The oxidation-reduction potential of the reaction-centre chlorophyll of Photosystem I (P700) in spinach chloroplasts was determined by using the ability of the reaction centre to photoreduce the bound ferredoxin and to photo-oxidize P700 on illumination at 20K as an indicator of the oxidation state of P700. This procedure shows that P700 is oxidized with Em (pH8.0) (mid-point redox potential at pH8.0) +375mV. Further oxidation of the chloroplast preparations by high concentrations ofK3Fe(CN)6 (10mM) in the presence of mediating dyes leads to the appearance of a large radical signal with an apparent Em. +470mVA second, light-inducible, radical also appears over the same potential range. We propose that these signals are due to bulk chlorophyll oxidation and not, as was previously thought [Knaff & Malkin (1973) induced absorbance change at 435nm, usually attributed to P700, showed a potential dependence similar to that of bulk chlorophyll oxidation. Determination of Em of P700 on the basis of the appearance of the P700 signal in oxidized-versus-reduced difference spectra showed E. (pH8.0) +360mV. Measurements of the effect of potential on the irreversible photo-oxidation of P700 at 77 K showed that P700 became oxidized in this potential range. We conclude that the reaction-centre chlorophyll of Photosystem I has Em (pH8.0) +375mV.The primary event in photosynthesis is thought to be the absorption of light, followed by the photooxidation of a chlorophyll molecule in a special environment, the reaction-centre chlorophyll. This photo-oxidation is coupled to the reduction of an electron acceptor. In 02-evolving organisms two photochemical reactions are involved in photosynthesis, each with a specialized reaction centre.