Sensing Coated Iron-Oxide Nanoparticles with Spectral Induced Polarization (SIP): Experiments in Natural Sand Packed Flow-Through Columns.

The development of nanoparticle-based soil remediation techniques is hindered by the lack of accurate in situ nanoparticle (NP) monitoring and characterization methods. Spectral induced polarization (SIP), a noninvasive geophysical technique, offers a promising approach to detect and quantify NPs in porous media. However, its successful implementation as a monitoring tool requires an understanding of the polarization mechanisms, the governing NP-associated SIP responses and their dependence on the stabilizing coatings that are typically used for NPs deployed in environmental applications. Herein, we present SIP responses (0.1-10 000 Hz) measured during injection of a poloxamer-coated superparamagnetic iron-oxide nanoparticle (SPION) suspension in flow-through columns packed with natural sand from the Borden aquifer. An advective-dispersive transport model is fitted to outflow SPION concentration measurements to compute average concentrations over the SIP spatial response domain (within the columns). The average SPION concentrations are compared with the real and imaginary components of the complex conductivity. Excellent correspondence is found between the average SPION concentrations in the columns and the imaginary conductivity values, suggesting that NP-mediated polarization (that is, charge storage) increases proportionally with increasing SPION concentration. Our results support the possibility of SIP monitoring of spatial and temporal NP distributions, which can be immediately deployed in bench-scale studies with the prospect of future real-world field applications.

Evaluation of antiphospholipid antibody assays using latent class analysis to address the lack of a reference standard.

BACKGROUND: Method evaluation of new assays for the detection of antiphospholipid antibodies (aPL) such as anti-cardiolipin (aCL) or anti-ß2-glycoprotein I (aß2-GPI) is challenging, as no internationally accepted reference material is available yet. Besides a lack of standardization, unacceptable inter-laboratory comparability of established tests is regularly observed. Owing to the absence of a commonly accepted reference standard, the evaluation of two research surface plasmon resonance (SPR) biosensor assays was performed using statistical methods from latent class analysis (LCA). METHODS: aCL and aß2-GPI IgG and IgM were measured in sera from 63 antiphospholipid syndrome patients, fulfilling the Sydney criteria, and in 34 healthy controls with four commercial assays. LCA was performed on the results and sera were assigned to the antibody-positive or antibody-negative group. Sera were subsequently evaluated in the SPR assays for aCL and aß2-GPI. Optimal cutoffs and diagnostic performances of the research systems were established employing the LCA-derived gold standard. RESULTS: With area under the curve results of 0.96 and 0.89 for the detection of aCL and aß2-GPI, the research SPR assays discriminated well between antibody-positive and antibody-negative sera. Their sensitivities and specificities were comparable to the investigated commercial immunoassays. CONCLUSIONS: SPR assays are a suitable tool for the detection of aCL and aß2-GPI with diagnostic performances not different from currently available commercial tests. LCA enabled the calculation of sensitivities and specificities for aPL assays in absence of a reference standard.

Organic Photovoltaic Materials--Design, Synthesis and Scale-Up.

This account describes the work of our group in the area of organic photovoltaics in the past six years. The emphasis is on our experiences in the development of the organic materials, their characterization, scale-up and application in devices. We share our insight into the relationship between synthetic methods, molecular properties, bulk material properties and device performance.

Plasmodium falciparum coronin organizes arrays of parallel actin filaments potentially guiding directional motility in invasive malaria parasites.

