Analytical and Bioanalytical Chemistry

, Volume 405, Issue 10, pp 3399–3406 | Cite as

Separation of dansylated 17β-estradiol, 17α-estradiol, and estrone on a single HPLC column for simultaneous quantitation by LC–MS/MS

  • Szabolcs Szarka
  • Vien Nguyen
  • Laszlo Prokai
  • Katalin Prokai-TatraiEmail author
Original Paper


We show here that baseline separation of dansylated estrone, 17β-estradiol, and 17α-estradiol can be done, contrary to previous reports, within a short run time on a single RP-LC analytical column packed with particles bonded with phenyl-hexyl stationary phase. The chromatographic method coupled with isotope dilution tandem MS offers a simple assay enabling the simultaneous analysis of these analytes. The method employs 13C-labeled estrogens as internal standards to eliminate potential matrix effects arising from the use of deuterated estrogens. The assay also offers adequate accuracy and sensitivity to be useful for biological samples. The practical applicability of the validated method is demonstrated by the quantitative analyses of in vivo samples obtained from rats treated with Premarin®.


Quantification of estrogens from rat samples by LC–MS/MS


Estrogens Phenyl-hexyl stationary phase LC–MS/MS Dansylation 13C-labeled internal standards 



We thank Ms. Shastazia White for her excellent technical assistance. This work was supported in part by the National Institutes of Health (grant number AG031535 to LP and AG031421 to KPT) and the Robert A. Welch Foundation (endowment BK-0031 to LP).

Supplementary material

216_2013_6710_MOESM1_ESM.pdf (62 kb)
ESM 1 (PDF 62 kb)

(MPG 16102 kb)


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Copyright information

© Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  • Szabolcs Szarka
    • 1
  • Vien Nguyen
    • 1
  • Laszlo Prokai
    • 1
  • Katalin Prokai-Tatrai
    • 2
    • 3
    Email author
  1. 1.Department of Molecular Biology and ImmunologyUniversity of North Texas Health Science CenterFort WorthUSA
  2. 2.Department of Pharmacology and NeuroscienceUniversity of North Texas Health Science CenterFort WorthUSA
  3. 3.Department of Pharmaceutical SciencesUniversity of North Texas System College of PharmacyFort WorthUSA

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