Quantification of 4-methylimidazole in class III and IV caramel colors: validation of a new method based on heart-cutting two-dimensional liquid chromatography (LC-LC).

4-Methylimidazole (4MeI) is a nitrogen compound formed during the manufacture of class III and IV caramel colors. The European Commission has limited its content to 250 ppm. Two methods were compared to perform 4MeI quantification in caramels. The first one, currently used and considered to be the reference method, consists of a hot extraction of caramel color with dichloromethane and an analysis of the acetyl derivative of the extract by gas chromatography coupled to mass spectrometry (GC-MS). The second method is based on the heart-cutting two-dimensional liquid chromatography technique (LC-LC) to directly separate 4MeI from the other components present in caramel color sample (diluted in water) in <30 min. The accuracy profile validation method and the comparison between the results obtained with the two methods show that the new and completely automated LC-LC method is usable to quantify 4MeI in caramels.