Iboga-Type Alkaloids from Ervatamia officinalis - American Chemical

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Iboga-Type Alkaloids from Ervatamia of ficinalis Ben-Qin Tang,†,‡,§ Wen-Jing Wang,‡,§ Xiao-Jun Huang,‡ Guo-Qiang Li,‡ Lei Wang,*,‡ Ren-Wang Jiang,‡ Ting-Ting Yang,‡ Lei Shi,‡ Xiao-Qi Zhang,*,‡ and Wen-Cai Ye*,†,‡ †

Department of Natural Medicinal Chemistry, China Pharmaceutical University, Nanjing 210009, People’s Republic of China Institute of Traditional Chinese Medicine and Natural Products, College of Pharmacy, Jinan University, Guangzhou 510632, People’s Republic of China

‡

S Supporting Information *

ABSTRACT: Seven new iboga-type alkaloids, ervaoffines A− D (1−4), (7S)-3-oxoibogaine hydroxyindolenine (5), ibogaine-5,6-dione (6), and 19-epi-5-oxovoacristine (7), and 10 known alkaloids were isolated from Ervatamia off icinalis. The absolute configurations of 1−7 were determined through X-ray diffraction and electronic circular dichroism (ECD) analyses. Ervaoffines A and B represent the first iboga-type pseudoindoxyl alkaloids in which the C-2 spiro carbon configuration is opposite to that of other members of this class, such as iboluteine (8). The relationship between the absolute configuration of the spiro carbons and the Cotton effect in the ECD spectrum is established for the first time for iboga-type pseudoindoxyl and oxindole alkaloids. Additionally, a plausible biogenetic pathway for these alkaloids is proposed.

Ervatamia off icinalis is a common shrub that is primarily distributed in the Guangdong and Hainan Provinces of China.1 Plants from the genus Ervatamia are a rich source of monoterpenoid indole alkaloids, particularly iboga-type alkaloids, which exhibit significant biological activities.2 Owing to their complex polycyclic structures and promising antiaddictive properties, iboga alkaloids have long been of considerable interest for organic chemists.3−5 However, the structural complexity and multiple stereogenic centers of iboga-type alkaloids have presented difficulties in determining their absolute configurations using spectroscopic methods. Accordingly, the absolute configurations of most iboga alkaloids have been established by correlation with the chemical structures of parent alkaloids and based on biogenetic considerations.6−8 In our investigation of structurally unique and biologically active substances from medicinal plants,9−11 seven new iboga-type alkaloids, ervaoffines A−D (1−4), (7S)-3-oxoibogaine hydroxyindolenine (5), ibogaine-5,6-dione (6), and 19-epi-5-oxovoacristine (7), and 10 known compounds, iboluteine (8),12 (7S)ibogaine hydroxyindolenine (9),13 ibogaine (10),12 ibogaline (11),14 voacangine (12),15 conopharyngine (13),16 voacristine (14),17 heyneanine (15),17 19S-hydroxyibogamine (16),18 and ibogaine N4-oxide (17),19 were isolated from E. off icinalis. The absolute configurations of 1−7 were determined using singlecrystal X-ray diffraction and electronic circular dichroism (ECD) analyses supported by time-dependent density functional theory (TDDFT) calculations. Ervaoffines A (1) and B (2) represent the first iboga-type pseudoindoxyl alkaloids in which the C-2 spiro carbon configuration is opposite to that of known members of this class, such as iboluteine (8). The relationship between the absolute configuration of the spiro © XXXX American Chemical Society and American Society of Pharmacognosy

carbon in iboga-type pseudoindoxyl and oxindole alkaloids and the Cotton effect in the associated ECD spectrum is established for the first time. The presence of 1−3, 8, and ervahainine A11 in plants of the genus Ervatamia suggested that these alkaloids may be formed through a pinacol rearrangement.

