Thiol-Dependent Reduction of the Triester and Triamide Derivatives of Finland Trityl Radical Triggers O2-Dependent Superoxide Production.

Tetrathiatriaylmethyl (trityl) radicals have found wide biomedical applications as magnetic resonance probes. Trityl radicals and their derivatives are generally stable toward biological reducing agents such as glutathione (GSH) and ascorbate. We demonstrate that the triester (ET-03) and triamide (AT-03) derivatives of the Finland trityl radical exhibit unique reduction by thiols such as GSH and cysteine (Cys) to generate the corresponding trityl carbanions as evidenced by the loss of EPR signal and appearance of characteristic UV-vis absorbance at 644 nm under anaerobic conditions. The trityl carbanions can be quickly converted back to the original trityl radicals by oxygen (O2) in air, thus rendering the reaction between the trityl derivative and biothiol undetectable under aerobic conditions. The reduction product of O2 by the trityl carbanions was shown to be superoxide radical (O2•-) by EPR spin-trapping. Kinetic studies showed that the reaction rate constants (k) depend on the types of both trityl radicals and thiols with the order of kET-03/Cys (0.336 M-1 s-1) > kET-03/GSH (0.070 M-1 s-1) > kAT-03/Cys (0.032 M-1 s-1) > kAT-03/GSH (0.027 M-1 s-1). The reactivity of trityl radicals with thiols is closely related to the para-substituents of trityl radicals as well as the pKa of the thiols and is further reflected by the rate of O2•- production and consumptions of O2 and thiols. This novel reaction represents a new metabolic process of trityl derivatives and should be considered in the design and application of new trityl radical probes.