key: cord-0008393-1flxylyn authors: HUANG, Sheng-Dian; ZHANG, Yu; HE, Hong-Ping; LI, Shi-Fei; TANG, Gui-Hua; CHEN, Duo-Zhi; CAO, Ming-Ming; DI, Ying-Tong; HAO, Xiao-Jiang title: A new Amaryllidaceae alkaloid from the bulbs of Lycoris radiata date: 2013-07-08 journal: Chin J Nat Med DOI: 10.1016/s1875-5364(13)60060-6 sha: 0ae08d570fd266cc5c5e7f7f21b52b068f67a505 doc_id: 8393 cord_uid: 1flxylyn AIM: To study the Amaryllidaceae alkaloids of the bulbs of Lycoris radiata. METHODS: The chemical constituents were isolated and purified by various chromatographic techniques, and the chemical structures were elucidated on the basis of spectroscopic methods. In addition, the antiviral activities of alkaloids 1–10 were evaluated using flu virus A. RESULTS: One new homolycorine-type alkaloid 2α-methoxy-6-O-ethyloduline (1), together with nine known alkaloids 2α-methoxy-6-O-methyloduline (2), trispherine (3), 8-O-demethylhomolycorine (4), homolycorine (5), 9-O-demethylhomolycorine (6), oduline (7), lycorenine (8), 6α-O-methyllycorenine (9) and O-ethyllycorenine (10) were obtained. CONCLUSION: Alkaloid 1 is a new compound, and 1–3 were major alkaloids in this plant. Alkaloids 1–3 showed weak antiviral activities against flu virus A with IC50 values of 2.06, 0.69, and 2.71 μg·mL-1 and CC50 values of 14.37, 4.79, and 80.12 μg·mL-1, respectively. Amaryllidaceae alkaloids are characteristic constituents of the Amaryllidaceae plant family, whose remarkable biological activities and unique skeletons have attracted great interest as challenging targets for total synthesis and diversity-oriented synthesis [1] [2] [3] [4] [5] [6] [7] [8] [9] . A series of new Amaryllidaceae alkaloids was isolated from Hosta plantaginea Asch. in a previous phytochemical investigation, which showed some inhibition activities against the tobacco mosaic plant virus (TMV) [10] [11] . As a continuation of that research work, an Comparison of the NMR data of 1 with 2 suggested that 1 exhibited structural similarities with 2. Notable differences in the NMR spectra with those of 2 inferred that 1 exhibited closely similar signals to 2, except for the presence of an OEt group and the disappearance of the OMe group in 2. The presence of an OEt group located at C-6 instead of the OMe group in 1 was confirmed by the HMBC correlation of H-6 to C-15 and 1 H-1 H COSY correlation from H-15 to H-17. Thus, the structure of 1 was established and named as 2α-methoxy-6-O-ethyloduline. [13] . The IR (KBr) spectrum displayed absorption bands for a phenyl functional group (1 656, 1 608 and 905 cm −1 ) and a double bond (1 562 cm −1 ). The 1 H NMR spectrum (Table) showed one methyl singlet peak at δ 2. 16 (3H, s, H-16) , two methoxyl groups at δ 3.55 (3H, s, H-15) and δ 3.43 (3H, s, H-14) , two aromatic protons at δ 6.96 (1H, s,H-10) and 6.76 (1H, s, H-7), a signal for the H-atom of C=C double bond [δ 5.46 (1H, s, H-6) ] and a signal for the O-CH 2 -O group [δ 5.93 (2H, br s, H-13) ], respectively. The 13 C NMR spectrum (Table) revealed 19 signals comprising of five sp 2 quaternary C-atoms, five sp 3 CH, three sp 2 CH and three sp 3 CH 2 groups, as well as two OMe groups and one NMe group. The five sp 2 quaternary C-atoms were assignable to four aromatic