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PubMed2001

Infrared and EPR Spectroscopic Studies of 2-C(2)H(2)F and 1-C(2)H(2)F Radicals Isolated in Solid Argon.

2-fluorovinyl radicals were generated in solid argon by solid-state chemical reactions of mobile F atoms with acetylene and its deuterated analogues. Highly resolved EPR spectra of the stabilized radicals CHF&dbond;(*)CH, CDF&dbond;(*)CD, CHF&dbond;(*)CD, and CDF&dbond;(*)CH were obtained for the first time. The observed spectra were assigned to cis-2-fluorovinyl radical based on excellent agreement between the measured (a(F) = 6.50, a(betaH) = 3.86, a(alphaH) = 0.25 mT) hyperfine constants and those calculated using density functional (B3LYP) theory. Analogous experiments carried out using infrared spectroscopy yielded a complete assignment of the vibrational frequencies. An unusual reversible photochemical conversion is observed in which cis-2-fluorovinyl radicals can be partially converted to 1-fluorovinyl radicals by pulsed laser photolysis at 532 nm. Photolysis at 355 nm converts 1-fluorovinyl back to cis-2-fluorovinyl. High-resolution EPR and infrared spectra of 1-fluorovinyl were obtained for the first time. The measured hyperfine constants (a(F) = 13.71, a(H1) = 4.21, a(H2) = 1.16 mT) are in good agreement with calculated values. Copyright 2001 Academic Press.

Journal Article↗

PubMed2003

How the confocal laser scanning microscope entered biological research.

A history of the early development of the confocal laser scanning microscope in the MRC Laboratory of Molecular Biology in Cambridge is presented. The rapid uptake of this technology is explained by the wide use of fluorescence in the 80s. The key innovations were the scanning of the light beam over the specimen rather than vice-versa and a high magnification at the level of the detector, allowing the use of a macroscopic iris. These were followed by an achromatic all-reflective relay system, a non-confocal transmission detector and novel software for control and basic image processing. This design was commercialized successfully and has been produced and developed over 17 years, surviving challenges from alternative technologies, including solid-state scanning systems. Lessons are pointed out from the unusual nature of the original funding and research environment. Attention is drawn to the slow adoption of the instrument in diagnostic medicine, despite promising applications.

Animals↗

PubMed2005

Morphological behavior of lipid bilayers induced by melittin near the phase transition temperature.

Morphological changes of DMPC, DLPC, and DPPC bilayers containing melittin (lecithin/melittin molar ratio of 10:1) around the gel-to-liquid crystalline phase transition temperatures (Tc) were examined by a variety of biophysical methods. First, giant vesicles with the diameters of approximately 20 microm were observed by optical microscopy for melittin-DMPC bilayers at 27.9 degrees C. When the temperature was lowered to 24.9 degrees C (Tc = 23 degrees C for the neat DMPC bilayers), the surface of vesicles became blurred and dynamic pore formation was visible in the microscopic picture taken at different exposure times. Phase separation and association of melittin molecules in the bilayers were further detected by fluorescent microscopy and mass spectrometry, respectively. These vesicles disappeared completely at 22.9 degrees C. It was thus found that the melittin-lecithin bilayers reversibly undergo their fusion and disruption near the respective Tcs. The fluctuation of lipids is, therefore, responsible for the membrane fusion above the Tc, and the association of melittin molecules causes membrane fragmentation below the Tc. Subsequent magnetic alignments were observed by solid-state (31)P NMR spectra for the melittin-lecithin vesicles at a temperature above the respective Tcs. On the other hand, additional large amplitude motion induced by melittin at a temperature near the Tc breaks down the magnetic alignment.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1992

A new endoscope for ophthalmic microsurgery.

We present an endoscope designed for ophthalmic microsurgery. The handpiece contains a charge-coupled device, a solid-state imager (250,000 pixels), and a probe (either 0.89 or 1.5 mm in diameter), including the light guide. The light source is a halogen lamp. The clear views the instrument provides of the ciliary sulcus and ciliary body behind the iris make it useful in secondary posterior chamber intraocular lens implantation using the ciliary sulcus suturing technique, and in endolaser photocoagulation of the ciliary body in end-stage glaucoma. It also is useful in performing vitrectomy in cases in which visualization is difficult because of corneal opacification or fluid-gas exchange.

