Charles River Laboratories Preclinical Services, Montreal, Canada.
J Pharm Biomed Anal. 2011 Apr 28;55(1):125-34. doi: 10.1016/j.jpba.2010.12.031. Epub 2010 Dec 30.
Liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods for the determination of unconjugated and total (conjugated plus unconjugated) S-equol in human plasma and urine were developed and validated. The separation of R and S enantiomers was achieved with a Chiracel OJ-H column operated in a normal phase mode using ethanol/hexane mobile phase components. Ionization of S-equol by negative ion electrospray generated the M-H ion whose response was augmented by post-column addition of ammonium hydroxide. A triple stage quadrupole mass spectrometer was used to measure the ion current generated from the dissociative transitions m/z 241→m/z 121 (S-equol) and m/z 245→m/z 123 (equol-d(4)). The determination of total S-equol included an additional deconjugation step involving incubation of the sample with sulfatase and glucuronidase. Average recovery for both unconjugated and total S-equol was 85% with no observable matrix effects. Linearity was established for unconjugated S-equol from 0.025ng/mL to 10ng/mL (plasma) and 0.20ng/mL to 200ng/mL (urine). The average coefficient of variation and accuracy per occasion was within ±15% of the theoretical concentration of S-equol. The method was used to measure the pharmacokinetics of S-equol in human plasma after an oral administration of a single 20mg dose of S-equol to three normal healthy volunteers.
建立并验证了用于测定人血浆和尿液中未结合和总（结合加未结合）S-雌马酚的液相色谱-串联质谱（LC-MS/MS）方法。采用 Chiracel OJ-H 柱，在正相模式下，以乙醇/己烷为流动相，实现了 R 和 S 对映体的分离。通过负离子电喷雾使 S-雌马酚离子化，生成M-H离子，通过柱后加入氨水溶液来增强其响应。采用三重四极杆质谱仪测量从解离跃迁 m/z 241→m/z 121（S-雌马酚）和 m/z 245→m/z 123（雌马酚-d(4)）产生的离子电流。总 S-雌马酚的测定包括一个额外的去结合步骤，涉及用硫酸酯酶和葡萄糖醛酸酶孵育样品。未结合和总 S-雌马酚的平均回收率为 85%，无明显基质效应。未结合 S-雌马酚的线性范围为 0.025ng/mL 至 10ng/mL（血浆）和 0.20ng/mL 至 200ng/mL（尿液）。每次测定的平均变异系数和准确度均在 S-雌马酚理论浓度的±15%范围内。该方法用于测量 3 名正常健康志愿者口服单剂量 20mg S-雌马酚后 S-雌马酚在人血浆中的药代动力学。
