Furanoflavones and other chemical constituents of Lonchocarpus obtusos

ARTICLE

Furanoflavones and other chemical constituents of Lonchocarpus obtusos

Maria Gilvânia B. Cavalcante^I; Rochanne M. Silva^I; Paulo N. Bandeira^I; Hélcio S. dos Santos^I; Otília Deusdênia L. Pessoa^II; Raimundo Braz-Filho^II; Maria Rose Jane R. Albuquerque^I,*

^ICoordenação de Química, Centro de Ciências Exatas e Tecnologia, Universidade Estadual Vale do Acaraú, CP D-3, 62040-340 Sobral-CE, Brazil

^IIDepartamento de Química Orgânica e Inorgânica, Universidade Federal do Ceará, 60021-940 Fortaleza-CE, Brazil

ABSTRACT

Two new furanoflavonoids, 5-hydroxy-2’’-isopropenyl-3-methoxyfurane-(2",3":7,8)-flavone and 5-hydroxy-2’’-(1-hydroxy-1-methylethyl)-3-methoxyfurane-(2",3":7,8)-flavone, along with thirteen other known compounds, were isolated from the root barks of Lonchocarpus obtusus (Leguminosae). The structures of all compounds were determined by spectrometric methods such as 1D and 2D NMR (COSY, HSQC and HMBC) and HREISMS beside comparison with spectral data for similar compounds.

Keywords: Lonchocarpus obtusos, Leguminosae, furanoflavonoids, flavones

RESUMO

Dois novos furanoflavonoides, 5-hidróxi-2’’-isopropenil-3-metoxifurano-(2",3":7,8)-flavona e 5-hidróxi-2’’-(1-hidróxi-1-metiletil)-3-metoxifurano-(2",3":7,8)-flavona, juntamente com treze compostos conhecidos, foram isolados das cascas das raízes de Lonchocarpus obtusos (Leguminosae). As estruturas de todos os compostos isolados foram determinadas usando métodos espectrométricos tais como RMN 1D e 2D (COSY, HSQC e HMBC) e EMAR, além da comparação com dados espectrais descritos na literatura para compostos de estruturas semelhantes.

Introduction

The flavonoids consist of a large group of low-molecular weight phenol compounds whose pharmacological and medicinal properties have been extensively investigated.^1,2 Several activities have been reported for these compounds, especially as antioxidant,³ antiproliferative,⁴ anti-inflammatory,⁵ analgesic⁶ and cytotoxic.⁷

The genus Lonchocarpus (Leguminosae, Papilionoideae), comprises approximately 135 species, twenty-four of which are native to Brazil.⁷Lonchocarpus is known as a prolific source of flavonoids (aurones, chalcones, flavones, flavonols, flavans, flavanones and stilbenes)⁸ has been the subject of incessant investigations searching for compounds of chemical and biological interest. Activities such as gastroprotective,⁹ trypanocidal and antimalarial,¹⁰ cytotoxic,¹¹ anti-inflammatory and antimicrobial¹² have been reported for compounds isolated from Lonchocarpus.

As part of an ongoing research for new secondary metabolites from plants of the northeastern Brazil flora we have investigated the extracts of Lonchocarpus obtusus. In the present work the isolation and characterization of two new flavonoids (6 and 7), from the roots of L. obtusos are described. Additionally, thirteen known flavonoids: derriobtusone A (1),¹³ obovatin (2),¹⁴ 3,6-dimethoxy-2",2"-dimethylchromene-(3",4":7,8)-flavone (3),¹⁵ derriobtusone B (4),¹³ (2",3":7,8)-furanoflavone (5),¹⁶ pongaglabol (8)¹⁷ and its methyl ether (9),¹⁷ lonchocarpin (10),¹⁸ 5-hydroxy-2",2"-dimethylchromene-(3",4":6:7)-flavone (11),¹⁹ medicarpin (12),²¹ maackiain (13),²² butein (14)²³and cycloeucalenol (15)²⁰ were isolated.

