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Improved Resolution (improved + resolution)
Selected AbstractsA modified CRISIS-HSQC for band-selective IMPRESSMAGNETIC RESONANCE IN CHEMISTRY, Issue 2 2005Scott A. Bradley Abstract CRISIS (Compensation of Refocusing Inefficiency with Synchronized Inversion Sweep) is a powerful technique for obtaining multiplicity-edited HSQC spectra without compromising sensitivity. However, the stringent requirement for the duration of the CRISIS waveforms makes them unsuitable for other functions, such as band selection or IMPRESS (IMProved REsolution using Symmetrically Shifted pulses). We report here a modified CRISIS-gHSQC pulse sequence employing time-reversed 13C ,/2 EBURP-2 pulses. This IC-bs-gHSQC (IMPRESS-CRISIS-bs-gHSQC) sequence was found to be equally useful for acquiring multiplicity-edited, band-selective spectra individually or in tandem with IMPRESS. Remarkably, the latter provides multiple spectra in significantly less time and is the preferred approach when several crowded regions need to be assigned unambiguously. The use of adiabatic sweeps and the CRISIS pulses enable IC-bs-gHSQC to give better sensitivity than the original IMPRESS sequence for band-selective spectra. Copyright © 2005 John Wiley & Sons, Ltd. [source] Orthogonal separations of nicotine and nicotine-related alkaloids by various capillary electrophoretic modesELECTROPHORESIS, Issue 9 2004Alex Marsh Abstract The migration behaviour of nicotine and related tobacco alkaloids was investigated using three different capillary electrophoretic (CE) modes. Novel separations were achieved both using microemulsion electrokinetic chromatography (MEEKC) and nonaqueous CE (NACE). Improved resolution compared to previous studies was obtained using free-solution CE (FSCE). Each technique resulted in different, orthogonal separation selectivity. The suitability of each method for application to the analysis of nicotine lozenges is discussed. The FSCE method was applied to the analysis of nicotine lozenges due to its compatibility with an established lozenge extraction solvent. The method used gave good injection precision and linearity. Good agreement of CE and high-performance liquid chromatography (HPLC) results was obtained. The CE method is therefore considered suitable for the quantitative determination of nicotine in nicotine lozenges. [source] Improved resolution with microchip-based enhanced field inversion electrophoresisELECTROPHORESIS, Issue 11 2003Christopher J. Backhouse Abstract We present an improvement of the field inversion electrophoresis (FIE) method in which the passage of sample such as DNA back and forth within a short length of a microchannel can provide a similar resolution to that of a significantly longer microchannel. In constant field FIE the application of an alternating potential (e.g., +/, V) over short periods of time (e.g., several Hz) can provide enhanced separations of DNA fragments. In contrast, the present method consists of a series of separations, each of much longer duration, under high and low fields in such a way that the resolution is enhanced. This method is readily modeled and allows improved resolution to be obtained from extremely short microchannels (e.g., 8 mm) while requiring relatively low applied voltages (e.g., less than 600 V). An additional advantage is that this method can allow for the same equipment to be used in a rapid, low-resolution mode or in a slower, high-resolution mode through what might be referred to as an automated "zoom" capability. We believe that this method may facilitate the integration of microfluidic devices and microelectronic devices by allowing these devices to be of a similar small scale (< 1 cm). [source] Improved resolution in two-dimensional 1H NMR spectra of peptides by band-selective, homonuclear decoupling during both the evolution and acquisition periods: application to characterization of the binding of peptides by heparinMAGNETIC RESONANCE IN CHEMISTRY, Issue 8 2006Jing Wang Abstract Two-dimensional 1H NMR experiments that achieve band-selective, homonuclear decoupling in both the indirectly detected F1 and directly detected F2 dimensions were used to assign the highly overlapped 1H NMR spectrum of the peptide Ac-SRGKARVRAKVKDQTK-NH2, both free in solution and bound to heparin. Band-selective, homonuclear decoupling during the evolution period was achieved using a double pulsed field gradient spin-echo (DPFGSE) with semi-selective shaped pulses; band-selective, homonuclear decoupling during the acquisition period was achieved by time-shared semi-selective shaped pulse decoupling. Regular TOCSY, ROESY and NOESY spectra and TOCSY, ROESY and NOESY spectra measured with band-selective, homonuclear decoupling in the evolution (F1) dimension (BASHD-TOCSY, ROESY and NOESY spectra) and with band-selective, homonuclear decoupling in both the F1 and F2 dimensions (D-(or Double)-BASHD-TOCSY, ROESY and NOESY spectra) are reported and compared for the peptide and its heparin complex. Complete assignment of the 1H-NMR spectra of the free and heparin-complexed peptide was achieved with the high resolution of the D-BASHD-TOCSY, ROESY and NOESY spectra. Characterization of the heparin-complexed peptide is of interest because of the ability of the