Laser-Excited Fluorescence of Dityrosine

Abstract

In this research, laser-excited fluorescence was examined for sensitive detection of aqueous dityrosine. Samples were excited with a 6.3-mW, 325-nm helium-cadmium laser focused into a small volume-fluorescence cell with a 10-cm lens. The resulting fluorescence emission was collected perpendicular to the excitation and detected with two different schemes. An optical bandpass filter was used with a photomultiplier tube for sensitive quantitative measurement, while a photodiode array detector was used in conjunction with a spectrograph for qualitative characterization of fluorescence emission spectra. Dityrosine detection on the order of 2 × 10−11 M was obtained with the use of the photomultiplier tube with bandpass optical filter. The dityrosine fluorescence yield is found to vary with the solution pH, the relative concentrations of ferric and ferrous iron, and the amount of dissolved oxygen. A maximum fluorescence yield is observed for iron-free, oxygen-free alkaline solutions. Fluorescence quenching by oxygen is a cumulative photolysis effect that diminished fluorescence yield with increased irradiation time. Flowing the solutions minimized photolysis effects in oxygenated solutions. Quenching by ferrous and ferric iron is found to be due primarily to complex formation. The ferrous iron complex appears to have a fluorescence efficiency of ∼20% of the free dityrosine. The ferric iron dityrosine complex appears to have two ferric ions per molecule at low iron concentration. Other complexes may form at different concentrations. Solvent effects on dityrosine absorption and fluorescence spectra were also investigated. A red shift in dityrosine fluorescence maximum was observed in 1 M trichloroacetic acid and in N, N-dimethylformamide. The fluorescence emission maximum was shifted to the blue in acetonitrile and glacial acetic acid. These shifts were attributed to typical solvochromic behavior.

A novel synthetic approach to tyrosine dimers based on pterin photosensitization

“…The emission decays, registered at 410 nm, were clearly monoexponential ( Figure 9) with a value of fluorescence lifetime (τF) of 4.0 (±0.1) ns and 4.4 (±0.1) ns, in acid and alkaline media, respectively. These results are in disagreement with previous works, in which biexponential fluorescence decay pattern of Tyr2 have been reported [39]. We attribute this difference to the purity of our samples using our method of synthesis and isolation.…”

“…The absorption and emission spectra of this new peak are red-shifted with respect to those of Tyr [38]. The absorption spectrum of this product, registered by the UV-Vis detector (see inset of Figure 5a), corresponded to the previously reported for the acidic form of Tyr2 [39,40]. Therefore, this new peak was safely assigned to the dimeric form of Tyr (Tyr2).…”

“…Moreover, fluorescence of Tyr2 is an unspecific marker of oxidative damage in proteins [6,15]. Despite the biological and medical importance of Tyr2 emission, the reports on fluorescence of isolated Tyr2 in aqueous solutions are rare [15,39].…”

Live tissue intrinsic emission microscopy using multiphoton-excited native fluorescence and second harmonic generation

“…The spectrum of an individual NFT shows an emission peak at 460 nm (Fig. 4b), which is redder than would be expected from ditryosine alone (400-nm emission) and occurs possibly because of higher-order polymerized tyrosine products (36). Lipofuscin granules are also brightly fluorescent but have an emission spectrum easily distinguishable from NFT emission (Fig.…”

The interstitial crystal-nucleating sheet in molluscan Haliotis rufescens shell: A bio-polymeric composite

“…Dityrosine has a maximum emission peaks at about 400 nm (Mahmoud and Bialkowski, 1995;Harms et al, 1997), however this position is strongly affected by the pH of the environment, the presence of close carboxylate groups, and the reorientational mobility of the peptide in solution (Bothwell et al, 2003). The green color and fluorescence pattern of the interstitial polymer sheets from the Haliotis rufescens shell both are likely to originate with the structures of some of the suggested cross-links of the tyrosine aromatic side-chains.…”