Development of combined micro -preparation, separation, and mass spectrometry methods and application in the microdialysis study of neuropeptide metabolism

Two sets of mass spectrometry-based methods were developed specifically for the in vivo study of extracellular neuropeptide biochemistry. First, an integrated micro-concentration/desalting/matrix-addition device was constructed for matrix-assisted laser desorption ionization mass spectrometry (MALDI MS) to achieve attomole sensitivity for microdialysis samples. Second, capillary electrophoresis (CE) was incorporated into the above micro-liquid chromatography (LC) and MALDI MS system to provide two-dimensional separation and identification (i.e. electrophoretic mobility and molecular mass) for the analysis of complex mixtures. The latter technique includes two parts of instrumentation: (1) the coupling of a preconcentration LC column to the inlet of a CE capillary, and (2) the utilization of a matrix-precoated membrane target for continuous CE effluent deposition and for automatic MALDI MS analysis (imaging) of the CE track.^ Initial in vivo data reveals a carboxypeptidase A (CPA) activity in rat brain involved in extracellular neurotensin metabolism. Benzylsuccinic acid, a CPA inhibitor, inhibited neurotensin metabolite NT1-12 formation by 70%, while inhibitors of other major extracellular peptide metabolizing enzymes increased NT1-12 formation. CPA activity has not been observed in previous in vitro experiments. Next, the validity of the methodology was demonstrated in the detection and structural elucidation of an endogenous neuropeptide, (L)VV-hemorphin-7, in rat brain upon ATP stimulation. Finally, the combined micro-LC/CE/MALDI MS was used in the in vivo metabolic study of peptide E, a mu-selective opioid peptide with 25 amino acid residues. Profiles of 88 metabolites were obtained, their identity being determined by their mass-to-charge ratio and electrophoretic mobility. The results indicate that there are several primary cleavage sites in vivo for peptide E in the release of its enkephalin-containing fragments. ^

Making genetic biodiversity measurable : a review of statistical multivariate methods to study variability at gene level

Measures of agro-ecosystems genetic variability are essential to sustain scientific-based actions and policies tending to protect the ecosystem services they provide. To build the genetic variability datum it is necessary to deal with a large number and different types of variables. Molecular marker data is highly dimensional by nature, and frequently additional types of information are obtained, as morphological and physiological traits. This way, genetic variability studies are usually associated with the measurement of several traits on each entity. Multivariate methods are aimed at finding proximities between entities characterized by multiple traits by summarizing information in few synthetic variables. In this work we discuss and illustrate several multivariate methods used for different purposes to build the datum of genetic variability. We include methods applied in studies for exploring the spatial structure of genetic variability and the association of genetic data to other sources of information. Multivariate techniques allow the pursuit of the genetic variability datum, as a unifying notion that merges concepts of type, abundance and distribution of variability at gene level.

An environmental magnetic study, sedimentological and numerical methods around Tauranga Harbour, New Zealand

Magnetic iron minerals are widespread and indicative sediment constituents in estuarine, coastal and shelf systems. We combine environmental magnetic, sedimentological and numerical methods to identify magnetite-enriched placer-like zones in a complex coastal system and delineate their formation mechanisms. Magnetic susceptibility and remanence measurements on 245 surficial sediment samples collected in and around Tauranga Harbour, the largest barrier-enclosed tidal estuary of New Zealand, reveal several discrete enrichment zones controlled by local hydrodynamic conditions. Active magnetite enrichment takes place in tidal channels, which feed into two coast-parallel nearshore magnetite-enriched belts centered at water depths of 6-10 m and 10-20 m. A close correlation between magnetite content and magnetic grain size was found, where higher susceptibility values are associated within coarser magnetic crystal sizes. Two key mechanisms for magnetite enrichment are identified. First, tide-induced residual currents primarily enable magnetite enrichment within the estuarine channel network. A coast-parallel, fine sand magnetite enrichment belt in water depths of less than 10 m along the barrier island has a strong decrease in magnetite content away from the southern tidal inlet and is apparently related to active coast-parallel transport combined with mobilizing surf zone processes. A second, less pronounced, but more uniform magnetite enrichment belt at 10-20 m water depth is composed of non-mobile, medium-coarse-grained relict sands, which have been reworked during post-glacial sea level transgression. We demonstrate the potential of magnetic methods to reveal and differentiate coastal magnetite enrichment patterns and investigate their formative mechanisms.

