Preparation and biodegradation of sugar-containing poly(vinyl acetate) emulsions

To accelerate the biodegradability of poly(vinyl acetate)-based emulsions, emulsion copolymerizations of vinyl sugars, including triacetylated N-acetyl-D-glucosamine (GlcNAc)-substituted 2-hydroxyethyl methacrylate (GlcNAc(Ac)3-substituted HEMA), glucose-substituted HEMA (GEMA) and 6-O-vinyladipoyl-D-glucose (6-O-VAG) with vinyl acetate (VAc), were carried out using poly(vinyl alcohol) as an emulsifying agent in the presence of poly[(butylene succinate)-co-(butylene adipate)] [poly(BS-co-BA)]. Copolymerization with GEMA produced a stable emulsion and that with 6-O-VAG also produced a homogeneous emulsion. Their biodegradation tests indicated that PVAc main chain scission was accelerated by copolymerization with vinyl sugars.     

On-chip fabrication of mutifunctional envelope-type nanodevices for gene delivery

Microfluidic devices may be highly beneficial to the rapid fabrication of small quantities of various nonviral vectors with different functionalities, which is indispensable for effective order-made gene therapy. We adapted a microfluidic chip-based approach for fabricating small quantities of nonviral vectors in a short time in preparation for order-made gene therapy applications. This approach permitted us to fabricate multifunctional envelope-type nanodevices (MENDs), composed of a compacted (or condensed) DNA core and a lipid bilayer membrane shell, which are considered as promising nonviral vectors for gene therapy applications. The on-chip fabrication of the MEND was very simple, rapid, convenient, and cost-effective compared with conventional methods. The size of the MEND showed strong dependence on the concentration and flow rate of the reaction precursors and could be controlled to be much smaller than that achievable by conventional methods. This, together with abovementioned merits, makes our microfluidic chip-based approach very attractive for the fabrication of MENDs for effective application to order-made gene therapy.     

Determination of human blood glucose levels using microchip electrophoresis

A high-performance monitoring system for human blood glucose levels was developed using microchip electrophoresis with a plastic chip. The combination of reductive amination as glucose labeling with fluorescent 2-aminoacridone (AMAC) and glucose-borate complex formation realized the highly selective detection of glucose even in a complex matrix such as a blood sample. The migration time of a single peak, observed on an electropherogram of AMAC-labeled plasma, closely resembled that of glucose standard solution. The treatment of plasma with hexokinase or glucokinase for glucose phosphorylation resulted in a peak shift from approximately 145 to 70 s, corresponding to glucose and glucose-6-phosphate, respectively. A double-logarithm plot revealed a linear relationship between glucose concentration and fluorescence intensity in the range of 1-300 microM of glucose (r(2) = 0.9963; p <0.01), and the detection limit was 0.92 microM. Furthermore, blood glucose concentrations estimated from the standard curves of three subjects were compared with results obtained by conventional colorimetric analysis using glucose dehydrogenase. Good correlation was observed between methods according to simple linear regression analysis (p <0.05). The reproducibility of the assay was about 6.3-9.1% (RSD) and the within-days and between-days reproducibility were 1.6-8.4 and 5.2-7.2%, respectively. This system enables us to determine blood glucose with high sensitivity and accuracy, and will be applicable to clinical diagnosis.     

Nanopatterning and fabrication of memory devices from layer-by-layer poly(3,4-ethylenedioxythiophene)-poly(styrene sulfonate) ultrathin films

A write-read-erasable memory device was fabricated on layer-by-layer (LbL) ultrathin films prepared from poly(3,4-ethylenedioxythiophene)-poly(styrene sulfonate) (PEDOT-PSS) and poly(diallyldimethylammonium chloride) (PDDA). By use of current-sensing atomic force microscopy (CS-AFM), nanopatterns were formed by applying a bias voltage between a conductive tip (Pt-coated Si3N4 cantilever) in contact with the polymer film and gold substrate. The dependence of the nanopatterns on film thickness, applied bias, and writing speed was studied. Moreover, the height of the patterns was 3-5 times higher than the original thickness of the films, opening the possibility for three-dimensional nanopatterning. The ability of the patterns to be erased after nanowriting was also investigated. By comparing the I-V characteristics under ambient conditions and under N2 environment, a joule-heating activated, water meniscus-assisted anion doping mechanism for the nanopatterning process was determined. Write-read-erase memory device capability was demonstrated on the nanopatterns.     

