Abstract:In order to understand the chemical constituents of Neoboletus brunneissimus, eighteen compounds were isolated. On the basis of spectral data, they were identified as 5α,8α-epidoxy-(22E,24R)-ergosta-6,22-dien- 3β-ol (1), 5α,8α-epidioxy-(22E,24R)-ergosta-6,9(11),22-trien-3β-ol (2), (22E,24R)-ergosta-7,22-dien-3β-ol (3), (24S)-ergosta-7-en-3b-ol (4), (22E,24R)-ergosta-7,22-dien-3β,5α,6β-triol (5), (24S)-ethylcholest-7-ene-3β,5α,6β- triol (6), (22E,24R)-ergosta-7,22-diene-3β,5α,6β,9α-tetrol (7), (22E,4R)-ergosta-5,7,22-trien-3β-ol (8), 3β-O- glucopyranosyl-5α,8α-epidioxy-(22E,24R)-ergosta-6,22-dien (9), monomethyl fumarate (10), fumaric acid (11), succinic acid (12), trans-2-decenedioic acid (13), nicotinic acid (14), nicotinamide (15), shikimic acid (16), linoleic acid-α-glyceride (17) and 1-O-β-d-glucopyranosyl-(2S,3R,4E,8E,2'R)-2-N-(2'-hydroxypalmitoyl)-9-methyl- 4,8-sphingadienine(18). All compounds were isolated from N. brunneissimus for the first time, compounds 6, 9, 13 and 17 were isolated from the Boletaceae fungi for the first time.