Results and Discussion

Compound 6 was isolated as a yellow resin. Its IR spectrum exhibited absorption bands at 3420 (hydroxyl), 1658 (carbonyl) and 1599 cm^-1 (benzene ring) compatible with a flavonoid skeleton. Its HRESIMS spectrum, in the positive mode, revealed a peak at m/z 349.0941 [M + H]⁺ indicating the molecular formula of C₂₁H₁₆O₅, corresponding to fourteen degrees of unsaturation. The ¹H NMR spectrum showed a signal at δ 12.64 (s) for a chelated hydroxyl group, two multiplets at δ 8.14 (m, H-2’/H-6’) and 7.57 (m, H-3’/H-4’/H-5’) appropriate for a monosubstituted benzene ring, two singlets at δ 6.88 (s, H-6) and 6.84 (s, H-3"), and a signal for one methoxyl group at δ 3.90 (3-OMe). In addition, the ¹H NMR spectrum showed characteristic signals for an isopropenyl moiety at δ 2.15 (s, 3H-5"), 5.79 (s, H-6a") and 5.21 (s, H-6b"). Apart from the signals typical of a flavonoidic skeleton, the ¹³C NMR spectrum of 6 showed the signals at δ 132.3 (C-4"), 113.8 (C-6") and 19.4 (C-5"), which were inferred to the isopropenyl moiety already suggested by the ¹H NMR data. The signals at δ 156.8 (C-2") and 99.5 (C-3"), were associated with a furan ring while a signal at δ60.6 with the methoxyl group located on C ring. The NMR spectral data (Table 1) of 6 combined with the molecular formula suggested a methoxy-furaneflavone.

In the HMBC spectrum, the correlations of the hydrogens at δ 6.84 (H-3"), 5.79/5.21 (2H-6") and 2.15 (3H-5") with the carbon at δ 156.8 (C-2") were in agreement with the presence of the isopropenyl moiety at C-2", while the long range correlation between the proton signal at δ 6.84 (H-3") with the carbon at δ 149.2 (C-9) confirmed the location of the furan ring at the C-7/C-8 position. In order to attend to the feature of a monosubstituted ring B in the structure of 6, the methoxyl group was located in the C ring at the C-3 position in accordance with the carbon chemical shift at δ 140.7 (C-3) and the correlation between the signal at δ 3.90 (O-Me) with that carbon. Based on all spectroscopic evidences the structure of 6 was established as the 5-hydroxy-2’’-isopropenyl-3-methoxyfurane-(2",3":7,8)-flavone.

Compound 7 was also isolated as a yellow resin. Its IR spectrum exhibited absorption bands at 3547 (hydroxyl), 1656 (carbonyl) and 1602 (benzene ring) compatible with the structure of a flavonoid. Its HRESIMS spectrum, in the positive mode, revealed the molecular peak at m/z 367.1064 [M + H]⁺ indicating the molecular formula of C₂₁H₁₈O₆, corresponding to thirteen degrees of unsaturation. A detailed analysis of the 1D and 2D NMR spectral data of 7 revealed high structural similarity to flavonoid 6. The main observed difference was related to an isopropyl alcohol moiety supported by the methyl signal at δ 1.89 (s), integrating to 6 hydrogen atoms (3H-5’’/3-H-6’’) and the carbon signals at δ 30.0 (C-5" and C-6") and δ 68.9 (C-4") (Table 1). The isopropyl alcohol moiety was positioned at C-2’’ through the HMBC correlations between the methyl signal at 1.89 (3H-5"/3-H-6") and the carbon signal at 166.0 (C-2’’), in accordance with the hydration of the propenyl side chain of 6. Additional long range correlations can be seen in Figure 1. Based on the aforementioned data the structure of 7 was established as 5-hydroxy-2’’-(1-hydroxy-1-methylethyl)-3-methoxyfurane-(2",3":7,8)-flavone, a new hydrated derivative of flavone 6.

Previous phytochemicals studies have demonstrated that Lonchocarpus is a prolific source of flavonoids with furan or pyran substituent at ring A in an angular or linear position. Flavonoids 6 and 7 were previously unknown, while 8, 9 and 14 are been reported for the first time in this genus.

Experimental

General experimental procedures

Melting points were measured on a digital Mettler Toledo FP90 apparatus and are uncorrected. IR spectra were recorded using a Perkin-Elmer FT-IR 1000 spectrometer. Electrospray ionization-high resolution mass spectra were run on a quadrupole LCMS-IT-TOF (Shimadzu) spectrometer equipped with an electrospray ionization source. 1D and 2D NMR experiments were performed on a Bruker Avance DRX-500 spectrometer (¹H 500 MHz, ¹³C 125 MHz) equipped with a 5 mm inverse detection z-gradient probe. ¹H and ¹³C NMR spectra were measured at 27 ºC using CDCl₃ and pyridine as the solvent (0.6 mL). Open column chromatography was run using silica gel 60 (70-230 mesh, Vetec), while flash column chromatography was run on silica gel (230-400 mesh, Merck). TLC was performed on precoated silica gel polyester sheets (Kieselgel 60 F₂₅₄, 0.20 mm, Merck) by detection with a spraying reagent (vanillin/perchloric acid/EtOH solution) followed by heating at 100 ºC.