Orbital control on carbonate-lignite cycles on sediment core KAP-107 in the Ptolemais Basin, northern Greece

Time control is essential for the reconstruction of geological processes. We use a combination of relative and absolute methods to establish the chronology and related paleoclimatic processes for Late Neogene lacustrine sediment from the Ptolemais Basin, northern Greece. We determined changes in magnetic polarity and correlated them to the global magnetic polarity time scale, which again is calibrated by radiometric methods, to provide a low-resolution age model for the Upper Miocene to Lower Pliocene (7 - 3 Ma). Sedimentary successions show rhythmic alterations of lignites, clays, and marls. Using photospetrometry we measured this variability at 1-cm resolution, and correlated the pattern to known changes in earth's orbital parameters, namely to eccentricity and precession. For 230-m long borehole KAP-107 from the Amynteon Sub-Basin we obtained a high-resolution age model that spans 2 myr from 5.1 to 3.1 Ma, with age control points at insolation maxima (20-kyr resolution). We recommend using photospectrometry as reliable tool to establish orbital-based chronologies and to reconstruct paleoclimate variability at high resolution.

Deep levels in a-plane, high Mg-content MgxZn1-xO epitaxial layers grown by molecular beam epitaxy

Deep level defects in n-type unintentionally doped a-plane MgxZn1−xO, grown by molecular beam epitaxy on r-plane sapphire were fully characterized using deep level optical spectroscopy (DLOS) and related methods. Four compositions of MgxZn1−xO were examined with x = 0.31, 0.44, 0.52, and 0.56 together with a control ZnO sample. DLOS measurements revealed the presence of five deep levels in each Mg-containing sample, having energy levels of Ec − 1.4 eV, 2.1 eV, 2.6 V, and Ev + 0.3 eV and 0.6 eV. For all Mg compositions, the activation energies of the first three states were constant with respect to the conduction band edge, whereas the latter two revealed constant activation energies with respect to the valence band edge. In contrast to the ternary materials, only three levels, at Ec − 2.1 eV, Ev + 0.3 eV, and 0.6 eV, were observed for the ZnO control sample in this systematically grown series of samples. Substantially higher concentrations of the deep levels at Ev + 0.3 eV and Ec − 2.1 eV were observed in ZnO compared to the Mg alloyed samples. Moreover, there is a general invariance of trap concentration of the Ev + 0.3 eV and 0.6 eV levels on Mg content, while at least and order of magnitude dependency of the Ec − 1.4 eV and Ec − 2.6 eV levels in Mg alloyed samples.

Estudio molecular de gluteninas de alto y bajo peso molecular en Triticum aestivum ssp. vulgare L. y su relación con la calidad panadera

