Determination of Saccharides Labelled with a Fluorescent Reagent, DBD-ProCZ, by Liquid Chromatography

✍ Scribed by Toshimasa Toyo'oka; Akiko Kuze

• Publisher: John Wiley and Sons
• Year: 1997
• Tongue: English
• Weight: 154 KB
• Volume: 11
• Category: Article
• ISSN: 0269-3879
• DOI: 10.1002/(sici)1099-0801(199705)11:3<132::aid-bmc633>3.0.co;2-n

✦ Synopsis

Saccharides were determined by high-performance liquid chromatography (HPLC) after precolumn tagging with 4-(2-carbazoylpyrrolidin-1-yl)-7-(N,N-dimethylaminosulphonyl)-2,1,3-benzoxadiazole (DBD-ProCZ). The tagging conditions were optimized with D-glucose. Monosaccharides and oligosaccharides were efficiently labelled under conditions of 65 °C for 180 min in water : acetonitrile mixture containing 0.5% trichloroacetic acid (TCA). The resulting derivatives were detected at 540 nm (excitation at 450 nm) after separation by HPLC. The separation of individual saccharides was possible using water : acetonitrile as the mobile phase using a reversedphase column (Inertsil ODS-80A) and a normal-phase column (TSKgel Amide-80). N-Acetyl-D-glucosamine (GluNAC), N-acetyllactosamine (LacNAC) and tri-N-acetyl-chitotriose (tri-N-AceChito), which are components of glycoconjugates, were completely separated with an isocratic elution using as Inertsil ODS column. The oncolumn detection limit with the proposed HPLC separation and fluorescence detection at the low pmol level.