高效液相色谱法测定人血中羟氯喹浓度方法学优化及应用性分析

摘要/Abstract

摘要： 目的：建立一种新的高效液相色谱法测定人全血中羟氯喹的浓度，并对方法进行优化。方法：取300 μL全血样本，加入25 μg·mL-1甲硝唑溶液10 μL作为内标，用900 μL乙酸乙酯进行液液萃取，氮气吹干后用流动相200 μL复溶，取20 μL上清液进样。使用Symmetry C18色谱柱（4.6 mm×250 mm，5 μm）进行分离，以20 mmol·L-1磷酸二氢钾-乙腈（85∶15，v/v，磷酸调pH=3.0）为流动相，流速：0.8 mL·min-1，柱温：35 ℃，检测波长：254 nm。考察该方法的特异性、标准曲线、精密度、准确度、稳定性和提取回收率。结果：羟氯喹在100~5 000 ng·mL-1浓度范围内线性关系良好，标准曲线方程为f=0.03×10-1C-0.12×10-1(r=0.999 6)；日内、日间精密度RSD均小于4.0%，准确度高于85%，全血样品提取回收率为（78.08±5.32）%（n=5）。结论：本方法灵敏度高、选择性高、操作简洁、检测时间短、结果准确可靠，能够满足检测人血中羟氯喹浓度的需求，经优化后缩短了检测时间，提升了检测效率，适合于临床诊疗监测及药效学研究。

关键词: font-size:medium, ">高效液相色谱法；人血；羟氯喹；浓度

Abstract: Objective：To develop a new high performance liquid chromatography（HPLC）method for the determination of the concentration of hydroxychloroquine in human blood and optimize the method to improve its efficiency. Methods： 300 μL of blood sample and 10 μL of 25 μg·mL-1metronidazole solution which was used as the internal standard were extracted with 900 μL of ethyl acetate， redissolved with 200 μL of mobile phase after drying with nitrogen， and took 20 μL of supernatant for the determination. Chromatographic separation was performed on a Symmetry C18 column（4.6 mm×250 mm，5 μm）at 35 ℃ using the mobile phase of 20 mmol·L-1 KH2PO4-acetonitrile （85∶15，v/v，pH was adjusted to 3 by H3PO4） at a flow rate of 0.8 mL·min-1 and the detection wavelength was 254 nm. The specificity， standard curve， precision and accuracy， stability and the extraction recovery rate of the method were investigated. Results： The linear range of hydroxychloroquine was 100-5 000 ng·mL-1， and the equation of standard curve was f=0.03×10-1C-0.12×10-1(r=0.999 6). The RSDs of intra-day and inter-day precision were all less than 4.0% and the accuracy was higher than 85%. The average extraction recovery of hydroxychloroquine in blood was （78.08±5.32）%（n=5）. Conclusion： The method established has high sensitivity and selectivity， and is simple， accurate and rapid. It can satisfy the demand of detecting the concentration of hydroxychloroquine in human blood and can be applied in pharmacokinetics and pharmacodynamics study and therapeutic drug monitoring.

Key words: font-size:medium, ">HPLC；Human blood；Hydroxychloroquine；Concentration

中图分类号:

刘思鲁, 胡蕾, 刘博宇, 陶依然, 赵海艳, 孙博洋, 黄琳, 封宇飞. 高效液相色谱法测定人血中羟氯喹浓度方法学优化及应用性分析[J]. 中国医药导刊, 2022, 24(1): 76-80.

LIU Silu, HU Lei, LIU Boyu, TAO Yiran, ZHAO Haiyan, SUN Boyang, HUANG Lin, FENG Yufei. An Optimized HPLC Method for the Determination of Hydroxychloroquine Concentration in Human Blood and its Application Analysis[J]. CHINESE JOURNAL OF MEDICINAL GUIDE, 2022, 24(1): 76-80.

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