A sensitive and robust liquid chromatography – tandem mass spectrometry (LC-MS/MS) method was developed and validated to simultaneously quantify the concentration levels of selective cyclo-oxygenase- 2 inhibitor, etoricoxib, and glutamate antagonist, riluzole, in rat plasma and brain tissue. Labeled internal standards (IS) etoricoxib D4 and [¹³C,¹⁵N₂] riluzole were used for the quantifications of etoricoxib and riluzole, respectively. Analytes riluzole and etoricoxib, as well as their IS's, were separated from rat plasma and brain tissue homogenate by means of liquid – liquid extraction using organic solvent of ethyl acetate. A Waters Acquity UPLC BEH C18 (1.7 µm) column was used for chromatographic separation of analytes on a SCIEX ExionLC AD using a binary solvent system. In conjunction, a SCIEX QTRAP 5500 mass spectrometer was used to quantify etoricoxib at m/z of 359 → 244 (IS of m/z 363 → 248) and riluzole at m/z 235 → 166 (IS of m/z 238 → 169) mass transitions using positive mode electrospray ionization (ESI) with multiple reaction monitoring (MRM). The bioanalytical method was validated over a linear concentration range of 0.25 (LLOQ at average signal to noise >10) – 600 ng/ml in plasma and 0.625 – 1500 ng/g in brain tissue for both analytes. The methods intra/inter-day accuracy and precision are well within the FDA validation recommendations range for chromatographic assays for both riluzole and etoricoxib in rat plasma. A partial validation was conducted within rat brain biomatrix, yielding an acceptable intra-day accuracy and precision for both riluzole and etoricoxib. For riluzole and etoricoxib the average matrix effect and recovery from rat plasma and brain tissue were negligible. This bioanalytical method was effectively applied to perform a pharmacokinetic study of each compound administered concurrently in healthy male Long-Evan (L.E.) rats. This method was developed as a pre-requisite for the pre-clinical pharmacokinetic and pharmacodynamic evaluations of co-treatment with riluzole and etoricoxib in rats with a controlled cortical impact (CCI) induced traumatic brain injury.

开发并验证了一种灵敏且稳健的液相色谱 - 串联质谱 (LC-MS/MS) 方法，可同时量化大鼠血浆和大脑中选择性环加氧酶 2 抑制剂依托考昔和谷氨酸拮抗剂利鲁唑的浓度水平组织。标记的内标 (IS) 依托考昔 D4 和 [ ¹³ C, ¹⁵ N ₂] 利鲁唑分别用于对依托考昔和利鲁唑进行定量。分析物利鲁唑和依托考昔，以及它们的 IS，通过使用乙酸乙酯有机溶剂的液-液萃取从大鼠血浆和脑组织匀浆中分离出来。Waters Acquity UPLC BEH C18 (1.7 µm) 色谱柱用于在 SCIEX ExionLC AD 上使用二元溶剂系统对分析物进行色谱分离。结合使用 SCIEX QTRAP 5500 质谱仪对 m/z 359 → 244（m/z 363 的 IS → 248）和利鲁唑 m/z 235 → 166（m/z 238 → 169 的 IS）进行定量) 使用带多反应监测 (MRM) 的正模式电喷雾电离 (ESI) 进行质量转换。生物分析方法在 0.25 的线性浓度范围内得到验证（平均信噪比的 LLOQ > 10) – 两种分析物的血浆浓度为 600 ng/ml，脑组织中浓度为 0.625 – 1500 ng/g。对于大鼠血浆中利鲁唑和依托考昔的色谱分析，该方法的日内/日间准确度和精密度完全在 FDA 验证建议范围内。在大鼠脑生物基质中进行了部分验证，利鲁唑和依托考昔均获得可接受的日内准确度和精密度。对于利鲁唑和依托考昔，大鼠血浆和脑组织的平均基质效应和回收率可以忽略不计。这种生物分析方法有效地应用于对健康雄性 Long-Evan (LE) 大鼠同时给药的每种化合物进行药代动力学研究。