Development and validation of a modified QuEChERS protocol coupled to UHPLC-APCI-MS/MS for the simple and rapid quantification of 16 heterocyclic aromatic amines in cooked beef

Heterocyclic aromatic amines (HAAs) are neo-formed compounds generated during the cooking of meats and are known or suspected to be mutagenic and carcinogenic. In this study, a novel, simple, and fast methodology combining salting-out liquid-liquid microextraction, solid-phase extraction (SPE), and UHPLC-APCI-MS/MS was developed for the analysis of 16 HAAs. The QuEChERS extraction (quick, easy, cheap, efficient, rugged, and safe) was revisited and modified using mixed-mode SPE purification to adapt the method to the particular physicochemical properties of HAAs and the fatty nature of the beef matrix. The UHPLC-MS/MS analysis was performed on a C8 column in less than 4 min using positive APCI ionisation and an internal standard. The method was validated according to the European Medicines Agency and Eurachem guidelines and was successfully applied to beef samples of various cooking degrees, showing HAA levels similar to those shown by previous studies.