madman
Super Moderator
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.
1. Introduction
Steroid hormones are biologically active substances having important roles in sexual maturation, sugar metabolism, and maintenance of homeostasis. Since the endogenous concentrations of steroid hormones reflect various physiological and pathological conditions, they are also used as a biomarker [1–3]. In most studies, blood (plasma, serum) saliva [4,5] and urine [6] were used frequently as samples for measurement. Recently, the number of studies on steroid hormones in hair has increased. Because the concentrations in a hair show the integrated values (average values) of hormone secretion over the past period [7,8] hair samples are highly useful compared with the conventional samples such that they cannot be influenced by a circadian rhythm of secretion [9] and can be collected non-invasively.
4. Discussion
We have developed a method by analyzing the sample in LC-MS/MS that can measure 10 steroid hormones in only 5 to 10 hair strands (about 1 mg) from the same sample. It was a combination of following the three techniques to establish a sensitive analysis method: (1) use of a novel, highly efficient solvent extraction instead of methanol; (2) derivatization of steroids with picolinic acid and fusaric acid; (3) high-resolution and high-sensitivity analysis using 2D-LC.
5. Conclusion
Herein, we reported the highly sensitive and low-invasive quantitation method for 10 steroid hormones in hair, covering from sex hormones to stress hormones. There have already been several reports on the assessment of chronic stress by measuring the steroid hormone concentrations in hair samples. However, the number of hair samples for the assay (50 to 100 hair strands = about 10 mg) would be a burden for the participants and be unrealistic when applied to large-scale cohort studies, examinations, and diagnosis. The novel method we have developed achieved a realistic hair usage (5 to 10 hair strands = about 1 mg) for cohort studies and hair diagnosis, so it will contribute to the convenience of future research. In future research, we hope that this method will clarify the relationship between steroid hormone levels and diseases that show alopecia such as chronic stress and AGA.
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.
1. Introduction
Steroid hormones are biologically active substances having important roles in sexual maturation, sugar metabolism, and maintenance of homeostasis. Since the endogenous concentrations of steroid hormones reflect various physiological and pathological conditions, they are also used as a biomarker [1–3]. In most studies, blood (plasma, serum) saliva [4,5] and urine [6] were used frequently as samples for measurement. Recently, the number of studies on steroid hormones in hair has increased. Because the concentrations in a hair show the integrated values (average values) of hormone secretion over the past period [7,8] hair samples are highly useful compared with the conventional samples such that they cannot be influenced by a circadian rhythm of secretion [9] and can be collected non-invasively.
4. Discussion
We have developed a method by analyzing the sample in LC-MS/MS that can measure 10 steroid hormones in only 5 to 10 hair strands (about 1 mg) from the same sample. It was a combination of following the three techniques to establish a sensitive analysis method: (1) use of a novel, highly efficient solvent extraction instead of methanol; (2) derivatization of steroids with picolinic acid and fusaric acid; (3) high-resolution and high-sensitivity analysis using 2D-LC.
5. Conclusion
Herein, we reported the highly sensitive and low-invasive quantitation method for 10 steroid hormones in hair, covering from sex hormones to stress hormones. There have already been several reports on the assessment of chronic stress by measuring the steroid hormone concentrations in hair samples. However, the number of hair samples for the assay (50 to 100 hair strands = about 10 mg) would be a burden for the participants and be unrealistic when applied to large-scale cohort studies, examinations, and diagnosis. The novel method we have developed achieved a realistic hair usage (5 to 10 hair strands = about 1 mg) for cohort studies and hair diagnosis, so it will contribute to the convenience of future research. In future research, we hope that this method will clarify the relationship between steroid hormone levels and diseases that show alopecia such as chronic stress and AGA.