Plant material

The roots of Lonchocarpus obtusus were collected, in April 2007, from Meruoca County (Ceará State, Brazil). The plant authentication was performed by Professor Afrânio Gomes Fernandes and a voucher specimen (No. 39550) has been deposited at the Herbário Prisco Bezerra (EAC) of the Departamento de Biologia, Universidade Federal do Ceará.

Extraction and isolation

Dried roots bark (720 g) and wood (750 g) of L. obtusus were separately, powdered and then extracted at room temperature with n-hexane (3 × 2.0 L) followed by ethanol (3 × 2.0 L). The solvents were distillated under reduced pressure to afford the crude n-hexane (root bark: 37.9 g; root wood: 56.1 g) and EtOH (root bark: 3.9 g; root wood: 20.5 g) extracts. During the distillation process of the n-hexane extract of the root bark was observed a yellowish precipitate which was filtrated and purified by crystallization in acetone to give 1 (21.3 mg). The n-hexane extract from the root bark (36.0 g) was subjected to silica gel column chromatography (36.6 g) by elution with n-hexane (800 mL), EtOAc (200 mL), CHCl₃ (500 mL) and MeOH (100 mL), yielding the following fractions, after solvent evaporation: n-hexane (9.8 g), CHCl₃ (23.0 g), EtOAc (643.0 mg) and MeOH (812.0 mg). The n-hexane fraction (9.8 g) was chomatographed over silica gel, eluting with n-hexane, n-hexane/EtOAc (9:1, 8:2, 7:3, 6:4, 1:1) and EtOAc to afford 7 fractions of 100 mL each (F_A1-F_A7). F_A3 (796.3 mg, n-hexane/EtOAc 8:2 mixture) was subjected to a silica gel flash column chromatography using n-hexane/EtOAc 9:1 mixture to yield 2 (5.6 mg).