Cataract Extraction↗

PubMed2003

Synthesis, electronic properties, and applications of indium oxide nanowires.

Single-crystalline indium oxide nanowires were synthesized using a laser ablation method and characterized using various techniques. Precise control over the nanowire diameter down to 10 nm was achieved by using monodisperse gold clusters as the catalytic nanoparticles. In addition, field effect transistors with on/off ratios as high as 10(4) were fabricated based on these nanowires. Detailed electronic measurements confirmed that our nanowires were n-type semiconductors with thermal emission as the dominating transport mechanism, as revealed by temperature-dependent measurements. Furthermore, we studied the chemical sensing properties of our In(2)O(3) nanowire transistors at room temperature. Upon exposure to a small amount of NO(2) or NH(3), the nanowire transistors showed a decrease in conductance of up to five or six orders of magnitude, in addition to substantial shifts in the threshold gate voltage. Our devices exhibit significantly improved chemical sensing performance compared to existing solid-state sensors in many aspects, such as the sensitivity, the selectivity, the response time and the lowest detectable concentrations. We have also demonstrated the use of UV light as a "gas cleanser" for In(2)O(3) nanowire chemical sensors, leading to a recovery time as short as 80 seconds.

Crystallization↗

PubMed2006

Proton-decoupled 15N and 31P solid-state NMR investigations of the Pf3 coat protein in oriented phospholipid bilayers.

The coat proteins of filamentous phage are first synthesized as transmembrane proteins and then assembled onto the extruding viral particles. We investigated the transmembrane conformation of the Pseudomonas aeruginosa Pf3 phage coat protein using proton-decoupled 15N and 31P solid-state NMR spectroscopy. The protein was either biochemically purified and uniformly labelled with 15N or synthesized chemically and labelled at specific sites. The proteins were then reconstituted into oriented phospholipid bilayers and the resulting samples analysed. The data suggest a model in which the protein adopts a tilted helix with an angle of approximately 30 degrees and an N-terminal 'swinging arm' at the membrane surface.

Capsid Proteins↗

PubMed2003

Use of confocal microscopy to follow the development of penetrative hyphae during growth of Rhizopus oligosporus in an artificial solid-state fermentation system.

Two methods were compared for determining the concentration of penetrative biomass during growth of Rhizopus oligosporus on an artificial solid substrate consisting of an inert gel and starch as the sole source of carbon and energy. The first method was based on the use of a hand microtome to make sections of approximately 0.2- to 0.4-mm thickness parallel to the substrate surface and the determination of the glucosamine content in each slice. Use of glucosamine measurements to estimate biomass concentrations was shown to be problematic due to the large variations in glucosamine content with mycelial age. The second method was a novel method based on the use of confocal scanning laser microscopy to estimate the fractional volume occupied by the biomass. Although it is not simple to translate fractional volumes into dry weights of hyphae due to the lack of experimentally determined conversion factors, measurement of the fractional volumes in themselves is useful for characterizing fungal penetration into the substrate. Growth of penetrative biomass in the artificial model substrate showed two forms of growth with an indistinct mass in the region close to the substrate surface and a few hyphae penetrating perpendicularly to the surface in regions further away from the substrate surface. The biomass profiles against depth obtained from the confocal microscopy showed two linear regions on log-linear plots, which are possibly related to different oxygen availability at different depths within the substrate. Confocal microscopy has the potential to be a powerful tool in the investigation of fungal growth mechanisms in solid-state fermentation.

Biofilms↗

PubMed2005

Powder-to-powder polycondensation of natural saccharides. Facile preparation of highly branched polysaccharides.

Solid-state polycondensation of a natural saccharide was found to take place in the presence of H3PO4(5 mol%) at 110 degrees C under a N2 flow, giving a highly branched polysaccharide (conv. 11-84%, Mw = 1400-19000, Mn = 1200-3700); the reaction mixture was powdery throughout the polymerization. Interestingly, alpha- and beta-anomers showed different polymerization behaviour; the former was polymerized more slowly, however, they gave comparable molecular weight polymers. The polysaccharide product was per-O-methylated and subjected to structure analyses. The acid-hydrolysis products, the partially O-methylated monosaccharides, suggested that the polysaccharide products have highly branched structures. MALDI-TOF mass analysis revealed that intramolecular glycosylation and acetal exchange reactions are involved in the polymerization mechanism.

