Next Article in Journal
Deciphering the Role of Phytoalexins in Plant-Microorganism Interactions and Human Health
Next Article in Special Issue
Anti-Inflammatory Effect of Methylpenicinoline from a Marine Isolate of Penicillium sp. (SF-5995): Inhibition of NF-κB and MAPK Pathways in Lipopolysaccharide-Induced RAW264.7 Macrophages and BV2 Microglia
Previous Article in Journal
Exogenous Spermidine Improves Seed Germination of White Clover under Water Stress via Involvement in Starch Metabolism, Antioxidant Defenses and Relevant Gene Expression
Previous Article in Special Issue
Flavonoids from Gynostemma pentaphyllum Exhibit Differential Induction of Cell Cycle Arrest in H460 and A549 Cancer Cells
Article Menu

Export Article

Molecules 2014, 19(11), 18025-18032; doi:10.3390/molecules191118025

Article
Dysidinoid A, an Unusual Meroterpenoid with Anti-MRSA Activity from the South China Sea Sponge Dysidea sp.
1
Key Laboratory for Marine Drugs, Department of Pharmacy, Renji Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai 200127, China
2
College of Food and Biological Engineering, Jimei University, Xiamen 361021, China
3
Department of Clinical Laboratory, Renji Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai 200127, China
4
Laboratory of Marine Drugs, Department of Pharmacy, Changzheng Hospital, Second Military Medical University, Shanghai 200003, China
These authors contributed equally to this work.
*
Author to whom correspondence should be addressed; Tel.: +86-21-6838-3346; Fax: +86-21-5873-2594.
Received: 19 October 2014; in revised form: 30 October 2014 / Accepted: 30 October 2014 / Published: 5 November 2014

Abstract

:
An unusual meroterpenoid, dysidinoid A (1), was isolated from the South China Sea sponge Dysidea sp. Its structure was elucidated by extensive spectroscopic methods including HRESIMS and 2D NMR, and its absolute configuration was determined by single-crystal X-ray diffraction analysis. Dysidinoid A (1) is the first meroterpenoid from Nature bearing a 9,4-friedodrime skeleton and a 2,5-dionepyrrole unit. Dysidinoid A (1) showed potent antibacterial activity against two strains of pathogenic bacteria methicillin-resistant Staphylococcus aureus (MRSA) with MIC90 values of 8.0 μg/mL against both.
Keywords:
dysidinoid A; meroterpenoid; anti-MRSA; Dysidea; marine sponge

1. Introduction

Infectious diseases are the leading cause of death worldwide. Emerging infections due to methicillin resistant Staphylococcus aureus (MRSA) pose a significant threat to patients [1,2]. It has been estimated that in the United States more people die from MRSA related infections than from HIV [3]. Infections involving drug resistant bacteria are more difficult to treat due to increased costs and decreased efficacies [4,5]. One important approach to drug discovery for the treatment of MRSA is through natural products research.
Marine sponges of the genus Dysidea (order Dictyoceratida, family Dydideidae) have proven to be prolific producers of structurally diverse secondary metabolites, such as sesequiterpene quinones [6,7,8], sesquiterpenoids [9], diterpenoids [10], sterols [11], and polychlorinated compounds [12,13,14]. These metabolites showed a spectrum of interesting biological activities, including antifungal [15], antibacterial [16], antitumor [17,18], anti-inflammatory [15,19], and antioxidative activities [20].
In our efforts to search for new anti-MRSA agents from marine sponges collected from the South China Sea, chemical investigation of an active fraction from the sponge Dysidea sp. resulted in the isolation of a novel meroterpenoid, dysidinoid A (1) (Figure 1). It is the first meroterpenoid from Nature bearing a 9,4-friedodrime skeleton and a 2,5-dionepyrrole. Antibacterial evaluation showed that dysidinoid A showed potent antibacterial activity against two strains of pathogenic bacteria MRSA with MIC90 values of 8.0 μg/mL against both. Details of structural elucidation and antibacterial activity of dysidinoid A (1) were reported herein.
Figure 1. The chemical structure of dysidinoid A (1).
Figure 1. The chemical structure of dysidinoid A (1).
Molecules 19 18025 g001

