To comprehensively understand the chemical constituents of the edible mushroom Agrocybe salicacola and their biological functions, a phytochemical separation of the cultural broth of A. salicacola led to the isolation of four new illudane sesquiterpenoids, agrocybins H-K (1-4), along with 10 known analogues (5-14). Compounds 2-4 were racemates of which 2 and 3 were further separated into single enantiomers as 2a/2b and 3a/3b. All new structures with absolute configurations were elucidated on the basis of an extensive spectroscopic analysis and quantum chemistry calculations. Compound 1 possesses a new carbon skeleton that might be derived from the protoilludane backbone. Compounds 1, 5, 8, and 9 show a certain degree of cytotoxicity to five human cancer cell lines. Compound 1 shows a mild inhibitory effect on nitric oxide production with an IC50 value of 31.4 μM. It is concluded that A. salicacola is rich in illudin derivatives with potential bioactivity prospects, which would make A. salicacola a good material of medicine and food homology.