Biopolymers↗

PubMed2006

Low-picomolar limits of detection using high-power light-emitting diodes for fluorescence.

Fluorescence detectors are ever more frequently being used with light-emitting diodes (LEDs) as the light source. Technological advances in the solid-state lighting industry have produced LEDs which are also suitable tools in analytical measurements. LEDs are now available which deliver 700 mW of radiometric power. While this greater light power can increase the fluorescence signal, it is not trivial to make proper use of this light. This new generation of LEDs has a large emitting area and a highly divergent beam. This presents a classic problem in optics where one must choose between either a small focused light spot, or high light collection efficiency. We have selected for light collection efficiency, which yields a light spot somewhat larger than the emitting area of the LED. This light is focused onto a flow cell. Increasing the detector cell internal diameter (i.d.) produces gains in (sensitivity)3. However, since the detector cell i.d. is smaller than the LED spot size, scattering of excitation light towards the detector remains a significant source of background signal. This can be minimized through the use of spectral filters and spatial filters in the form of pinholes. The detector produced a limit of detection (LOD) of 3 pM, which is roughly three orders of magnitude lower than other reports of LED-based fluorescence detectors. Furthermore, this LOD comes within a factor of six of much more expensive laser-based fluorescence systems. This detector has been used to monitor a separation from a gel filtration column of fluorescently labeled BSA from residual labeling reagent. The LOD of fluorescently labeled BSA is 25 pM.

Journal Article↗

PubMed2005

Impurity formation studies with peptide-loaded polymeric microspheres Part II. In vitro evaluation.

Since acylated peptide impurities were isolated from octreotide microspheres following incubation in an in vivo environment, the present investigation was undertaken to determine the dosage form dynamics responsible for facilitating acylation. In particular, microsphere batches made with poly(L-lactide) (PLA) and poly(lactide-co-glycolide) (PLGA) 85:15 were studied for in vitro drug release, mass balance relationships, mass loss behavior, hydration uptake, and solid-state stability. Furthermore, native octreotide was incubated in a varying pH stability model (heat treated lactic acid solutions 42.5%, w/w) to determine the effects of acidity on impurity formation. From a review of the experimental results, the appearance of octreotide impurities or related substances occurred with the onset of polymeric mass loss. In fact, the significant formation of acylated peptide did not appear until >90% mass loss, which was observed at 14 days. It was surmised that because of water uptake, the hydrolytic cleavage of the polymeric backbone created an acidic microenvironment to facilitate the covalent coupling of peptide with polymer. The lactic acid solution stability model corroborated with greater evidence of acylation at pH 2.25 where the presence lactoyl (+72 m/z) derivatives of octreotide were confirmed by MALDI-TOF mass spectrometry.

Acylation↗

PubMed2003

Overexpression, purification, and characterization of recombinant Ca-ATPase regulators for high-resolution solution and solid-state NMR studies.

Phospholamban (PLB) and Sarcolipin (SLN) are integral membrane proteins that regulate muscle contractility via direct interaction with the Ca-ATPase in cardiac and skeletal muscle, respectively. The molecular details of these protein-protein interactions are as yet undetermined. Solution and solid-state NMR spectroscopies have proven to be effective tools for deciphering such regulatory mechanisms to a high degree of resolution; however, large quantities of pure recombinant protein are required for these studies. Thus, recombinant PLB and SLN production in Escherichia coli was optimized for use in NMR experiments. Fusions of PLB and SLN to maltose binding protein (MBP) were constructed and optimal conditions for protein expression and purification were screened. This facilitated the large-scale production of highly pure protein. To confirm their functionality, the biological activities of recombinant PLB and SLN were compared to those of their synthetic counterparts. The regulation of Ca-ATPase activity by recombinant PLB and SLN was indistinguishable from the regulation by synthetic proteins, demonstrating the functional integrity of the recombinant constructs and ensuring the biological relevance of our future structural studies. Finally, NMR spectroscopic conditions were established and optimized for use in investigations of the mechanism of Ca-ATPase regulation by PLB and SLN.

