Development and Evaluation of a Candidate Reference Measurement Procedure for the Determination of Progesterone in Human Serum Using Isotope-Dilution Liquid Chromatography-Tandem Mass Spectrometry

Published

September 1, 2006

Author(s)

Susan S. Tai, Bei Xu, Michael J. Welch

Abstract

Progesterone is a steroid hormone that is involved in regulating female reproductive processes. Its concentration in blood is measured to determine ovarian function. A candidate reference measurement procedure for progesterone in human serum involving isotope-dilution (ID) coupled with liquid chromatography/tandem mass spectrometry (LC/MS/MS) has been developed and critically evaluated. The progesterone along with its internal standard (progesterone -13C2) was extracted from the serum matrix using liquid-liquid extraction prior to reversed-phase LC/MS/MS. The accuracy of the measurement was evaluated by a comparison of results of this method on a lyophilized human serum reference material for progesterone [Certified Reference Material (CRM) 347] with the certified values determined by gas chromatography/mass spectrometry (GC/MS) reference methods, and by a recovery study for the added progesterone. The results of this method for progesterone agreed with the certified value within the uncertainty of the measurements for the CRM 347. The recovery of the added progesterone ranged from 100.1% to 100.9%. This method was applied to the determination of progesterone in frozen serum samples from three individual female donors with the progesterone concentrations ranged from 0.151ng/g to 24.42 ng/g. Excellent reproducibility was obtained with within-set coefficients of variations (CVs) ranging from 0.1% to 1.4%, and between-set CVs ranging from 0.3% to 0.5%. Excellent linearity was also obtained with correlation coefficients of all linear regression lines (measured intensity ratios vs. (continued) mass ratios) ranging from 0.9998 to 1.0000. The detection limit at a signal-to-noise ratio of ~3 was 1.8 pg of progesterone. This well-characterized LC/MS/MS method for serum progesterone, which demonstrates good accuracy and precision, low susceptibility to interferences, and comparability with GC/MS reference methods, qualifies as a reference measurement procedure that can be used to provide an accuracy base to which routine methods for progesterone can be compared and that will serve as a standard of higher order for measurement traceability.

Citation

Analytical Chemistry

Volume

78 No 18

Pub Type

Journals

Keywords

electrospray ionization, isotope dilution, LC/MS/MS, liquid-liquid extraction, Progesterone, reference measurement procedure

Citation

Tai, S. , Xu, B. and Welch, M. (2006), Development and Evaluation of a Candidate Reference Measurement Procedure for the Determination of Progesterone in Human Serum Using Isotope-Dilution Liquid Chromatography-Tandem Mass Spectrometry, Analytical Chemistry (Accessed April 25, 2024)