Maytensifolone, a new triterpene from Maytenus distichophylla Mart. ex Reissek

SHORT REPORT

Maytensifolone, a new triterpene from Maytenus distichophylla Mart. ex Reissek

Marcelo Cavalcante Duarte^I; Josean Fechine Tavares^I; Sara Alves L. Madeiro^I; Vicente Carlos O. Costa^I; José Maria Barbosa Filho^I; Maria de Fátima Agra^II; Raimundo Braz Filho^III; Marcelo Sobral da Silva^I,*

^IDepartamento de Ciências Farmacêuticas and ^IICentro de Biotecnologia, Departamento de Biotecnologia, Universidade Federal da Paraíba, CP 5009, 58051-970 João Pessoa-PB, Brazil

^IIILaboratório de Ciências Químicas, Universidade Estadual do Norte Fluminense, Campos dos Goytacazes, 28013-602 Rio de Janeiro-RJ, Brazil

ABSTRACT

Phytochemical study of the leaves of Maytenus distichophylla Mart. ex Reissek led to the isolation of the new triterpene 3,16,21-trioxo-6β,12α-dihydroxy-1-en-friedelane, named maytensifolone, along with the known triterpenes 3-oxofriedelane, 3,12-dioxofriedelane, 3β-hydroxyfriedelane, 3-oxo-29-hydroxyfriedelane, 3-oxo-12α-hydroxyfriedelane and 3-oxo-30-hydroxyfriedelane. Their structural identification was based on spectroscopic methods and comparison with literature data.

Keywords: Celastraceae, Maytenus distichophylla, maytensifolone, friedelane triterpene

RESUMO

O estudo fitoquímico das folhas de Maytenus distichophylla Mart. ex Reissek levou ao isolamento de um novo triterpeno 3,16,21-trioxo-6β,12α-dihidroxi-1-en-friedelano, nomeado maytensifolona, juntamente com os triterpenos conhecidos, 3-oxofriedelano, 3,12-dioxofriedelano, 3β-hidroxifriedelano, 3-oxo-29-hidroxifriedelano, 3-oxo-12α-hidroxifriedelano e 3-oxo-30-hidroxifriedelano. A identificação estrutural foi baseada em métodos espectroscópicos e comparação com dados da literatura

Introduction

The family Celastraceae is composed of 90 genera and approximately 1300 species.¹ The genus Maytenus comprises about 80 species distributed throughout Brazil.² They are chemically characterized mainly by the presence of flavonoids and pentacyclic triterpenes, which are considered taxonomic markers for this genus.^3-5 Various pharmacological activities have been reported for triterpenes isolated from Maytenus, such as antiulcerogenic and antifungal.^6,7

In previous works, we reported the isolation and structural characterization of triterpenes and flavonoids from M. obtusifolia,³ as well as their toxicity and antiulcerogenic activity.⁸ In continuing our work on Maytenus sp., we conducted a phytochemical study of M. distichophylla Mart. ex Reissek, a species that was not previously subjected to chemical or pharmacological studies. Accordingly, seven triterpenes of the friedelane group were isolated and characterized (Figure 1), including the new 3,16,21-trioxo-6β,12α-dihydroxy-1-en-friedelane, here named as maytensifolone (1), and known triterpenes 3-oxofriedelane (2),⁹ 3,12-dioxofriedelane (3),¹⁰ 3β-hydroxyfriedelane (4),¹¹ 3-oxo-29-hydroxyfriedelane (5),¹² 3-oxo-12α-hydroxyfriedelane (6)¹³ and 3-oxo-30-hydroxyfriedelane (7)¹⁴ which are being reported for the first time in this species.

