Department of Pharmacology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
Acta Pharmacol Sin. 2010 Nov;31(11):1508-14. doi: 10.1038/aps.2010.122. Epub 2010 Sep 27.
To improve and validate analytical methods based on HPLC and liquid chromatography coupled to electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) for the quantitative measurement of sinomenine in rat plasma and brain tissue.
The separation of analytes and the internal standard (IS), chloramphenicol, was performed on an Agilent TC-C18 column (250×4.6 mm, 5 μm). Blood samples were measured with a Surveyor photodiode array (PDA) detector at a wavelength of 263 nm. The LCQ DECA XP(Plus) mass spectrometer was operated in the multiple reactions monitoring mode using positive electrospray ionization, and the transition from the precursor ion (m/z 279) to the product ion (m/z 224) for sinomenine was measured in brain tissue.
Measurements were linear over the concentration range of 0.1-100 μg/mL for sinomenine in plasma and over the range of 0.01-5.00 μg/g for sinomenine in brain tissue. The intra- and inter-day variabilities were less than 10% of the relative standard deviation (RSD), and the extraction and recovery of sinomenine was 72.48%-80.26% from plasma and 73.75%-80.26% from brain tissue. The limit of quantification (LOQ) was 0.1 μg/mL for plasma, and 0.01 μg/g for brain tissue. Identification of sinomenine was reproducible at 0.5, 5, and 50 μg/mL in the plasma and at 0.05, 0.50, and 2.00 μg/g in brain tissue. The concentration of sinomenine measured in brain tissue after a single ip dose had a neuroprotective effect on H₂O₂-induced injury in PC12 cells in vitro.
Our methods offered a sensitivity within a wide linear concentration range for sinomenine. These methods were successfully applied to evaluate sinomenine pharmacokinetics over time in rat brain tissue after a single ip dose of 30 mg/kg.
改进和验证基于高效液相色谱和液相色谱-电喷雾串联质谱(LC-ESI-MS/MS)的分析方法,用于定量测定大鼠血浆和脑组织中的盐酸青藤碱。
采用 Agilent TC-C18 柱(250×4.6mm,5μm)对分析物和内标(氯霉素)进行分离。采用 Surveyor 光电二极管阵列(PDA)检测器在 263nm 波长处测量血样。LCQ DECA XP(Plus)质谱仪在正电喷雾电离模式下以多反应监测模式运行,测量脑组织中盐酸青藤碱的前体离子(m/z 279)到产物离子(m/z 224)的跃迁。
盐酸青藤碱在血浆中的浓度范围为 0.1-100μg/mL 时,在脑组织中的浓度范围为 0.01-5.00μg/g 时,测定结果均呈线性。日内和日间变异系数(RSD)小于 10%,盐酸青藤碱在血浆中的提取回收率为 72.48%-80.26%,在脑组织中的提取回收率为 73.75%-80.26%。血浆中的定量下限(LOQ)为 0.1μg/mL,脑组织中的 LOQ 为 0.01μg/g。在血浆中浓度为 0.5、5 和 50μg/mL,在脑组织中浓度为 0.05、0.50 和 2.00μg/g 时,盐酸青藤碱的重现性良好。单次腹腔注射 30mg/kg 后,大鼠脑组织中盐酸青藤碱的浓度对 H₂O₂诱导的 PC12 细胞损伤具有神经保护作用。
本方法提供了一种在较宽线性浓度范围内测定盐酸青藤碱的灵敏度。这些方法成功地应用于评估单次腹腔注射 30mg/kg 后大鼠脑组织中盐酸青藤碱的药代动力学。