Chemical Constituents from the Leaves of Annona Rugulosa (Annonaceae)

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Biochemical Systematics and Ecology 49 (2013) 152–155

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Biochemical Systematics and Ecology

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Chemical constituents from the leaves of Annona rugulosa (Annonaceae)

Mayara Evelyn Vendramin a, Emmanoel Vilaça Costa b, Élide Pereira dos Santos c, Maria Lúcia Belém Pinheiro d, Andersson Barison a, Francinete Ramos Campos e,*

a Departamento de Química, Universidade Federal do Paraná, Centro Politécnico, Jardim das Américas, CEP 81531-990, P.O. Box 19081, Curitiba, PR, Brazil b Departamento de Química, Universidade Federal de Sergipe, Rosa Elze, CEP 49100-000, São Cristóvão, SE, Brazil c Departamento de Botânica, Universidade Federal do Paraná, Centro Politécnico, Jardim das Américas, CEP 81531-990, P.O. Box 19031, Curitiba, PR, Brazil d Departamento de Química, Universidade Federal do Amazonas, CEP 69077-000, Manaus, AM, Brazil e Departamento de Farmácia, Universidade Federal do Paraná, Campus Jardim Botânico, Jardim Botânico, CEP 80210-170, Curitiba, PR, Brazil

article info

Article history: Received 6 November 2012 Accepted 9 March 2013 Available online 28 April 2013

Keywords: Annonaceae Annona rugulosa Magnococline NMR Alkaloids Steroids

1. Subject and source

Annona L. belongs to the family Annonaceae and includes approximately 175 species of tree and shrubs, with the recent inclusion of the species from the genus Rollinia proposed by Heimo Rainer (Rainer, 2007), found predominantly in lowland tropical regions. Economically, this genus is the most important of the family Annonaceae due to its edible fruits such as “araticum”, “graviola”, “fruta-do-conde”, “pinha”, “ata”, “biribá”, “condessa”, “marolo” and others, and its medicinal prop- erties (Corrêa, 1984). Annona rugulosa (Schltdl.) H. Rainer (basynonymy Rollinia rugulosa Schltdl.) is a tree up to 12 m high and 40 cm diameter (Maas et al., 1992; Rainer, 2007; Maas et al., 2011), found in South and Southeast of Brazil in “Mata Atlântica” (Mass et al., 2012). It is popularly known as “araticum-de-porco”, “araticum-verde”, “araticum-de-comer”, “araticum” and “anona”.In folk medicine the tea from leaves is used against kidney infection and sore throat (Garlet and Irgang, 2001). In the present investigation, the botanical material (leaves) of A. rugulosa was collected in September 2009, in Curitiba, Paraná state, Brazil [coordinates: 2523056.1000 S and 4915008.8100 W] and identiﬁed by Prof. Dr. E. P. Santos from Department of Botany of Paraná

* Corresponding author. Tel: þ55 41 3360 4076; fax: þ55 41 3360 4106. E-mail address: [email protected] (F.R. Campos).

Federal University. A voucher specimen (# 1249) was deposited in the UPCB herbarium of Paraná Federal University (UFPR) Brazil. The molecular authentication of A. rugulosa was performed by the sequence of two short coding regions of plastid DNA (matK and rbcL), used as standard barcodes for land plants to identify species (CBOL Plant Working Group, 2009; Chase and Fay, 2009; Li et al., 2011). Total genomic DNA was extracted from fresh material of A. rugulosa, followed by PCR ampliﬁcation and DNA sequencing, according to CBOL Plant Working Group (2009). The sequences are deposited in GenBank, under numbers JX880394 (matK) and JX880395 (rbcL).

2. Previous work

There are no previous phytochemical investigations on A. rugulosa.

