Sbd f

2 ml blood from the antecubital vein were immediately placed on ice and separated into plasma and cells within 30 min, then stored at – 80 °C until analysis. Plasma glucose measurements were performed by glucose oxidase method on the Cobas 8000 Modular Analyzer Series (Roche, Mannheim). Fasting plasma insulin (FPI) was determined by enzyme-linked immunosorbent assay (ELISA) with the Insulin ELISA 10-1113-10 kit (Mercodia, Sweden, with intraassay CV of 5.91% and between-assay CV of 7.15%). The absorbance was measured at 450 nm using the Automatic Microplate Reader (Syngene, BioTek, USA). Homeostasis model of insulin resistance (HOMA-IR) was used to evaluate insulin resistance. HOMA-IR was calculated as fasting glucose × fasting insulin/22.5. HOMA-beta cell function (HOMA-f) was calculated as 20 × fasting insulin/(fasting glucose − 3.5).
Plasma total homocysteine (tHcy: the sum of all Hcy forms) was measured by high-performance liquid chromatography (HPLC, Waters 1525, USA) with fluorescence detection, applying 7-fluorobenzo-2oxa-1,3-diazole-4-sulfonate (SBD-F, Sigma, USA) as derivating agent and tris(2-carboxyethyl)phosphine (TCEP, Sigma, USA) as reducing agent^{27 (link)}. The intraassay CV was 3.60%, and between-assay CV was 4.49%.