2. Results and Discussion

Dysidinoid A (1) was obtained as colorless needles with Molecules 19 18025 i001 +35.4 (c 0.50, MeOH). Its IR spectrum showed absorption bands assignable to amide (3276 cm−1) and carbonyl (1775 and 1714) functionalities. The positive ESIMS of 1 exhibited quasimolecular ion peaks at m/z 302.2 [M+H]+ and 324.2 [M+Na]+, respectively. The molecular formula of C19H27NO2 with seven degrees of unsaturation, was deduced from HRESIMS at m/z 324.1941 [M+Na]+ (calcd. for C19H27NO2, 324.1939), which was supported by the 1H- and 13C-NMR data (Table 1). The 1H-NMR spectrum of 1 showed resonances attributable to two olefinic protons at δH 5.16 (H-3) and 6.26 (H-18), three tertiary methyl groups at δH 1.55 (H3-11), 1.00 (H3-12), and 0.88 (H3-14), a secondary methyl group at δH 0.95 (H3-13). In addition, the spectrum showed resonances due to an exchangable amine proton at δH 7.33 (20-NH), as well as partially overlapping signals with complex coupling patterns between δH 1.08 and 2.61 that could be attributed to several aliphatic methylene and methine units. The 13C-NMR and DEPT spectra of 1 showed 19 carbon resonances, corresponding to two carbonyl groups (δC 171.7 and 170.4), two olefinic quaternary carbons (δC 143.9 and 147.9), two aliphatic quaternary carbons (δC 38.3 and 42.4), two olefinic methine carbons (δC 120.5 and 130.4), two aliphatic methine carbons (δC 37.4 and 47.0), five aliphatic methylene carbons (δC 19.0, 26.3, 36.2, 27.4, and 32.5), and four methyl carbons (δC 17.7, 19.8, 16.3, and 18.0). The above spectroscopic signatures suggested the presence of a 9,4-friedodrime sesquiterpene moiety and accounted for four degrees of unsaturation, indicating three rings in the structure of 1.
Table 1. The 1H- (600 MHz) and 13C- (150 MHz) NMR data of compound 1 in CDCl3. a
Table 1. The 1H- (600 MHz) and 13C- (150 MHz) NMR data of compound 1 in CDCl3. a
PositionδCδH (J in Hz)HMBC (H→C)NOESY
19.0, CH21.83, mC-2, 3, 5, 9, 10H-1β, 2β, 10
1.53, mC-2, 5, 10H3-12, 14, H-1α, 2β
26.3, CH21.93, mH-1α, 1β, 10
2.07, mC-3, 4, 10H-1α, 1β, 2α
3120.5, CH5.16, br sC-5, 11H3-11, H-2α, 2β
4143.9, C
538.3, C
36.2, CH21.08, mC-8H-6β, 7a, 8, 10
1.68, dt (12.8, 3.4)C-7, 8, 10, 12H3-11, 12, H-6α, 7b
7a27.4, CH21.41, mC-5, 6, 9, 13H-6, 8
7b1.40, dd (6.9, 3.5)H-6, 8, H3-12, 13, 14
837.4, CH1.28, mC-7, 9, 13H-7b, 10, H3-13
942.4, C
1047.0, CH1.12, dd (12.4, 1.6)C-2, 4, 5, 9, 12, 14, 15H-1α, 2α, 8, 15α, 15β
1117.7, CH31.55, br sC-3, 4, 5H3-12, H-3
1219.8, CH31.00, sC-4, 5, 6, 10H3-11, 14, H-6β, 7β
1316.3, CH30.95, d (6.7)C-7, 8, 9H3-14, H-7β, 8
1418.0, CH30.88, sC-8, 9, 10, 15H3-12, 13, H-1β, 7β
15α32.5, CH22.61, d (14.1)C-8, 9, 10, 14, 16, 17, 18H3-14, H-1α, 10
15β2.43, dd (14.1, 1.2)C-8, 9, 10, 14, 16, 17, 18H3-13
16147.9, C
17171.7, C
18130.4, CH6.26, d (1.0)C-15, 16, 19H3-13, H-10, 15α, 15β
19170.4, C
20-NH7.33, br s
a Assignments of the 13C and 1H signals were made on the basis of HSQC spectroscopic data.
