Phytochemistry and evaluation of the photoinduced biological effect of extracts and isolated substances from Annonaceae (plant in nature and obtained by cell culture)

Author(s)

Caroline Caramano de Lourenço

• Document type: Doctoral Thesis
• Press: Campinas, SP.
• Institution: Universidade Estadual de Campinas (UNICAMP). Instituto de Biologia
• Defense date: 2016-02-05

Examining board members

Marcos José Salvador; Elfreiede Marianne Bacchi; Stella Regina Zamuner; Felipe Rabello Lourenço; Silmara Marques Allegretti

• Advisor: Marcos José Salvador

Abstract

he aim of the study chemical, biological and biotechnological of two Annonaceae¿s species (Annona Cacans, Annona mucosa, Annona squamosa or Guatteria hispida) went for bioactive substances and photoactivated these plant matrices (plants in nature and obtained by cell culture). The study of methanolic crude extract was carried out by monitoring the absorption spectrum in the region of interest in photodynamic therapy (600 to 800 nm), the fluorescence spectrum analysis (mode emission and excitation), the evaluation of the photochemical efficiency and the results of bioassays including assessment of the biological effect photoinduced by laser radiation at 660 nm in an experimental model in vitro antimicrobial photodynamic therapy (PDT-antimicrobial). Therefore, we proceeded to fit methodology for biological testing PDT-antimicrobial and development of analytical procedures including the study bioactivity Phytochemistry, obtaining initially a chemical profile (CCDC, GC, HPLC, NMR and ESI-MS ). The results allowed the identification of 11 alkaloids already described in literature for Annonaceae species directly in crude extracts (dereplication). After the step of screening and by comparing the results obtained it was possible to define G hispida and. A. squamosa as the most promising species for which continued the study. The other species showed low absorption and fluorescence in the region of interest in PDT and low photochemical efficiency. The extracts of the two select species with substances that could not be identified in dereplication stage underwent biomonitored fractionation and isolated substances and purified held its structural determination using spectroscopic methods of analysis which enabled the identification of 5 alkaloids: alkaloid Isomoschatoline and 9-methoxy-isomoschatoline were isolated only from samples G. hispida and Palmatine alkaloid isolated only from A. squamosa extract while Anonaina and liriodenine alkaloids have been isolated from two species of the extracts. In biotechnology study proceeded to the establishment and development of cell cultures of the two most promising species in bioassays, aimed beyond the phytochemical study, the handling of physical and chemical factors for the production of secondary metabolites and the comparative study of bioactive metabolites in cell cultures and those obtained from the fresh plant parts. Friable callus and oxidation free were obtained for the two species. The comparative analysis of ESI-MS / MS spectra of the bioactive extracts showed similarities between the composition of the plant extracts in nature to that obtained by cultivating cells and allowed the quantitation UHPLC-MS some alkaloids already described for species of Annonaceae in samples studied, including Asimilobine alkaloids, Liriodenine, Lysicamine and Palmatine that have potential for use in antimicrobial PDT observing similarities in results in chemical and biological terms. In collaboration with the "Laboratoire Jean Perrin, Campus Jussieu, Université Pierre et Marie Curie in Paris, France," were carried out some studies of isoquinoline alkaloids, such as Isomoschatoline (GH1) and Palmatine (AS3) alone and associated with the cyclodextrin. Studies suggest that these alkaloids are best directed when complexed within tumor cells and predominantly concentrated in the lysosomes (AU)

• FAPESP's process: 11/18315-2 - CHEMICHAL, BIOLOGICAL AND BIOTECHNOLOGICAL STUDIES BY ANNONACEAE SPECIES (Annona cacans, Annona squamosa, Annona mucosa or Guatteria hispida)
• Grantee: Caroline Caramano de Lourenço
• Support Opportunities: Scholarships in Brazil - Doctorate