Amino Acid Sequence↗

PubMed2006

In search of excited-state proton transfer in the lumichrome dimer in the solid state: theoretical and experimental approach.

Quantum chemical density functional theory (DFT) calculations and spectral data were employed to investigate the possibility of the excited-state double proton transfer (ESDPT) in lumichrome crystals. The calculations in a lumichrome dimer predict a transfer of a proton in the first excited state, leading to a cation-anion pair. The presently reported X-ray structure of 1,3-dimethyllumichrome and its complex solid-state luminescence indicate that also in this molecule intermolecular hydrogen bonds might be involved in the photophysics. The long-wavelength emission in lumichrome crystals peaked at 530 nm is attributed to excited-state proton transfer, whereas a wider emission band in methylated lumichrome derivatives peaked at 560 nm is attributed to ions formed upon photoexcitation of the crystals.

Algorithms↗

PubMed2004

Single-crystal X-ray diffraction studies of photo-induced molecular species.

This tutorial review gathers together the recent developments in single-crystal X-ray diffraction that are starting to enable one to quantify directly the nature of light-induced electronic perturbations in chemical structures. Such structural information is key to understanding many photo-activated chemical processes and physical properties, and a description of the scientific impetus behind this incipient area of structural science, from academic and industrial perspectives, is given. Photoisomerism processes, solid-state photochemical reactions and spin-cross-over magnetic transitions, that have long-lived or irreversible light-induced states, are best understood by unravelling their three-dimensional structures measured in situ in their photo-converted state. A review of steady-state laser-induced single-crystal X-ray diffraction studies conducted, to date, and the experimental methodologies used in order to realise such structures, is presented. The structural characterisation of more transient photo-induced species (down to picosecond lifetimes) is paramount to a better understanding of the materials that undergo high-speed electronic switching, which make operative much of the electronics and optics industry, since there exists an inherent relationship between the excited-state structure and the physical properties exhibited. Prime examples include excited-state structures of molecular conductors and luminescent materials with potential applications as molecular wires, light-emitting diodes, non-linear optics, triboluminescence and electroluminescence. Previously, only indirect and qualitative interpretations of the nature of these excited-states could be formulated via spectroscopic techniques, but the developments in ms-ps time-resolved laser pump, X-ray probe single-crystal diffraction techniques, described herein, are overcoming this barrier, affording results that are entirely quantitative via a three-dimensional structural representation. In this regard, a review of structures of transient species studied to date is presented along with a discussion of the key experimental parameters that are required for a successful experiment, in terms of the X-ray, laser and sample characteristics. The importance of auxiliary spectroscopic work and complementary theoretical calculations is also briefly discussed. The paper concludes with a future outlook on new possible X-ray sources that will facilitate such work and extend it to structural studies on even more ephemeral species in the future.

Journal Article↗

PubMed2006

Optically directed molecular transport and 3D isoelectric positioning of amphoteric biomolecules.

We demonstrate the formation of charged molecular packets and their transport within optically created electrical force-field traps in a pH-buffered electrolyte. We call this process photoelectrophoretic localization and transport (PELT). The electrolyte is in contact with a photoconductive semiconductor electrode and a counterelectrode that are connected through an external circuit. A light beam directed to coordinates on the photoconductive electrode surface produces a photocurrent within the circuit and electrolyte. Within the electrolyte, the photocurrent creates localized force-field traps centered at the illuminated coordinates. Charged molecules, including polypeptides and proteins, electrophoretically accumulate into the traps and subsequently can be transported in the electrolyte by moving the traps over the photoconductive electrode in response to movement of the light beam. The molecules in a single trap can be divided into aliquots, and the aliquots can be directed along multiple routes simultaneously by using multiple light beams. This photoelectrophoretic transport of charged molecules by PELT resembles the electrostatic transport of electrons within force-field wells of solid-state charge-coupled devices. The molecules, however, travel in a liquid electrolyte rather than a solid. Furthermore, we have used PELT to position amphoteric biomolecules in three dimensions. A 3D pH gradient was created in an electrolyte medium by controlling the illumination position on a photoconductive anode where protons were generated electrolytically. Photoelectrophoretic transport of amphoteric molecules through the pH gradient resulted in accumulation of the molecules at their apparent 3D isoelectric coordinates in the medium.