The CHCl₃ fraction (23.0 g) was chromatographed over silica gel (81.3 g), eluting with n-hexane/CHCl₃ (6:4, 4:6, 2:8), CHCl₃, EtOAc and MeOH to afford 6 fractions of 200 mL each (F_B1-F_B6). F_B5 (7.1 g, EtOAc) was subjected to silica gel column chromatography using n-hexane/CHCl₃ (1:1, 2.5:7.5), CHCl₃, CHCl₃/EtOAc (9:1, 8:2, 7:3), EtOAc and MeOH, to afford 9 fractions of 125 mL each (F_C1-F_C9). F_C1 (3.0 g, n-hexane/CHCl₃ 1:1) was chromatographed over a silica gel column, using n-hexane with increasing amounts of CHCl₃ (9:1 to 1:9) as eluent to yield 102 fractions (10 mL each), that after TLC analysis yielded 9 fractions (F_D1-F_D9). F_D4 (1.4 g, n-hexane/CHCl₃ 1:1 and 4:6) was subjected to silica gel column chromatography using n-hexane/EtOAc gradient (9.5:0.5 to 0.5:9.5), EtOAc and MeOH to give 115 fractions, which after TLC analysis were combined into 7 fractions (F_E1-F_E7). F_E4 (60.6 mg, n-hexane/EtOAc 9.5:0.5 to 8:2) was rechromatographed using the same solvent system to afford 3 (34.1 mg) and 4 (16.4 mg). F_C2 (2.1 g, n-hexane/CHCl₃ 2.5:7.5) was subjected to silica gel column chromatography using n-hexane/EtOAc gradient (8:2, 7.5:2.5, 7:3) EtOAc, and MeOH. Fractions n-hexane/EtOAc 8:2 and 7.5:2.5 (364 mg) were combined and subjected to repeated silica gel column chromatography using n-hexane/EtOAC gradient to yield 5 (24.3 mg). F_C3 (1.1 g, CHCl₃) was chromatographed over a silica gel column, eluting with n-hexane/EtOAc gradient (8:2 to 2:8), EtOAc and MeOH, to give 119 fractions which after TLC analysis were combined into eight fractions (F_F1-F_F8). F_F3 (98.4 mg, n-hexane/EtOAc 8:2) was subjected to successive purifications using Sephadex LH-20 (CH₂Cl₂/MeOH 1:1) to yield 6 (6.5 mg) and 7 (3.0 mg). Chromatography of subfraction F_F7 (142.5 mg, EtOAc) over silica gel eluting with CHCl₃ yielded 8 (52.6 mg). The EtOH extract from the root bark (76.1 g) was subjected to silica gel column chromatography eluting with CH₂Cl₂, CH₂Cl₂/EtOAc (7.5:2.5, 1:1), EtOAc, EtOAc/MeOH (7.5:2.5) and MeOH (1000 mL of the each solvent), to afford the following fractions: CH₂Cl₂ I (2.0 g), CH₂Cl₂ II (8.8 g), CH₂Cl₂/EtOAc 7.5:2.5 (12.7 g), CH₂Cl₂/EtOAc 1:1 (4.2 g), EtOAc (2.6 g), EtOAc/MeOH 7.5:2.5 (9.5 g) and MeOH (7.7 g). CH₂Cl₂ I fraction (2.0 g) was subjected to repeated silica gel flash column chromatography using n-hexane/CH₂Cl₂ 9:1 to afford compounds 9 (10.7 mg) and 10 (12.3 mg). The n-hexane extract from the root wood (3.9 g) was subjected to silica gel column chromatography eluting with n-hexane (920 mL), EtOAc (150 mL), CHCl₃ (450 mL) and MeOH (100 mL) to yield the fractions: n-hexane (0.8 g), CHCl₃ (2.5 g), EtOAc (0.3 g) and MeOH (0.02 g). CHCl₃ fraction (2.5 g) was fractioned over silica gel, eluting with n-hexane/EtOAc (9:1, 8:2, 7:3, 6:4, 1:1, 7.5:2.5), EtOAc and MeOH, to afford 100 fractions (10 mL each) which after TLC analysis were combined to 10 fractions (F_G1-F_G10). Fraction F_G3 (0.6 g, n-hexane/EtOAc 9:1 and 8:2) was subjected to successive silica gel column chromatography to yield compounds 1 (12.0 mg), 5 (6.2 mg) and 11 (7.2 mg). The EtOH extract from the root wood (15.9 g) was subjected to silica gel column chromatography eluted with CH₂Cl₂ (500 mL), CH₂Cl₂/EtOAc 1:1 (850 mL), EtOAc (600 mL), EtOAc/MeOH 1:1 (600 mL) and MeOH (800 mL), yielding the respective fractions: CH₂Cl₂ (2.9 g), CH₂Cl₂/EtOAc 1:1 (4.0 g), EtOAc (1.6 g), EtOAc/MeOH 1:1 (6.2 g) and MeOH (1.3 g). The CH₂Cl₂ fraction (2.9 g) was subjected to silica gel column chromatography using n-hexane/EtOAc gradient (95:0.5 to 1:1, 2.5:7.5), EtOAc and MeOH to give 103 fractions (10 mL) that after TLC analysis were pooled into 16 fractions (F_H1-F_H16). Fraction F_H7 (261.1 mg, n-hexane/EtOAc 8:2 and 7.5:2.5) yielded the mixture of compounds 12/13 (21.6 mg) and 3 (5.3 mg). Fraction CH₂Cl₂ III (0.7 g) was subjected to slica gel column chromatography using n-hexane/EtOAc (9:1, 8.5:1.5, 8:2, 7.5:2.5, 7:3, 4:6), EtOAc and MeOH resulting in 16 fractions (F_I1-F_I16), after TLC analysis. F_I5 (n-hexane/EtOAc 8:2, 130.5 mg) was fractionated on Sephadex LH-20 column using acetone/MeOH 1:1 to produce the mixture of 13/14 (6.6 mg). Fraction CH₂Cl₂/EtOAc I (3.0 g) was chromatographed over silica gel, eluting with n-hexane/EtOAc (9:1, 8:2, 7:3, 1:1, 4:6), EtOAc and MeOH to afford 7 fractions (F1-F7, 100 mL each). F6 (n-hexane/EtOAc 1:1, 258.3 mg) after purification over Sephadex LH-20 (CH₂Cl₂/MeOH 1:9) yielded 15 (3.5 mg).

Supplementary Information

Supplementary data for compounds 6 and 7 are available free of charge at http://jbcs.sbq.org.br as PDF file.

