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

Background: Simultaneous measurement of testosterone (T) and 5 alpha-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 46.5 nmol/L, intra- and interassay imprecision and inaccuracy < 15% and recoveries in spiked samples > 90% for both analytes. The limit of quantitation was 0.117 nmol/L for T and 0.168 nmol/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.