and β-HCH, and 0.04 μg/L for all other compounds. The average percent recoveries were greater than 95%. Coefficients of variations (CVs) based on repeated analyses of a standard reference material (SRM 1589; N = 15) ranged from 3.9% to 18.5%, except for PCB 105 and p, p'-DDT, for which CVs were 31.6% and 26.2%, respectively.
Plasma OC concentrations were expressed on a lipid basis. We calculated the total plasma lipid concentration from the concentrations of cholesterol esters, free cholesterol, triglycerides and phospholipids [28 (link)], which were measured using standard enzymatic procedures.
Urine samples were analysed for cadmium by graphite furnace atomic absorption spectrometry and concentrations were corrected for urinary creatinine content to take into account differences in urinary output between participants. Duplicates were run every 10 samples. The CV of the method based on repeated analyses of a standard reference material from CTQ interlaboratory comparison program (N = 34) was 2.0%, and the limit of detection was 0.2 μg/L.
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