Acknowledgments

The authors thank to Prof. Edilberto R. Silveira (UFC) for the plant material collection and for the providing the NMR facilities, and to CNPq/CAPES/FUNCAP and PRONEX for the financial support.

Submitted: June 26, 2011

Published online: December 6, 2011

[Supplementary material]

• 1. Paul, M. D.; Medicinal Natural Products: A Biosyntthetic Approach, 2^nd ed., John Wiley & Sons: New York, 2003.
• 2. Lim, C.-G.; Koffas, M. A. G.; Curr. Org. Chem 2010, 14, 1727.
• 3. Ferreira, J. F. S.; Luthria, D. L.; Sasaki, T.; Heyerick, A.; Molecules 2010, 15, 3135.
• 4. Magalhães, A. F.; Tozzi. A. M. G. A.; Magalhães, E. G.; Sannomiya, M.; Soriano, M. D. P. C.; Perez, M. A. F.; An. Acad. Bras. Cienc. 2007, 79, 351.
• 5. Garcia-Lafuente, A.; Guillamon, E.; Villares, A.; Rostagno, M. A.; Martinez, J. A.; Inflammation Res. 2009, 58, 537.
• 6. Parveen, Z.; Deng, Y.; Saeed, M. K.; Dai, R.; Ahamad, W.; Yu, Y. H.; YaKugaku Zasshi 2007, 127, 1275.
• 7. Patel, B.; Das, S.; Prakash, R.; Yasir, M.; Int. J. Adv. Pharm. Sci. 2010, 1, 32.
• 8. Lima, A. F.; Mileo, P. G.; Andrade-Neto, M.; Braz-Filho, R.; Silveira, E. R.; Pessoa, O. D. L.; Magn. Reson. Chem. 2009, 47, 165.
• 9. Campos, D. A.; de Lima, A. F.; Ribeiro, S. R. L.; Silveira, E. R.; Pessoa, O. D. L.; Rao, V. S.; Santos, F. A.; J. Pharm. Pharmacol. 2008, 60, 391.
• 10. Santos, D. A. P.; Braga, P. A. CVieira, P. C.; Magalhaes, A. F.; Magalhaes, E. G.; Marsaioli, A. J.; Moraes, V. R. S; Rattray, L.; Croft, S. L.;J. Pharm. Pharmacol. 2009, 61, 257.
• 11. Cassidy, C. E.; Setzer, W. N.; J. Mol. Model. 2010, 16, 311.
• 12. Alencar, N. M. N.; Cavalcante, C. F.; Vasconcelos. M. P.; Leite, K. B.; ragao, K. S.; Assreuy, A. M. S.; Nogueira, N. A.P.; Cavada, B. S.; Vale, M. R.; J. Pharm. Pharmacol. 2005, 57, 919.
• 13. Nascimento, M. C.; Dias, R. L. V.; Mors, W. B.; Phytochemistry 1976, 15, 1553.
• 14. Andrei, C. C.; Ferreira, D. T.; Faccione, M.; Moraes, L. A. B.; Carvalho, M. G.; Braz-Filho, R.; Phytochemistry 2000, 55, 799.
• 15. Arriaga, A. M. C.; Gomes, G. A.; Braz-Filho, R.; Fitoterapia 2000, 71, 211.
• 16. Garcez, F. R.; Scramin, S.; Nascimento, M. C.; Mors, W. B.; Phytochemistry 1988, 27, 1079.
• 17. Talapatra, S. K.; Mallik, A. K.; Talapatra, B.; Phytochemistry 1980, 19, 1199.
• 18. Monache, F. D.; Suarez, L. E. C.; Marini-Bettolo, G. B.; Phytochemistry 1978, 17, 1812.
• 19. Borges-Argáez, R.; Díaz, M. E. P.; Waterman, P. G.; Peña-Rodríguez, L. M.; J. Braz. Chem. Soc. 2005, 16, 1078.
• 20. Paula, J. R.; Vieira, I. J. C.; Silva, M. F. G. S., Fo, E. R.; Fernandes, J. B.; Vieira, P. C.; Pinheiro, A. L.; Vilela, E. F.; Phytochemistry 1997, 44, 1449.

Publication Dates

• Publication in this collection
  02 Mar 2012
• Date of issue
  Feb 2012

History

• Received
  26 June 2011
• Accepted
  06 Dec 2011