Precursor-directed Biosynthesis in Tabernaemontana catharinensis as a New Avenue for Alzheimer's Disease-modifying Agents.

Plants produce a high diversity of metabolites that can act as regulators of cholinergic dysfunction. Among plants, the potential of species of the genus Tabernaemontana to treat neurological disorders has been linked to iboga-type alkaloids that are biosynthesized by those species. In this context, precursor-directed biosynthesis approaches were carried out using T. catharinensis plantlets to achieve new-to-nature molecules as promising agents against Alzheimer's disease. Aerial parts of T. catharinensis, cultured in vitro, produced 7 unnatural alkaloids (5-fluoro-ibogamine, 5-fluoro-voachalotine, 5-fluoro-12-methoxy-Nb-methyl-voachalotine, 5-fluoro-isovoacangine, 5-fluoro-catharanthine, 5-fluoro-19-(S)-hydroxy-ibogamine, and 5-fluoro-coronaridine), while root extracts showed the presence of the same unnatural iboga-type alkaloids and 2 additional ones: 5-fluoro-voafinine and 5-fluoro-affinisine. Moreover, molecular docking approaches were carried out to evaluate the potential inhibition activity of T. catharinensis' natural and unnatural alkaloids against AChE and BChE enzymes. Fluorinated iboga alkaloids (5-fluoro-catharanthine, 5-fluoro-voachalotine, 5-fluoro-affinisine, 5-fluoro-isovoacangine, 5-fluoro-corinaridine) were more active than natural ones and controls against AchE, while 5-fluoro-19-(S)-hydroxy-ibogamine, 5-fluoro-catharanthine, 5-fluoro-isovoacangine, and 5-fluoro-corinaridine showed better activity than natural ones and controls against BChE. Our findings showed that precursor-directed biosynthesis strategies generated "new-to-nature" alkaloids that are promising Alzheimer's disease drug candidates. Furthermore, the isotopic experiments also allowed us to elucidate the initial steps of the biosynthetic pathway for iboga-type alkaloids, which are derived from the MEP and shikimate pathways.

Unnatural spirocyclic oxindole alkaloids biosynthesis in Uncaria guianensis.

Spiro-oxindole scaffolds have been studied due to their promising therapeutic potential. In the Amazon rainforest there are two important Uncaria species known as "cat's claw", which biosynthesize spirocyclic oxindole alkaloids; Uncaria tomentosa (Willd. ex Schult.) DC. and Uncaria guianensis (Aublet) Gmell. We carried out a precursor-directed biosynthesis approach with U. guianensis and successfully obtained oxindole alkaloid analogues with molecular mass corresponding to the addition of a methyl or fluorine group on the oxindole ring using tryptamine analogue precursors. Two of these novel oxindole alkaloid analogues (3b-7-methyl-isomitraphylline and 3c-6-fluoro-isomitraphylline) were isolated and characterized by NMR spectroscopy and ESI-QTOF-MS/MS. Having established a substrate feeding protocol for these plantlets, the biosynthetic route for mitraphylline (1), rhynchophylline (2), isomitraphylline (3) and isorhynchophylline (4) was also investigated using 13C-precursors (1-13C-D-glucose, 2-13C-tryptophan, 1-13C-DL-glyceraldehyde, and methyl-13C-D-methionine).

Genetic diversity in natural populations of Jacaranda decurrens Cham. determined using RAPD and AFLP markers

Jacaranda decurrens (Bignoniaceae) is an endemic species of the Cerrado with validated antitumoral activity. The genetic diversity of six populations of J. decurrens located in the State of São Paulo was determined in this study by using molecular markers for randomly amplified polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP). Following optimization of the amplification reaction, 10 selected primers generated 78 reproducible RAPD fragments that were mostly (69.2 percent) polymorphic. Two hundred and five reproducible AFLP fragments were generated by using four selected primer combinations; 46.3 percent of these fragments were polymorphic, indicating a considerable level of genetic diversity. Analysis of molecular variance (AMOVA) using these two groups of markers indicated that variability was strongly structured amongst populations. The unweighted pair group method with arithmatic mean (UPGMA) and Pearson's correlation coefficient (RAPD -0.16, p = 0.2082; AFLP 0.37, p = 0.1006) between genetic matrices and geographic distances suggested that the population structure followed an island model in which a single population of infinite size gave rise to the current populations of J. decurrens, independently of their spatial position. The results of this study indicate that RAPD and AFLP markers were similarly efficient in measuring the genetic variability amongst natural populations of J. decurrens. These data may be useful for developing strategies for the preservation of this medicinal species in the Cerrado.

Variabilidade sazonal de constituintes químicos (triterpenos, flavonóides e polifenóis) das folhas de Maytenus aquifolium Mart. (Celastraceae) / Seasonal variability of chemical compounds (triterpenes, flavonoids and polyphenols) from Maytenus aquifolium Mart (Celastraceae) leaves

Estudou-se a variabilidade química sazonal em Maytenus aquifolium Mart. (Celastraceae), conhecida como "espinheira santa", em exemplares cultivados em Ribeirão Preto, SP, através da análise das substâncias presentes nas folhas, ao longo das quatro estações em dois anos. O teor de fenóis totais variou de 21,96 a 45,92 mg / g (expresso em mg ácido tânico / g folhas secas), de 55,88 a 93,20 mg triterpenos totais / g folhas secas, e de 1,349 a 3,859 mg flavonóides / g folhas secas. A amostra Primavera 92 apresentou o maior teor de flavonóides e fenóis totais, e a amostra Inverno 94 apresentou o maior teor de triterpenos.

The chemical seasonal variability of Maytenus aquifolium Mart. (Celastraceae), a Brazilian medicinal plant known as "espinheira santa", was investigated on specimens cultivated at Ribeirão Preto SP, Brazil, at the four seasons of two years. The content of total phenolics, flavonoids and triterpenes varied from 21.96 to 45.92 mg total phenolics (expressed as tanic acid) / g dried leaves, 55.88 - 93.20 mg total triterpenes / g dried leaves, and 1.349 - 3.859 mg flavonoid / g dried leaves. Sample Spring 92 had the higher content of flavonoids and total phenolics, while sample Winter 94 had the highest content of triterpenes.