Cytotoxic Bryostatin Derivatives from the South China Sea Bryozoan

May 1, 2015 - (5−7), have been isolated from an extract of the South China. Sea bryozoan ... in the South China Sea, led to the isolation of bryosta...
0 downloads 0 Views 714KB Size
Download PDF
Note pubs.acs.org/jnp

Cytotoxic Bryostatin Derivatives from the South China Sea Bryozoan Bugula neritina Hao-Bing Yu,†,‡,§ Fan Yang,‡,§ Yan-Yun Li,‡,§ Jian-Hong Gan,† Wei-Hua Jiao,‡ and Hou-Wen Lin*,†,‡ †

Laboratory of Marine Drugs, Department of Pharmacy, Changzheng Hospital, Second Military Medical University, 415 Fengyang Road, Shanghai 200003, People’s Republic of China ‡ Marine Drugs Research Center, Department of Pharmacy, State Key Laboratory of Oncogenes and Related Genes, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, 160 Pujian Road, Shanghai 200127, People’s Republic of China S Supporting Information *

ABSTRACT: Four new macrocyclic lactones, bryostatin 21 (1) and 9-O-methylbryostatins 4, 16, and 17 (2−4), together with three known related compounds, bryostatins 4, 16, and 17 (5−7), have been isolated from an extract of the South China Sea bryozoan Bugula neritina. The structures of all compounds were unambiguously elucidated using detailed spectroscopic analysis. Structurally, the presence of a single methyl group at C-18 in compound 1 has not been observed before for known bryostatins. The isolated macrolides exhibited inhibitory effects against a small panel of human cancer cell lines.

T

Herein, we report the details of the purification and structure elucidation of these compounds, as well as the evaluation of cytotoxic activities against a variety of different human cancer cell lines.

he marine bryozoan Bugula neritina is a colonial suspension feeder widely distributed in the temperate marine environment. It is the source of bioactive metabolites, including bryostatins,1 alkaloids,2 fatty aldehydes,3 ceramides,4 cerebrosides,4 and sterols.5 Bryostatins, possessing a 26membered macrocyclic lactone with three imbedded highly functionalized pyran rings,6 have attracted much interest. During a search for cytotoxic metabolites from B. neritina initiated in 1968, potent cytotoxic activity of the extracts against lymphocytic leukemia cell lines attracted considerable interest.6 Many years of research led to the isolation and structure elucidation of the first example of this family, bryostatin 1, which was reported by Pettit and co-workers in 1982.6 To date, only 20 natural bryostatins have been obtained,7 and they exhibit a remarkable range of biological activities, including cognition and memory enhancement, cytotoxic, and synergistic chemotherapeutic activitives.1 Among the 20 known macrolides, bryostatin 1, as a unique PKC modulator, has been studied most extensively and has undergone phase I and phase II clinical trials.8 Due to their excellent bioactive properties but scarcity in nature, 9 the biological activities, structure modification, and structure−activity relationships of the bryostatins have attracted considerable interest over the past two decades.1 Our previous chemical investigation of B. neritina, collected in the South China Sea, led to the isolation of bryostatin 19.10 Recently, we demonstrated that bryostatin 5 can induce apoptosis by activating PUMA and caspases in acute monocytic leukemia cells.11 As a continuation of our studies on bryostatins, we carried out further chromatographic purification on the extracts of B. neritina, identifying four new bryostatin macrolides, 1−4, along with bryostatins 4, 16, and 17 (5−7). © XXXX American Chemical Society and American Society of Pharmacognosy

Bryostatin 21 (1) was isolated as a colorless, amorphous powder. The molecular formula was determined as C45H68O17 by an HRESIMS adduct ion peak at m/z 903.4347 ([M + Na]+), implying the presence of 12 degrees of unsaturation. The UV spectrum of 1 exhibited an absorption at 225 nm. The IR spectrum indicated the presence of hydroxy (3459 cm−1) and carbonyl (1723 cm−1) groups. These spectroscopic characteristics and the initial inspections of the 1H and 13C NMR spectra indicated that 1 is a member of the bryostatin class.12 The COSY and key HMBC correlations confirmed this hypothesis (Figure 1). The 1H NMR and 13C NMR spectra of compound 1 (Table 1) were similar to those of bryostatin 4 Received: January 27, 2015

