Globalization facilitated the spread of invasive alien species (IAS), undermining the stability of the world’s ecosystems. We investigated the metabolomic profiles of three IAS species: Chromolaena odorata (Asteraceae) Datura stramonium (Solanaceae), and Xanthium strumarium (Asteraceae), comparing metabolites of individual plants in their native habitats (USA), to their invasive counterparts growing in and around Kruger National Park (South Africa, ZA). Metabolomic samples were collected using RApid Metabolome Extraction and Storage (RAMES) technology, which immobilizes phytochemicals on glass fiber disks, reducing compound degradation, allowing long-term, storage and simplifying biochemical analysis. Metabolomic differences were analyzed using ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) of samples eluted from RAMES disks. Partial Least Squares-Discriminant Analysis (PLS-DA) of metabolomes of individual plants allowed statistical separation of species, native and invasive populations of each species, and some populations on the same continent. Invasive populations of all species were more phytochemically diverse than their native counterparts, and their metabolomic profiles were statistically distinguishable from their native relatives. These data may elucidate the mechanisms of successful invasion and rapid adaptive evolution of IAS. Moreover, RAMES technology combined with PLS-DA statistical analysis may allow taxonomic identification of species and, possibly, populations within each species.

Study of plant metabolome is a growing field of science that catalogs vast biochemical and functional diversity of phytochemicals. However, collecting and storing samples of plant metabolome, sharing these samples across the scientific community and making them compatible with bioactivity assays presents significant challenges to the advancement of metabolome research. We have developed a RApid Metabolome Extraction and Storage (RAMES) technology that allows efficient, highly compact, field-deployable collection and storage of libraries of plant metabolome. RAMES technology combines rapid extraction with immobilization of extracts on glass microfiber filter discs. Two grams of plant tissue extracted in ethanol, using a specially adapted Dremel^® rotary tool, produces 25–35 replicas of 10 mm glass fiber discs impregnated with phytochemicals. These discs can be either eluted with solvents (such as 70% ethanol) to study the metabolomic profiles or used directly in a variety of functional assays. We have developed simple, non-sterile, anti-fungal, anti-bacterial, and anti-oxidant assays formatted for 24-multiwell plates directly compatible with RAMES discs placed inside the wells. Using these methods we confirmed activity in 30 out of 32 randomly selected anti-microbial medicinal plants and spices. Seven species scored the highest activity (total kill) in the anti-bacterial (bacteria from human saliva) and two anti-fungal screens (Fusarium spp. and Saccharomyces cerevisiae), providing functional validation of RAMES technology. RAMES libraries showed limited degradation of compounds after 12 months of storage at -20°C, while others remained stable. Fifty-eight percent of structures characterized in the extracts loaded onto RAMES discs could be eluted from the discs without significant losses. Miniaturized RAMES technology, as described and validated in this manuscript offers a labor, cost, and time-effective alternative to conventional collection of phytochemicals. RAMES technology enables creation of comprehensive metabolomic libraries from various ecosystems and geographical regions in a format compatible with further biochemical and functional studies.