RP-HPLC法测定急性淋巴细胞白血病患儿血清中甲氨蝶呤浓度.docVIP

RP-HPLC法测定急性淋巴细胞白血病患儿血清中甲氨蝶呤浓度 刘文? ? 通讯作者：刘文（1978～ ），男，硕士，主管药师，研究方向：临床药学与药代动力学研究 Tel: (0731) Fax: (0731 E-mail: liuwen0917@163.com （湖南省人民医院药学部，410005） 摘　要 目的：建立一快速、准确的RP-HPLC方法测定人血清中甲氨蝶呤的浓度。 方法：以阿魏酸内标，采用Agilent 1100系列高效液相色谱仪，以大连依利特Kromasil C18柱（(250mm×4. 6mm ,5μm)为分析柱；流动相为乙腈：磷酸缓冲盐溶液（pH 6.8）=(15∶85 ,v/ v)；流速1.0 mL·min-1；检测波长：306 nm；柱温30℃。血浆样品用乙腈沉淀蛋白后，取上清液，在60℃下经氮气挥干，残渣用流动相定容，取20μL进样分析。结果：色谱峰分离良好，无干扰。线性方程为：Y=-1.65×10-2X+0.355，r=0.9995，高、中、低三浓度的样品萃取回收率均大于75%。结论：本方法是一种可靠、快速灵敏的检测方法，适用于MTX的血药浓度监测。 关键词：甲氨蝶呤，高效液相色谱，儿童急性淋巴细胞白血病 Determinate the concentration of MTX in the ALL Children by HPLC Liu wen, Deng Nan, Dai Ying-Chun (Department of Pharmacy of People’s Hospital of Hunan Province, Changsha 410005, China) Abstract Objective: To develop a HPLC method to determine the concentration of methotrexatate (MTX) in human plasma. Method: The ferulic acid was used as inter standard (I.S.) The analytical column was a Kromasil C18 column connected with the Agilent 1100 HPLC system. The isocratic mobile phase was the acetonitrile and buffer of phosphate acid (pH 6.8) with the rate of 15:85(V/V). The detective UV wavelength was 306 nm and the column temperature was 30℃. The plasma samples, added ferulic acid as internal standard, were deposited down the protein with acetonitrile and the upper phase was evaporated by nitrogen at 60℃. The remains was dissolved with mobile phase, 20μLwas injected into the sampler. Results: The chromatography was good and not interfered by the components of the plasma. The linear equation was Y=-1.65×10-2X+0.355,r=0.9995,r=0.9995. Conclusion: This is a accurate, sensitive and conventient method for the dertermination of MTX in the therapeutic drug monito Keywords: MTX, HPLC, ALL 甲氨蝶呤 (methot rexate，MTX) 是一种二氢叶酸还原酶抑制剂，是通过抑制细胞内二氢叶酸酶达到杀灭肿瘤细胞的作用。目前临床采用大剂量甲氨蝶呤配合甲酰四氢叶酸钙解救 (HDMTX+CF) 治疗儿童急性淋巴细胞白血病(ALL)[1]。在该方案实施过程中，甲氨蝶呤(MTX)血药浓度对甲酰四氢叶酸钙(CF)剂量及用药次数的选择起着指导作用。MTX在体内部分经肝细胞代谢转化为谷氨酸盐，部分通过胃肠道细菌代谢；MTX 主要经肾排泄，大部分为药物原型。另外有少量药物原型及代谢产物以结合型形式储存于肾脏和肝