UPLC-MS/MS法同时测定人血浆中5-FU及其代谢产物5-FUH2的浓度及应用
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篇名: UPLC-MS/MS法同时测定人血浆中5-FU及其代谢产物5-FUH2的浓度及应用
TITLE: Simultaneous Determination of 5-FU and Its Metabolites 5-FUH2 in Human Plasma by UPLC-MS/MS and Appli- cation
摘要: 目的:建立同时测定人血浆中5-氟尿嘧啶(5-FU)及其代谢产物5-氟-5,6-二氢尿嘧啶(5-FUH2)浓度的方法,并应用于临床。方法:血浆样品经乙酸乙酯二次萃取处理后,以5-溴尿嘧啶(5-Bru)为内标,采用超高效液相色谱-串联质谱技术(UPLC-MS/MS)测定。色谱柱为AcquityUPLCHSST3,流动相为甲醇-水(30∶70,V/V),流速为0.3mL/min,柱温为20℃,进样量为5μL。采用电喷雾离子源,以多反应监测进行负离子扫描,毛细管电压为1.5kV,锥孔电压为20V,脱溶剂温度为450℃,脱溶剂气流为850L/h,锥孔气流速为50L/h;用于定量分析的离子对分别为m/z129.00→41.90(5-FU)、m/z130.87→82.92(5-FUH2)、m/z189.00→42.10(5-Bru)。选择2020年8-10月哈尔滨医科大学附属肿瘤医院接受5-FU持续静脉滴注46h为基础化疗方案治疗的晚期结直肠癌患者10例,测定其持续静脉滴注18~30h时的5-FU稳态血药浓度及5-FUH2血药浓度,计算5-FU药-时曲线下面积(AUC)和5-FUH2/5-FU浓度比值。结果:5-FU、5-FUH2检测质量浓度的线性范围均为20~1000ng/mL(R2均大于0.990),定量下限均为20ng/mL;精密度试验的RSD均小于20%,相对误差在±10%范围内;提取回收率、基质效应均不影响待测物的测定。10例晚期结直肠癌患者中,5-FU稳态血药浓度为180.04~622.83ng/mL,AUC为8.28~28.65mg·h/L;5-FUH2血药浓度为336.48~948.43ng/mL;5-FUH2/5-FU浓度比值为0.93~4.21;10例患者的5-FUAUC存在约3~4倍的个体差异。结论:所建方法精密度、准确性均较好,灵敏度较高且操作简便,可用于晚期结直肠癌患者体内5-FU血药浓度的监测。
ABSTRACT: OBJECTIVE:To establish the method for simultaneous determination of 5-fluorouracil(5-FU)and its metabolites 5-fluoro-5,6-dihydrouracil (5-FUH2) in human plasma ,and apply it in the clinic. METHODS :After plasma samples were processed twice by ethyl acetate ,UPLC-MS/MS method was adopted using 5-bromouracil (5-Bru) as internal standard. The determination was performed on Acquity UPLC HSS T 3 column with mobile phase consisted of methanol-water (30 ∶ 70,V/V)at the flow rate of 0.3 mL/min. The column temperature was 20 ℃,and sample size was 5 μL. An electrospray ion source was used to carry out negative ion scanning with multiple reaction monitoring. The capillary voltage was 1.5 kV;the taper hole voltage was 20 V;the desolvent temperature was 450 ℃;the desolvent air flow was 850 L/h;the cone hole gas velocity was 50 L/h. The ion transitions for quantitative analysis were m/z 129.00→41.90(5-FU),m/z 130.87→82.92(5-FUH2),m/z 189.00→42.10(5-Bru), respectively. From Aug. to Oct. 2020,10 patients with advanced colorectal cancer were treated with continuous intravenous drip of 5-FU for 46 hours were collected from Harbin Medicinal University Cancer Hospital. Steady-state plasma concentration of 5-FU and plasma concentration of 5-FUH2 were determined at 18-30 h of continuous intravenous drip. The area under the curve (AUC)for 5-FU and concentration ratio of 5-FUH2/5-FU were calculated. RESULTS :The linear range of 5-FU and 5-FUH2 were 20 to 1 000 μg/mL(R 2>0.990). The quantification limits were 20 ng/mL. RSDs of precision test were all lower than 20%,and relative error ranged ±10%. The extraction recovery and matrix effects didn ’t affect the determination of substance to be measured. Among 10 patients with advanced colorectal cancer ,the steady-state concentration of 5-FU were 180.04-622.83 ng/mL,and AUC of 5-FU ranged from 8.28 to 28.65 mg·h/L. The concentration of 5-FUH2 ranged 336.48-948.43 ng/mL,and concentration ratio of 5-FUH2/ 5-FU ranged 0.93-4.21. AUC of 5-FU in 10 patients had about 3-4 fold individual differences. CONCLUSIONS :The established method has good precision and accuracy ,high sensitivity ,and simple operation. It can be used for plasma monitoring of 5-FU in patients with advanced colorectal cancer.
期刊: 2021年第32卷第04期
作者: 佟志强,吴东媛,张娟,程梦飞,张硕,董梅
AUTHORS: TONG Zhiqiang ,WU Dongyuan ,ZHANG Juan,CHENG Mengfei ,ZHANG Shuo,DONG Mei
关键字: 超高效液相色谱-串联质谱法;5-氟尿嘧啶;5-氟-5,6-二氢尿嘧啶;血药浓度
KEYWORDS: UPLC-MS/MS; 5-FU; 5-FUH2; Plasma
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