Bullen W W, Lathia C D, Abel R B, Hayes R N
Parke-Davis Pharmaceutical Research Division, Warner-Lambert Company, Ann Arbor, MI 48105, USA.
J Pharm Biomed Anal. 1998 Sep;17(8):1399-413. doi: 10.1016/s0731-7085(98)00034-x.
A liquid chromatographic/mass spectrometric (LC/MS/MS) method to quantitate CI-1011 in rat plasma has been validated and compared to an LC/UV assay. The analyte and internal standard were isolated from the plasma matrix by using liquid/liquid extraction with diethyl ether. The ether layer was evaporated to dryness and the residue reconstituted in acetonitrile-water (70:30, v/v). A 2.1 x 150 mm x 5 microns Zorbax RX-C18 column with a mobile phase of acetonitrile-ammonium acetate (pH 8.0; 5 mM)-triethylamine (70:30:0.03, v/v/v) delivered at a flow rate of 0.2 ml min-1 was used for chromatography. Analyte and internal standard ion chromatograms were obtained by operating the mass spectrometer in the negative ion multiple reaction monitoring mode to detect the presence of a precursor-product ion pair for both the analyte and the internal standard. Samples were introduced into the mass spectrometer using electrospray ionization. Retention times of CI-1011 and of the internal standard (IS), [13C6]CI-1011, were approximately 4.2 min. No peaks interfering with the quantitation of CI-1011 were observed throughout the validation process. Mean recoveries of CI-1011 from rat plasma ranged from 98.2 to 105%. The recovery of the IS was 100%. Assay precision for CI-1011, based on the percent relative standard deviation of replicate quality controls, was less than or equal to 5.60% with an accuracy of +/- 8.80%. The lower limit of quantitation for CI-1011 was 0.500 ng ml-1 for a 0.2-ml sample aliquot. CI-1011 is stable in rat plasma for 24 h at room temperature and for at least 34 days at -20 degrees C. This assay has been proven suitable for routine quantitation of CI-1011 in rat plasma at concentrations from 0.500 (100 pg on-column) to 500 ng ml-1. The applicability of this method to determine CI-1011 concentrations in rat plasma is reported in this manuscript. CI-1011 concentrations, in plasma samples from cholesterol- and chow-fed rats administered single daily oral doses of CI-1011 in a CMC/Tween suspension, obtained using a validated LC/UV assay were compared to concentrations obtained using the reported LC/MS/MS assay over the concentration range 0.0806-12.3 micrograms ml-1. The concordance correlation coefficient determined for this comparison was 0.9977, suggesting that the CI-1011 concentrations obtained by the two assays are in excellent agreement.
已验证一种用于定量大鼠血浆中CI-1011的液相色谱/质谱联用（LC/MS/MS）方法，并将其与LC/UV测定法进行了比较。通过使用乙醚液-液萃取从血浆基质中分离出分析物和内标。将乙醚层蒸发至干，残留物用乙腈-水（70:30，v/v）复溶。使用2.1×150 mm×5微米的Zorbax RX-C18色谱柱，流动相为乙腈-醋酸铵（pH 8.0；5 mM）-三乙胺（70:30:0.03，v/v/v），流速为0.2 ml min-1进行色谱分析。通过在负离子多反应监测模式下操作质谱仪以检测分析物和内标的前体-产物离子对的存在，获得分析物和内标离子色谱图。使用电喷雾电离将样品引入质谱仪。CI-1011和内标[13C6]CI-1011的保留时间约为4.2分钟。在整个验证过程中未观察到干扰CI-1011定量的峰。CI-1011从大鼠血浆中的平均回收率为98.2%至105%。内标的回收率为100%。基于重复质量控制的相对标准偏差百分比，CI-1011的测定精密度小于或等于5.60%，准确度为±8.80%。对于0.2 ml的样品等分试样，CI-1011的定量下限为0.500 ng ml-1。CI-1011在大鼠血浆中于室温下稳定24小时，在-20℃下至少稳定34天。该测定法已被证明适用于常规定量大鼠血浆中浓度为0.500（柱上100 pg）至500 ng ml-1的CI-1011。本手稿报道了该方法用于测定大鼠血浆中CI-1011浓度的适用性。将使用经过验证的LC/UV测定法获得的，在CMC/Tween悬浮液中每日单次口服给予CI-1011的胆固醇喂养和普通饲料喂养大鼠的血浆样品中的CI-1011浓度，与在0.0806 - 12.3微克ml-1浓度范围内使用报道的LC/MS/MS测定法获得的浓度进行比较。该比较确定的一致性相关系数为0.9977，表明两种测定法获得的CI-1011浓度具有极好的一致性。
