HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY OF GLYCOPEPTIDES AND OLIGOSACCHARIDES ON GRAPHITIZED CARBON COLUMNS

High-performance liquid chromatography of carbohydrate materials on gr aphitized carbon columns (GCC) has some advantages over other types of chromatography. Oligosaccharides and glycopeptides with few amino aci ds are barely retained on reversed-phase columns even under high salt or low pH conditions, but can be retained effectively on a graphitized carbon column. Moreover, elution of GCC requires concentrations of or ganic solvents lower than that required for normal-phase columns. The usefulness of graphitized carbon columns is exemplified by the followi ng results: (i) Man(9)GlcNAc(2) with only Asn or Asn-Phe (derived from soybean agglutinin) was not retained by a C-18 reversed-phase column, but could be separated on a GCC with a gradient of 10-45% CH8CN in 30 min. (ii) Ribonuclease B glycopeptides obtained by Pronase digestion could be separated on GCC with a gradient of 10-30% CH3CN, but they we re not retained on a C-18 reversed-phase column even with water as elu ent. (iii) Oligosaccharides released from ribonuclease B by endo-beta- N-acetylglucosaminidase were separated from each other and peptides on GCC with a linear gradient of 10 mM NH4OH to 10 mM NH4OH-12.5% CH3CN in 50 min at 70 degrees C. Silica-based columns do not allow such an a lkaline eluent. (iv) Chito-oligosaccharides (DP 1-9) are well separate d within 40 min on GCC with a gradient (10 mM NH4OH-10 mM NH4OH with 2 5% CH3CN) at 50 degrees C. Chito-oligosaccharides could not be separat ed by high-performance anion exchange columns such as Carbopac PA-1. ( C) 1994 Academie Press, Inc.