Antimalarial and Antiproliferative Cassane Diterpenes of Caesalpinia

Article pubs.acs.org/jnp

Antimalarial and Antiproliferative Cassane Diterpenes of Caesalpinia sappan Guoxu Ma,†,⊥ Haifeng Wu,†,⊥ Deli Chen,§ Nailiang Zhu,† Yindi Zhu,† Zhonghao Sun,† Pengfei Li,† Junshan Yang,† Jingquan Yuan,*,‡ and Xudong Xu*,† †

Key Laboratory of Bioactive Substances and Resource Utilization of Chinese Herbal Medicine, Ministry of Education, Institute of Medicinal Plant Development, Peking Union Medical College and Chinese Academy of Medical Sciences, Beijing 100193, People’s Republic of China ‡ National Engineering Laboratory of Southwest Endangered Medicinal Resource Development, National Development and Reform Commission, Guangxi Botanical Garden of Medicinal Plants, Nanning 530023, People’s Republic of China § Hainan Branch Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Wanning 571533, People’s Republic of China S Supporting Information *

ABSTRACT: Bioassay-guided fractionation of a methanol extract of the seeds of Caesalpinia sappan led to the isolation of 12 new cassane-type diterpenes, caesalsappanins A−L (1−12). Their structures were elucidated on the basis of NMR and HRESIMS analysis, and the absolute configuration of compound 1 was determined by single-crystal X-ray crystallography. All isolated compounds were tested against a chloroquine-resistant Plasmodium falciparum strain for antiplasmodial activities and against a small panel of human cancer cell lines for antiproliferative activities. Compounds 7 and 8 displayed antimalarial activity against the chloroquine-resistant K1 strain of P. falciparum with IC50 values of 0.78 and 0.52 μM and selectivity indices of 17.6 and 16.4, respectively. Compound 10 showed antiproliferative activity against the KB cancer cell line with an IC50 value of 7.4 μM.

M

inhibit in vitro the multidrug-resistant K1 strain of P. falciparum, with IC50 value of 0.38 μg/mL. Purification of this bioactive fraction resulted in the isolation of 12 new cassane diterpenes, caesalsappanins A−L (1−12). The structure and absolute configuration of compound 1 were determined by single-crystal X-ray crystallography. Herein are reported the isolation and structure elucidation of the new isolates as well as the evaluation of their bioactivities against the P. falciparum strain K1 and a small panel of human cancer cell lines.

alaria is one of the most life-threatening infectious diseases worldwide, claiming the lives of millions of persons each year.1−3 Although antimalarial medication continues to play an important role in the control and elimination of the disease, the appearance of drug-resistant Plasmodium falciparum has made the treatment of malaria increasingly problematic.4,5 Thus, it is urgent to search for new alternatives to currently available drugs. Natural products remain important potential sources of new and selective agents for the treatment of malaria. The genus Caesalpinia (Leguminosae) is distributed widely in the tropical and subtropical regions of Southeast Asia, consisting of more than 70 species.6−8 Caesalpinia sappan L. is a shrubby tree, with its heartwood being used as an antibacterial and anti-inflammatory agent in folk medicine.9,10 Previous phytochemical investigations on this plant have disclosed the presence of cassane-type diterpenoids, flavonoids, lignans, steroids, and triterpenoids.11,12 Among these components, cassane diterpenes have shown potent cytotoxicity and antimalarial activities.13,14 The highly variable tricyclic skeletons of cassane diterpenes, along with their varied biological properties, have made this family of diterpenoids especially attractive targets for lead compound discovery.15−17 In a continuing search for biologically active natural products from the genus Caesalpinia,18,19 the CHCl3-soluble fraction from the methanol extract of C. sappan seeds was found to © XXXX American Chemical Society and American Society of Pharmacognosy

■

RESULTS AND DISCUSSION The air-dried and powdered seeds of C. sappan were extracted three times with MeOH. The resulting extract was separated via column chromatography (CC) and semipreparative HPLC purification, affording 12 new cassane-type diterpenes (1−12). Caesalsappanin A (1) was obtained as colorless crystals with [α]25 D −64.6. The HRESIMS showed a quasimolecular ion at m/ z 443.2045 [M + Na]+ (calcd for C23H32NaO7, 443.2046). Thus, in conjunction with 13C NMR data, the molecular formula was established as C23H32O7, representing eight indices of hydrogen deficiency. The UV and IR spectra showed absorptions for a hydroxy group (3500 cm−1) and an α,βReceived: April 10, 2015

A

DOI: 10.1021/acs.jnatprod.5b00317 J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products

Article

Hz) and 3.77 (d, J = 12.0 Hz). Except for the methoxy (δC 51.7) and ethoxy (δC 59.2, 15.1) substituents, the 13C APT NMR spectrum displayed 20 resonances due to one methyl (δC 11.2), seven methylenes (δC 21.0, 24.0, 29.2, 35.8, 37.4, 37.5, and 61.5), six methines (δC 37.6, 41.3, 41.8, 45.1, 97.0, and 115.4), and six quaternary carbons (δC 38.6, 45.7, 108.2, 170.7, 172.6, and 175.7). All the proton signals were assigned to the corresponding carbons through direct 1H and 13C correlations in the HSQC spectrum. The overall 1H and 13C NMR spectroscopic data confirmed that 1 is an oxygenated diterpenoid possessing a fused butenolide unit.22,23 Partial structures were deduced from the 1H−1H COSY spectrum, and the whole structure was connected on the basis of long-range correlations observed in the HMBC spectrum (Figure 1). The methine protons at δH 4.82 (d, J = 2.4 Hz, H-20) showed longrange correlations with the carbons at δC 37.4 (C-1) and 41.3 (C-9), indicating that C-1, C-9, and C-20 are connected through the quaternary carbon C-10. Moreover, the connectivity of C-3 (δC 35.8), C-5 (δC 45.1), C-18 (δC 175.7), and C-19 (δC 61.5) with the quaternary carbon C-4 (δC 45.7) was established on the basis of HMBC correlations from the methylene protons (H2-19) to the carbons C-3, C-4, C-5, and C-18. Likewise, the methylene protons (H2-11) and methine

unsaturated butenolide moiety (214 nm; 1749 cm−1).20,21 The olefinic proton signal at δH 5.76 (H-15, s) and downfield carbon signals at δC 108.2 (C-12), 115.4 (C-15), 170.7 (C-16), and 172.6 (C-13) in the 1H and 13C APT NMR spectra (Tables 1 and 2) also confirmed the presence of the α,β-unsaturated butenolide ring. Additionally, the 1H NMR spectrum exhibited signals for a methyl group at δH 1.07 (d, J = 7.2 Hz, H3-17), a methoxy group at δH 3.65 (s), an ethoxy group at δH 3.24, 3.52 (qd, J = 7.2, 1.8 Hz, OCH2CH3) and 1.15 (d, J = 7.2 Hz, OCH2CH3), an oxygenated methylene at δH 4.82 (d, J = 2.4 Hz), and an oxygenated methyl at δH 4.31 (dd, J = 12.0, 2.4

