Rapid separation and quantification of imidazole dipeptides in meats using a PBr column packed with 3-(pentabromobenzyloxy)propyl group modified silica gel

Abstract

Meats are rich in imidazole dipeptides (IDPs) such as carnosine, anserine, and balenine, known for their antioxidant and antifatigue properties. The concentrations and types of these IDPs vary significantly among different animal species, necessitating a quantitative method for the precise measurement of individual IDPs. Simultaneous analysis of multiple compounds is typically conducted using reversed-phase high-performance liquid chromatography (RP–HPLC). However, C₁₈ columns, which are commonly employed in RP–HPLC, fail to adequately retain highly hydrophilic IDPs, making separation and quantification challenging. Previously, we developed a PBr column packed with 3-(pentabromobenzyloxy)propyl group modified silica gel, which effectively retains various highly hydrophilic compounds in RP mode. In this study, we established a method for the rapid separation and quantification of IDPs within 10 min under simplified conditions (isocratic mode) using a single quadrupole liquid chromatography–mass spectrometer (LC–MS) equipped with a PBr column, without the need for derivatization. Linear calibration curves for each IDP were generated using glycyl-L-leucine as the internal standard, with the desolvation temperature of the MS instrument set at 500 °C. The proposed method achieved extraction and recovery rates of IDPs ranging from 100.0 % to 113.5 % at three spiking levels, with no carryover observed, even in samples with high concentrations. Additionally, matrix effects ranged from 95.5 % to 109.6 %, with negligible ion suppression and enhancement effects. Furthermore, the method enabled accurate analysis of IDPs with a relative standard deviation of <15 % in meats from various animal species, including chicken, pork, beef, lamb, mutton, deer, horse, and kangaroo.