A novel GC-MS method in urinary estrogen analysis from postmenopausal women with osteoporosis

Ju-Yeon Moon,Kwang Joon Kim,Myeong Hee Moon,Bong Chul Chung,Man Ho Choi

doi:10.1194/jlr.d016113

Abstract

Estrogen metabolites play important roles in the development of female-related disorders and homeostasis of the bone. To improve detectability, a validated gas chromatography-mass spectrometry (GC-MS) method was conducted with two-phase extractive ethoxycarbonlyation (EOC) and subsequent pentafluoropropionyl (PFP) derivatization was introduced. The resulting samples were separated through a high-temperature MXT-1 column within an 8 min run and were detected in the selected ion monitoring (SIM) mode. The optimized analytical conditions led to good separation with a symmetric peak shape for 19 estrogens as their EOC-PFP derivatives. The limit of quantification (LOQ) was from 0.02 to ∼0.1 ng/ml for most estrogens analyzed, except for 2-hydroxyestriol (0.5 ng/ml). The devised method was found to be linear (r² > 0.995) in the range from the LOQ to 40 ng/ml, whereas the precision (% CV) and accuracy (% bias) ranged from 1.4 to 10.5% and from 91.4 to 108.5%, respectively. The good sensitivity and selectivity of this method even allowed quantification of the estrogen metabolites in urine samples obtained from the postmenopausal female patients with osteoporosis. The present technique can be useful for clinical diagnosis as well as to better understand the pathogenesis of estrogen-related disorders in low-level quantification.

Highlights

Estrogen metabolites play important roles in the development of female-related disorders and homeostasis of the bone
gas chromatography-mass spectrometry (GC-MS) characteristics of the derivatives. To enhance both specificity and sensitivity, a comprehensive derivatization for polar functional groups of estrogen analysis was carried out with the extractive EOC with ECF in the aqueous phase, which was applied successfully to protect the active hydrogens of the phenolic hydroxy group in estrogen molecules as the direct-derivatization technique [17]
The results showed that the EOC-PFP derivatives of estrogens were quite stable when the prepared samples were injected one day after being placed in the sample-tray (< ±12% relative standard deviation (RSD))

Summary

Introduction

Estrogen metabolites play important roles in the development of female-related disorders and homeostasis of the bone. The good sensitivity and selectivity of this method even allowed quantification of the estrogen metabolites in urine samples obtained from the postmenopausal female patients with osteoporosis. In contrast to many cancer studies [1,2,3,4], it is difficult to measure the concentrations of estrogen metabolites in postmenopausal women with osteoporosis to provide accurate quantification using conventional gas chromatography-mass. Endogenous estrogens play an important role in the development of human cancers, such as breast, endometrial, ovary, thyroid, and prostate cancer, as well as in bone homeostasis [1,2,3,4,5,6]. Higher levels of estrogens are associated with an increased risk of breast cancer, but an

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