Table 1. 1H NMR Spectroscopic Data (600 MHz) for Compounds 1−6 (δH in ppm, J in Hz) 1a

position 1α 1β 2α 2β 3α 3β 5 6α 6β 7 8 9 11α 11β 14 15 17 19α 19β 20 OCH3-12 OCH2CH3-12 OCH2CH3-12 OCH3-18 OCH3-19 OCH2CH3-19 OCH2CH3-19 OCH2CH3-20 OCH2CH3-20 OCH3-20 a

1.56, 2.03, 1.57, 2.30, 1.89, 2.04, 1.61, 1.24, 2.09, 1.36, 1.54, 2.23, 1.51, 1.28, 2.56, 2.87, 5.76, 1.07, 3.77, 4.31, 4.82,

m m m m m m m m m m m m m m d (12.6, 2.4) qd (7.2, 2.4) s d (7.2) d (12.0) dd (12.0, 2.4) d (2.4)

3.24, 3.52, 1.15, 3.65,

qd (7.2, 1.8) qd (7.2, 1.8) t (7.2) s

2a

3a

4a

0.97, m 2.52, m 1.48, m 2.22, m 1.61 m 1.92, m 1.74, m 1.99, m 1.76, m 1.38, m 1.50, m 1.58, m 1.47, m 1.56, m 2.70, dd (12.6, 2.4) 2.92, qd (7.2, 2.4) 5.74, s 1.09, d (7.2) 4.82, s

1.22, 2.23, 1.52, 2.20, 1.85, 2.21, 1.58, 1.13, 1.94, 1.24, 1.52, 2.07, 1.50, 1.33, 2.58, 2.86, 5.77, 1.05, 5.24,

5.02, s

4.38, s

4.40, s

3.29, 3.53, 1.16, 3.65, 3.36,

3.26, 3.52, 1.19, 3.67,

3.26, 3.51, 1.15, 3.68, 3.54,

qd (7.2, 1.8) qd (7.2, 1.8) t (7.2) s s

m m m m m m m m m m m m m m dd (12.0, 3.0) m s d (7.2) s

qd (7.2, 1.8) qd (7.2, 1.8) t (7.2) s

1.25, 2.08, 1.57, 2.19, 1.88, 2.22, 1.58, 1.16, 1.96, 1.27, 1.52, 2.09, 1.51, 1.32, 2.56, 2.86, 5.78, 1.05, 5.13,

m m m m m m m m m m m m m m dd (12.0, 3.0) qd (7.2, 2.4) s d (7.2) s

qd (7.2, 1.8) qd (7.2, 1.8) t (7.2) s s

5a

6b

1.34, 1.98, 1.56, 2.28, 1.90, 1.99, 1.58, 1.20, 1.97, 1.38, 1.51, 2.13, 1.53, 1.31, 2.53, 2.86, 5.76, 1.05, 3.64, 4.08, 4.20,

m m m m m m m m m m m m m m dd (12.6, 3.0) qd (7.2, 2.4) s d (7.2) d (12.0) dd (12.0, 1.2) d (1.2)

3.23, 3.51, 1.15, 3.64,

qd (7.2, 1.8) qd (7.2, 1.8) t (7.2) s

1.28, 2.10, 1.58, 2.32, 1.90, 2.05, 1.62, 1.21, 2.00, 1.37, 1.57, 2.15, 1.52, 1.31, 2.60, 2.93, 5.88, 1.09, 3.61, 4.10, 4.29, 3.24,

m m m m m m m m m m m m td (12.6, 2.4) m dd (12.6, 3.0) qd (7.2, 2.4) s d (7.2) d (12.0) d (12.0) s s

3.65, s

3.67, qd (7.2, 1.8) 3.89, qd (7.2, 1.8) 1.23, t (7.2) 3.41, qd (7.2, 2.4) 3.87, qd (7.2, 2.4) 1.16, t (7.2)

3.34, qd (7.2, 1.8) 3.52, qd (7.2, 1.8) 1.17, t (7.2) 3.30, s

3.32, s

3.20, s

Spectra were recorded in CDCl3. bSpectra were recorded in methanol-d4. B



Article

Table 2. 13C NMR Spectroscopic Data (150 MHz) for Compounds 1−12 (δC in ppm)

a

position

1a

2a

3a

4a

5a

6b

7a

8a

9a

10a

11a

12a

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 OCH3-12 OCH2CH3-12 OCH2CH3-12 OCH3-18 OCH3-19 OCH2CH3-19 OCH2CH3-19 OCH2CH3-20 OCH2CH3-20 OCH3-20

37.4 21.0 35.8 45.7 45.1 24.0 29.2 41.8 41.3 38.6 37.5 108.2 172.6 37.6 115.4 170.7 11.2 175.7 61.5 97.0

32.9 20.8 36.6 49.8 45.7 24.0 30.4 42.2 42.6 39.8 39.6 108.2 172.2 37.3 114.9 170.7 12.3 174.7 103.6 97.4

37.9 20.6 29.0 49.9 48.0 24.7 29.4 41.8 41.1 38.8 37.7 108.0 172.4 37.5 115.4 170.6 11.3 174.7 96.0 105.2

37.8 20.6 28.9 49.7 47.9 24.5 29.3 41.7 41.0 38.7 37.6 107.9 172.4 37.5 115.5 170.6 11.3 174.7 97.6 105.1

37.4 21.0 35.8, 45.5 45.3 23.8 29.1 41.9 41.2 38.8 37.6 108.0 172.4 37.5 115.3 170.5 11.3 175.6 61.5 103.5

37.6 21.0 35.8 45.7 45.1 24.0 29.3 41.5 41.6 38.6 38.0 105.9 173.6 37.3 113.6 171.2 12.0 175.6 61.7 97.1

37.8 20.6 28.6 50.4 47.2 24.2 29.5 41.1 41.3 38.7 38.1 105.9c 173.7 37.1 113.8 170.7 12.1 175.5 90.1 105.4

30.4 18.7 37.0 47.8 49.5 23.8 30.4 41.6 45.1 40.8 39.5 105.8 173.3 37.0 113.4 171.0 12.9 179.3 17.4 61.4

