Isolation of γ-Lactam Alkaloids from the Macrolepiota neomastoidea

Abstract

Macrolepiota neomastoidea Hongo (Agaricaceae) is a poisonous mushroom, which is widely distributed throughout Korea and other East Asian countries. M. neomastoidea has been known to cause severe gastrointestinal symptoms including intestinal irritation, vomiting and profuse diarrhea. To the best of our knowledge, the active principles of this mushroom are unknown except for two compounds, lepiotins A and lepiotins B. Therefore, as part of our systematic study of Korean toxic mushrooms, we investigated the constituents of M. neomastoidea collected at Mt. Jiri, Namwon of Jeonbuk province, in Korea in November, 2005. Half dried aerial parts of M. neomastoidea were extracted with 80% aqueous MeOH at room temperature. The concentrated MeOH extract was partitioned with n-hexane, CHCl3 and nBuOH. Purification of the n-BuOH fraction by repeated column chromatography furnished four γ-lactam pyrrolidinone alkaloids, lepiotins A (1), lepiotins B (2), lepiotins C (3) and (R)-5-hydroxypyrrolidin-2-one (4). Although compound 3 has previously been reported as a synthetic compound, here we have isolated it for the first time from a natural source and named it lepiotins C. Moreover, the spectral data of isolated lepiotins C (3) were little different from those of synthetic compound. Lepiotins A (1) and lepiotins B (2) were isolated from this mushroom by Tomihisa et al., but their absolute configurations of these compounds at C-5 were not clarified. The structures of the isolated metabolites, Lepiotins A (1), lepiotins B (2) and (R)-5-hydroxypyrrolidin-2-one (4) were determined by comparison of spectral data with those reported previously. The compound 4 was for the first time isolated from this mushroom. This paper describes the isolation and structure elucidation of compound 3, as well as the determination of absolute configurations of 1 and 2 by the convenient Mosher’s method and Circular Dichroism (CD) study. Compound 3 was obtained as a colorless gum, which tested positive against Dragendorff reagent. Its molecular formula was determined to be C10H11NO2 from the [M + H] peak at m/z 178.0866 (C10H12NO2, calcd. for 178.0868) in the positive-ion high resolution (HR)-FAB-MS spectrum. The IR spectrum indicated that 3 possessed hydroxyl (3443 cm−1) and carbonyl (1662 cm−1) groups. The H-NMR spectrum (Table 1) of 3 displayed signals for the presence of two methylene groups at δH: 1.89-1.98 (2H, m) and at δH: 2.42-2.45 (2H, t, J = 7.0 Hz), and one methylene group adjacent to the nitrogen function at δH: 3.21-3.23 (2H, t, J = 7.0 Hz). The Hand C-NMR spectra of 3 exhibited signals for two sets of methine groups (δH/δC; 6.67/114.9, 7.35/120.8) on a 1,4-disubstituted aromatic ring (Figure 1). The C-NMR spectrum displayed ten carbon signals, composed of a carbonyl carbon of amide, one benzene ring, and three methylene carbons (Table 1). The C-NMR resonances were similar to those of the related alkaloid, lepiotins A (1), except for the replacement of the hydroxylated methine group at C-5 (δC: 87.3) in 1 with the methylene group at C-5