C-atoms [δ 147.3 (C-8) , 147.1 (C-9), 130.8 (C-6a), 126.9 (C-10a)] and a C-atom of C=C double bond [δ 145.4 (C-4) ], 1 of 3 sp 3 CH 2 groups was attributable to a O-CH 2 -O group [δ 101.1 (C-13) ], another two sp 3 CH 2 groups corresponded to a -CH 2 CH 2 − group. The above data proved that 2 was a homolycorine-type Amaryllidaceae alkaloid. The NMR data of 2 were closely similar to those of the known alkaloid 2α-hydroxy-6-O-methyloduline [14] . The minor difference between them was the presence of an additional methoxyl group signal at δ C (57.2) in 2. The HMBC correlation of H-2 to C-14, as well as the downfield shift of C-2 (δ C 78.1, Δδ C +9.8) [14] confirmed the structure. Thus, the planar structure of 2 was elucidated as shown in Fig. 2 . The relative configuration of 2 was assigned as identical to that of 2α-hydroxy-6-O-methyloduline on the basis of the ROESY experiment and coupling constants [14] . Detailed analysis of the 2D NMR data established the structure of compound 2 as 2α-methoxy-6-O-methyloduline. This is the first time to report the NMR data of this alkaloid [15] . Eight known homolycorine-type alkaloids (3) (4) (5) (6) (7) (8) (9) (10) were also isolated and identified as trispherine (3) [14] , 8-O-demethylhomolycorine (4) [16] , homolycorine (5) [17] , 9-O-demethylhomolycorine (6) [18] , oduline (7) [19] , lycorenine (8) [20] , 6α-O-methyllycorenine (9) [21] , O-ethyllycorenine (10) [22] by comparison of their 1D-NMR data with those in the literature. All of the alkaloids were evaluated for inhibitory activities against flu virus A in vitro [23] , and alkaloids 1-3 showed weak antiviral activities with IC 50 values of 2.06, 0.69 and 2.71 μg·mL −1 , and CC 50 values of 14.37, 4.79 and 80.12 μg·mL −1 , respectively. Perkin-Elmer model 241 polarmeter; Bio-Rad FTS-135 spectrometer; Shimadzu UV-2401A spectrometer; Bruker AM-400 or a DRX-500 instrument (using TMS as internal standard); Finnigan MAT 90 instrument and VG Auto Spec-3000 spectrometer; silica gel (SiO 2 , 300−400 mesh; Qingdao Marine Chemical Co., Ltd., China); MCI gel (CHP20P, 75−150 µm; Mitsubishi Chemical Industries Ltd., Japan); Sephadex LH-20 (40−70 µm; Amersham Pharmacia Biotech AB, Uppsala, Sweden); Merck, Darmstadt, Germany) . Semi-Preparative HPLC was performed on a Zorbax SB-C-18 column (i.d. 9.4 mm × 250 mm; Agilent Co. Ltd., Santa Clara, USA). TLC plates were pre-coated with silica gel GF-254 and HF-254 (Qingdao Marine Chemical Co., Ltd., China). The bulbs of Lycoris radiata were bought from Hengyang, Hunan Province, China, in July 2008, and were identified by GONG Xun. A voucher specimen has been deposited with the Kunming Institute of Botany, Chinese Academy of Sciences, China. The air-dried and powdered sample (180 kg) was extracted with MeOH three times to give a crude extract. The crude extract was adjusted to pH 2−3 by dissolving in 0.5% HCl soln. The aqueous phase was extracted with EtOAc, and then the acidic H 2 O-soluble was adjusted to pH 9−10 with 10% aq.NH 3 soln. and extracted with CHCl 3 to give an alkaline extract (1.4 kg). The alkaline extract was subjected to CC (SiO 2 ; CHCl 3 /MeOH gradient 1 : 0→0 : 1) to afford seven fractions (A 1 −A 7 ). Fraction A 1 (78 g) was applied to