Chemical constituents of Cordia piauhiensis: Boraginaceae

Abstracts

A new monodesmoside triterpenoid saponin characterized as 3beta-O-alpha-L-rhamnopyranosyl-(1->2)-beta-D-glucopyranosyl pomolic acid, was isolated from Cordia piauhiensis Fresen (Boraginaceae). Its structure was determined by extensive NMR analysis, including ¹H, ¹H-COSY, HMQC, HMBC, and NOESY experiments. In addition, four known triterpenoids: quinovic acid, cincholic acid, cincholic 3beta-O-6-deoxy-beta-D-glucopyranoside acid and quinovic 3beta-O-beta-D-glucopyranoside acid were also isolated.

Cordia piauhiensis; Boraginaceae; triterpenes; saponins

Uma nova saponina triterpênica monodesmosídica, caracterizada como ácido 3beta-O-alfa-L-ramnopiranosil-(1->2)-beta-D-glicopiranosil pomólico, foi isolada de Cordia piauhiensis Fresen (Boraginaceae). Sua estrutura foi determinada através de extensiva análise de métodos de RMN, incluindo os experimentos ¹H, ¹H-COSY, HMQC, HMBC e NOESY. Os triterpenóides ácido quinóvico, ácido cinchólico, ácido 3beta-O-6-deoxi-beta-D-glicopiranosídeo cinchólico e 3beta-O-beta-D-glicopiranosídeo quinóvico foram também isolados.

SHORT REPORT

Chemical constituents of Cordia piauhiensis – Boraginaceae

Renata Paiva dos Santos^I; Telma L. G. Lemos^I; Otilia Deusdênia L. Pessoa^I,*; Raimundo Braz-Filho^II; Edson Rodrigues-Filho^III; Francisco Arnaldo Viana^IV; Edilberto R. Silveira^I

^IDepartamento de Química Orgânica e Inorgânica, Centro de Ciências, Universidade Federal do Ceará, CP 12200, 60021-970 Fortaleza - CE, Brazil

^IISetor de Química de Produtos Naturais - LCQUI - CCT, Universidade Estadual do Norte Fluminense, 28015-620 Campos - RJ, Brazil

^IIIDepartamento de Química, Universidade Federal de São Carlos, 14040-970 São Carlos - SP, Brazil

^IVUniversidade do Estado do Rio Grande do Norte, 59610-210 Mossoró - RN, Brazil

ABSTRACT

A new monodesmoside triterpenoid saponin characterized as 3b-O-a-L-rhamnopyranosyl-(1®2)-b-D-glucopyranosyl pomolic acid, was isolated from Cordia piauhiensis Fresen (Boraginaceae). Its structure was determined by extensive NMR analysis, including ¹H, ¹H-COSY, HMQC, HMBC, and NOESY experiments. In addition, four known triterpenoids: quinovic acid, cincholic acid, cincholic 3b-O-6-deoxy-b-D-glucopyranoside acid and quinovic 3b-O-b-D-glucopyranoside acid were also isolated.

Keywords:Cordia piauhiensis, Boraginaceae, triterpenes, saponins

RESUMO

Uma nova saponina triterpênica monodesmosídica, caracterizada como ácido 3b-O-a-L-ramnopiranosil-(1®2)-b-D-glicopiranosil pomólico, foi isolada de Cordia piauhiensis Fresen (Boraginaceae). Sua estrutura foi determinada através de extensiva análise de métodos de RMN, incluindo os experimentos ¹H, ¹H-COSY, HMQC, HMBC e NOESY. Os triterpenóides ácido quinóvico, ácido cinchólico, ácido 3b-O-6-deoxi-b-D-glicopiranosídeo cinchólico e 3b-O-b-D-glicopiranosídeo quinóvico foram também isolados.

Introduction

The genus Cordia (Boraginaceae), a known source of benzoquinones,¹ naphthoquinones,² hydroquinones, cromenes,³ triterpenes,⁴ sesquiterpenes,⁵ polyphenols,⁶ and flavonoids⁷ comprises about 250 species distributed throughout the New World.⁸ Many compounds originally isolated from Cordia species have been reported as presenting several biological activities such as antifungal, larvicidal, anti-inflammatory and anti-androgenic.^2,4,6,7

As part of our current interest in Cordia species from Northeastern Brazil flora we have investigated C. piauhiensis Fresen (syn.: Cordia rufescens A. DC.), an endemic shrub distributed in the South, Southeast and Northeast regions of Brazil.⁸ In a previous paper we reported the isolation and characterization of a triterpenoid bidesmoside saponin from the stems of this plant.⁹ Continuing with our phytochemical research we now report the isolation and structural elucidation of 3b-O-a-L-rhamnopyranosyl-(1®2)-b-D-glucopyranosyl pomolic acid (1), a novel monodesmoside saponin, along with four triterpenoids and other known compounds.

