HETEROCYCLES

Note | Regular issue | Vol. 78, No. 8, 2009, pp. 2053-2059
Received, 31st January, 2009, Accepted, 14th April, 2009, Published online, 16th April, 2009.
DOI: 10.3987/COM-09-11671

■ New Alkaloids from Conchocarpus gaudichaudianus

Lucia E. Ranieri Cortez, Diógenes A. Garcia Cortez,* João B. Fernandes, Paulo C. Vieira, Antonio G. Ferreira, and M. Fátima das G. F. da Silva

Department of Pharmacy and Pharmacology, State University of Maringá, Av. Colombo, 5790
CEP 87020-090, Maringá, Paraná, Brazil

Abstract

The methanol extract from stems of Conchocarpus gaudichaudianus yielded four new alkaloids: 3-(2-(7,7-dimethyl-3,7-dihydropyrano[3,2-e]indol-1-yl)ethyl)quinazoline-2,4(1H,3H)-dione (1), 3-(2-(7,7-dimethyl-3,7-dihydropyrano[3,2-e]indol-1-yl)ethyl)-1-hydroxyquinazoline-2,4(1H,3H)-dione (2), 3-(2-(7,7-dimethyl-3,7-dihydropyrano[3,2-e]indol-1-yl)ethyl-1-methylquinazoline-2,4(1H,3H)-dione (3), (E)-3-(6,7-dihydroxy-3,7-dimethyloct-2-enyl)-4-methoxy-1-methylquinolin-2(1H)-one (6), in addition to the known alkaloids N-trans-coumaroyltyramine and N-trans-feruloyltyramine. The structures of these compounds were identified by IR, UV, 1H and 13C NMR spectroscopy, gHSQC and gHMBC spectral analysis and comparison of data from the literature.

Conchocarpus gaudichaudianus, a tree, is used by the aboriginal natives in northern Brazil.1 Of the 45 species in the genus Conchocarpus, only C. heterophyllus, C. inopinatus and C. paniculata have been investigated by spectroscopic analysis. Extracts from leaves of C. heterophyllus and the dichloromethane fraction of C. inopinatus show in vitro trypanocidal activity2,3 and C. paniculata exhibits molluscicidal activity.4
Indolopyridoquinazoline type alkaloids, like rutaecarpine isolated from Evodia rutaecarpa, shows a variety of pharmacological effects including anti-inflammatory5 antithrombotic and vasorelaxant effects.6
We report studies on the stems of C. gaudichaudianus, which afforded three new indoloquinazolone alkaloids (1, 2 and 3) and one 2-quinolone alkaloid (6).
The methanol extract from stems of Conchocarpus gaudichaudianus yielded six alkaloids 1 - 6, including the known N-trans-coumaroyltyramine (4) and N-trans-feruloyltyramine (5).

