Olivetonide

Lichens produce a diversity of secondary metabolites, among them depsides comprised of two or more hydroxybenzoic acid units linked by ester, ether, or CC-bonds. During classic solid support-based purification processes, depsides are often hydrolyzed and in many cases time, consuming procedures result only in the isolation of decomposition products. In an attempt to avoid extensive purification steps while maintaining metabolite structure integrity, we propose an alternative method to identify the major depsides of a lichen crude extract (Pseudevernia furfuracea var. ceratea (Ach.) D. Hawksw., Parmeliaceae) directly within mixtures. Exploiting the acidic character of depsides and differences in polarity, the extract was fractionated by centrifugal partition chromatography in the pH-zone refining mode resulting in twelve simplified mixtures of depsides. After (13)C nuclear magnetic resonance analysis of the produced fractions, the major molecular structures were directly identified within the fraction series by using a recently developed pattern recognition method, which combines spectral data alignment and hierarchical clustering analysis. The obtained clusters of (13)C chemical shifts were assigned to their corresponding molecular structures with the help of an in-house (13)C NMR chemical shift database, resulting in six unambiguously identified compounds, namely methyl β-orcinolcarboxylate (1), atranorin (2), 5-chloroatranorin (3), olivetol carboxylic acid (4), olivetoric acid (5), and olivetonide (6).

This study reports for the first time in the chemical composition of acetone, ether, ethyl acetate and dichloromethane extracts of Hypogymnia tubulosa determined by HPLC-UV, GC-FID and GC-MS as well as effect of H. tubulosa acetone extract on micronucleus distribution on human lymphocytes and on cholinesterase activity. Additionally, antioxidant (estimated via DPPH, ABTS, TRP, CUPRAC and TPC assays) and antibacterial activity against two Gram-positive and three Gram-negative bacteria were also determined. The HPLC-UV analysis revealed the presence of depsidones, 3-hydroxyphysodic, 4-O-methyl physodic acid, physodic and physodalic acid together with two depsides, atranorin and chloroatranorin. GC-FID and GC-MS analyses enabled the identification of atranol, chloroatranol, atraric acid, olivetol, olivetonide and 3-hydroxyolivetonide as the main components. The results of present study show that H. tubulosa acetone extract is a promising candidate for in vivo experiments considering antioxidant activity.