High-performance liquid chromatographic separation of monosaccharides as their peracetylated ketoximes and aldononitriles

Abstract

A wide variety of chromatographic methods have been applied to monosaccharides. The most common method is gas chromatography (GC), alone or combined with mass spectrometry (MS). However, this technique presents some problems. On the one hand, sugars must be derivatized to increase their volatility [1 7] and in most instances the derivatization process leads to multiple products (anomeric mixtures, furanose or pyranose forms) that complicate the analysis; to avoid this problem, derivatives such as alditols [8,9] and oximes [10 -13] which give rise to acyclic structures have been prepared. On the other hand, GC is generally a destructive method, in which the complete structural elucidation must be made by using reference samples. By the use of GC MS, it is easy to establish ifa compound is a C~or an Cs-aldose. However, the only way to distinguish among stereoisomers is to use reference samples and compare their retention times. An alternative to GC is high-performance liquid chromatography (HPLC) [14,15], in which different none-destructive detection systems can be used. HPLC permits amounts of pure substance to be collected, which can then be studied by use of another analytical technique [16]. HPLC has been applied extensively in carbohydrate chemistry [14,15,17-20]. In spite of the vast literature in which a variety of analytical conditions are described, most of them are of restrictive application and there is none that could separate simultaneously all monosaccharides. With the latter aim, we have studied the HPLC of mixtures of peracetylated aldononitriles and peracetylated ketoximes. The reasons for the choice of those derivatives were (a) both of the two families of sugar derivatives have been the subject of extensive previous GC MS studies [21 23], making possible rapid control of our work by an alternative method; (b) they are easy to prepare, purify and store, and are reasonably stable for long periods of time; and (c) the intermediate polarity of the peracetylated aldononitrile (PAAN) and peracetylated ketoxime (PAKO) derivatives allows their HPLC analysis by using either polar, intermediate or apolar stationary phases. In this paper we report the behaviour of PAAN and PAKO sugar derivatives.