A pair of enantiomeric polyketides, (+)- and (-)-alternamgin (1), featuring an unprecedented 6/6/6/6/5/6/6 seven ring backbone, were isolated from the endophytic fungi Alternaria sp. MG1. The relative configuration of 1 was determined using X-ray diffraction, and the absolute configurations of (±)-1 were confirmed by comparing the experimental and calculated ECD data. Plausible biosynthetic pathways for 1 were proposed. Compound (-)-1 exhibited moderate necrosis rates to Hela and HepG2 cells, but (+)-1 only showed similar necrosis rates to HepG2 cells.