Estabelecimento de culturas de raízes in vitro e avaliação da produção de metabólitos secundários de Cleome spinosa Jacq. (Cleomaceae)

• Autor(a) principal: Moraes, Brenda Amaral de
• Data de Publicação: 2015
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: Biblioteca Digital de Teses e Dissertações da UERJ
• Texto Completo: http://www.bdtd.uerj.br/handle/1/7959
• topic: Auxin; Phenolic compounds; Chromatography; Crude extract; Terpenes; Auxinas; Compostos fenólicos; Cromatografia; Extrato bruto; Terpenos.; Fisiologia vegetal; Tecidos vegetais - Cultura e meios de cultura; Fitoquímicos; Plantas medicinais; CNPQ::CIENCIAS BIOLOGICAS::BOTANICA

Cleome spinosa is a herbaceous species used in folk medicine, mainly in the Northeast of Brazil. Cultures of adventitious roots were established with root segments (0.5 to 1.0 cm) obtained from two sources of explants: in vitro propagated plants and plants obtained from in vitro germination process. The explants were inoculated in MS liquid medium supplemented with or without the auxin NAA, IAA and IBA in different concentrations (0, 0.5, 1.0, 1.5, 3.0, 5.0 mg.L-1). Cultures were maintained in a growth room under agitation (100 rpm) and photoperiod of 16h light or in the dark, with subcultures every 45 days. Explants from in vitro propagated plants were also cultured in medium containing BAP in combination with auxins, in the presence of sorbitol (alone or in combination with sucrose), in MS medium with a reduction in the total concentration of mineral salts (MS1/2 and MS1/4) and on solidified medium. The results showed that hormonal supplementation was essential for roots multiplication, once cultures maintained in MS0 medium only developed shoots. The medium supplementation with NAA was not efficient for root production and resulted in callus induction. The presence of IAA and IBA was efficient to root multiplication. A significant root proliferation capacity was observed in cultures initiated from explants originating from plants obtained by in vitro germination process. On the other hand, cultures initiated with explants originating from in vitro propagated plants, regardless the different manipulations performed in the culture medium, showed low root multiplication capacity. The highest biomass production was achieved by cultures initiated with explants from in vitro germination plants cultured in medium supplemented with 3.0 mg.L-1 IBA and kept in the dark. Cultures with the highest biomass accumulation were subcultured at each 45 days or 60 days interval to fresh medium with the same composition during three - four subcultures. Cultures maintained at 45 days interval reached the highest biomass accumulation during the second subculture, while cultures maintained at 60 days interval, despite the maintenance of root multiplication along the subcultures, reached the highest production during the first 60 days in culture. Samples from in vivo and in vitro material were extracted with increasingly polar solvents (hexane, dichloromethane, ethyl acetate, methanol) and the extracts were submitted to chromatographic analysis. The analysis by thin layer indicated the presence of terpenes in hexane and dichloromethane extracts obtained with field plants and in vitro materials, and the presence of phenolic componds in ethyl acetate extracts obtained with field plants. Analysis by liquid chromatography coupled to mass spectrometry showed the in vitro production of metabolites not found in field plants, especially flavonoids. The gas chromatography coupled to mass spectrometry indicated the presence of steroids in the extracts obtained with hexane from field plants and root cultures. The results demonstrated the feasibility in the production of in vitro root cultures of C. spinosa and the potential of these biotechnological materials to phytochemical studies, mainly considering the presence under in vitro cultures of metabolites not produced by field plants