■

RESULTS AND DISCUSSION Ervaoffine A (1) was isolated as yellow crystals via crystallization from 80% aqueous methanol. The molecular formula, C20H24N2O3, was established based on its 13C NMR data and an m/z 363.1680 [M + Na]+ ion in the HRESIMS. The IR spectrum exhibited absorptions of amino (3374 cm−1), carbonyl (1671 cm−1), and aromatic (1593, 1496 cm−1) functionalities. The 1H NMR spectrum (Table 1) exhibited resonances for a 1,2,4-trisubstituted benzene ring [δH 7.13 (1H, dd, J = 8.8, 2.6 Hz, H-11), 6.95 (1H, d, J = 2.6 Hz, H-9), and 6.88 (1H, d, J = 8.8 Hz, H-12)], a bridgehead proton adjacent to a nitrogen [δH 3.05 (1H, d, J = 3.2 Hz, H-21)], a methoxy group [δH 3.74 (3H, s)], as well as an ethyl side chain [δH 1.54 (1H, overlapped, H-19a), 1.50 (1H, overlapped, H-19b), and 0.95 (3H, t, J = 7.4 Hz, H-18)]. The 13C NMR and DEPT−135 spectra of 1 showed 20 resonances comprising a carbonyl, six olefinic carbons, a quaternary carbon, six methine, four methylene, and two methyl carbons. The 13C NMR data of 1 closely matched those of iboluteine (8),12 except for the absence of resonances for two methylenes [δC 52.0 (C-3) and 22.6 (C-6)] and the presence of resonances for two oxygenated Received: March 13, 2014

A

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Chart 1

Table 1. 1H (600 MHz) and 13C (150 MHz) NMR Data for Compounds 1−3 (δ in ppm, J in Hz)a 1b δH

position 2 3 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 OCH3 NH a

4.85 α β

3.45 d (12.0) 2.89 dd (12.0, 3.9) 3.86 d (3.9)

6.95 d (2.6) 7.13 dd (8.8, 2.6) 6.88 d (8.8)

α β α β a b

2.20 1.83 1.13 1.59 1.05 3.07 0.95 1.54 1.50 1.44 3.05 3.74

dd (5.2, 5.1) m m dd (9.8, 3.2) dd (13.4, 9.8) t (7.4)

d (3.2) s

2c δC

mult

71.1 96.6

C CH

δH

57.0

CH2

75.4 204.4 121.8 105.0 154.7 129.0 115.3 158.0 31.3 29.8

CH C C CH C CH CH C CH CH2

43.5 19.2

CH CH2

12.0 27.6

CH3 CH2

1.80 1.73 1.12 1.80 1.45 2.60 0.93 1.56

37.5 59.1 56.2

CH CH CH3

1.51 2.74 3.75 s

a b α β

2.87 dt (11.5, 2.7) 3.16 2.77 3.13 4.45 dd (11.5, 5.3)

7.01 d (2.7) 7.13 dd (8.8, 2.7) 6.97 d (8.8)

α β α β

m m m m t (7.1)

3c δC

mult

72.1 53.8

C CH2

57.0

CH2

62.1 202.3 122.5 104.5 154.1 127.4 115.2 156.4 25.2 32.4

CH C C CH C CH CH C CH CH2

40.7 23.8

CH CH2

11.9 28.7

CH3 CH2

38.6 53.6 55.7

CH CH CH3

δH 4.95 d (5.0) α β

4.40 d (11.5) 2.76 dd (11.5, 3.8) 4.04 d (3.8)

7.42 d (2.4) 6.79 dd (8.4, 2.4) 6.76 d (8.4)

α β α β a b

2.29 1.79 1.19 1.75 1.24 2.44 0.92 1.57 1.51 1.33 3.55 3.82 7.78

dd (5.6, 5.0)

dd (13.9, 5.6) t (7.4)

m d (3.2) s br s

δC

mult

178.9 95.5

C CH

52.2

CH2

75.9 55.5 133.7 115.3 155.6 112.6 109.2 133.3 30.2 28.8

CH C C CH C CH CH C CH CH2

41.5 24.1

CH CH2

11.8 26.7

CH3 CH2

35.9 54.6 56.0

CH CH CH3

Overlapped resonances are reported without designating multiplicity. bMeasured in methanol-d4. cMeasured in CDCl3.

