Anti-Tuberculosis Constituents from the Stem Bark of Micromelum hirsutum

Abstract

Anti-TB bioassay-directed fractionation led to the isolation of six carbazole alkaloids, as well as the γ-lactone derivative of oleic acid, from the CH₂Cl₂ extract of the stem bark of Micromelum hirsutum. The carbazoles include the new micromeline (2) and five known alkaloids: lansine (3), 3-methylcarbazole (4), methyl carbazole-3-carboxylate (5), 3-formylcarbazole (6), and 3-formyl-6-methoxycarbazole (7). Compound 1 was identified as the lactone derivative of oleic acid, (-)-Z-9-octadecene-4-olide, for which the trivial name micromolide (1) is suggested. It showed potent in vitro anti-TB activity against H37Rv (MIC: 1.5 μg/mL), a selectivity index (SI) of 63, and exhibited activity against the Erdman strain of M. tuberculosis in a J774 mouse macrophage model (EC₉₀ : 5.6 μg/mL). Thus, 1 appears worthy of further evaluation as a potential new anti-TB agent. Isolates 2, 3, 6 and 7 had anti-TB MIC values between 14.3 and 42.3 μg/mL, while compounds 4 and 5 were considered inactive (MIC > 128 μg/mL). Structure elucidation and identification were based on spectroscopic analysis, including MS, 1D/2D NMR, and a full ¹H spin system analysis of 1.

Key words

Micromelum hirsutum - Rutaceae - (-)-Z-9-octadecene-4-olide - micromeline - micromolide - carbazole alkaloids - anti-TB activity

References

• 1 Smith C V, Sharma V, Sacchettini J C. TB drug discovery: addressing issues of persistence and resistance.  Tuberculosis. 2004;  84 45-55
• 2 Soejarto D D, Gyllenhaal C, Regalado J C, Pezzuto J M, Fong H HS, Tan G T. et al . Studies on biodiversity of Vietnam and Laos: The UIC-based ICBG program.  Pharm Bio. 1999;  37 (Supplement) 100-13
• 3 Collins L S, Franzblau S G. Microplate Alamar blue assay versus BACTEC 460 system for high-throughput screening of compounds against Mycobacterium tuberculosis and Mycobacterium avium .  Antimicrob Agents Ch. 1997;  41 1004-9
• 4 Cantrell C L, Lu T, Fronczek F R, Fischer N H, Adams L B, Franzblau S G. Antimycobacterial cycloartanes from Borrichia frutescens .  J Nat Prod. 1996;  59 1131-6
• 5 Bohlmann F, Abraham W R. Neue prenylfavanone aus Helichrysum hypocephalum .  Phytochemistry. 1979;  18 1851-3
• 6 Cosse A A, Bartelt R J, James D G, Petroski R J. Identification of a female-specific, antennally active volatile compound of the currant steam girdler.  J Chem Ecol. 2001;  27 1841-53
• 7 Jaki B, Franzblau S G, Pauli G F. An NMR method towards routine chiral determination of natural products. Phytochemical Analysis 2004: in press
• 8 Jakupovic J, Zdero C, Grenz M, Tsichritzis F, Lehmann L, Hashemi-Nejad S M, Bohlmann F. Twenty-one acylphloroglucinol derivatives and further constituents from south African Helichrysum species.  Phytochemistry. 1989;  28 1119-31
• 9 Gunstone F D. High-resolution ¹³C NMR spectroscopy of lipids. 1993 The Oily Press Dundee; p 35
• 10 Li W S, McChesney J D, El-Feraly F S. Carbazole alkaloids from Clausena lansium .  Phytochemistry. 1991;  30 343-6
• 11 Wu T S, Huang S C, Wu P L, Teng C M. Carbazole alkaloids from Clausena excavata and their biological activity.  Phytochemistry. 1996;  43 133-40
• 12 Prakash D, Raj K, Kapil R S, Popli S P. Chemical constituents of Clausena lansium: part 1 - structure of lansamide-I and lansine.  Indian J Chem. 1980;  19B 1075-6
• 13 Chakrabarty M, Batabyal A. Indolisation of cyclohexanone phenylhydrazones using phosphorous trichloride.  Indian J Chem. 1992;  31B 199-201
• 14 Chakraborty D P, Das K C, Basak S P. New syntheses of isomeric methyl carbazoles.  J Indian Chem Soc. 1968;  45 84-6
• 15 Sunthitikawinsakul A, Kongkathip N, Kongkathip B, Phonnakhu S, Daly J W, Spande T F, Nimit Y, Rochanaruangraj S. Coumarins and carbazoles from Clausena excavata exhibited antimycobacterial and antifungal activities.  Planta Medica. 2003;  69 155-7
• 16 Chadwick L R. Ph. D. Dissertation: Estrogens and congeners from spent hops. University of Illinois at Chicago: Department of Medicinal Chemistry and Pharmacognosy Chicago; 2004
• 17 Orme I. Search for new drugs for treatment of tuberculosis.  Antimicrob Agents Ch. 2001;  45 1943-6
• 18 Pauli G F. qNMR - A versatile concept for the validation of natural product reference compounds.  Phytochemical Analysis. 2001;  12 28-42