Anthropometric and Metabolic Measurements in Longitudinal Study

At the baseline and 1-year follow-up visits, waist circumference, weight, and height were measured. The body mass index (BMI) was calculated as kg/m². Peripheral venous blood samples were obtained, at both time points, after 12 hours of fasting. Serum glucose, total cholesterol, triglycerides, and high-density lipoprotein (HDL) cholesterol were quantified by standard enzymatic methods. The low-density lipoprotein (LDL) cholesterol was calculated by the Friedewald equation.^{38 (link)} Glycated hemoglobin was determined using a chromatographic method, and 25-hydroxyvitamin D [ELISA kit (Immundiagnostik, Bensheim, Germany)] as previously described.^{39 (link)} Fecal samples were collected at baseline and 1-year time points in a sterile hermetic flask and immediately stored at − 80°C until analysis.