Measurement of steroid hormones by liquid chromatography-tandem mass spectrometry with small amounts of hair

Abstract

Steroid hormone levels in hair reflect the integrated values (average values) of hormone secretion over the past few months. We have used a method to evaluate diseases and chronic stress, discrimination of banned drug use, and so on. In contrast, the hair analysis methods reported so far required at least 10 mg (about 50 to 100 hair strands) of hair to analyze multiple steroid hormones from the same sample. Here, we developed a new method for measuring steroid hormones in hair by liquid chromatography-tandem mass spectrometry, which identifies multiple steroid hormones from 5 to 10 (about 1 mg) hair strands. Ten steroid hormones (cortisol, cortisone, testosterone, dihydrotestosterone, dehydroepiandrosterone, androstenedione, progesterone, pregnenolone, androstenediol and estradiol) covering from sex hormones to stress hormones were derivatized and measured by four different measuring systems. The method showed good linearity for all steroids with correlation coefficients of 0.999 or more. The accuracy and precision of intra- and inter-assay ranged from 96.0 to 106.4% and 4.8 to 8.1% for intra-assay, and from 96.9 to 104.9% and 6.9 and 10.6% for inter-assay, respectively. A mixed solution containing 0.1 M trifluoroacetic acid and 50% acetonitrile was used to extract hair and to enhance the cortisol extraction efficiency approximately twice compared to the previously reported extraction with methanol. This method has the potential to clarify the relationship between steroid hormone levels and diseases that show alopecia such as chronic stress and androgenetic alopecia.