Metabolomic analysis of Cyrtopodium glutiniferum extract by UHPLC-MS/MS and in vitro antiproliferative and genotoxicity assessment.

Extracts of orchids have been traditionally used as human phytotherapeutics. Cyrtopodium flavum, for example, is used to heal skin lesions in the Brazilian folk medicine, and it is focus of research due to the analgesic and anti-inflammatory properties. The pseudobulbs of Cyrtopodium glutiniferum, an orchid species found in Brazilian southeastern rainforest, are known to synthesize anti-inflammatory compounds, such as glucomannans and other potentially therapeutic compounds.We have reported the first metabolomic analysis focused on the phenols expression of the neotropical orchid Cyrtopodium glutiniferum Raddi, besides free radical scavenging, anti-inflammatory and antiproliferative activities, and the genotoxicity properties of the aqueous extract.

The metabolomics of C. glutiniferum aqueous extract was performed through UHPLC-MSⁿ acquisition. We have detected the scavenging potential of the extract using DPPH assay. The genotoxic potential was performed by Ames Test (0-5000 μg mL^-1) and micronucleous assay (0-5000 μg mL^-1) in RAW264.7 cells. The cytotoxic potential of the extract against RAW264.7 was tested by WST-1 assay (0-500 μg mL^-1). And after all, the RAW264.7 cells were treated with non-cytotoxic concentrations of C. glutiniferum (0-50 μg mL^-1) to evaluate the antiproliferative and anti-inflammatory potential, besides the mitochondrial activity.

From the 55 molecules identified, 45.5% belonged to the phenolic compounds database from Phenol Explorer, 29% to an in-house built Orchidaceae molecules database, and 25.5% to both. Among the identified phenolic compounds, 18 subclasses were discriminated, being phenanthrenes the most abundant. Doses-dependent of C. glutiniferum extract were able to induce DPPH free radicals scavenging and also to increase TA100 His⁺ revertants, in metabolic environment, showing mutagenicity just in the highest concentration, of 5000 μg/plate. On Eukaryotic cell models, the extract also has induced dose-response and time-response cytotoxicity against RAW264.7 macrophages, mainly after 48 h and 72 h, even though the extract has not been able to induce the increase of micronucleated cells and mitotic index alteration on Micronucleus assay. The activation and proliferation of macrophages cultures were downregulated after 24 h and 48 h by the non-cytotoxic concentrations of the extract in a dose-dependent manner.

The Cyrtopodium glutiniferum metabolomics, anti-inflammatory and anti-proliferative properties observed in this study have suggested a therapeutic efficacy of the orchid extract applied in folk medicine.