Proposed definitive methods for measurement of plasma testosterone and 17 alpha-hydroxyprogesterone.
OBJECTIVE
This report provides the results of the development and evaluation of definitive isotope dilution/mass spectrometry (ID/MS) methods for the determination of testosterone and 17 alpha-hydroxyprogesterone in human plasma at concentrations ranging from 1.4 to 37.9 nmol/L and 1.5 to 45.4 nmol/L, respectively. The internal standards were 16, 16, 17-2H-testosterone and 21,21,21, -2H-17 alpha-hydroxyprogesterone. The development of optimum extraction and derivatization procedures, and studies of storage time, temperature effects, accuracy, and precision are presented.


RESULTS
The results indicate that the methods employing the TBDMS derivative of testosterone and MO-TMS derivative of 17 alpha-hydroxyprogesterone are capable of generating accurate and precise data at the inherently low concentrations given, with recovery greater than 95%. Accuracy of testosterone in the fortified steroid-free and pooled plasma by ID/MS measurement was good because the relative error ranged from +3.1% to -0.7, with a mean of 0.9% over the concentration levels of 1.4 to 37.9 nmol/L testosterone, and the imprecision ranged from 4.2% to 0.7% CV, with a mean of 1.9%. Accuracy of 17 alpha-hydroxyprogesterone in the fortified steroid-free and pooled plasma was also good, considering the inherently low concentrations. The relative error ranged from -2.1% to +1.5%, with a mean of 1.1%, and the imprecision ranged from 3.8% to 0.8% CV with a mean of 1.3% over concentration levels of 1.5 to 15.1 nmol/L. The high precision and accuracy and absence of statistically significant bias qualifies these methods as candidate definitive methods for plasma testosterone and 17 alpha-hydroxyprogesterone.
Massé
R
R
INRS-Santé, Pointe-Claire, Québec, Canada.
Wright
L A
LA
D019326:17-alpha-Hydroxyprogesterone / Q000097:blood*; D005260:Female; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D006801:Humans; D008297:Male; D013739:Testosterone / Q000097:blood*
D013739:Testosterone; D019326:17-alpha-Hydroxyprogesterone