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目的:建立一种同时测定人血清样品中茶碱及其代谢物1,3-二甲基尿酸血药浓度的HPLC方法。方法:血清样品经乙腈沉淀除去蛋白,选择对乙酰氨基酚为内标,真空浓缩、复溶后采用HPLC分析。色谱柱为:Agilent ZORBAX SB-C18柱(150 mm×4.6 mm,5μm),流动相为乙腈∶0.3%甲酸溶液(5∶95),检测波长为280 nm,流速为1 mL·min-1,进样量为20μL。结果:茶碱、1,3-二甲基尿酸的血清浓度线性范围分别为0.50~40.00μg·mL-1和0.10~12.80μg·mL-1,两者的线性关系均良好(R分别为0.998 8和0.999 7),最低定量下限分别为0.50,0.10μg·mL-1。茶碱和1,3-二甲基尿酸低、中、高质量浓度的提取回收率分别为80.22%,87.25%,89.23%和87.03%,80.01%,87.95%,二者批内和批间RSD均小于15%。结论:该方法简便、准确,特异性和灵敏度能够满足临床测定人血清中茶碱及其代谢物1,3-二甲基尿酸浓度的要求,适用于评价和研究人体内茶碱的药动学影响,指导临床用药。
Objective: To establish a HPLC method for the simultaneous determination of theophylline and its metabolites 1,3-dimethyl uric acid in human serum samples. Methods: Serum samples were removed by acetonitrile precipitation. Acetaminophen was selected as internal standard and concentrated in vacuo. The reconstituted samples were analyzed by HPLC. The column was Agilent ZORBAX SB-C18 column (150 mm × 4.6 mm, 5 μm) with a mobile phase of acetonitrile: 0.3% formic acid (5:95) at a detection wavelength of 280 nm and a flow rate of 1 mL · min- Injection volume of 20μL. Results: The linear range of theophylline and 1,3-dimethyluric acid was 0.50-40.00μg · mL-1 and 0.10-12.80μg · mL-1, respectively, with good linearity (R = 0.998 8 and 0.999 7), respectively. The lowest limits of quantification were 0.50 and 0.10 μg · mL -1, respectively. The recoveries of theophylline and 1,3-dimethyl uric acid at low, middle and high concentrations were 80.22%, 87.25%, 89.23%, 87.03%, 80.01%, 87.95%, respectively. Less than 15%. Conclusion: The method is simple, accurate, specific and sensitive to meet the clinical determination of the concentration of theophylline and its metabolites 1,3-dimethyl uric acid in human serum requirements for the evaluation and study of human pharmacokinetics of theophylline Influence, guide clinical medication.