Msd 5973 system with capillary column

Plasma glucose concentration was determined using the glucose oxidase method (Yellow Spring Instruments Co.). Plasma insulin concentrations were measured by using radioimmunoassay kits (Linco Research, St Louis, MO). Plasma proteins were precipitated with ice-cold acetone, and hexane was used to extract plasma lipids. The aqueous phase, containing glucose, was dried by speed-vac centrifugation (Savant Instruments, Farmingdale, NY). The plasma glucose tracer-to-tracee ratio (TTR) was determined by gas-chromatography/mass-spectrometry (MSD 5973 system with capillary column; Hewlett-Packard, Palo Alto, CA) after heptafluorobutyric (HFB) anhydride was used to form an HFB derivative of glucose. Glucose TTR were determined by selectively monitoring ions at mass-to-charge ratios (m/z) 519 and 521 (Korenblat et al., 2008 (link)). Hepatic insulin sensitivity was calculated as the inverse of the product of plasma insulin concentration and the endogenous glucose rate of appearance (Ra) into the systemic circulation, determined by dividing the glucose tracer infusion rate by the average plasma glucose TTR (Korenblat et al., 2008 (link)). The homeostasis model assessment of insulin resistance (HOMA-IR) was calculated by dividing the product of the plasma concentrations of insulin (in μU/mL) and glucose (in mmol/L) by 22.5 (Matthews et al., 1985 (link)).