| Congresso Brasileiro de Microbiologia 2023 | Resumo: 662-2 | ||||
Resumo:Amaryllidaceae J. St.-Hil. is a family of plants native to Central Asia and North America, which can be found in various biomes around the world, including the Cerrado. It is a family rich in alkaloids of great pharmacological interest, such as galantamine and licorine. However, plant-derived secondary metabolites may not be the only natural source of promising substances: endophytic fungi, microorganisms that symbiotically inhabit plant tissues, are potential sources of bioactive compounds, although studies on them are still limited. The aim of this study was to prospect endophytic fungi from leaves and bulbs of Crinum americanum (Amaryllidaceae) and identify, among the isolated fungi, the presence of species that produce alkaloids related to the host plant. The leaves and bulbs used were collected in the Federal District in July 2022. After collecting the plant material, one leaf and one bulb were randomly selected and washed with running water. Then, the plant material was completely immersed in 70% ethanol for 60 seconds and in three containers containing 2% sodium hypochlorite for 60, 90, and 180 seconds, respectively. The material was immersed again in 70% ethanol for 60 seconds and then in three containers with distilled water for 30 seconds. The leaf and bulb were cut and inoculated on Petri dishes containing Sabouraud dextrose agar (SDA) medium enriched with 2% malt extract. A negative control was performed with a sample containing 100 μL of the last wash water, inoculated in enriched SDA medium, and another control consisted of gently pressing the surface of the freshly washed material onto another plate containing the same medium. All plates were incubated at 28 ºC until fungal growth appeared. In the end, three fungi were isolated from the bulb (BCEL1, BCEL2, and BCEL3) and three fungi were isolated from the leaves (FCEL9, FCEL10, and FCEL11) of C. americanum. The isolated fungi were cultivated in SPY medium (2% sucrose, 0.5% peptone, and 0.5% yeast extract) at 28 ºC, 120 rpm for 3 days. After this period, the culture was filtered and the biomass transferred to SPY medium (1% sucrose, 0.2% peptone, and 0.2% yeast extract), and cultivated at 28 ºC, 120 rpm for 15 days. Finally, the medium was filtered, and the mycelium was macerated in methanol for 9 days, with solvent renewal every 3 days, resulting in the methanolic mycelium extract (MME). The liquid phases were partitioned with hexane, ethyl acetate, and n-butanol, generating the hexane fraction of the liquid phase (HFF), ethyl acetate fraction of the liquid phase (EAFF), and n-butanol fraction of the liquid phase (nBFF). The obtained fractions and extracts were analyzed by thin-layer chromatography, eluted in chloroform: acetone: diethylamine (5:4:1), and visualized under UV light (Figure 1). The n-butanol fractions BCEL2, FCEL9, FCEL10, and FCEL11, as well as the methanolic extracts FCEL9 and FCEL10, showed bands similar to the lycorine standard, indicating the presence of alkaloids related to the host plant in the fungal fractions and extracts. However, the method used is not sufficient to confirm the production of such alkaloids by the isolated endophytic fungi. Another methodology for separation and analysis of substances, such as gas chromatography coupled with mass spectrometry (GC-MS), is required to confirm the presence of alkaloids of interest in these fungal fractions and extracts. Palavras-chave: Endophytic fungi, Crinum americanum, alkaloid, lycorine Agência de fomento:Fundação de Apoio à Pesquisa do Distrito Federal – FAPDF; Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – CAPES | |||||