Quantitative Determination of 2-Oxo-Imidazole-Containing Dipeptides by High-Performance Liquid Chromatography/Tandem Mass Spectrometry

Abstract

2-Oxo-imidazole-containing dipeptides (2-oxo-IDPs), novel imidazole-containing dipeptide (IDP) derivatives, exhibit a much higher antioxidant capacity than that of IDPs. However, quantitative methods have only been developed for IDPs, and methods for the quantitative analysis of 2-oxo-IDPs are needed. In this study, we developed methods for the quantitative analysis of 2-oxo-IDPs by high-performance liquid chromatography with online electrospray ionization-tandem mass spectrometry (HPLC-ESI-MS/MS) coupled with a stable isotope dilution method. First, we prepared stable isotope-labeled IDP and 2-oxo-IDP standards for MS analyses. Next, using these standards, we established highly sensitive, selective, and absolute quantitative analysis methods for five IDPs and five 2-oxo-IDPs by HPLC-ESI-MS/MS, achieving a limit of detection in the fmol range. Finally, we applied the method to various types of meat, such as beef, pork, chicken, and whale meat, demonstrating the detection of both IDPs and 2-oxo-IDPs. Furthermore, we provide the first evidence for the endogenous production of 2-oxo-balenine in meats. The methods developed in this study enable the detection of trace levels of 2-oxo-IDPs in biological samples and could be helpful for understanding the biological relevance of 2-oxo-IDPs.

Antioxidant Activity of Beef, Pork and Chicken Burgers before and after Cooking and after In Vitro Intestinal Digestion

“…Bioactive peptides are amino acid sequences that are inactive within the parent protein sequence but are activated following proteolytic processes (enzymatic or chemical hydrolysis), microbial fermentation or gastrointestinal digestion [21]. In addition, some endogenous antioxidants specific to meat, such as dipeptides that contain 2-oxoimidazole (2-oxo-IDP), were formed, in turn, from histidine or its methylated derivatives, which had a marked antioxidant activity even greater than other dipeptides, and have been observed in different meats (beef, pork and chicken) [22].…”

Monocarboxylate Transporter 13 (MCT13/SLC16A13) Functions as a Novel Plasma Membrane Oligopeptide Transporter

“…The samples were passed through a 0.45-μm PVDF filter (FastRemover for Protein; GL Sciences, Tokyo, Japan), and the filtrates were used for analysis. Analytes were separated by the LC system at a 0.6 mL/min flow rate on an Intrada Amino acid analytical column (50 × 3 mm, 3 μm) (Imtakt, Kyoto, Japan), which can separate dipeptide and peptide derivatives including anserine and carnosine, especially [ 32 ]. The separation was achieved by using a gradient program composed of solvent A (ammonium formate buffer (20 mM)) and solvent B (acetonitrile).…”