Generalized eikonal of partially coherent beams and its use in quantitative imaging

The generalized eikonal of a partially coherent paraxial wave is introduced via a differential equation describing the evolution of the time-averaged intensity. The theoretical formalism provides an analytical tool for the study of partially coherent imaging systems. It also makes possible quantitative phase retrieval and compositional mapping of weakly absorbing samples using phase-contrast imaging with broadband polychromatic radiation of known spectral distribution. An experimental demonstration is presented of the quantitative reconstruction of the projected thickness of a sample, given a phase-contrast image obtained using a polychromatic microfocus x-ray source.     

Refractive microlens array for wave-front analysis in the medium to hard x-ray range

We report an alternative approach to x-ray wave-front analysis that uses a refractive microlens array as a Shack-Hartmann sensor. The sensor was manufactured by self-assembly and electroplating techniques and is suitable for high-resolution wave-front analysis of medium to hard x rays. We demonstrate its effectiveness at an x-ray energy of 3 keV for analysis of x-ray wave-front perturbations caused by microscopic objects. The sensor has potential advantages over other methods for x-ray phase imaging and will also be useful for the characterization of x-ray beams and optics.     

A designed protein interface that blocks fibril formation

Protein fibril formation is implicated in many diseases, and therefore much effort has been focused toward the development of inhibitors of this process. In a previous project, a monomeric protein was computationally engineered to bind itself and form a heterodimer complex following interfacial redesign. One of the protein monomers, termed monomer-B, was unintentionally destabilized and shown to form macroscopic fibrils. Interestingly, in the presence of the designed binding partner, fibril formation was blocked. Here we describe the complete characterization of the amyloid properties of monomer-B and the inhibition of fiber formation by the designed binding partner, monomer-A. Both proteins are mutants of the betal domain of streptococcal protein-G. The free monomer-B protein forms amyloid-type fibrils, as determined by transmission electron microscopy and the change in fluorescence of Thioflavin T, an amyloid-specific dye. Fibril formation kinetics are influenced by pH, protein concentration, and seeding with preformed fibrils. Under all conditions tested, monomer-A was able to inhibit the formation of monomer-B fibrils. This inhibition is specific to the engineered interaction, as incubation of monomer-B with wild-type protein-G (a structural homologue) did not result in inhibition under the same conditions. Thus, this de novo-designed heterodimeric complex is an excellent model system for the study of protein-based fibril formation and inhibition. This system provides additional insight into the development of pharmaceuticals for amyloid disorders, as well as the potential use of amyloid fibrils for self-assembling nanostructures.     

Radical performance enhancements for combinatorial optimization algorithms based on the dead-end elimination theorem

Recent advances in protein design have demonstrated the effectiveness of optimization algorithms based on the dead-end elimination theorem. The algorithms solve the combinatorial problem of finding the optimal placement of side chains for a set of backbone coordinates. Although they are powerful tools, these algorithms have severe limitations when the number of side chain rotamers is large. This is due to the high-order time dependence of the aspect of the calculation that deals with rotamer doubles. We present three independent algorithmic enhancements that significantly increase the speed of the doubles computation. These methods work by using quantities that are inexpensive to compute as a basis for forecasting which expensive calculations are worthwhile. One of the methods, the comparison of extrema, is derived from analytical considerations, and the remaining two, the “magic-bullet” and the “q_rs” and “q_uv” metrics, are based on empirical observation of the distribution of energies in the system. When used together, these methods effect an overall speed improvement of as much as a factor of 47, and for the doubles aspect of the calculation, a factor of 95. Together, these enhancements extend the envelope of inverse folding to larger proteins by making formerly intractable calculations attainable in reasonable computer time. © 1998 John Wiley & Sons, Inc. J Comput Chem 19: 1505–1514, 1998     

Aging and degradation of polyolefins. I. Peroxide-initiated oxidations of atactic polypropylene

Efficiencies of polymer radical production by thermal decomposition of di-tert-butylperoxy oxalate (DBPO) have been measured in bulk atactic polypropylene (PP) at 25–55°C; they range from 1 to 26%, depending on [DBPO], temperature, and presence of oxygen. Most of the polymer radicals thus produced disproportionate in the absence of oxygen but form peroxy radicals in its presence. Most of the pairs of peroxy radicals interact by a first-order reaction in the polymer cage. The fraction that escapes gives hydroperoxide in a reaction that is half order in rate of initiation. In interactions of polymer peroxy radicals, in or out of the cage, about one-third give dialkyl peroxides and immediate chain termination, two-thirds give alkoxy radicals. About one-third of the later cleave at 45°C; the rest abstract hydrogen to give hydroxy groups and new polymer and polymer peroxy radicals. The primary peroxy radicals from cleavage account for the rest of the chain termination. Cleavage of alkoxy radicals and crosslinking of PP through dialkyl peroxides nearly compensate. Up to 70% of the oxygen absorbed has been found in hydroperoxides. The formation of these can be completely inhibited, but cage reactions are unaffected by inhibitors. Concentrations of free polymer peroxy radicals have been measured by electron spin resonance and found to be very high, about 10^?3M at 58–63°C. Comparison with results on 2,4-dimethylpentane indicate that rate constants for both chain propagation and termination in the polymer are much smaller than those for the model hydrocarbon but that the ratio, k_p/(2k_t)^½, is about the same.     