Results and Discussion

Compound 1 was isolated in the form of a white amorphous solid, mp 310-312 ºC, [α]_D²⁵ +1.4, (c. 0.001 in CHCl₃). The high resolution mass spectrum utilizing the ESI⁺ ionization mode showed a quasi-molecular peak at m/z 485.3306 [M + H]⁺, compatible with molecular formula C₃₀H₄₄O₅ (calc. 485.3261). The infrared (IR) spectrum showed absorptions in the region of 3504 cm^-1 (hydroxyl group), 1681 cm^-1 (ketone carbonyl) and 1662 cm^-1 (α,β unsaturated carbonyl). The ¹³C APT (attached proton test) NMR spectrum (125 MHz, CDCl₃ + C₅D₅N) showed 29 signals corresponding to 30 carbon atoms: eight methyl, five methylene, eight methine and nine non-hydrogenated, in line with a triterpene skeleton of the friedelane type.⁴ The signals at d_C 200.87, 146.28 and 130.14 were attributed to C-3, C-1 and C-2, respectively, consistent with an α,β unsaturated carbonyl system of friedelane triterpenes.⁹ The signals at d_C 77.25 and 69.32 were attributed to C-6 and C-12, respectively.^4,15 The location of the hydroxyl groups at C-6 and C-12 was corroborated by the chemical shift at 8.69, corresponding to the methyl groups CH₃-24 and CH₃-27, both subject to a γ effect. Also, this spectrum displayed signals at d_C 214.24 and 218.13 attributed to carbons C-16 and C-21.¹⁵ The ¹H NMR spectrum showed signals at d_H 0.74 (s), 0.84 (s), 0.89 (s), 0.91 (s), 1.04 (s), 1.09 (s), 1.11 (s), and 1.17 (d, 1H, J 6.5 Hz), attributed to eight methyl groups. In agreement with literature, the doublet at d_H 1.17 is consistent with CH₃-23 of Δ¹ friedelanes.¹⁶ In addition, signals were observed at d_H 6.65 (d, 1H, J 10.5 Hz, H-1), 5.92 (dd, 1H, J 10.5, 3.0 Hz, H-2), 3.63 (dd, 1H, J 9.5, 5.0 Hz, H-6ax) and 3.94 (dd, 1H, J 11.0, 4.0 Hz, H-12ax), thereby inferring the equatorial orientation of the hydroxyl groups bound to C-6 and C-12. Observed HMQC (heteronuclear multiple quantum coherence) correlations are shown in Table 1. In the long-range (HMBC) ¹H-¹³C NMR correlation spectrum, the following correlations were observed: d_H 2.23 (H-4) with carbons at d_C 200.87, 49.10, 60.35, 8.69 and 9.80 confirming the attributions of C-3, C-5, C-10, CH₃-24 and CH₃-23, respectively; d_H 0.74 (CH₃-24) with carbons at d_C 77.25 and 57.72, corresponding to C-6 and C-4, respectively; d_H 2.08 (H-10) with the carbon at d_C 19.57 (CH₃-25) and d_H 0.89 (CH₃-25) with carbons at d_C 48.19, 36.66 and 45.37, which were attributed to C-8, C-9 and C-11, respectively; d_H 1.40 (H-8) with carbons at d_C 19.93 and 49.56 (CH₃-26 and C-15), respectively; d_H 1.82/1.17 (2H-11) with the carbon at d_C 69.32 (C-12); and d_H 0.84 (CH₃-27) with carbons at d_C 44.66 (C-18) and 69.32 (C-12). The other correlations are given in Table 1. The relative stereochemistry was determined by NOESY (nuclear Overhauser effect spectroscopy) and is shown in Figure 2, confirming the equatorial orientation of the hydroxyls at C-6 and C-12. The above findings support the structure of 1 as 3,16,21-trioxo-6β,12α-dihydroxy-1-en-friedelane, a new natural product named as maytensifolone.

The other compounds were identified by comparison of their spectroscopic data with those described in literature, as 3-oxofriedelane (2),⁹ 3,12-dioxofriedelane (3),¹⁰ 3β-hydroxyfriedelane (4),¹¹ 3-oxo-29-hydroxyfriedelane (5),¹² 3-oxo-12α-hydroxyfriedelane (6)¹³ and 3-oxo-30-hydroxyfriedelane (7),¹⁴ and are reported here for the first time in M. distichophylla.

Experimental

General experimental procedures

Melting points were determined with a digital apparatus model MQAPF-302 from Microchemical and were not corrected. IR spectra were recorded on a BOMEM-MB 100 spectrophotometer. One-dimensional (¹H and ¹³C) and two-dimensional (gHMQC, gHMBC, gCOSY and gNOESY) NMR analyses were performed on a Varian-System spectrometer operating at 500 MHz (¹H) and 125 MHz (¹³C). CDCl₃ and C₅D₅N were used as solvents with TMS (tetramethylsilane) as an internal standard. HRESIMS (high resolution electrospray ionization mass spectrometry) was obtained using a micrOTOF-II system from Bruker. Silica gel 60 (0.063-0.20 and 0.04-0.063 mm; Merck), was used in column chromatography (CC), whereas silica gel TLC (thin layer chromatography) plates PF254 7749 (Merck) stained with iodine vapor and viewed under UV light (254/366 nm) were used to monitor chromatographic purification procedures.