Animals↗

PubMed2003

Molecular weight, tertiary structure, water binding and colon behaviour of ispaghula husk fibre.

Molecular variables, using aqueous and alkaline extracts, of the polysaccharide from ispaghula husk (IH) were examined using gel-permeation chromatography linked to multi-angle laser light scattering. Progressive extraction can yield a component with a molecular weight (MW)value up to about 7 x 106 Da, and gels, which accompany the extraction, have MW ranging from 10-20 x 106 Da. To mimic the polysaccharide degradation, particularly in the colon, the solid IH was degraded progressively using ionising radiation. A chain break occurs every 7.5 kGy in NaOH and every 15 kGy in water. The solid-state matrix is opened by the radiation to yield increased visco-elasticity of the aqueous extracts at critical radiation doses, before further degradation occurs after about 12 kGy. Differential scanning calorimetry is used to study the mechanism of interaction of water with IH. The first water to be taken up is non-freezing water and represents about twelve water molecules/disaccharide unit of the polysaccharide. As the water content is increased, the water becomes bound to the polysaccharide and freezes and melts at a temperature different from free water. This water is thermodynamically distinguishable from free water. It forms amorphous ice on cooling which crystallises exothermically and subsequently melts endothermically. Saturation occurs at a water content of 2-3 g water/g polymer, showing that about 60% of the water in the system is 'bound'. The most surprising conclusion is that despite the fact that the IH swells in water to form a solid and stiff gel, the greater part of that water in the gel is still free and behaves like liquid water.

Chromatography, Gel↗

PubMed2001

A nifedipine coground mixture with sodium deoxycholate. I. Colloidal particle formation and solid-state analysis.

Sodium deoxycholate (DCNa) is a bile salt that forms multimolecular inclusion compounds with a variety of organic substances. In this study, complex formulation of DCNa with nifedipine, a poorly water soluble drug, by grinding was investigated. The coground mixture was prepared with a vibration rod mill, and its solid state was characterized using powder X-ray diffraction, differential scanning calorimetry (DSC), and Fourier transform infrared (FTIR) spectroscopy. A laser diffraction particle size analyzer was also used to determine the particle size distribution curve in solution. When a nifedipine-DCNa (1:2 w/w) mixture coground for 30 min was dispersed into water and a pH 6.8 buffer solution, a semitransparent colloidal solution occurred immediately; 90% of the total particles formed in solution had a diameter less than 600 nm. Both powder X-ray diffraction peaks and DSC endothermic peak of nifedipine crystals were not found for the coground mixture, whereas a new exothermic peak was observed on DSC thermograms. The magnitude of this exothermic peak depended on the weight fraction of DCNa and the grinding time, indicating that nifedipine crystals changed into an amorphous state by complex formation with DCNa during the grinding process. In the FTIR spectrum of the coground mixture, the peaks of aromatic CH out-of-plane bend and dihydropyridine NH stretch of nifedipine were considerably weakened, suggesting that van der Waals interaction may be present between the drug and DCNa molecules. From these results, it is clear that the cogrinding method with DCNa is very useful for the formation of amorphous nifedipine in the solid state and the production of colloidal particles of the drug in solution.

Buffers↗

PubMed2002

Scalable solid-state quantum processor using subradiant two-atom states.

We propose a realization of a scalable, high-performance quantum processor whose qubits are represented by the ground and subradiant states of effective dimers formed by pairs of two-level systems coupled by resonant dipole-dipole interaction. The dimers are implanted in low-temperature solid host material at controllable nanoscale separations. The two-qubit entanglement either relies on the coherent excitation exchange between the dimers or is mediated by external laser fields.

Journal Article↗

PubMed2003

Broadband saturable absorber with cobalt-doped tellurite glasses.

A robust solid-state saturable absorber was developed with cobalt-ion-doped tellurite glasses that has a broad saturable-absorption band in the near-infrared. A nonlinear optical absorber was measured by the Z-scan technique. Numerically fitting the Z-scan experimental data yields excited-state absorption cross sections at a 1064-nm laser wavelength of sigma(es) = 1.45 x 10(-20) and sigma(es) = 1.15 x 10(-20) cm2 for 1.0- and 2.5-at. % Co2+-doped glasses, respectively.

Journal Article↗