Larvicidal, Antimycobacterial and Antifungal Compounds from the Bark of the Peruvian Plant Swartzia polyphylla DC

Abstract

Perú is a country with a large number of medicinal plants, many of which are used for the treatment of infectious diseases , 1) although only few studies have been conducted to prove their efficacy and safety. 2,3) Due to the emergence of microorganism resistance to the common antibiotics 4) and its worldwide impact on health, our research aims to identify new natural products that may lead to the discovery of new antibacterial agents with higher efficiency and lower toxicity. As part of our continuing work on bioactive compounds from Peruvian medicinal plants, 5-7) the in vitro antimycobac-terial activity of 102 ethanol extracts from 84 plants-used traditionally in Perú for the treatment of inflammatory or infectious disorders-was screened using a tetrazolium mi-croplate assay (TEMA). 8) In a separate screening, the anti-fungal and larvicidal activities of over 100 plants, including those previously tested for antimycobacterial activity, were bio-assayed. As a result of these screenings, Swartzia polyphylla DC (Fabaceae) was found to exhibit powerful antimycobacterial action against the sensitive H 37 Rv and multidrug-resistant Mycobacterium tuberculosis. It also inhibited the in vitro growth of the dermatophyte Trichophyton mentagrophytes, and was active against the larvae of the mosquito Culex quin-quefasciatus. We are now pleased to report the isolation of the larvicidal, antimycobacterial and antifungal principles present in the crude extract of S. polyphylla. A solvent-partition of the 95% ethanol extract showed that the larvicidal and antimycobacterial activities were concentrated in the hexane fraction, while the 90% methanol fraction was active only in the antifungal assay. The hexane fraction (28 g) was chromatographed on a silica gel column using a hexane-chloroform-methanol gradient. Each fraction (F1-F7) was evaluated for larvicidal and antimycobacterial activity in vitro. The most active fraction (F6, 8.7 g) was purified by column chromatography using a hexane-dichloromethane-ethanol gradient and then by MPLC (Lobar Lichroprep silica gel RP-8, 40-63 mm, 31025 mm, Merck) with acetonitrile-methanol-water (3 : 2 : 2) yielding the most active fraction F-6-4-2 (51.4 mg). This fraction was finally purified by HPLC (Waters Nova-pak H R silica 6 mm, 3.9300 mm, Waters Model 600E with Waters 2996 PDA detector) using hexane-chloroform gradient (0 to 70%) to obtain T-cadinol (1, 9 mg) (Fig. 1). 9,10) The 90% methanol fraction (25.5 g) was subjected to column chromatography (silica gel, 0.063-0.200 mm) using a hexane-chloroform-methanol gradient. Each fraction (F1-F9) was tested for antifungal activity in vitro. The most active fraction (F3) yielded fraction F3-7-5 (756 mg) after repeated column chromatography using hexane-ethylacetate-methanol as eluent. This fraction was finally purified by MPLC with methanol-water (6 : 4) to afford biochanin A (2, 15 mg) and dihydrobiochanin A (3, 59 mg) (Fig. 1). 11) The bark of S. polyphylla contains various flavonoids and isoflavones, some of them with strong activity against carcinogenic bacteria. 11-13) A bioassay-guided isolation of the 95% ethanol extract of S. polyphylla afforded the compound T-cadinol, which showed a moderate anti-Mycobacterium tuberculosis activity (MIC50 mg/ml for the sensitive and multidrug-resistant strains) and strong larvicidal activity. T-cadinol, at a concentration of 300 mg/ml, produces 100% mortality of the larvae of C. quinquefasciatus after 1 h exposure. Biochanin A (2) and dihydrobiochanin A (3) are responsible for the antifungal activity present in the ethanol extract of S. polyphylla (Table 1). Both compounds are very active especially against filamentous fungi. The remaining fractions obtained through the bioassay-guided isolation studies were devoid of larvicidal, antimycobacterial or antifungal activities. 278 Notes Chem. The 95% ethanol extract of the bark of Swartzia polyphylla DC (Fabaceae) possesses important larvicidal, antimycobacterial and antifungal activity in vitro. Bioassay-guided studies performed on the crude ethanol extract afforded T-cadinol as the larvicidal and anti-Mycobacterium tuberculosis principle, while the antifungal activity of the extract is due to the presence of the flavonoids biochanin A and dihydrobiochanin A.