The páramo lakes, Colombia, situated in the neotropical region, are of great value both as natural heritage and as water source to the most populated areas of the Andes, but are threatened by expanding agriculture, livestock, mining, and landscape fragmentation. Nonetheless, a general assessment of the lakes’ distribution and biogeochemical characteristics was lacking. We made a complete inventory of the Colombian páramo lakes and characterized their morphometry and water chemistry based on a survey of 51 lakes in the Eastern Cordillera. There are 3250 lakes distributed across 28 páramo complexes in Colombia, mainly located between 3600 and 4400 m a.s.l. The lakes are usually small (<10 ha) and shallow (<10 m). Most of them are slightly acidic (average pH~6), with high nutrient (total phosphorus ~0.6 mg/L, total nitrogen ~1 mg/L) and total organic carbon (~6 mg/L) concentrations and low oxygen (~3.5 mg/L) at the bottom. Water chemistry varies according to two main independent gradients related to watershed bedrock geology and trophic state. Global change pressures may be challenging the preservation of these unique ecosystems. Increasing the protection of more lake watersheds should reduce these potential impacts by mitigating negative synergies with local pressures.