Molecular Recognition Research Center, Korea Institute of Science and Technology, Seoul, 02792, Republic of Korea.
Department of Pathology, Tohoku University School of Medicine, Sendai, 980-8575, Japan.
J Steroid Biochem Mol Biol. 2018 Apr;178:185-192. doi: 10.1016/j.jsbmb.2017.12.012. Epub 2017 Dec 18.
Sex steroids in clinical endocrinology have been mainly investigated with peripheral blood and urine samples, while there is limited information regarding the local levels within tissues. To improve analytical properties of sex steroids from trace amounts of tissue samples, two-phase extractive ethoxycarbonlyation and subsequent pentafluoropropionyl derivatization coupled to gas chromatography-tandem mass spectrometry (GC-MS/MS) was developed. The optimized analytical conditions led to excellent chromatographic separation of 15 estrogens, 6 androgens, and 2 progestins. The quantitative results were calculated based on in-house control samples as the steroid-free tissues, and the precision and accuracy were 4.2%-26.8% and 90.8%-116.4%, respectively. The on-column limit of quantification was from 180 fg to 0.5 pg for androgens and estrogens, and 1.25 pg for progestins, which were found to be linear (r> 0.990). The validated method was then applied to quantify 7 sex steroids from three 100-μm-thick frozen breast tissue slices from postmenopausal patients with breast cancer. This is the first report on the improved GC-MS/MS method for the detection of androgens and pregnenolone from breast cancer tissues, and it can be a useful technique to measure the local levels of sex steroids, thus, enhancing our understanding of the pathophysiological significances of steroidogenesis.
甾体性激素在临床内分泌学中主要通过外周血和尿液样本进行研究，而关于组织内局部水平的信息有限。为了提高痕量组织样本中甾体性激素的分析性能，开发了两相萃取乙氧羰基化和随后的五氟丙酰化衍生化，与气相色谱-串联质谱（GC-MS/MS）相结合。优化的分析条件导致 15 种雌激素、6 种雄激素和 2 种孕激素实现了极好的色谱分离。定量结果基于无类固醇组织的内部对照样品进行计算，精密度和准确度分别为 4.2%-26.8%和 90.8%-116.4%。雄激素和雌激素的柱上进样定量限为 180 fg 至 0.5 pg，孕激素为 1.25 pg，均呈线性（r>0.990）。然后，将经过验证的方法应用于从 3 位绝经后乳腺癌患者的 3 个 100μm 厚冷冻乳腺组织切片中定量 7 种甾体性激素。这是首次报道从乳腺癌组织中检测雄激素和孕烯醇酮的改进 GC-MS/MS 方法，它可以是一种有用的技术来测量甾体性激素的局部水平，从而增强我们对类固醇生成的病理生理意义的理解。
