Evaluation of plasma enzyme activities using gas chromatography-mass spectrometry based steroid signatures.
The simultaneous quantification of 65 plasma steroids, including 22 androgens, 15 estrogens, 15 corticoids and 13 progestins, was developed using gas chromatography-mass spectrometry (GC-MS). The extraction efficiency of the catechol estrogens was improved by the addition of l-ascorbic acid in several steps. All steroids, as their trimethylsilyl derivatives, were well separated with good peak shapes within a 50min run. The devised method provided good linearity (correlation coefficient, r(2)>0.993), while the limit of quantification ranged from 0.2 to 2.0ngmL(-1). The precision (% CV) and accuracy (% bias) were 2.0-12.4% and 93.5-109.2%, respectively. The metabolic changes were evaluated by applying this method to plasma samples obtained from 26 healthy male subjects grouped according to the pre- and post-administration of dutasteride, which inhibits 5alpha-reductase isoenzyme types 1 and 2. The levels of three plasma steroids, such as dihydrotestosterone, 5alpha-androstanedione and allotetrahydrocortisol, were decreased significantly after drug administration, while the levels of testosterone and 5beta-androstane-3beta,17alpha-diol were increased. In addition, the ratios of the steroid precursors and their metabolites, which represent the activities of the related enzymes, were z-score transformed for visualization in heat maps generated using supervised hierarchical clustering analysis. These results validated the data transformation because 5alpha-reductase is an indicator for the biological actions of dutasteride. GC-MS base quantitative visualization might be found in the integration with the mining biomarkers in drug evaluations and hormone-dependent diseases.
Ha
Young Wan
YW
Life Sciences Division, Korean Institute of Science and Technology, 39-1 Hawolkok-dong, Seoul 136-791, Republic of Korea.
Moon
Ju-Yeon
JY
Jung
Hyun-Jin
HJ
Chung
Bong Chul
BC
Choi
Man Ho
MH
D000328:Adult; D001378:Azasteroids / Q000494:pharmacology; D000068538:Dutasteride; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D006801:Humans; D008297:Male; D010949:Plasma / Q000201:enzymology*; D012680:Sensitivity and Specificity; D013256:Steroids / Q000097:blood*
D001378:Azasteroids; D004791:Enzyme Inhibitors; D013256:Steroids; D000068538:Dutasteride