BACKGROUND: Propolis is a relevant research subject worldwide. However, there is no information so far on Bolivian propolis. Ten propolis samples were collected from regions with high biodiversity in the main honey production places in Bolivia and were analyzed for their total phenolics (TP), flavonoids (TF) and antioxidant activity. The chemical profiles of the samples were assessed by TLC, HPLC-DAD, HPLC-DAD-MS/MSn and NMR analysis. RESULTS: TP, TF, TLC and NMR analysis showed significant chemical differences between the samples. Isolation of the main constituents by chromatography and identification by HPLC-DAD-MS/MSn achieved more than 35 constituents. According to their profiles, the Bolivian propolis can be classified into phenolic-rich and triterpene-rich samples. Propolis from the valleys (Cochabamba, Chuquisaca and Tarija) contained mainly prenylated phenylpropanoids, while samples from La Paz and Santa Cruz contained cycloartane and pentacyclic triterpenes. Phenolic-rich samples presented moderate to strong antioxidant activity while the triterpene-rich propolis were weakly active. CONCLUSION: High chemical diversity and differential antioxidant effects were found in Bolivian propolis. Our results provide additional evidence on the chemical composition and bioactivity of South American propolis.