Those women with clinical diagnosis of diabetes mellitus, who used vitamin supplements, with presence of renal, liver, or heart failure, and who were pregnant and lactating were excluded. General information about age, smoking habit, socioeconomic status, family history, medical history, Panobinostat solubility dmso and the use of supplements and drugs were obtained through a questionnaire developed by the researchers. The usual dietary intakes of folic acid, cobalamin, pyridoxine, cholesterol, fiber, alcohol, and coffee were assessed through the FFQ (which contains 81 food items) [14] because these can influence Hcy levels in accordance

to scientific literature. In the prefortification group, food not fortified with folic acid was not considered in the analysis of the FFQ, whereas in the postfortification group, fortified food with folic acid was considered. Nutrient analysis was performed using the program “The Food Processor” (Esha Research, Salem, Mass, USA) [16], adapted to the Brazilian reality. The evaluation of the

folic acid content in the prefortification group, selected in the program Food Processor food, was not fortified with this vitamin. For the postfortification group, we used food fortified with folic acid. Body weight (kilograms) and height (meters) were measured using the Filizola platform scale and a Filizola vertical stadiometer, respectively [17]. Body mass index was calculated as weight divided by square height (kg/m2) [15]. Waist circumference was measured at the midpoint between the last rib and Apoptosis Compound Library the iliac crest using an inelastic metric tape [18]. The pressure levels were measured with a mercury sphygmomanometer [19]. Blood samples were drawn after a 12-hour overnight fast and were placed into tubes that did not contain anticoagulant or with ethylene diamine tetra-acetic acid (Vacutainer, Becton Dickinson, NJ, USA). Aliquots of serum and plasma samples were obtained by centrifugation at 4000 rpm for 15 minutes (Centrifuge Excelsa Baby I; Fanem, São Paulo, Brazil) and stored at −20°C until analysis. The

analyses of the prefortification group were performed at the end of the blood draw from all participants in 2003, and the analyses of the postfortification group were performed at the end of the blood draw of all participants in 2009. Serum concentrations of glucose [20], triglycerides Sunitinib concentration [21], high-density lipoprotein cholesterol (HDL-C) [22], and total cholesterol [23] were determined by enzymatic method, according to the manufacturer’s instructions (CELM and KATAL kits; Katal Biotechnologica, Ind, Com, Ltda, Minas Gerais, Brazil, and CELM-Cia; Equipadora de Laboratories, Moderneros-Sao Paulo, Brazil). The following values were considered normal as indicated by the manufacturers: glucose less than 100 mg/dL, triglycerides less than 150 mg/dL, HDL-C greater than 50 mg/dL, and total cholesterol less than 200 mg/dL. Low-density lipoprotein cholesterol (LDL-C) was calculated [24], the ideal reference value being less than 100 mg/dL.