The over-exploitation of coastal aquifers has led to seawater intrusion issues in many parts of the globe; this problem, which is associated with water recharge deficit and anthropogenic pollution, represents the main source of groundwater degradation in Santiago Island in the Republic of Cape Verde’s archipelago. Brackish groundwater for agriculture and human consumption is being provided to several areas on Santiago Island as the only type of available water. Chemical and isotopic data obtained in three main groundwater systems were used in the characterization of the groundwater resources and in the identification of the main source responsible for their degradation. The obtained results indicate water–rock interaction as the major process responsible for the groundwater quality reflecting its lithological composition. Carbonatite dissolution can be partially responsible for the calcium increase along the groundwater flow path. Isotopic data (δ2H, δ18O; 3H and 14C) combined with the water chemistry provided a wide characterization of the groundwater recharge and identification of salinization processes (like seawater intrusion and marine aerosols dissolution in different sectors of the island). In the eastern part of Santiago Island, a different isotopic pattern (2H-18O) was observed in the groundwater samples, which was likely ascribed to different climate conditions. Carbon-14 determinations indicate apparent groundwater ages between 3.5 and 5.1 ka BP.