A

DOI: 10.1021/acs.jnatprod.5b00081 J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products

Note

They were readily purified by HPLC and rapidly interconverted into a mixture of the Δ21,34 E and Δ21,34 Z forms at room temperature (rt) with an equilibrium ratio of 3:2 (Figure S31), which is similar to the phenomenon observed for bryostatins 16 (6) and 17 (7).13 The molecular formula of the mixture was established as C43H64O14 based on the HRESIMS data (m/z 827.4189 [M + Na]+), indicating the presence of 11 degrees of unsaturation for both compounds. In the 1H NMR spectrum belonging to compound 3, the presence of one isolated methoxy proton resonance at 9-OCH3 (δH 3.19) indicated that compound 3 was an O-methyl product of bryostatin 16.13 The key HMBC correlation of 9-OCH3 with C-9 (δC 103.3) established the presence of the methoxy group at C-9. Moreover, the structure of compound 4 was unambiguously assigned based on the HMBC correlation from 9-OCH3 (δH 3.19) to C-9 (δC 103.2). A comprehensive set of NOESY correlations observed in 3 and 4 indicated that the relative configurations in 3 and 4 were the same as for bryostatins 16 and 17, respectively (Figure S27). Because MeOH was used for the extractions, it is probable that compounds 3 and 4 are artifacts. In addition to the four new compounds 1−4, three known compounds, bryostatin 4 (5), bryostatin 16 (6), and bryostatin 17 (7), were also obtained and elucidated by comparing their physical and spectroscopic features with the data reported in the literature.12,13 The cytotoxicities of compounds 1, 2, and 5, as well as the mixtures of 3 and 4 (ratio = 3:2) and 6 and 7 (ratio = 3:2) were evaluated against four human cancer cell lines: U937 (human histiocytic lymphoma), K562 (human chronic leukemia), SGC7901 (human gastric carcinoma), and HeLa (human cervical adenocarcinoma). Doxorubicin was used as a positive control. The isolated compounds showed cytotoxic effects against all of the human cancer cell lines tested (Table 2). In the previous studies, all bryostatins were mainly assessed for their cytotoxicities against the murine P388 lymphocytic leukemia cell line.6,12,13 Compared to the reported potent cytotoxicity against the P388 cell line, compounds 1−7 exhibited weaker cytotoxicities against the above four cancer cell lines. These results indicated that the P388 cell line may be more sensitive to the bryostatins. The biological evaluation indicated that Omethylation at C-9 correlated with a loss of inhibitory activity, as was observed for the mixtures of 3 and 4, and 6 and 7. In addition, the presence of a single methyl group at C-18, as demonstrated by compound 1, positively affected cytotoxic activity, as determined via comparison with compound 5. Moreover, the inhibitory effects of the bryostatins were more profound in leukemia cell lines than in solid tumor cell lines.

Figure 1. COSY, key HMBC, and selected NOESY correlations of 1.

