Published:Journal of Chromatographic Science,
ISSN 0021-9665 Volume
47, Number 1, January 2009, pp. 52-56
Investigation of HPLC–MS Using a Monolithic
Column to Separate a Diverse Suite of Steroids
Dominic M. Colosi1, John A. Bowden1,
Diana Mora-Montero1,
Timothy J. Garrett2, and Richard A. Yost1, 1Department of Chemistry, University of Florida, Gainesville,
FL 32611 and 2Biomedical Mass Spectrometry Laboratory, College
of Medicine, University of Florida, Gainesville, FL 32611
A rapid method for profiling steroids with a wide
range of polarity has been developed using high-performance liquid
chromatography–mass spectrometry with a monolithic LC column.
Steroids are detected using tandem mass spectrometry (MSn) with
a quadrupole ion trap and quantified using testosterone-d3 as
the internal standard. The method is compared to two similar
methods using a traditional particulate column in terms of number
of steroids eluted, peak area reproducibility, limits of detection,
and overall analysis time. The monolithic method elutes the steroids
in a 20-min analysis time, whereas the particulate methods elute
the steroids in 30 and 45 min, respectively. The monolithic column
also allows for improved reproducibility (relative standard deviations
from 5–23%, as opposed to 14–42% for the shorter
particulate method) and lower limits of detection (typically
2–5 times lower) when compared to the particulate column.
Finally, the method is evaluated with unextracted, spiked alligator
plasma, giving responses within 80–90% of those expected
for standards for all steroids tested (except androstenedione).
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