Chemical composition of the leaves of Periploca sepium

Abstract

The genus Periploca (Asclepiadaceae) contains 12 species, of which there are four species in China. Periploca sepium Bage, widely distributed in the northeast, southeast, northwest, and southwest of China, is one of the four species. The root of Periploca sepium Bage has been used as a traditional Chinese herbal medicine for more than two thousand years [1]. Previous phytochemical studies of this species have isolated, from the root of P. sepium, cardenoides, pregnane glycosides, etc. [2, 3]. For many years, research on the plant has mainly concentrated on the underground part. To continue searching for novel agents from P. sepium, the leaves of P. sepium were chosen for phytochemical investigation. In this paper, we report the isolation and identification of four compounds. The leaves of P. sepium Bage were collected from Qinling Mountain, Shaanxi, China, in July, 2006 and identified by Prof. E. X. Han of Northwest A & F University. The voucher specimen was deposited in the Forestry College, Northwest A & F University. Air-dried and powdered leaf of P. sepium Bage (3.9 kg) was extracted exhaustively with 95% ethanol at room temperature. The ethanol extract was concentrated in vacuum at 60 C to give a residue. The residue was suspended in water and successively extracted with petroleum ether, ethyl acetate, and n-butanol. The ethyl acetate extract (47.0 g) was subjected to repeated silica gel column chromatography with Sephadex LH-20 using different percentages of chloroform–methanol mixtures, and compounds 1 and 2 were obtained. Repeated chromatography of the n-butanol extract (47.3 g) on silica gel with chloroform–methanol and Sephadex LH-20 afforded compounds 3 and 4. The structures of these compounds were elucidated by a combination of spectral analysis and were determined as oleanolic acid (1) [4], quercetin (2) [5], 6 -methyl ester of quercetin-3-O-D-glucuronide (3) [6], and isoquercitrin (4) [7] by comparison with the data in the literature. All these four compounds were isolated for the first time from P. sepium, and compound 2 was isolated for the first time from the genus of Periploca. Oleanolic acid (1) white powder, mp > 300 C. IR (KBr, max, cm –1): 3446 (OH), 2941, 2861, 1693 (C=O), 1636 (C=C), 1462, 1385, 1274, 1084, 1036, 749. 1H NMR (300 MHz, CDCl3, , ppm): 0.74 (3H, s, CH3, H-26), 0.77 (3H, s, CH3, H-23), 0.90 (3H, s, CH3, H-24), 0.91 (3H, s, CH3, H-29), 0.92 (3H, s, CH3, H-30), 0.98 (3H, s, CH3, H-25), 1.13 (3H, s, CH3, H-27), 5.28 (1H, br.s, H-12), 3.24 (1H, br.s, H-3), 2.81 (1H, br.s, H-18). Quercetin (2) yellow needle crystals, mp > 300 C. UV (CH3OH, max, nm): 381.0, 257.0, (CH3OH + NaOH, max, nm): 419.0, 330.0, 280.0, (CH3OH + AlCl3, max nm): 448.0, 271.0. 1H NMR (300 MHz, DMSO-d6, , ppm, J/Hz): 6.19 (1H, d, J = 1.8, H-6), 6.41 (1H, d, J = 1.8, H-8), 6.88 (1H, d, J = 8.4, H-5 ), 7.54 (1H, dd, J = 2.0, 8.4, H-6 ), 7.68 (1H, d, J = 2.0, H-2 ), 9.34 (1H, s, 3 -OH), 9.40 (1H, s, 3-OH), 9.62 (1H, s, 4 -OH), 10.80 (1H, s, 7-OH), 12.51 (1H, s, 5-OH). 13C NMR (75 MHz, DMSO-d6, ): 147.24 (C-2), 136.19 (C-3), 176.29 (C-4), 161.17 (C-5), 98.63 (C-6), 164.34 (C-7), 93.79 (C-8), 156.58 (C-9), 103.46 (C-10), 122.40 (C-1 ), 115.51 (C-2 ), 145.51 (C-3 ), 148.15 (C-4 ), 116.05 (C-5 ), 120.42 (C-6 ). 6 -Methyl ester of quercetin-3-O-D-glucuronide (3) yellow needle crystals, mp 181–183 C. 1H NMR (300 MHz, DMSO-d6, , ppm, J/Hz): 6.21 (1H, d, J = 1.1, H-6), 6.41 (1H, d, J = 1.1, H-8), 6.83 (1H, d, J = 8.4, H-5 ), 7.51 (1H, d, J = 2.5, H-2 ), 7.56 (1H, dd, J = 2.5, 8.4, H-6 ), 5.46 (IH, d, J = 6.9, H-1 ), 3.56–3.73 (m, sugar protons), 3.57 (3H, s, COOCH3).