(5). Instead of one quaternary carbon at δC 44.9 (C-18) in 5, one sp3 methine (δH/δC 2.66/39.7, CH-18) in 1 was detected. Taken together with a doublet methyl signal at δH 0.90 (d, J = 6.6 Hz, H3-32), the significant difference was the presence of a single methyl group at C-18 in compound 1. These results were confirmed by the COSY correlations of H2-14/H-15/H-16/H17/H-18/H3-32, as well as the HMBC correlations from H-18 (δH 2.66) to C-16 (δC 132.4) and C-17 (δC 132.5) and from H3-32 (δH 0.90) to C-17, C-18 (δC 39.7), and C-19 (δC 99.5) (Figure 1). Although an HMBC correlation between H-25 (δH 5.21) and C-1 was not observed, the deshielded resonance of C-25 (δC 73.5) suggested the attachment between C-1 and C25 via an oxygen atom. Finally, the planar structure of 1 was established as shown and named bryostatin 21. Structurally, the presence of a single methyl group at C-18 in compound 1 has not been observed previously for all of the known bryostatins. The relative configuration of 1 was established by coupling constant analysis and NOESY experiments. The observed similarity in the NMR chemical shift values and coupling constants indicated that the relative configurations in 1 were the same as for other bryostatins.7 The conjugated C-16/C-17 double bond was assigned to be trans due to its large coupling constant (16.2 Hz). The trans configuration for the C-21/C-33 double bond was deduced based on the strong NOESY correlation between H-20 and H-33, as well as the lack of NOESY correlation between H-33 and H-22. NOESY correlations of H-5/H-7, H-7/H3-29, H3-29/H-11, H-11/H15, H-15/H-17, H-17/H-18, and 3-OH/26-OH indicated that these protons were α-oriented, while the NOESY cross-peaks of H3-32/19-OH, H3-32/H-20, H-20/19-OH, H-23/H-26, and H-23/H-3 indicated that they were cofacial and β-oriented (Figure 1). Therefore, the relative configuration of 1 was assigned as 3R*, 5R*, 7S*, 9S*, 11S*, 15R*, 18S*, 19S*, 20S*, 23S*, 25R*, 26R*. The molecular formula of 9-O-methylbryostatin 4 (2) was determined as C47H72O17 according to its HRESIMS, which corresponded to 12 degrees of unsaturation. The 1H and 13C NMR spectra of 2 were similar to those of bryostatin 4 (5),12 except for the presence of a methoxy group (δH/δC 3.36/52.0). The extra methoxy group was located at C-9, as determined by the HMBC correlations from 9-OCH3 (δH 3.36) to C-9 (δC 104.4). The unambiguous COSY and key HMBC correlations confirmed the postulated methoxy homologue of 5 (Figure S32). On the basis of the 1H−1H coupling constant values and NOESY data, the relative configuration of 2 was assigned as 3R*, 5R*, 7S*, 9S*, 11S*, 15R*, 19S*, 20S*, 23S*, 25R*, 26R*. Compound 2, however, could possibly be an artifact from the extraction process with MeOH as a solvent. 9-O-Methylbryostatin 16 (3) and 9-O-methylbryostatins 17 (4) were isolated as a colorless, amorphous powder mixture.

■

EXPERIMENTAL SECTION

General Experimental Procedures. Optical rotation data were recorded using a PerkinElmer model 341 polarimeter with a 1 dm cell. UV and IR (KBr) spectra were obtained using a Hitachi U-3010 spectrophotometer and Jasco FTIR-400 spectrometer, respectively. 1 H, 13C, DEPT135, COSY, HSQC, HMBC, and NOESY NMR spectra were acquired at rt using a Bruker AVANCE-600 instrument. HRESIMS and ESIMS data were obtained using a Waters Q-Tof micro YA019 mass spectrometer. Reversed-phase HPLC was performed on YMC-Pack Pro C18 RS (5 μm) columns using a Waters 1525 separation module equipped with a Waters 2998 photodiode array (PDA) detector. Column chromatographic purifications were performed using silica gel 60 (200−300 mesh; Yantai, China), Sephadex LH-20 (18−110 μm, Pharmacia Co.), and ODS (50 μm, YMC Co.). Analytical thin-layer chromatography (TLC) was carried B

DOI: 10.1021/acs.jnatprod.5b00081 J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products

Note

Table 1. 1H (600 MHz) and 13C (150 MHz) NMR Spectroscopic Data of 1−4 in CDCl3 1 position

δC, type

1 2a 2b 3 3-OH 4a 4b 5 6a 6b 7

172.4, C 42.1, CH2

8 9 10a 10b 11 12a 12b 13 14a 14b 15 16

41.3, C 101.8, C 42.1, CH2

17 18 19 19-OH 20 21 22a 22b 23 24a 24b 25 26 26-OH 27 28 29 30 31 32 33 34 35 36 37 9-OCH3 1′ 2′ 3′ 4′ 5′ 1″ 2″ 3″ 4″

68.3, CH 39.8, CH2 65.6, CH 33.1, CH2 72.5, CH

71.3, CH 44.1, CH2 156.5, C 36.4, CH2 78.7, CH 132.4, CH 132.5, CH 39.7, CH 99.5, C 73.2, CH 150.9, C 31.6, CH2 64.6, CH 35.6, CH2 73.5, CH 70.2, CH 19.6, 17.0, 21.0, 114.3, 166.7, 10.9, 120.5, 166.7, 51.1, 51.0,

CH3 CH3 CH3 CH C CH3 CH C CH3 CH3

178.0, 39.0, 27.1, 27.1, 27.1, 172.4, 36.4, 18.5, 13.6,

C C CH3 CH3 CH3 C CH2 CH2 CH3

2 δH, (J in Hz)

