Steroids and related products. LIV. The synthesis of 11-oxa steroids. VI. The synthesis of 11-oxatestosterone

The synthesis of 11-oxatestosterone from 11-oxa-5 alpha-androstane-3,17-dione, which is available from hecogenin, is described. The product shows, in comparison with the natural hormone, diminished androgenic and anabolic activities.     

Evaluation of orange-fleshed sweet potato (<Emphasis Type="Italic">Ipomoea batatas</Emphasis> L.) genotypes for salt tolerance through shoot apex culture under in vitro NaCl mediated salinity stress conditions

Fifteen genotypes of sweet potato were evaluated for salinity stress tolerance under in vitro NaCl mediated salinity stress conditions (MS, MS + 0.5% and MS + 1.0% NaCl). The growth parameters such as number of leaves, number of shoots, number of roots, length of plantlets and length of roots decreased significantly among the genotypes with increase in level of salinity. Of the 15 genotypes tested, six genotypes (108X1, 90/606, 90/696, CIP 8, S-30X15 and SP-61) were unable to sprout even at 0.5% NaCl and were characterized as susceptible to salt stress, three genotypes (CIP 6, 90/774 and CIP 3) which could tolerate 0.5% NaCl as moderately tolerant and six genotypes (CIP 12, CIP 13, JO 14, JP 13, SB-198/115 and Gouri) as tolerant to salinity at 1.0% NaCl. Amongst the six genotypes showing tolerance to 1.0% NaCl, the exotic genotypes––JP 13, CIP 12 and indigenous one SB-198/115 continued to exhibit significant higher values for growth parameters over the susceptible one. Based on the performance under NaCl mediated salinity stress (1.0%), the pattern of salinity tolerance in the genotypes through shoot apex culture was JP 13 > SB-198/115 > JO 14 > Gouri > CIP 12 > CIP 13. The effect of salt stress on the activity of antioxidative enzymes was studied in leaves of 8-week-old plantlets of those six genotypes, which responded at higher NaCl stress along with a susceptible genotype 90/606. In leaves of salt stressed plants, superoxide dismutase (SOD), guaiacol peroxidase (GPX) and catalase (CAT) activities increased when compared with the stress free control. The increase was more pronounced in the tolerant genotypes than that in the susceptible one. These results indicate that oxidative stress may play an important role in salt stressed sweet potato plants and that the greater protection of tolerant plants from salt induced oxidative damage results, at least in part, through the increase in the activity of antioxidant enzymes.     

Estimating dataset size requirements for classifying DNA microarray data.

A statistical methodology for estimating dataset size requirements for classifying microarray data using learning curves is introduced. The goal is to use existing classification results to estimate dataset size requirements for future classification experiments and to evaluate the gain in accuracy and significance of classifiers built with additional data. The method is based on fitting inverse power-law models to construct empirical learning curves. It also includes a permutation test procedure to assess the statistical significance of classification performance for a given dataset size. This procedure is applied to several molecular classification problems representing a broad spectrum of levels of complexity.     

Monitoring of biofilms grown on differentially structured metallic surfaces using confocal laser scanning microscopy

Imaging of biofilms on opaque surfaces is a challenge presented to researchers especially considering pathogenic bacteria, as those typically grow on living tissue, such as mucosa and bone. However, they can also grow on surfaces used in industrial applications such as food production, acting as a hindrance to the process. Thus, it is important to understand bacteria better in the environment they actually have relevance in. Stainless steel and titanium substrata were line structured and dotted surface topographies for titanium substrata were prepared to analyze their effects on biofilm formation of a constitutively green fluorescent protein (GFP)‐expressing Escherichia coli strain. The strain was batch cultivated in a custom built flow cell initially for 18 h, followed by continuous cultivation for 6 h. Confocal laser scanning microscopy (CLSM) was used to determine the biofilm topography. Biofilm growth of E. coli GFPmut2 was not affected by the type of metal substrate used; rather, attachment and growth were influenced by variable shapes of the microstructured titanium surfaces. In this work, biofilm cultivation in flow cells was coupled with the most widely used biofilm analytical technique (CLSM) to study the time course of growth of a GFP‐expressing biofilm on metallic surfaces without intermittent sampling or disturbing the natural development of the biofilm.     

E,E,E-1-(4-Arylamino-4-oxo-2-butenoyl)-3,5-bis(arylidene)-4-piperidones: a topographical study of some novel potent cytotoxins

A series of E,E,E-3,5-bis(arylidene)-1-(4-arylamino-4-oxo-2-butenoyl)-4-piperidones 4 (phenylidene) and 5 (4-nitrophenylidene) were prepared in order to explore the structural features of the N-acyl group which affects the cytotoxic potency. Evaluation toward human Molt 4/C8 and CEM T-lymphocytes revealed that many of the IC(50) figures were submicromolar and lower than melphalan. Marked inhibitory potencies toward murine leukemia L1210 cells were also noted. When evaluated against a panel of human tumor cell lines, three representative compounds in series 4 displayed selective toxicity to leukemia and colon cancer cell lines and were significantly more potent than the reference drug melphalan. Molecular modeling of representative compounds in both series 4 and the analogs, in which the configuration of the olefinic double bond was changed from E to Z (series 3), revealed that the torsion angles of the arylidene aryl rings and locations of the terminal arylaminocarbonyl groups may have contributed to the greater cytotoxic properties displayed in 3. Compounds 4c (3,4-dichlorophenylamino), d (4-methylphenylamino) and 5c (3,4-dichlorophenylamino), d (4-methylphenylamino) inhibited the activity of human N-myristoyltransferase by approximately 50% at concentrations of 50-100 microM. The compounds in series 4 and 5 were well tolerated in a short-term toxicity study in mice.     

