Determination of carbohydrates by high-performance capillary electrophoresis with indirect absorbance detection

Abstract

High-performance capillary electrophoresis (HPCE) methods with indirect absorbance detection (IAD) have been developed for the determination of carbohydrates, e.g., glucose, fructose, rhamnose, ribose, maltose, lactose, sucrose and gluconic acid. The suitability and performance of six background electrolytes (BGEs), i.e., 1-naphthylacetic acid (NAA), 2-naphthalenesulfonic acid, 1,3-dihydroxynaphthalene, phenylacetic acid, p-cresol and sorbic acid, for the IAD method were investigated. The effects of the concentration of the BGE, pH and temperature on the CE separation of these analytes were evaluated. NAA was found to be best suited as the carrier buffer and background absorbance provider for the detection at 222 nm. The optimal CE performance was found when employing 2 m M NAA, pH 12.2, at 25°C. In comparison with the previous method that used sorbate as the BGE, the present method utilizing NAA shows a 3–6 fold increase in the separation efficiency and a 2–5 fold improvement in the detection limit. The calculated number of theoretical plates is in the range of 1.0–3.0·10 5. The precision of the present method for most sugar analytes, measured by the coefficient of variation (C.V.), typically, is less than 1% for the migration time and better than 3% for the peak height and peak area ( n=6). The detection limit is about 0.1 m M for all analytes, except for ribose for which it is about 0.2 m M. This new method is fast, accurate and can be readily applied to real biological samples for quantitative determination of selected carbohydrates.