Estabelecimento e multiplicação in vitro de Eriope blanchetii (Benth.) Harley para quantificação da produção de podofilotoxina

Abstract

The lignans, are a secondary metabolites class with great biological importance, which are found in some species of the Lamiaceae family. Among the species variety within this family, the outstanding Eriope blanchetii, endemic of Brazil, occurs on the restingas from Bahia and Sergipe. The medicinal value of this species was already proven by a phytochemical and cytotoxic study, on which was detected podophyllotoxin presence, a secondary metabolite that is a precursor of two semisynthetic derivatives (etoposide and teniposide) with an anticancer effect. Considering the vulnerable status given to this species, and being aware that the in natura the biosynthesis of secondary metabolites presents a large variation due to the enviromental conditions, the use of vegetal tissues culture technology is here highlighted. Thus, this work aimed to establish E. blanchetii in vitro cultivation and analyze abiotics factors that might influence over podophyllotoxin expression after the in vitro cultivation. Different desinfestation, germination, conservation and seed dormancy breaking protocols were tested, as well as the salts concentrations influence during the in vitro establishment culture medium. Also, a new seeds desinfestation protocol was proposed for E. blanchetii, that was composed by a pre-germinative stage (at a seed level) and a pos-germinative stage (at a plant level). The most efficient protocol for overcoming seed dormancy procedure was the gibberellin solution at 200mg.L-1 soaking for 1h. The E. blanchetii seeds can be preserved at -80ºC for up to 90 days without compromising its germination potential. The biggest matter production of E. blanchetii can be obtained by the use of a MS (MURASHIGE; SKOOG, 1962) culture medium supplemented with active charcoal 2g.L-1. For in vitro multiplication of E. blanchetii, node regions explants were submitted to different in vitro cultivation conditions, aiming to stimulate a overproduction of podophyllotoxin. The best rooting data were obtained in the absence of regulators. For the aerial parts, fresh matter and dry matter development, the use of 8,88μM of BAP is recommended. The ideal temperature for E. blanchetii in vitro multiplication is 25ºC. Finally, the in vitro cultivation conditions, as well as the abiotic factors used as a stress source, made the podophyllotoxin expression on E. blanchetii impossible.