B

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a 6-hydroxy iboga-type pseudoindoxyl alkaloid. This was supported by the HMBC correlations between H-5β and C2/-3/-21 and between H-6 and C-5/-7. The α-orientation of 6OH was deduced by the NOESY correlations between H-6 and H-3a/-17β. Furthermore, the ECD spectrum of 2 exhibited Cotton effects similar to those observed for 1 (Supporting Information). Thus, its absolute configuration was defined as 2S,6R,14R,16R,20S,21S. Ervaoffine C (3) has the same molecular formula as 1, C20H24N2O3, established by the 13C NMR and HRESIMS data. Comparison of the 13C NMR spectra of 3 and 1 revealed considerable structural similarity, except for the conspicuous deshielding of the carbonyl resonance (from δC 204.4 to 178.9) and the spiro carbon resonance (from δC 71.1 to 55.5), indicating that the pseudoindoxyl moiety in 1 was replaced by an oxindole group in 3. This was confirmed by the HMBC correlations between H-5β/-9/-17α/-17β and C-7, between H6 and C-8, and between H-16 and C-2. Additionally, the HMBC correlations between H-5α/-6 and C-3 confirmed the presence of a 1,3-oxazolidine ring, similar to 1. The NOESY data revealed that H-5α and -21, as well as H-15α, -16, -17α, and -20, were cofacial. Correlations between H-9 and H-17β suggested a 7R* configuration. The absolute configuration of 3 was determined by comparison of its experimental and calculated ECD data. The experimental ECD spectrum of 3 exhibited negative Cotton effects at 305 (Δε −4.8) and 260 (Δε −6.9) nm and positive Cotton effects at 233 (Δε +20.2) and 206 (Δε +18.5) nm, which were similar to those in the calculated spectrum for the isomer with 3S,6R,7R,14R,16S,20S,21S configurations (Figure 5). Thus, the absolute configuration of 3 was established as 3S,6R,7R,14R,16S,20S,21S. Ervaoffine D (4) exhibited a molecular formula of C20H24N2O4, as deduced from its 13C NMR and HRESIMS data. The 1H NMR spectrum of 4 (Table 2) exhibited resonances for three aromatic protons [δH 7.06 (2H, s, H-11/ H-12), 6.79 (1H, br s, H-9)], a bridgehead proton adjacent to a nitrogen [δH 4.08 (1H, br s, H-21)], a methoxy [δH 3.76 (3H, s)], and an ethyl [δH 1.28 (1H, m, H-19a), 1.15 (1H, m, H19b), and 0.91 (3H, t, J = 7.3 Hz, H-18)] group. The 13C NMR data of 4 resembled those of ibogaine (10),12 except for the absence of resonances for two olefinic carbons [δC 143.0 (C-2) and 108.9 (C-7)] and a methylene at δC 49.9 (C-3) and the presence of three carbonyl resonances (δC 202.4, 173.0, 172.7). The resonance at δC 202.4 was assigned to C-7 based on the HMBC correlations between H-9/-5α/-5β and C-7. Additionally, the lactam carbonyl resonances at δC 173.0 and 172.7 were assigned to C-3 and C-2, respectively, based on the HMBC

methines, which were assigned to C-3 (δC 96.6) and C-6 (δC 75.4) based on the 1H−1H COSY correlations between H-6 and H-5β and between H-3 and H-14 in addition to the HMBC correlations between H-6 and C-7/-16 and between H-3 and C-5/-17/-21 (Figure 1). On the basis of the molecular formula,

Figure 1. 1H−1H COSY and selected HMBC correlations of 1 and 3.

C-3 and C-6 were anticipated to be connected by an ether function to form a 1,3-oxazolidine moiety, and this expectation was confirmed by the HMBC correlations between H-3 and C6 (Figure 1). The relative configuration of 1 was deduced using NOESY analysis (Figure 2). The NOESY cross-peaks between H-5α and H-6/-21 suggested that H-5α, -6, and -21 were cofacial. Correlations between H-16 and H-17α/-20 and between H-20 and H-15α suggested that these protons possessed identical orientations. Similarly, H-3, -5β, -14, and -15β were also cofacial based on their NOESY correlations. A sample of 1 was dissolved in DMSO-d6 and showed clear NOESY correlations between NH and H-5α/-21, which suggested a 2S* configuration. Crystals suitable for X-ray crystallographic analysis (Figure 3) were obtained from MeOH−H2O, the X-ray data permitting definition of the absolute configuration of 1 based on a Flack parameter of 0.02(16). Additionally, the ECD spectra calculations for (2S,3S,6S,14R,16R,20S,21S)-1 and (2R,3S,6S,14R,16R,20S,21S)-1 using the TDDFT method were performed (Supporting Information). The experimental ECD spectrum of 1 exhibited positive Cotton effects at 334 (Δε +4.2), 245 (Δε +11.7), and 221 (Δε +14.1) nm, which were similar to those in the calculated spectrum for the isomer with 2S,3S,6S,14R,16R,20S,21S configurations (Figure 4). Accordingly, the structure of 1 was unambiguously determined. Ervaoffine B (2) was assigned a molecular formula of C20H26N2O3 according to the 13C NMR and HRESIMS data. The NMR spectra of 2 and 1 (Table 1) exhibited similar resonances, except for the presence of a methylene (δC 53.8) rather than an oxymethine (C-3, δC 96.6), indicating that 2 was

Figure 2. Key NOESY correlations of 1 and 3. C

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Figure 3. ORTEP drawings of 1, 4, 8, 9, and 10.