2.47, 2.53, 4.14, 4.24, 1.58, 1.94, 4.20, 1.44, 1.72, 5.11,

1.70, 2.12, 3.96, 2.22, 2.11,

dd (12.0, 2.4) d (12.0) m d (12.6) bt (14.4, 3.6) t (12.6) t (17.4) q (12.0) m dd (4.8, 11.4)

m m t (8.4) d (8.4) d (8.4)

1.94, d (10.2) 3.68, d (10.2) 4.16, m 5.41, ddd (1.2, 7.2, 15.6) 5.92, dd (4.8, 16.2) 2.66, m 5.46, s 4.99, s 2.02, 3.70, 4.05, 1.84, 1.99, 5.21, 3.78, 3.20, 1.23, 1.05, 0.95, 5.69,

m m m m m m m br s d (6.6) s s s

0.90, d (6.6) 6.05, s 3.72, s 3.69, s

1.20, s 1.20, s 1.20, s 2.34, m 1.65, m 0.95, t (7.2)

δC, type 172.0, C 42.5, CH2 68.4, CH 41.0, CH2 66.3, CH 33.8, CH2 71.7, CH 42.4, C 104.4, C 39.7, CH2 73.2, CH 43.0, CH2 156.5, C 36.4, CH2 79.6, CH 128.9, CH 139.3, CH 44.8, C 98.9, C 74.1, CH 151.9, C 31.2, CH2 64.7, CH 35.7, CH2 73.9, CH 70.1, CH 19.5, 17.6, 20.6, 114.4, 166.7, 19.6, 24.5, 119.6, 166.9, 51.1, 51.0, 52.0, 177.9, 38.9, 27.1, 27.1, 27.1, 171.9, 36.5, 18.2, 13.6,

CH3 CH3 CH3 CH C CH3 CH3 CH C CH3 CH3 CH3 C C CH3 CH3 CH3 C CH2 CH2 CH3

3 δH, (J in Hz)

171.7, C 42.7, CH2

2.54, m 4.16, 4.20, 1.68, 2.00, 4.06, 1.35, 1.83, 5.14,

1.75, 2.17, 4.02, 2.17,

δC, type

m d (7.2) m m m q (12.0) m m

65.2 CH 41.3, CH2 64.9, CH 32.1, CH2 73.3, CH 42.0, C 103.3, C 39.9, CH2

m m m m

72.4, CH 43.1, CH2

2.00, m 3.63, d (13.8) 3.99, m 5.34, dd (9.0, 16.2) 5.78, d (16.2)

5.37, s 5.15, s

157.7, C 36.2, CH2 77.4, CH 127.6, CH 137.5, CH 41.0, C 150.5, C 100.7, CH 167.8, C 31.7, CH2

2.01, 3.66, 4.01, 1.85, 1.94, 5.16, 3.82,

m m m m m m m

1.23, 1.01, 0.89, 5.68,

d (6.6) s s s

72.9, CH 35.8, CH2 73.2, CH 69.3, CH

1.01, s 1.56, s 6.00, s 3.70, s 3.68, s 3.36, s

1.18, s 1.18, s 1.18, s

19.4, 17.2, 20.3, 114.4, 166.8, 25.9, 24.5, 107.6, 169.5, 51.0, 50.7, 49.0, 178.0, 38.9, 27.1, 27.1, 27.1,

CH3 CH3 CH3 CH C CH3 CH3 CH C CH3 CH3 CH3 C C CH3 CH3 CH3

4 δH, (J in Hz)

2.41, m 2.47, m 3.94, m 1.56, m 1.63, m 4.35, m 1.45, m 1.5, m 5.16, dd (4.8, 11.4)

1.70, 2.17, 3.82, 2.16, 2.24,

m m m m m

1.86, d (13.2) 3.76, d (13.8) 3.93, m 5.47, dd (6.6, 16.2) 5.83, d (16.2)

