2.3. Analysis of Plasma Lipids and Metabolites

Nuclear magnetic resonance (NMR) spectroscopy experiments quantitating plasma lipids and metabolites were as previously described [7,21]. In brief, we used a 14.0 T Bruker magnet equipped with a Bruker AV-III console operating at 600.13 MHz. Experiment conditions included: sample temperature of 310 K, 96 k data points, 30 ppm sweep width, a recycle delay of 4 s, a mixing time of 150 ms, and 32 scans. All spectra were acquired in 3 mm NMR tubes using a Bruker 5 mm QCI cryogenically cooled NMR probe. Plasma samples were prepared and analyzed according to the Bruker In-Vitro Diagnostics research (IVDr) protocol. Sample preparation consisted of combining 50 uL of plasma with 150 uL of the buffer supplied by Bruker Biospin specifically for the IVDr protocol. For 1D ¹H NMR, data was acquired using the 1D-NOE experiment, which filters NMR signals associated with broad line widths. Lipoprotein subclass analysis was performed using regression analysis of the NMR data, which is done automatically as part of the IVDr platform as previously described.