Plant material

The botanical material utilized was collected in the Matureia city (Paraíba, Brazil) in June 2009, and identified by Prof. Dra. Maria de Fatima Agra. A dried specimen is deposited in the Herbarium Professor Lauro Pires Xavier at Universidade Federal da Paraíba (Paraíba, Brazil) under No. 7448.

Extraction and isolation

The leaves of M. distichophylla (3.5 kg), dried and pulverized, were extracted with 95% ethanol at room temperature for 3 days. The obtained extract was concentrated in a rotary evaporator under reduced pressure at 40 ºC, yielding 685.0 g of ethanolic extract. A portion (100.0 g) was suspended in MeOH:H₂O (7:3) and partitioned successively with n-hexane, CHCl₃ and EtOAc to obtain the n-hexane (2.5 g), chloroform (5.4 g) and ethyl acetate (6.5 g) fractions. The ethyl acetate fraction (5.4 g) was separated by CC, on silica gel 60 (0.063-0.200 mm), eluted with n-hexane, EtOAc and MeOH, pure or in binary mixtures, in increasing order of polarity, resulting in 110 fractions of 100 mL each, which were submitted to analytical NMR. Fraction 14 eluted with n-hexane:EtOAc (7:3) yielded compound 2 (50.3 mg). Fractions 1-2 (168.3 mg) were submitted to another CC utilizing similar conditions as before, providing 25 subfractions of 10 mL each. Subfractions 10-15 eluted with n-hexane:EtOAc (8:2) gave the triterpene friedelane 1 (13.4 mg). Fractions 27-35 (99.3 mg) were rechromatographed as before, resulting in 55 subfractions of 10 mL each. Subfractions 33-37 eluted with n-hexane:EtOAc (6:4) yielded 3 (26.2 mg).

The chloroform fraction (5.0 g) was submitted to CC, on silica gel 60 (0.063-0.200 mm), eluted with hexane, EtOAc and MeOH, pure or in binary mixtures and with increasing order of polarity, resulting in 110 fractions of 100 mL each, which were concentrated in a rotary evaporator and submitted to analytical NMR. Fractions 42-45 eluted with n-hexane:EtOAc (8:2) provided compound 4 (22.7 mg). Fractions 67-81 (675.3 mg) were submitted to another CC, utilizing a column packed with silica gel 60 (0.04-0.063 mm) and the eluents n-hexane and EtOAc and MeOH, resulting in 53 subfractions of 100 mL each, which were submitted to analytical NMR and combined into 10 groups. Subfractions 2-3 eluted with n-hexane:EtOAc (85:15) yielded compounds 5 (8.5 mg), 6-7 eluted with n-hexane:EtOAc (6:4) yielded compound 6 (5.0 mg) and 9-10 eluted with n-hexane:EtOAc (1:1) yielded compound 7 (9.0 mg).

3,16,21-Trioxo-6β,12α-dihydroxy-1-en-friedelane (1)

White amorphous powder; mp 310-312 ºC; IR (KBr) ν_max/cm^-1 at 3504, 1681, 1662; HRESIMS m/z 485.3306 [M + H]⁺, (calc. for C₃₀H₄₄O₅, 485.3261); ¹H NMR (500 MHz, CDCl₃ + C₅D₅N) and ¹³C NMR (125 MHz, CDCl₃ + C₅D₅N), see Table 1.

Supplementary Information

Supplementary data associated with this paper are available free of charge at http://jbcs.sbq.org.br as a PDF file.

Acknowledgments

The authors thank CNPq, CAPES, FAPESQ-PB and INSA for financial support and LMCA-Central Analitica of UFPB for providing the spectra. Dr. A. Leyva helped with English revision.

Submitted: May 17, 2013

Published online: August 23, 2013

Supplementary Information

The supplementary material is available in pdf: [Supplementary material]

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Publication Dates

• Publication in this collection
  02 Oct 2013
• Date of issue
  Oct 2013

History

• Received
  17 May 2013
• Accepted
  23 Aug 2013