Mitochondria-Targeted Spin Traps: Synthesis, Superoxide Spin Trapping, and Mitochondrial Uptake

Abstract

Development of reliable methods and site-specific detection of free radicals is an active area of research. Here, we describe the synthesis and radical-trapping properties of new derivatives of DEPMPO and DIPMPO, bearing mitochondria-targeting triphenylphosphonium cationic moiety or guanidinium cationic group. All the spin traps prepared have been observed to efficiently trap superoxide radical anion in cell-free system. The superoxide spin adducts exhibited similar spectral properties indicating no significant differences in the geometry of the cyclic nitroxide moieties of the spin adducts. The superoxide adduct stability was measured and observed to be highest (t1/2 = 73 min) for DIPPMPO nitrone linked to triphenylphosphonium moiety via a short carbon chain (Mito-DIPPMPO). The experimental results and DFT quantum chemical calculations indicate that the cationic property of the triphenylphosphonium group may be responsible for increased superoxide trapping efficiency and adduct stability of Mito-DIPPMPO, as compared to DIPPMPO spin trap. The studies of uptake of the synthesized traps into isolated mitochondria indicated the importance of both cationic and lipophilic properties, with the DEPMPO nitrone linked to triphenylphosphonium moiety via a long carbon chain (Mito10-DEPMPO) exhibiting the highest mitochondrial uptake. We conclude that of the synthesized traps, Mito-DIPPMPO and Mito10-DEPMPO are the best candidates as the potential mitochondria-specific spin traps for use in biologically-relevant systems.

Antiproliferative effects of mitochondria-targeted cationic antioxidants and analogs: Role of mitochondrial bioenergetics and energy-sensing mechanism

“…Blunting the antioxidant functions of Mito-CP (by changing the nitroxide group to an acetamido group) and Mito-chromanol (by estrification of the phenolic hydroxyl group) did not affect the antiproliferative effects of Mito-CP-Ac and Mito-chromanol acetate in several cancer cells [11]. Mito-CP and Mito-chromanol exhibit superoxide dismutating and radical scavenging abilities [16]. The fact that both Mito-CP (O 2 •− dismutating agent) and Mito-CP-Ac (lacking O 2 •− dismutating ability) mitigate tumor proliferation to the same extent argues against mitochondrial ROS intermediacy as a regulatory factor in tumor proliferation mechanism.…”

“…DIPPMPO spin trap was synthesized, as reported previously [15,16]. Mito-CP was synthesized according to the previously published procedure [17].…”

Toward selective detection of reactive oxygen and nitrogen species with the use of fluorogenic probes – Limitations, progress, and perspectives

“…Several ROS-sensitive probes producing easily detectable and relatively stable products have been developed. In addition to spin trapping techniques [5,6], luminescent probes have also become widely used tools in the studies on oxidative stress.…”

Taking up the cudgels for the traditional reactive oxygen and nitrogen species detection assays and their use in the cardiovascular system

“…much more sensitive electrochemical detection of the DHE products) or new DHE analogs allowing sitespecific detection of superoxide (e.g. mito-SOX for mitochondrial superoxide and hydropropidine for extracellular superoxide) or the most advanced new spin traps providing higher stability in biological samples and thereby allow the EPR-based detection of superoxide and/ or peroxynitrite-derived free radicals in cells and tissues (some candidates might be found in [32,[135][136][137]). High performance liquid chromatography (HPLC)-based detection of superoxide formation in the extracellular space, cytosol and mitochondria can be established by hydropropidine, a positively-charged water-soluble analogue of dihydroethidium (DHE) for extracellular superoxide formation [138], DHE for cytosolic superoxide formation and mitoSOX, a mitochondria-targeted DHE analogue for mitochondrial superoxide formation (Fig.…”

“…The use of classical spin traps such as DMPO or DEPMPO in biological samples is limited by their instability in the presence of cellular antioxidants but they are suitable for detection of superoxide in isolated immune cells [63]. Candidates for new spin traps are Mito-DIPPMPO and Mito10-DEPMPO for mitochondrial superoxide formation [136] or FDMPO, DPPMPO, DBPMPO, DEHPMPO and DIPPMPO with superior half-life of HOOand HO-spin adducts in biological samples [152].…”