Furthermore, X-ray diffraction analysis using Cu Kα radiation unambiguously established the absolute configuration of 4 with a Flack parameter of −0.13(17) (Figure 3). (7S)-3-Oxoibogaine hydroxyindolenine (5) was found to be an oxidized derivative of (7S)-ibogaine hydroxyindolenine (9)13 based on the HRESIMS and NMR analyses (Table 2). The HMBC correlations between H-5α/-5β and C-7 (δC 86.8) and between H-5α/-5β/-21 and C-3 (δC 175.0) suggested that the carbonyl group was located at C-3. The NOESY spectrum indicated that the configuration of 5 was consistent with that of (7S)-ibogaine hydroxyindolenine (9). In particular, the αorientation of 7-OH was deduced by NOESY correlations between 7-OH and H-5α/-6α/-16/-21. The agreement of the ECD curve of 5 with those of 9 and 10 allowed assignment of the absolute configuration of 5 as 7S,14R,16R,20S,21S (Figure 6). The molecular formula of ibogaine-5,6-dione (6) was determined to be C20H22N2O3 by its 13C NMR and HRESIMS data. Analysis of the NMR data of 6 (Table 2) and comparison with those of ibogaine (10) revealed that the C5/C6 tryptamine bridge in the known compound was replaced with two carbonyls, which was substantiated by their chemical shifts (δC 184.3 and 169.6) and the HMBC correlations between H21 (δH 4.04) and C-5 (δC 169.6)/C-2 (δC 155.4). Biosynthetically, 6 is expected to be derived from ibogaine (10) via oxidation at C-5 and C-6. Additionally, the ECD spectrum of 6 was in good agreement with the calculated spectrum for the (14R,16R,20S,21S)-6 diastereoisomer (Supporting Information), indicating that the absolute configuration of 6 was identical to that of 10. The NMR data of 7 (Table 2) were closely similar to those of voacristine (14),17 except that a methylene in 14 was replaced with a carbonyl moiety, as well as some differences at C-15 and C-21. The HMBC cross-peaks between H-3a/-6α/6β/-21 and C-5 (δC 178.2) suggested that the carbonyl group

Figure 4. Experimental ECD spectra of 1 and 8 and the calculated ECD spectra of (2S,3S,6S,14R,16R,20S,21S)-1 and (2R,3S,6S,14R,16R,20S,21S)-1.

Figure 5. Experimental ECD spectra of 3 and ervahainine A and the calculated ECD spectra of (3S,6R,7R,14R,16S,20S,21S)-3 and (3S,6R,7S,14R,16S,20S,21S)-3.

correlations between H-5α/-5β/-21/-15α/-15β/-17α/-17β and C-3 and between H-21/-17α/-17β/NH and C-2. The NOESY spectrum of 4 revealed that the relative configurations of C-14, -16, -20, and -21 in 4 were identical to those in ibogaine (10).12 D

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a

position

E

a b

α β

α β

α β α β

dd (13.0, 3.8) t (13.0) t (16.6) dd (16.6, 3.8)

br s t (12.0) dd (12.0, 2.1) d (9.5) pseudo t (12.9) d (12.9) t (7.3) m m m br s s

10.01 br s

2.36 1.91 1.02 2.87 1.45 2.26 0.91 1.28 1.15 1.78 4.08 3.76

7.06 7.06

6.79 br s

2.68 4.69 3.30 2.32

δH

δC

38.5 57.7 55.6

11.6 27.9

43.8 23.0

202.4 140.8 111.9 157.5 116.7 128.1 128.2 38.0 32.1

39.1

38.5

172.7 173.0

CH CH CH3

CH3 CH2

CH CH2

C C CH C CH CH C CH CH2

CH2

CH2

C C

mult

a b

α β

α β

α β α β

dd (13.1, 10.5) m t (7.4) m m m br s s

br s m m

6.27 br s

2.33 1.90 1.11 3.11 2.22 2.01 0.90 1.40 1.28 1.72 4.44 3.75

6.83 dd (8.3, 2.5) 7.26 d (8.3)

6.93 d (2.5)

3.13 4.04 dd (13.3, 4.5) 2.06 d (13.8) 1.21 dt (13.8, 4.5)

