The Effect of Famotidine on the Metabolism of Metoprolol Tartrate in Human Liver Microsomes
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摘要:
目的 通过体外人肝微粒体孵育体系考察酒石酸美托洛尔的酶促反应动力学，并探究法莫替丁对酒石酸美托洛尔代谢的影响。 方法 建立LC-MS/MS方法测定人肝微粒体孵育体系中酒石酸美托洛尔的浓度。构建人肝微粒体孵育体系，确定最佳孵育时间和蛋白浓度；采用底物减少法，计算酒石酸美托洛尔的酶促反应动力学参数（Vmax和Km）；将系列浓度的法莫替丁与酒石酸美托洛尔共同孵育，并测定酒石酸美托洛尔的浓度，考察法莫替丁对酒石酸美托洛尔代谢的影响。 结果 酒石酸美托洛尔在人肝微粒体的最佳孵育时间为60 min，最佳蛋白浓度为1.0 mg/mL；酶促反应动力学参数Vmax = 0.07 µmol·min-1·（mg·protein）-1，Km = 7.84 µmol/L。 结论 法莫替丁对酒石酸美托洛尔在人肝微粒体孵育体系中的代谢未产生显著的抑制作用，提示两药联合使用较安全。 Abstract:Objective To explore the enzymatic reaction kinetics of metoprolol tartrate in an in vitro human liver microsome incubation experiment and to explore the effect of famotidine on the metabolism of metoprolol tartrate. Methods An LC-MS/MS method was developed to accurately measure the concentration of metoprolol tartrate in a human liver microsomal incubation system. A human liver microsome incubation system was established to determine the optimal incubation time and protein concentration; using the substrate depletion method, the enzymatic reaction kinetics parameters of metoprolol tartrate （such as Vmax and Km） were calculated. Series concentrations of famotidine were co-incubated with metoprolol tartrate, and the concentration of metoprolol tartrate was measured to evaluate the effect of famotidine on its metabolism. Results The optimal incubation time for metoprolol tartrate in human liver microsomes was found to be 60 minutes, with an optimal protein concentration of 1.0 mg/mL. The enzymatic reaction kinetics parameters were Vmax = 0.07 μmol·min-1·（mg·protein）-1 and Km = 7.84 µmol/L. Conclusion The results indicated that famotidine did not produce a significant inhibitory effect on the metabolism of metoprolol tartrate in human liver microsome incubation system, suggesting that the combination of these two drugs may be relatively safe. -
Key words:
- Metoprolol tartrate /
- Famotidine /
- Human liver microsomes /
- LC-MS/MS /
- Kinetics of enzymatic reactions
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图 1 酒石酸美托洛尔（Ⅰ）和D6-酒石酸美托洛尔（Ⅱ）的典型色谱图
A：空白孵育样品；B：酒石酸美托洛尔 + 内标样品；C：肝微粒体孵育样品。
Figure 1. Typical chromatograms of metoprolol tartrate （Ⅰ） and D6-metoprolol tartrate （Ⅱ）
图 2 孵育时间对酒石酸美托洛尔在肝微粒体中代谢的影响
Figure 2. Effect of incubation time on the metabolism of metoprolol tartrate in liver microsomes
图 3 蛋白浓度对酒石酸美托洛尔在肝微粒体中代谢的影响
Figure 3. Effect of protein concentration on the metabolism of metoprolol tartrate in liver microsomes
图 4 酒石酸美托洛尔在肝微粒体中的酶促反应动力学曲线
Figure 4. Enzymatic reaction kinetics curve of metoprolol tartrate in liver microsomes
图 5 法莫替丁对酒石酸美托洛尔在人肝微粒体中代谢的影响
Figure 5. Effect of famotidine on the metabolism of metoprolol tartrate in human liver microsomes
表 1 酒石酸美托洛尔的精密度与准确度结果（$ \bar x \pm s $，n = 6）
Table 1. Results of precision and accuracy for metoprolol tartrate（$ \bar x \pm s $，n = 6）
理论浓度（µmol/L） 日内精密度 日间精密度 准确度 实测浓度 RSD（%） 实测浓度 RSD（%） 0.36 0.37 ± 0.02 4.60 0.38 ± 0.02 2.51 3.08 0.73 0.70 ± 0.03 4.12 0.71 ± 0.05 0.52 −2.65 5.84 5.81 ± 0.16 2.77 5.76 ± 0.26 9.59 −1.31 9.92 9.97 ± 0.23 2.26 10.04 ± 0.39 4.86 1.13 
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表 2 酒石酸美托洛尔的提取回收率与基质效应结果（$ \bar x \pm s $）
Table 2. Results of extraction recovery and matrix effect for metoprolol tartrate（$ \bar x \pm s $）
理论浓度
（µmol/L）提取回收率 RSD（%） 基质效应 RSD（%） 0.73 94.36 ± 0.04 4.37 97.22 ± 0.01 1.35 5.84 94.75 ± 0.03 2.95 97.13 ± 0.02 1.99 9.92 91.16 ± 0.04 4.15 95.41 ± 0.01 1.10
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表 3 酒石酸美托洛尔的稳定性试验结果（$ \bar x \pm s $，n = 6）
Table 3. Stability test results of metoprolol tartrate（$ \bar x \pm s $，n = 6）
理论浓度 （µmol/L） 实测浓度 RSD（%） 0.73 0.69 ± 0.02 2.76 5.84 5.74 ± 0.01 1.05 9.92 9.81 ± 0.08 0.86
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