The genus Michelia (Magnoliaceae) consists of about 30 species. Michelia alba is an evergreen tree, especially distributed in Taiwan and China. Michelia species have been used by indigenous peoples for the treatment of cancer. For example, Michelia champaca has been used in India for the treatment of abdominal tumors and M. hypoleuca and M. officinalis for carcinomatous sores and leukemia, respectively, in China [1]. Previous phytochemical studies on this species isolated, from the roots and flowers of M. alba, ushinsunine, oxoushinsunine, salicifoline, michelalbine, limonene, benzyl acetate, linalool, nerol, hydroxycitronellal, benzaldehyde, benzyl benzoate, and methyl eugenol [2, 3]. To further understand the chemotaxonomy and to continue searching for novel agents from Magnoliaceous plants, the leaves of M. alba were chosen for the first time for phytochemical investigation. In this paper, we report the isolation of 21 pure substances. The compounds included four aporphines, (−)-anonaine ( 1 ) [4], (−)-norushinsunine ( 2 ) [5], (−)-ushinsunine ( 3 ) [5], and (−)-N-acetylanonaine ( 4 ) [6]; two oxoaporphines, liriodenine ( 5 ) [5] and oxoxylopine (6) [7]; three sesquiterpene lactones, michelenolide ( 7 ) [8], costunolide ( 8 ) [8], and 11,13-dehydrolanuginolide ( 9 ) [9]; one amide, N-trans-feruloyltyramine ( 10 ) [10]; one lignan, (+)-syringaresinol ( 11 ) [11]; three benzenoids, 4-hydroxybenzaldehyde ( 12 ) [12], 4-hydroxybenzoic acid ( 13 ) [12], and methylparaben ( 14 ) [12]; two steroids, β-sitosterol and stigmasterol [13]; three aliphatic compounds, palmitic acid [14], stearic acid [15], and linoleic acid [16]; and two chlorophylls, pheophorbide a [17] and aristophyll-C [18]. In addition to 3 and 5, all of these compounds were isolated for the first time from this source.

figure 1

The leaves of M. alba were collected from Fooyin University, Kaohsiung County, Taiwan in March, 2005. A voucher specimen (Michelia 2) was characterized by Dr. Yen-Ray Hsui of the Division of Silviculture, Taiwan Forestry Research Institute, Taipei, Taiwan and deposited in the Basic Medical Science Education Center, Fooyin University, Kaohsiung County, Taiwan. The air-dried leaves of M. alba (6.0 kg) were extracted with MeOH (70 L × 6) at room temperature, and a MeOH extract (367.8 g) was obtained upon concentration under reduced pressure. The MeOH extract, suspended in H2O (1 L), was partitioned with CHCl3 (2 L × 5) to give fractions soluble in CHCl3 (154.3 g) and H2O (144.1 g). The CHCl3-soluble fraction was chromatographed over silica gel (800 g, 70–230 mesh) using n-hexane/CHCl3/MeOH mixtures as eluents to produce five fractions. Part of fraction 1 (43.12 g) was subjected to silica gel chromatography by eluting with n-hexane-EtOAc (5:1) and enriched gradually with EtOAc to furnish five fractions (1-1-1-4). Fraction 1-1 (12.89 g) was further purified on a silica gel column using n-hexane/EtOAc mixtures to obtain palmitic acid (52 mg), stearic acid (27 mg), and linoleic acid (37 mg).