δH

5b δC

35.8 57.5 55.5

11.3 27.4

43.3 32.3

86.8 144.2 108.5 158.0 113.2 120.0 145.1 37.8 30.6

37.3

43.1

188.2 175.0

CH CH CH3

CH3 CH2

CH CH2

C C CH C CH CH C CH CH2

CH2

CH2

C C

mult

a b

α β

α β

a b

br s s

m dd (11.5, 5.9) t (7.3) m

br s

10.92 br s

2.17 1.93 1.47 3.26 2.36 1.69 0.94 1.56 1.47 1.91 4.04 3.76

6.80 dd (8.8, 2.3) 7.34 d (8.8)

7.66 d (2.3)

3.87 d (8.8) 3.29

δH

6c δC

38.7 55.4 55.7

11.9 28.3

37.7 31.1

110.5 127.4 103.3 156.6 113.4 112.8 130.6 28.6 29.7

184.3

169.6

155.4 49.7

Overlapped resonances are reported without designating multiplicity. bMeasured in DMSO-d6. cMeasured in CDCl3. dMeasured in methanol-d4.

20 21 OCH3 COOCH3 COOCH3 NH OH

18 19

16 17

7 8 9 10 11 12 13 14 15

6

5

2 3

4b

Table 2. 1H (600 MHz) and 13C (150 MHz) NMR Data for Compounds 4−7 (δ in ppm, J in Hz)a

CH CH CH3

CH3 CH2

CH CH2

C C CH C CH CH C CH CH2

C

C

C CH2

mult

α β

α β

α β

a b

dq (13.8, 1.8) dt (13.8, 2.0) d (6.2) m

3.71 s

1.73 m 4.62 d (2.5) 3.81 s

3.03 1.55 1.29 3.65

2.19 br s 1.79 m 1.86 m

6.73 dd (8.7, 2.3) 7.14 d (8.7)

6.98 d (2.3)

4.13 d (15.4) 3.62 d (15.4)

3.56 dt (11.9, 2.6) 3.14 d (11.9)

δH

7d δC

43.1 53.7 56.3 174.5 53.3

22.5 71.3

51.8 35.2

102.7 128.8 100.8 155.6 113.1 112.8 132.4 29.1 27.5

33.1

178.2

139.0 50.2

CH CH CH3 C CH3

CH3 CH

C CH2

C C CH C CH CH C CH CH2

CH2

C

C CH2

mult

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X-ray crystallographic analyses (Cu Kα) with indicative Flack parameters of −0.02(18) and 0.09(15), respectively. Interestingly, opposite absolute configurations of the C-2 spiro centers in 1 and 8 were observed. Comparison of the ECD curves of 1 and 8 (Figure 4) suggested that the Cotton effects between 200 and 280 nm were dominated by the C-2 absolute configuration. Similarly, comparison of the calculated ECD curve of 3 with that of its 7S-epimer indicated that the absolute configuration of the C-7 spiro carbon of oxindole alkaloids is the basis for the Cotton effects between 200 and 220 nm, a finding supported by the experimental ECD spectrum of ervahainine A, which is a related compound previously isolated from E. hainanensis11 (Figure 5). The ECD spectra of compounds 1−8 may be useful for the identification of the absolute configurations of ibogatype alkaloids. The discovery of these new iboga-type alkaloids expands the family of monoterpenoid indole alkaloids. Compounds 1−5, 8, and 9 were considered to be derived from the precursor ibogaine (10), which is a major constituent of Ervatamia plants (Scheme 1). Briefly, 10 may be oxidized to produce an intermediate, which may subsequently undergo a pinacol rearrangement to yield pseudoindoxyl and oxindole alkaloids with opposite spiro carbon configurations. If 2-OH is protonated and then eliminated, the intermediate may undergo a 1,2-alkyl shift to produce pseudoindoxyl alkaloids 1, 2, and 8. Alternatively, if the leaving group is 7-OH, oxindole alkaloids 3 and ervahainine A may be generated. Previously, 8 was believed

Figure 6. Experimental ECD spectra of 5, 9, and 10.

was located at C-5. Comparison of the chemical shift values of C-15 and C-21 in 7 with those of other 19-hydroxyiboga alkaloids indicated a 19R* configuration,20 which was confirmed by the NOESY cross-peaks between H-19 and H3b/-15β and between H-18 and H-21. The 14R,16S,19R,20S,21S configuration of 7 was defined based on the agreement between the experimental and calculated ECD curves (Supporting Information). ECD and X-ray crystallographic diffraction are accessible and powerful techniques to determine the absolute configuration of organic compounds. In this study, the absolute configurations of the known compounds iboluteine (8) and (7S)-ibogaine hydroxyindolenine (9) were determined for the first time using Scheme 1. Plausible Biosynthetic Pathways of 1−5, 8, and 9

