Methodologies In Conjunction Research Articles

Dereplication and chemotaxonomical studies of marine algae of the Ochrophyta and Rhodophyta phyla.

Dereplication and chemotaxonomic studies of six marine algae of the Ochrophyta and one of the Rhodophyta phyla resulted in the detection of 22 separate compounds. All 16 secondary metabolites, including four new compounds (16–19), could be rapidly dereplicated using HPLC-NMR and HPLC-MS methodologies in conjunction with the MarinLit database. This study highlights the advantages of using NMR data (acquired via HPLC-NMR) for database searching and for the overall dereplication of natural products.

Numerical Prediction of Differences in Micromechanical Fracture Sequence in Notched and Un-Notched Wires for Civil Engineering Applications

The need to employ simulation-based investigations of fracture of civil engineering structures has been emphasised recently in literature. In this paper, three dimensional finite element simulations revealed that the cup and cone fracture process in channel-shaped notched wire for civil engineering applications begins with a slant fracture at the root of the outer edge of the channel-shaped notch and follows a slant to flat fracture sequence. These results demonstrate that the fracture origin in cup and cone fracture in notched wires for civil engineering applications does not necessarily begin with a flat fracture at the center of the wire and does not necessarily follows the flat to slant fracture sequence or propagation generally reported in literature. These results further demonstrate the need to employ simulation-based methodologies in conjunction with or as an alternative to purely experimental fractographic analysis for an accurate failure analysis of wires used for civil engineering applications.

Precise Synthesis of Block Polymers Composed of Three or More Blocks by Specially Designed Linking Methodologies in Conjunction with Living Anionic Polymerization System

This article reviews the successful development of two specially designed linking methodologies in conjunction with a living anionic polymerization system for the synthesis of novel multiblock polymers, composed of three or more blocks, difficult to be synthesized by sequential polymerization. The first methodology with the use of a new heterofunctional linking agent, 2-(4-chloromethylphenyl)ethyldimethylchlorosilane (1), was developed for the synthesis of multiblock polymers containing poly(dimethylsiloxane) (PDMS) blocks. This methodology is based on the selective reaction of the chain-end silanolate anion of living PDMS, with the silyl chloride function of 1, and subsequent linking reaction of the resulting ω-chain-end-benzyl chloride-functionalized polymer with either a living anionic polymer or living anionic block copolymer. With this methodology, various multiblock polymers containing PDMS blocks, up to the pentablock quintopolymer, were successfully synthesized. The second methodology using an α-phenylacrylate (PA) reaction site was developed for the synthesis of multiblock polymers composed of all-vinyl polymer blocks. In this methodology, an α-chain-end-PA-functionalized polymer or block copolymer, via the living anionic polymerization, was first prepared and, then, reacted with appropriate living anionic polymer or block copolymer to link the two polymer chains. As a result, ACB (BCA), BAC (CAB), (AB)n, (AC)n, ABA, ACA, BCB, and ABCA multiblock polymers, where A, B, and C were polystyrene, poly(2-vinylpyridine), and poly(methyl methacrylate) segments, could be successfully synthesized. The synthesis of triblock copolymers, BAB, CAC, and CBC, having molecular asymmetry in both side blocks, was also achieved. Furthermore, the use of living anionic polymers, derived from many other monomers, categorized as either of styrene, 2-vinylpyridine, or methyl methacrylate in monomer reactivity, in the linking methodology enabled the number of synthetically possible block polymers to be greatly increased. Once again, all of the block polymers synthesized by these methodologies are new and cannot be synthesized at all by sequential polymerization. They were well-defined in block architecture and precisely controlled in block segment.

Screening for Modulatory Effects on Steroidogenesis Using the Human H295R Adrenocortical Cell Line: A Metabolomics Approach