Unambiguous assignment of NMR data of 1 was achieved by a combination of COSY, HSQC, and HMBC experiments, as depicted in Figure 2. In the 1H-1H COSY spectrum, the correlations of H2-1/H2-2/H-3, H2-6/H2-7/H-8/H3-13, and allylic coupling correlations of H-3/H3-11 revealed the presence of two fragments (thick lines in Figure 2). The two spin systems and their connectivity with the remaining atoms enabled assembly into the final planar structure based upon the HMBC spectrum of 1. The HMBC correlations from H3-11 to C-3, C-4, and C-5, from H3-12 to C-4, C-5, C-6 and C-10, from H3-13 to C-7, C-8, and C-9, and H3-14 to C-8, C-9, C-10, and C-15 indicated the presence of 9,4-friedodrime sesquiterpene skeleton with four methyl groups at C-4, C-5, C-8, and C-9, respectively. This assignment was confirmed by the HMBC correlations from H-10 to C-2, C-4, C-5, C-9, C-12, C-14, and C-15. Furthermore, the olefinic proton H-18 showed HMBC correlations with C-15, C-16, and C-19, in combination with the chemical shifts of the proton and carbon resonances, suggested the presence of a 2,5-dionepyrrole substructure. In addition, HMBC correlations from the methylene protons H2-15 to C-8, C-9, C-10, C-14, C-16, C-17, and C-18 supported the linkage of C-9 and C-16 via the methylene CH2-15 between the 9,4-friedodrime sesquiterpene moiety and 2,5-dionepyrrole substructure. Therefore, the gross structure of 1 was determined as shown in Figure 2.
Figure 2. Key COSY, HMBC, and NOESY correlations of dysidinoid A (1).
Figure 2. Key COSY, HMBC, and NOESY correlations of dysidinoid A (1).
Molecules 19 18025 g002
The relative configuration of 1 was deduced from NOESY correlations in combination with coupling constant values. The large coupling constant between H-1β and H-10 (J = 12.4 Hz) and the NOESY correlations of H-1β/H3-12 and H3-14 indicated the axial orientations of these protons and methyls and also revealed the trans fusion of the two six-numbered rings [15,18]. The NOESY correlation of H3-13/H3-14 and H3-12/H3-14 revealed the three methyl groups are all β-orientation, while NOESY correlations from H-8 to H-6α, and H-10 suggested the three protons were α-orientation.
Fortunately, crystals of 1 suitable for single crystal X-ray diffraction analysis were obtained from a methanol solution. The relative configuration of 1 was unambiguously established by its X-ray crystal structure (Figure 3). Besides, a final refinement of the CuKa diffraction data resulted in the assignment of the absolute configuration of 1 as 5S, 8S, 9R, and 10S.
Minimal inhibitory concentration (MIC) was detected to evaluate the antimicrobial activities of dysidinoid A (1) toward two strains of hospital-acquired methicillin-resitant Staphylococcus aureus (MRSA H0556 and MRSAH0117). Dysidinoid A (1) showed potent inhibitory activity against MRSA with MIC90 values of 8 μg/mL, and chloromycetin was used as positive control (MIC90 2 μg/mL), while methicillin was used as negative control (MIC90 128 μg/mL).
Figure 3. X-ray ORTEP drawing of dysidinoid A (1).
Figure 3. X-ray ORTEP drawing of dysidinoid A (1).
Molecules 19 18025 g003

