高效液相色谱法同时测定人血清中茶碱及其代谢物1,3-二甲基尿酸浓度

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，5 μm），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20 μL。结果：茶碱、1，3-二甲基尿酸的血清浓度线性范围分别为0.50~40.00 μg·mL^-1和0.10~12.80 μg·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0.50，0.10 μg·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。

Simultaneous detection of concentrations of theophylline and its metabolite 1, 3-dimethyluric acid in human serum by HPLC

OBJECTIVE To establish a method for simultaneous detection of serum concentrations of theophylline and its metabolite 1,3-dimethyluric acid in human.METHODS Serum protein was removed by acetonitrile precipitation,4-Acetamidophenol was added as internal standard substance,then its concentration and dissolution were detected by HPLC.HPLC was performed on Agilent ZORBAX SB-C₁₈ reversed phase column (150 mm×4.6 mm,5 μm) with a mobile phase consisting of acetonitrile and 0.3% formic acid (5:95).The flow rate was 1 mL·min^-1 and the detection wavelength was 280 nm,and injection volume was 20 μL.RESULTS The calibration graphs were linear in range of 0.50-40.00 μg·mL^-1 for theophylline and 0.10-12.80 μg·mL^-1 for 1,3-dimethyluric acid (R=0.998 8 and 0.999 7,respectively).Limits of quantification of theophylline and 1,3-dimethyluric acid were 0.50 and 0.10 μg·mL^-1,respectively.Extraction recoveries of low,medium and high concentrations were 80.22%,87.25% and 89.23% for theophylline,and 87.03%,80.01% and 87.95% for 1,3-dimethyluric acid,respectively.Either intra-or inter-batch RSD was less than 15%.CONCLUSION HPLC is simple,sensitive and reliable for detection of serum concentrations of theophylline and its metabolite 1,3-dimethyluric acid in human.Therefore,it is suitable to evaluate and research effects of drug on pharmacokinetics of theophylline in human,and direct reasonable medication in clinical practice.