Diatoms are frequently exposed to high light (HL) levels, which can result in photoinhibition and damage to PSII. Many microalgae can photoreduce oxygen using the Mehler reaction driven by PSI, which could protect PSII. The ability of Nitzschia epithemioides Grunow and Thalassiosira pseudonana Hasle et Heimdal grown at 50 and 300 μmol photons · m−2 · s−1 to photoreduce oxygen was examined by mass spectrometric measurements of 18O2. Both species exhibited significant rates of oxygen photoreduction at saturating light levels, with cells grown in HL exhibiting higher rates. HL-grown T. pseudonana had maximum rates of oxygen photoreduction five times greater than N. epithemoides, with 49% of electrons transported through PSII being used to reduce oxygen. Exposure to excess light (1,000 μmol photons · m−2 · s−1) produced similar decreases in the operating quantum efficiency of PSII (Fq′/Fm′) of low light (LL)- and HL-grown N. epithemoides, whereas HL-grown T. pseudonana exhibited much smaller decreases in Fq′/Fm′ than LL-grown cells. HL-grown T. pseudonana and N. epithemioides exhibited greater superoxide and hydrogen peroxide production, higher activities (in T. pseudonana) of superoxide dismutase (SOD) and ascorbate peroxidase (APX), and increased expression of three SOD- and one APX-encoding genes after 60 min of excess light compared to LL-grown cells. These responses provide a mechanism that contributes to the photoprotection of PSII against photodamage.