Arturo Navarro-Ocaña

Application of metabolic engineering to the production of scopolamine.

Scopolamine is an alkaloid widely used in medicine for its anticholinergic activity. The aim of this review is to show that metabolic engineering techniques constitute a suitable tool to improve the production of tropane alkaloids, focusing in particular on scopolamine. We present an overview of results obtained by various research groups, including our own, who have studied the overexpression of genes involved in the biosynthesis of scopolamine in different plant species that produce tropane alkaloids. Experiments carried out to improve production in hairy root cultures will also be described, as well as those attempting to biotransform hyoscyamine into scopolamine in roots and transgenic tobacco cells.

The Pentacyclic Triterpenes , -amyrins: A Review of Sources and Biological Activities

Pentacyclic triterpenes are ubiquitously distributed throughout the plant kingdom, in a free form as aglycones or in combined forms, and have long been known to have a number of biological effects. The compounds-amyrin and -amyrin are commonly found in medicinal plants and oleo-resin obtained by bark incision of several species of Bursera or Protium of the Burseraceae family. Both in vitro and in vivo studies have shown that amyrin also has important biological functions.

ANCUT2, an Extracellular Cutinase from Aspergillus nidulans Induced by Olive Oil

Cutinases are versatile carboxylic ester hydrolases with great potential in many biocatalytic processes, including biodiesel production. Genome sequence analysis of the model organism Aspergillus nidulans reveals four genes encoding putative cutinases. In this work, we purified and identified for the first time a cutinase (ANCUT2) produced by A. nidulans. ANCUT2 is a 29-kDa protein which consists of 255 amino acid residues. Comparison of the amino acid sequence of ANCUT2 with other microbial cutinase sequences revealed a high degree of homology with other fungal cutinases as well as new features, which include a serine-rich region and conserved cysteines. Cutinase production with different lipidic and carbon sources was also explored. Enzyme activity was induced by olive oil and some triacylglycerides and fatty acids, whereas it was repressed by glucose (1%) and other sugars. In some conditions, a 22-kDa post-translational processing product was also detected. The cutinase nature of the enzyme was confirmed after degradation of apple cutin.

Immobilization and Biochemical Properties of the Enantioselective Recombinant NStcI Esterase of Aspergillus nidulans

The recombinant NStcI A. nidulans esterase was adsorbed on Accurel MP1000, where protein yield and immobilization efficiency were 42.48% and 81.94%, respectively. Storage stability test at 4°C and RT showed 100% of residual activity after 40 days at both temperatures. The biocatalyst retains more than 70% of its initial activity after 3 cycles of repeated use. Biochemical properties of this new biocatalyst were obtained. Maximum activity was achieved at pH 11 and 30°C, while the best stability was observed with the pH between 9 and 11 at 40°C. NStcI thermostability was increased after immobilization, as it retained 47.5% of its initial activity after 1 h at 60°C, while the free enzyme under the same conditions displayed no activity. NStcI preserved 70% of its initial activity in 100% hexane after 72 h. Enzymatic kinetic resolution of (R,S)-1-phenylethanol was chosen as model reaction, using vinyl acetate as acyl donor. After optimization of reaction parameters, the highest possible conversion (42%) was reached at 37°C, a w of 0.07, and 120 h of bioconversion in hexane with an enantiomeric excess of 71.7%. NStcI has selectivity for (R)-enantiomer. The obtained E value (31.3) is in the range considered useful to resolve enantiomeric mixtures.

Growth Inhibition of Streptococcus from the Oral Cavity by α-Amyrin Esters

Five terpenoids were tested by the macrodilution broth method to determine their inhibition activity on cariogenic bacterial growth. In general, α-, β-amyrin and α-amyrin phenylacetate proved to be active, reducing the bacterial viability to less than 20%.

Effect of postharvest UV-C treatment on the bacterial diversity of Ataulfo mangoes by PCR-DGGE, survival of E. coli and antimicrobial activity

Since Mexico is the second largest exporter of mangoes, its safety assurance is essential. Research in microbial ecology and knowledge of complex interactions among microbes must be better understood to achieve maximal control of pathogens. Therefore, we investigated the effect of UV-C treatments on bacterial diversity of the Ataulfo mangoes surface using PCR-DGGE analysis of variable region V3 of 16S rRNA genes, and the survival of E. coli, by plate counting. The UV-C irradiation reduced the microbial load on the surface of mangoes immediately after treatment and the structure of bacterial communities was modified during storage. We identified the key members of the bacterial communities on the surface of fruits, predominating Enterobacter genus. Genera as Lactococcus and Pantoea were only detected on the surface of non-treated (control) mangoes. This could indicate that these genera were affected by the UV-C treatment. On the other hand, the treatment did not have a significant effect on survival of E. coli. However, genera that have been recognized as antagonists against foodborne pathogens were identified in the bands patterns. Also, phenolic compounds were determined by HPLC and antimicrobial activity was assayed according to the agar diffusion method. The main phenolic compounds were chlorogenic, gallic, and caffeic acids. Mango peel methanol extracts (UV-C treated and control mangoes) showed antimicrobial activity against strains previously isolated from mango, detecting significant differences (P < 0.05) among treated and control mangoes after 4 and 12 days of storage. Ps. fluorescens and Ps. stutszeri were the most sensitive.

Reductive cyclization with baker's yeast of 4-alkyl-2-nitroacetanilides to 6-alkylbenzimidazoles and 1-hydroxy-2-methyl-6-alkylbenzimidazoles

Reduction of 4-substituted 2-nitroacetanilides by bakers yeast in acid media effected cyclization, resulting in the formation of 6-substituted 2-methylbenzimidazoles and 6-substituted 1-hydroxy-2-methylbenzimidazole via the chemo- and regioselective reduction of the 2-nitro aromatic group to amine or hydroxylamine.

Evaluation of the hydroxynitrile lyase activity in cell cultures of capulin (Prunus serotina).

Enzymatic preparations obtained from young plants and cell cultures of capulin were screened for hydroxynitrile lyase activity. The three week old plants, grown under sterile conditions, were used to establish a solid cell culture. Crude preparations obtained from this plant material were evaluated for the transformation of benzaldehyde to the corresponding cyanohydrin (mandelonitrile). The results show that the crude material from roots, stalks, and leaves of young plants and calli of roots, stalks, internodes and petioles biocatalyzed the addition of hydrogen cyanide (HCN) to benzaldehyde with a modest to excellent enantioselectivity.