HPLC梯度洗脱法同时测定人血浆中对乙酰氨基酚和维生素C浓度

Simultaneous Determination of Paracetamol and Vitamin C in Human Plasma by HPLC Method with Gradient Elution

目的建立一种同时测定血浆中对乙酰氨基酚（PAC）和维生素C（vitC）浓度的梯度洗脱高效液相色谱法，以用于二者组成的复方制剂的体内评价研究。方法血浆样品以高氯酸直接沉淀蛋白后采用HPLC-UV结合梯度洗脱法在Hypersil ODS柱上分离测定，流动相为磷酸缓冲盐（A）和甲醇（B），流速1．0mL·min^-1。洗脱条件：t=0，A=100％；t=1．5min，A=82％，B=18％；t=-7．0min，A=100％，检测波长245nm。结果对乙酰氨基酚与维生素c分离度好，内源性杂质不干扰测定。在浓度0．625-40mg·L^-1内，对乙酰氨基酚和维生素C的浓度（C）与各自的峰面积（Y）线行关系良好，最低定量限均为0．0625mg·L^-1。绝对回收率分别为对乙酰氨基酚：84．9％～97．2％m=5）；维生素C：81．9％～91.3％（n=5）。方法回收率分别为PAC：94．9％～108．2％（n=15）；维生素C：105．7％-108．9％（n=15）。对乙酰氨基酚和VitC的E1内RSD分别为0．53％-4．23％（n=15）和0．57％-3．79％（n=15）；日间RSD分别为3．15％～9．79％（n=15）和6．79％～7．72％（n=15）。结论建立的高效液相梯度洗脱法灵敏、简便、快速、准确，适用于两者组成的复方制剂的体内过程研究。

OBJECTIVE To develop a high performance liquid chromatographic （HPLC） method with gradient elution for simultaneous determination ofparacetamol and vitamin C in human plasma, and to study the pharmacokinetics ofparacetamol and vitamin C. METHODS The samples were separated on a Hypersil ODS colunm by a gradient elution. The mobile phase of phosphate buffer （A） and methanol （B） at a flow rate of 1.0 mL·min^-1. RESULTS Paracetamol and vitamin C were separated well, the calibration curves were linear with the range from 0.625 to 40 mg.Ll, and the limit detection was 0.062 5 mg·-L^-1 .The absolute recoveries of paracetamol and vitamin C respectively were in the range of 84.9%-97.2%（n=5） and 81.9%-91.3%（n=5）, the method recoveries were in the range of 94.9%-108.2%（n=5） and 105.7%-108.9%（n=5） respectively. The intra-day RSDs were 0.53%-4.23%（n=5） and 0.57%-3.79%（n=5）, inter-day RSDs were 3.15%-9.79%（n=5） and 6.79%-7.72%（n=5）, respectively. CONCLUSION The established HPLC method with gradient elution was accurate, sensitive, and was suitable for the pharmacokinetic study ofparacetamol and vitamin C in human.