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This study was designed to characterize the phenolic profile and evaluate the antibacterial, antimutagenic and antitumor activities of Veronica urticifolia Jacq. methanolic extract. HPLC-DAD/ESI-MS analysis revealed the presence of phenolic acids, flavonoids and phenylethanoids, with acteoside as the main component (14.9 mg/g of extract). Antibacterial effect was determined using the microbroth dilution assay and Staphylococcus aureus was the most sensitive strain (MIC and MBC = 7.5 mg/mL). Antimutagenic activity was evaluated by Ames mutagenicity assay. At 1 mg/plate, the tested extract afforded high protection against the mutagenicity of nitroquinoline-N-oxide (4NQO) to Salmonella typhimurium strain TA100 (inhibition rate 48.3%). Antitumor activity was screened in Ehrlich ascites carcinoma (EAC) model. Pretreatment with 2 mg/kg body weight showed statistically significant decrease in tumor cell viability, while ascites volume and tumor cell count became slightly decreased, but not to a statistically significant extent. Above results indicate that V. urticifolia deserves further research into chemoprevention. ; Ministry of Education, Science and Technological Development of Serbia for financial support, project number 173015. The authors are grateful to Fundação para a Ciência e a Tecnologia (FCT, Portugal) for financial support to CIMO (strategic project PEst-OE/AGR/UI0690/2011). J.C.M. Barreira thanks FCT, POPH-QREN and FSE for his grant (SFRH/BPD/72802/2010). The GIP-USAL is financially supported by the Spanish Government through the Consolider-Ingenio 2010 Programme (FUN-C-FOOD, CSD2007-00063) and project ref. BFU2012-35228