In the late 1950's free radicals and antioxidants were almost unheard of in the clinical and biological sciences but chemists had known about them for years in the context of radiation, polymer and combustion technology. Daniel Gilbert, Rebeca Gerschman and their colleagues related the toxic effects of elevated oxygen levels on aerobes to those of ionizing radiation, and proposed that oxygen toxicity is due to free radical formation, in a pioneering paper in 1956. Biochemistry owes much of its early expansion to the development and application of chromatographic and electrophoretic techniques, especially as applied to the study of proteins. Thus, superoxide dismutase (SOD) enzymes (MnSOD, CuZnSOD, FeSOD) were quickly identified. By the 1980's Molecular Biology had evolved from within biochemistry and microbiology to become a dominant new discipline, with DNA sequencing, recombinant DNA technology, cloning, and the development of PCR representing milestones in its advance. As a biological tool to explore reaction mechanisms, SOD was a unique and valuable asset. Its ability to inhibit radical reactions leading to oxidative damage in vitro often turned out to be due to its ability to prevent reduction of iron ions by superoxide. Nitric oxide (NO.) provided the next clue as to how SOD might be playing a critical biological role. Although NO. is sluggish in its reactions with most biomolecules it is astoundingly reactive with free radicals, including superoxide. Overall, this high reactivity of NO. with radicals may be beneficial in vivo, e.g. by scavenging peroxyl radicals and inhibiting lipid peroxidation. If reactive oxygen species are intimately involved with the redox regulation of cell functions, as seems likely from current evidence, it may be easier to understand why attempts to change antioxidant balance in aging experiments have failed. The cell will adapt to maintain its redox balance. Indeed, transgenic animals over-expressing antioxidants show some abnormalities of function. There must therefore be a highly complex interrelationship between dietary, constitutive, and inducible antioxidants with the body, under genetic control. The challenge for the new century is to be able to understand these relationships, and how to manipulate them to our advantage to prevent and treat disease.