Investigation of HPLC-MS using a monolithic column to separate a diverse suite of steroids.
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 (MS(n)) with a quadrupole ion trap and quantified using testosterone-d(3) 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).
Colosi
Dominic M
DM
Department of Chemistry, University of Florida, Gainesville, FL 32611, USA.
Bowden
John A
JA
Mora-Montero
Diana
D
Garrett
Timothy J
TJ
Yost
Richard A
RA
D000489:Alligators and Crocodiles / Q000097:blood*; D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation / Q000379:methods*; D006801:Humans; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013256:Steroids / Q000032:analysis* / Q000097:blood; D053719:Tandem Mass Spectrometry / Q000379:methods*; D013739:Testosterone / Q000737:chemistry; D013997:Time Factors
D013256:Steroids; D013739:Testosterone