Ferroptosis is a regulated form of cell death driven by iron-mediated lipid peroxidation. Its execution is initiated by the peroxidation of polyunsaturated fatty acid–containing phospholipids, ultimately leading to membrane rupture. Cells possess multiple defense systems to suppress ferroptosis. The cystine/GSH/GPX4 axis detoxifies and reduces lipid hydroperoxides, whereas several GPX4-independent pathways—such as the FSP1–CoQ10 pathway, the GCH1–BH4 pathway, and the vitamin K cycle—suppress ferroptosis by scavenging lipid radicals and preventing propagation of lipid peroxidation. In addition, endogenous antioxidants including vitamin E, squalene, and 7-dehydrocholesterol also contribute to ferroptosis suppression. Beyond antioxidant defenses, factors such as membrane phospholipid composition and iron homeostasis also determine cellular susceptibility to ferroptosis. Elucidating these regulatory mechanisms provides fundamental insight into ferroptosis biology and also highlights promising therapeutic target against ferroptosis-associated disease conditions, such as cancer, neurodegeneration, and acute organ injuries.

As its name suggests, ferroptosis is fundamentally dependent on the presence of iron. While iron is an essential nutrient for the human body, excess iron or metabolic dysfunction of iron can cause various forms of cellular damage. In ferroptosis, lipid peroxidation serves as the trigger for ferroptosis, and iron, particularly ferrous iron (Fe²⁺), plays a central role in the lipid damage. The reaction between peroxides and ferrous iron, historically known as the Fenton reaction, generates radicals that lead to cell death when they overwhelm the cellular defense mechanisms. Many ferroptosis-inducing agents function by inhibiting the protective mechanisms against damage caused by ferrous iron and lipid peroxides. Consequently, compounds that act as iron chelators or radical scavengers function as ferroptosis inhibitors. Although the relationship between peroxidized lipids and ferroptosis has been extensively studied since the discovery of ferroptosis, the roles and subcellular dynamics of ferrous iron have only recently begun to be investigated. This delay occurred because fluorescent probes capable of detecting ferrous iron were not available at the time of ferroptosis discovery. This review introduces the principles underlying the ferrous iron fluorescent probes developed by our group and presents imaging applications of these probes, including their use in ferroptosis research.

Recently, lipid-derived radicals and oxidized lipids have been reported to play critical roles in cell death, particularly ferroptosis, as well as in inflammatory responses, and have therefore attracted considerable attention. However, because of their extremely high reactivity, techniques for accurately measuring these species have been limited. To accelerate research in this field, direct detection of these molecules is indispensable. Furthermore, the development of appropriate inhibitors will also be required. In this article, we describe methods for the detection and structural analysis of lipid-derived radicals, which serve as the chain reaction centers of lipid peroxidation, as well as analytical techniques for elucidating the structures of oxidized phospholipids.