Endoceutics Inc., 2795 Laurier Blvd, Suite 500, Quebec G1V 4M7, QC, Canada.
Endoceutics Inc., 2795 Laurier Blvd, Suite 500, Quebec G1V 4M7, QC, Canada.
J Chromatogr B Analyt Technol Biomed Life Sci. 2019 Apr 15;1113:30-36. doi: 10.1016/j.jchromb.2019.03.007. Epub 2019 Mar 7.
Sensitive and accurate measurement of androstane-3β,17β-diol and androstane-3α,17β-diol in the circulation is important for clinical research and accurate clinical diagnosis. This report describes a highly sensitive, specific, precise and reliable assay for the simultaneous accurate measurement of serum androstane-3α,17β-diol and androstane-3β,17β-diol in postmenopausal women. The LLOQ of 1 pg/mL has been achieved with nicotinic acid derivatization, which is superior to picolinic acid by a factor of 5 to 10 in terms of signal to noise ratio. The difference is attributed to the higher acidity of picolinic acid which forms a more stable intermediate, thus decreasing derivatization efficiency. Potential interference from androstane-3α, 17α-diol, androstane-3β, 17α-diol, and 5-androstenediol has been well separated from the two target diols. The high level of specificity has been determined by well-developed chromatography and ion ratio monitoring. A good linearity in the range of 1 pg/mL to 200 pg/mL (0.03 pg to 6 pg on column) was obtained for both compounds at R > 0.998. The bias and coefficients of variation of all the QC levels are within the range of 10% while the recovery in both charcoal-stripped and unstripped human serum is around 85%. The matrix effect was evaluated and the results well met the acceptance criteria according to the guidelines of bioanalytical method development and validation. Using this newly developed method, the concentrations of both androstane-3α,17β diol and androstane-3β,17β diol were measured in normal postmenopausal serum, where the concentrations range from 2 pg/mL to 32 pg/mL for androstane-3α,17β diol and from 1 pg/mL to 10 pg/mL for androstane-3β,17β diol, respectively.
准确测量循环中的雄烷二醇 3α,17β 和雄烷二醇 3β,17β 对于临床研究和准确的临床诊断非常重要。本报告介绍了一种高度敏感、特异、精确和可靠的测定绝经后妇女血清中雄烷二醇 3α,17β 和雄烷二醇 3β,17β 的方法。采用烟酸衍生化法，LLOQ 达到 1pg/mL，与吡啶甲酸相比，信噪比提高了 5 至 10 倍。这一差异归因于吡啶甲酸的酸度较高，形成更稳定的中间产物，从而降低了衍生化效率。雄烷二醇 3α,17α 和雄烷二醇 3β,17α 以及 5-雄烯二醇的潜在干扰已从两种目标二醇中得到很好的分离。通过完善的色谱和离子比率监测，确定了高特异性。两种化合物在 1pg/mL 至 200pg/mL 范围内（柱上 0.03pg 至 6pg）均具有良好的线性，相关系数 R>0.998。所有 QC 水平的偏差和变异系数均在 10%范围内，而在去蛋白和未去蛋白的人血清中的回收率均在 85%左右。已评估基质效应，结果根据生物分析方法开发和验证指南，符合可接受标准。使用新开发的方法，测量了正常绝经后血清中雄烷二醇 3α,17β 和雄烷二醇 3β,17β 的浓度，其中雄烷二醇 3α,17β 的浓度范围为 2pg/mL 至 32pg/mL，雄烷二醇 3β,17β 的浓度范围为 1pg/mL 至 10pg/mL。
