Prenylated Naphthoquinone Dimers from Lippia microphylla.

Transcription

1 Vol., Nº, 00 and C NMR Application and C NMR Application to Structure Elucidation of Prenylated Naphthoquinone Dimers from Lippia microphylla. élcio Silva dos Santos, Telma Lêda Gomes Lemos*, tília Deusdênia Loiola Pessoa, Sônia Maria liveira Costa, Daniel Esdras de Andrade Uchôa and Edilberto Rocha Silveira. Centro Nordestino de Aplicação e Uso da RMN (CENAUREMN), Curso de Pós-Graduação em Química rgânica e Inorgânica, Departamento de Química rgânica e Inorgânica, Universidade Federal do Ceará. Cx. Postal.00, Fortaleza-CE, Brazil. tlemos@dqoi.ufc.br Keywords: spectroscopic data, naphthoquinone dimer, Lippia microphylla, Verbenaceae. Abstract: Two prenylated naphthoquinone dimers, a known and a new one, were isolated from roots of Lippia microphylla. Their structures were determined by spectroscopic methods (IR and MS) including detailed D and D NMR (BBD, DEPT, MQC, MBC and NESY) analysis. The unambiguous assignment of all NMR data with all hydrogens and carbons of the novel compound is provided. Resumo: Duas naftoquinonas diméricas preniladas, uma conhecida e outra inédita, foram isoladas das raízes de Lippia microphylla. Suas estruturas foram determinadas por métodos espectroscópicos (IV and EM) incluindo detalhada análise de RMN D e D (BBD, DEPT, MQC, MBC e NESY). A correlação inequívoca de todos os dados de RMN com todos os átomos de hidrogênio e carbono da estrutura proposta para o novo composto é sugerida. Introduction The Lippia genus (Verbenaceae) is a prolific source of flavouring plants, most of them with folk medicinal use. In the Northeast of Brazil two Lippia species are widely used: L. sidoides, popularly known as alecrim-pimenta of topical use as a general antiseptic, and L. alba, popularly designated as erva cidreira, whose leaves infusion is used as a soothing tea. A literature survey revealed several papers about Lippia species but no phytochemical report was found for L. microphylla, popularly known as alecrim-de-tabuleiro. Despite not being currently used in popular medicinal it was chosen as the subject of the present work. Experimental Plant material Roots of L. microphylla (Verbenaceae) were collected in April, in Várzea Alegre, Ceará State Brazil, and authenticated by Prof. Afrânio Gomes Fernandes of the Departamento de Biologia, Universidade Federal do Ceará. A voucher specimen (No ) has been deposited in the erbarium Prisco Bezerra - UFC. Extraction and isolation The dried, ground biomass (0 g) from the roots of L. microphylla was extracted at room temperature with Et. After solvent evaporation under reduced pressure, a brown residue (0 g) was obtained. This material was fractioned over a Silica gel column with hexane/ccl (:), CCl, EtAc and Me. The hexane/ccl (:) fraction was chromatographed over Silica gel by elution with 0 00% EtAc-hexane mixtures. The residue from the % EtAc-hexane fraction was subjected to repeated column chromatography to yield ( mg, m.p. C) and (00 mg, m.p. 0 C) as pure compounds. NMR spectra and C spectra were determined on a Bruker DRX 00 spectrometer at C operating at 00. Mz for and. Mz for C. The samples [ ( mg) and (0 mg)] were dissolved in 0. ml of CDCl or DMS-d and set into.0 mm NMR tubes (Norell, Inc., #0-UP RB). Internal lock and residual CCl (δ =.) and CDCl (C =.00) signals were used as references for and.