The recently OECD validated H295R steroidogenesis assay provides an in vitro alternative to evaluate the potential interference of exogenous compounds with endogenous steroid hormone synthesis. Currently, this assay is used for a simple negative-positive screening of compounds using testosterone and estradiol levels as end points, measured with specific enzyme immunoassays (EIAs) or targeted liquid chromatography (LC) and gas chromatography (GC)-mass spectrometry (MS) methods. However, recent developments in LC-MS and bioinformatics allow for more comprehensive approaches to evaluate changes in steroid profiles. In the current work, the H295R cell model was combined with a metabolomics approach to monitor changes in metabolite profiles in both a targeted and untargeted way. H295R cells were exposed for 48 h to model compounds, i.e., forskolin, abiraterone, prochloraz, ketoconazole, trilostane, formestane, aminoglutethimide, fadrozole, etomidate, and metyrapone, known to affect steroidogenesis. After exposure, the levels of 9 natural steroids were determined by a quantitative targeted GC-MS/MS method and compared to a metabolomics method using Ultra Performance Liquid Chromatography-Time-of-Flight-Mass Spectrometry (UPLC-ToF-MS). Like the EIAs, both methods were suited for negative-positive screening, but the MS methods also generated specific fingerprints, allowing chemical class prediction of the compound under investigation. Although the targeted GC-MS/MS was more sensitive, which was an advantage regarding analysis of the estrogens 17β-estradiol and estrone, the untargeted UPLC-ToF-MS was able to evaluate effects on the synthesis of the corticosteroids. Moreover, untargeted comparison of the aligned chemical profiles allowed identification of all m/z-values that are differential between exposed and nonexposed H295R cells. In conclusion, application of a comprehensive metabolite profiling methodology not only provides a tool to screen compounds for steroidogenic modulating properties, but also allows chemical class prediction. As such, steroid profiling methodologies in conjunction with the H295R assay can contribute to the prioritization of chemicals for additional safety testing.

Asymptotic Tracking for Aircraft via Robust and Adaptive Dynamic Inversion Methods

Two asymptotic tracking controllers are designed in this paper, which combine model reference adaptive control and dynamic inversion methodologies in conjunction with the robust integral of the signum of the error (RISE) technique for output tracking of an aircraft system in the presence of parametric uncertainty and unknown, nonlinear disturbances, which are not linearly parameterizable (non-LP). The control designs are complicated by the fact that the control input is multiplied by an uncertain, non-square matrix. A robust control design is presented first, in which partial knowledge of the aircraft model along with constant feedforward estimates of the unknown input parameters are used with a robust control term to stabilize the system. Motivated by the desire to reduce the need for high-gain feedback, an adaptive extension is then presented, in which feedforward adaptive estimates of the input uncertainty are used. These results show how asymptotic tracking control can be achieved for a nonlinear system in the presence of a non-square input matrix containing parametric uncertainty and nonlinear, non-LP disturbances. Asymptotic output tracking is proven via Lyapunov stability analysis, and high-fidelity simulation results are provided to verify the efficacy of the proposed controllers.

Theoretical Study of the Structure and Electronic Properties of Si3On(-) and Si6On(-) (n = 1-6) Clusters. Fragmentation and Formation Patterns.

A theoretical study of two series of small clusters, Si3On(-) and Si6On(-) (n = 1-6), has been carried out. The minimum energy structures were produced adding an electron to neutral species followed by relaxation at the B3LYP-6-311G(2d) level. The vertical ionization energies (VIEs) were computed using the electron propagator theory (EPT) in two approximations, Unrestricted Outer Valence Green Functions (UOVGF) and partial third-order approximation (P3). In the series Si3On(-) the theoretical VIEs of the minimum energy structures agree well with experimental data. For the second series there are not experimental VIEs, and the theoretical results are predictions. The performance of EPT methodologies in conjunction with all-electron or pseudopotentials (PP) calculations is analyzed. The conjunction of P3 and PP approximation proves to be the most efficient and economical methodology to calculate the VIEs of small anionic silicon oxide clusters. In the series Si6On(-) different channels of fragmentation have been calculated. The results suggest that the fragments do not have drastic geometric changes and the anionic fragment corresponds to the atoms where the spin density of the initial large cluster is localized. The Fukui function calculated over selected optimized fragments predicts adequately the interaction between them to form large stable clusters.

Recent Advances in Performance Evaluation and Flexural Response of Existing Bridges

This paper presents a comprehensive overview of recent developments in the performance evaluation and flexural response of existing bridges from fundamental concepts to advanced topics. The focus is particularly on the structural evaluation to enhance the quality assessment of constructed bridge superstructures, taking into account a synthesis of the most important contributions to the flexural behavior of existing bridges, such as dynamic responses and lateral load distributions. This state-of-the-art paper provides a critical review of published literature and existing assessment methodologies in conjunction with corresponding analysis techniques, including technical comments on the code provisions. Detailed descriptions of the important parameters influencing flexural responses of existing bridges are discussed, such as the geometric effect, loading configuration, present condition of a bridge, and contribution of secondary structural elements. The review also includes the investigation methodologies, namely, the diagnostic and proof load tests. Finally, the current research needs to further advance the performance evaluation technologies for existing bridge superstructures are recommended.

