Endocrine Abstracts

Introduction: Progesterone is a steroid hormone commonly measured during assessment of female ovarian function. However, there is a lack of validated methods with the sensitivity to quantitate progesterone at concentrations found in males, post-menopausal females or patients on steroid hormone lowering medications. Here, we aimed to develop a high sensitivity LC-MS/MS method for this purpose. We also applied this method to derive a reference interval in a male cohort.

Methods: The method was developed on a Waters TQ-XS mass spectrometer. Calibrators and quality controls were gravimetrically prepared from separate stocks of certified reference material. The method utilised a 96-well plate format, with a progesterone-2,3,4-¹³C₃ internal standard. Extraction was performed using supported liquid extraction with MTBE. Male samples (n = 52), collected between 8 – 10 AM from patients aged 25 - 81, were retrieved prior to disposal, anonymised and analysed.

Results: Progesterone and its internal standard eluted with a symmetrical peak at 3.3 minutes. Concentrations ranging from 25 to 54000 pmol/l could be quantitated. The method demonstrated good recovery and minimal matrix effects. Male samples (n = 19) were excluded from the reference interval (n =14 due to medications that may cause adrenal suppression, n = 5 due to outlying progesterone concentrations). Mean progesterone was 170 pmol/L, with 2^nd and 97.5^th percentiles were 71 and 316 pmol/L, respectively.

Conclusions: We present a high sensitivity LC-MS/MS method to measure serum progesterone. This may benefit clinical applications such as assessment of infertility or breast cancer risk in post-menopausal women. We have also applied this method to derive a male reference interval for progesterone.