Biology session

The black locust tree (Robinia pseudoacacia, family Fabaceae) is native to South America, and it was first introduced in Hungary in the 18th century as ornamental. Later it was used to replanting the saline soil and to stop the drift. Nowadays one fourth of Hungarian forests are black locust forests. The tree is utilized as honey plant as well as in the building and furniture industries. It is a species with low ecological needs, its seeds keep the germinability for long time, when they are in the soil. The plant can also reproduce vegetatively from shoots. Thanks to these, it is invasive, it can quickly occupy large areas.

Recently a vast of study focuses on the search for natural occurring bioactive materials and substances, so the test of different plants is in focus again. The black locust, since it can provide large biomass, could be a candidate as an easily exploitable herb. So far, in the black locust, mainly its flavonoids (e.g. acacetin, apigenin, diosmetin, luteolin) and toxic substances (e.g. robin) were examined.

The aim of our research was the search for antioxidant and antibacterial compounds from black locust and their characterization. We investigated the parts of the tree: bark, leaves, flowers and fruit. The collected samples were dried in dark at room temperature and after they were grinded and extracted with different solvents. The components of the extracts were tested by thin-layer chromatography (TLC) coupled with bioactivity assays. Therefore, TLC methods were developed for the appropriate separation of the bioactive plant compounds.

The antibacterial activity of the components separated by TLC were detected in situ in the adsorbent bed by direct bioautography using various bacterial strains. During the experiment the layer was immersed into bacterial cell suspension and after sufficient incubation time the antibacterial zones were visualized e.g. by vital dye reagent. This method is fast, reliable and suitable for high-throughput test as well as for effect-directed isolation. The antioxidants were visualized by dipping the developed layer into methanol solution of the relatively stable radical DPPH (2,2-diphenyl-1-picrylhydrazyl). The detected bioactive compounds were further characterized by TLC reagents, successfully isolated from the layer and analysed by HPLC-DAD-ESI-MS(/MS). Based on the preliminary results one of the antibacterial components is a fatty acid, probably linolenic acid.

The gallic acid equivalent antioxidant capacity of methanol extract of bark, leaf, flower and fruit samples collected from different place at the same time and from the same place at different time (from spring to autumn) were determined and compared by the use of 96-well microplates and DPPH.