Rhombomere-specific analysis reveals the repertoire of genetic cues expressed across the developing hindbrain

BackgroundThe Hox family of homeodomain transcription factors comprises pivotal regulators of cell specification and identity during animal development. However, despite their well-defined roles in the establishment of anteroposterior pattern and considerable research into their mechanism of action, relatively few target genes have been identified in the downstream regulatory network. We have sought to investigate this issue, focussing on the developing hindbrain and the cranial motor neurons that arise from this region. The reiterated anteroposterior compartments of the developing hindbrain (rhombomeres (r)) are normally patterned by the combinatorial action of distinct Hox genes. Alteration in the normal pattern of Hox cues in this region results in a transformation of cellular identity to match the remaining Hox profile, similar to that observed in Drosophila homeotic transformations.ResultsTo define the repertoire of genes regulated in each rhombomere, we have analysed the transcriptome of each rhombomere from wild-type mouse embryos and not those where pattern is perturbed by gain or loss of Hox gene function. Using microarray and bioinformatic methodologies in conjunction with other confirmatory techniques, we report here a detailed and comprehensive set of potential Hox target genes in r2, r3, r4 and r5. We have demonstrated that the data produced are both fully reflective and predictive of rhombomere identity and, thus, may represent some the of Hox targets. These data have been interrogated to generate a list of candidate genes whose function may contribute to the generation of neuronal subtypes characteristic of each rhombomere. Interestingly, the data can also be classified into genetic motifs that are predicted by the specific combinations of Hox genes and other regulators of hindbrain anteroposterior identity. The sets of genes described in each or combinations of rhombomeres span a wide functional range and suggest that the Hox genes, as well as other regulatory inputs, exert their influence across the full spectrum of molecular machinery.ConclusionWe have performed a systematic survey of the transcriptional status of individual segments of the developing mouse hindbrain and identified hundreds of previously undescribed genes expressed in this region. The functional range of the potential candidate effectors or upstream modulators of Hox activity suggest multiple unexplored mechanisms. In particular, we present evidence of a potential new retinoic acid signalling system in ventral r4 and propose a model for the refinement of identity in this region. Furthermore, the rhombomeres demonstrate a molecular relationship to each other that is consistent with known observations about neurogenesis in the hindbrain. These findings give the first genome-wide insight into the complexity of gene expression during patterning of the developing hindbrain.

Vegetable Waste Treatment: Comparison and Critical Presentation of Methodologies

Vegetable industries have been considered responsible for a great amount of pollution; hence, there has been a strong need for the optimization of vegetable waste treatment systems. The currently employed systems are numerous and fall in the following large categories; thermal processes, evaporation, membrance processes, anaerobic digestion, anaerobic co-digestion, biodiesel spraying, combustion, transesterification, coagulation, and composting. Respective methodologies in conjunction with waste treatment methods were presented per waste treatment method. The comparative presentation of the various vegetable waste treatment methodologies showed that though anaerobic digestion stands for the most enviromentally friendly technique, its required longer treatment time in conjuction with its weakness to deal with elemental contaminants makes imperative the employment of a second alternative technique which could either be a membrance process (low energy cost, reliability, reduced capital cost) or a coagulation/flocculation method because of its low cost and high effectiveness. Biogas production appears to be another promising and energy effective waste treatment method. On the other hand, methods like distillation and ozonation (high cost) and electrolysis (experimental level) have not been employed in the field. Finally, new waste management technologies have been described.

