Determination of erythrocyte methotrexate polyglutamates by liquid chromatography/tandem mass spectrometry after low-dose methotrexate therapy in Chinese patients with rheumatoid arthritis
Determination of erythrocyte methotrexate polyglutamates by liquid chromatography/tandem mass spectrometry after low-dose methotrexate therapy in Chinese patients with rheumatoid arthritis
复制标题
液相色谱/串联质谱法测定中国类风湿关节炎患者低剂量甲氨蝶呤治疗后红细胞甲氨蝶呤聚谷氨酸含量。
DOI：
10.1016/j.jchromb.2012.08.032
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发表时间：
2012-10-15
影响因子：
3
通讯作者：
Chen, Xiao
中科院分区：
文献类型：
--
作者：
Mo, Xiaolan;Wen, Yuguan;Chen, Xiao
甲氨蝶呤（MTX）是目前应用最广泛的治疗类风湿性关节炎（RA）的药物之一，通过甲氨蝶呤多谷氨酸（MTXPGs）的聚谷氨酸化，将序列γ -连锁谷氨酸残基附着在MIX上。建立了一种新的、灵敏的LC/MS/MS测定全血MIX和总MTX （MTX + MTXPGs）的方法，并进行了验证，计算MTXPGs的浓度。测定全血MTX，全血用50%三氟乙酸沉淀，用乙酰乙酸乙酯提取。采用正电喷雾电离法对分析物进行LC/MS/MS分析。为了测定全血总MIX，将全血与抗坏血酸（200 mM）在37℃下孵育3小时，酶促mtxpg转化为MIX，然后用上述方法处理。在ppb （ng/mL）水平上加标MIX的加标回收率为26.2% ~ 37.8%，日内精密度小于15.8%，日内精密度小于11.8%。检测下限为0.5 ng/mL，定量下限为1 ng/mL。灵敏的LC/MS/MS方法具有高选择性和可接受的准确度和精密度，成功应用于RA患者红细胞甲氨蝶呤多谷氨酸的测定。(C) 2012 Elsevier B.V.版权所有
Methotrexate (MTX) is currently one of the most widely used drugs for treatment of rheumatoid arthritis (RA) through polyglutamation of methotrexate polyglutamates (MTXPGs), a process attaching sequential gamma-linked glutamic residues to MIX. A new and sensitive LC/MS/MS method was developed and validated for determination of whole-blood MIX and total MTX (MTX + MTXPGs), and then concentration of MTXPGs was calculated. To determine whole-blood MTX, whole blood was precipitated with 50% trifluoroacetic acid, and extraction was performed using ethyl acetoacetate. Analytes were subjected to LC/MS/MS analysis using positive electrospray ionization. To determine whole-blood total MIX, whole blood was incubated with ascorbic acid (200 mM) at 37 degrees C for 3 h to enzymatically convert the MTXPGs to MIX, and then processed with the same method mentioned above. Recoveries of spiked MIX at ppb (ng/mL) level were between 26.2% and 37.8% with intra- and inter-day precision less than 15.8% and 11.8%, respectively. The lower limit of detection (LLOD) and lower limit of quantitation (LLOQ) were 0.5 ng/mL and 1 ng/mL, respectively. The sensitive LC/MS/MS method was fully validated with high selectivity and acceptable accuracy and precision, which was successfully applied to determine the erythrocyte methotrexate polyglutamates in patients with RA. (C) 2012 Elsevier B.V. All rights reserved.