Profiling intact steroid sulfates and unconjugated steroids in biological fluids by liquid chromatography-tandem mass spectrometry (LC-MS-MS)
Open accessAbstract
Within the combined DFG research project "Sulfated Steroids in Reproduction" an analytical method was needed for determining sulfated and unconjugated steroids with highest specificity out of different biological matrices such as aqueous solution, cell lysate and serum. With regard to this analytical challenge, LC-MS-MS presents the technique of choice because it permits (1) analysis of the intact steroid conjugate, (2) allows for simultaneous determination of multiple analytes (profiling, targeted metabolomics approach) and (3) is independent of phenomena such as cross-reactivity. Sample work up consisted of incubation of sample with internal standards (deuterium labeled steroids) followed by solid phase extraction. Only serum samples required a protein precipitation step prior to solid phase extraction. The extract was divided in two parts: six steroid sulfates (E1S, E2S, AS, 16-OH-DHEAS, PREGS, DHEAS) were analyzed by C18aQ-ESI-MS-MS in negative ion mode and eleven unconjugated steroids (E3, 16-OH-DHEA, E1, E2, (4)A, DHEA, T, 17-OH-PREG, Prog, An, PREG) were analyzed by C18-APCI-MS-MS in positive ion mode. For steroid sulfates, we found high sensitivities with LoQ values ranging from 0.08 to 1 ng mL(-1). Unconjugated steroids showed LoQ values between 0.5 and 10 ng mL(-1). Calibration plots showed excellent linearity. Mean intra- and inter-assay CVs were 2.4% for steroid sulfates and 6.4% for unconjugated steroids. Accuracy - determined in a two-level spike experiment - showed mean relative errors of 5.9% for steroid sulfates and 6.1% for unconjugated steroids. In summary, we describe a novel LC-MS-MS procedure capable of profiling six steroid sulfates and eleven unconjugated steroids from various biological matrices.
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To date only a limited number of methods for the simultaneous measurement of free, sulfated and glucuronidated androgens have been established, although significant progress has been made.
“but to date only a limited number of methods for the simultaneous measurement of free, sulfated and glucuronidated androgens have been established, although significant progress has been made (Badoud et al., 2011, Doue et al., 2015, Galuska et al., 2013).”
From Intracrine androgen biosynthesis, metabolism and action revisited (2018) · cites this paper as Galuska et al., 2013
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- Authors
- Galuska, C. E., Hartmann, M. F., Sánchez-Guijo, A., Bakhaus, K., Geyer, J., Schuler, G., Zimmer, K.-P., & Wudy, S. A.
- Affiliations
- Justus-Liebig-Universität Gießen (DE)
- Corresponding author
- Hartmann, Michaela F.
- Venue
- The Analyst
- Publisher
- Royal Society of Chemistry (RSC)
- Published
- 2013
- Volume
- 138
- Issue
- 13
- Pages
- 3792
- ISSN
- 0003-2654 (ISSN-L)
- 1364-5528
- OpenAlex
- W2086120516
- Cited by
- 64
- References
- 31
- Citations by year
- 2026: 3 · 2025: 4 · 2024: 2 · 2023: 4 · 2022: 3 · 2021: 5 · 2020: 2 · 2019: 3 · 2018: 9 · 2017: 10 · 2016: 6 · 2015: 4 · 2014: 6 · 2013: 3
- FWCI
- 2.83
- Type
- journal article
- Language
- en
- Open access
- green
- doi.org
- Locations
- The Analyst (publishedVersion)
- PubMed (publishedVersion)
- University writing server of the Justus Liebig University Giessen (Justus-Liebig-Universität Gießen) (submittedVersion)
- Universitätsbibliothek Gießen
- Funders
- Deutsche Forschungsgemeinschaft
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Metabolomics and Mass Spectrometry Studies
- Physical Sciences › Chemistry › Spectroscopy › Mass Spectrometry Techniques and Applications
- Physical Sciences › Chemistry › Spectroscopy › Analytical Chemistry and Chromatography
- Keywords
- steroid sulfates, steroid profiling, biological fluids, serum, solid-phase extraction, deuterated internal standards, targeted metabolomics, reproductive endocrinology, dehydroepiandrosterone sulfate, androgens, progestins
- MeSH terms
- Blood Chemical Analysis, Calibration, Chromatography, Liquid, Humans, Steroids, Sulfates, Reproducibility of Results, Tandem Mass Spectrometry, HEK293 Cells
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