Aging and degradation of polyolefins. II. γ-initiated oxidations of atactic polypropylene

Oxidations of bulk atactic polypropylene (PP) have been carried out at 22 and 45°C, and the dependence of rate of formation of each product on rate of initiation has been determined. The principal product is PP hydroperoxide, formed in a half-order reaction. One termination product is polymeric dialkyl peroxide, formed in a first-order reaction. Other termination and propagation products, alcohols and carbonyl compounds, are formed in reactions that are mostly first-order in initiation. At 22°C, G is 9–63. G is about three times as great at 45°C as at 22°C. Experiments with 2,6-di-tert-butyl-p-cresol shows that it can inhibit all non-cage propagation and all formation of PP hydroperoxide, but that it does not affect cage reactions of initiating radicals and their successors. Only about 16% of the initiating PPO₂· radicals escape the cage at 45°C. Oxidations of PP, n-hexane, and their mixture with both peroxide and γ-ray initiation show that nearly all the initiating radicals escape the cage in solution but that the concentration of PPO₂· radicals is much less than in bulk because of a much faster chain termination. Both the propagation and termination constants for PP oxidation are much faster in solution, but the changes compensate so that k_p/(2k_t)^½ is about the same in solution as in bulk.     

Development and validation of a single HPLC method for determination of α‐tocopherol in cell culture and in human or mouse biological samples

A straightforward and common analytical method for α‐tocopherol (αT) determination in various biological samples, including plasma, red blood cells (RBC), tissues and cultured cell lines, was developed and validated, using a reverse phase‐chromatographic method (RP‐HPLC). Even though many chromatographic methods for αT determination have been reported, most of them require readjustment when applied to different types of samples. Thus, an effective and simple method for αT determination in different biological matrices is still necessary, specifically for translational research. This method was applied using a C₁₈ column (250 × 4.6 mm, 5 µm particle size) under isocratic elution with MeOH:ACN:H₂O (90:9:1 v/v/v) at a flow rate of 1 mL/min and detected using photodiode array at 293 nm. Linearity (r >0.9997) was observed for standard calibration with inter‐ and intraday variation of standard <4%. Lower limits of detection and quantification for αT in this assay were 0.091 and 0.305 µg/mL respectively. Validation proved the method to be selective, linear, accurate and precise. The method was successfully applied in great variety of biological samples, that is, human and mouse plasma, RBCs, murine tissues and human/mouse/rat cultured cell lines. More importantly, a single protocol of extraction and detection can be applied, making this method very convenient for standardization of different types of samples. Copyright © 2014 John Wiley & Sons, Ltd.     

Selective analysis of histamine in food by means of solid-phase extraction cleanup and chromatographic separation

A simple, practical technique is presented for the selective determination and measurement of histamine (HA) levels in fermented food. The method involved a solid-phase extraction cleanup using a Sep-Pak Plus C-18 cartridge and ion-paired reversed-phase high-performance liquid chromatographic (IP-RP-HPLC) separation, followed by detection of HA at its UV absorbance wavelength of 220nm. After evaporating a methanolic extract from the food sample, the resulting residue was reconstituted with 0.2M phosphate buffer (pH 3.0), and subsequently passed through the cartridge. The aliquot of the solution which came out of the cartridge was chromatographed in IP-RP mode on a C-18 column, as the stationary phase, and with a solution of 0.2M phosphate buffer (pH 3.0)-acetonitrile-water (1:24:166, v/v) containing 2mM sodium 1-octane sulfonic acid, as the mobile phase. When this method was applied to a mixture of HA, Cadaverine (Cad), Putrescine (Put), Serotonin (5HT), and Tyramine (Tyr), only HA was detected at 16.4min of retention time. The method was fully validated and validation parameters were: linearity range 2-1000ppm; correlation coefficient >0.991; mean recovery >99.5%; limit of quantification 2ppm and limit of detection 0.5ppm. The method was next applied to 12 brands of Miso (fermented soybean paste), 9 brands of Sake (rice wine), and 5 brands of Shouyu (Japanese soy sauce) to verify its ability to detect the presence of HA in a variety of fermented foods. The method proved to be both rapid and accurate and is therefore recommended for use in HA pollution surveys and in the routine practice of food-quality control.