The Simultaneous measurement of serum testosterone and 5α-dihydrotestosterone by gas chromatography-mass spectrometry (GC-MS)

Clin Chim Acta. 2018 Jan;476:15-24. doi: 10.1016/j.cca.2017.10.030. Epub 2017 Nov 6.


Background: Simultaneous measurement of testosterone (T) and 5α-dihydrotestosterone (DHT) is important for diagnosing androgen deficiency states and hyperandrogenism in males and females, respectively. However, immunoassays used for T and DHT determination suffer from inadequate specificity and sensitivity, while tandem mass spectrometry is expensive and demanding in use.

Methods and results: We developed a selective gas chromatography-mass spectrometry (GC-MS) method for parallel T and DHT measurement. The assay showed a linear response up to 46.5nmol/L, intra- and interassay imprecision and inaccuracy <15% and recoveries in spiked samples >90% for both analytes. The limit of quantitation was 0.117nmol/L for T and 0.168nmol/L for DHT. Comparison with immunoassays revealed good agreement for T in males, but a bias in favour of immunoassays at low concentrations for T in females and DHT in both sexes. We established reference ranges for T and DHT and suggest interval partitioning for T according to age in men and menstrual cycle in women. Assay validation in a clinical setting suggests that measuring DHT or T/DHT ratio may help identify patients with polycystic ovary syndrome.

Conclusion: We developed a selective, simple and inexpensive GC-MS method for parallel measurement of T and DHT with potential use in the clinical laboratory.

Keywords: Dihydrotestosterone; Gas chromatography–mass spectrometry; Polycystic ovary syndrome (PCOS); Reference intervals; Testosterone.

MeSH terms

  • Adult
  • Dihydrotestosterone / blood*
  • Female
  • Gas Chromatography-Mass Spectrometry
  • Humans
  • Male
  • Testosterone / blood*
  • Young Adult


  • Dihydrotestosterone
  • Testosterone