RESUMO
An efficient nicotine-degrading fungus was isolated from tobacco leaves and identified as Aspergillus oryzae 112822 based on morphological characteristics and sequence analysis of 18S rDNA, 5.8S rDNA and the internal transcribed spacer (5.8S-ITS region). When the strain was cultured in a medium with tobacco leaf extract for 40 h, the maximum amount of cell growth was 3.6 g l(-1) and nicotine degradation was 2.19 g l(-1). The intermediates of nicotine degradation by resting cells were isolated by preparative TLC or semi-preparative HPLC, and identified by TLC, MS, NMR, Fourier-transform (FT)-IR and GC-MS analysis. The pathway for nicotine degradation in A. oryzae 112822 was proposed to be from nicotine to 2,3-dihydroxypyridine through the intermediates nornicotine, myosmine, N-methylnicotinamide and 2-hydroxy-N-methylnicotinamide. The ring of 2,3-dihydroxypyridine was opened between the 2- and 3-hydroxy positions to yield succinic acid. N-methylnicotinamide and 2,3-dihydroxypyridine were satisfactorily verified as metabolites of nicotine degradation. This is the first elucidation of a pathway for nicotine degradation in fungi.
Assuntos
Aspergillus oryzae/metabolismo , Nicotiana/metabolismo , Nicotiana/microbiologia , Nicotina/metabolismo , Alcaloides/metabolismo , Aspergillus oryzae/isolamento & purificação , Biodegradação Ambiental , DNA Ribossômico , DNA Espaçador Ribossômico , Cromatografia Gasosa-Espectrometria de Massas , Redes e Vias Metabólicas , Modelos Biológicos , Niacinamida/análogos & derivados , Niacinamida/metabolismo , Nicotina/análogos & derivados , Folhas de Planta/microbiologia , Piridinas/metabolismo , RNA Ribossômico 18SRESUMO
BACKGROUND & OBJECTIVE: Solanum indicum L., an anti-inflammatory and wound-healing herb in traditional Chinese medicine, is enriched of unique dioscins, that is, indiosides. Our previous studies revealed that several synthetic indiosides from Solanum indicum L. have potent anticancer effects. This study was to investigate the anticancer mechanism of synthetic indiosides I from Solanum indicum L. METHODS: Human hepatocarcinoma Bel-7402 cells were treated with different concentrations of indiosides. The inhibitory rate of cell proliferation and 50% inhibitory concentration (IC50) of indiosides were detected by the acid phosphatase assay (APA). Cell morphology was observed under optical microscope with crystal violet staining. The expression of apoptosis-related proteins was detected by Western blot. RESULTS: Indiosides significantly inhibited proliferation of Bel-7402 cells: when treated with indioside I for 72 h, the IC50 value was 4.2 microg/ml, cell density was decreased, and bubbles were observed in cytoplasm. Western blot showed that the cytoplasmic level of cytochrome c was increased significantly, Caspase-3 was activated, and poly (ADR-ribose) polymerase (PARP) was cleavaged after indioside I treatment. CONCLUSION: Indiosides have dose-dependent inhibitory effect on proliferation of Bel-7402 cells, and can induce cell apoptosis through mitochondria-dependent pathway.
