Kawabata Kiyoshi, Samata Naozumi, Urasaki Yoko
Drug Metabolism and Pharmacokinetics Research Laboratories, Sankyo Co., Ltd., 1-2-58, Hiromachi, Shinagawa-ku, Tokyo 140-8710, Japan.
J Chromatogr B Analyt Technol Biomed Life Sci. 2005 Feb 25;816(1-2):73-9. doi: 10.1016/j.jchromb.2004.11.014.
A quantitative method was developed and validated for rapid and sensitive analysis of pravastatin and R-416, the main metabolite of pravastatin, in human plasma. The analytes were extracted from plasma samples by a solid phase extraction method using a Bond Elut C(8). The method involved the use of liquid chromatography coupled with atmospheric pressure chemical ionization (APCI) and selected reaction monitoring (SRM) mass spectrometry. A pravastatin analog, R-122798, was used as the internal standard (I.S.). Separation of pravastatin, R-416 and the I.S. was accomplished using a reverse-phase column (C(18)). The components eluted were ionized by the APCI source (negative ion) and subsequently detected by a highly selective triple quadrupole mass spectrometer in the SRM mode. Linear standard curves were obtained from 0.1 ng/mL (lower limit of quantification, LLOQ) to 100 ng/mL. The intra-assay precisions (coefficient of variation) for the samples at the LLOQ were 1.8% for pravastatin and 1.6% for R-416. The intra-assay accuracy values were 95.8-107.6% for pravastatin, and 92.6-109.0% for R-416, respectively. Precision and accuracy of quality control (QC) samples were determined at concentrations of 0.5, 10 and 80 ng/mL for all analytes. The intra- and inter-assay precision calculated from QC samples were within 10% for pravastatin and within 11% for R-416. The overall recoveries for pravastatin and R-416 were 75.7-82.1% and 68.6-74.3%, respectively. Pravastatin and R-416 were stable in human plasma for 3 weeks at -20 degrees C in a freezer, up to 6h at room temperature, and up to 48 h at 6 degrees C. This assay method was successfully used to evaluate the pravastatin and R-416 levels in healthy volunteers following oral administration of Mevalotin.
开发并验证了一种定量方法，用于快速、灵敏地分析人血浆中普伐他汀及其主要代谢物R - 416。采用Bond Elut C(8)固相萃取法从血浆样品中提取分析物。该方法采用液相色谱与大气压化学电离(APCI)和选择反应监测(SRM)质谱联用。普伐他汀类似物R - 122798用作内标(I.S.)。使用反相柱(C(18))实现普伐他汀、R - 416和内标的分离。洗脱的组分由APCI源(负离子)电离，随后由高选择性三重四极杆质谱仪在SRM模式下检测。线性标准曲线的范围为0.1 ng/mL(定量下限，LLOQ)至100 ng/mL。LLOQ水平样品的批内精密度(变异系数)，普伐他汀为1.8%，R - 416为1.6%。批内准确度值，普伐他汀为95.8 - 107.6%，R - 416为92.6 - 109.0%。所有分析物的质量控制(QC)样品在0.5、10和80 ng/mL浓度下测定精密度和准确度。从QC样品计算的批内和批间精密度，普伐他汀在10%以内，R - 416在11%以内。普伐他汀和R - 416的总回收率分别为75.7 - 82.1%和68.6 - 74.3%。普伐他汀和R - 416在人血浆中于-20℃冷冻条件下可稳定3周，室温下可达6小时，6℃下可达48小时。该测定方法成功用于评估健康志愿者口服美伐他汀后的普伐他汀和R - 416水平。
