Development of a quantitative method for determination of steroids in human plasma by gas chromatography-negative chemical ionization-tandem mass spectrometry.

Sex steroids are involved in biological functions that encompass from the complete sexual development of individuals up to the deregulation of metabolic pathways leading to some pathologies. Steroids are present in blood at low concentration levels from pg mL to ng mL. For this reason, a high sensitive and selective method based on gas chromatography-negative chemical ionization-tandem mass spectrometry (GC-NCI-MS/MS) is here proposed to quantify either androgens (androstenedione, dehydroepiandrosterone, dihydrotestosterone and testosterone), estrogens (estrone and estradiol) and a progestogen (progesterone) in human plasma. The sample preparation steps, protein precipitation and solid phase extraction, were optimized to ensure the sample matrix removal and to extract steroids with high efficiency. The NCI-MS/MS detection approach was compared with that based on electron impact to evaluate the incidence of the ionization source in the determination of steroids. The quantification limits for determination of these analytes were in a range from 10 pg mL to 5 ng mL, with a high sensitivity for estrogens, typically found at low concentrations. The proposed method was tested for the determination of steroids in male blood samples, in which 6 out of 7 steroids were detected and quantified to report concentration values in agreement with those described in the literature.