peptide to neutralize the anticoagulant activity of heparin. Copyright © 2006 John Wiley & Sons, Ltd. [source] Cyclodextrin-based nonaqueous electrokinetic chromatography with UV and mass spectrometric detection: Application to the impurity profiling of amiodarone,ELECTROPHORESIS, Issue 17 2008Roelof Mol Abstract The potential of nonaqueous electrokinetic chromatography (NAEKC) using cyclodextrins (CD) for the analysis of basic drugs and related compounds was evaluated. Both UV absorbance and mass spectrometric (MS) detection were employed. Addition of neutral CD to the NA background electrolyte did not significantly enhance the separation of a test mixture of basic drugs, and no change in selectivity was observed. In contrast, anionic single-isomer-sulfated CD strongly added to the selectivity of the NAEKC system inducing an improved resolution among the test compounds and increasing the migration time window. The applicability of the NAEKC system using anionic CD is demonstrated by the profiling of a sample of the drug amiodarone that had been stored for 1,year at room temperature. Amiodarone is poorly soluble in water. NAEKC-UV analysis indicated the presence of at least seven impurities in the amiodarone sample. In order to identify these compounds, the NAEKC system was coupled directly to electrospray ionization (ESI) ion-trap MS. The total of detected impurities increased to 12 due to the added sensitivity and selectivity of MS detection. Based on the acquired MS/MS data, three sample constituents could be identified as ,known' impurities (British Pharmacopoeia), whereas for three unknown impurities molecular structures could be proposed. Estimated limits of detection for amiodarone using the NAEKC method were 1,,g/mL with UV detection and 15,ng/mL with ESI-MS detection (full-scan). Based on relative responses, the impurity content of the stored drug substance was estimated to be 0.33 and 0.47% using NAEKC-UV and NAEKC-ESI-MS, respectively. [source] A simple polyacrylamide gel electrophoresis procedure for separation of polyamidoamine dendrimersELECTROPHORESIS, Issue 16 2003Ajit Sharma Abstract A simple, inexpensive, and rapid electrophoresis technique was developed for use as a routine tool for evaluating purity of polyamidoamine (PAMAM) dendrimers. A variety of factors influencing migration of generations 0,7 dendrimers on nongradient polyacrylamide gels were evaluated. The low generation dendrimers were found to be very sensitive to diffusion during or after electrophoresis. The proposed method incorporates steps that minimize diffusion, in order to obtain improved resolution and sensitivity, especially for the lower-molecular-weight dendrimers. This was accomplished by inclusion of a dendrimer fixation step with glutaraldehyde and performing the electrophoresis separation, fixation, staining, and destaining at 4°C. PAMAM dendrimer separation was studied under basic and acidic conditions. Electrophoresis under acidic conditions gave increased resolution and sensitivity over separation at alkaline pH. Oligomers and trailing generations could be clearly separated and visualized under these conditions. The smallest PAMAM dendrimer, generation 0, was visible at 1.5 ,g under the optimized acidic conditions. With slight modifications, this technique should be applicable to separation of other water-soluble dendrimers. [source] Improved resolution with microchip-based enhanced field inversion electrophoresisELECTROPHORESIS, Issue 11 2003Christopher J. Backhouse Abstract We present an improvement of the field inversion electrophoresis (FIE) method in which the passage of sample such as DNA back and forth within a short length of a microchannel can provide a similar resolution to that of a significantly longer microchannel. In constant field FIE the application of an alternating potential (e.g., +/, V) over short periods of time (e.g., several Hz) can provide enhanced separations of DNA fragments. In contrast, the present method consists of a series of separations, each of much longer duration, under high and low fields in such a way that the resolution is enhanced. This method is readily modeled and allows improved resolution to be obtained from extremely short microchannels (e.g., 8 mm) while requiring relatively low applied voltages (e.g., less than 600 V). An additional advantage is that this method can allow for the same equipment to be used in a rapid, low-resolution mode or in a slower, high-resolution mode through what might be referred to as an automated "zoom" capability. We believe that this method may facilitate the integration of microfluidic devices and microelectronic devices by allowing these devices to be of a similar small scale (< 1 cm). [source] Non-uniqueness with refraction inversion , the Mt Bulga shear zoneGEOPHYSICAL PROSPECTING, Issue 4 2010Derecke Palmer ABSTRACT The tau-p inversion algorithm is widely employed to generate starting models with many computer programs that implement refraction tomography. However, this algorithm can frequently fail to detect even major lateral variations in seismic velocities, such as a 50 m wide shear zone, which is the subject of this study. By contrast, the shear zone is successfully defined with the inversion algorithms of the generalized reciprocal method. The