36.7 18.5 32.2 47.6 49.8 23.4 30.2 42.0 43.9 50.4 38.1 107.1 170.8 37.1 115.9 169.9 11.5 178.2 16.2 207.3

36.9 19.3 35.4 47.8 50.2 23.6 29.5 40.6 42.8 48.0 38.6 107.4 171.6 37.0 115.6 170.4 11.7 179.1 15.6 178.4

32.3 19.3 34.3 58.5 49.6 24.5 29.9 42.2 43.9 50.9 38.2 107.0 170.8 37.1 116.0 169.9 11.6 172.9 171.4 207.8

59.2 15.1 51.7

59.2 15.1 51.7 55.7

59.2 15.1 51.6

59.2 15.1 51.7 57.5

59.1 15.0 51.6

38.8 22.3 37.0 46.9 46.8 25.1 30.3 43.6 42.7 40.1 38.6 109.7 174.7 38.8 116.3 172.6 11.8 177.1 62.5 104.9 54.8 54.8 52.2

51.7

52.3

59.1 15.0 52.3

59.1 14.9 52.1

59.2 15.0 52.9 52.3

51.7

65.7 15.4 64.6 15.5

60.3 15.4 54.8

54.8

54.6

55.7

b

c

Spectra were recorded in CDCl3. Spectra were recorded in methanol-d4. The signal was observed from HMBC correlations.

17 (δH 1.07) indicated that the hydroxy group at C-20 is βoriented, while the ethoxy group at C-12 and the methyl group at C-14 are both α-oriented. A single-crystal X-ray diffraction analysis using the anomalous scattering of Cu Kα radiation yielded a Flack parameter of 0.10 (0.19), which confirmed the structural assignment of 1 as well as its absolute configuration (Figure 2). Thus, the structure of 1 was established as shown. Compound 1 is representative of a new cassane diterpene skeleton with an oxygen bridge between C-19 and C-20. Caesalsappanin B (2) was obtained as a white, amorphous powder, with its molecular formula assigned as C26H38O8 on the basis of the positive HRESIMS (m/z 501.2457 [M + Na]+).

Figure 1. Key 1H−1H COSY (bonds), HMBC (arrows), and NOESY (↔) correlations for compound 1.

proton (H-14) showed long-range correlations with the carbons at δC 108.2 (C-12) and 172.6 (C-13), suggesting that the α,β-unsaturated butenolide moiety is connected to C-11 and C-14. The methyl group (C-17) was located at C-14 from the HMBC correlation between H3-17 (δC 1.07, d, J = 7.2 Hz) and C-14 (δC 37.6). The methoxy and ethoxy groups could be located at C-18 and C-12, respectively, on the basis of the HMBC correlations from δH 3.65 (s, OCH3) to δC 175.7 (C18) and from δH 3.24 and 3.52 (qd, J = 7.2, 1.8 Hz, OCH2CH3) to δC 108.2 (C-12). Also, the oxygen bridge between C-19 and C-20 was confirmed by the HMBC correlations between H-19 and C-20 together with the downfield chemical shifts of C-19 (δC 61.5) and C-20 (δC 97.0). The relative configuration of 1 was determined by a NOESY experiment. In the NOESY spectrum (Figure 1), the key correlations between H-20 (δH 4.82) and H-11α (δH 1.28), between CH3-14 (δH 2.87) and H9 (δH 1.51), and between the proton signal at δH 3.24 and H3-

Figure 2. ORTEP plot for the molecular structure of 1 drawn with 30% probability displacement ellipsoids. C



Article

Table 3. 1H NMR Spectroscopic Data (600 MHz, in CDCl3) for Compounds 7−12 (δH in ppm, J in Hz) position 1α 1β 2α 2β 3α 3β 5 6α 6β 7 8 9 11α 11β 14 15 17 19α 19β 20

7 1.18, 2.01, 1.58, 2.25, 1.88, 1.97, 1.62, 1.24, 2.08, 1.40, 1.59, 2.31, 1.55, 1.62, 2.53, 2.90, 5.68, 1.14, 3.68, 4.33, 4.83,

m m m m m m m m m m m m m m dd (12.0, 2.4) qd (7.2, 2.4) s d (7.2) d (12.0) dd (12.0, 1.8) d (1.8)

8 1.27, 2.04, 1.58, 2.22, 1.81, 2.25, 1.69, 1.17, 1.98, 1.25, 1.55, 2.12, 1.58, 1.41, 2.52, 2.90, 5.70, 1.13, 5.61,


4.39, s

9

10

0.98, m 2.22, m 1.49, m 1.69, m 1.64, m 1.82, m 1.89, m 1.13, m 1.39, m 1.45, m 1.61, m 1.87, m 1.62, m 1.66,m 2.52, dd (12.0, 2.4) 2.94, qd (7.2, 2.4) 5.67, s 1.18, d (7.2) 1.26, s

1.04, 2.49, 1.60, 1.72, 1.61, 2.25, 2.14, 1.60, 1.71, 1.59, 1.82, 1.91, 1.78, 1.16, 2.58, 2.93, 5.76, 1.11, 1.01,

3.83, d (12.0) 3.87, d (12.0)

10.06, s

OCH2CH3-12 OCH2CH3-12 OCH3-18 OCH3-19 OCH3-20

3.67, s

3.71, s

3.21, 3.50, 1.15, 3.69,

3.67, s

m m m m m m dd (7.2, 1.2) m m m m m m m dd (12.0, 2.4) qd (7.2, 2.4) s d (7.2) s

qd (7.2, 1.8) qd (7.2, 1.8) t (7.2) s

11 1.51, 1.77, 1.57, 1.79, 1.04, 2.49, 1.88, 1.07, 2.18, 1.41, 1.53, 1.89, 1.67, 1.00, 2.58, 2.88, 5.75, 1.05, 1.01,

12


1.02, 2.49, 1.59, 1.68, 1.61, 2.40, 2.12, 1.63, 1.70, 1.59, 1.79, 1.90, 1.88, 1.13, 2.58, 2.98, 5.79, 1.12,

m m m m m m dd (7.2, 1.2) m m m m m m m dd (12.0, 2.4) qd (7.2, 2.4) s d (7.2)