Results and Discussion

Compound 1 was isolated as an amorphous solid, mp 218-220 ºC. The ESIMS (negative ion mode) showed a molecular ion peak at m/z 779 ([M]^-), consistent with a molecular formula C₄₂H₆₈O₁₂, which was confirmed by its ¹H and ¹³C NMR spectral data (Tables 1). Its IR spectrum showed strong absorption bands at n_max 3428 (hydroxyl) and 1695 cm^-1 (carboxyl). The ¹³C NMR BB and DEPT spectra of 1 (Table 1) indicated 42 carbon atoms, related to 8 methyls, 10 methylenes, 16 methines and 8 non-hydrogenated carbons. Besides the 30 carbons assigned to the aglycone, 12 carbons were assigned to two sugar units. The ¹³C NMR spectra demonstrated the presence a trisubstituted double bond (d 129.6 and 140.1), a carboxylic acid group (d 182.6), an oxymethine carbon (d 90.5), an oxygenated non-hydrogenated carbon (d 73.9) and seven methyl groups between d 16.2 and 28.7, consistent with an aglycone, which was established to be the 3b,19a-dihydroxyurs-12-en-28-oic acid, also known as pomolic acid, after appropriate NMR and mass spectrometry (negative ion peak at m/z 471) analysis, and comparison with literature data.¹⁰ The two sugar units were evidenced by two anomeric carbon signals at d 105.7 and 102.0 correlated with the proton signals at d 4.41 (d, J 7.2 Hz) and 5.36 (d, J 1.3 Hz), respectively, in the HMQC spectrum. To determine the identity of each monosaccharide interglycoside linkage, as well as to establish the linkage of the disaccharide chain to the aglycone, ¹H and ¹³C NMR assignments were unambiguously made by a combination of several 2D experiments such as ¹H, ¹H-COSY, HMQC, HMBC and NOESY. In the ¹H NMR spectrum of 1 the presence of a glucose unit was supported by the anomeric proton signal at d 4.41 and two double doublets for the oxymethylene protons at d 3.66 and 3.83. Similarly, the presence of a M rhamnose unit was readily supported by the characteristic methyl doublet at d 1.21 and by the anomeric proton signal at d 5.36.¹¹ The anomeric configurations for the sugar moieties were defined as b for the glucose and a for the rhamnose from their coupling constants of 7.2 and 1.3 Hz, respectively and by comparison with data from literature.¹¹ The linkage between the two sugar units was established from the HMBC correlation between the signals of the anomeric proton of the rhamnose at d 5.36 and the carbon signal of the glucose at d 79.1 (C-2). The attachment of the disaccharide chain was particularly determined based on the relevant correlation between C-1' (d 105.7) and H-3 (d 3.18) in the HMBC spectrum, and by NOE correlation between H-1'(d 4.41) and H-3 (d 3.18), observed in the NOESY experiment as shows in Figure 1. This also was confirmed by the conspicuous deshielding of C-3 (d 90.5) as well as by analogy with the NMR data of the known saponins.^9,12 Thus, the structure of 1 was determined as the 3b-O-a-L-rhamnopyranosyl-(1®2)-b-D-glucopyranosyl pomolic acid.

The known triterpenoids were identified by spectroscopic analysis and comparison with published data to be quinovic acid (3) and cincholic acid (4),¹³ obtained as a 3:1 mixture, respectively, cincholic 3b-O-6-deoxy-b-D-glucopyranoside acid (5)¹³ and quinovic 3b-O-b-D-gluycopyranoside acid (6).¹⁴ Mannitol¹⁵ (2) and b-sitosterol-b-D-glucoside (7)¹⁶ were also isolated.