3. Present study

Air-dried (40 C, forced ventilation) and powdered leaves of A. rugulosa (728.0 g) were extracted with EtOH:H2O (70:30, v/v) on a turbo extractor at 60 Hz for 2 h at room temperature. The hydroalcoholic extract was then filtered through a filter paper (No. 1) follow by solvent reduction to 1/6 of initial volume, under reduced pressure, and freeze-dried yielding ethanolic extract (133.5 g). TLC investigations indicated a high concentration of alkaloids in the EtOH extract. Therefore, an aliquot of this extract (40.0 g) was initially subjected to an acid-base extraction (Costa et al., 2006) to give alkaloid (1.24 g) and neutral (15.79 g) fractions. An aliquot of alkaloid fraction (1.13 g) was initially subjected to silica gel column chromatography previously treated with a 10% NaHCO3 solution (Costa et al., 2006), and eluted with increasing concentrations of CH2Cl2 in hexane (100:0 to 10:90, v/v), followed by EtOAc in CH2Cl2 (100:0 to 10:90, v/v), and MeOH in EtOAc (100:0 to 50:50, v/v), giving 129 fractions (10 mL each) that were evaluated and pooled according to TLC analysis to yield 19 fractions. Fraction 9 (18.0 mg) resulted in a mixture of b-sitosterol (1, Habib et al., 2007) and stigmasterol (2, Habib et al., 2007). Fraction 13 (164.0 mg) was subjected to preparative TLC eluted with CHCl3:MeOH (90:10, v/v) affording anonaine (3, 6.0 mg, Hu et al., 2010). Fraction 14 (71.0 mg) was further purified on preparative TLC eluted with CHCl3:MeOH (95:5, v/v) yielding anonaine (3, 9.0 mg, Chen et al., 1997; Chang et al., 2000; Hu et al., 2010; Da Cruz et al., 2011), nornantenine (4, 9.0 mg, Guinaudeau et al., 1983), N-nornuciferine (5, 9.0 mg, Guinaudeau et al., 1983; Chang et al., 2000), xylopine (6, 13.0 mg, Guinaudeau et al.,1983), norisocorydine (7, 4.0 mg, Lee et al., 1992) and litseferine (8, 4.0 mg, Guinaudeau et al., 1988). Fractions 15 (112.0 mg) and 16 (261.0 mg) were subjected to preparative TLC eluted with the same solvent system as above, giving lanuginosine (9, 6.0 mg, Wijeratne et al., 1996), liriodenine (10, 5.0 mg; Wijeratne et al., 1996; Costa et al., 2011a) and magnococline (11, 6.0 mg, Yang and Liu, 1971). Fractions 17 and 18 were pooled and resulted in asimilobine (12, 11.0 mg, Chen et al., 1997; Chang et al., 2000; Guo et al., 2011; Da Cruz et al., 2011). Fraction 19 (374.0 mg) was further purified on preparative TLC eluted with CHCl3:MeOH (80:20, v/v) resulting in isoboldine (13, 7.0 mg, Iwasa et al., 2008), reticuline (14, 7.0 mg, Lee et al., 2007; Da Cruz et al., 2011) and N-methylcoclaurine (15, 3.0 mg, Tomita et al., 1965). All isolated compounds (Fig. 1) were identified by a series of spectrometric methods, mainly MS and NMR (1D and 2D), as well as comparison with data reported in the literature. Although magnococline (11) has been described a long time ago, its NMR dataset is incomplete (Yang and Liu, 1971), and are given in Tau (s). Therefore, the complete and unequivocal NMR data for magnococline (11) were revised according to 1D and 2D NMR experiments, and are described as follows: 1H NMR (400 MHz, CDCl3) d 7.20 (2H, d, J ¼ 8.6 Hz, H-11 and H-15), 6.87 (2H, d, J ¼ 8.6 Hz, H-12 and H-14), 6.76 (1H, d, J ¼ 8.3 Hz, H-6), 6.63 (1H, d, J ¼ 8.3 Hz, H-5), 3.89 (3H, s, 7-OCH3), 3.79 (3H, s, 13-OCH3), 4.55 (1H, m, H-1), 3.53 (2H, m, H-3), 3.35 (2H, m, H-9), 13 2.66 (2H, m, H-4); C NMR (100 MHz, CDCl3) d 158.3 (C-13), 144.4 (C-7), 141.9 (C-8), 130.4 (C-11 and C-15), 130.3 (C-10), 126.9 (C-4a), 122.9 (C-8a), 119.5 (C-5), 114.2 (C-12 and C-14), 109.3 (C-6), 56.1 (7-OCH3), 55.4 (13-OCH3), 52.4 (C-1), 42.3 (C-3), 36.8 (C-9), 27.4 (C-4).