Fraction 1–2 (6.23 g) was further purified on a silica gel column using n-hexane/EtOAc mixtures to obtain pheophorbide a (25 mg) and aristophyll-C (51 mg). Fraction 1–3 (15.31 g) was further purified on a silica gel column using n-hexane/EtOAc/MeOH mixtures to obtain a mixture of β-sitosterol and stigmasterol (144 mg). Fraction 1-4 (4.11 g) was further purified on a silica gel column using n-hexane/EtOAc/MeOH mixtures to obtain methylparaben ( 14 ) (15 mg). Part of fraction 2 (9.21 g) was subjected to silica gel chromatography by eluting with n-hexane-EtOAc (1:1) enriched with EtOAc to furnish five further fractions (2-1-2-4). Fraction 2-2 (1.43 g) was purified on a silica gel column (200 g, 230–400 mesh) using CHCl3/MeOH mixtures to obtain 4-hydroxybenzaldehyde ( 12 ) (21 mg) and 4-hydroxybenzoic acid ( 13 ) (18 mg). Part of fraction 3 (13.25 g) was subjected to silica gel chromatography by eluting with n-hexane-EtOAc (1:1) and enriched gradually with EtOAc to furnish five fractions (3-1-3-3). Fraction 3-1 (5.53 g) was further purified on a silica gel column using n-hexane/EtOAc mixtures to obtain michelenolide (7) (12 mg). Fraction 3-3 (4.87 g) was further purified on a silica gel column using n-hexane/EtOAc mixtures to obtain 11,13-dehydrolanuginolide ( 9 ) (34 mg). Part of fraction 4 (38.54 g) was subjected to silica gel chromatography by eluting with CHCl3-MeOH (40:1), enriched with MeOH to furnish five fractions (4-1-4-5). Fraction 4-1 (3.02 g) was subjected to further silica gel chromatography by eluting with CHCl3-MeOH (100:1) and enriched gradually with MeOH to obtain four fractions (4-1-1-4-1-4). Fraction 4-1-1 (0.52 g) was further purified by passage over another silica gel column using n-hexane/EtOAc mixtures to obtain costunolide ( 8 ) (11 mg). Fraction 4-1-2 (0.74 g), eluted with n-hexane-EtOAc (1:7), was further separated using silica gel column chromatography and preparative TLC (CHCl3-MeOH (100:1) and gave (+)-syringaresinol ( 11 ) (15 mg). Fraction 4-2 (5.73 g) was subjected to further silica gel chromatography by eluting with CHCl3-MeOH (100:1) and enriched gradually with MeOH, to obtain four fractions (4-2-1-4-2-3). Fraction 4-2-1 (2.11 g) was further purified by passage over another silica gel column using EtOAc/MeOH mixtures to obtain liriodenine ( 5 ) (25 mg). Fraction 4-2-2 (0.44 g), eluted with EtOAc/MeOH (20:1), was further separated using silica gel column chromatography and preparative TLC (EtOAc-MeOH (100:1) and gave oxoxylopine ( 6 ) (5 mg). Fraction 4-3 (15.12 g) was subjected to further silica gel chromatography by eluting with CHCl3-MeOH (50:1) and enriched gradually with MeOH to obtain three fractions (4-3-1-4-3-3). Fraction 4-3-2 (3.15 g), eluted with CHCl3-MeOH (100:1), was further separated using silica gel column chromatography and preparative TLC (CHCl3-MeOH (100:1) and gave (−)-N-acetylanonaine ( 4 ) (14 mg). Fraction 4-3-3 (7.75 g), eluted with CHCl3-MeOH (40:1), was further separated using silica gel column chromatography and gave (−)-anonaine ( 1 ) (46 mg). Fraction 4-5 (8.74 g) was subjected to further silica gel chromatography by eluting with CHCl3-MeOH (80:1) and enriched gradually with MeOH to obtain four fractions (4-5-1-4-5-5). Fraction 4-5-2 (4.13 g) was further purified on a silica gel column using CHCl3/MeOH mixtures to obtain (−)-norushinsunine ( 2 ) (48 mg). Fraction 4-5-4 (1.27 g) was further purified on a silica gel column using CHCl3/MeOH mixtures to obtain (−)-ushinsunine ( 3 ) (58 mg). Part of fraction 5 (15.01 g) was subjected to silica gel chromatography by eluting with CHCl3-MeOH (50:1) and enriched gradually with MeOH, to furnish five fractions (5-1-5-4). Fraction 5-2 (4.07 g) was further purified on a silica gel column using CHCl3/MeOH mixtures to obtain N-trans-feruloyltyramine ( 10 ) (26 mg).