3. Experimental Section

3.1. General Experimental Procedures

Optical rotations were recorded on an Autopol I polarimeter (No. 30575, Rudolph Research Analytical, Perkin-Elmer, Inc., Waltham, MA, USA) with a 10 cm length cell at room temperature. UV and IR (KBr) spectra were recorded on a Hitachi U-3010 spectrophotometer (Hitachi, Inc., Tokyo, Japan) and Jasco FTIR-400 spectrometer (Jasco Inc., Tokyo, Japan), respectively. 1H, 13C, DEPT135, COSY, HSQC, HMBC, and NOESY NMR spectra were recorded at room temperature on a Bruker Avance DRX-600 MHz NMR spectrometer ((Bruker Biospin Corp., Billerica, MA, USA) with CDCl3 as the solvent. HRESIMS spectra were measured on an Agilent 6210 LC/MSD TOF mass spectrometer (Agilent, Milford, MA, USA). Column chromatography was conducted using pre-coated silica gel (65 × 250 or 230 × 400 mesh). Sephadex LH-20 was purchased from Amersham Pharmacia Biotech AB (Pharmacia Fine Chemicals, Piscataway, NJ, USA). Purification of the compounds was performed using a Waters Alliance 2695 separation module equipped with a Waters 2998 Photodiode Array (PDA) detector (Waters Corp., Milford, MA, USA).

3.2. Animal Material

Samples of Dysidea sp. were collected along the coast of Yongxing Island in Xiasha on 12 April 2010. The voucher number for this collection is XD10401, and a voucher sample is maintained at the Key Laboratory for Marine Drugs, Department of Pharmacy, Renji Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai, China. The sponge was identified by Professor Jin-He Li (Institute of Oceanology, Chinese Academy of Science).

3.3. Extraction, Isolation and Characterization

The animals (200 g, dry weight) were soaked in EtOH (250 mL, 25 °C, 72 h) repeatedly to give 24.6 g of a crude EtOH extract after solvent removal. The extract was dissolved in 250 mL H2O, and partitioned five times with the same volume of CH2Cl2 to yield after concentration 12.0 g of a CH2Cl2 solvent extract, The CH2Cl2-soluble fraction was subjected silica gel column chromatography eluting with a gradient of CH2Cl2 and MeOH, yielding four subfractions (D1–D4). Fraction D3 (1.4 g) was passed through an ODS chromatography column eluted with a gradient of aqueous MeOH, size-exclusion chromatography Sephadex LH-20 eluted with CH2Cl2/MeOH (1:1), and then purified by reversed-phase HPLC (YMC-Park Pro C18, 10 mm × 250 mm, 2 mL/min, 280 nm) with 65% CH3CN, to give dysidinoid A (1, 4.3 mg, tR 26.5 min); colorless needles (MeOH); Molecules 19 18025 i002 +35.4 (c 0.5, MeOH); UV (MeOH) λmax (log ε) 209 (4.05), 235 (398) nm; 1H and 13C-NMR, see Table 1; IR (KBr) υmax 3276, 2961, 2928, 2857, 1775, 1714, 1621, 1453, 1344, 1124, 1075, 871, 626 cm−1; positive ESIMS m/z 302.2 [M+H]+, 324.2 [M+Na]+; positive HRESIMS m/z 324.1941 [M+Na]+ (calcd for C19H27NO2, 324.1939).

3.4. X-ray Crystallographic Analysis of Dysidinoid A (1)

C19H27NO2, colorless blocks, M = 301.42, Orthorhombic, P21, a = 7.4098(2) Å, b = 14.0638(3) Å, c = 16.2014(3) Å, α = β = γ = 90°, V = 1688.35(7) Å3, Z = 4, Dx = 1.186 mg/m3, F (000) = 656, μ(Cu-Kα) = 0.594 mm−1, crystal dimensions 0.30 × 0.16 × 0.10 mm3 were used for measurement on a SMART CCD using graphite monochromated radiation (λ = 1.54178 Å); 5416 unique reflections were collected to θmax = 69.73°. The structure was solved by direct methods (Shelxs97) and refined by full-matrix least-squares on F2. Hydrogen atoms were located by the geometric calculation method and difference Fourier method. The final R1 = 0.0358, wR2 = 0.1083 (w = 1/σ|F|2) and S = 1.012. Crystallographic data for dysidinoid A (1) have been deposited with the Cambridge Crystallographic Data Center as supplementary publication No. CCDC 1029972. Copies of the data can be obtained, free of charge, on application to the Director, CCDC, 12 Union Road, Cambridge CB2 1EZ, UK (Fax: +44-(0)1223-336033, or E-Mail: deposit@ccdc.cam.ac.uk).

