Direct detection of biomolecules in a capillary electrophoresis-chemiluminescence detection system.
Direct detection of biomolecules in a capillary electrophoresis-chemiluminescence detection system.
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DOI：
10.1021/ac030344i
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发表时间：
2004-08
影响因子：
7.4
通讯作者：
K. Tsukagoshi;Koji Nakahama;R. Nakajima
中科院分区：
文献类型：
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作者：
K. Tsukagoshi;Koji Nakahama;R. Nakajima
使用毛细管电泳-化学发光检测系统直接检测生物分子，例如α-氨基酸、肽和蛋白质，其中利用鲁米诺-过氧化氢-Cu(II)催化的化学发光反应。生物分子在毛细管中迁移，与鲁米诺和运行缓冲液中包含的 Cu(II) 催化剂混合。将毛细管出口插入具有补充过氧化氢的电解质溶液的间歇式化学发光检测池中。在毛细管出口尖端观察到化学发光。由于Cu(II)对鲁米诺-过氧化氢化学发光的催化活性增强，出现了生物分子的化学发光峰。当 Cu(II) 与生物分子相互作用形成 Cu(II)-生物分子复合物时，其催化活性更高。本研究采用毛细管电泳-化学发光检测系统直接分离和检测生物分子。检查了 20 种 α-氨基酸、4 种肽和 11 种蛋白质。其中大多数都具有令人满意的 CL 强度响应。谷氨酸是一种 α-氨基酸，检测浓度范围为 2.0 x 10(-7) 至 1.2 x 10(-5) M，检测限 (S/N = 3) 为 1.0 x 10(-7) M (0.6 fmol)。甘氨酰甘氨酸是一种肽，检测浓度范围为 1.7 x 10(-7) 至 1.2 x 10(-5) M，检测限 (S/N = 3) 为 1.7 x 10(-7) M (0.9 fmol)。血红蛋白是一种血红素蛋白，其中血红素结构具有独立的催化活性，检测浓度范围为 1.2 x 10(-7) 至 1.0 x 10(-5) M，检测限 (S/N = 3) 为 1.2 x 10(-7) M (0.6 fmol)。 α-氨基酸和肽的代表性混合物能够以优异的分离度被很好地检测到。
Direct detection of biomolecules, such as alpha-amino acids, peptides, and proteins, was accomplished using a capillary electrophoresis-chemiluminescence detection system, in which a luminol-hydrogen peroxide-Cu(II)-catalyzed chemiluminescence reaction was utilized. Biomolecules migrated in the capillary, where they mixed with luminol and the Cu(II) catalyst included in the running buffer. The capillary outlet was inserted into a batch-type chemiluminescence detection cell with hydrogen peroxide-supplemented electrolyte solution. Chemiluminescence was observed at the tip of the capillary outlet. The chemiluminescence peak from biomolecules appeared due to the enhancement of Cu(II) catalytic activity for luminol-hydrogen peroxide chemiluminescence. The Cu(II) was more catalytically active when it interacted with biomolecules forming Cu(II)-biomolecule complexes. In this study, biomolecules were directly separated and detected in a capillary electrophoresis-chemiluminescence detection system. Twenty alpha-amino acids, 4 peptides, and 11 proteins were examined. Most of them were detected with satisfactory CL intensity response. Glutamic acid, an alpha-amino acid, was detected at concentrations ranging from 2.0 x 10(-7) to 1.2 x 10(-5) M with a detection limit (S/N = 3) of 1.0 x 10(-7) M (0.6 fmol). Glycylglycine, a peptide, was detected at concentrations ranging from 1.7 x 10(-7) to 1.2 x 10(-5) M with a detection limit (S/N = 3) of 1.7 x 10(-7) M (0.9 fmol). Hemoglobin, a heme protein, in which the heme structure was independently catalytically active, was detected at concentrations ranging from 1.2 x 10(-7) to 1.0 x 10(-5) M with a detection limit (S/N = 3) of 1.2 x 10(-7) M (0.6 fmol). Representative mixtures of alpha-amino acids and peptides were well detected with superior separation.