Prospection and identification of nematotoxic compounds from Canavalia ensiformis seeds effective in the control of the root knot nematode Meloidogyne incognita

Abstract

Meloidogyne incognita is responsible for significant crop losses worldwide. The main strategy to control this phytopathogen is still based on synthetic nematicides that are harmful to human health and the environment. In this context, aqueous seed extracts of antagonistic plants were screened for molecules effective toward the infective stage (J2) of M. incognita. The aqueous crude extract of Canavalia ensiformis (ACECe) showed the highest nematicidal activity (87±3% mortality). ACECe dialysis fractionation allowed the collection of an external dialysate (EDCe) containing molecules smaller than 3.5kDa effective against J2 (96±3.0% mortality); innocuous toward non targeted organisms as saprophytic nematodes, fungi, bacterium and insects larvae; active against J2 (96±2% mortality) after heating (50°C); no cytotoxic for bovine red blood cells; reduction of M. incognita eggs masses by 82.5% in tomato plants at green house conditions. Fractionation of the EDCe by reversed-phase high-performance liquid chromatography (RP-HPLC) separated five nematotoxic fractions. Analyses of those fractions based on gas chromatography–mass spectrometry (GC–MS), liquid chromatography–mass spectrometry (LCMS) and nuclear magnetic resonance (NMR) identified nine specific metabolites, follow-up testing of the individual authentic standards of each metabolite as the compounds: d-glucose, l-canavanine, xanthotoxin, cis-aconitic acid, trans-aconitic acid, malic acid, citric acid, palmitic acid and S-carboxymethylcysteine; all them confirmed to possess nematotoxic properties. However, some of those metabolites were not described previously demonstrating biological action against M. incognita.