MCI gel (MeOH−H 2 O, 30/70−100/0), Sephadex LH-20 (MeOH), and then to silica gel CC eluting with petroleum ether(PE)/acetone/diethylamine (5 : 1 : 0.05) to yield 3 (17 g), 4 (41 mg), 7 (20 mg), and 9 (13 mg), respectively. Fraction A 2 (170 g) was subjected to repeated column chromatography (silica gel, Sephadex LH-20 (MeOH), and HPLC) to yield 1 (27 g), 2 (33 g), 10 (11 mg), respectively. Fraction A 3 (15 g) was purified by silica gel and HPLC to give 5 (14 mg), 6 (44 mg) and 8 (8 mg Homolycorine ( 4.81 (1H, ddd, J = 4.8, 1.8, 1.7 Hz, 2.49 (1H, m, 5.50 (1H, m, 2.72 (1H, dd, J = 9.6, 2.0 Hz, 7.57 (1H, s, , 6.99 (1H, s, 2.64 (1H, dd, J = 9.6, 1.8 Hz, 2.63 (2H, m, 3.14 (1H, ddd, J = 10.0, 7.0, 3.5 Hz, 2.24 (1H, dd, J = 18.0, 9.2 Hz, Oduline ( 3.14 (1H, ddd, J = 9.2, 6.3, 3.8 Hz, 2.25 (1H, dd, J = 18.7, 9.5 Hz, 5.97 (2H, d, J = 14.7 Hz, -OCH 2 O-), 3.67 (1H, s, -OH), 2.11 (3H, s, -NCH 3 ); 13 C NMR (100 MHz, MeOD) δ: 66.7 (C-1), 31.7 (C-2), 115.7 C-10a), 44.0 (C-10b) MeOD 1 : 1) δ: 4.53 (1H, d, J = 5.4 Hz, H-1), 2.50 (1H, m, H-2), 5.70 (1H, s, H-3), 2.68 (1H, br. d, J = 9 MeOD 1 : 1) δ: 67.1 (C-1) 113.0 (C-10), 128.1 (C-10a), 43.6 (C-10b), 28.2 (C-11), 57.1 (C-12) 27 (2H, d, J = 8.0 Hz, H-12), 3.82 (3H, s, -OCH 3 ), 3.85 (3H, s A white amorphous powder 51 (1H, s, H-6), 6.86 (1H, s, H-7), 6.97 (1H, s, H-10), 2.43 (1H, s, H-10b), 2.45 (2H, m, H-11), 3.17 (1H, s, H-12), 3.82 (3H, s, -OCH 3 ), 3.84 (3H, s, -OCH 3 ), 2.43 (2H, q, J = 9.05, 1.45 Hz, -OCH 2 CH 3 ), 2.31 (3H, t, J = 2.50 Hz, -OCH 2 CH 3 ), 2..08 (3H, s, -NCH 3 ); 13 C NMR (100 MHz General methods for alkaloid synthesis. Total synthesis of racemic lycoramine Total synthesis of dl-lycoramine The total synthesis of (±)-lycoramine. Part II Total synthesis of (+)-lycoramine. I Phenol oxidation and biosynthesis.V. Synthesis of galanthamine A short stereospecific synthesis of (dl)-lycoramine. Control of relative stereochemistry by dipole effects Oxidative intramolecular phenolic coupling reaction induced by a hypervalent iodine (III) reagent: Leading to galanthamine-type Amaryllidaceae alkaloids Total synthesis of racemic lycoramine Identification of natural compounds with antiviral activities against SARS-associated coronavirus Benzylphenethylamine Alkaloids from Hosta plantaginea with inhibitory activity against tobacco mosaic virus and acetylcholinesterase Structure elucidation and biomimetic synthesis of hostasinine A, a new benzylphenethylamine alkaloid from Hosta plantaginea Flora Reipublicae Popularis Sinicae New alkaloids from Pancratium maritimum L Alkaloids from Narcissus cv Extracts of Lycoris aurea induce apoptosis in murine sarcoma S180 cells Revised NMR data for incartine: an alkaloid from Galanthus elwesii Alkaloid N-oxides of Amaryllidaceae Narcissus alkaloids. Part III. 9-O-demethylhomolycorine from Narcissus confusus 2D NMR studies of lycorenine as a model for the structural assignment of lycorenine-type alkaloids Alkaloids from Narcissus munozii-garmendiae Capillary gas chromatography-mass spectrometry of Amaryllidaceae alkaloids A review on 2009 influenza A virus O-ethyllycorenine (10)。结论：化合物 1 为新的高石蒜碱类型的石蒜生物碱, 生物碱 1−3 为该植物中的主要成分, 且对流感甲型 病毒显示了较弱的抗病毒活性 We thank Prof. GONG Xun for the identification of the plant material.