Jerilynn C. Prior
Canada
Research Article
Quantitation of Urinary 6ß-Hydroxycortisol and Free Cortisol by Ultra-Performance Liquid Chromatography-Quadrupole Time-of-Flight Mass Spectrometry
Author(s): Jun Han, Shirin Kalyan, Jerilynn C. Prior and Christoph H. BorchersJun Han, Shirin Kalyan, Jerilynn C. Prior and Christoph H. Borchers
A new UPLC-MS method that combined 0.01% formic acid/isopropanol as the mobile phase and an ESI-QTOF instrument has been developed for reliable quantitation of urinary 6β-hydroxycortisol (6βHC) and free cortisol (FC). The use of 0.01% formic acid/isopropanol for binary gradient elution on a reversed-phase C18 column resulted in 3.1 and 5.0 times the peak areas for 6βHC and FC, respectively, as compared to 0.1% formic acid/acetonitrile. Liquid-liquid extraction with ethyl acetate at pH 5 was superior to solid-phase extraction for sample preparation. A mass window of 50 ppm was used for quantitative monitoring of 6βHC (m/z 379.212) and FC (m/z 363.217) using full-mass detection, with the on-column limits of detection as 4.0 and 1.4 fmol, and the lower limits of quantitation as 13.6 and 6.9 fmol for 6βHC and FC. The accuracy of quantitation ranged from 93.3% to 1.. View More»
DOI:
10.4172/2161-1459.S5-005