9.84, s 3.19, 3.47, 1.12, 3.63,

qd (7.2, 1.8) qd (7.2, 1.8) t (7.2) s

3.21, 3.51, 1.16, 3.73, 3.68,

qd (7.2, 1.8) qd (7.2, 1.8) t (7.2) s s

3.32, s

H-8 (δH 2.07, m) and H-11β. Accordingly, the structure of compound 3 was established as depicted. Caesalsappanin D (4) gave a molecular formula of C25H36O8 on the basis of HRESIMS analysis (m/z 487.2302 [M + Na]+). A comparison of the NMR data of 4 and 3 (Tables 1 and 2) indicated that the ethoxy group at C-19 in 3 is replaced by a methoxy group in 4. This difference was supported by the HMBC correlations of H-19 (δH 5.13, s) with δC 57.5 (−OCH3). Together with supportive evidence from the NOE spectrum, accordingly, the structure of 4 was defined as shown. The molecular formulas of 5 and 6 (caesalsappanins E and F) were both determined as C24H34O7, or 14 mass units more than that of 1. Except for the presence of an additional methoxy signal, their 1H and 13C NMR spectra (Tables 1 and 2) resembled those of 1, suggesting that 5 and 6 are methyl derivatives of 1. The downfield-shifted C-20 (δC 103.5) resonance of 5, in contrast to the C-20 (δC 97.0) signal of 1, as well as the HMBC correlations of H-20 (δH 4.20) with −OCH3 (δC 54.6), suggested that a methoxy group is attached at C-20. The relative configuration of 5 was determined to be the same as that of 1 by analysis of its NOESY spectrum. The 1 H NMR and 13C APT spectroscopic data of 6 were similar to analogous data of 5 except for the downfield-shifted C-12 (Δ1.7 ppm) and C-19 (Δ1.0 ppm) along with the upfieldshifted C-20 (Δ1.4 ppm), caused by the change of the location of methoxy and ethoxy groups in 6. In the HMBC spectrum, the proton signals at δH 3.24 (3H, s) exhibited a long-range correlation with C-12 (δC 109.7), and the resonances at δH 3.34 and 3.52 (2H, dd, J = 7.2, 1.8 Hz) correlated with C-20 (δC 104.9), suggesting the methoxy and ethoxy groups to be located at C-12 and C-20, respectively. NOE correlations from δH 3.24

The NMR data for this compound were similar to those of 1 (Tables 1 and 2), except for the additional ethoxy group signals at δH 3.41, 3.87 (2H, qd, J = 7.2, 2.4 Hz), and δH 1.16 (3H, d, J = 7.2 Hz) and a methoxy group signal at δH 3.36 (3H, s). In the HMBC spectrum, the correlations of δH 3.36 with C-19 (δC 103.6) and of δH 3.41, 3.87 with C-20 (δC 97.4) suggested that the methoxy and ethoxy groups are located at C-19 and C-20, respectively. All cassane-type diterpenes isolated so far from the genus Caesalpinia share the same carbon skeleton, with a trans/ anti/trans system of the three six-membered rings, A, B, and C, and a β-oriented proton at C-8 and the α-oriented protons at C-5/C-9 being well established.24−26 From biogenetic considerations, 2 was inferred as having an identical absolute configuration to 1. The acetal protons at δH 4.82 (H-19) showed NOE enhancements with H-3β and H-2β, and δH 5.02 (H-20) with H-6α and H-11α, which suggested a β-orientation for H-19 and an α-orientation for H-20. Therefore, compound 2 was determined as shown. Caesalsappanin C (3) exhibited the molecular formula C26H38O8, according to HRESIMS (m/z 501.2461 [M + Na]+). The 1H NMR and 13C APT NMR data were closely related to those of 2 (Tables 1 and 2). The differences were in the positions of the methoxy and ethoxy groups, which were at C-19 and C-20 in 2, respectively, but were placed at C-20 and C-19 in 3. The HMBC correlations from H-19 (δH 5.24, s) to δC 65.7 (−OCH2−) and from H-20 (δH 4.38, s) to δC 54.8 (OCH3) confirmed the locations of the ethoxy group at C-19 and the methoxy group at C-20. The α-orientation of H-19 and β-orientation of H-20 were evident from the NOE correlations of H-19 (δH 5.24, s) with H-3α and of H-20 (δH 4.38, s) with D



Article

3.87, d, J = 12.0 Hz) with C-1 (δC 30.4) and C-10 (δC 40.8) were observed. From this information and in conjunction with observations from the NOE spectrum (Figure 3), the structure of 9 was established as shown. Caesalsappanin J (10) exhibited the molecular formula C23H32O6 through HRESIMS analysis (m/z 427.2150 [M + Na]+). The 1H and 13C APT NMR spectra (Tables 2 and 3) were similar to those of 9, except for the presence of ethoxy and aldehyde group signals in 10. In the HMBC spectrum, the ethoxy group protons at δH 3.21 and 3.50 (2H, dd, J = 7.2, 1.8 Hz) showed long-range correlations with C-12 (δC 107.1), indicating that the hydroxy group at C-12 in 9 is replaced with an ethoxy group in 10. Furthermore, the HMBC correlation from the aldehyde group signal (δH 10.06, s) to C-10 (δC 50.4) suggested the hydroxymethyl group at C-20 in 9 to be oxidized to an aldehyde group at C-20 in 10. By inferring a similar biosynthetic pathway for 10 and 9, the configuration of C-20 in 10 was considered to be β, which was also supported by the NOESY correlations between H-20 (δH 10.06, s) and H3-19 (δH 1.01, s) and H-11β (δH 2.58, dd, J = 12.0, 2.4 Hz). Thus, the structure of 10 was assigned as shown. Caesalsappanin K (11) was determined to have a molecular formula of C23H32O7, on the basis of the 13C APT data and by HRESIMS at m/z 443.2052 [M + Na]+ (calcd for C23H32NaO7, 443.2046), accounting for eight indices of hydrogen deficiency. An examination of the 1H and 13C APT NMR data (Tables 2 and 3) showed the structure of 11 to be similar to that of 10. Further analysis of the NMR data of 11 indicated that the aldehyde signal at C-20 of 10 was absent. In contrast, an additional carbonyl signal at δC 178.4 in 11 that could be located at C-20 was observed. In the HMBC spectrum, the correlation signals of H2-1 and H-9 with C-10 (δC 48.0) and C20 (δC 178.4) confirmed the location of a carbonyl group at C20. The carbonyl group in 11 can be inferred as being the result of further oxidation of the aldehyde group at C-20 in 10, so the relative configuration of C-20 in 10 was assigned as β-oriented. On the basis of the above evidence, the structure of 11 was established as shown. Caesalsappanin L (12) was assigned the molecular formula C24H32O8 from the 13C APT NMR and positive-ion mode HRESIMS data (m/z 471.1999 [M + Na]+). The 1H and 13C APT NMR spectroscopic data of 12 were similar to those of 11, implying that these compounds share the same carbon skeleton. Analysis of its 1D (Tables 2 and 3) and 2D NMR data (Supporting Information, Figures S69−72) indicated that the methyl functionality at C-19 in 11 was replaced by a methyl carboxylate group (δH 3.68, δC 52.3, 172.8) in 12. Taken together with information from the NOE spectrum, the structure of 12 was defined as shown. The isolated compounds were tested against the chloroquine-resistant strain K1 of P. falciparum and were also tested against a small panel of human cancer cell lines. Compounds 7 and 8 displayed antimalarial activity in vitro with IC50 values of 0.78 and 0.52 μM and selectivity indices of 17.6 and 16.4, respectively. Compounds 1 and 9 showed moderate antimalarial activity with IC50 values of 7.4 and 2.5 μM, respectively, whereas the other compounds showed only weak activity against the chloroquine-resistant K1 strain of P. falciparum (Table 4). It appears that the presence of a hydroxy group at C12 in cassane-type diterpenoids may play an important role in enhancing activity against the chloroquine-resistant K1 strain of P. falciparum in vitro. In addition, compound 10 showed

