氢氧化钡水解HPLC法测定碘化酪蛋白中甲状腺素的含量

High Performance Liquid Chromatographic Determination of Thyroxine in Iodinated Casein with Barium Hydroxide Method

建立了碘化酪蛋白中甲状腺素 (T4)的氢氧化钡水解HPLC测定方法。实验发现 ,每毫升碘化酪蛋白溶液加入0.5～1.0g的八水合氢氧化钡 ,110℃水解8～12h ,控制用硫酸沉淀钡离子后的pH12～13,碘化酪蛋白释放的甲状腺素达到相对最高稳定得率。使用WatersSymmetryShieldTMRP18色谱柱 (5μm ,3.9mm×150mm) ,乙腈 (B ,0.1 %的甲酸 )和水 (A ,0.1%的甲酸 )为流动相 ,0.8mL·min -1的流速 ,B从10 %到50 %线性洗脱40min ,240nm的紫外吸收波长 ,能使碘化酪蛋白水解液中甲状腺素得到完全分离并定量测定。方法在考察的T4 质量浓度范围内 (0.5～25mg·L-1)线性良好 ,r>0.99 ,检出限可达0.2mg·g-1,实验条件下测定的商品碘化酪蛋白中T4含量为1.0 %

A high performance liquid chromatographic method was developed for the determination of thyroxˉine(T 4 )in iodinated casein(IC).The IC was hydrolyzed with bariumhydroxde solution.It was found that the relative highest stable yield of T 4 from IC was obtained under the following conditions:barium octahydrate amount added into per milliliter of IC solution ranged from0.5to1.0g;the hydrolyzing time at110℃ranged from8to12hours;and the pH range was controlled within12to13after precipitating the bariumions with sulphuric acid.The separation was performed on a Waters Symmetry shield TM RP18column(5μm,3.9mm×150mm).T 4 could be separated completely using phase A(water,0.1%formic acid)and phase B(acetonitrile,0.1%formic acid)as mobile phase at a flowrate of0.8mL/min,and10%to50%of phase B as eluant for40min.The T 4 was then quantitatively determined by external standard method at240nm.A good linear calibration curve was obtained for0.5to25mg·L -1 T 4 ,with a detection limit of0.2mg·g -1 .