shear zone is confirmed with a 2D analysis of the head wave amplitudes, a spectral analysis of the refraction convolution section and with numerous closely spaced orthogonal seismic profiles recorded for a later 3D refraction investigation. Further improvements in resolution, which facilitate the recognition of additional zones with moderate reductions in seismic velocity, are achieved with a novel application of the Hilbert transform to the refractor velocity analysis algorithm. However, the improved resolution also requires the use of a lower average vertical seismic velocity, which accommodates a velocity reversal in the weathering. The lower seismic velocity is derived with the generalized reciprocal method, whereas most refraction tomography programs assume vertical velocity gradients as the default. Although all of the tomograms are consistent with the traveltime data, the resolution of each tomogram is comparable only with that of the starting model. Therefore, it is essential to employ inversion algorithms that can generate detailed starting models, where detailed lateral resolution is the objective. Non-uniqueness can often be readily resolved with head wave amplitudes, attribute processing of the refraction convolution section and additional seismic traverses, prior to the acquisition of any borehole data. It is concluded that, unless specific measures are taken to address non-uniqueness, the production of a single refraction tomogram that fits the traveltime data to sufficient accuracy does not necessarily demonstrate that the result is either correct, or even the most probable. [source] Controlling diffusion of 3He by buffer gases: A structural contrast agent in lung MRIJOURNAL OF MAGNETIC RESONANCE IMAGING, Issue 6 2006Rodolfo H. Acosta PhD Abstract Purpose To study the influence of admixing inert buffer gases to laser-polarized 3He in terms of resulting diffusion coefficients and the consequences for image contrast and resolution. Materials and Methods The diffusion coefficient of 3He was altered by admixing buffer gases of various molecular weights (4He, N2, and SF6). The influence of the pulse sequence and the diffusion coefficient on the appearance of MRI of (laser-polarized) gases was analyzed by comparison of basic theoretical concepts with demonstrative experiments. Results Excellent agreement between theoretical description and observed signal in simple gradient echoes was observed. A maximum signal gain can be predicted and was experimentally validated. Images acquired under such conditions revealed improved resolution. The nature and concentration of the admixed gas defines a structural threshold for the observed apparent diffusion coefficient (ADC) as demonstrated with diffusion-weighted MRI on a pig's lung flooded with suitable gas mixtures. Conclusion A novel procedure is proposed to control the diffusion coefficient of gases in MRI by admixture of inert buffer gases. Their molecular mass and concentration enter as additional parameters into the equations that describe structural contrast. This allows for setting a structural threshold up to which structures contribute to the image. For MRI of the lung this enables images of very small structural elements (alveoli) only, or in the other extreme, all airways can be displayed with minimal signal loss due to diffusion. J. Magn. Reson. Imaging 2006. © 2006 Wiley-Liss, Inc. [source] High-resolution H/D exchange studies on the HET-s218,295 prion proteinJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 5 2005Alexis Nazabal Abstract In a search for improved resolution of hydrogen/deuterium (H/D) exchange experiments analyzed by mass spectrometry (HXMS), we evaluated two methodologies for a detailed structural study of solvent accessibility in the case of the HET-s218,295 prion protein. For the first approach, after incubation in the deuterated solvent, aggregated HET-s218,295 was digested with pepsin and the generated peptides were analyzed by nanospray mass spectrometry in an ion trap, with and without collision-induced dissociation (CID). We compared deuterium incorporation in peptides as determined on peptide pseudomolecular ions and on b and y fragments produced by longer peptides under CID conditions. For both b and y fragment ions, an extensive H/D scrambling phenomenon was observed, in contrast with previous studies comparing CID-MS experiments and 1H NMR data. Thus, the spatial resolution of HXMS experiments could not be improved by means of MS/MS data generated by an ion trap mass spectrometer. In a second approach, the incorporation of deuterium was analyzed by MS for 76 peptides of the HET-s218,289 peptide mass fingerprint, and the use of shared boundaries among peptic peptides allowed us to determine deuteration levels of small regions ranging from one to four amino acids. This methodology led to evidence of highly protected regions along the HET-s218,295 sequence. Copyright © 2005 John Wiley & Sons, Ltd. [source] Application of 31P NMR spectroscopy and chemical derivatization for metabolite profiling of lipophilic compounds in human serumMAGNETIC RESONANCE IN CHEMISTRY, Issue S1 2009M. Aruni DeSilva Abstract New methods for obtaining metabolic fingerprints of biological samples with improved resolution and sensitivity are highly sought for