LC-MS/MS法测定人乳汁中非诺贝特酸浓度的方法学建立及其在高甘油三酯血症产妇中的应用

Determination of Fenofibric Acid in Breast Milk by LC-MS/MS and Its Application in Breastfeeding Women with Hypertriglyceridemia

关键词:  非诺贝特  非诺贝特酸  乳汁  液相色谱-串联质谱  高甘油三酯血症

Key words:  Fenofibrate  Fenofibric acid  Breast milk  Liquid chromatography-tandem mass spectrometry  Severe gestational hypertriglyceridemia

摘要:

目的 建立测定人乳汁中非诺贝特酸浓度的高效液相色谱-串联质谱法并评估其临床应用。方法 以非诺贝特酸-D6为内标物，蛋白沉淀法预处理乳汁，采用ACE Excel 3 C18色谱柱，以甲酸溶液(甲酸:水=100 : 0.1)和甲醇为流动相进行梯度洗脱，流速0.4 mL·min?1，进样量1.0 μL，分析时间5 min。以电喷雾离子源，正离子多反应监测模式扫描，定量分析离子对分别为m/z 319.1→233.1(非诺贝特酸)和m/z 325.3→233.0(内标物)。对所建LC-MS/MS法进行方法学验证，并用于检测5例产妇乳汁中非诺贝特酸浓度。结果 非诺贝特酸在1~500 ng·mL-1范围内线性关系良好(r2=0.9988)；质控样本的批内、批间精密度RSD≤5.8%，平均提取回收率和基质效应分别为96.74%~108.50%和101.01%~102.70%。该方法成功应用于5例产妇乳汁中药物浓度测定，结果发现产妇首次分泌的乳汁中非诺贝特酸浓度范围为8.38~442.00 ng·mL-1，停药5 d后浓度范围为2.67~60.63 ng·mL-1。结论 本研究建立的乳汁中非诺贝特酸浓度检测方法简单、经济且灵敏，有助于评估非诺贝特在哺乳期应用的安全性。

Abstract:

OBJECTIVE To establish a high-performance liquid chromatography-tandem mass spectrometry method for the determination of fenofibrate acid concentration in human breast milk and asses the postpartum exposure to fenofibric acid. METHODS Fenofibrate acid-D6 was used as internal standard and protein precipitation was used for extraction, followed by separation on an ACE Excel 3 C18 column with gradient elution of formic acid solution (formic acid: water=100:0.1) and methanol. The flow rate was 0.4 mL·min?1 ,injection volume was 1.0 μL and analysis time was 5 min. The detection of the analytes was performed by electrospray ionization in positive mode by multiple reaction monitoring with the transition of m/z 319.1→233.1 (fenofibrate acid) and m/z 325.3→233.0 (internal standard). The established LC-MS/MS method was researched in methodology and used to determine the fenofibrate acid concentrations in the breast milk of 5 lactating patients. RESULTS The linear range of fenofibrate acid was 1-500 ng·mL-1 (r2=0.9988). The intra-assay and inter-assay precisions of quality control samples were both≤5.8%. The mean recovery and matrix effect were 96.74%-108.50% and 101.01%-102.70%, respectively. This method was successfully applied to the determination of fenofibrate acid concentration in breast milk of 5 lactating patients. Fenofibrate acid concentration ranged from 8.38 to 442 ng·mL-1 in the first breast milk sample, and 2.67 to 60.63 ng·mL-1 on day 5 after fenofibrate withdrawal. CONCLUSION This established method for quantification of fenofibrate acid in human milk is simple, economical and sensitive, which helps us to understand the safety of fenofibrate use during lactation.