El trigo blando (Triticum aestivum ssp vulgare L., AABBDD, 2n=6x=42) presenta propiedades viscoélasticas únicas debidas a la presencia en la harina de las prolaminas: gluteninas y gliadinas. Ambos tipos de proteínas forman parte de la red de gluten. Basándose en la movilidad en SDS-PAGE, las gluteninas se clasifican en dos grupos: gluteninas de alto peso molecular (HMW-GS) y gluteninas de bajo peso molecular (LMW-GS). Los genes que codifican para las HMW-GS se encuentran en tres loci del grupo 1 de cromosomas: Glu-A1, Glu-B1 y Glu-D1. Cada locus codifica para uno o dos polipéptidos o subunidades. La variación alélica de las HMW-GS es el principal determinante de de la calidad harino-panadera y ha sido ampliamente estudiado tanto a nivel de proteína como de ADN. El conocimiento de estas proteínas ha contribuido sustancialmente al progreso de los programas de mejora para la calidad del trigo. Comparadas con las HMW-GS, las LMW-GS forman una familia proteica mucho más compleja. La mayoría de los genes LMW se localizan en el grupo 1 de cromosomas en tres loci: Glu-A3, Glu-B3 y Glu-D3 que se encuentran estrechamente ligados a los loci que codifican para gliadinas. El número de copias de estos genes ha sido estimado entre 10-40 en trigo hexaploide, pero el número exacto aún se desconoce debido a la ausencia de un método eficiente para diferenciar los miembros de esta familia multigénica. La nomenclatura de los alelos LMW-GS por electroforesis convencional es complicada, y diferentes autores asignan distintos alelos a la misma variedad lo que dificulta aún más el estudio de esta compleja familia. El uso de marcadores moleculares para la discriminación de genes LMW, aunque es una tarea dificil, puede ser muy útil para los programas de mejora. El objetivo de este trabajo ha sido profundizar en la relación entre las gluteninas y la calidad panadera y desarrollar marcadores moleculares que permitan ayudar en la correcta clasificación de HMW-GS y LMW-GS. Se han obtenido dos poblaciones de líneas avanzadas F4:6 a partir de los cruzamientos entre las variedades ‘Tigre’ x ‘Gazul’ y ‘Fiel’ x ‘Taber’, seleccionándose para los análisis de calidad las líneas homogéneas para HMW-GS, LMW-GS y gliadinas. La determinación alélica de HMW-GS se llevó a cabo por SDS-PAGE, y se complementó con análisis moleculares, desarrollándose un nuevo marcador de PCR para diferenciar entre las subunidades Bx7 y Bx7*del locus Glu-B1. Resumen 2 La determinación alélica para LMW-GS se llevó a cabo mediante SDS-PAGE siguiendo distintas nomenclaturas y utilizando variedades testigo para cada alelo. El resultado no fue concluyente para el locus Glu-B3, así que se recurrió a marcadores moleculares. El ADN de los parentales y de los testigos se amplificó usando cebadores diseñados en regiones conservadas de los genes LMW y fue posteriormente analizado mediante electroforesis capilar. Los patrones de amplificación obtenidos fueron comparados entre las distintas muestras y permitieron establecer una relación con los alelos de LMW-GS. Con este método se pudo aclarar la determinación alélica de este locus para los cuatro parentales La calidad de la harina fue testada mediante porcentaje de contenido en proteína, prueba de sedimentación (SDSS) y alveógrafo de Chopin (parámetros P, L, P/L y W). Los valores fueron analizados en relación a la composición en gluteninas. Las líneas del cruzamiento ‘Fiel’ x ‘Taber’ mostraron una clara influencia del locus Glu-A3 en la variación de los valores de SDSS. Las líneas que llevaban el nuevo alelo Glu-A3b’ presentaron valores significativamente mayores que los de las líneas con el alelo Glu-A3f. En las líneas procedentes del cruzamiento ‘Tigre ’x ‘Gazul’, los loci Glu-B1 y Glu-B3 loci mostraron ambos influencia en los parámetros de calidad. Los resultados indicaron que: para los valores de SDSS y P, las líneas con las HMW-GS Bx7OE+By8 fueron significativamente mejores que las líneas con Bx17+By18; y las líneas que llevaban el alelo Glu-B3ac presentaban valores de P significativamente superiores que las líneas con el alelo Glu-B3ad y significativamente menores para los valores de L . El análisis de los valores de calidad en relación a los fragmentos LMW amplificados, reveló un efecto significativo entre dos fragmentos (2-616 y 2-636) con los valores de P. La presencia del fragmento 2-636 estaba asociada a valores de P mayores. Estos fragmentos fueron clonados y secuenciados, confirmándose que correspondían a genes del locus Glu-B3. El estudio de la secuencia reveló que la diferencia entre ambos se hallaba en algunos SNPs y en una deleción de 21 nucleótidos que en la proteína correspondería a un InDel de un heptapéptido en la región repetida de la proteína. En este trabajo, la utilización de líneas que difieren en el locus Glu-B3 ha permitido el análisis de la influencia de este locus (el peor caracterizado hasta la fecha) en la calidad panadera. Además, se ha validado el uso de marcadores moleculares en