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May 6, 2026Analytical Science Advances0 citationsOpen Access

Comprehensive Quantitative Urinary Steroid Profiling of 29 Steroids Using Liquid Chromatography‐Tandem Mass Spectrometry

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JBJoshua BainFSFozia ShaheenAPAlessandro Prete

Key Points

  • This research aims to develop a reliable method for quantifying multiple urinary steroids using LC-MS/MS.
  • Utilized liquid chromatography-tandem mass spectrometry for steroid separation and quantification.
  • Analyzed urine samples from 40 healthy volunteers to assess steroid levels and variations.
  • Measured metrics such as recovery, precision, and matrix effects during assay validation.
  • Most steroids were detected above the lower limit of quantification in over 95% of samples.
  • Tetrahydro-aldosterone, 5-pregnenediol, and pregnanetriol-one showed lower quantification rates (85%, 68%, 59% respectively).
  • Significant diurnal and sex-based variations were observed in excretion levels.

Abstract

column for their separation. Lower limit of quantification (LLOQ) ranged from 2 to 20 ng/mL, witha ccuracy (bias) ranging from -18.7% to 19.9%, and precision (percentage coefficient of variation %CV) ranging from 4.0% to 18.6%. Matrix effects remained within the ideal range <±15% for all steroids. Recovery ranged from 76% to 103%, and intra- and inter-assay imprecision (CV) ranged from 0.8% to 14.9%. In 40 healthy volunteers, most analytes were detected above the LLOQ in over 95% of samples, although tetrahydro-aldosterone (85%), 5-pregenediol (68%), and pregnanetriol-one (59%) demonstrated lower quantification rates. Diurnal and sex-based variations were observed, with excretion levels significantly higher during daytime and in males. This robust, high-throughput LC-MS/MS method facilitates the simultaneous quantification of multiple steroid classes, enhancing its utility for clinical and research applications in endocrine science.

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Cite This Study

Bain et al. (2026) studied this question.

synapsesocial.com/papers/69fada7f03f892aec9b1e4d5https://doi.org/10.1002/ansa.70087
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