5.41, s 2.34, 3.63, 3.88, 1.93, 3.63, 5.22, 3.84,

m m m m m m m

1.22, 0.99, 0.87, 5.69,

d (6.6) s s s

1.19, s 1.22, s 5.43, s 3.71, s 3.65, s 3.19, s

1.17, s 1.17, s 1.17, s

δC, type 171.7, C 42.8, CH2 64.8 CH 41.3, CH2 64.5, CH 32.2, CH2 73.3, CH 41.9, C 103.2, C 40.0, CH2 72.4, CH 42.8, CH2 157.4, C 36.5, CH2 77.6, CH 127.9, CH 137.2, CH 41.2, C 148.6, C 96.7, CH 167.4, C 36.7, CH2 72.6, CH 35.7, CH2 73.2, CH 69.2, CH 19.5, 17.3, 20.3, 114.2, 166.9, 26.7, 24.2, 107.3, 170.1, 51.1, 50.8, 48.9, 177.9, 38.9, 27.1, 27.1, 27.1,

CH3 CH3 CH3 CH C CH3 CH3 CH C CH3 CH3 CH3 C C CH3 CH3 CH3

δH, (J in Hz) 2.39, m 2.49, m 4.38, m 1.55, m 1.63, m 3.88, m 1.44, m 1.74, m 5.16, dd (4.8, 11.4)

1.80, 2.15, 3.82, 2.13, 2.24,

m m m m m

1.84, d (13.2) 3.78, d (13.8) 3.95, m 5.46, dd (6.6, 16.2) 5.84, d (15.6)

6.84, m 2.22, 2.50, 3.93, 1.92, 2.00, 5.22, 3.91,

m m m m m m m

1.21, 1.01, 0.86, 5.69,

d (6.6) s s s

1.20, s 1.29, s 5.20, s 3.69, s 3.68, s 3.19, s

1.18, s 1.18, s 1.18, s

2.30, m 1.65, m 0.94, t (7.2) C

DOI: 10.1021/acs.jnatprod.5b00081 J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products

Note

(KBr) νmax 3362, 3208, 2923, 2853, 1724, 1658, 1635, 1608, 1466, 1431, 1378, 1262, 1156, 1100, 804, 739 cm−1; 1H and 13C NMR data, Table 1; HRESIMS m/z 827.4189 [M + Na]+ (calcd for C43H64O14, 827.4194). Bryostatin 4 (5): colorless, amorphous powder; [α]25D +32.9 (c 0.10, MeOH); lit. [α]27D +93.6 (c 0.32, MeOH).12 Mixture of bryostatin 16 (6) and bryostatin 17 (7): colorless, amorphous powder; [α]25D +43.9 (c 0.10, MeOH); lit. [α]D +84.0 (c 0.43, MeOH) for bryostatin 16 and [α]D +231.0 (c 0.34, MeOH) for bryostatin 17.13 Cytotoxicity Assay. The cytotoxicity of compounds 1−7 against U937, K562, SGC-7901, and HeLa human cancer cell lines was determined using the MTT method.14 The cells were cultured in RPMI-1640 or DMEM medium (Hyclone) supplemented with 10% fetal bovine serum (Hyclone) at 37 °C in a humidified atmosphere with 5% CO2. Briefly, 100 μL of adherent cells was seeded into each well of a 96-well microplate and allowed to adhere for 12 h before drug addition. Suspended cells were seeded at an initial density of 1 × 105 cells/mL before drug addition. Each cancer cell line was treated with the indicated test compound at various concentrations, in triplicate, for 72 h, and doxorubicin was used as a positive control. DMSO (100 μL for each well) was used to solubilize the cells, and the optical density (OD) of the lysate was measured at 490 nm in a 96-well microtiter plate reader (Spectra MAX 340). IC50 values were obtained from the curves of average OD values of the triplicate tests versus drug concentrations.

Table 2. Cytotoxicities of Compounds 1−7 to Human Cancer Cell Lines IC50 (at 72 h, μM)

a

compound

U937

K562

SGC-7901

HeLa

1 2 3 and 4 5 6 and 7 doxorubicin

7.7 NA 13.0 NA 1.1 0.1

9.9 NA 4.9 NA 2.1 0.2

NAa NA NA NA 11.0 0.2

NA NA NA NA 11.0 0.1

NA = not active at a concentration of 20 μg/mL.

