Synthesis and Characterization of the Perthiatriarylmethyl Radical and Its Dendritic Derivatives with High Sensitivity and Selectivity to Superoxide Radical

Abstract

EPR spectroscopy, coupled with the use of tetrathiatriarylmethyl (TAM) radicals, has been a reliable method to detect the superoxide radical (O ). However, the specificity and biocompatibility of TAM radicals need to be further improved. Although derivatization may overcome the drawbacks of current TAM radicals, esterification or amidation through the carboxylic groups greatly changes their redox properties and makes them inert to O . Herein, the synthesis of a perthiatriarylmethyl (PST) radical and its dendritic derivatives, PST-TA and PST-NA, in which PST is covalently linked with dendrons containing three (TA) and nine (NA) carboxylic acids, respectively. The results show that PST rapidly reacts with O to yield a unique quinone methide product. Dendritic modification of PST slightly decreases the reactivities of PST-TA and PST-NA, but notably increases their biostability toward various oxidoreductants. The detection limit of PST-NA to O was estimated to be 2.1 nm min over 60 min of detection. Importantly, PST-NA shows threefold higher sensitivity to O in the presence and absence of ascorbic acid than that of the classic spin-trapping technique. In addition, the application of PST-NA to detect extracellular O generation in stimulated RAW 264.7 macrophages was also explored. This study demonstrates that PST-NA has great potential for specific detection and quantitation of O in extracellular sites.

Site Selective and Efficient Spin Labeling of Proteins with a Maleimide-Functionalized Trityl Radical for Pulsed Dipolar EPR Spectroscopy

“…Since the introduction of 5 [46] many synthesis improvements [33,47,48,49,50] and derivatization strategies [44,49,50,51,52,53,54,55,56,57,58,59] as well as applications of trityl compounds in medicinal probing [60,61], imaging [62,63], as magnetic materials [64], and as spin labels in structural biology [28,33,35,36,37,43,44,45,65] have been reported. Recent examples for the trityl labeling of cysteine residues in proteins used butene ( 4 , Figure 1) [33] or methanethiosulfonate ( 6 and 7 , Figure 2) [33,35,36] derivatives of 5 to establish the bioconjugation via thioether bonds for in cell studies or via disulfide bonds for in vitro studies, respectively.…”

Molecular Probes for Evaluation of Oxidative Stress by In Vivo EPR Spectroscopy and Imaging: State-of-the-Art and Limitations

“…Besides, trityl-based probes are currently being developed as selective and quantitative sensors for superoxides, but have not yet been tested in vivo [110,111]. Their selectivity relies on the specific transformation of tetrathiatriarylmethyl radicals by a superoxide to quinone-methide products [112] ( Figure 11).…”

“…Their narrow EPR linewidth (≈0.02 mT under anaerobic conditions) makes them very promising probes for imaging. Dendritic encapsulation enhances the selectivity for a superoxide and increases the resistance against reduction to the corresponding triarylmethanes, while only moderately decreasing the reaction rate with a superoxide (k ≈ 10 4 M −1 s −1 ) and reducing albumin binding [110]. These sensors are, however, not able to permeate cells and will thus be limited to extracellular detection of a superoxide.…”

Synthesis and Redox Properties of Water-Soluble Asymmetric Trityl Radicals

“…•− in air and further undergo O 2 •− -promoted decay. 16,42 Moreover, ascorbate was also reported to be another reducing agent for the decay of trityl radicals. 37,43,44 Despite these studies, there is no report regarding substituent effect on redox properties of trityl radicals.…”