(OCH3-12) to H3-17 (δH 1.09, d, J = 7.2 Hz) and from H3-17 to H-9 (δH 1.52, td, J = 12.6, 2.4 Hz) indicated that these hydrogens are cofacial and α-oriented, while the correlations between δH 4.29 (s, H-20) and H-1β (δH 2.10) and H-11β (δH 2.60) and of H-11β with H-8 (δH 2.15) suggested that these protons are β-oriented. Therefore, the structures of 5 and 6 were established as shown. Caesalsappanin G (7) exhibited the molecular formula C21H28O7, according to HRESIMS (m/z 415.1737 [M + Na]+). The 1H and 13C APT NMR spectra (Tables 2 and 3) of 7 were comparable with those of 6. The differences evident were that both the methoxy and ethoxy group signals in 6 were absent in 7. The upfield-shifted C-12 (δC 105.9) and C-20 (δC 97.1) signals of 7, in contrast to resonance for C-12 (δC 109.7) and C-20 (δC 104.9) of 6, as well as the HMBC correlations of H-20 (δH 4.83) with C-19 (δC 61.7) and C-1 (δC 37.6), together with the above molecular formula, suggested the presence of hydroxy groups at C-12 and C-20 in 7. A correlation between H-20 and H-11α observed in the NOESY spectrum of 7 indicated the β-orientaton of OH-20. Accordingly, the structure of 7 was assigned as shown. Caesalsappanin H (8) showed a [M + Na]+ ion peak at m/z 445.1836 in the HRESIMS, corresponding to the molecular formula C22H30O8. The obtained 1H and 13C APT NMR (Tables 2 and 3) spectra were closely related to those of 7, with the exception of signals for an extra methoxy group. The downfield-shifted C-20 dioxymethine carbon (δC 105.4) of 8, in contrast to the C-20 (δC 97.1) signal of 7, as well as the HMBC correlation of H-20 (δH 4.39, s) with −OCH3 (δC 55.7), suggested the additional methoxy group to be located at C-20. In the NOESY spectrum, the dioxymethine protons at δH 5.61 (H-19) showed enhancements with H-6β (δH 1.98), and δH 4.39 (H-20) with H-1β (δH 2.10) and H-11β (δH 2.52), suggesting the α-orientation for OH-19 and OCH3-20. Thus, the structure of 8 was characterized as shown. Caesalsappanin I (9) was obtained as an amorphous, white powder. HRESIMS analysis showed a quasimolecular ion peak at m/z 401.1921 [M + Na]+ (calcd for 401.1940) in the positive-ion mode. In conjunction with the 1H and 13C APT NMR data (Tables 2 and 3), the molecular formula was deduced as C21H30O6, representing seven indices of hydrogen deficiency. The olefinic proton signal at δH 5.67 (H-15, s) and downfield carbon signals at δC 105.8 (C-12), 113.4 (C-15), 171.0 (C-16), and 173.3 (C-13) indicated the presence of an α,β-unsaturated butenolide moiety.17−19 The two methyl signals at δH 1.18 (H3-17, d, J = 7.2 Hz) and 1.26 (H3-19, s) together with the above-mentioned degree of unsaturation suggested that 9 is a typical tetracyclic cassane-type diterpene possessing a fused butenolide unit.27,28 In the HMBC spectrum (Figure 3), correlations of H3-19 (δH 1.26) with C-4 (δC 47.8), C-5 (δC 49.5), and C-18 (δC 179.3) and of H2-20 (δH 3.83,

Figure 3. Selected HMBC (→) and NOESY (↔) correlations for compound 9. E



Article

Table 4. In Vitro Antimalarial Activities of Compounds 1−12 against the Chloroquine-Resistant K1 Strain of Plasmodium falciparum compound 1 2 3 4 5 6 chloroquinec

IC50 (μM)a 7.4 34.8 32.1 28.4 15.7 19.2 0.37

± ± ± ± ± ± ±

0.25 1.53 2.4 1.05 0.64 0.82 0.02

LC50 (μM)a

SIb

compound

± ± ± ± ± ± ±

12.3 11.7 14.3 9.6 5.2 4.8 129.5

7 8 9 10 11 12

91.0 407 459 272 81.6 92.2 47.9

6.9 20.8 7.3 11.6 10.1 5.8 4.7

IC50 (μM)a 0.78 0.52 2.5 29.7 41.2 38.9

± ± ± ± ± ±

0.32 0.15 1.2 5.7 3.2 10.3

LC50 (μM)a

SIb

± ± ± ± ± ±

17.6 16.4 10.5 3.6 7.2 4.1

13.7 8.50 26.3 106 296 159

3.1 2.7 3.1 9.6 6.2 31.7

IC50 = inhibitory concentration 50%; LC50 = lethal concentration 50%. bSI = selectivity index. Values are means ± SD of triplicate experiments. Positive control substance.

