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本研究建立并验证了测定糖尿病大鼠血浆中二甲双胍浓度的高效液相色谱方法,以阿替洛尔为内标,加入10%高氯酸沉淀蛋白来制备血浆样品。色谱条件:AQ-C18极限色谱柱(250mm×4.6mm,5μm),流动相(pH5.05)为乙腈-水(31:69,v/v,水相中含有0.002M十二烷基磺酸钠,0.0125M磷酸二氢钾,0.015M三乙胺),流速1.0mL/min。二甲双胍在7.5-4000ng/mL范围内具有良好的线性关系(r>0.994),最低检测限(LLOQ)为7.5ng/mL,描述精密度的相对标准偏差(RSD)范围为1.87%-15.70%,准确度为93.98%-106.89%。二甲双胍和内标的回收率分别为95.40%和95.31%。不同实验条件下质控样品的稳定性相对偏差范围在±9.00%之内。该方法成功地运用到糖尿病大鼠单剂量灌胃10mg/kg二甲双胍后的药代动力学研究中,结果显示本研究采用AQ-C18极限色谱柱建立的离子对色谱法对强极性化合物如二甲双胍具有良好的分离效果。
In this study, we established and validated the HPLC method for the determination of metformin in the plasma of diabetic rats. Atenolol was used as internal standard, and 10% perchloric acid precipitated protein was added to prepare plasma samples. Chromatographic conditions: AQ-C18 limit column (250 mm × 4.6 mm, 5 μm) with a mobile phase (pH5.05) of acetonitrile-water (31: 69, v / v, 0.002 M dodecylsulfonic acid Sodium, 0.0125 M potassium dihydrogen phosphate, 0.015 M triethylamine) at a flow rate of 1.0 mL / min. Metformin had a good linearity (r> 0.994) with a LLOQ of 7.5 ng / mL in the range of 7.5-4000 ng / mL, with a relative standard deviation (RSD) ranging from 1.87% to 15.70% The accuracy is 93.98% -106.89%. Metformin and internal standard recoveries were 95.40% and 95.31% respectively. The relative deviation of the stability of the control samples under different experimental conditions is within ± 9.00%. The method was successfully applied to the pharmacokinetics of diabetic rats after single-dose oral administration of 10mg / kg metformin, the results showed that this study using AQ-C18 limit column ion-pair chromatography for strongly polar compounds such as metformin Has a good separation effect.