Detailed chemical, spectroelectrochemical and computational studies have been used to investigate the mechanism of hypoxia selectivity of a range of copper radiopharmaceuticals. A revised mechanism involving a delicate balance between cellular uptake, intracellular reduction, reoxidation, protonation and ligand dissociation is proposed. This mechanism accounts for observed differences in the reported cellular uptake and washout of related copper bis(thiosemicarbazonato) complexes. Three copper and zinc complexes have been characterised by X-ray crystallography and the redox chemistry of a series of copper complexes has been investigated by using electronic absorption and EPR spectroelectrochemistry. Time-dependent density functional theory (TD-DFT) calculations have also been used to probe the electronic structures of intermediate species and assign the electronic absorption spectra. DFT calculations also show that one-electron oxidation is ligand-based, leading to the formation of cationic triplet species. In the absence of protons, metal-centred one-electron reduction gives the reduced anionic copper(I) species, [CuIATSM]−, and for the first time it is shown that molecular oxygen can reoxidise this anion to give the neutral, lipophilic parent complexes, which can wash out of cells. The electrochemistry is pH dependent and in the presence of stronger acids both chemical and electrochemical reduction leads to quantitative and rapid dissociation of copper(I) ions from the mono- or diprotonated complexes, [CuIATSMH] and [CuIATSMH2]+. In addition, a range of protonated intermediate species have been identified at lower acid concentrations. The one-electron reduction potential, rate of reoxidation of the copper(I) anionic species and ease of protonation are dependent on the structure of the ligand, which also governs their observed behaviour in vivo.