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The Journal of Biochemistry

Year: 2010  |  Volume: 147  |  Issue: 4  |  Page No.: 501 - 509

A comprehensive HPLC analytical system for the identification and quantification of hexoses that employs 2-aminobenzamide coupling

K Hasehira, S. i Nakakita, N Miyanishi, W Sumiyoshi, S Hayashi, K Takegawa and J. Hirabayashi


Rare sugars are defined as monosaccharides with extremely low natural abundances. Their natural distributions and biological functions remain to be clarified. To establish a robust analytical system that can separate, identify and quantify rare sugars, 12 d-hexoses—including five rare aldohexoses and three rare ketohexoses—were labelled with 2-aminobenzamide (2-AB), and the fluorescently tagged monosaccharides were separated by high-performance liquid chromatography (HPLC). Because the ketohexoses were much less reactive than were the aldohexoses, the reaction conditions were optimized to achieve the maximum yields (>75%) for both aldohexoses and ketohexoses. The calibration curve determined for the rare ketohexose, d-psicose (Psi), was linear between 1 pmol and 1 µmol and had a correlation coefficient of 0.9999. Using the developed method, the psicose content in a leaf of Itea virginica, in which the presence of psicose has previously been reported, was found to be 2.7 mg psicose/g leaf. The result proved feasibility of the method even for natural products. Because the system is a comprehensive tool, it should help answer questions concerning the biosyntheses and functions of rare hexose sugars.

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