2-Oxo-Imidazole-Containing Dipeptides Play a Key Role in the Antioxidant Capacity of Imidazole-Containing Dipeptides

Abstract

There is substantial evidence for the antioxidant functions of imidazole-containing dipeptides (IDPs), including carnosine and anserine, under physiological and pathological conditions in vivo. However, the detailed mechanism underlying the antioxidant functions is still poorly understood. Recently, we discovered the endogenous production of 2-oxo-imidazole-containing dipeptides (2-oxo-IDPs), such as 2-oxo-carnosine and 2-oxo-anserine, as novel derivatives of IDPs in mouse tissues and revealed that the antioxidant capacity of 2-oxo-carnosine was much greater than that of carnosine. However, the antioxidant capacity of 2-oxo-IDPs still remains unclear. In this study, we evaluated 2-oxo-carnosine and 2-oxo-anserine by multiple in vitro assays, such as 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging, ferric reducing/antioxidant power, and oxygen radical absorbance capacity assays in comparison with the corresponding IDPs, carnosine and anserine. All the assays employed herein demonstrated that 2-oxo-carnosine and 2-oxo-anserine exhibited a greater antioxidant capacity than that of the corresponding IDPs. Quantitative high-performance liquid chromatography tandem mass spectrometry revealed that commercial IDPs standards were contaminated with a certain amount of 2-oxo-IDPs, which was correlated with the antioxidant capacity. DPPH radical scavenging assay revealed that the elimination of contaminated 2-oxo-IDPs from the IDPs standards caused a significant decrease in the antioxidant capacity compared to the original IDPs standards. These results suggest that the main driver of the antioxidant capacity of IDPs is 2-oxo-IDPs; accordingly, the conversion of IDPs to 2-oxo-IDPs may be a critical step in the antioxidant functions.

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

“…However, no study has used isotope-labeled balenine. The application of HPLC-ESI-MS/MS coupled with a stable isotope dilution method for 2-oxo-IDPs has only been demonstrated by our previous reports for the detection of 2-oxo-carnosine [ 7 , 24 , 25 ], 2-oxo-anserine [ 7 , 24 , 25 ], 2-oxo-homocarnosine [ 7 ], and 2-oxo-homoanserine [ 23 ]. However, no study has demonstrated about 2-oxo-balenine.…”

“…Previous studies have demonstrated that 2-oxo-IDPs (i.e., 2-oxo-carnosine, 2-oxo-anserine, and 2-oxo-homoanserine) exhibit a greater antioxidant capacity, compared to the corresponding IDPs [ 7 , 23 , 25 ]. Further, it has been reported that 2-oxo-carnosine shows significant cytoprotective effects on rotenone-induced neuronal cell death, while there are no significant effects by the treatment with the precursor carnosine [ 7 ].…”

“…The DPPH radical scavenging assay was carried out as previously described [ 7 , 25 ], with slight modifications. In brief, in a 96-well plate, 100 μM DPPH was incubated in 12 mM sodium phosphate buffer (pH 7.4) in the presence or absence of 2-oxo-IDPs (final 20–40 μM) or IDPs (final 200–1000 μM) for 20 min at room temperature (25 °C).…”

“…Trolox solutions (final 5–30 μM) were used for defining the standard curve. Radical scavenging capacity was evaluated by the inhibition ratio (%), which was calculated by the following formula: Inhibition ratio (%) = (Ac − As)/Ac × 100, where Ac and As indicate the absorbances of the blank control (water) and sample, respectively [ 25 ]. The radical scavenging capacity was expressed as Trolox equivalent antioxidant capacity (TEAC), which was calculated using the Trolox standard curve.…”

“…2-Oxo-carnosine and 2-oxo-anserine have also been detected in several types of meat samples [ 24 ]. Notably, multiple in vitro assays have demonstrated that 2-oxo-carnosine and 2-oxo-anserine exhibit a greater antioxidant capacity than that of corresponding IDPs, suggesting that 2-oxo-IDPs play a key role in the antioxidant capacity of IDPs [ 25 ]. In addition, 2-oxo-homoanserine also have showed potent radical scavenging capacity than the precursor, homoanserine [ 23 ].…”