Screening of panamanian plant extracts for agricultural and cosmetic activities, and metabolomic study of "Isatis tinctoria" accessions

Abstract

The FP7 project AGROCOS aimed at the discovery of new natural product scaffolds for possible uses in agrochemistry and as cosmetic ingredients. In addition, novel tools for metabolomics and multivariate statistics were applied to Isatis tinctoria as a model plant. Nature provides compounds that are not only important for pharmaceutical industry, but also for agrochemistry and cosmetics. Extracts and pure natural compounds are widely used in cosmetics. For agrochemistry, natural products rather play a role for lead discovery, i.e. the discovery of a model compound that is further developed to a synthetic agrochemical. In the first part of the project, a library of 600 Panamanian plant extracts was screened for agrochemical and cosmetic properties. Selected extracts were submitted to HPLC-based activity profiling whereby five extracts showed activity in the agrochemistry screens, and two extracts in the cosmetic screens. For the agrochemical part of the project, extracts and microfractions were screened for fungicidal, insecticidal, and herbicidal properties. Compounds from Bocconia frutescens, Miconia affinis, Myrcia splendens, Combretum affinis laxum, and Erythroxylum macrophyllum were isolated, identified, and submitted to screening for the corresponding indication. Benzophenanthridine alkaloids and arjunolic acid from B. frutescens and M. affinis, respectively, showed moderate to good fungicidal activity. Myricetin-3-O-(6''-O-galloyl)-beta-galactopyranoside from M. splendens showed moderate insecticidal activity. From the two plants selected for herbicidal activity of their extracts, no herbicidal compounds could be identified. For the cosmetic part, phenolic compounds and flavonoids were isolated from methanolic extracts of leaves from Mosquitoxylum jamaicense and Combretum cacoucia. All isolated compounds showed either activity in the DPPH assay, in the UV-B protection assay, or in both. Gallic acid derivatives were the most active in the DPPH assay (IC50 < 0.01 mg/mL), while protocatechuic acid and isoquercitrin showed the best activity in the UV-B protection assay (< 10% UV-B-induced cell death). Metabolomics of plant extracts combined with multivariate statistics is increasingly used for the differentiation of plant samples and assessment of quality traits. Metabolomics provides a metabolic fingerprint, in contrast to the classical quantitative analysis of single compounds, or a single class of compounds. The second part of the AGROCOS project comprised a metabolomic study on Isatis tinctoria of five different accessions, which were grown under identical and controlled conditions on experimental field plots. Leaves were harvested at six time points during the vegetation period, whereby sampling was done, on the one hand, by collection of single leaves from intact plants, and by repeated mowing of leaf rosettes, on the other hand. The latter regimen was to simulate repeated harvesting of annual plants. Extracts were prepared by PLE with ethyl acetate and 70% aqueous methanol, submitted to 1H NMR spectroscopy, and analyzed by the PCA/CA/k-NN classification method. PCA/CA/k-NN analysis of accessions and time points of harvesting showed differences in both cases. Using pairwise 1D-STOCSY, compounds such as unsaturated fatty acids, porphyrins, carbohydrates, indole derivatives, isoprenoids, phenylpropanoids, and minor aromatic compounds could be identified as metabolites contributing to differences of accessions and harvesting dates. In addition, an effect of repeated harvesting could be observed from the PCA/CA/k-NN plot, and the 1D-STOCSY plot revealed a decrease of 1,5-anhydroglucitol, sucrose, unsaturated fatty acids, porphyrins, isoprenoids, and a flavonoid upon repeated harvesting.