Niche-based prediction of establishment of biocontrol agents: an example with Gratiana boliviana and tropical soda apple

Successful establishment of a biological control agent is a prerequisite for effective reduction of an invasive weed. Niche-based species distribution models can generate valuable information about the potential spread of a biological control agent and help to predict its distribution. The Maximum Entropy Species Distribution Model was used in our study to predict distributions of the leaf beetle Gratiana boliviana Spaeth (Coleoptera: Chrysomelidae) and its target weed, tropical soda apple (TSA), Solanum viarum Dunal (Solanaceae). The specific objectives of this study were to (1) assess the climatic suitability for establishment of this insect across the invasive range of the weed by overlapping predicted current distributions and, (2) examine the niche-related restriction in distribution of the insect by visualising the predictive niche in multivariate space. Accuracy of predicted distributions was tested using binomial tests and area under the curve scores. The results of statistical tests confirmed that the predictions were significantly better than random. The predictions indicated that the potential distribution of G. boliviana in the USA will be more restricted than that of its host plant. Consequently, the beetle's ability to inflict damage to TSA will be geographically limited. Niche visualisation, using a PCA-based analysis, provided evidence of the niche imposed restriction on the distribution of G. boliviana. Overall, this study proposes a new approach for understanding the spatial limitations in establishment of biological control agents, allowing researchers to establish more realistic expectations of success.     

Potent Dengue Virus Neutralization by a Therapeutic Antibody with Low Monovalent Affinity Requires Bivalent Engagement

We recently described our most potently neutralizing monoclonal antibody, E106, which protected against lethal Dengue virus type 1 (DENV-1) infection in mice. To further understand its functional properties, we determined the crystal structure of E106 Fab in complex with domain III (DIII) of DENV-1 envelope (E) protein to 2.45 Å resolution. Analysis of the complex revealed a small antibody-antigen interface with the epitope on DIII composed of nine residues along the lateral ridge and A-strand regions. Despite strong virus neutralizing activity of E106 IgG at picomolar concentrations, E106 Fab exhibited a ∼20,000-fold decrease in virus neutralization and bound isolated DIII, E, or viral particles with only a micromolar monovalent affinity. In comparison, E106 IgG bound DENV-1 virions with nanomolar avidity. The E106 epitope appears readily accessible on virions, as neutralization was largely temperature-independent. Collectively, our data suggest that E106 neutralizes DENV-1 infection through bivalent engagement of adjacent DIII subunits on a single virion. The isolation of anti-flavivirus antibodies that require bivalent binding to inhibit infection efficiently may be a rare event due to the unique icosahedral arrangement of envelope proteins on the virion surface.     

A new nitrilase from Bradyrhizobium japonicum USDA 110. Gene cloning, biochemical characterization and substrate specificity

A nitrilase gene blr3397 from Bradyrhizobium japonicum USDA110 was cloned and over-expressed in Escherichia coli, and the encoded protein was purified to give a nitrilase with a single band of about 34.5kD on SDS-PAGE. The molecular weight of the holoenzyme was about 340kD as determined by light scattering analysis, suggesting that nitrilase blr3397 self-aggregated to an active form with the native structure being a decamer. The V(max) and K(m) for phenylacetonitrile were 3.15U/mg and 4.36mM, respectively. The catalytic constant k(cat) and specificity constant k(cat)/K(m) were 111min(-1) and 2.6x10(4)min(-1)M(-1). This nitrilase is most active toward the hydrolysis of hydrocinnamonitrile among the tested substrates (4.3 times that of phenylacetonitrile). The nitrilase blr3397 shows higher activity towards the hydrolysis of aliphatic nitriles than that for the aromatic counterparts, and can be characterized as an aliphatic nitrilase in terms of activity. This nitrilase also possesses distinct features from the nitrilase bll6402 of the same microbe.     

Genomic relations among 31 species of Mammillaria haworth (Cactaceae) using random amplified polymorphic DNA

Thirty-one species of Mammillaria were selected to study the molecular phylogeny using random amplified polymorphic DNA (RAPD) markers. High amount of mucilage (gelling polysaccharides) present in Mammillaria was a major obstacle in isolating good quality genomic DNA. The CTAB (cetyl trimethyl ammonium bromide) method was modified to obtain good quality genomic DNA. Twenty-two random decamer primers resulted in 621 bands, all of which were polymorphic. The similarity matrix value varied from 0.109 to 0.622 indicating wide variability among the studied species. The dendrogram obtained from the unweighted pair group method using arithmetic averages (UPGMA) analysis revealed that some of the species did not follow the conventional classification. The present work shows the usefulness of RAPD markers for genetic characterization to establish phylogenetic relations among Mammillaria species.