Determination of free and reversibly bound sulphite in foods by reverse-phase, ion-pairing high-performance liquid...

The reaction of sulphite with formaldehyde to form hydroxymethylsulphonate (HMS), which is very stable under the controlled conditions of this assay, was used as the first step in an analytical procedure to determine foodborne sulphite. The effect of mobile-phase pH on the stability of HMS during high-performance liquid chromatography was studied. It was found that on-column HMS dissociation to formaldehyde and bisulphite increased with the pH of the mobile phase; therefore the relatively low pH 4.7, at which the dissociation of HMS was approximately 2%, was selected for the analysis. In addition, the release of sulphite from its reversibly bound forms in wine and other foods was examined as a function of the pH of the extraction medium by following the appearance of HMS formed from the reaction of the freed sulphite with formaldehyde. The rate of dissociation of the reversibly bound sulphite was relatively slow at pH 3 but very rapid at pH 7. This difference in kinetics was exploited to develop a procedure to determine free and reversibly bound sulphite in food. The method was challenged by post-reagent spiking studies, i.e. adding the sulphite spike after the food has been blended with the sulphite-protective formaldehyde solution but before proceeding with the remainder of the assay. An average recovery of 100% with a standard deviation of 5.2% (n = 45) was realized at levels of 5, 10 and 20 ppm by weight as sulphur dioxide. Recovery of the sulphite added as the bisulphite addition product of acetaldehyde, a model compound for reversibly bound sulphite, was 95%.