title: Comprehensive and highly sensitive urinary steroid hormone profiling method based on stable isotope-labeling liquid chromatography-mass spectrometry.
abstract: Steroid hormones are crucial substances that mediate a wide range of vital physiological functions of the body. Determination of the levels of steroid hormones plays an important role in understanding the mechanism of the steroid hormone-related diseases. In this study, we present a novel targeted metabolic profiling method based on the introduction of an easily protonated stable isotope tag to a hydroxyl-containing steroid hormone with a synthesized derivatization reagent, deuterium 4-(dimethylamino)-benzoic acid (d(4)-DMBA), and liquid chromatography-mass spectrometry (LC-MS). Different from other reported derivatization reagents that have been used to enhance the sensitivities for estrogens or androgens, our method is comprehensive with the capability of covering hydroxyl-containing androgens, estrogens, corticoids, and progestogens. Furthermore, the nonderivatized steroid hormones (e.g., 17α-hydroxyprogesterone, progesterone, and androstenedione) were not destroyed during the derivatization process, and their levels could still be obtained in one LC-MS run. We were able to detect 24 steroid hormones at subng/mL levels (the lower limit of detection could reach 5 pg/mL for estrone and 16α-hydroxy estrone, which is equivalent to 0.1 pg on column) with maximum sensitivity enhancement factors of more than 10(3)- to 10(4)-fold after derivatization. The method was successfully applied to the measurement of free (unconjugated) steroid hormones in urine samples of males, females, and pregnant women. Because the significant role the steroid hormone pathway plays in humans, a comprehensive, sensitive, specific, and accurate method for profiling the steroid hormone metabolome shall offer new insights into hormone-related diseases.
authors.affiliation: CAS Key Laboratory of Separation Science for Analytical Chemistry, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, China.
authors.forename: Weidong
authors.initials: W
authors.lastname: Dai
authors.forename: Qiang
authors.initials: Q
authors.lastname: Huang
authors.forename: Peiyuan
authors.initials: P
authors.lastname: Yin
authors.forename: Jia
authors.initials: J
authors.lastname: Li
authors.forename: Jia
authors.initials: J
authors.lastname: Zhou
authors.forename: Hongwei
authors.initials: H
authors.lastname: Kong
authors.forename: Chunxia
authors.initials: C
authors.lastname: Zhao
authors.forename: Xin
authors.initials: X
authors.lastname: Lu
authors.forename: Guowang
authors.initials: G
authors.lastname: Xu
mesh_terms: D000328:Adult; D002853:Chromatography, Liquid / Q000379:methods*; D003903:Deuterium; D005260:Female; D012739:Gonadal Steroid Hormones / Q000652:urine*; D006801:Humans; D007553:Isotope Labeling; D008297:Male; D010446:Peptide Fragments / Q000032:analysis*; D011247:Pregnancy; D052616:Solid Phase Extraction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D014357:Trypsin / Q000378:metabolism; D055815:Young Adult
chemical_list: D012739:Gonadal Steroid Hormones; D010446:Peptide Fragments; D003903:Deuterium; D014357:Trypsin