Discussion
Site-specific chemical lineages of rooibos ecotypes were confirmed via multivariate data analyses. Important features identified via PLS-DA disclosed higher relative abundances of certain tentative metabolites (e.g., rutin, aspalathin and apiin) present in the Dobbelaarskop, Blomfontein, Welbedacht and Eselbank sites, in comparison to other locations. Several unknown novel metabolites (e.g., m/z 155.0369, 231.0513, 443.1197, 695.2883) are responsible for metabolomic separation of the populations, four of which showed higher amounts of key metabolites and were thus selected for bioactivity analysis. The Welbedacht and Eselbank site 2 populations consistently displayed higher antioxidant activities, with 2,2-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) radical scavenging activities of 679.894 ± 3.427 µmol Trolox/g dry matter and 635.066 ± 5.140 µmol Trolox/g dry matter, respectively, in correlation with a high number of phenolic and flavonoid compounds. The contribution of the individual metabolites to the pharmacological effectiveness of rooibos remains unknown and as such, further structural elucidation and phytopharmacological testing is thus urgently needed.
Methods
Despite the commercial popularity of the cultivated variety, the uncultivated ecotypes are largely understudied. To address gaps in knowledge about the biochemical constituency, ultra-performance liquid chromatography-mass spectrometry analysis of fifteen populations was performed, enabling high-throughput metabolomic fingerprinting of 50% (v/v) methanolic extracts. Antioxidant screening of selected populations was performed via three assays and antimicrobial activity on two microbial species was assessed. The metabolomic