Liquid chromatography-mass spectrometry (LC-MS) of steroid hormone metabolites and its applications.
Advances in liquid chromatography-mass spectrometry (LC-MS) can be used to measure steroid hormone metabolites in vitro and in vivo. We find that LC-electrospray ionization (ESI)-MS using a LCQ ion trap mass spectrometer in the negative ion mode can be used to monitor the product profile that results from 5alpha-dihydrotestosterone (DHT)-17beta-glucuronide, DHT-17beta-sulfate, and tibolone-17beta-sulfate reduction catalyzed by human members of the aldo-keto reductase (AKR) 1C subfamily and assign kinetic constants to these reactions. We also developed a stable isotope dilution LC-electron capture atmospheric pressure chemical ionization (ECAPCI)-MS method for the quantitative analysis of estrone (E1) and its metabolites as pentafluorobenzyl (PFB) derivatives in human plasma in the attomole range. The limit of detection for E1-PFB was 740attomole on column. Separations can be performed using normal-phase LC because ionization takes place in the gas phase rather than in solution. This permits efficient separation of the regioisomeric 2- and 4-methoxy-E1. The method was validated for the simultaneous analysis of plasma E2 and its metabolites: 2-methoxy-E2, 4-methoxy-E2, 16alpha-hydroxy-E2, estrone (E1), 2-methoxy-E1, 4-methoxy-EI, and 16alpha-hydroxy-E1 from 5pg/mL to 2000pg/mL. Our LC-MS methods have sufficient sensitivity to detect steroid hormone levels in prostate and breast tumors and should aid their molecular diagnosis and treatment.
Penning
Trevor M
TM
Centers of Excellence in Environmental Toxicology and Cancer Pharmacology, Department of Pharmacology, University of Pennsylvania School of Medicine, Philadelphia, PA 19104-6084, USA. penning@upenn.edu
Lee
Seon-Hwa
SH
Jin
Yi
Y
Gutierrez
Alejandro
A
Blair
Ian A
IA
D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006728:Hormones / Q000097:blood*; D006801:Humans; D007700:Kinetics; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013256:Steroids / Q000097:blood*
D006728:Hormones; D013256:Steroids