a c

Subfraction Fr. E4 (821 mg) was purified via Kromasil RP-18 CC eluting with 70% MeOH in H2O followed by recrystallization using MeOH as solvent to produce 10 (24.7 mg). Compounds 6 (6.1 mg, tR 24.7 min) and 8 (5.2 mg, tR 28.1 min) were purified from subfraction Fr. E5 (934 mg) by semipreparative Kromasil RP-18 HPLC using MeOH−H2O (68:32, v/v) as the mobile phase. Fraction F (33.1 g) was chromatographed by silica gel CC eluting with CH2Cl2−MeOH (from 40:1 to 1:1, v/v) to yield 10 subfractions (Fr. F1−10). Frs. F4 and F5 were separated via semipreparative HPLC with a MeOH−H2O gradient solvent system (from 40% to 100% MeOH) using a Kromasil RP-18 column. Finally, compounds 7 (4.9 mg) and 12 (6.3 mg) were obtained at tR 22.5 and 26.4 min, respectively, using a MeOH−H2O (62:38) system from Fr. F4, and compound 9 (3.5 mg) was obtained at tR 30.5 min using a MeOH−H2O (65:45) solvent system from Fr. F5. Caesalsappanin A (1): colorless crystals (MeOH); [α]20 D −64.6 (c 0.1, MeOH); UV (MeOH) λmax (log ε) 214 (3.89) nm; IR (film) νmax 3500, 1749 cm−1; 1H and 13C NMR data, see Tables 1 and 2; HRESIMS m/z 443.2045 [M + Na]+, calcd for C23H32O7Na, 443.2046. Caesalsappanin B (2): amorphous, white powder; [α]20 D +16.7 (c 0.1, MeOH); UV (MeOH) λmax (log ε) 214 (3.21) nm; IR (film) νmax 2923, 2867, 1729 cm−1; 1H and 13C NMR data, see Tables 1 and 2; HRESIMS m/z 501.2457 [M + Na]+, calcd for C26H38O8Na, 501.2464. Caesalsappanin C (3): amorphous, white powder; [α]20 D −44.2 (c 0.1, MeOH); UV (MeOH) λmax (log ε) 213 (4.02) nm; IR (film) νmax 2928, 2871, 1766 cm−1; 1H and 13C NMR data, see Tables 1 and 2; HRESIMS m/z 501.2461 [M + Na]+, calcd for C26H38O8Na, 501.2464. Caesalsappanin D (4): amorphous, white powder; [α]20 D −49.3 (c 0.2, MeOH); UV (MeOH) λmax (log ε) 215 (3.12) nm; IR (film) νmax 2936, 2837, 1729 cm−1; 1H and 13C NMR data, see Tables 1 and 2; HRESIMS m/z 487.2302 [M + Na]+, calcd for C25H36O8 Na, 487.2308. Caesalsappanin E (5): amorphous, white powder; [α]20 D −77.1 (c 0.2, MeOH); UV (MeOH) λmax (log ε) 214 (4.94) nm; IR (film) νmax 2932, 2886, 1766 cm−1; 1H and 13C NMR data see Tables 1 and 2; HRESIMS m/z 457.2204 [M + Na]+, calcd for C24H34O7 Na, 457.2202. Ccaesalsappanin F (6): amorphous, white powder; [α]20 D −19.5 (c 0.1, MeOH); UV (MeOH) λmax (log ε) 213 (3.75) nm; IR (film) νmax 2932, 2886, 1729 cm−1; 1H and 13C NMR data, see Tables 1 and 2; HRESIMS m/z 457.2195 [M + Na]+, calcd for C24H34O7 Na, 457.2202. Caesalsappanin G (7): amorphous, white powder; [α]20 D −17.3 (c 0.1, MeOH); UV (MeOH) λmax (log ε) 214 (4.37) nm; IR (film) νmax 3452, 1726 cm−1; 1H and 13C NMR data, see Tables 2 and 3; HRESIMS m/z 415.1737 [M + Na]+, calcd for C21H28O7Na, 415.1733. Caesalsappanin H (8): amorphous, white powder; [α]20 D −39.3 (c 0.1, MeOH); UV (MeOH) λmax (log ε) 213 (3.38) nm; IR (film) νmax 3430, 1727 cm−1; 1H and 13C NMR data, see Tables 2 and 3;

antiproliferative activity against the KB cancer cell line, with an IC50 value of 7.4 μM.

■

EXPERIMENTAL SECTION

General Experimental Procedures. Optical rotation data were obtained using a PerkinElmer 341 digital polarimeter. UV data were recorded with a Shimadzu UV2550 spectrometer. IR data were recorded using a FTIR-8400S spectrometer. NMR spectra were obtained using a Bruker AV III 600 NMR spectrometer with the chemical shift values presented as δ values having TMS as an internal standard. HRESIMS was performed using an LTQ-Obitrap XL spectrometer. HPLC separations were conducted using a Lumiere K-1001 pump, a Lumiere K-2501 single λ absorbance detector, and a Kromasil semipreparative column packed with C18 reversed-phase silica gel (250 × 10 mm, 5 μm). Sephadex LH-20 (Pharmacia, Uppsala, Sweden), MCI gel (CHP 20P, 75−150 μm, Mitsubishi Chemical Corporation, Tokyo, Japan), C18 reversed-phase silica gel (40−63 μm, Merck, Darmstadt, Germany), and silica gel (100−200 and 300−400 mesh, Qingdao Marine Chemical Inc., Qingdao, People’s Republic of China) were used for CC, and precoated silica gel GF254 plates (Zhi Fu Huang Wu Pilot Plant of Silica Gel Development, Yantai, People’s Republic of China) were used for TLC. All solvents employed were of analytical grade (Beijing Chemical Plant, People’s Republic of China). Plant Material. Casealpinia sappan seeds were collected in Nanning, Guangxi Province, People’s Republic of China, in April 2013 and identified by Prof. Jing-Quan Yuan of the Department of Pharmaceutical Chemistry, Guangxi Botanical Garden of Medicinal Plants. A voucher specimen (no. 13418) was deposited at the Guangxi Botanical Garden of Medicinal Plants. Extraction and Isolation. The air-dried and powdered seeds of C. sappan (5.0 kg) were extracted with MeOH (3 × 40 L, 2 h each). Removal of the MeOH under reduced pressure yielded a MeOHsoluble extract (1267 g). The residue was suspended in H2O (3 L) and partitioned with petroleum ether (3 × 3 L), CHCl3 (3 × 3 L), EtOAc (3 × 3 L), and n-BuOH (3 × 3 L), successively. The CHCl3 fraction (201 g) was subjected to CC over silica gel (100−200 mesh, 15 × 60 cm) eluting with a stepwise gradient of CH2Cl2−MeOH (from 1:0 to 0:1) to afford fractions A−K. Fr. C (2.8 g) was subjected to CC over silica gel (300−400 mesh, 2 × 80 cm) eluting with petroleum ether− CHCl3 (1:1, 1:2, 1:2.5, 1;3, 1:4, 1:5, 0:1, v/v) followed by CHCl3− MeOH (100:1, 80:1, 40:1, v/v), yielding compounds 1 (106.4 mg) and 5 (8.5 mg). Fr. D (20.6 g) was purified using a Sephadex LH-20 column (3 × 135 cm) eluting with CHCl3−MeOH (30:70, v/v) and then subjected to MPLC using C18 reversed-phase silica gel, by elution with MeOH−H2O (30:70; 60:40; 70:30; 80:20; 90:10; 100:0, v/v), to yield six fractions (Fr. D1−6). Fr. D2 (3.1 g) was subjected to CC on silica gel (300−400 mesh), eluting with CHCl3−MeOH (100:1; 80:1; 60:1; 50:1; 40:1; 30:1; 20:1; 0:1, v/v), to afford compounds 4 (6.8 mg) and 11 (79.6 mg). Fr. D3 (1.2 g) was separated using semipreparative HPLC using a mobile phase of MeOH−H2O (78:22, v/v) and a Kromasil RP-18 column to afford compounds 2 (4.3 mg, tR 32.5 min) and 3 (4.7 mg, tR 35.7 min). Fr. E (12.1 g) was applied to silica gel CC to give five subfractions (Fr. E1−5). F



Article

values of 1.2, 0.68, 1.4, and 0.83 μM against the HeLa, HT-29, KB, and MCF-7 cell lines, respectively.