3.5. Antimicrobial Assays

Minimum inhibitory concentration (MIC) was determined according to Clinical and Laboratory Standards Institute (CLSI) guidelines. The MIC90 values were recorded using a spectrophotometer. For antibiotic sensitivity assays, bacteria in 96-well plates (Corning) were incubated with dysidinoid A (1) or antibiotic standards at final concentrations of 0 to 256 mg/mL. The plates were incubated at 37 °C and read at 24 h.

4. Conclusions

Marine sponges provide a rich source on drug discovery for the treatment of MRSA infectious diseases. In this paper, dysidinoid A (1), an unusual meroterpenoid, was isolated from the South China Sea sponge Dysidea sp. Its structure was determined based on extensive spectroscopic data, and the absolute configuration of 1 was established by single-crystal X-ray diffraction analysis. Dysidinoid A (1) showed potent antibacterial activity against two strains of hospital-acquired pathogenic MRSA with MIC90 values of 8.0 μg/mL against both.

Supplementary Materials

Supplementary materials can be accessed at: http://www.mdpi.com/1420-3049/19/11/18025/s1.

Supplementary Files

Supplementary File 1

Acknowledgments

This research was supported by the National Natural Science Fund for Distinguished Young Scholars of China (81225023), the Shanghai Rising-Star program (14QA1402800), the National Natural Science Fund of China (No. 41106127, 81302691, and 81172978), the Shanghai Subject Chief Scientist program (12XD1400200), the Fund of the Science and Technology Commission of Shanghai Municipality (14431901300), We are also grateful for the financial support of the National High Technology Research and Development Program of China (863 Projects, No. 2013AA092902).