Experimental

General experimental procedures

Melting points were determined using a digital Mettler Toledo FP90 apparatus. The optical rotations were measured on a Perkin-Elmer 341 digital polarimeter. IR spectra (KBr pellets) were recorded using a Perkin-Elmer FT-IR 1000 spectrometer. ESIMS were measured on a Micromass Quatro LC instrument. NMR spectra were recorded on a Bruker Avance DRX-500 (500 MHz for ¹H and 125 MHz for ¹³C) or DPX-300 (300 MHz for ¹H and 75 MHz for ¹³C) spectrometers using pyridine-d₅, CD₃OD or D₂O as solvents. Chemical shifts, given on the d scale, were referenced to internal DSS for D₂O solution, and to the residual undeuterated portion of the deuterated organic solvent, for proton (pyridine, d_H 8.74, 7.58, 7.22; CD₃OD, d_H 2.31), and the center peak of the deuterated solvent (pyridine, d_C 150.35, 135.91, 123.87; CD₃OD, d_C 49.15). Column chromatography was run using silica gel 60 (70 - 230 mesh, Vetec) and Sephadex LH-20 (Pharmacia). TLC was performed on precoated silica gel poliester sheets (kieselgel 60 F₂₅₄, 0.20 mm, Merck). Saponins were detected by spraying with vanillin/perchloric acid/EtOH solution followed by heating at 120 ºC, while the sugar was detected by spraying the orcinol reagent.

Plant material

Cordia piauhiensis was collected in August 1999, from Barreiro Grande – Crato County, State of Ceará, and identified by Prof. Edson Paula Nunes. A herborized specimen (# 29.104) has been stored at the Herbario Prisco Bezerra (EAC) of the Departamento de Biologia, Universidade Federal do Ceará.

Extraction and isolation

Air-dried and powdered roots (2.8 Kg) and stems (1.7 Kg) were individually extracted exhaustively with EtOH at room temperature. After evaporation of the solvents under reduced pressure the crude extracts were obtained. Upon concentration of the EtOH extract from roots a precipitate was obtained, which was filtered and washed successively with acetone and EtOH to yield 2 (15 g). The remaining EtOH liquors were evaporated (198 g) and coarsely fractionated over silica gel by elution with n-hexane, CHCl₃ followed by EtOAc and finally MeOH. The EtOAc fraction (27 g) was subjected to silica gel CC eluting with n-hexane-EtOAc (8:2, 6:4, 4:6, 2:8 and 0:10) followed by EtOAc-MeOH gradients (8:2, 5:5 and 0:10). A precipitate was obtained from a n-hexane-EtOAc (4:6) subfraction, which was filtered and recrystallized from MeOH to yield a binary mixture of 3 and 4 (102 mg). The EtOAc subfraction was further chromatographed over silica gel to give 5 (95 mg) and 6 (450 mg) by elution with n-hexane-EtOAc (1:9) and EtOAc, respectively. Similarly, the EtOH extract of the stems (103 g) was fractionated over silica using CHCl₃, EtOAc, acetone and MeOH as eluent. The EtOAc fraction (8.1 g) was chromatographed over silica gel eluting with CH₂Cl₂, CHCl₃-EtOAc gradients (8:2, 5:5, 3:7 and 0:10) and MeOH. The subfraction CHCl₃ yielded a precipitate which was purified by recrystallization in MeOH to yield 3 (21 mg). Likewise compound 7 (32 mg) was isolated from subfraction CHCl₃-EtOAc (3:7). The acetone fraction (39.7 g) was dissolved in H₂O-MeOH (8:2) and partitioned with EtOAc and n-BuOH. The EtOAc fraction (10.9 g) was subjected to silica gel CC eluting with EtOAc-MeOH (3:7, 5:5 and 0:10). Subfraction EtOAc-MeOH (3:7, 590 mg) was chromatographed twice over Sephadex LH-20 eluting with MeOH to afford 1 (43 mg).

3b-O-a-L-rhamnopyranosyl-(1®2)-b-D-glucopyranosyl pomolic acid (1)

White amorphous solid; mp 218-220 ºC; [a]_D²⁰ - 0.44º (c 0.05, MeOH); IR (KBr) n_max/cm^-1 : 3428, 2932, 2883, 1695, 1653, 1457, 1131, 1050; ¹H (300 MHz) and ¹³C NMR (75 MHz) data, see Table 1.

Acknowledgments

The authors thank Prof. Edson P. Nunes (Departamento de Biologia – UFC) for plant identification. This work was supported by grants from the Brazilian National Agencies CAPES/CNPq/FUNCAP and PRONEX.

Received: June 10, 2004

Published on the web: March 15, 2005

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Publication Dates

• Publication in this collection
  09 Aug 2005
• Date of issue
  June 2005

History

• Received
  10 June 2004
• Accepted
  15 Mar 2005