HRESIMS m/z 445.1836 [M + Na]+, calcd for C22H30O8Na, 445.1838. Caesalsappanin I (9): amorphous, white powder; [α]20 D −62.8 (c 0.1, MeOH); UV (MeOH) λmax (log ε) 213 (4.38) nm; IR (film) νmax 3433, 1735 cm−1; 1H and 13C NMR data, see Tables 2 and 3; HRESIMS m/z 401.1921 [M + Na]+, calcd for C21H30O6Na, 401.1940. Caesalsappanin J (10): amorphous, white powder; [α]20 D −74.3 (c 0.11, MeOH); UV (MeOH) λmax (log ε) 214 (3.62) nm; IR (film) νmax 2728, 1738 cm−1; 1H and 13C NMR data, see Tables 2 and 3; HRESIMS m/z 427.2150 [M + Na]+, calcd for C23H32O6Na, 427.2097. Caesalsappanin K (11): amorphous, white powder; [α]20 D −53.7 (c 0.1, MeOH); UV (MeOH) λmax (log ε) 212 (3.59) nm; IR (film) νmax 2938, 2884, 1742 cm−1; 1H and 13C NMR data, see Tables 2 and 3; HRESIMS m/z 443.2052 [M + Na]+, calcd for C23H32O7Na, 443.2045. Caesalsappanin L (12): amorphous, white powder; [α]20 D −48.6 (c 0.1, MeOH); UV (MeOH) λmax (log ε) 215 (3.72) nm; IR (film) νmax 2939, 2885, 1740 cm−1; 1H and 13C NMR data, see Tables 2 and 3; HRESIMS m/z 471.1999 [M + Na]+, calcd for C24H32O8Na, 471.1995. X-ray Diffraction Analysis of 1. A colorless crystal of 1 was obtained from MeOH. Intensity data were collected at room temperature on a Bruker APEX DUO diffractometer equipped with an APEX II CCD, using Cu Kα radiation. Cell refinement and data reduction were performed with Bruker’s SAINT program. The structures were solved by direct methods using SHELXS-97.29 Refinements were performed with SHELXL-97 using full-matrix least-squares, with anisotropic displacement parameters for all the non-hydrogen atoms. The H atoms were placed in calculated positions and refined using a riding model. Molecular graphics were computed with PLATON. Crystallographic data for compound 1 have been deposited with the Cambridge Crystallographic Data Center as supplementary publication (CCDC 1037844). Crystal data: C23H32O7, M = 420.49, orthorhombic, crystal size 0.29 × 0.24 × 0.21 mm3, space group P212121, a = 9.7720(10) Å, b = 13.5369(2) Å, c = 16.5519(2) Å, α = 90.00°, β = 90.00°, γ = 90.00°, V = 2189.53(5) Å3, T = 298(2) K, space room P212121, Z = 4, μ(Cu Kα) = 0.770 mm−1, Dcalcd = 1.276 mg/m3, 3692 reflections measured, 3645 independent reflections (Rint = 0.0215). The final R1 value was 0.0351 (I > 2δ(I)). The final wR(F2) value was 0.1044 (I > 2δ(I)). The final R1 value was 0.0356 (all data). The final wR(F2) value was 0.1048 (all data). The goodness of fit on F2 was 1.046. Flack parameter = 0.10(19). Antimalarial Assays. Antimalarial activity in vitro was determined by means of the microculture radioisotope technique based on the method described by Desjardins et al.30 The parasite Plasmodium falciparum (K1, mutidrug-resistant strain), which was obtained from the Academy of Military Medical Sciences (Beijing, People’s Republic of China), was cultured continuously according to the method of Trager and Jensen.31 Three replications were used for each experiment. Data are presented as means ± SEM. Statistical analysis was performed by means of the Student t-test. A p value of less than 0.05 was considered a significant difference. Antiproliferative Assays. Cytotoxicity was assessed against HeLa (cervical cancer), HT-29 (colon cancer), KB (oral cancer), and MCF7 (breast adenocarcinoma cancer) human cancer cell lines by the MTT method. Cells were grown in DMEM medium supplied with 10% fetal bovine serum and cultured at a density of 6 × 104 cells/mL per well in a 96-well microtiter plate. Then, five different concentrations of each compound dissolved in dimethyl sulfoxide (DMSO) were added to each well. Each concentration was tested in triplicate. After incubation at 37 °C in 5% CO2 for 48 h, 10 mL of MTT (4 mg/mL) was added to each well and incubated for another 4 h. The medium was then removed, and cells were then lysed with 200 μL of DMSO. The absorbance was recorded on a microplate reader at a wavelength of 570 nm. The experiments were conducted a minimum of three times. Doxorubicin was used as the positive control with IC50

■

ASSOCIATED CONTENT

S Supporting Information *

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jnatprod.5b00317. Crystallographic data for compound 1 (CIF) NMR spectra of compounds 1−12 (Figures S1−S48) (PDF)

■

AUTHOR INFORMATION

Corresponding Authors

*Tel/Fax: 86-771-5601290. E-mail: [email protected] (J.-Q. Yuan). *Tel/Fax: 86-10-57833296. E-mail: [email protected] (X.-D. Xu). Author Contributions ⊥

G. Ma and H. Wu contributed equally to this work.

Notes

The authors declare no competing financial interest.

■

ACKNOWLEDGMENTS The work was financially supported by the Technological Large Platform for Comprehensive Research and Developmentof New Drugs in the Twelfth Five-Year “Significant New Drugs Created” Science and Technology Major Projects (No. 2012ZX09301-002-001-026), the National Science and Technology Support Program (No. 2012BA127B06), the Innovation Capacity Building in Guangxi Science and Technology Agency (No. 10100027-3), the National Natural Sciences Foundaton of China (No. 81502945), and the PUMC Youth Fund (No. 3332015141).

