A liquid chromatography-mass spectrometry method for the simultaneous measurement of 15 urinary estrogens and estrogen metabolites: assay reproducibility and interindividual variability.
Level 4 - case-series / case-control
Cross-sectional analytical validation study using human urine samples
PubMed 19064556 · doi:10.1158/1055-9965.EPI-08-0355
What was done
Researchers developed an HPLC-tandem mass spectrometry (HPLC-MS/MS) assay to simultaneously measure 15 urinary estrogens and estrogen metabolites across parent, catechol, 16alpha-pathway, and methoxy estrogen classes. Assay reproducibility was tested using blinded, randomized duplicate aliquots of overnight urine from 20 individuals (5 follicular-phase premenopausal women, 5 luteal-phase premenopausal women, 5 postmenopausal women, and 5 men) run across four batches. An additional 25 participants (total n = 45) were evaluated to assess interindividual variability and metabolite profiles across groups.
What was found
Assay coefficients of variation (CV) were ≤10% for all 15 metabolites, and intraclass correlation coefficients (ICCs) within menstrual/sex groups were generally ≥98%. Five metabolites (estriol, 2-hydroxyestrone, estrone, estradiol, and 16-ketoestradiol) comprised 60% to 75% of total urinary metabolites across all groups. Within each group, absolute concentrations showed marked interindividual variability, spanning a 10- to 100-fold range for nearly all metabolites.
Why it matters
This assay provides a highly reproducible, multi-pathway LC-MS/MS panel to quantify endogenous estrogen exposures and metabolic profiles for large-scale breast cancer and endocrine epidemiological research.
Limits
The total sample size was small (n = 45; n = 20 for the core reproducibility assessment). The study evaluated single overnight urine samples and did not measure blood or breast tissue levels.
Cited by
- supports Estradiol can be converted to estrone (E1) and estriol (E3), but estriol cannot be converted back to estradiol or estrone; estrone metabolizes into 2-, 4-, and 16-hydroxyestrone, whereas estriol can metabolize into 2-hydroxyestrone but not 4- or 16-hydroxyestrone.