out using HSGF 254 plates and visualized by spraying with anisic aldehyde reagent. Animal Material. The bryozoan samples were collected off Daya Bay in the South China Sea in June 2012 and identified as Bugula neritina by Prof. Ru-xing Cai (Biology Department, Hangzhou University, China) and Prof. Yang-hua Yi (Research Center for Marine Drugs, School of Pharmacy, Second Military Medical University, China). A voucher sample (No. CTC1201-DY) has been deposited in the Laboratory of Marine Drugs, Department of Pharmacy, Changzheng Hospital, Second Military Medical University, China. Extraction and Isolation. Air-dried bryozoan (10.0 kg, dry weight) was powdered and extracted with 95% aqueous EtOH at rt, with each extraction taking 1 week. The combined extracts were concentrated under reduced pressure to yield the crude extract (102.1 g), which was suspended in H2O and sequentially extracted with EtOAc and n-BuOH to afford EtOAc- and n-BuOH-soluble extracts. The EtOAc-soluble extract (97.8 g) was partitioned between 90% aqueous MeOH and n-hexane, and the n-hexane layer was collected and concentrated under reduced pressure to afford the n-hexanesoluble extract (72.0 g). The 90% aqueous MeOH phase was diluted to 60% aqueous MeOH with H2O, which was extracted with CH2Cl2 to afford the CH2Cl2-soluble extract (18.7 g). The CH2Cl2-soluble extract was subjected to vacuum liquid chromatography (VLC) on silica gel by gradient elution using n-hexane/acetone (50:1, 30:1, 20:1, 10:1, 8:1, 5:1, 1:1, 0:1, v/v) as the solvent to give seven fractions (A− G). Fraction E (1.1 g) was subjected to column chromatography (CC) on Sephadex LH-20 using CH2Cl2/MeOH (1:1) as the eluting solvent to afford four subfractions (E1−E4). Subfraction E2 (0.1 g) was also separated by reversed-phase HPLC eluting with 87% MeOH/H2O to afford compound 2 (2.0 mL/min, 228 nm, tR = 38.9 min, 1.4 mg). Subfraction E3 (0.8 g) was further separated on an ODS (50 μm) column, followed by stepwise gradient elution with MeOH/H2O (1:1, 3:2, 4:1, 1:0) to afford seven fractions (Fr. E31−E37). Fraction E36 was also purified by reversed-phase HPLC with an elution of 87% MeOH/H2O detected at a wavelength of 228 nm to give compounds 7 (2.0 mL/min, tR = 22.8 min, 5.9 mg), 4 (2.0 mL/min, tR = 34.2 min, 2.1 mg), 1 (2.0 mL/min, tR = 36.5 min, 3.5 mg), 5 (2.0 mL/min, tR = 38.0 min, 9.7 mg), 6 (2.0 mL/min, tR = 39.4 min, 8.5 mg), and 3 (2.0 mL/min, tR = 64.6 min, 2.0 mg). Bryostatin 21 (1): colorless, amorphous powder; [α]25D +7.3 (c 0.10, MeOH); UV (MeOH) (log ε) λmax 225 (4.46); IR (KBr) νmax 3459, 3365, 2926, 2855, 1723, 1662, 1463, 1437, 1408, 1376, 1286, 1255, 1236, 1159, 1084, 990.863, 805, 706 cm−1; 1H and 13C NMR data, Table 1; HRESIMS m/z 903.4347 [M + Na]+ (calcd for C45H68O17Na, 903.4354). 9-O-methylbryostatin 4 (2): colorless, amorphous powder; [α]25D +24.4 (c 0.10, MeOH); UV (MeOH) (log ε) λmax 228 (4.69) nm; IR (KBr) νmax 3467, 3330, 2970, 2933, 1723, 1658, 1461, 1436, 1380, 1285, 1160, 1101, 988, 861, 737 cm−1; 1H and 13C NMR data, Table 1; HRESIMS m/z 931.4672 [M + Na]+ (calcd for C47H72O17Na, 931.4667). Mixture of 9-O-methylbryostatin 16 (3) and 9-O-methylbryostatin 17 (4): colorless, amorphous powder; [α]25D +33.6 (c 0.10, MeOH); UV (MeOH) (log ε) λmax 225 (4.09), 304 (4.06) nm; IR

■

ASSOCIATED CONTENT

* Supporting Information S

Copies of 1D and 2D NMR, HRESIMS, UV, and IR spectra for compounds 1−4 are available. The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jnatprod.5b00081.

