Ahmed Rezk

• The evolution of bacterial resistance to current antibiotics is one of the biggest threats to human health. There is an increasing interest to identify novel antimicrobial compounds from various natural sources. Plants-derived compounds are used as ingredients in traditional treatment of numerous human disorders including infectious diseases caused by pathogenic microorganisms. In planta, secondary metabolites like the polyphenols are known to act as potent antimicrobial agents against several plant pathogens. Plants of the genus Rhododendron are typically used in a range of ethno-medical applications. There are more than 1,000 Rhododendron species growing/spreading/distributed particularly in the Northern hemisphere. The City of Bremen harbors the Rhododendron-Park in which approximately 600 different species of Rhododendron and hybrids are grown. This enables research with about two thirds of all known Rhododendron species. The aim of this study is to identify novel compounds from the secondary metabolite pool synthesized by Rhododendron that can be used as antimicrobial treatments of human diseases/maladies in form of ectopic application or as orally administered drugs. Therefore, leaf extracts of a total of 120 different Rhododendron species were tested using the agar diffusion assay towards twenty-six bacterial species representing different taxonomic clades of non-pathogenic strains Gram-positive and Gram-negative bacteria. The possible cytotoxic effects of the most promising, antimicrobial bioactive extracts from Rhododendron species were assessed in a concentration-dependent manner using epidermal keratinocytes of the skin and epithelial cells of the intestinal mucosa, respectively, as target cell systems. The leaf extracts of 17 Rhododendron species exhibited significant growth-inhibiting activities against Gram-positive bacteria. In contrast, only very few of the leaf extracts affected the growth of Gram-negative bacteria. All leaf extracts with antimicrobial bioactivity were extracted from representatives of the subgenus Rhododendron, with 15 from the sub-section Rhododendron and two belonging to the section Pogonanthum. Six Rhododendron species out of the 17 species showed the lowest minimum inhibitory concentration with 50 µg leaf extract powder per mL solvent. Equally low and moderate concentrations (50 µg/ml) of leaf extracts from three of these plant species were non-toxic towards both mammalian cell lines, i.e. HaCaT keratinocytes and IEC6 enterocytes. However, higher doses such as 500 µg/mL of Rhododendron leaf extracts were potent in negatively affecting both, keratinocytes and, particularly, the intestine epithelial cells. We conclude that bioactive compounds with high antimicrobial activities can be extracted from the leaves of Rhododendron species mainly belonging to the subgenus Rhododendron, and that they acted mostly against Gram-positive organisms. The leaf extracts from R. minus, R. ferrugineum, and R. racemosum applied at a concentration of 50 µg/ml proved safe to be used in 24-h-incubations of monolayer cultures of both, HaCaT keratinocytes and IEC6 intestine epithelial cells. In contrast, high doses of most leaf extracts induced apoptosis evidenced by a significant increase in the levels of active caspase-3 in IEC6 intestine epithelial cells. Finally, keratinocytes proved more resistant than intestine epithelial cells against cytotoxicity exerted by leaf extracts of a large variety of Rhododendron species.