Author Contributions

W.H.J. and J.L. contributed to the structural determination and manuscript writing. W.H.J. and T.T.X. conducted the isolation work for the new compound. J.L. and Q.L. performed the antimicrobial activity of the new compound against two strains of MRSA. G.H.S., H.B.Y., F.Y., B.N.H., M.L. gave the constructive suggestions for the experiments and manuscript writing. W.H.J. and H.W.L. conceived and designed the research.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Figueroa, M.; Jarmusch, A.K.; Raja, H.A.; El-Elimat, T.; Kavanaugh, J.S.; Horswill, A.R.; Cooks, R.G.; Cech, N.B.; Oberlies, N.H. Polyhydroxyanthraquinones as quorum sensing inhibitors from the guttates of penicillium restrictum and their analysis by desorption electrospray ionization mass spectrometry. J. Nat. Prod. 2014, 77, 1351–1358. [Google Scholar]
  2. Li, T.; Song, Y.; Zhu, Y.; Du, X.; Li, M. Current status of Staphylococcus aureus infection in a central teaching hospital in Shanghai, China. BMC Microbiol. 2003, 13. [Google Scholar] [CrossRef]
  3. Payne, D.J. Desperately seeking new antibiotics. Science 2008, 321, 1644–1645. [Google Scholar]
  4. Plaza, A.; Keffer, J.L.; Bifulco, G.; Lloyd, J.R.; Bewley, C.A. Chrysophaentins A–H, antibacterial bisdiarylbutene macrocycles that inhibit the bacterial cell division protein FtsZ. J. Am. Chem. Soc. 2010, 132, 9069–9077. [Google Scholar]
  5. Li, M.; Du, X.; Villaruz, A.; Diep, B.; Wang, D.; Song, Y.; Tian, Y.; Hu, J.; Yu, F.; Lu, Y.; et al. MRSA epidemic linked to a quickly spreading colonization and virulence determinant. Nat. Med. 2012, 18, 816–819. [Google Scholar]
  6. Jiao, W.H.; Xu, T.T.; Yu, H.B.; Chen, G.D.; Huang, X.J.; Yang, F.; Li, Y.S.; Han, B.N.; Liu, X.Y.; Lin, H.W. Dysideanones A–C, unusual sesquiterpene quinones from the South China Sea sponge Dysidea avara. J. Nat. Prod. 2014, 77, 346–350. [Google Scholar]
  7. Jiao, W.H.; Xu, T.T.; Yu, H.B.; Mu, F.R.; Li, J.; Li, Y.S.; Yang, F.; Han, B.N.; Lin, H.W. Dysidaminones A–M, cytotoxic and NF-κB inhibitory sesquiterpene aminoquinones from the South China Sea sponge Dysidea fragilis. RSC Adv. 2014, 4, 9236–9246. [Google Scholar]
  8. Marcos, I.S.; Conde, A.; Moro, R.F.; Basabe, P.; Diez, D.; Urones, J.G. Quinone/hydroquinone sesquiterpenes. Mini-Rev. Org. Chem. 2010, 7, 230–254. [Google Scholar]
  9. Ueda, K.; Kadekaru, T.; Siwu, E.R.; Kita, M.; Uemura, D. Haterumadysins A–D, sesquiterpenes from the Okinawan Marine sponge Dysidea c hlorea. J. Nat. Prod. 2006, 69, 1077–1079. [Google Scholar]
  10. Lu, Q.; Faulkner, D.J. Three dolabellanes and a macrolide from the sponge Dysidea sp. from Palau. J. Nat. Prod. 1998, 61, 1096–1100. [Google Scholar]
  11. De Almeida Leone, P.; Redburn, J.; Hooper, J.N. A.; Quinn, R.J. Polyoxygenated Dysidea sterols that inhibit the binding of [I125] IL-8 to the human recombinant IL-8 receptor type A. J. Nat. Prod. 2000, 63, 694–697. [Google Scholar]
  12. Bai, R.; Paull, K.; Herald, C.; Malspeis, L.; Pettit, G.; Hamel, E. Halichondrin B and homohalichondrin B, marine natural products binding in the vinca domain of tubulin. Discovery of tubulin-based mechanism of action by analysis of differential cytotoxicity data. J. Biol. Chem. 1991, 266, 15882–15889. [Google Scholar]
  13. Fu, X.; Ferreira, M.L.G.; Schmitz, F.J.; Kelly-Borges, M. New diketopiperazines from the sponge Dysidea chlorea. J. Nat. Prod. 1998, 61, 1226–1231. [Google Scholar]
  14. Kazlauskas, R.; Murphy, P.; Warren, R.; Wells, R.; Blount, J. New quinones from a dictyoceratid sponge. Aust. J. Chem. 1978, 31, 2685–2697. [Google Scholar]
  15. Ciavatta, M.L.; Lopez Gresa, M.P.; Gavagnin, M.; Romero, V.; Melck, D.; Manzo, E.; Guo, Y.W.; van Soest, R.; Cimino, G. Studies on puupehenone-metabolites of a Dysidea sp.: Structure and biological activity. Tetrahedron 2007, 63, 1380–1384. [Google Scholar]
  16. Urban, S.; Capon, R.J. 5-epi-Isospongiaquinone, a new sesquiterpene/quinone antibiotic from an Australian Marine sponge, Spongia hispida. J. Nat. Prod. 1992, 55, 1638–1642. [Google Scholar]
  17. Pérez-García, E.; Zubía, E.; Ortega, M.J.; Carballo, J.L. Merosesquiterpenes from two sponges of the genus Dysidea. J. Nat. Prod. 2005, 68, 653–658. [Google Scholar]
  18. Jiao, W.H.; Huang, X.J.; Yang, J.S.; Yang, F.; Piao, S.J.; Gao, H.; Li, J.; Ye, W.C.; Yao, X.S.; Chen, W.S.; et al. Dysidavarones A–D, new sesquiterpene quinones from the marine sponge Dysidea avara. Org. Lett. 2012, 14, 202–205. [Google Scholar]
  19. McNamara, C.E.; Larsen, L.; Perry, N.B.; Harper, J.L.; Berridge, M.V.; Chia, E.W.; Kelly, M.; Webb, V.L. Anti-inflammatory sesquiterpene-quinones from the New Zealand sponge Dysidea cf. cristagalli. J. Nat. Prod. 2005, 68, 1431–1433. [Google Scholar]
  20. Utkina, N.K.; Denisenko, V.A.; Krasokhin, V.B. Sesquiterpenoid aminoquinones from the marine sponge Dysidea sp. J. Nat. Prod. 2010, 73, 788–791. [Google Scholar]
  • Sample Availability: Sample of the compound 1is available from the authors.
Molecules EISSN 1420-3049 Published by MDPI AG, Basel, Switzerland RSS E-Mail Table of Contents Alert
Back to Top