F

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Kα radiation (λ = 1.541 78 Å). The structures were refined by fullmatrix least-squares on F2 using SHELXL-97 package software. The crystallographic data have been deposited at the Cambridge Crystallographic Data Centre as CCDC 987650 for 1 and CCDC 987651 for 4. Crystal data of 1: C20H24N2O3 (fw = 340.41); orthorhombic, space group P212121; a = 12.54080(10) Å, b = 12.59410(10) Å, c = 21.8561(3) Å, a = β = γ = 90.00°; V = 3451.95(6) Å3, T = 160(2) K, Z = 8, Dc = 1.310 mg/mm3, F(000) = 1456. A total of 12 377 reflections were collected in the range 4.05 ≤ θ ≤ 61.25, of which 4992 unique reflections with I > 2σ(I) were collected for the analysis. Final R = 0.0319, Rw = 0.0764, and S = 1.050. Flack parameter = 0.02(16). Crystal data of 4: C20H24N2O4 (fw = 356.41); monoclinic, space group P21; a = 9.5735(2) Å, b = 9.44199(19) Å, c = 10.4447(2) Å, a = γ = 90.00°, β = 109.162(2)°; V = 891.82(3) Å3, T = 173(2) K, Z = 2, Dc = 1.327 mg/mm3, F(000) = 380. A total of 10 716 reflections were collected in the range 4.89 ≤ θ ≤ 62.73, of which 2769 unique reflections with I > 2σ(I) were collected for the analysis. Final R = 0.0276 and Rw = 0.0724, and S = 1.040. Flack parameter = −0.13(17). Ervaoffine A (1): yellow crystals (MeOH/H2O), mp 195−196 °C; [α]25 D +125 (c 0.9, MeOH); UV (MeOH) λmax (log ε) 227 (4.12) nm; ECD (MeOH, Δε) λmax 358 (0), 334 (+4.2), 245 (+11.7), 221 (+14.1) nm; IR (KBr) νmax 3374, 2934, 1671, 1593, 1496, 1456, 1350, 1273, 1223, 1159, 1085, 1037, 1001, 936, 893, 818, 778, 704 cm−1; 1H and 13C NMR data, see Table 1; HRESIMS m/z 363.1680 [M + Na]+ (calcd for C20H24N2O3Na, 363.1679). Ervaoffine B (2): yellow oil; [α]25 D +27 (c 0.7, MeOH); UV (MeOH) λmax (log ε) 227 (4.29) nm; ECD (MeOH, Δε) λmax 338 (0), 309 (−0.6), 275 (0), 242 (+9.0), 214 (0), 210 (−1.6), 204 (0) nm; IR (KBr) νmax 3330, 2928, 2867, 1672, 1587, 1498, 1462, 1385, 1276, 1224, 1111, 1075, 1029, 970, 823, 786, 729, 540 cm−1; 1H and 13 C NMR data, see Table 1; HRESIMS m/z 343.2019 [M + H]+ (calcd for C20H27N2O3, 343.2016). Ervaoffine C (3): yellow oil; [α]25 D +125 (c 1.0, MeOH); UV (MeOH) λmax (log ε) 205 (4.17), 259 (3.71) nm; ECD (MeOH, Δε) λmax 330 (0), 305 (−4.8), 260 (−6.9), 249 (0), 233 (+20.2), 206 (+18.5) nm; IR (KBr) νmax 3430, 2952, 2933, 2874, 1694, 1594, 1483, 1456, 1307, 1263, 1205, 1158, 1083, 1034, 994, 890, 814, 778, 637 cm−1; 1H and 13C NMR data, see Table 1; HRESIMS m/z 341.1861 [M + H]+ (calcd for C20H25N2O3, 341.1860). Ervaoffine D (4): colorless crystals (MeOH), mp 208−209 °C; [α]25 D −22 (c 1.4, MeOH); UV (MeOH) λmax (log ε) 223 (4.19), 300 (2.95) nm; ECD (MeOH, Δε) λmax 317 (+3.3), 274 (0), 225 (−23.8), 209 (0) nm; IR (KBr) νmax 3453, 3256, 2961, 1698, 1664, 1649, 1577, 1490, 1456, 1417, 1360, 1304, 1279, 1234, 1171, 1029, 1002, 825 cm−1; 1H and 13C NMR data, see Table 2; HRESIMS m/z 379.1629 [M + Na]+ (calcd for C20H24N2O4Na, 379.1628). 7S-3-Oxoibogaine hydroxyindolenine (5): yellow oil; [α]25 D +32 (c 1.3, MeOH); UV (MeOH) λmax (log ε) 201 (4.08), 226 (3.98), 283 (3.53) nm; ECD (MeOH, Δε) λmax 392 (0), 315 (+1.0), 306 (0), 289 (−4.4), 279 (0), 258 (18.6), 239 (0), 230 (−9.0), 217 (0) nm; IR (KBr) νmax 3393, 2957, 1648, 1559, 1474, 1429, 1359, 1272, 1165, 1104, 1027, 977, 824 cm−1; 1H and 13C NMR data, see Table 2; HRESIMS m/z 341.1859 [M + H]+ (calcd for C20H25N2O3, 341.1860). Ibogaine-5,6-dione (6): yellow oil; [α]25 D +201 (c 1.1, MeOH); UV (MeOH) λmax (log ε) 211 (4.12), 258 (3.81), 285 (3.60), 334 (3.42) nm; ECD (MeOH, Δε) λmax 400 (0), 341 (+5.7), 263 (+1.6), 253 (0), 245 (−1.1), 238 (0), 221 (+4.1), 209 (0) nm; IR (KBr) νmax 3197, 2930, 1636, 1466, 1272, 1214, 1145, 1108, 1034, 805 cm−1; 1H and 13 C NMR data, see Table 2; HRESIMS m/z 361.1524 [M + Na]+ (calcd for C20H22N2O3Na, 361.1523). 19-epi-5-Oxovoacristine (7): yellow oil; [α]25 D +8 (c 1.1, MeOH); UV (MeOH) λmax (log ε) 208 (4.24), 279 (3.64) nm; ECD (MeOH, Δε) λmax 306 (+2.4), 295 (0), 276 (−6.1), 257 (0), 252 (+0.9), 248 (0), 241 (−1.3), 236 (0), 226 (+5.2), 207 (+13.9) nm; IR (KBr) νmax 3408, 2936, 2879, 1733, 1639, 1488, 1456, 1384, 1313, 1246, 1217, 1149, 1082, 1027, 934, 803, 744 cm−1; 1H and 13C NMR data, see Table 2; HRESIMS m/z 399.1915 [M + H]+ (calcd for C22H27N2O5, 399.1914).