la determinación alélica de las LMW-GS y su relación con la calidad panadera. Summary 3 Bread wheat (Triticum aestivum ssp vulgare L., AABBDD, 2n=6x=42) flour has unique dough viscoelastic properties conferred by prolamins: glutenins and gliadins. Both types of proteins are cross-linked to form gluten polymers. On the basis of their mobility in SDS-PAGE, glutenins can be classified in two groups: high molecular weight glutenins (HMW-GS) and low molecular weight glutenins (LMW-GS). Genes encoding HMW-GS are located on group 1 chromosomes in three loci: Glu-A1, Glu-B1 and Glu-D1, each one encoding two polypeptides, named subunits. Allelic variation of HMW-GS is the most important determinant for bread making quality, and has been exhaustively studied at protein and DNA level. The knowledge of these proteins has substantially contributed to genetic improvement of bread quality in breeding programs. Compared to HMW-GS, LMW-GS are a much more complex family. Most genes encoded LMW-GS are located on group 1 chromosomes. Glu-A3, Glu-B3 and Glu-D3 loci are closely linked to the gliadin loci. The total gene copy number has been estimated to vary from 10–40 in hexaploid wheat. However, the exact copy number of LMW-GS genes is still unknown, mostly due to lack of efficient methods to distinguish members of this multigene family. Nomenclature of LMW-GS alleles is also unclear, and different authors can assign different alleles to the same variety increasing confusion in the study of this complex family. The use of molecular markers for the discrimination of LMW-GS genes might be very useful in breeding programs, but their wide application is not easy. The objective of this work is to gain insight into the relationship between glutenins and bread quality, and the developing of molecular markers that help in the allele classification of HMW-GS and LMW-GS. Two populations of advanced lines F4:6 were obtained from the cross ‘Tigre’ x ‘Gazul’ and ‘Fiel’ x ‘Taber’. Lines homogeneous for HMW-GS, LMW-GS and gliadins pattern were selected for quality analysis. The allele classification of HMW-GS was performed by SDS-PAGE, and then complemented by PCR analysis. A new PCR marker was developed to undoubtedly differentiate between two similar subunits from Glu-B1 locus, Bx7 and Bx7*. The allele classification of LMW-GS was initially performed by SDS-PAGE following different established nomenclatures and using standard varieties. The results were not completely concluding for Glu-B3 locus, so a molecular marker system was applied. DNA from parental lines and standard varieties was amplified using primers designed in conserved domains of LMW genes and analyzed by capillary electrophoresis. The pattern of amplification products obtained was compared among samples and related to the protein allele classification. It was possible to establish a correspondence between specific amplification products and almost all LMW alleles analyzed. With this method, the allele classification of the four parental lines was clarified. Flour quality of F4:6 advanced lines were tested by protein content, sedimentation test (SDSS) and alveograph (P, L, P/L and W). The values were analyzed in relation to the lines prolamin composition. In the ‘Fiel’ x ‘Taber’ population, Glu-A3 locus showed an influence in SDSS values. Lines carrying new allele Glu-A3b’, presented a significantly higher SDSS value than lines with Glu-A3f allele. In the ‘Tigre ’x ‘Gazul’ population, the Glu-B1 and Glu-B3 loci also showed an effect in quality parameters, in SDSS, and P and L values. Results indicated that: for SDSS and P, lines with Bx7OE+By8 were significantly better than lines with Bx17+By18; lines carrying Glu-B3ac allele had a significantly higher P values than Glu-B3ad allele values. lines with and lower L The analysis of quality parameters and amplified LMW fragments revealed a significant influence of two peaks (2-616 y 2-636) in P values. The presence of 2-636 peak gave higher P values than 2-616. These fragments had been cloned and sequenced and identified as Glu-B3 genes. The sequence analysis revealed that the molecular difference between them was some SNPs and a small deletion of 21 nucleotides that in the protein would produce an InDel of a heptapeptide in the repetitive region. In this work, the analysis of two crosses with differences in Glu-3 composition has made possible to study the influence of LMG-GS in quality parameters. Specifically, the influence of Glu-B3, the most interesting and less studied loci has been possible. The results have shown that Glu-B3 allele composition influences the alveograph parameter P (tenacity). The existence of different molecular variants of Glu-B3 alleles have been assessed by using a molecular marker method. This work supports the use of molecular approaches in the study of the very complex LMW-GS family, and validates their application in the analysis of advanced recombinant lines for quality studies.

