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

Abstract

Ethnopharmacological relevanceExtracts of orchids have been traditionally used as human phytotherapeutics. Cyrtopodium flavum, for example, due to the analgesic and anti-inflammatory properties, beside the capacity of heal skin lesions has been focus of research. Also Cyrtopodium glutiniferum, an orchid found in the Brazilian southeastern rainforest, is known to synthesize anti-inflammatory glucomannans in the pseudobulbs, as other potentially therapeutic compounds. Aim of the studyWe 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. Material and MethodsThe 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. ResultsFrom the 55 molecules identified, 45.5% belonged to the phenolic compounds database from Phenol Explorer, 29% to an in-house 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 extracts 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 5 mg/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. ConclusionsThe Cyrtopodium glutiniferum metabolomics, anti-inflammatory and anti-proliferative properties observed in this study suggest a therapeutic efficacy of the orchid extract applied in folk medicine.