A new iridoid, 10-acetylborreriagenin (1), and five known iridoid glycosides (2-6), were isolated from the aerial parts of Hedyotis pilulifera. Their structures were elucidated by spectral analyses, including 1D- and 2D-NMR, and HR-ESI-MS, and comparisons with the NMR data reported in the literature. The isolated compounds 1-6 were tested against six bacterial species. Among them, 10-acetylborreriagenin (1) showed antibacterial activity against Staphylococcus aureus, with an MIC value of 100 µg/mL.