A method for rapid quantification of sucrose and fructan oligosaccharides suitable for enzyme and physiological studies

Abstract

This paper presents a method for rapid quantification of sucrose and fructans by thin-layer chromatography (TLC) and densitometry. In the optimized protocol, glass-backed TLC plates were developed in a solution of propan-1-ol: ethyl acetate : water (45:35:20, v/v/v) for 3·5 h at 30 °C. In this system, sucrose, the fructan trisaccharides: 1-kestose, 6G-kestose and 6-kestose, fructan tetrasaccharide and fructan pentasaccharide were separated. An internal standard of stachyose was useful for most of the applications examined. The fructans were detected by spraying with urea-phosphoric acid and heating at 110 °C for 6 min. The blue colour developed was stable for 18 h provided that plates were kept in the dark. Quantification was achieved by reflectance densitometry and comparison with standards. The standard curves for the oligosaccharides were not linear, owing to the spreading of spots on the TLC plates, but were highly repeatable. The TLC quantification method provided a more rapid analysis of sucrose and fructan oligosaccharides than was possible by high-performance liquid chromatography analysis because time-consuming pre-treatment of samples was not needed. The method will be useful for both enzyme and physiological studies.