群特異的新規高性能充填剤の開発とその生理活性ペプチド・タンパク質の高選択的分離・定量システム構築への応用

概要

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Anhydrochyntotrypsin (AHC), β-cyclodextrin sulfate (β-CD・SUL) and hyaluronic acid (HA) are known to exhibit affinity toward peptides having an aromatic amino acid sequence at their C-termini, heparin-binding proteins and gelatin-binding proteins, respectively. In this study, AHC, β-CD・SUL- and HA-immobilized adsorbents were developed for high-performance affinity chromatography. A column-switching HPLC system consisting of precolumns packed with these novel group-specific adsorbents and a reversed-phase analytical column was also developed forselective on-line enrichment and to separation of bioactive peptides and proteins. Among 14 peptides which contained C-terminal aromatic amino acid sequence and more than 5 amino acidresidues, 10 peptides were retained on the AHC-precolumn almost quantitatively. All of the 10 proteins having heparin-binding activity were retained on the β-CD・SUL-precolumn almost quantitatively, while 5 proteins having no heparin-binding activity were not retained. Similarly, 4 proteins having gelatin-binding activity were retained selectively on the HA-immobilized precolumn. Calibration graphs for these peptides and proteins were observed to be linear, and good reproducibility was obtained. In addition to the above characteristics, their precolumns were stable after analyzing more than 200〜450 samples during at least a period of 1 year.
社団法人日本分析化学会の論文
1999-05-05