Murko Simona, Milacic Radmila, Kralj Bogdan, Scancar Janez
Department of Environmental Sciences, Jozef Stefan Institute, Jamova 39, 1000 Ljubljana, Slovenia.
Anal Chem. 2009 Jun 15;81(12):4929-36. doi: 10.1021/ac9006232.
A new analytical procedure using separation support based on Convective Interaction Media (CIM) was developed for speciation of Al in human serum at normal concentration levels. The separation of proteins was performed on a weak anion exchange CIM diethylamine monolithic column. Isocratic elution with buffer A (0.05 M tris(hydroxymethyl)aminomethane-hydrochloric acid + 0.03 M sodium hydrogen carbonate) was applied for 5 min, followed by linear gradient elution from 100% buffer A to 100% buffer B (buffer A + 1 M ammonium chloride) for the next 40 min. Separation of proteins was followed by UV detection at 278 nm. Separated Al species were detected online by inductively coupled plasma mass spectrometry. It was experimentally proven that 91 +/- 7% of Al in human serum was eluted under the transferrin peak. Transferrin was identified on the basis of the retention volume and by ACQUITY ultraperformance liquid chromatography-electrospray ionization mass spectrometry. The problem of extraneous contamination with Al was successfully overcome by using efficient cleaning procedures of eluents and chromatographic supports. The efficient cleaning was of paramount importance to perform Al speciation at extremely low concentration levels. The repeatability of measurement tested for six consecutive separations of unspiked serum was +/-8.6%. The limits of detection and quantification (based on 3 and 10 s of the blank) were 0.15 and 0.49 ng mL(-1) Al bound to transferrin, respectively. This is the first report on quantitative and reliable speciation of Al in human serum at normal concentration levels.
开发了一种基于对流相互作用介质（CIM）分离支持物的新分析方法，用于在正常浓度水平下对人血清中的铝进行形态分析。蛋白质的分离在弱阴离子交换CIM二乙胺整体柱上进行。用缓冲液A（0.05 M三（羟甲基）氨基甲烷 - 盐酸 + 0.03 M碳酸氢钠）等度洗脱5分钟，随后在接下来的40分钟内从100%缓冲液A到100%缓冲液B（缓冲液A + 1 M氯化铵）进行线性梯度洗脱。蛋白质分离后在278 nm处进行紫外检测。分离出的铝形态通过电感耦合等离子体质谱在线检测。实验证明，人血清中91±7%的铝在转铁蛋白峰下洗脱。根据保留体积并通过ACQUITY超高效液相色谱 - 电喷雾电离质谱法鉴定转铁蛋白。通过使用高效的洗脱液和色谱支持物清洁程序，成功克服了铝的外来污染问题。高效清洁对于在极低浓度水平下进行铝形态分析至关重要。对未加标血清连续六次分离测试的测量重复性为±8.6%。与转铁蛋白结合的铝的检测限和定量限（基于空白的3倍和10倍标准偏差）分别为0.15和0.49 ng mL⁻¹。这是关于正常浓度水平下人血清中铝的定量和可靠形态分析首份报告。