Jupiter power generation with electrodynamic tethers at constant orbital energy

An electrodynamic tether system for power generation at Jupiter is presented that allows extracting energy from Jupiter's corotating plasmasphere while leaving the system orbital energy unaltered to first order. The spacecraft is placed in a polar orbit with the tether spinning in the orbital plane so that the resulting Lorentz force, neglecting Jupiter's magnetic dipole tilt, is orthogonal to the instantaneous velocity vector and orbital radius, hence affecting orbital inclination rather than orbital energy. In addition, the electrodynamic tether subsystem, which consists of two radial tether arms deployed from the main central spacecraft, is designed in such a way as to extract maximum power while keeping the resulting Lorentz torque constantly null. The power-generation performance of the system and the effect on the orbit inclination is evaluated analytically for different orbital conditions and verified numerically. Finally, a thruster-based inclination-compensation maneuver at apoapsis is added, resulting in an efficient scheme to extract energy from the plasmasphere of the planet with minimum propellant consumption and no inclination change. A tradeoff analysis is conducted showing that, depending on tether size and orbit characteristics, the system performance can be considerably higher than conventional power-generation methods.

Structuring research methods and data with the research object model: genomics workflows as a case study

Background: One of the main challenges for biomedical research lies in the computer-assisted integrative study of large and increasingly complex combinations of data in order to understand molecular mechanisms. The preservation of the materials and methods of such computational experiments with clear annotations is essential for understanding an experiment, and this is increasingly recognized in the bioinformatics community. Our assumption is that offering means of digital, structured aggregation and annotation of the objects of an experiment will provide necessary meta-data for a scientist to understand and recreate the results of an experiment. To support this we explored a model for the semantic description of a workflow-centric Research Object (RO), where an RO is defined as a resource that aggregates other resources, e.g., datasets, software, spreadsheets, text, etc. We applied this model to a case study where we analysed human metabolite variation by workflows. Results: We present the application of the workflow-centric RO model for our bioinformatics case study. Three workflows were produced following recently defined Best Practices for workflow design. By modelling the experiment as an RO, we were able to automatically query the experiment and answer questions such as “which particular data was input to a particular workflow to test a particular hypothesis?”, and “which particular conclusions were drawn from a particular workflow?”. Conclusions: Applying a workflow-centric RO model to aggregate and annotate the resources used in a bioinformatics experiment, allowed us to retrieve the conclusions of the experiment in the context of the driving hypothesis, the executed workflows and their input data. The RO model is an extendable reference model that can be used by other systems as well.