to be derived from 9 via a Wagner−Meerwein rearrangement.8,13,21 However, compounds 1 and 2, with spiro carbon configurations that are opposite to those of other members of this class, would not be obtained through this reaction mechanism.8,13 The assumption of a pinacol rearrangement reasonably accounts for the co-occurrence of iboga-type alkaloids with opposite spiro carbon configurations in Ervatamia species. Additionally, ibogaine (10) may undergo oxidative cleavage of the indole moiety to afford ketoamide 4.22 Hydroxyindolenine derivatives 5 and 9 may also be formed by oxidation of 10.13

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EXPERIMENTAL SECTION

General Experimental Procedures. Optical rotations were determined using a JASCO P-1020 digital polarimeter at 25 °C. Melting points were measured using an X-5 micro melting point apparatus (uncorrected). UV data were recorded using a JASCO V550 UV/vis spectrophotometer. ECD data were measured with a JASCO J-810 spectrometer. IR spectra were obtained by a JASCO FT/ IR-480 Plus infrared spectrometer using KBr pellets. HRESIMS were recorded on an Agilent 6210 ESI-TOF mass spectrometer. NMR spectra were acquired using Bruker AV-400 or AV-600 spectrometers. Single-crystal data were performed using an Agilent Gemini S Ultra diffractometer and Cu Kα radiation (λ = 1.541 78 Å). Column chromatography (CC) utilized Sephadex LH-20 (Pharmacia Biotech AB, Uppsala, Sweden), silica gel (200−300 mesh; Qingdao Marine Chemical Inc., Qingdao, P. R. China), and ODS (YMC, Kyoto, Japan). Preparative HPLC (pHPLC) was carried out on an Agilent 1260 system (G1310B Iso pump and G1365D MWD VL detector) with a CAPCELL PAK MGII C18 reversed-phase column (20 × 250 mm, 5 μm, Shiseido Fine Chemicals Ltd., Japan). Plant Material. The twigs and leaves of E. of f icinalis were collected in Diaoluo Mountain, Hainan Province, P. R. China, in November 2010 and identified by Dr. Shi-Man Huang, Hainan University, Haikou, P. R. China. A voucher specimen (No. 2010113001) was deposited at the herbarium of the College of Pharmacy, Jinan University, Guangzhou, P. R. China. Extraction and Isolation. The twigs and leaves of E. of f icinalis (10.0 kg) were percolated at room temperature with 95% EtOH (24 h × 5) to afford a residue (0.5 kg), which was treated with 2.8 L of 0.5% HCl and then extracted with CHCl3. The acidic aqueous phase was basified with ammonia to pH 9−10 and partitioned with CHCl3 to afford the crude alkaloids (39.1 g). The crude alkaloid extract was subjected to silica gel CC and eluted with CHCl3−MeOH (10:0 to 0:10, v/v) to obtain 15 fractions (Fr. A−O). Fr. B was separated by repeated Sephadex LH-20 CC eluting with CHCl3−MeOH (1:1, v/v) to afford 3 (5.2 mg), 12 (8.6 mg), and 13 (8.4 mg). Fr. C was chromatographed on an ODS column with MeOH−H2O (3:7 to 10:0, v/v), followed by pHPLC with MeOH−H2O−Et2NH (6.5:3.5:0.001, v/v), to afford 1 (7.0 mg, tR 39.6 min), 5 (3.1 mg, tR 33.7 min), 14 (10.2 mg, tR 44.5 min), and 15 (7.1 mg, tR 61.1 min). Fr. D was successively separated on Sephadex LH-20 CC (MeOH) and purified by pHPLC with MeOH−H2O−Et2NH (5:5:0.001, v/v) to afford 6 (5.3 mg, tR 13.8 min). Fr. E was purified using Sephadex LH-20 CC with CHCl3−MeOH (1:1, v/v) to yield 7 (1.9 mg). Fr. F was separated by ODS CC with MeOH−H2O (1:9 to 10:0, v/v), then subjected to Sephadex LH-20 CC (MeOH), to yield 4 (8.2 mg). Fr. G was purified using Sephadex LH-20 CC with CHCl3−MeOH (1:1, v/ v) to yield 10 (420.8 mg). Fr. I was purified using Sephadex LH-20 CC (MeOH) to afford 8 (10.3 mg). Fr. K was subjected to pHPLC with MeCN−H2O−Et2NH (6:4:0.001, v/v) to afford 9 (5.2 mg, tR 21.8 min), 11 (6.8 mg, tR 53.5 min), and 16 (4.9 mg, tR 22.9 min). Fr. L was subjected to Sephadex LH-20 CC with CHCl3−MeOH (1:1, v/ v) to yield 2 (2.3 mg). Fr. M was successively purified using Sephadex LH-20 CC (MeOH) and pHPLC with MeOH−H2O−Et2NH (8:2:0.001, v/v) to afford 17 (8.0 mg, tR 12.5 min). X-ray Crystallographic Analysis of 1 and 4. All X-ray data were collected on an Agilent Gemini S Ultra CCD diffractometer with Cu G

dx.doi.org/10.1021/np500240b | J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products

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Computation Section. Conformational searches were carried out by Sybyl 8.1 software using the MMFF94S force field with an energy cutoff of 10 kcal/mol. The single-crystal X-ray diffraction data and the obtained conformers were used for geometry reoptimizatons at the B3LYP/6-31+G(d) level with a PCM solvent model for methanol in the Gaussian 09 software.23 TDDFT ECD calculations for the optimized conformers were performed at the B3LYP/6-31+G(d) level in vacuo. The overall ECD curves of all the compounds were weighted by Boltzmann distribution after UV correction. The ECD curves were produced by SpecDis 1.6 software.24

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ASSOCIATED CONTENT

S Supporting Information *

UV, IR, NMR, MS, and ECD spectra of compounds 1−7; CIFs of compounds 1, 4, 8, 9, and 10. This material is available free of charge via the Internet at http://pubs.acs.org.

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AUTHOR INFORMATION

Corresponding Authors

*E-mail: [email protected] (L. Wang). *E-mail: [email protected] (X.-Q. Zhang). *Tel: +86-20-85220936. Fax: +86-20-85221559. E-mail: [email protected] (W.-C. Ye). Author Contributions §

B.-Q. Tang and W.-J. Wang contributed equally to this work.

Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS This research was financially supported by the National Natural Science Foundation of China (81373935, 81202428), the Ministry of Science and Technology of China (2013BAI11B05, 2013DFM30080, 2012ZX09103201-056), the Fundamental Research Funds for the Central Universities (21612203), and the Program for New Century Excellent Talents in University (NCET-12-0676).

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REFERENCES

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