Twenty-two new highly oxygenated bisabolane-type sesquiterpenoids, pararubin A-V (1-22), were isolated from the whole plant of Parasenecio rubescens. The structure determination of 1-22, including the assignment of their relative configurations, was accomplished by extensive 1D and 2D NMR spectroscopy. The absolute configuration of pararubin A (1) was established by single-crystal X-ray crystallographic analysis. The isolated compounds were evaluated for their cytotoxic effects against three cancer cell lines (B16 mouse melanoma cells, HepG2 human hepatocellular carcinoma cells, and MCF7 human breast adenocarcinoma cells) and for their antimicrobial effects against Staphylococcus aureus, Escherichia coli, and Monilia albicans.