Pigments are chemical compounds that impart color to materials via absorption, reflection, and refraction. Traditional sources of natural pigments include plant and insect tissues; however, microorganisms such as bacteria, yeasts, algae, and filamentous fungi also hold potential for pigment production. In this study, we investigated pigment production in 20 isolates from our in-house Actinobacteria strain collection, obtained from the banks of the Guaviare and Arauca Rivers in Colombia. Isolates were identified using 16S rRNA gene sequencing. We evaluated different culture media and determined their various bioactivities, including antioxidant, antibacterial, and cytotoxic properties. Promising pigmented hydroalcoholic extracts exhibiting the above-mentioned properties were identified and their chemical compositions were examined using LC-MS. Our findings highlighted a growth medium-dependent effect, as pigment production, coloration, and bioactivity outcomes were influenced by growth media. These results demonstrate the significant potential of Streptomyces isolates as sources of bioactive pigments for diverse applications.