【摘要】 采用液相色谱-四极杆-飞行时间质谱（LC-QTOF-MS）技术分析了5种甾体激素的肝微粒体体外孵育产物，包括孕酮、睾酮、雄烯二酮、17α-羟基孕酮及脱氧皮质酮的羟基化产物。研究发现，D环羟基化3-酮-4-烯型甾体激素的碎片离子m/z 97.06与109.06在质谱图中的相对丰度显著高于其它碎片离子，而其它羟基化位点的产物呈现独特的丰度分布特征。利用甾体激素的质谱特征碎片及其相对离子丰度，共鉴定了31种单羟基化产物，包括8种孕酮单羟基化产物、6种睾酮单羟基化产物、5种雄烯二酮单羟基化产物、8种17α-羟基孕酮单羟基化产物和4种脱氧皮质酮单羟基化产物。通过分析和总结3-酮-4-烯型甾体激素的单羟基化产物的质谱裂解途径，提高了对羟基化产物的识别精度。本研究为进一步探究甾体激素的代谢途径及其在疾病中的作用提供了重要参考，也为甾体激素组学的全面表征奠定了基础。更多还原

【Abstract】 In this study, the mass fragmentation routes responsible for the generation of m/z 97.06, 109.06,121.06, and 123.08 fragment ions from testosterone in positive ion mode were proposed in depth. Then the impactsof hydroxyl substitution site on relative ion intensity(RII) properties of those featured fragment ions weresummarized through incorporating the fragmentation pattern of testosterone and the findings described in theprevious articles. RII ratio between m/z 109.06 and 97.06 signals exhibited tight correlation with D-ringhydroxylation. Greater RII of m/z 121.06 and 97.06 usually corresponded to hydroxylation at C-2 and C-7 sites,respectively. RII of m/z 121.06 displayed the lowest value in MS2spectrum when C-6 site was hydroxylated. Themonohydroxylated products of five steroid hormones, namely progesterone, testosterone, androstenedione, 17α-hydroxyprogesterone and deoxycorticosterone, were characterized using liquid chromatography-hybrid triplequadrupole-tandem time of flight-mass spectrometry(LC-QTOF-MS) following in vitro incubation with human livermicrosomes in the presence of the whole NADPH-regeneration system. As a result, a total of 37 kinds of metaboliteswere detected and structurally deciphered based on the high-resolution m/z values of both precursor ions andfragment ions. Particularly, 31 ones, including 8, 6, 5, 8, and 4 kinds of monohydroxylated metabolites forprogesterone, testosterone, androstenedione, 17α-hydroxyprogesterone, and deoxycorticosterone, respectively,were confirmatively identified through fortifying the correlations between RII patterns and hydroxylation site to thehigh-resolution mass profiles. Moreover, the structures of two metabolites, such as 21-hydroxyprogesterone and 17α-hydroxyprogesterone, were justified by matching with authentic compounds. Above all, the identificationconfidences of hydroxylated products of 3-keto-4-ene steroid hormones were dramatically elevated through takingboth RII features and high-resolution mass values into consideration. These findings offered a robust approach forthe structural elucidation of steroid hormone metabolites, providing valuable insights into the metabolic pathwaysof steroid hormones and their roles in diseases, and laid the foundation for a comprehensive characterization of thesteroid hormone-targeted submetabolome.更多还原