Samples of natural groundwater (with low turbidity, neutral pH and 0.3 mg L-1 iron concentration) inoculated with Escherichia coli K-12 were exposed to simulated solar light both in the presence and in the absence 10 mg L-1 of H2 O2. Results demonstrated that the viability of E. coli (by DVC-FISH) was grounded to zero after 360 min of irradiation. This abatement could be caused by the oxidative stress induced by ·OH radicals or another photo-induced reactive oxygen species. Two 23 factorial experimental designs enabled the evaluation of the effects of chemical factors on the inactivation of E. coli. The first experimental design considered the pH, iron and H2 O2 , while the second evaluated the ions fluoride, carbonate and phosphate found in groundwater. pH was found to play a key role in the inactivation of E. coli. The best reduction in viability was obtained at the lower pH (6.75), while a nonsignificant effect was observed when iron or H2 O2 concentrations were raised. At higher concentrations, anions, such as carbonate and phosphate, negatively affected the E. coli abatement. However, a higher concentration of fluoride accelerated it. In all experiments, the pH was observed to rise to values higher than 8.0 units after 360 min of treatment.