The extracts of 14 Julianaceae and 5 Clusiaceae species growing in Mexico were tested in vitro (50 µ g/mL) against Mycobacterium tuberculosis H37Rv and HIV reverse transcriptase (HIV-RT). The Julianaceae bark and leaf extracts inhibited M. tuberculosis (>84.67%) and HIV-RT (<49.89%). The Clusiaceae leaves extracts also inhibited both targets (>58.3% and >67.6%), respectively. The IC 50 values for six selected extracts and their cytotoxicity (50 µ g/mL) to human macrophages were then determined. Amphipterygium glaucum , A. molle , and A. simplicifolium fairly inhibited M. tuberculosis with IC 50 of 1.87–2.35 µ g/mL; but their IC 50 against HIV-RT was 59.25–97.83 µ g/mL. Calophyllum brasiliense , Vismia baccifera , and Vismia mexicana effect on M. tuberculosis was noteworthy (IC 50 3.02–3.64 µ g/mL) and also inhibited RT-HIV (IC 50 26.24–35.17 µ g/mL). These 6 extracts (50 µ g/mL) presented low toxicity to macrophages (<23.8%). The HPLC profiles of A. glaucum , A. molle , and A. simplicifolium indicated that their antimycobacterial activity cannot be related to masticadienonic, 3 α , or 3 β -hydromasticadienonic acids, suggesting that other compounds may be responsible for the observed activity or this might be a synergy result. The anti-HIV-RT and antimycobacterial activities induced by C. brasiliense can be attributed to the content of calanolides A, B, as well as soulatrolide.