2 Vol., Nº, 00 and C NMR Application C C C 0 ' ' 0' ' 0 ' ' 0' ' ' ' ' ' ' ' ' ' ' ' ' ' ' ' C C ' ' C C ' ' The acquisition was performed by standard Bruker pulse programs [zg0 ( ), zgpg0 ( C- BBD), dept ( C-DEPT ), cosy0 (, - CSY), invbtp (MQC), invlplrnd (MBC) using either a mm dual C/ probe for normal ( C) detection or mm multinuclear inverse Z-grad. probe for inverse ( ) detection. and K data point sets, with spectral width of and Kz, The NESY spectrum was obtained with Kz spectral width for both F and F. K x K data blocks, with phase adjustment for both F and F, followed by final data matrix symmetrization were used for processing. ther parameters were: increments in t, transients and relaxation delay of s and a mixing time of 0 ms. Results and Discussion Silica gel column chromatography of the hexane/ccl fraction from the Et extract of roots of L. microphylla, using n-hexane with increasing concentrations of EtAc as eluent, yielded the two naphthoquinones, and. Naphthoquinone, was isolated as a green powder, m.p. C. Spectrometric analysis, particularly by one and two-dimensional NMR, allowed its identification as tecomaquinone I (), by comparison to the data already reported in the literature. Tecomaquinone I had been previously isolated from L. sidoides, a congener of L. microphylla. Naphthoquinone, was isolated as colorless prisms, m.p. 0 C. Its IR spectrum showed absorption bands for hydroxyl ( cm - ) and were collected for and C one-dimensional spectra, respectively. Inverse detected D heteronuclear correlated spectra were collected over K data points, with spectral width of Kz and points in F, and Kz in F. Data processing were performed with K x w blocks, using backward linear prediction in F to generate the final data matrix. conjugated carbonyl groups (0 cm - ). The molecular ion peak with m/z 0 Daltons, in conjunction with the and BB spectra (Table ) showing signals, allowed the determination of its molecular formula as C 0. The DEPT spectrum revealed three methyl groups, one of which was a methoxy group (δ.), one methine, nine monohydrogenated sp carbons, and by comparison to the BB spectrum, fourteen nonhydrogenated sp carbons, including three carbonyl carbons (δ.,. and.) and two oxygenated carbons (δ. and.). and C NMR data comparison of both and, revealed a striking similarity, except for the missing cyclic isoprene moiety for, compared to and, in addition, the presence of a carbomethoxy group for. This strongly supports the oxidation at C-, generating the carbomethoxy group, and the consequent elimination of the remaining four carbons unity of that side chain in. Even though a more deshielded absorption for C- (conjugated with the carbomethoxy) was expected, initially the complete C data assignment was a difficult task because the absence of any hydrogen long-range correlation with C- or C- (this one is ortho to the

3 Vol., Nº, 00 and C NM hydroxy, but is conjugated to the quinone C-`) did not allow an unambiguous assignment of those carbons. Fortunately, the MBC data obtained in dry DMS-d revealed long-range correlations of the hydroxy hydrogen (δ 0.) with C- (δ.), C- (δ.) and C- (δ 0.). In addition, the distinction between C-`(δ.) and C-` (δ.), both conjugated to the quinone carbonyls, was done by the correlations of the carbinolic like, and doubly allylic -`(δ.) with C- (δ.), C-` (δ.), C-` (δ.) and C-` (δ.), as well as by the correlations of the vinyl hydrogen -` (δ.0) with C-`(δ.0) and C- `(δ.0). Figure shows some significant correlations observed through the MBC experiment. Finally the relative stereochemistry suggested for, named microphyllaquinone, similarly to, was based on the NESY analysis, Figure. C C 0 0 ' ' ' ' ' ' 0' ' ' ' 0 ' ' ' ' ' ' 0' ' ' ' ' ' C C ' ' C C ' ' Figure. Interpreted MBC spectrum of (00/ Mz, DMS-d ):numbered arrows indicate long-range heteronuclear C- correlations whose signals are marked with the same number. Figure. Interpreted NESY spectrum of (00 Mz, DMS-d ):numbered double arrows indicate observed dipolar spatial correlations whose cross-peaks are marked with the same number on the structure.

4 Vol., Nº, 00 and C NMR Application Table C NMR ( Mz) and NMR (00 Mz) Data for and. (CDCl ) (CDCl ) (DMS-d ) Position δ C δ δ C δ δ (d, J=.).0. (d, J=,). (m).. (t, J=.).. (m).0 (m).0. (t, J=.).00. (m).0 (m).. (d, J=.).. (d, J=.). (m) (d, J=.). -.. (d, J=.) (s) - -. (s) (d, J=.).0 (m) (m). (m).... (m). (m)..0.. (d, J=.).0 (m) (d, J=.).. (d, J=.). (d, J=.0).. (d, J=.)..0 (d, J=.).0 (d, J=.0) (s). (s) (s).0 (s) Me (s). (s) (s) 0. (s)

5 Vol., Nº, 00 and C NMR Application Conclusion Two prenylated naphthoquinone dimers were isolated from L. microphylla. Beside other spectroscopic techniques, extensive NMR studies allowed their identification as the known tecomaquinone I, previously isolated from L. sidoides, and a novel one we have designated microphyllaquinone. The unambiguous assignment of all NMR data with all hydrogen and carbons was also performed. The alecrim-de-tabuleiro should maybe have the same properties of L. sidoides, which shows a similar chemical composition. Acknowledgments The authors express their appreciation to A. G. Fernandes for plant material identification and to R. Braz-Filho for the EIMS data and his expert assistance. To CNPq / CAPES / FINEP / FUNCAP/ PRNEX for financial support. E. R. S. e T. L. G. L. acknowledge CNPq for their Research Fellowships. References. Matos, F. J. A. Rev. Bras. Farm.,, -.. Matos, F. J. A. Rev. Bras. Farm.,, -.. Lemos, T. L. G.; Costa, S. M..; Pessoa,. D. L.; Braz-Filho, R. Magn. Reson. Chem.,, 0-.