■

AUTHOR INFORMATION

Corresponding Author

*Tel: +86-21-68383346. Fax: +86-21-58732594. E-mail: [email protected] (H. W. Lin). Author Contributions §

H.-B. Yu, F. Yang, and Y.-Y. Li contributed equally to this paper. Notes

The authors declare no competing financial interest.

■

ACKNOWLEDGMENTS This research was supported by the National Natural Science Fund for Distinguished Young Scholars of China (81225023), the National High Technology Research and Development Program of China (863 Projects, Nos. 2013AA092902 and 2011AA09070107), and the National Science & Technology Major Project (2013ZX09103002-017). We are also grateful for the financial support of the National Natural Science Fund of China (Nos. 41476121, 81302691, 81172978, and 41106127), the Innovation Program of Shanghai Municipal Education Commission (No. 14YZ037), Shanghai Rising-Star Program (14QA1402800), and Science & Technology Support Project of Shanghai Science and Technology Commission (12431900804).

■

REFERENCES

(1) Ruan, B. F.; Zhu, H. L. Curr. Med. Chem. 2012, 19, 2652−2664. (2) Anthoni, U.; Larsen, C.; Nielsen, P. H.; Christophersen, C.; Lidgren, G. Comp. Biochem. Physiol. B 1989, 92, 711−713. (3) Berdyshev, E.; Getmanova, O.; Svetashev, V.; Kubanin, A. Comp. Biochem. Physiol. B 1992, 102, 639−641.

D

DOI: 10.1021/acs.jnatprod.5b00081 J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products

Note

(4) Tian, X. R.; Tang, H. F.; Li, Y. S.; Lin, H. W.; Ma, N.; Zhang, W.; Yao, M. N. J. Asian Nat. Prod. Res. 2009, 11, 1005−1012. (5) Tian, X. R.; Tang, H. F.; Li, Y. S.; Lin, H. W.; Fan, X. P.; Feng, J. T.; Zhang, X. Phytochem. Lett. 2014, 9, 1−6. (6) Pettit, G. R.; Herald, C. L.; Doubek, D. L.; Herald, D. L.; Arnold, E.; Clardy, J. J. Am. Chem. Soc. 1982, 104, 6846−6848. (7) Lopanik, N.; Gustafson, K. R.; Lindquist, N. J. Nat. Prod. 2004, 67, 1412−1414. (8) Kollár, P.; Rajchard, J.; Balounová, Z.; Pazourek, J. Pharm. Biol. 2013, 52, 237−242. (9) Schaufelberger, D. E.; Koleck, M. P.; Beutler, J. A.; Vatakis, A. M.; Alvarado, A. B.; Andrews, P.; Marzo, L. V.; Muschik, G. M.; Roach, J.; Ross, J. T.; Lebherz, W. B.; Reeves, M. P.; Eberwein, R. M.; Rodgers, L. L.; Testerman, R. P.; Snader, K. M.; Forenza, S. J. Nat. Prod. 1991, 54, 1265−1270. (10) Lin, H. W.; Yi, Y. H.; Li, W. L.; Yao, X. S.; Wu, H. M. Chin. J. Mar. Drugs 1998, 17, 1−3. (11) Wang, Y. W.; Zhang, J. B.; Wang, Q. X.; Zhang, T.; Yang, Y.; Yi, Y. H.; Gao, G. X.; Dong, H. J.; Zhu, H. F.; Li, Y.; Lin, H. W.; Tang, H. F.; Chen, X. Q. Eur. J. Pharmacol. 2013, 718, 340−349. (12) Pettit, G. R.; Kamano, Y.; Herald, C. L.; Tozawa, M. J. Am. Chem. Soc. 1984, 106, 6768−6771. (13) Pettit, G. R.; Gao, F.; Blumberg, P. M.; Herald, C. L.; Coll, J. C.; Kamano, Y.; Lewin, N. E.; Schmidt, J. M.; Chapuis, J. C. J. Nat. Prod. 1996, 59, 286−289. (14) Jiao, W. H.; Xu, T. T.; Yu, H. B.; Chen, G. D.; Huang, X. J.; Yang, F.; Li, Y. S.; Han, B. N.; Liu, X. Y.; Lin, H. W. J. Nat. Prod. 2014, 77, 346−350.

E

DOI: 10.1021/acs.jnatprod.5b00081 J. Nat. Prod. XXXX, XXX, XXX−XXX