BACKGROUND: Gliding motility in Plasmodium parasites, the aetiological agents of malaria disease, is mediated by an actomyosin motor anchored in the outer pellicle of the motile cell. Effective motility is dependent on a parasite myosin motor and turnover of dynamic parasite actin filaments. To date, however, the basis for directional motility is not known. Whilst myosin is very likely orientated as a result of its anchorage within the parasite, how actin filaments are orientated to facilitate directional force generation remains unexplained. In addition, recent evidence has questioned the linkage between actin filaments and secreted surface antigens leaving the way by which motor force is transmitted to the extracellular milieu unknown. Malaria parasites possess a markedly reduced repertoire of actin regulators, among which few are predicted to interact with filamentous (F)-actin directly. One of these, PF3D7_1251200, shows strong homology to the coronin family of actin-filament binding proteins, herein referred to as PfCoronin. METHODS: Here the N terminal beta propeller domain of PfCoronin (PfCor-N) was expressed to assess its ability to bind and bundle pre-formed actin filaments by sedimentation assay, total internal reflection fluorescence (TIRF) microscopy and confocal imaging as well as to explore its ability to bind phospholipids. In parallel a tagged PfCoronin line in Plasmodium falciparum was generated to determine the cellular localization of the protein during asexual parasite development and blood-stage merozoite invasion. RESULTS: A combination of biochemical approaches demonstrated that the N-terminal beta-propeller domain of PfCoronin is capable of binding F-actin and facilitating formation of parallel filament bundles. In parasites, PfCoronin is expressed late in the asexual lifecycle and localizes to the pellicle region of invasive merozoites before and during erythrocyte entry. PfCoronin also associates strongly with membranes within the cell, likely mediated by interactions with phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2) at the plasma membrane. CONCLUSIONS: These data suggest PfCoronin may fulfil a key role as the critical determinant of actin filament organization in the Plasmodium cell. This raises the possibility that macro-molecular organization of actin mediates directional motility in gliding parasites.

Medium-Ring Effects on the Endo/Exo Selectivity of the Organocatalytic Intramolecular Diels-Alder Reaction.

The intramolecular Diels-Alder reaction has been used as a powerful method to access the tricyclic core of the eunicellin natural products from a number of 9-membered-ring precursors. The endo/exo selectivity of this reaction can be controlled through a remarkable organocatalytic approach, employing MacMillan's imidazolidinone catalysts, although the mechanistic origin of this selectivity remains unclear. We present a combined experimental and density functional theory investigation, providing insight into the effects of medium-ring constraints on the organocatalyzed intramolecular Diels-Alder reaction to form the isobenzofuran core of the eunicellins.

Development of a new parallelized, optical biosensor platform for label-free detection of autoimmunity-related antibodies.

Autoimmune diseases are characterized by the presence of autoantibodies in serum of affected patients. The heterogeneity of autoimmune relevant antigens creates a variety of different antibodies, which requires a simultaneous detection mode. For this reason, we developed a tool for parallelized, label-free, optical detection that accomplishes the characterization of multiple antigen-antibody interactions within a single measurement on a timescale of minutes. Using 11-aminoundecyltrimethoxysilane, we were able to immobilize proteinogenic antigens as well as an amino-functionalized cardiolipin on a glass surface. Assay conditions were optimized for serum measurements with a single spot antigen chip on a single spot 1-λ detection system. Minimized background signal allows a differentiation between patients and healthy controls with a good sensitivity and specificity. Applying polarized imaging reflectometric interference spectroscopy, we evaluated samples from three APS patients and three control subjects for this proof-of-principle and already obtained good results for ß2-glycoprotein I and cardiolipin.

Covalent attachment of functionalized cardiolipin on a biosensor gold surface allows repetitive measurements of anticardiolipin antibodies in serum.

Antiphospholipid antibodies (aPL) are a relevant serological indicator of antiphospholipid syndrome (APS). A solid-state surface with covalently bound ω-amine-functionalized cardiolipin was established and the binding of ß2-glycoprotein I (ß2-GPI) was investigated either by use of surface plasmon resonance (SPR) biosensor, by electrically switchable DNA interfaces (switchSENSE) and by scanning tunneling microscopy (STM). STM could clearly visualize the attachment of ß2-GPI to the cardiolipin surface. Using the switchSENSE sensor, ß2-GPI as specific ligand could be identified by increased hydrodynamic friction. The binding of anti-cardiolipin antibodies (aCL) was detected against the ω-amine-functionalized cardiolipin-modified SPR biosensor (aCL biosensor) using sera from healthy donors, APS patients and syphilis patients. Our results showed that the aCL biosensor is a much more sensitive diagnostic device for APS patients compared to previous methods. The specificity between ß2-GPI-dependent autoimmune- and ß2-GPI-independent infection-associated types of aPLs was also studied and they can be distinguished by the different binding kinetics and patterns.

Photophysics and aggregation effects of a triphenylamine-based dye sensitizer on metal-oxide nanoparticles suspended in an ion trap.

