Most saccharides form anionic complexes in alkaline borate buffers and therefore are amenable to separation by ion-exchange chromatography. Khym and Zill (1, 2) demonstrated the utility of ion-exchange chromatography of sugar-borate complexes and HalIen (3) further developed the technique so that separation and quantitation of micromole quantities of neutral monosaccharide could be achieved. More recently several systems were described for the automated analysis of saccharides using alkaline borate buffers (4-7).
In order to minimize alkaline rearrangement reactions an ion-exchange chromatographic system utilizing elution with boric acid/glycerol buffers at neutral pH was developed (8). Since 2,3-butanediol forms a weak complex with borate (9), it is possible to substitute 2,3-butanediol for glycerol. Such a substitution has proved valuable in developing a twobuffer stepwise elution system.
MATERIALS
Columns. Jacketed glass columns having an internal diameter of 6 mm and a length of 100 cm were obtained from Glass Engineering Co., Inc., Houston, Texas (catalog No. 3200-6100).
Resin. Dowex l-X4 (minus 400 mesh, total capacity 3.5 meq/dry gm, control No. 5407-45) was obtained from Bio-Rad Laboratories, Richmond, California.
In order to obtain more uniform particle size, the resin was fractionated by settling. Moist resin (500 gm) was suspended in 3.5 liters of water in a 4 liter beaker and the resin allowed to settle 'Portions of this research were presented at the 155th National Meeting of the American Chemical Society, April, 1968.