4. Chemotaxonomic signiﬁcance

The present work reports the isolation and identiﬁcation of two steroids b-sitosterol (1) and stigmasterol (2); eight aporphine alkaloids, anonaine (3), nornantenine (4), N-nornuciferine (5), xylopine (6), norisocorydine (7), litseferine (8), asimilobine (12) and isoboldine (13); two oxoaporphine alkaloids, lanuginosine (9) and liriodenine (10); and tree benzyl- tetrahydroisoquinoline alkaloids, magnococline (11), reticuline (14) and N-methylcoclaurine (15). All isolated compounds are reported here for the ﬁrst time from this plant. b-sitosterol and stigmasterol are two compounds reported to occur in many plants (Dewick, 2009). Anonaine (3), N-nornuciferine (5), xylopine (6), liriodenine (10), and asimilobine (12) are ubiquitous in Annonaceae, and are found in all most genera of this family, mainly in the Annona, Artabotrys, Desmos, Duguetia, Enantia, Fissistigma, Goniothalamus, Guatteria, Polyalthia and Xylopia (Guinaudeau et al., 1975, 1979, 1983, 1988, 1994; Leboeuf et al., 1982; Costa et al., 2009; Costa et al., 2006; Pinheiro et al., 2009; Costa et al., 2010, 2011a,b). They are considered as chemotaxonomic markers of the genera of Annonaceae, especially Annona (Da Cruz et al., 2011). In Annona, they have been reported in the leaves of Annona sericea (Campos et al., 2008), fresh fruits of Annona glabra (Chang et al., 2000), stems of Annona cherimola (Chen et al., 1997), bark of Annona salzmannii (Da Cruz et al., 2011), stems of Annona amazonica (Pinheiro et al., 2009), bark (Costa et al., 2006) and branches of Annona foetida (Costa et al., 2011a) and leaves of Annona pickelii (Dutra et al., 2012). Isoboldine (13) and reticuline (14) were recently described in the leaves of A. sericea (Campos et al., 2008). N- 154 M.E. Vendramin et al. / Biochemical Systematics and Ecology 49 (2013) 152–155

Δ22,23

Fig. 1. Chemical constituents isolated from the leaves of Annona rugulosa.

methylcoclaurine (15) was described in only two species in genus, in A. sericea (Campos et al., 2008) and Annona squamosa (Wu et al., 1994). Therefore, the present findings support the view that Rollinia cannot be maintained as a separate genus and must be placed in Annona as proposed by Rainer (2007). Norisocoridine (7) and litseferine (8) are been described for the first time in the genus Annona. Magnococline (11) is described for the first time in the Annonaceae, and was reported only once elsewhere in Lauraceae, a family phylogenetically close to Annonaceae (Yang and Liu, 1971).

Acknowledgements

The authors are grateful to CNPq, CAPES, FINEP, Fundação Araucária, FAPITEC/SE and UFPR for ﬁnancial support and fellowships.

References

Campos, F.R., Batista, R.L., Batista, C.L., Costa, E.V., Barison, A., Santos, A.G., Pinheiro, M.L.B., 2008. Biochem. Syst. Ecol. 36, 804. CBOL Plant Working Group, 2009. Proc. Natl. Acad. Sci. U S A 106, 12794. Chase, M.W., Fay, M.F., 2009. Science 35, 682–683. Chang, F.-R., Chen, C.-Y., Hsieh, T.-J., Cho, C.-P., Wu, Y.-C., 2000. J. Chin. Chem. Soc. 47, 913. Chen, C.-Y., Chang, F.-R., Wu, Y.-C., 1997. J. Chin. Chem. Soc. 44, 313. Corrêa, M.P., 1984. Dicionário das plantas úteis do Brasil e das exóticas cultivadas. IBDF, Rio de Janeiro, RJ. Costa, E.V., Pinheiro, M.L.B., Souza, A.D.L., Barison, A., Campos, F.R., Valdez, R.H., Ueda-Nakamura, T., Filho, B.P.D., Nakamura, C.V., 2011a. Molecules 16, 9714. Costa, E.V., Marques, F.A., Pinheiro, M.L.B., Braga, R.M., Delarmelina, C., Duarte, M.C.T., Ruiz, A.L.T.G., Carvalho, J.E., Maia, B.H.L.N.S., 2011b. J. Braz. Chem. Soc. 22, 1111. Costa, E.V., Pinheiro, M.L.B., Marques, F.A., Braga, R.M., Maia, B.H.L.N.S., 2009. Biochem. Syst. Ecol. 37, 43. Costa, E.V., Pinheiro, M.L.B., Barison, A., Campos, F.R., Salvador, M.J., Maia, B.H.L.N.S., Cabral, E.C., Eberlin, M.N., 2010. J. Nat. Prod. 73, 1180. Costa, E.V., Pinheiro, M.L.B., Xavier, C.M., Silva, J.R.A., Amaral, A.C.F., Barison, A., Campos, F.R., Ferreira, A.G., Machado, G.M.C., Leon, L.L.P.J., 2006. J. Nat. Prod. 69, 292. Da Cruz, P.E.O., Costa, E.V., Moraes, V.R.S., Nogueira, P.C.L., Vendramin, M.E., Barison, A., Ferreira, A.G., Prata, A.P.N., 2011. Biochem. Syst. Ecol. 39, 872. Dewick, P.M., 2009. Medicinal Natural Products: a Biosynthetic Approach, third ed. John Wiley & Sons, Ltd, United Kingdom. Dutra, L.M., Costa, E.V., Moraes, V.R.S., Nogueira, P.C.L., Vendramin, M.E., Barison, A., Prata, A.P.N., 2012. Biochem. Syst. Ecol. 41, 115. Garlet, T., Irgang, B., 2001. Revista Brasileira de Plantas Medicinais 4, 9. Guinaudeau, H., Leboeuf, M., Cavé, A., 1975. Lloydia 38, 275. Guinaudeau, H., Leboeuf, M., Cavé, A., 1979. J. Nat. Prod. 42, 325. Guinaudeau, H., Leboeuf, M., Cavé, A., 1983. J. Nat. Prod. 46, 761. Guinaudeau, H., Leboeuf, M., Cavé, A., 1988. J. Nat. Prod. 51, 389. Guinaudeau, H., Leboeuf, M., Cavé, A., 1994. J. Nat. Prod. 57, 1033. M.E. Vendramin et al. / Biochemical Systematics and Ecology 49 (2013) 152–155 155

