Measurement of amylose/amylopectin ratio by high-performance liquid chromatography

Amylose content has been measured by iso-amylase digestion of starch followed by quantitation of the amounts of the higher molecular weight chains (from amylose) and the lower molecular weight chains (from amylopectin). The separatrion and quantitation has been performed by size-exclusion high performance liquid chromatography an hydrophilic columns. Three columns were tested and an Ultrahydrogel 250 was found to provide the optimum separation. Starches of up to 30-35 per cent amylose may be analysed in this way without the need for first removing the lipids. With starch of higher amylose content, precipitation of the amylose in the sample may occur while it is awaiting HPLC analysis. This precipitation can be prevented in starches of up to 40 per cent amylose by the use of dimethyl sulfoxide in the reaction mixture. Samples with amylose content higher than this always show some precipitation and the amylose contents obtained are similar for all starches with amylose in the range from 40 to 80 per cent, measured by iodine determination. The method gives results which are more repeatable than those obtained by spectrophotometric determination of iodine binding, and which appear to be unaffected by the lipid content of the starch.