GC／MS法测定人血浆中地芬尼多药物浓度及其在药代动力学研究中的应用

目的建立GC/MS法测定人血浆中地芬尼多的含量,用于药代动力学研究中血药浓度的测定。方法液-液萃取法进行血样前处理,分析采用（15 m×0.25 mm×0.25μm）DB-17MS毛细管柱,载气为氦气,载气流速为1.0 ml/m in;进样口温度250℃,炉温升程：100℃→280℃（按25℃/m in的速度升温,且在280℃保持1 min）再升至300℃（按50℃/min的速度升温,且在300℃保持1.6 m in）;MS源温度230℃,四级杆温度150℃,电子轰击能量为70 eV。结果地芬尼多及内标的保留时间分别为7.8 min及7.97 min,其定量碎片基峰分别为m/z98和138。地芬尼多在1300 ng/ml浓度范围内,浓度与响应呈良好的线性关系（r2=0.999）;定量下限为1 ng/ml,准确度在82%104%范围内;地芬尼多的提取回收率为72.7%90.0%;批内和批间RSD分别在2.7%和3.9%以内,准确度在101.5%114.1%范围内;地芬尼多人血浆样品在-20℃保存10 d及在室温放置24 h稳定;其甲醇溶液在4℃冰箱保存21 d稳定。结论本研究建立的GC/MS方法测定人血浆中地芬尼多浓度,特异性好、灵敏度高,适用于盐酸地芬尼多口崩片在健康受试者体内的药代动力学研究。

GC/MS Method for Determination of Difenidol in Chinese Healthy Volunteers and its application in pharmacokinetics

Objective To establish a GC/MS method for determination of Difenidol in Chinese healthy volunteers.Methods Difenidol and the internal standard were extracted from plasma by liquid-liquid extraction and separated on the DB-17MS column（15 m×0.25 mm×0.25 μm）.Helium was used as the carrier gas at a flow-rate of 1 ml/min.An Agilent 5973 MS was used to analyze samples and was operated in the EI mode.The base peak was performed to quantify Difenidol.The oven temperature program was as follows： isothermal at 100℃,was heated at 25℃/min to 280℃ and maintained for 1 min,before being heated at 50℃/min to 300℃ and maintained for 1.6 min.The injection temperature was 250℃,MS source temperature 230℃ and Quadrupole temperature was 150℃.The EI energy was set at 70 eV.Results Linear calibration curves were obtained within the concentration range of 1-300 ng/ml.The regression equation of this curve and its correlation coefficient（r2） were calculated as follows： Y=2.05X-2.36×10-3,r2=0.999.The limit of quantification was 1 ng/ml（n=5） for Difenidol,and acceptable accuracy and precision（below 15%） at this level were obtained.The intra-and inter-day relative standard deviation（RSD） was less than 3.9%.The accuracy of the assay was determined at three concentration levels and was found in the range from 101.5% to 114.1%.Conclusions The validation results showed that the method is sensitive,specific and reproducible,proved to be suitable for pharmacokinetic investigations of Difenidol in Chinese healthy volunteers.