Determination of estrone and 17β-estradiol in human hair by gas chromatography–mass spectrometry

Abstract

An efficient procedure is described for the determination of estrone and 17 beta-estradiol in hair by gas chromatography-mass spectrometry (GC-MS). The method involves alkyloxycarbonylation with isobutyl chloroformate (isoBCF) of phenolic hydroxy groups after alkaline digestion of hair samples. The resulting isobutyloxycarbonyl derivatives of estrone and 17 beta-estradiol are extracted with hexane and subjected to chlorodifluoroacetyl derivatization in order to protect the remaining alcoholic hydroxy groups. When GC-MS with selected ion monitoring (SIM) was used, the quantitative ions were at m/z 270 and 384 in the electron ionization mass spectra for estrone and 17 beta-estradiol, respectively. The detection limits for SIM of the steroids were 1 and 2 pg, respectively, and the SIM responses were linear with correlation coefficients varying from 0.991 to 0.994 in the concentration range 0.2-4.0 ng g-1 for the estrogens studied. The detection of estrone and 17 beta-estradiol in hair samples was possible in the concentration range of 0.24-1.30 ng g-1. The concentrations of the two estrogens detected were different in male and female hair samples.

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

“…spectrometry (GC-MS)-based estrogen methods. These have been developed to measure the endogenous estrogen metabolites in biological specimens using RIA or enzyme immunoassay (EIA) ( 13,14 ), and GC-MS ( 4,(15)(16)(17)(18) or LC-MS (19)(20)(21). Although immunoassays have limited applicability due to an overestimation by cross reactions (22)(23)(24)(25), LC-MS-based profi ling is a proven technique with high sensitivity and selectivity in estrogen analysis combined with chemical derivatization for the phenolic hydroxyl groups of estrogens as follows: a ) pentafl uorobenzyl (PFB) derivatization with ECAPCI-MS ( 21 ), b ) dansyl ( 20 ) and picolinoyl ( 19 ) derivatizations with LC-ESI/MS, and c ) N-methyl-2-pyridyl ( 26 ), 1-(2,4-dinitro-5-fl uorphenyl)-4,4,-dimethylpiperazine ( 27 ), or N-methyl-nicotinyl derivatizations with LC-ESI/MS ( 28 ).…”

“…In high-resolution GC-MS analysis, estrogen analysis is essential in blocking active hydrogens in both phenolic and aliphatic hydroxy groups. For this purpose, perfl uoroacylation and trimethylsilylation are mainly performed ( 4,(15)(16)(17)(18). As a different approach, alkoxycarbonylation, which is carried out with highly reacting agents for amino-, thiol-, imidazole-or phenolic hydroxy groups, can be suitable for direct reactions in aqueous media ( 17,(30)(31)(32)(33)(34).…”

“…For this purpose, perfl uoroacylation and trimethylsilylation are mainly performed ( 4,(15)(16)(17)(18). As a different approach, alkoxycarbonylation, which is carried out with highly reacting agents for amino-, thiol-, imidazole-or phenolic hydroxy groups, can be suitable for direct reactions in aqueous media ( 17,(30)(31)(32)(33)(34). This direct derivatization to make analytes extractable by nonpolar organic solvents can be achieved simultaneously to provide rapid and reproducible results with excellent purifi cation ( 17,31,32 ).…”

“…To enhance both specifi city 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 ). In the subsequent derivatization to block the remaining aliphatic hydroxy and ketone groups prior to GC analysis, TMS derivatization was initially tested, and the results of the EOC-TMS derivatives were compared with the commonly used TMS derivatives (supplementary Fig.…”

“…When high-resolution capillary GC-MS is used for estrogen analysis (15)(16)(17)(18), it is essential to block the active hydrogens in these polar functional groups. Although TMS derivatives are commonly used in GC-MSbased estrogen analysis ( 15,16 ), TMS derivatives are not sensitive enough for quantifi cation of urinary estrogen metabolites and the derivatization process is time consuming.…”