■

REFERENCES

(1) Snow, R. W.; Guerra, C. A.; Noor, A. M.; Myint, H. Y.; Hay, S. I. Nature 2005, 434, 214−217. (2) Greenberg, P. L.; Gordeuk, V.; Issaragrisil, S.; Siritanaratkul, N.; Fucharoen, S.; Ribeiro, R. C. Hematology 2001, 2001, 479−498. (3) Miller, L. H.; Baruch, D. I.; Marsh, K.; Doumbo, O. K. Nature 2002, 415, 673−679. (4) Nosten, F.; White, N. J. Am. J. Trop. Med. Hyg. 2007, 77, 181− 192. (5) Buendia-Orozco, J.; Guerrero, A.; Pastor, N. Arch. Med. Res. 2005, 36, 317−330. (6) He, S. Z. Chin. Tradit. Herb. Drugs 2000, 31, 225−226. (7) Zheng, Y.; Zhang, S. W.; Cong, H. J.; Huang, Y. J.; Xuan, L. J. J. Nat. Prod. 2013, 76, 2210−2218. (8) Wu, L.; Wang, X. B.; Shan, S. M.; Luo, J.; Kong, L. Y. Chem. Pharm. Bull. 2014, 62, 729−733. (9) Badami, S.; Rai, S. R.; Moorkoth, S.; Rajan, S.; Suresh, B. Anc. Sci. Life 2003, 23, 100−107. (10) Wang, D.; Chen, C.; Zhou, Y. Inf. Tradit. Chin. Med. 2003, 20, 15−16. (11) Zhang, J. Y.; Wu, F. H.; Qu, W.; Liang, J. Y. Zhongguo Tianran Yaowu 2012, 10, 218−221. (12) Zhang, J. Y.; Abdel-Mageed, W. M.; Liu, M. M.; Huang, P.; He, W. N.; Li, L.; Song, F. H.; Dai, H. Q.; Liu, X. T.; Liang, J. Y.; Zhang, L. X. Org. Lett. 2013, 15, 4726−4729. (13) Matsuno, Y.; Deguchi, J.; Hirasawa, Y.; Ohyama, K.; Toyoda, H.; Hirobe, C.; Ekasari, W.; Widyawaruyanti, A.; Zaini, N. C.; Morita, H. Bioorg. Med. Chem. Lett. 2008, 18, 3774−3777.

G



Article

(14) Yodsaoue, O.; Cheenpracha, S.; Karalai, C.; Ponglimanont, C.; Chantrapromma, S.; Fun, H. K.; Kanjana-Opas, A. Phytochemistry 2008, 69, 1242−1249. (15) Linn, T. Z.; Awale, S.; Tezuka, Y.; Banskota, A. H.; Kalauni, S. K.; Attamimi, F.; Ueda, J. Y.; Asih, P. B.; Syafruddin, D.; Tanaka, K.; Kadota, S. J. Nat. Prod. 2005, 68, 706−710. (16) Maurya, R.; Ravi, M.; Singh, S.; Yadav, P. P. Fitoterapia 2012, 83, 272−280. (17) Kalauni, S. K.; Awale, S.; Tezuka, Y.; Banskota, A. H.; Linn, T. Z.; Asih, P. B.; Syafruddin, D.; Kadota, S. Biol. Pharm. Bull. 2006, 29, 1050−1052. (18) Ma, G.; Sun, Z.; Sun, Z.; Yuan, J.; Wei, H.; Yang, J.; Wu, H.; Xu, X. Fitoterapia 2014, 95, 234−239. (19) Ma, G.; Yuan, J.; Wu, H.; Cao, L.; Zhang, X.; Xu, L.; Wei, H.; Wu, L.; Zheng, Q.; Li, L.; Zhang, L.; Yang, J.; Xu, X. J. Nat. Prod. 2013, 76, 1025−1031. (20) Kinoshita, T.; Haga, Y.; Narimatsu, S.; Shimada, M.; Goda, Y. Chem. Pharm. Bull. 2005, 53, 717−720. (21) Ma, G.; Xu, X.; Cao, L.; Yuan, J.; Yang, J.; Ma, L. Planta Med. 2012, 78, 1363−1369. (22) Suthiwong, J.; Pitchuanchom, S.; Wattanawongdon, W.; Hahnvajanawong, C.; Yenjai, C. J. Nat. Prod. 2014, 77, 2432−2437. (23) Wu, Z.; Zhang, Y.; Zhu, K.; Luo, C.; Zhang, J.; Cheng, C.; Feng, R.; Yang, W.; Zeng, F.; Wang, Y.; Xu, P.; Guo, J.; Liu, X.; Guan, S.; Guo, D. J. Nat. Prod. 2014, 77, 2342−2351. (24) Cota, B. B.; Oliveira, D. M. D.; Siqueira, E. P. D.; SouzaFagundes, E. M.; Pimenta, A. M. C.; Santos, D. M.; Rabello, A.; Zani, C. L. Fitoterapia 2011, 82, 969−975. (25) Gómez-Hurtado, M. A.; Á lvarez-Esquivel, F. E.; RodríguezGarcía, G.; Martínez-Pacheco, M. M.; Espinoza-Madrigal, R. M.; Pamatz-Bolaños, T.; Salvador-Hernández, J. L.; García-Gutiérrez, H. A.; Cerda-García-Rojas, C. M.; Joseph-Nathan, P.; Río, R. E. D. Phytochemistry 2013, 96, 397−403. (26) Yodsaoue, O.; Karalai, C.; Ponglimanont, C.; Tewtrakul, S.; Chantrapromma, S. Tetrahedron 2011, 67, 6838−6846. (27) Matsuno, Y.; Deguchi, J.; Hosoya, T.; Hirasawa, Y.; Hirobe, C.; Shiro, M.; Morita, H. J. Nat. Prod. 2009, 72, 976−979. (28) Yin, Y.; Ma, L.; Hu, L. Helv. Chim. Acta 2008, 91, 972−977. (29) Sheldrick, G. M. Acta Crystallogr., Sect. A: Found. Crystallogr. 2008, A64, 112−122. (30) Desjardins, R. E.; Canfield, C. J.; Haynes, J. D.; Chulay, J. D. Antimicrob. Agents Chemother. 1979, 16, 710−718. (31) Trager, W.; Jensen, J. B. Science 1976, 193, 673−675.

H


Antimalarial and Antiproliferative Cassane Diterpenes of Caesalpinia

Recommend Documents