Reversed-phase high-performance liquid chromatographic/mass spectrometric method for separation of 4-methylimidazole and 2-acetyl-4-(1,2,3,4-tetrahydroxybutyl)imidazole at pg levels Original Research Article

A high-performance liquid chromatographic/mass spectrometric method (HPLC/MS) has been developed to determine both 4-methylimidazole (4MeI) and 2-acetyl-4-(1,2,3,4-tetrahydroxybutyl)imidazole (THI) in one run. Among three sorbents tested, the best peak shape of 4MeI was achieved on a reversed-phase MetaChem Polaris C18-A at pH 9.5. The sensitivity and the range of UV detection at 215 and 290 nm for 4MeI and THI, respectively, were compared to the parameters achieved by the electrospray ionization mass spectrometric (ESI/MS) detection in the presence of 5 mmol l−1 ammonium hydroxide using selected ion-monitoring (SIM) mode. The on-column limits of detection were 0.147 ng of 4MeI and 0.084 ng of THI using UV detection and 1 pg of 4MeI and 3 pg of THI using ESI/MS detection, respectively. The effect of both ammonium ion concentration and energy of fragmentation on 4MeI and THI ionization is discussed. The method could be applied for a fast and sensitive determination of 4MeI and THI in different biological materials as well as in Class III Caramels.