The photophysical behaviour of a triphenylamine-based organic dye sensitizer (Carbz-PAHTDTT) attached to alumina and titania nanoparticles (labelled Carbz-Al and Carbz-Ti, respectively) is examined in the absence and presence of the chenodeoxycholic acid (CDCA) coadsorber. The experiments are conducted in vacuo by suspending the target dye-sensitized nanoparticles within a quadrupole ion trap, where they are probed with laser radiation to obtain emission spectra and time-resolved excited state decay curves. For Carbz-Al, the dye's emission band is blue-shifted and the excited state lifetime is increased upon the coabsorption of CDCA, effects attributed to reduced dye aggregation. Compared to Carbz-Al, the Carbz-Ti excited state lifetimes are significantly shorter due to excited dye molecules injecting electrons into the titania conduction band. For Carbz-Ti, the electron injection quantum yields for the surfaces with CDCA (CDCA : dye = 25 : 1) and without CDCA are estimated to be 0.87 and 0.71, respectively. The gas-phase results demonstrate that Carbz-PAHTDTT dye aggregates are detrimental to the performance of a dye-sensitized solar cell.

Enhancement of efficiency in organic photovoltaic devices containing self-complementary hydrogen-bonding domains.

Self-complementary hydrogen-bonding domains were incorporated as the electron deficient unit in "push-pull", p-type small molecules for organic photovoltaic active layers. Such compounds were found to enhance the fill factor, compared with similar non-self-organized compounds reported in the literature, leading to higher device efficiencies. Evidence is presented that the ability of these molecules to form one-dimensional hydrogen-bonded chains and subsequently exhibit hierarchical self-assembly into nanostructured domains can be correlated with improved device efficiency.

Causation in a cascade: the origins of selectivities in intramolecular nitrone cycloadditions.

The factors controlling chemo-, regio-, and stereoselectivity in a cascade of reactions starting from a bis(cyanoalkenyl)oxime and proceeding via nitrone cycloadditions have been unraveled through a series of density functional theory calculations with several different functionals. Both kinetic and thermodynamic control of the reaction cascade are important, depending upon the conditions. Kinetic control was analyzed by the distortion/interaction model and found to be dictated by differences in distortions of the cycloaddends in the transition states. A new mechanism competing with that originally proposed in the application of these reactions to the histrionicotoxin synthesis was discovered in these studies.

Reverse cope elimination of hydroxylamines and alkenes or alkynes: theoretical investigation of tether length and substituent effects.

Quantum mechanical calculations have been used to study the intramolecular additions of hydroxylamines to alkenes and alkynes ("reverse Cope eliminations"). In intermolecular reverse Cope eliminations, alkynes are more reactive than alkenes. However, competition experiments have shown that tethering the hydroxylamine to the alkene or alkyne can reverse the reactivity order from that normally observed. The exact outcome depends on the length of the tether. In agreement with experiment, a range of density functional theory methods and CBS-QB3 calculations predict that the activation energies for intramolecular reverse Cope eliminations follow the order 6-exo-dig < 5-exo-trig < 5-exo-dig ≈ 7-exo-dig. The order of the barriers for the 5-, 6-, and 7-exo-dig reactions of alkynes arises mainly from differences in tether strain in the transition states (TSs), but is also influenced by the TS interaction between the hydroxylamine and alkyne. Cyclization onto an alkene in the 5-exo-trig fashion incurs slightly less tether strain than a 6-exo-dig alkyne cyclization, but its activation energy is higher because the hydroxylamine fragment must distort more before the TS is reached. If the alkene terminus is substituted with two methyl groups, the barrier becomes so much higher that it is also disfavored compared to the 5- and 7-exo-dig cyclizations.

Intramolecular nitrone dipolar cycloadditions: control of regioselectivity and synthesis of naturally-occurring spirocyclic alkaloids.

The intramolecular nitrone dipolar cycloaddition of in situ-generated nitrones such as compound 26 has been used for the synthesis of cyclic isoxazolidines 27 and 29. The regioselectivity of the intramolecular cycloaddition depends on the nature of the terminal substituent on the dipolarophile. The influence of the substituent on the regioselectivity of the cycloaddition has been examined using several model systems and two methods of nitrone formation. These studies demonstrated that the cyano-substituent plays a special role in favouring the formation of the 6,6,5-ring fused adduct 27 under thermodynamically controlled conditions. The utility of the cyclo-adduct 57 (see Scheme 12) as a precursor for the naturally occurring histrionicotoxins is illustrated by the synthesis of three "unsymmetrical" (i.e. with each side chain bearing different functional groups) members of the histrionicotoxin family HTX-259A, HTX-285C and HTX-285E (2, 3 and 4 respectively).

