Department of Physiological Sciences, Florida Racing Laboratory, University of Florida, College of Veterinary Medicine, Gainesville, USA.
J Anal Toxicol. 2011 Nov;35(9):656-64. doi: 10.1093/anatox/35.9.656.
A rapid, sensitive, and specific ultra-high-performance liquid chromatography with heated electrospray ionization-tandem mass spectrometry (UHPLC-HESI-MS-MS) method to detect and quantify glycopyrrolate in horse plasma has been developed and validated. We also determined glycopyrrolate in plasma after oral and intravenous administration of clinically relevant doses to Thoroughbred horses. Calibration was accomplished by weighted, linear regression analysis using a deuterated analogue of glycopyrrolate as internal standard (IS). Glycopyrrolate (GLY) and the IS (GLY-d(3)) were isolated from plasma matrices via weak cation exchange using a simple solid-phase extraction technique. Chromatographic analysis was achieved by reversed-phase UHPLC on a C(18) Acquity™ column. Extracts were analyzed in positive electrospray ionization mode and precursor and product ions were detected and quantified by MS-MS using a triple-stage quadrupole (TSQ) instrument. The method was characterized by a linear range of 0.125-25 pg/mL (R(2) > 0.998), a lower limit of quantification of 0.125 pg/mL and a lower limit of detection of 0.025 pg/mL. Recovery of GLY ranged from 78% to 96%, and intra- and interbatch precision were 3.3-14.4%CV and 3.4-14.4%CV, respectively. Glycopyrrolate was stable in plasma for up to 170 days at -80°C, through three freeze/thaw cycles, and for up to 48 h after extraction under 20°C autosampler conditions.
已建立并验证了一种快速、灵敏、特异的超高效液相色谱-加热电喷雾串联质谱（UHPLC-HESI-MS-MS）法，用于检测和定量马血浆中的硫酸氢吡哆醇。我们还测定了口服和静脉给予临床相关剂量的硫酸氢吡哆醇后马血浆中的硫酸氢吡哆醇。通过使用硫酸氢吡哆醇氘代物作为内标（IS）的加权线性回归分析来完成校准。通过使用简单的固相萃取技术，从血浆基质中通过弱阳离子交换分离硫酸氢吡哆醇（GLY）和 IS（GLY-d（3））。通过反相 UHPLC 在 C（18） AcquityTM 柱上进行色谱分析。通过正电喷雾电离模式提取，通过三重四极杆（TSQ）仪器的 MS-MS 检测和定量前体和产物离子。该方法的线性范围为 0.125-25 pg/mL（R（2）>0.998），定量下限为 0.125 pg/mL，检测下限为 0.025 pg/mL。GLY 的回收率为 78%-96%，批内和批间精密度分别为 3.3%-14.4%CV 和 3.4%-14.4%CV。硫酸氢吡哆醇在-80°C 下可稳定 170 天，在 20°C 自动进样器条件下提取后可稳定 48 小时，经过三个冻融循环后仍稳定。