Vibrational population relaxation of carbon monoxide in the heme pocket of photolyzed carbonmonoxy myoglobin: Comparison of time-resolved mid-IR absorbance experiments and molecular dynamics simulations

The vibrational energy relaxation of carbon monoxide in the heme pocket of sperm whale myoglobin was studied by using molecular dynamics simulation and normal mode analysis methods. Molecular dynamics trajectories of solvated myoglobin were run at 300 K for both the δ- and ɛ-tautomers of the distal His-64. Vibrational population relaxation times of 335 ± 115 ps for the δ-tautomer and 640 ± 185 ps for the ɛ-tautomer were estimated by using the Landau–Teller model. Normal mode analysis was used to identify those protein residues that act as the primary “doorway” modes in the vibrational relaxation of the oscillator. Although the CO relaxation rates in both the ɛ- and δ-tautomers are similar in magnitude, the simulations predict that the vibrational relaxation of the CO is faster in the δ-tautomer with the distal His playing an important role in the energy relaxation mechanism. Time-resolved mid-IR absorbance measurements were performed on photolyzed carbonmonoxy hemoglobin (Hb13CO). From these measurements, a T1 time of 600 ± 150 ps was determined. The simulation and experimental estimates are compared and discussed.

Molecular cloning, sequence, expression, and processing of the interleukin 16 precursor

Interleukin 16 (IL-16) has been shown to function as chemoattractant factor, as a modulator of T-cell activation, and as an inhibitor of immunodeficiency virus replication. The recent identification of inconsistencies in published IL-16 cDNA nucleotide sequences led to the proposal that IL-16 is synthesized in the form of a large precursor protein (pro-IL-16). To identify the true transcriptional start of the IL-16 mRNA rapid amplification of cDNA ends methods were applied. The complete pro-IL-16 cDNA was subsequently molecularly cloned, sequenced, and expressed in COS-7 cells. We report here that pro-IL-16 is most likely synthesized as a 67-kDa protein and is encoded from a major 2.6-kb transcript. Recombinant pro-IL-16 polypeptides are specifically cleaved in lysates of CD8(+) cells, suggesting that the naturally secreted bioactive form of IL-16 is smaller than the originally published 130 amino acids fragment. Moreover, in contrast to other interleukins such as IL-15, IL-16 mRNA expression is almost exclusively limited to lymphatic tissues underlining the potential of IL-16 as an immune regulatory molecule.

Simultaneous A8344G heteroplasmy and mitochondrial DNA copy number quantification in Myoclonus Epilepsy and Ragged-Red Fibers (MERRF) syndrome by a multiplex Molecular Beacon based real-time fluorescence PCR

The association of a particular mitochondrial DNA (mtDNA) mutation with different clinical phenotypes is a well-known feature of mitochondrial diseases. A simple genotype–phenotype correlation has not been found between mutation load and disease expression. Tissue and intercellular mosaicism as well as mtDNA copy number are thought to be responsible for the different clinical phenotypes. As disease expression of mitochondrial tRNA mutations is mostly in postmitotic tissues, studies to elucidate disease mechanisms need to be performed on patient material. Heteroplasmy quantitation and copy number estimation using small patient biopsy samples has not been reported before, mainly due to technical restrictions. In order to resolve this problem, we have developed a robust assay that utilizes Molecular Beacons to accurately quantify heteroplasmy levels and determine mtDNA copy number in small samples carrying the A8344G tRNALys mutation. It provides the methodological basis to investigate the role of heteroplasmy and mtDNA copy number in determining the clinical phenotypes.

Alternate protein frameworks for molecular recognition.

In an effort to determine whether proteins with structures other than the immunoglobulin fold can be used to mimic the ligand binding properties of antibodies, we generated a library from the four-helix bundle protein cytochrome b562 in which the two loops were randomized. Panning of this library against the bovine serum albumin (BSA) conjugate of N-methyl-p-nitrobenzylamine derivative 1 by phage display methods yielded cytochromes in which residues Trp-20, Arg-21, and Ser-22 in loop A and Arg-83 and Trp-84 in loop B were conserved. The individual mutants, which fold into native-like structure, bind selectively to the BSA-1 conjugate with micromolar dissociation constants (Kd), in comparison to a monoclonal antibody that binds selectively to 1 with a Kd of 290 nM. These and other antibody-like receptors may prove useful as therapeutic agents or as reagents for both intra- and extracellular studies.