Selective photocatalytic reduction of selenate over TiO2 in the presence of nitrate and sulfate in mine-impacted water.

Selenium contamination is a critical global issue across numerous industries. Industrial waters such as mine-impacted water (MIW) can contain toxic levels of selenate, in addition to varying concentrations of many different dissolved species from the underlying strata, such as sulfate, carbonate, nitrate, organic matter, and many dissolved metals. The removal of selenate from MIW is desired, due to selenate's acute and chronic toxicity in aquatic ecosystems at elevated concentrations. However, due to the complexity of the water matrix and the presence of many other dissolved constituents, this is often very challenging. In this study, we present for the first time the reduction of selenate in a real industrial wastewater, namely MIW, and reveal a significant advantage of photocatalytic reduction; the ability to selectively reduce selenate from >500 µg L-1 to <2 µg L-1 in the presence of the more energetically favourable electron acceptor, nitrate (250× molar concentration of selenate) and high concentrations of sulfate (1,940× molar concentration of selenate). The presence and impacts of sulfate, chloride, carbonate, and nitrate on the competitive adsorption and reduction of selenate on TiO2 are thoroughly investigated for the first time, using formic acid as an electron hole scavenger. The electron transfer mechanism proposed follows TiO2 conduction band electrons are responsible for the reduction of selenate to elemental Se (Se0) and both carbon dioxide radicals (CO2·-) and Se conduction band electrons are responsible for the further reduction of Se0 to hydrogen selenide (H2Se).

Hierarchical self-assembly of semiconductor functionalized peptide α-helices and optoelectronic properties.

To determine the ability of semiconductors templated by α-helical polypeptides to form higher order structures and the charge carrier properties of the supramolecular assemblies, L-lysine was functionalized with a sexithiophene organic semiconductor unit via iterative Suzuki coupling and the click reaction. The resultant amino acid was incorporated into a homopolypeptide by ring-opening polymerization of an amino acid N-carboxyanhydride. Spectroscopic investigation of the polypeptide revealed that it adopted an α-helical secondary structure in organic solvents that underwent hierarchical self-assembly to form higher order structures. In cyclohexane, the polymer formed organogels at 2% (w/v). Organic photovoltaic and organic field effect transistor devices were fabricated by deposition of the PCBM blended active layer from chlorobenzene at concentrations shown to induce self-assembly of the polymer. Compared with control compounds, these devices showed significantly greater hole mobility, short circuit current, and efficiency. This work establishes the potential of this previously unreported bioinspired motif to increase device performance.

Fluorenyl hexa-peri-hexabenzocoronene-dendritic oligothiophene hybrid materials: synthesis, photophysical properties, self-association behaviour and device performance.

Apart from molecular properties, intermolecular forces play a vital role in defining the performance of organic electronic devices. This is particularly relevant in bulk heterojunction (BHJ) solar cells in which the arrangement of electron-donor and -acceptor materials into distinct crystalline phases of ideal size and distribution can lead to better power conversion efficiencies. In this study, a series of fluorenyl hexa-peri-hexabenzocoronenes (FHBC) decorated with thiophene dendrons (DOT) of variable size was obtained by using a convergent synthetic approach. With such variety of molecular sizes and shapes in hand, the objective of this study is to highlight the relationships between molecular properties, bulk properties and device performance. Correlations between π-π stacking ability and dendrimer generation were established from self-organisation studies in solution and solid state. The synergistic combination of molecular organisation at the nanoscale and photophysical characteristics derived from the FHBC and DOT moieties leads to a notable improvement of the photovoltaic performance.

Continuous flow synthesis of fullerene derivatives.

Various fullerene-based electron acceptor materials for organic photovoltaic applications were prepared via [3+2] and [4+2] cycloadditions using a continuous flow approach. The 1,3-dipolar cycloaddition of the tosylhydrazone precursor and the Diels-Alder cycloaddition of indene to either C(60) or C(70) under conventional batch reaction conditions were translated to the continuous flow process. By varying the residence time, temperature, and equivalents of cycloaddition reagent, significant improvements in yields and reaction times were achieved over conventional batch processes.

Dithienothiophene (DTT)-based dyes for dye-sensitized solar cells: synthesis of 2,6-dibromo-DTT.