early disease detection, studies of human health and pathophysiology, and for better understanding systems biology. Considering the complexity of biological samples, interest in biochemical class selection through the use of chemoselective probes for improved resolution and quantitation is increasing. Considering the role of lipids in the pathogenesis of a number of diseases, in this study fingerprinting of lipid metabolites was achieved by 31P labeling using the derivatizing agent 2-chloro-4,4,5,5-tetramethyldioxaphospholane. Lipids containing hydroxyl, aldehyde and carboxyl groups were selectively tagged with 31P and then detected with good resolution using 31P NMR by exploiting the 100% natural abundance and wide chemical shift range of 31P. After standardizing the reaction conditions using representative compounds, the derivatization approach was used to profile lipids in human serum. The results show that the 31P derivatization approach is simple, reproducible and highly quantitative, and has the potential to profile a number of important lipids in complex biological samples. Copyright © 2009 John Wiley & Sons, Ltd. [source] Methyl TROSY: explanation and experimental verificationMAGNETIC RESONANCE IN CHEMISTRY, Issue 10 2003Jason E. Ollerenshaw Abstract In TROSY experiments, relaxation interference effects are exploited to produce spectra with improved resolution and signal-to-noise. Such experiments cannot be explained using the standard product operator formalism, but must instead be analyzed at the level of individual density matrix elements. Herein we illustrate this point using an example from our recent work on a TROSY 1H,13C correlation experiment for methyl groups in large proteins. Methyl groups are useful spectroscopic probes of protein structure and dynamics because they are found throughout the critical core region of a folded protein and their resonances are intense and well dispersed. Additionally, it is relatively easy to produce highly deuterated protein samples that are 1H,13C labeled at selected methyl positions, facilitating studies of high molecular weight systems. Methyl groups are relaxed by a network of 1H,1H and 1H,13C dipolar interactions, and in the macromolecular limit the destructive interference of these interactions leads to unusually slow relaxation for certain density matrix elements. It is this slow relaxation that forms the basis for TROSY experiments. We present a detailed analysis of evolution and relaxation during HSQC and HMQC pulse schemes for the case of a 13C1H3 spin system attached to a macromolecule. We show that the HMQC sequence is already optimal with respect to the TROSY effect, offering a significant sensitivity enhancement over HSQC at any spectrometer field strength. The gain in sensitivity is established experimentally using samples of two large proteins, malate synthase G (81.4 kDa) and ClpP protease (305 kDa), both highly deuterated and selectively 1H,13C-labeled at isoleucine , methyl positions. Copyright © 2003 John Wiley & Sons, Ltd. [source] Use of terminal restriction fragment length polymorphism (T-RFLP) for identification of phytoplasmas in plantsPLANT PATHOLOGY, Issue 3 2007J. Hodgetts A terminal restriction fragment analysis (T-RFLP) technique was developed for the simple and rapid detection and diagnosis of phytoplasmas in plants. The selected primers amplified part of the 23S rRNA gene to provide improved resolution between the taxonomic groups compared to conventional restriction enzyme analysis of the 16S rRNA. Using the restriction enzymes Bsh12361 and MseI on the PCR products, and fragment analysis in the range 68,640 bp, the technique was tested on 37 isolates from 10 of the 16Sr groups. Distinct and unambiguous T-RFLP profiles were produced for nine of the 10 taxonomic groups, such that almost all isolates within a group shared the same profile and could be distinguished from isolates in other groups. The technique also identified the presence of mixtures of phytoplasmas from different groups in samples. Furthermore, the primers were devised to amplify a terminal restriction fragment (TRF) product of a specific defined size (461 bp) from the host plant chloroplast DNA, so that there was a built-in internal control in the procedure to show that the absence of a phytoplasma peak in a sample was the result of no detectable phytoplasma being present, not the result of PCR inhibition. This method offers the possibility of simultaneously detecting and providing a taxonomic grouping for phytoplasmas in test samples using a single PCR reaction. [source] Optimization of paper bridge loading for 2-DE analysis in the basic pH region: Application to the mitochondrial subproteomePROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 21 2006Lesley A. Kane Abstract Separation of basic proteins with 2-DE presents technical challenges involving protein precipitation, load limitations, and streaking. Cardiac mitochondria are enriched in basic proteins and difficult to resolve by 2-DE. We investigated two methods, cup and paper bridge, for sample loading of this subproteome into the basic range (pH,6,11) gels. Paper bridge loading consistently produced improved resolution of both analytical and preparative protein loads. A unique benefit of this technique is that