In order to explore the potential of endophytic actinomycetes to produce secondary metabolites with applications in the cosmeceutical industry, 138 potential candidate endophytic actinomycetes originated from Spanish biodiverse arid areas were selected to be studied. In order to broadly express the production of different secondary metabolites, OSMAC approach was applied and four nutritional conditions have been used for the cultivation of the selected strains in liquid media. In total 552 extracts have been generated and cell-free and cell-based bioassays have been incorporated for the evaluation of their skin-whitening activity. In parallel cytotoxicity evaluation was performed to the selected bioactive extracts in order to exclude those that exert cytotoxic effects.
The novel actinomycete strain CA287887 was selected as the most promising and was cultivated in large scale using the optimum fermentation medium (MO16). The scaled-up fermentation broth (3lt) was extracted with acetone (3lt). The active extract was loaded on a column packed with SP207SS reversed-phase brominated resin, while the aqueous phase was concentrated until dryness (aqueous extract). The loaded column was eluted using a linear gradient from 10 to 100% acetone in water. Bioassays revealed the active fractions, while the aqueous extract demonstrated the most significant activity. Sequential preparative & semi-preparative HPLC fractionations and liquid-liquid extraction with EtOAc were performed in order to purify the bioactive metabolites from the active fractions and the aqueous extract respectively.
The full set of spectroscopic data (MS and NMR) were recorded for all isolated compounds in order to unambiguously elucidate their structure. Special attention was given to the isolated alkaloid derivatives Cyclo-(histidine-proline) (C11H14N4O2) and Cyclo(Pro-Tyr) (C14H16N2O3) due to their remarkable whitening activity and to one novel alkaloid with molecular formula C30H49N7O10. Therefore, this can serve as a proof of concept that microbial ingredients can have successful applications in cosmeceutical industry.