Morfogênese in vitro, análise fitoquímica e caracterização anatômica de nim (Azadirachta indica A. Juss)

• Autor(a) principal: Rodrigues, Marcelo
• Data de Publicação: 2009
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: LOCUS Repositório Institucional da UFV
• Texto Completo: http://locus.ufv.br/handle/123456789/2516
• topic: Azadirachta indica; Nim; HPLC; Azadirachtina; Anatomia; Azadirachta indica; Neem; HPLC; Azadirachtina; Anatomy; CNPQ::CIENCIAS BIOLOGICAS::BOTANICA

The Azadirachta indica A. Juss species, popularly known as neem, belongs to the Meliaceae family, which has great potential for azadirachtin (AZA) production; however, neem has recalcitrant seeds, cross-pollination and variable content of AZA. Four experiments were conducted in order to understand the in vitro growth limiting factors of these plants and changes in levels of AZA: i) In vitro propagation of neem by culture of collected nodal segments; ii) Evaluation of leaf anatomy of plants grown in three different environments (in vivo, in vitro and acclimatized), using the micromorphometric technique; iii) Evaluation of levels of AZA, which were determined by High Performance Liquid Chromatography (HPLC) in cotyledonary callus, grown in WPM liquid medium with absence and presence of glucose (carbon source), hydrolyzed casein (nitrogen source) and methyl jasmonate (elicitor agent); and, iv) The propagation of somatic embryos from cotyledonary callus. It was found that MS medium has presented the best results for shooting rate, leaflet number and average size of the explants as compared to others culture media (WPM and JADS). It was not induced in vitro rooting of shoots; however, the root system has developed ex vitro during acclimatization, with seedling survival rate of 80%. It was also assayed the general content of lipids, essential oils, resins and oils, tannins and mucilage. It was found that there are significant differences in height between the evaluated leaf tissues, adaxial and abaxial epidermis, as well as in the mesophyll from three environmental conditions. The histochemical assay showed that there is general production of lipids, essential oils and tannins in leaf idioblasts of neem, meanwhile the mucilage test showed negative results for the three evaluated locations. The highest concentrations of AZA (mean 0.2470 mg g-1) were produced in the first and second cultivation week in the presence of these three substances, demonstrating the synergistic effect when the cell mass was cultured in medium supplemented with these. Calli showed high propagation capacity by both regenerative routes organogenesis and embryogenesis starting from the fourth subculture in WPM medium presenting the highest shooting and somatic embryos germination rates. Calli went from a smooth to granular appearance during the subcultures, while color went from dark brown to light brown and white. From the seventh subculture, there was dramatic reduction of in vitro regeneration rate for both types of neem cotyledonary callus. Therefore, this work has contributed for a better understanding of biotechnology techniques aiming the in vitro propagation, cell cultivation for AZA production and anatomical aspects of A. indica.