Stabilization of APIs and nutraceuticals through cocrystallization and molecular confinement

Cocrystallization of the molecule of interest could be a smart and dainty way to tune solubility properties of solid phases leaving the molecule chemically unchanged, hence it is widely investigated by companies and by solid state scientists. Despite of this extremely high interest towards cocrystallization no particular emphasis has been paid to using it as a means to stabilize liquid molecules. In this work we define a benchmark of relevant molecules for human health that have been combined with suitable partners according to crystal engineering methods in order to obtain cocrystals. Solubility properties in different solvents of cocrystals new solid phases have been tested and compared to the properties of the drugs. A further approach to deal with volatile compounds is molecular confinement inside molecular scaffold. Nowadays metal organic frameworks (MOFs) are studied in many fields ranging from catalysis to trapping or storage of gases, such as hydrogen, methane, CO2 thanks to their extremely high porosity. Our goal is to confine liquid guests of biological relevance inside MOF pores, monitoring via X-ray diffraction, spectroscopy and thermal analysis the stabilization of the molecule of interest inside the cavities.

QSAR e dinâmica molecular no estudo de sistemas biomoleculares: predição da atividade biológica de antagonistas do receptor sigma-1 e simulações de bicamadas lipídicas

Diferentes abordagens teóricas têm sido utilizadas em estudos de sistemas biomoleculares com o objetivo de contribuir com o tratamento de diversas doenças. Para a dor neuropática, por exemplo, o estudo de compostos que interagem com o receptor sigma-1 (Sig-1R) pode elucidar os principais fatores associados à atividade biológica dos mesmos. Nesse propósito, estudos de Relações Quantitativas Estrutura-Atividade (QSAR) utilizando os métodos de regressão por Mínimos Quadrados Parciais (PLS) e Rede Neural Artificial (ANN) foram aplicados a 64 antagonistas do Sig-1R pertencentes à classe de 1-arilpirazóis. Modelos PLS e ANN foram utilizados com o objetivo de descrever comportamentos lineares e não lineares, respectivamente, entre um conjunto de descritores e a atividade biológica dos compostos selecionados. O modelo PLS foi obtido com 51 compostos no conjunto treinamento e 13 compostos no conjunto teste (r² = 0,768, q² = 0,684 e r²teste = 0,785). Testes de leave-N-out, randomização da atividade biológica e detecção de outliers confirmaram a robustez e estabilidade dos modelos e mostraram que os mesmos não foram obtidos por correlações ao acaso. Modelos também foram gerados a partir da Rede Neural Artificial Perceptron de Multicamadas (MLP-ANN), sendo que a arquitetura 6-12-1, treinada com as funções de transferência tansig-tansig, apresentou a melhor resposta para a predição da atividade biológica dos compostos (r²treinamento = 0,891, r²validação = 0,852 e r²teste = 0,793). Outra abordagem foi utilizada para simular o ambiente de membranas sinápticas utilizando bicamadas lipídicas compostas por POPC, DOPE, POPS e colesterol. Os estudos de dinâmica molecular desenvolvidos mostraram que altas concentrações de colesterol induzem redução da área por lipídeo e difusão lateral e aumento na espessura da membrana e nos valores de parâmetro de ordem causados pelo ordenamento das cadeias acil dos fosfolipídeos. As bicamadas lipídicas obtidas podem ser usadas para simular interações entre lipídeos e pequenas moléculas ou proteínas contribuindo para as pesquisas associadas a doenças como Alzheimer e Parkinson. As abordagens usadas nessa tese são essenciais para o desenvolvimento de novas pesquisas em Química Medicinal Computacional.