Alvi Syed N, Dgither Saleh Al, Al-Odaib Ali
Environmental Health Program, Cell Biology Department, King Faisal Specialist Hospital & Research Center, P.O. Box 3354, MBC-03, Riyadh 11211, Saudi Arabia.
Research Administrative Operations, Research and Innovation, King Faisal Specialist Hospital & Research Center, P.O. Box 3354, MBC-03, Riyadh 11211, Saudi Arabia.
Adv Pharmacol Pharm Sci. 2025 Jan 17;2025:3859670. doi: 10.1155/adpp/3859670. eCollection 2025.
A simple and efficient validated assay for quantifying 21-deoxycortisol (21-DOC), 17-hydroxyprogesterone (17-OHP), cortisol, and cortisone in human plasma has been developed using ultra-high performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS). Analysis of plasma samples were performed on Atlantis dC18 (3 m) column using a mobile phase of 20.0 mM ammonium acetate and acetonitrile (50:50, : ) that was delivered at isocratic flow rate 0.3 mL/minute. After addition of d4-cortisol as an internal standard (IS), plasma samples containing 21-DOC, 17-OHP, cortisol, and cortisone were extracted with mixture of dichloromethane and tert-butylmethyl ether 1:2 (/). Analytes were detected and quantified in the positive ion mode of electrospray ionization using multiple reaction monitoring transition set at mass to charge (m/z): 347.17 ⟶ 311.12, 331.17 ⟶ 96.97, 363.11 ⟶ 121.00, 361.18 ⟶ 163.11, and 367.19 ⟶ 121.24 for 21-DOC and 17-OHP, cortisol, cortisone, and cortisol-d4 (IS), respectively. The relationship between concentration and peak area response (analyte/IS) were linear over the range of 0.25-50, 0.5-100, 1-200, and 2-400 ng/mL for 21-DOC, 17-OHP, cortisone, and cortisol, respectively. The mean extraction recovery of the analytes was in the range of 83%-96%. The coefficient of variation within and between days was less than 13.6%, and the bias ranged from -9.2% to 12%. The measured level of cortisol, cortisone, and 17-OHP was in the range of 21.9-110, 4.33-12.71, and 0.37-1.4 ng/mL, respectively. Furthermore, the measured value of cortisone-cortisol ratio was in the range of 0.08-0.21.
已开发出一种简单高效的经过验证的测定方法，用于使用超高效液相色谱-串联质谱法（UHPLC-MS/MS）定量测定人血浆中的21-脱氧皮质醇（21-DOC）、17-羟孕酮（17-OHP）、皮质醇和可的松。血浆样品在Atlantis dC18（3μm）柱上进行分析，流动相为20.0 mM乙酸铵和乙腈（50:50，v/v），等度流速为0.3 mL/分钟。加入d4-皮质醇作为内标（IS）后，含有21-DOC、17-OHP、皮质醇和可的松的血浆样品用二氯甲烷和叔丁基甲基醚1:2（v/v）的混合物萃取。使用电喷雾电离的正离子模式，通过设置质荷比（m/z）的多反应监测转换对分析物进行检测和定量：对于21-DOC和17-OHP、皮质醇、可的松和皮质醇-d4（IS），分别为347.17⟶311.12、331.17⟶96.97、363.11⟶121.00、361.18⟶163.11和367.19⟶121.24。21-DOC、17-OHP、可的松和皮质醇的浓度与峰面积响应（分析物/内标）之间的关系在0.25 - 50、0.5 - 100、1 - 200和2 - 400 ng/mL范围内呈线性。分析物的平均萃取回收率在83% - 96%范围内。日内和日间变异系数小于13.6%，偏差范围为 - 9.2%至12%。测得的皮质醇、可的松和17-OHP水平分别在21.9 - 110、4.33 - 12.71和0.37 - 1.4 ng/mL范围内。此外，可的松 - 皮质醇比值的测量值在0.08 - 0.21范围内。