proteins retained in the paper bridge after loading basic gels can be reloaded onto lower pH gradients (pH,4,7), allowing valued samples to be analyzed on multiple pH ranges. [source] Recent developments in gravity-wave effects in climate models and the global distribution of gravity-wave momentum flux from observations and modelsTHE QUARTERLY JOURNAL OF THE ROYAL METEOROLOGICAL SOCIETY, Issue 650 2010M. J. Alexander Abstract Recent observational and theoretical studies of the global properties of small-scale atmospheric gravity waves have highlighted the global effects of these waves on the circulation from the surface to the middle atmosphere. The effects of gravity waves on the large-scale circulation have long been treated via parametrizations in both climate and weather-forecasting applications. In these parametrizations, key parameters describe the global distributions of gravity-wave momentum flux, wavelengths and frequencies. Until recently, global observations could not define the required parameters because the waves are small in scale and intermittent in occurrence. Recent satellite and other global datasets with improved resolution, along with innovative analysis methods, are now providing constraints for the parametrizations that can improve the treatment of these waves in climate-prediction models. Research using very-high-resolution global models has also recently demonstrated the capability to resolve gravity waves and their circulation effects, and when tested against observations these models show some very realistic properties. Here we review recent studies on gravity-wave effects in stratosphere-resolving climate models, recent observations and analysis methods that reveal global patterns in gravity-wave momentum fluxes and results of very-high-resolution model studies, and we outline some future research requirements to improve the treatment of these waves in climate simulations. Copyright © 2010 Royal Meteorological Society and Crown in the right of Canada [source] Inclusion of weak high-resolution X-ray data for improvement of a group II intron structureACTA CRYSTALLOGRAPHICA SECTION D, Issue 9 2010Jimin Wang It is common to report the resolution of a macromolecular structure with the highest resolution shell having an averaged I/,(I) , 2. Data beyond the resolution thus defined are weak and often poorly measured. The exclusion of these weak data may improve the apparent statistics and also leads to claims of lower resolutions that give some leniency in the acceptable quality of refined models. However, the inclusion of these data can provide additional strong constraints on atomic models during structure refinement and thus help to correct errors in the original models, as has recently been demonstrated for a protein structure. Here, an improved group II intron structure is reported arising from the inclusion of these data, which helped to define more accurate solvent models for density modification during experimental phasing steps. With the improved resolution and accuracy of the experimental phases, extensive revisions were made to the original models such that the correct tertiary interactions of the group II intron that are essential for understanding the chemistry of this ribozyme could be described. [source] Systematic study on crystal-contact engineering of diphthine synthase: influence of mutations at crystal-packing regions on X-ray diffraction qualityACTA CRYSTALLOGRAPHICA SECTION D, Issue 10 2008Hisashi Mizutani It is well known that protein crystallizability can be influenced by site-directed mutagenesis of residues on the molecular surface of proteins, indicating that the intermolecular interactions in crystal-packing regions may play a crucial role in the structural regularity at atomic resolution of protein crystals. Here, a systematic examination was made of the improvement in the diffraction resolution of protein crystals on introducing a single mutation of a crystal-packing residue in order to provide more favourable packing interactions, using diphthine synthase from Pyrococcus horikoshii OT3 as a model system. All of a total of 21 designed mutants at 13 different crystal-packing residues yielded almost isomorphous crystals from the same crystallization conditions as those used for the wild-type crystals, which diffracted X-rays to 2.1,Å resolution. Of the 21 mutants, eight provided crystals with an improved resolution of 1.8,Å or better. Thus, it has been clarified that crystal quality can be improved by introducing a suitable single mutation of a crystal-packing residue. In the improved crystals, more intimate crystal-packing interactions than those in the wild-type crystal are observed. Notably, the mutants K49R and T146R yielded crystals with outstandingly improved resolutions of 1.5 and 1.6,Å, respectively, in which a large-scale rearrangement of packing interactions was unexpectedly observed despite the retention of the same isomorphous crystal form. In contrast, the mutants that provided results that were in good agreement with the designed putative structures tended to achieve only moderate improvements in resolution of up to 1.75,Å. These results suggest a difficulty in the rational prediction of highly effective mutations in crystal engineering. [source] | ||||||||||||||||