Oxidized ascorbic acid and reaction products between ascorbic and amino acids might constitute part of age pigments

Fluorescence and non-enzymatic browning were observed in reactions between ascorbic acid (AH2) and amino acids (AA) as well as in reactions involving AH2 autoxidation and/or polymerization in the presence of trace amounts of adventitious iron (less than or equal to 10 microM). These reaction products exhibited fluorescent spectra (400-490 nm) akin to those of extracts from lipofuscin-rich tissues. Lengthy incubation of AH2 at 37 degrees C in phosphate buffer (pH 7.0) caused a concentration- and time-dependent increase in absorbance at 412 nm, paralleling increase in 377/440 nm fluorescence, due to formation of autoxidation/polymerization products. The fluorescences of these substances were increased by acidity and quenched at alkaline conditions but restored by neutralization. The reactions between AH2 (0.1-2.0 mM) and a number of AA (1.0-4.0 mM) were also found to result in products with blue fluorescence. Following TBA test, the AH2 autoxidation/polymerization products and AH2/AA reaction products showed only moderate and slight absorbance at 535 nm, respectively, indicating a little or minute formation of aldehydes with MDA-like reactivity. The findings in this study, nevertheless, suggest possible misinterpretations of results in previous studies dealing with AH2-dependent, oxygen free radical induced, 'lipofuscin-related fluorescence'. Thus, similar to nonenzymatic glycosylation (Maillard) reactions, AH2 autoxidation as well as reactions between AH2 and AA may result in 'lipofuscin-like material', as judged from their fluorescence spectral patterns.