An Object-oriented Methodology for Embedded Real-time Systems

The usage of object-oriented methodologies in conjunction with formal description techniques has arisen as a promising way of dealing with the increasing complexity of embedded real-time systems. These methodologies are currently well supported by a set of tools that allow the specification, simulation and validation of the functional aspects of these systems. However, most of these methodologies do not take into account non-functional aspects such as hardware interaction and real-time constraints, which are especially important in the context of this kind of system. Based on our experiences in developing embedded real-time systems, we present a new methodology to design them. This methodology is based on a combination of ideas from different existing methodologies (UML, OCTOPUS, etc.) together with the integration of rate-monotonic analysis in the context of the SDL formal description technique development cycle. Additionally, in order to get this integration, a real-time execution model for SDL is presented to allow us to express hard real-time constraints. The methodology pays special attention to the transition from the object model to the task model, taking into account real-time and hardware integration issues. We also illustrate our proposal by applying it to the development of a multi-handset cordless telephone.

Estimating the plastic collapse of pressure vessels using plasticity contours

This investigation concerns the use of non-linear finite element analysis (FEA) to predict the plastic collapse of pressure vessels with and without defects. Computational non-linear plastic analysis of pressure vessels to estimate the collapse pressure is an iterative process. As a consequence, if the models are large and complex, considerable computer resources and time are required to generate solutions. As normal materials tend not to be ideally elastic–plastic (no sudden plastic collapse), a number of techniques are used to estimate the collapse pressure based on the FEA solution. These techniques often rely on plotting the pressure–strain curve; however, other techniques are also possible and are considered here. The aim of this paper is to assess the different methodologies in conjunction with FEA. Firstly, to consider their accuracy and secondly, to ascertain whether a particular technique would result in a reduction in the analysis time necessary to recover the plastic collapse pressure. Of the methods analysed, one based on the measurement (as a percentage) of the leading edge of the effective plastic strain (EPS) contour band showed promise. Under increasing pressures, regardless of geometry or material yield curve, the %EPS contour area coverage displays linear behaviour before through-wall plasticity is achieved. Based on this it is possible to extrapolate from low levels of plastic deformation up to plastic collapse pressures without requiring additional non-linear analyses, resulting in significant cost reductions in computational analysis times.

Eukaryotic genome organization analyzed by electron microscope in situ hybridization

In situ hybridization is a powerful tool for the localization of DNA/RNA sequences in nuclei and chromosomes. The introduction of nonisotopic labelling methodologies in conjunction with fluorescent or enzyme-linked detection have resulted in a dramatic increase in the application of this technique at the light microscope (LM) level and has placed it in a pivotal role in cell biology, development and genetics. Development of equivalent mapping protocols at the EM level offers increased spatial resolution. We have combined the use of nonisotopic probes with invmunogold labelling to investigate eukaryotic genome organization at high resolution.Metaphase chromosomes released from mitotically-arrested cells are deposited on gold EM grids by centrifugation through a sucrose cushion. After fixation (0.1% glutaraldehyde, 20 min) and DNA denaturation, chromosomes are hybridized to cloned probes enzymatically labelled with biotin-dUTP, digoxigenin-dUTP, dinitrophenyl-dUTP or covalently coupled to N-acetoxyacetoaminofluorene. Hybrid sites typically are detected by a two-step antibody incubation and 1-30 nm colloidal gold particles.

Resolution of Indeterminate HIV-1 Test Data Using the Department of Defense HIV-1 Testing Program

The Department of Defense HIV-1 Test Program was designed to provide timely, accurate, and cost-effective HIV-1 testing services for military personnel. This program incorporated a comprehensive quality assurance and quality improvement program in conjunction with large-scale commercial contracts. Definitive diagnostic data were provided for all sera, and nondiagnostic data were resolved as either reactive or nonreactive using the described guidelines and test system. The program used Food and Drug Administration (FDA)-approved HIV-1 ELISA and Western blot methodologies in conjunction with CBre3 recombinant protein and radioimmunoprecipitation (RIPA) assays. The program has evaluated over 4.7 million serum samples. The ELISA repeat reactive rate was approximately 0.6%, and when Western blot and CBre3 tests were performed with the specified criteria, nondiagnostic Western blot data were reduced from 10% to 1%. The remaining nondiagnostic serum samples were analyzed with the third confirmatory procedure, RIPA, which successfully resolved all but 19 samples (0.0004%). These remaining samples were eventually resolved when additional serum samples were retested as previously described. The overall HIV-1 prevalence was determined to be 0.33/1,000, and the program at an average test cost of $4.55 successfully identified over 2,500 infected individuals. This testing program, which required the analysis of two separate specimens, had a false-positive rate estimated at 1 in 1,000,000.