Eliciação de metabólitos secundários na cultura in vitro de Duroia macrophylla e Duroia saccifera

Abstract

Species of the Duroia genus are producers of terpenes, flavonoids and alkaloids, presenting several biological activities, with emphasis on the antitumor activity conferred by the novel alkaloid isolated from D. macrophylla; in addition to other alkaloids with activity against Mycobacterium tuberculosis that resulted in another patent already granted to previous research. In order to increase and constant the production of active substances, several species have been established in vitro and subjected to elicitation techniques, including species of the Duroia genus. Based on the above, the present research aimed to evaluate the effect of precursors and elicitors in D. macrophylla and D. saccifera cultures on the production of secondary metabolites, aiming at the production of alkaloids and/or intermediates. For biological elicitation, mycelia and methanolic extracts (MeOH) of Xylaria sp. and Colletotrichum dianesei were used; Physical elicitation was performed with Light Emitting Diodes (LEDs) in yellow, red and blue colors; tryptophan and iridoid monotropein methyl ester were used as precursors. From the results obtained, it was verified that the D. macrophylla calli remained multiform, and no cell suspension was obtained. The difference in the composition of the culture medium interfered with the production of D. macrophylla metabolites, and the WPM medium added with 2 mg.L-1 of naphthalene acetic acid (NAA) and 2 mg.L-1 of benzylaminopurine (BAP) was established as the most effective combination for the production of the iridoid monotropein methyl ester. From maintenance cultures, it was verified that in vitro Duroia calli are effective producers of phenolics, and it was possible to isolate and characterize scopoletin in D. saccifera and chlorogenic acid in D. macrophylla using chromatographic techniques and H nuclear magnetic resonance and mass spectrometry analyses. The precursors tryptophan and tryptamine did not modify aspects of growth and secondary metabolism of D. saccifera and D. macrophylla calli. The application of endophytes as elicitors did not alter the chemical production of D. saccifera cell suspensions, but intensified the production of previously produced metabolites, corroborating the literature. The elicitation of D. saccifera calli with red LED interfered with aspects of growth, but chemical production was altered by the growth phase and not by the incidence of light; the data were grouped according to their similarities, through chemometric analysis. D. macrophylla plants did not undergo changes in the production of new metabolites, but the red LED provided a constant production of the iridoid monotropein methyl ester, an intermediate in the pathway of the desired indole alkaloids.