A one-pot synthesis of 2,6-dibromodithieno[3,2-b;2',3'-d]thiophene (dibromo-DTT, 4) was developed. A key step was bromodecarboxylation of DTT-2,6-dicarboxylic acid, obtained by saponification of the diester 1. The donor-acceptor dye DAHTDTT (13), based on a central 2,6-bis[2'-(3'-hexylthienyl)]dithieno[3,2-b;2',3'-d]thiophene core (9), was prepared and incorporated in a dye-sensitized solar cell (DSC), which exhibited an energy conversion efficiency of 7.3% with V(oc) of 697 mV, J(sc) of 14.4 mA/cm(2), and ff of 0.73 at 1 sun.

A gram-scale batch and flow total synthesis of perhydrohistrionicotoxin.

The total synthesis of the spiropiperidine alkaloid (-)-perhydrohistrionicotoxin (perhydro-HTX) 2 has been accomplished on a gram scale by employing both conventional batch chemistry as well as microreactor techniques. (S)-(-)-6-Pentyltetrahydro-pyran-2-one 8 underwent nucleophilic ring opening to afford the alcohol 10, which was elaborated to the nitrone 13. Protection of the nitrone as the 1,3-adduct of styrene and side-chain extension to the unsaturated nitrile afforded a precursor 17, which underwent dipolar cycloreversion and 1,3-dipolar cycloaddition to give the core spirocyclic precursor 18 that was converted into perhydro-HTX 2. The principal steps to the spirocycle 18 have successfully been transferred into flow mode by using different types of microreactors and in a telescoped fashion, allowing for a more rapid access to the histrionicotoxins and their analogues by continuous processing.

Analysis of cellular phosphatidylinositol (3,4,5)-trisphosphate levels and distribution using confocal fluorescent microscopy.

We have developed an immunocytochemistry method for the semiquantitative detection of phosphatidylinositol (3,4,5)-trisphosphate (PI(3,4,5)P3) at the cell plasma membrane. This protocol combines the use of a glutathione S-transferase-tagged pleckstrin homology (PH) domain of the general phosphoinositides-1 receptor (GST-GRP1PH) with fluorescence confocal microscopy and image segmentation using cell mask software analysis. This methodology allows the analysis of PI(3,4,5)P3 subcellular distribution in resting and epidermal growth factor (EGF)-stimulated HEK293T cells and in LIM1215 (wild-type phosphoinositide 3-kinase (PI3K)) and LIM2550 (H1047R mutation in PI3K catalytic domain) colonic carcinoma cells. Formation of PI(3,4,5)P3 was observed 5min following EGF stimulation and resulted in an increase of the membrane/cytoplasm fluorescence ratio from 1.03 to 1.53 for HEK293T cells and from 2.2 to 3.3 for LIM1215 cells. Resting LIM2550 cells stained with GST-GRP1PH had an elevated membrane/cytoplasm fluorescence ratio of 9.8, suggesting constitutive PI3K activation. The increase in the membrane/cytoplasm fluorescent ratio was inhibited in a concentration-dependent manner by the PI3K inhibitor LY294002. This cellular confocal imaging assay can be used to directly assess the effects of PI3K mutations in cancer cell lines and to determine the potential specificity and effectiveness of PI3K inhibitors in cancer cells.

Synthesis and biological evaluation of phosphatidylinositol phosphate affinity probes.

The synthesis of the complete family of phosphatidylinositol phosphate analogues (PIPs) from five key core intermediates A-E is described. These core compounds were obtained from myo-inositol orthoformate 1 via regioselective DIBAL-H and trimethylaluminium-mediated cleavages and a resolution-protection process using camphor acetals 10. Coupling of cores A-E with phosphoramidites 34 and 38, derived from the requisite protected lipid side chains, afforded the fully-protected PIPs. Removal of the remaining protecting groups was achieved via hydrogenolysis using palladium black or palladium hydroxide on carbon in the presence of sodium bicarbonate to afford the complete family of dipalmitoyl- and amino-PIP analogues 42, 45, 50, 51, 58, 59, 67, 68, 76, 77, 82, 83, 92, 93, 99 and 100. Investigations using affinity probes incorporating these compounds have identified novel proteins involved in the PI3K intracellular signalling network and have allowed a comprehensive proteomic analysis of phosphoinositide interacting proteins.