The molecular formula of compound (1) was established to be C23H21N3O3 by analysis of its HR-MS (ESI) [m/z: 410.1519 (M+Na)+] and NMR data (Table 1). IR spectrum showed absorption of a tertiary amide function at 1660 cm-1. The UV results (see Experimental) were also in agreement with a highly aromatic conjugation system. The 1H NMR spectrum (300 MHz) of 1 showed two symmetrical triplets at δ 4.13 and δ 3.02, corresponding to protons H-7 and H-8, an NH indolic group at δ 10.44, and six aromatic hydrogens at δ 6.56 (d, J = 8.7 Hz, H-11); 7.08 (d, J = 8.7 Hz, H-12) and at δ 7.19 (d, J = 8.1 Hz, H-1); 7.65 (td, J = 8.1 and 1.3 Hz, H-2); 7.21 (t, J = 8.1 Hz, H-3); 7.95 (dd, J = 8.1 and 1.3 Hz, H-4). The presence of a 2,2-dimethylchromene ring was evidenced by a methyl singlet at δ 1.35 integrating for six hydrogens and two doublets H-9’ and H-10’ at δ 5.72 (d, J = 9.8 Hz, H-10’) and 7.28 (d, J = 9.8 Hz, H-9’).
Full assignments of the hydrogens and protonated carbons of alkaloid 1 were attained by 1H-1H gCOSY and 1H-13C heteronuclear correlated 2D NMR spectra (gHMQC). Also, by a hydrogen detected multiple bond 1H-13C correlation spectrum (gHMBC) all of the quaternary carbon signals were assigned (Table 1). Based on the above data and by comparison with data from goshuyuamide-II8 (3a) structure 1 was assigned to 3-(2-(7,7-dimethyl-3,7-dihydropyrano[3,2-e]indol-1-yl)ethyl)-1-hydroxyquinazoline- 2,4(1H,3H)-dione.
Compound (2) was obtained as a yellow oil. The UV spectrum of 2 showed it to be related to 1. In the IR spectrum, the absorption at 3397 cm-1 indicated the presence of a hydroxyl group. Evidence for a hydroxyl at N-14 was given by the mass spectrum, which exhibited a fragment at m/z 226 [m/z 404-178 (C15H16NO)] and 179 [m/z 404-225(C8H7N2O3)] and by correlation for a HN-13 at δ10.80 d, J=2.40 Hz with H13a obtained by 1H-1H gCOSY. The new alkaloid was therefore identified as 3-(2-(7,7-dimethyl-3,7-dihydropyrano[3,2-e]indol-1-yl)ethyl) -1-hydroxyquinazoline-2,4(1H,3H)-dione .
Compound 3 differed from 1 by the presence of a methyl group at N-14. The correlation for methyl protons with C-14a and C-1a, which indicated the location of the methyl group shown by the gHSQC and gHMBC experiments and the signal at δ10.76 d, J=2.40 Hz, HN-13 and obtained by 1H-1H gCOSY. Thus, the structure of the new alkaloid was characterized as 3-(2-(7,7-dimethyl-3,7-dihydropyrano[3,2-e]indol- 1-yl)ethyl) -1-methylquinazoline-2,4(1H,3H)-dione.
The known N-trans-coumaroyltyramine (4) and N-trans-feruloyltyramine (5) were also isolated and identified by 1H and 13C NMR, mass spectrometry, and comparison of published spectral data.9,10,11
Compound 6 was obtained as an amorphous yellow solid. The [α]D 0° obtained indicated that this compound is a raceme mixture. Its UV and IR spectra showed the characteristic absorption of a 2-quinolones9. The ESI-MSMS mass spectrum indicated the [M+H]+ at m/z 360 (base peak), and the characteristic fragment ion of the 4-methoxy-1-methyl-2-quinolone alkaloids the [M+H]+ at m/z 203.12 The 1H NMR spectrum revealed the presence of two signals at δ 3.94 s and 3.69 s due to methoxyl and N-methyl groups (Table 2), four aromatics at δ 7.84, (dd, J=7.8 and 1.5 Hz), 7.27, (td, J=7.8 and 0.9 Hz), 7.55, (td, J=7.8 and 1.5 Hz) and 7.38, (dd, J=7.8 and 0.9 Hz). There were two methyl singlets at δ 1.07, thus placing an isopropyl group at the end of the side chain, and one signal at δ 1.80, br s, H-10’. The observed signals for a hydroxyl-tertiary carbon at δ 72.8 and three methyl singlets at δH 1.07 (s, 6H), 1.80 (brs, 3H), a signal at δ 5.30 (td, J=6.6 and 1.1 Hz), 3.37 (d, J=6.6 Hz), in the 13C (Table 2) and 1H NMR, and complete assignment of chemical shifts for all carbons, confirmed the presence of a 3’,7’-dimethyl-6’,7’dihydroxy-2’-octenyl chain. Using the prenylquinoline alkaloid11 as a model, the structure of this new alkaloid was established as (E)-3-(6,7-dihydroxy-3,7-dimethyloct-2-enyl)-4- methoxy-1-methylquinolin-2(1H)-one.

EXPERIMENTAL

General
The NMR spectra were obtained in BRUKER DRX400 (9.4 T) and VARIAN GEMINI300 (7.05 T) spectrophotometers, using TMS as the internal standard. For gHMBC the coupling constants were optimized for 4, 6, 8 and 12 Hz. IR: film NaCl plates; High-resolution ESI-MS were recorded on a Micromass Q-Tof mass spectrometer, ESI-MSMS: low resolution on a triple quadrupole were recorded on a Micromass Quattro LC instrument equipped with a “Z-spray” ion source, and EI-MS on a CG/EM-SHIMADZU QP 2000 A. CC: silica gel 60 (70-230 and 230-400 mesh); TLC: silica gel plates F254 (0.25 mm in thickness).

