Analysis of formose sugar and formaldehyde by high-performance liquid chromatography

Formose sugar and formaldehyde (HCHO) were analyzed using high-performance liquid chromatography (HPLC) utilizing a CarboPac PA1 column (Dionex) and pulsed amperometric detection. This HPLC system was unsuitable for the analysis of formose sugar and HCHO and thus reducing sugars and unconverted HCHO were determined by endowing them with charges through a derivatization method using 2,4-dinitrophenylhydrazine. The separation and detection of compounds were performed by three Chromolith RP-C18 columns (Merck) and diode array detection, at a wavelength of 360 nm ultraviolet light, respectively. Lower sugars (except HCHO) showed some instabilities when the derivatized samples were kept for the extended periods of time. For C5 and consecutive higher sugars, a certain derivatization time was necessary. In the present case (formose reaction with partial HCHO conversion), approximately 18 h may be a reasonable compromise for the derivatization reaction. A derivatization agent to compound mole ratio of up to 100:1 was required to complete the derivatization of C4 and higher sugars. However, the analysis of C4 and consecutive higher sugars is problematic for example due to overlapping of peaks or branched-chain sugars.