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Determination of 4(5)-methylimidazole in sugar-amino acid aqueous systems by UPLC-Q-ToF-MS
1, 2 , 1 , 3 , 1 , * 1
1  Department of Food Science and Human Nutrition, Chemical Laboratories, Agricultural University of Athens, EU-CONEXUS European University, 11855 Athens, Greece
2  Department of Food Science and Technology, University of West Attica, Ag. Spyridonos str, 12243 Egaleo, Athens, Greece
3  Laboratory of Quality and Safety of Agricultural Products, Landscape and Environment, Department of Agriculture, Hellenic Mediterranean University, Stavromenos PC, Heraklion, 71410 Crete, Greece Institute of Agri-Food and Life Sciences Agro-Health, Helle
Academic Editor: Dirk W. Lachenmeier


4(5)-Methylimidazole (4(5)MEI) has been classified by the International Agency for Research on Cancer (IARC) as possibly carcinogenic to humans [1]. It can be formed during the caramelization procedure and the Maillard reaction as a result of the thermal processing of foods [2]. Although several studies concerning the formation of 4(5)MEI in Maillard reaction model systems exist (mainly sugars with ammonia), data concerning the formation of 4(5)MEI by the reaction between amino acids and sugars are still scarce [2]. In this study, an Ultra-high Performance Liquid Chromatography-Quadrupole Time-of-Flight Mass Spectrometry (UPLC-Q-ToF-MS) method was developed for the determination of 4(5)MEI in aqueous model systems containing sugars (glucose and fructose) and amino acids (proline, phenylalanine, lysine, and tyrosine) after thermal processing at 100 °C. Since studies on the formation of 4(5)MEI in honey are scarce, the above-mentioned compounds were intentionally chosen as they are some of the main components of honey. The results showed that the 4(5)MEI was formed in all model systems, with the highest concentrations to be determined in fructose-proline (3.5 μg mL-1) and fructose-tyrosine (3.0 μg mL-1) aqueous model systems. It is concluded that further research is required to examine whether this compound can be formed under any circumstance in honey.


  1. IARC Working Group on the Evaluation of Carcinogenic Risks to Humans. 4-Methylimidazole. IARC Monogr. Eval. Carcinog.
    Risks Hum. 2013, 101, 447–459.
  2. Revelou, P.K.; Xagoraris, M.; Alissandrakis, A.; Pappas, C.S.; Tarantilis, P.A. A review of the analytical methods for the determination of 4(5)-methylimidazole in food matrices. Chemosensors 2021, 9(11), 322.
Keywords: 4(5)-Methylimidazole; honey; Maillard reaction; liquid chromatography; mass spectrometry