Nucleophilicity of NH(2), N3, and O(2) centers of cytosine toward a model quinone methide (o-QM) as alkylating agent has been studied using DFT computational analysis [at the B3LYP/6-311+G(d,p) level]. Specific and bulk effects of water (by C-PCM model) on the alkylation pathways have been evaluated by analyzing both unassisted and water-assisted reaction mechanisms. An ancillary water molecule, H-bonded to the alkylating agent, may interact monofunctionally with the o-QM oxygen atom (passive mechanisms) or may participate bifunctionally in cyclic hydrogen-bonded structures as a proton shuttle (active mechanisms). A comparison of the unassisted with the water-assisted reaction mechanisms has been made on the basis of activation Gibbs free energies (DeltaG(++)). The gas-phase alkylation reaction at N3 does proceed through a passive mechanism that is preferred over both the active (by -6.3 kcal mol(-1)) and the unassisted process. In contrast, in the gas phase, the active assisted processes at NH(2) and O(2) centers are both favored over their unassisted counterparts by -4.0 and -2.2 kcal mol(-1), respectively. The catalytic effect of a water molecule, in gas phase, reduces the gap between the TSs of the O(2) and NH(2) reaction pathways, but the former remains more stable. Water bulk effect significantly modifies the relative importance of the unassisted and water-assisted alkylation mechanisms, favoring the former, in comparison to the gas-phase reactions. In particular, the unassisted alkylation becomes the preferred mechanism for the reaction at both the exocyclic (NH(2)) and the heterocyclic (N3) nitrogen atoms. By contrast, alkylation at the cytosine oxygen atom is a water-catalyzed process, since in water the active water-assisted mechanism is still favored. As far as competition, among all the possible mechanisms, our calculations unambiguously suggest that the most nucleophilic site both in gas phase (naked reagents: N3 >> O(2) >or= NH(2)) and in water solution (solvated reagents: N3 >> NH(2) >> O(2)) is the heterocyclic nitrogen atom (N3) (DeltaG(++)(gas) = +7.1 kcal mol(-1), and DeltaG(++)(solv) = +13.7 kcal mol(-1)). Our investigation explains the high reactivity and selectivity of the cytosine moiety toward o-QM-like structures both in deoxymononucleoside and in a single-stranded DNA, on the basis of strong H-bonding interactions between reactants and solvent bulk effect. It also offers two general reactivity models in water, uncatalyzed and active water-catalyzed mechanisms (for nitrogen and oxygen nucleophiles, respectively), which should provide a general tool for the planning of nucleic acid modification.