The source of hydrogens for the quenching of the phenylene diradical produced by esperamicin A (espA) and by esperamicin C (espC) in the presence of specifically deuteriated, double-stranded DNA (dsDNA) has been determined. Remarkably, both espA and espC were quenched at both positions of the diphenylene radical by exclusive abstraction of hydrogen from dsDNA. EspC, a predominantly ds-cleaver, afforded results consistent with 4’- and 5’-hydrogen transfer. EspA, a predominantly ss-cleaver, revealed no 4’-hydrogen transfer; however, results were consistent with 5’-hydrogen transfer and with the recently proposed 1’-hydrogen transfer (Yu, L.; Golik, J.; Harrison, R.; Dedon, P. J. Am. Chem. Sec. 1994, 116, 9733-9738). For espA, insufficient double strand DNA damage was produced to account for the role of DNA as the exclusive hydrogen source. Tn order to resolve this discrepancy, several reductants were used to activate espA and espC. The results indicated that a substantial portion of radical lesions produced in DNA by the esperamicins is subjected to repair by hydrogen transfer from the reductant. The efficiency of repair depended on the structural features of the reductant. The findings demonstrate that caution must be exercised when evaluating the propensity of DNA cleavers for ss- and ds-cleavage in the presence of reductants.