Plant Material
The aerial parts of Conchocarpus gaudichaudianus subsp. bahiensis were collected in Itacaré, Bahia, Brazil, in February 1993. The specimen was identified by Dr. Jose Pirani from the Department of Botany of the University of São Paulo, Brazil, and the voucher herbarium specimen was deposited in this University.

Extraction and Isolation
The stems (200 g) from Conchocarpus gaudichaudianus were exhaustively extracted with MeOH at rt, and the extract was evaporated in vacuo to yield 3.8 g. The extract was submitted to vacuum chromatography over a silica gel support and eluted with n-hexane, n-hexane-CHCl3 (1:1), CHCl3, CHCl3-EtOAc (1:1), EtOAc, and MeOH, giving 11 fractions. Fraction 8 (480 mg) was chromatographed over a gel filtration Sephadex LH-20 column using MeOH and MeOH-CHCl3 (1:1) successively to give 1 (1.0 mg), 2 (4.8 mg) and 3 (1.2 mg). Fraction 9 (520 mg) was twice chromatographed (Sephadex LH-20 MeOH and MeOH-CHCl3 (1:1); then silica gel, hexane-CH2Cl2-EtOAc-MeOH gradient) to give compounds 1 (2.8 mg), 4 (5.3 mg) and 5 (6.8 mg). Fraction 2 (1.82 g) was chromatographed on silica gel columns (hexane-CH2Cl2-EtOAc-MeOH gradient solvents) to yield Fr 89 (540 mg), and was purified by Sephadex LH-20 eluting with MeOH and MeOH-CHCl3 (1:1) respectively to yield the majority compound 6 (340 mg).

3-(2-(7,7-Dimethyl-3,7-dihydropyrano[3,2-e]indol-1-yl)ethyl)quinazoline-2,4(1H,3H)-dione (1).
Amorphous yellow solid; [UV MeOH λmax nm (log ε) ]: 242 (3.97), 312 (3.78). IRνmax (CHCl3) cm-1: 3341, 1660,1454 and 964. 1H NMR and 13C NMR: see Table 1. ESI-MSMS, m/z (rel. int.): 386 [M-H]- (20), 161(100). HR-MS (ESI) [M+Na]+ Found m/z: 410.1519 (calc. for C23H21N3O3Na: 410.1481).

3-(2-(7,7-Dimethyl-3,7-dihydropyrano[3,2-e]indol-1-yl)ethyl)-1-hydroxyquinazoline-2,4(1H,3H)-dio- ne (2).
Amorphous yellow solid. [UV MeOH λmax nm (log ε) ]: 242 (4.02), 312 (3.78). IRνmax (CHCl3) cm-1: 3397, 1645, 1450, and 982. 1H NMR and 13C NMR: see Table 1. ESI-MSMS, m/z (rel. int.): 404 [M+H]+ (100), 389 (8), 336 (4), 226 (19), 179 (8). HR-MS (ESI) [M-H]- at m/z: 402.1431 (calc. for C23H20N3O4 402.1453).

3-(2-(7,7-Dimethyl-3,7-dihydropyrano[3,2-e]indol-1-yl)ethyl)-1-methylquinazoline-2,4(1H,3H)-dione (3).
Amorphous yellow solid. [UV MeOH λmax nm (log ε) ]: 244 (4.10), 313 (3.92). IRνmax (CHCl3) cm-1: 3367, 2923, 1663, 1466 and 1022. 1H NMR and 13C NMR: see Table 1. ESI-MSMS, m/z (rel. int.): 400 [M-H]- (20), 175(100).
(E)-3-(6,7-Dihydroxy-3,7-dimethyloct-2-enyl)-4-methoxy-1-methylquinolin-2(1H)-one (6).
Amorphous yellow solid. [α]D 0° (CHCl3, c 0.01) [UV MeOH λmax nm (log ε) ]: 244 (4.21), 327 (3.83). IRνmax (CHCl3) cm-1: 3425, 2929, 1634, 1103 and 756. 1H NMR and 13C NMR: see Table 2. ESI-MSMS, m/z (rel. int.): 360 [M+H]+ (100), 341 (20), 203 (65). HR-MS (ESI) [M-H]- at m/z: 358.2032 (calc. for C21H28NO4 358.2019);

ACKNOWLEDGMENT
The authors thank Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP), and Coordenação de Aperfeiçoamento de Pessoal de Ensino Superior (CAPES) for financial suporte.

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