The one-electron reduction of 8-hydroxy-5-deazaisoalloxazine (HMDI) has been studied in aqueous solution in the acidity range pH 0 to 13 using the reducing species CO2.-, eaq- and (CH3)2COH radicals. The spectral and other properties of the HMDI radicals were found to be independent of the reductant used. Four protolytic forms of the radical were distinguished with associated pKa values of 2.3 +/- 0.3, 6.0 +/- 0.3 and 10.1 +/- 0.3.