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目的建立测定海马神经细胞内外地西泮、氟硝西泮和劳拉西泮含量的方法,并比较海马神经细胞模型对这3种药物的吸收利用差异。方法采用HPLC-CAD法测定含量。色谱柱:Capcell C_(18)(250 mm×4.6 mm,5μm),流动相:乙腈-水(44∶56),柱温30℃,流速1.0 m L·min~(-1),CAD检测器,检测波长254 nm,采用外标法定量。结果 3种苯二氮类药物与培养基及细胞中的其他成分能很好分离。地西泮、氯硝西泮与劳拉西泮分别在质量浓度1.83~183.16、1.81~190.04、1.74~173.56 mg·L~(-1)时呈线性关系(r>0.999 0),方法平均加样回收率分别为99.99%、100.42%和99.98%。给药培养24 h的海马神经细胞对地西泮、氟硝西泮和劳拉西泮的平均吸收率:正常组分别为45.11%、32.60%和29.73%,模型组分别为31.30%、25.98%和21.81%;平均转化利用率:正常组分别为72.98%、73.16%和74.90%,模型组分别为78.89%、83.04%和85.01%。结论正常组对3种药物的吸收效率比模型组更高,而模型组对吸收的药物转化利用率比正常组更高。HPLCCAD法同时测定海马神经细胞内外液中3种苯二氮药物浓度的方法准确、快速、简便,可为苯二氮类药物在海马神经细胞内的药物动力学研究提供检测方法。
Objective To establish a method for determination of diazepam, flunitrazepam and lorazepam in hippocampal neurons, and to compare the absorption and utilization of these three drugs in hippocampal neuronal models. Methods HPLC-CAD method was used to determine the content. The mobile phase consisted of acetonitrile-water (44:56), the column temperature was 30 ℃ and the flow rate was 1.0 mL · min -1. , Detection wavelength 254 nm, using external standard method. Results The three benzodiazepines were well separated from other components of the culture medium and cells. Diazepam, clonazepam and lorazepam showed a linear relationship (r> 0.999 0) at the mass concentrations of 1.83 ~ 183.16, 1.81 ~ 190.04, 1.74 ~ 173.56 mg · L -1, The sample recovery rates were 99.99%, 100.42% and 99.98% respectively. The mean absorption rates of diazepam, flunitrazepam and lorazepam in hippocampal neurons cultured for 24 h were 45.11%, 32.60% and 29.73% respectively in the normal group, 31.30% and 25.98% in the model group, respectively And 21.81%, respectively. The average transformation utilization rate was 72.98%, 73.16% and 74.90% in the normal group and 78.89%, 83.04% and 85.01% in the model group respectively. Conclusion The absorption efficiency of the three drugs in the normal group is higher than that in the model group, while that in the model group is higher than that in the normal group. The method of simultaneous determination of three benzodiazepines in hippocampal neurons by HPLCCAD method is accurate, rapid and simple, and can be used to detect the pharmacokinetics of benzodiazepines in hippocampal neurons.