Article pubs.acs.org/jnp
Acetylated Dammarane-Type Bisdesmosides from Combretum inflatum Russell B. Williams,* Vanessa L. Norman, Matt G. Goering, Mark O’Neil-Johnson, Gary R. Eldridge, and Courtney M. Starks Lead Discovery and Rapid Structure Elucidation Group, Sequoia Sciences, Inc., 1912 Innerbelt Business Center Drive, St. Louis, Missouri 63114, United States S Supporting Information *
ABSTRACT: The first study of the chemical constituents of Combretum inf latum has resulted in the isolation of seven new acetylated dammarane-type bisdesmosides (1−7). Their structures were determined from microgram quantities on hand using Bruker BioSpin TCI 1.7 mm MicroCryoProbe technology, ESIMS, and comparison to data found in the literature. Compounds 1−7 were screened for inhibition of an Escherichia coli strain UTI89 biofilm, MRSA inhibition, and cytotoxicity in NCI-H460 human lung cancer cells. Compounds 3−7 reduced the growth of MRSA at 16 μg/mL by 71−45%, and compound 7 had an IC50 value of 3.9 μM in NCI-H460.
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he genus Combretum (Combretaceae) has been a popular focus for natural product researchers owing to the extensive use of these plants in traditional African medicine and the subsequent identification of the combretastatins.1 One species native to Central Africa, C. inf latum Jongkind, was described in 1991, and until now no studies on its chemical constituents have been published.2 Screening of the library of plant natural product preparative HPLC fractions of Sequoia Sciences against an Escherichia coli (UTI89) biofilm revealed two consecutive antimicrobial fractions from the leaves of C. inf latum collected from the Haut-Ogooué region of Gabon by Missouri Botanical Garden representatives. Seven compounds were further purified from the active fractions. Despite the small amounts isolated (80% at 60 μg/mL). The F5 library was collected using a preparative HPLC method that employed a 5% to 10% (2−12 min) to 40% (12−36 min) to 85% (36−42 min) CH3CN gradient in H2O (20 mL/min) on an Aquasil C18 column (Thermo Scientific, 21.2 mm × 100 mm, 5 μm). HPLC solvents contained 0.05% TFA. Forty fractions were collected from 2 to 42 min to form the F5 library. Active fractions 38 and 39 (F5S38 and F5S39) eluted at 39−41 min. The two fractions contained different ratios of the same compounds based on analytical HPLC, so they were combined. The combined fraction (F5S38 and 39) was then further separated using semipreparative C18 HPLC eluted at 3 mL/min with a 55% to 95% CH3CN in H2O gradient from 5 to 50 min (Synergi Hydro, Phenomenex 250 mm × 10 mm, 4 μm). HPLC solvents contained 0.05% TFA. Serial collections afforded 1 (tR 14.9 min, 99 μg), 2 (tR 15.5 min, 82 μg), 3 (tR 28.5 min, 35 μg), 4 (tR 29.3 min, 29 μg), 7 (tR 30.6 min, 38 μg), 5 (tR 36.1 min, 187 μg), and 6 (tR 36.9 min, 163 μg). Combretaside A (1). Amorphous white solid; insufficient material was available to obtain specific rotation; 1H and 13C NMR, see Table 1 and 2; LRESIMS m/z 827 [M − H]−; HRESIMS m/z 827.5178 [M − H]− (calcd for C44H75O14, 827.5157). Combretaside B (2). Amorphous white solid; insufficient material was available to obtain specific rotation; 1H and 13C NMR, see Table 1 and 2; LRESIMS m/z 827 [M − H]−; HRESIMS m/z 827.5203 [M − H]− (calcd for C44H75O14, 827.5157). Combretaside C (3). Amorphous white solid; insufficient material was available to obtain specific rotation; 1H and 13C NMR, see Table 1 and 2; LRESIMS m/z 810 [M − H]−; HRESIMS m/z 809.5005 [M − H]− (calcd for C44H73O13, 809.5051). Combretaside D (4). Amorphous white solid; insufficient material was available to obtain specific rotation; 1H and 13C NMR, see Table 1 E
dx.doi.org/10.1021/np4002652 | J. Nat. Prod. XXXX, XXX, XXX−XXX
Journal of Natural Products
Article
screened as above at 60 μg/mL. Fractions demonstrating inhibition greater than 80% compared to control wells were considered active.
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ASSOCIATED CONTENT
S Supporting Information *
NMR spectra for compounds 1−7. This material is available free of charge via the Internet at http://pubs.acs.org.
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AUTHOR INFORMATION
Corresponding Author
*Tel: +1-314-373-5181. Fax: +1-314-373-5186. E-mail:
[email protected]. Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS Sequoia Sciences gratefully acknowledges the government of Gabon and Madam Nze at IPHA-METRA/CENAREST for permission to collect plants in Gabon. Sequoia Sciences acknowledges J. Stone, A. Bradley, G. Walters, J. Miller (now at the New York Botanical Garden), and R. Bussmann from Missouri Botanical Garden for collection and identification of the plant source of the natural products. The authors acknowledge the strategic collaboration between Bruker Biospin and Sequoia involving the Bruker BioSpin TCI 1.7 mm MicroCryoProbe. We also acknowledge the scientific collaboration with Advanced Chemistry Development, Inc. (ACD Laboratories) and the use of ACD/SpecManager and ACD/Structure Elucidator.
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REFERENCES
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dx.doi.org/10.1021/np4002652 | J. Nat. Prod. XXXX, XXX, XXX−XXX
