イオン交換クロマトグラフィーによるヌクレオチドの迅速分析法
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概要
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The rapid chromatographic methods for the separation of nucleotides such as CMP, UMP, IMP, GMP, AMP, ADP, ATP and Ino have been studied and applied to the determination of nucleotides in fish tissue.<BR>The rapid separations were achieved at high flow velocities of 0.71.1 cm/sec, by using the capillary columns of φ1.02.5 mm×150450 mm packed with AG 1-X4(Cl, -400 mesh) or Dowex 1-X4(Cl, 200400 mesh) and pH-concentration gradient elution with hydrochloric acid-sodium chloride buffers. The gradients from 10<SUP>-3</SUP><I>M</I> sodium chloride at pH 3 to 3×10<SUP>-1</SUP><I>M</I> at pH 2, formed in a multi-chambered gradient device, were used. The column effluent was monitored by an ultraviolet flow spectrophotometer with 3 wavelengths selector(250, 260, 280 mμ).<BR>Under the separating conditions of methods 1 and 1' shown in Table III, using a column of φ 2.5 mm×270 mm and gradient G-2, a mixture of Ino, IMP, CMP, AMP, ADP and ATP was completely separated within approximately 9' minutes. Under the conditions of methods 2 and 2' using a short column of φ 2.5 mm×150 mm and gradient G-3 made with small volumes of eluting agents, the time required for the complete spearation was approximately 60 minutes. For separating a mixture of Ino, IMP, CMP, AMP, ADP, ATP, UMP and GMP, the conditions of methods 4 and 5 using gradients of a type different from G-2 were suitable, and the time required was approximately 120 minutes. Under the condition of method 5 using a column of φ 1 mm×450 mm, the sensitivity of the nucleotides determined by the height-width method was 4.4 times higher than method 4.
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