Guo, Z.-F., Wang, X.-B., Luo, J.-G., Luo, J., Wang, J.-S., Kong, L.-Y., 2011. Fitoterapia 82, 637. Habib, M.R., Nikkon, F., Rahman, M., Haque, Z.M.E., Karim, M.R., 2007. Pak. J. Biol. Sci. 10, 4174. Hu, R., Dai, X., Lu, Y., Pan, Y., 2010. J. Chromatogr. B 878, 1881. Iwasa, K., Takahashi, T., Nishiyama, Y., Moriyasu, M., Sugiura, M., Takeuchi, A., Tode, C., Tokuda, H., Takeda, K., 2008. J. Nat. Prod. 71, 1376. Leboeuf, M., Cavé, A., Bhaumik, P.K., Mukherjee, B., Mukherjee, R., 1982. Phytochemistry 21, 2783. Lee, S.S., Chen, C.K., Chen, I.S., Liu, K.C.S., 1992. J. Chin. Chem. Soc. 39, 453. Lee, S.S., Lai, Y.C., Chen, C.K., Tseng, L.H., Wang, C.Y., 2007. J. Nat. Prod. 70, 637. Li, M., Cao, H., But, P.P.H., Shaw, P.C., 2011. J. Syst. Evol. 49, 271. Maas, P.J.M., Westra, L.Y.T., et al., 1992. Rollinia – Fl. Neotrop. Monogr. 57, 1–188. Maas, P.J.M., Westra, L.Y.T., Rainer, H., Lobão, A.Q., Erkens, R.H.J., 2011. Nord. J. Bot. 29, 257. Maas, P., Rainer, H., Lobão, A., 2012. Annonaceae in Lista de Espécies da Flora do Brasil. Jardim Botânico do Rio de Janeiro. http://ﬂoradobrasil.jbrj.gov.br/ 2012/FB110261. Pinheiro, M.L.B., Xavier, C.M., Souza, A.D.L., Rabelo, D.M., Batista, C.L., Batista, R.L., Costa, E.V., Campos, F.R., Barison, A., Valdez, R.H., Ueda-Nakamura, T., Nakamura, C.V., 2009. J. Braz. Chem. Soc. 20, 1095. Rainer, H., 2007. Ann. Naturhist. Mus. Wien. 108B, 191. Tomita, M., Shingu, T., Fujitani, K., Furukawa, H., 1965. Chem. Pharm. Bull. 13, 921. Wijeratne, E.M.K., Hatanaka, Y., Kikuchi, T., Tezuka, Y., Gunatilaka, A.A.L., 1996. Phytochemistry 42, 1703. Wu, Y.-C., Chang, F.-R., Chen, K.S., Ko, F.-N., Teng, C.-M., 1994. Chin. J. Pharmaceuticals 46, 439. Yang, T.-H., Liu, S.-C., 1971. J. Chin. Chem. Soc. 18, 91.