Two New Chemical Constituents from Leaves of Perilla frutescens var. acuta

Experimental Section

Plant Material. The leaves of P. frutescens var. acuta (25 kg) were collected from Namwon in Jeollanam-do, Korea, in April 2012. The plants were authenticated by one of the authors (K.R. Lee). A voucher specimen (SKKU-NPL-1207) of the plant was deposited in the herbarium of the School of Pharmacy, Sungkyunkwan University, Suwon, Korea.

Extraction and Isolation. The dried leaves of P. frutescens var. acuta (25 kg) were extracted with petroleum ether, and methanol, successively and evaporated under reduced pressure to give residues (264 g and 2 kg, respectively). The MeOH extracts (1 kg) were suspended in distilled water (2.4 L) and then successively partitioned with n-hexane, CHCl3, EtOAc, and hydrated n-BuOH, 190 g, 144 g, 60 g, and 95 g, respectively. The purification of 36 compounds (1-36) is described in Supplementary material.

Perillascens (1). Colorless gum. + 1.4 (c = 0.30 MeOH); UV (MeOH) λmax (log ε) 225 (4.33), 284 (4.27), 314 (4.44) nm; IR (KBr) νmax 3359. 2938, 2844, 1728, 1652, 1598, 1516, 1454, 1372, 1273, 1125, 1032, 872 cm-1; 1H and 13C NMR: see Table 1. Positive-ion HR-FAB-MS m/z = 344.1497 [M + H]+ (calcd. for C19H22NO5: 344.1498).

Perillaside (36). Colorless gum. IR (KBr) νmax 3307, 2916, 1991, 1671, 1561, 1509, 1371, 1155, 1074, 1035, 698 cm-1; 1H and 13C NMR: see table 2. Positive-ion HR-FABMS m/z = 345.1548 [M+ H]+ (calcd. for C16H25O8: 345.1549).

Acidic Hydrolysis and Sugar Identification. Compound 36 (1.0 mg) was heated at 60 ℃ and stirred with 1 mL of 7% HCL for 2 h. After cooling, the reaction mixture was neutralized with an Amberlite IRA400 column, and the eluate was extracted. The sugars residue obtained from the hydrolysis were dissolved in anhydrous pyridine (0.5 mL) and L-cysteine methyl ester hydrochloride (2 mg) was added. The mixture was stirred at 60 ℃ for 1.5 h. After the reaction mixture was dried in vacuo, the residue was trimethylsilylated with 1-trimethylsilylimidazole (0.1 mL) for 2 h. The mixture was partitioned between hexane and H2O (1 mL each), and organic layer (1 μL) was analyzed by GCMS. Identification of D-glucopyranoside for 36 was detected in each case by co-injecion of hydrolysate with standard silylated samples, giving single peaks at D-glucopyranose (10.12 min) for 36. Retention times of authentic samples treated in the same way with 1-trimethylsilylimidazole in pyridine, were D-glucopyranose (10.14 min).

D-Glucopyranose was identified by co-TLC with standard samples and comparison of Rf values with the literature; Dglucopyranose (CHCl3-MeOH-H2O = 6:4:1. Rf value : 0.31).