Two new cytochalasans, periconiasin I (1) with 9/6/5 tricyclic ring, and periconiasin J (2) with 5/6/6/5 tetracyclic ring, were isolated from the endophytic fungus Periconia sp. F-31. Their structures including absolute configurations were elucidated through extensive spectroscopic analyses, calculated ECD, and the circular dichroism data of the [Rh2(OCOCF3)4] complex. Compound 1 showed in vitro cytotoxicity against human MCF-7 tumor cell line with an IC50 value of 4.8 lM, compound 2 exhibited weak anti-HIV activity with an IC50 value of 25.0 lM.