Mitochondria are recognized as one of the most important targets for new drug design in cancer, cardiovascular, and neurological diseases. Currently, the most effective way to deliver drugs specifically to mitochondria is by covalent linking a lipophilic cation such as an alkyltriphenylphosphonium moiety to a pharmacophore of interest. Other delocalized lipophilic cations, such as rhodamine, natural and synthetic mitochondria-targeting peptides, and nanoparticle vehicles, have also been used for mitochondrial delivery of small molecules. Depending on the approach used, and the potentials of cell and mitochondrial membranes, more than 1000-fold higher mitochondrial concentration can be achieved. Mitochondrial targeting has been developed to study mitochondrial physiology and dysfunction and the interaction between mitochondria and other subcellular organelles and for treatment of a variety of diseases such as neurodegeneration and cancer. In this review, we discuss efforts to target small-molecule compounds to mitochondria for probing mitochondria function, as diagnostic tools and potential therapeutics. We describe the physicochemical basis for mitochondrial accumulation of lipophilic cations, synthetic chemistry strategies to target compounds to mitochondria, mitochondrial probes and sensors, and examples of mitochondrial targeting of bioactive compounds. Finally, we review published attempts to apply mitochondria-targeted agents for the treatment of cancer and neurodegenerative diseases.

In this study, we show that boronates, a class of synthetic organic compounds, react rapidly and stoichiometrically with peroxynitrite (ONOO−) to form stable hydroxy derivatives as major products. Using stopped-flow kinetic technique, we measured the second order rate constants for the reaction with ONOO−, hypochlorous acid (HOCl), and hydrogen peroxide (H2O2), and found that ONOO− reacts with 4-acetylphenylboronic acid nearly a million times (k = 1.6 × 106 M−1 s−1) faster than H2O2 (k = 2.2 M −1 s−1) and over two hundred times faster than HOCl (k = 6.2 × 103 M−1 s−1). Nitric oxide (•NO) and superoxide (O •2−) together, but not alone, oxidized boronates to the same phenolic products. Similar reaction profiles were obtained with other boronates. Results from this study will likely help develop a novel class of fluorescent probes for detection and imaging of ONOO− formed in cellular and cell-free systems.

Background: Recently, new "targeted" fluorescent probes that react selectively with reactive oxygen and nitrogen species to yield specific products have been discovered. Results: High-throughput fluorescence and HPLC-based methodology for global profiling of ROS/RNS is described. Conclusion: This methodology enables real-time monitoring of multiple oxidants in cellular systems. Significance: The global profiling approach using different ROS/RNS-specific fluorescent probes will help establish the identity of oxidants in redox regulation and signaling.

Boronates, a group of organic compounds, are emerging as one of the most effective probes for detecting and quantifying peroxynitrite, hypochlorous acid and hydrogen peroxide. Boronates react with peroxynitrite nearly a million times faster than with hydrogen peroxide. Boronate-containing fluorogenic compounds have been used to monitor real time generation of peroxynitrite in cells and for imaging hydrogen peroxide in living animals. This Perspective highlights potential applications of boronates and other fluorescent probes to high-throughput analyses of peroxynitrite and hydroperoxides in toxicological studies.

There is much interest in the nitration. , Reaction 1,The early indication of the occurrence of this reaction in biological systems came from the report on the inhibitory effect of O 2 . on the activity of endothelium-derived relaxing factor (7).After endothelium-derived relaxing factor identity was established as ⅐ NO (8, 9), its scavenging by O 2 . was first proposed as a contributing factor to endothelial injury (10). Reaction 1 has great physiological significance as both ⅐ NO and hydrogen peroxide (H 2 O 2 , the product of dismutation of O 2 . ) act as important second messengers in redox cell signaling (11,12). In the absence of scavengers, ONOO Ϫ decomposes at neutral pH via protonation to peroxynitrous acid (pK a ϭ 6.7, Reaction 2) to yield nitrate (NO 3 Ϫ ) and free radical intermediates:hydroxyl radical ( ⅐ OH) and nitrogen dioxide ( ⅐ NO 2 ) (Reaction 3, k 3 ϭ 1.25 s Ϫ1 ) (2, 13).In most biological systems, carbon dioxide is a likely scavenger of ONOO Ϫ , yielding a short-lived nitrosoperoxycarbonate anion (ONOOCO 2 Ϫ , Reaction 4, k 4 ϭ 2.9 ϫ 10 4 M Ϫ1 s Ϫ1 (14)). During the decomposition of ONOOCO 2 Ϫ , nitrate and carbon dioxide are formed, as well as nitrogen dioxide radical and carbonate radical anion (Reaction 5) (2, 13, 15).Due to the occurrence of Reactions 4 and 5, as well as the scavenging by peroxiredoxins or oxyhemoglobin in specific subcellular compartments, the lifetime of ONOO Ϫ in biological systems is limited to only a few milliseconds (2, 13). The current methodologies for detection of ONOO Ϫ are based on the detection of radical species formed from ONOO Ϫ decomposition, i.e. ⅐ NO 2 and CO 3 . or ⅐ OH, using tyrosine that forms nitrotyrosine (TyrNO 2 ) as a marker product of intracellular ⅐ NO 2 and dihydrorhodamine 123 (DHR) 2 as a fluorogenic probe for oxidants ( ⅐ NO 2 , ⅐ OH, CO 3 . ). However, ⅐ NO 2 radical formed from the ONOO Ϫ -independent processes, e.g. via myeloperoxidasecatalyzed oxidation of nitrite by H 2 O 2 (16), could make data interpretation more tenuous (17, 18). Additional problems with this indirect approach may arise from alternate mechanisms through which TyrNO 2 can be formed without the involvement of ⅐ NO 2 radicals (19). DHR can be oxidized to the fluorescent rhodamine molecule by various one-electron oxidants, including compounds I and II of peroxidases (20,21 2 The abbreviations used are: DHR, dihydrorhodamine 123; CBA, coumarin-7-boronic acid; CBE, coumarin-7-boronic acid, pinacolate ester; COH, 7-hydroxycoumarin; PAPA-NONOate, (Z)-1-[N-(3-ammoniopropyl)-N-(n-propyl)amino]diazen-1-ium-1,2-diolate; X, xanthine; XO, xanthine oxidase; SOD, superoxide dismutase; DTPA, diethylenetriaminepentaacetic acid; HPLC, high pressure liquid chromatography.