High Performance Anion Exchange Chromatography Of Carbohydrates

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In order for pulsed amperometry to have utility for the chromatographic determination of carbohydrates, a high resolution separation method is required that is also compatible with high pH. This requirement has been met by the development of column technology which allows high resolution separations of carbohydrates by anion exchange chromatography at high pH.

Carbohydrates are often thought of as neutral compounds, but in actuality they are weak acids with pKa's in the range of 12-14 (Table 1). At these pH levels, oxyanion formation occurs (Figure 6) allowing separation by ion exchange. Ion exchange separations are based on the relative affinity of the analyte ion in competition with the eluent ion for the same exchange sites (Figure 7). The greater the affinity of the ion, the longer the retention time. For carbohydrates, subtle structural differences and small differences in pKa can cause significant differences in retention characteristics.

Table 1 Dissociation constants of some common carbohydrates in water at 25°C

Sugar

Fructose

Mannose

Xyolse

Glucose

Galactose

Dulcitol

Sorbitol a-Methyl glucoside

Henshall

Figure 7 Ion exchange separations are based on the relative affinites of the analyte ions in competition with the eluent ion for the same exchange sites.

HPAE-PAD

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