Speciation of Some Biometals in Salmon Egg Cell Cytoplasm by Two-Dimensional HPLC/ICP-MS
Speciation of Some Biometals in Salmon Egg Cell Cytoplasm by Two-Dimensional HPLC/ICP-MS
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DOI：
10.11299/brte.15.345
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发表时间：
2004-12
期刊：
影响因子：
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通讯作者：
Hirotaka Matsuura;Kohei Takatani;Masaru Sugita;Takuya Hasegawa;T. Umemura;H. Haraguchi
中科院分区：
文献类型：
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作者：
Hirotaka Matsuura;Kohei Takatani;Masaru Sugita;Takuya Hasegawa;T. Umemura;H. Haraguchi
建立了一种二维（2D） HPLC/ICP-MS联用系统，并将其应用于鲑鱼卵细胞细胞质中生物金属的形态形成。二维高效液相色谱是基于表面活性剂介导的高效液相色谱和粒径排除色谱相结合的方法。本实验采用双阴离子胆汁酸衍生物CHAPS（3-[（3-胆酰胺丙基）二甲酰胺]-1-丙磺酸）包被的ODS柱作为一维表面活性剂介导的高效液相色谱分离柱。表面活性剂介导的高效液相色谱可以同时分离大分子和小分子。在二维高效液相色谱中，从第一维无机盐中分离出来的蛋白质被SEC柱进一步分离。建立的二维高效液相色谱/ICP-MS系统可提高鲑鱼卵细胞质中金属蛋白的SEC分离的分辨率。用SEC/ICP-MS分析时，在SEC柱总排除限（bbb300 kDa）处，鲑鱼卵细胞细胞质中含铁和锌的金属蛋白只有一个大峰。然而，在目前的二维HPLC/ICP-MS测量中，这些金属蛋白可以被检测到两个峰；铁蛋白的分子量分别为100 ~ 300 kDa和100 kDa，锌蛋白的分子量分别为150 ~ 300 kDa和300 kDa。
A two-dimensional (2D) HPLC/ICP-MS hyphenated system was developed and applied to speciation of biometals in salmon egg cell cytoplasm. The 2D HPLC was based on the combination of surfactant-mediated HPLC and size exclusion chromatography (SEC). In the present experiment, an ODS column coated with CHAPS (3-[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonate), which is a zwitterionic bile acid derivative, was employed as a separation column in the first-dimensional surfactant-mediated HPLC. The surfactant-mediated HPLC allowed simultaneous separation of large and small molecules. In the second-dimensional HPLC, proteins thus separated from inorganic salts in the first dimension were further separated by the SEC column. The present 2D HPLC/ICP-MS system was available for the improvement of resolution in SEC separation of metalloproteins in salmon egg cell cytoplasm. Metalloproteins including Fe and Zn in salmon egg cell cytoplasm were detected as only one large peak at the volume of total exclusion limit of SEC column (> 300 kDa), when analyzed by SEC/ICP-MS. However, these metalloproteins could be detected as two peaks in the present 2D HPLC/ICP-MS measurements; the molecular weights of Fe-proteins were> 300 kDa and 100 kDa, and those of Zn-proteins were> 300 kDa and 150 kDa.