Accelerated purification process development of monoclonal antibodies for shortening time to clinic. Design and case study of chromatography processes

For accelerating the purification process development of human monoclonal antibodies (hmAbs) for pharmaceutical drugs, we designed a standardized method for setting the conditions of the purification process, which could be applied to hmAbs for the early phase of pharmaceutical development. The process includes three sequential chromatography steps: Protein A affinity chromatography (AFC), anion-exchange chromatography (AIEC) and cation-exchange chromatography (CIEC), and also includes a low pH virus inactivation step after the AFC step. We predicted the elution pH in the AFC and elution salt concentration in the CIEC from the amino acid sequences of hmAbs, as described in our previous paper. The mobile phase pH in AIEC and the pH for virus inactivation were also predicted based on the amino acid sequence of hmAb. As a case study, six hmAbs (two of IgG(1), two of IgG(2) and two of IgG(4)) were purified with the standardized method. The recovery, purity and clearance of impurities (DNA, host cell proteins (HCP), and Protein A) were examined. All the six hmAbs were purified with high recovery and high clearance of the impurities. Factors affecting the impurities level in the purified products are also discussed.     

Application of a chromatography model with linear gradient elution experimental data to the rapid scale-up in ion-exchange process chromatography of proteins

We applied the model described in our previous paper to the rapid scale-up in the ion exchange chromatography of proteins, in which linear flow velocity, column length and gradient slope were changed. We carried out linear gradient elution experiments, and obtained data for the peak salt concentration and peak width. From these data, the plate height (HETP) was calculated as a function of the mobile phase velocity and iso-resolution curve (the separation time and elution volume relationship for the same resolution) was calculated. The scale-up chromatography conditions were determined by the iso-resolution curve. The scale-up of the linear gradient elution from 5 to 100mL and 2.5L column sizes was performed both by the separation of beta-lactoglobulin A and beta-lactoglobulin B with anion-exchange chromatography and by the purification of a recombinant protein with cation-exchange chromatography. Resolution, recovery and purity were examined in order to verify the proposed method.     

Synthesis of new spiro‐[isothiochromene‐3,5′‐isoxazolidin]‐4(1H)‐ones

A series of seven new 2′,3′,4′‐substituted spiro[isothiochromene‐3,5′‐isoxazolidin]‐4(1H)‐ones ( 7‐13 ) has been prepared in the reaction of benzylidene(phenyl)azane oxide ( 5 ) or benzylidene(methyl)azane oxide ( 6 ) with (3Z)‐3‐(4‐substituted‐benzylidene)‐1H‐isothio‐ chromen‐4(3H)‐one ( 1‐4 ). The reaction occurs by a 1,3‐dipolar cycloaddition mechanism that leads to the regiospecific formation of various spiroisoxazolidines ( 7‐13 ).     

Two new megastigmanes from the leaves of Cucumis sativus

Two new megastigmane, cucumegastigmanes I (1) and II (2), together with a known megastigmane, (+)-dehydrovomifoliol (3), and five other known compounds were isolated from the leaves of Cucumis sativus. The structures of the new compounds were elucidated from spectroscopic analysis and their absolute stereochemistries were determined in detail using the chemical conversion and a modified Mosher's method.     

Topological building blocks of hydrogen bond network in water

Basic three-dimensional units of the network, called fragments, are introduced to characterize the hydrogen bond (HB) network structure of water. Topological differences among normal liquid water, water at low temperature, and water under high pressure are elucidated by their fragment statistics. Water at low temperature has almost defect-free network and is filled with stable fragments with small distortion. It is found that there exists a certain way on how fragments mutually aggregate. Well-formed aggregates heterogeneously constitute very stable network structures. HB network rearrangements occur scarcely inside these aggregated domains but take place in their surface areas. The heterogeneity of HB structure and rearrangement in water is thus explained in terms of the fragment structure and its rearrangements. The fragment analysis thus elucidates the intermediate-range order in water HB network.     

Kinetics of photoinduced e to z isomerization of azobenzene in polystyrene films: thickness, molecular weight and temperature effects

E to Z (trans-->cis) photoisomerization of azobenzene (Az) chromophores tagged to polystyrene (PS) in thin films was studied as functions of thickness, PS molecular weight, and temperature, using the change in absorption at 336 nm arising from the Az E isomer remaining upon ultraviolet light irradiation at 350+/-5 nm. The photoisomerization in solid films exhibited fast and slow modes. The fractional amount of the fast mode (I1) started to increase with decreasing film thickness once the films were thinner than a threshold value. This was explained in terms of a surface layer in which the photoisomerization reaction proceeds quickly, the effect of which becomes more noticeable with decreasing thickness due to a larger surface to volume ratio. The thickness dependence of the I1 fraction was insensitive to the molecular weight of the PS used. The thickness of the surface layer, estimated through a layer model analysis, increased with rising temperature up to 298 K. Interestingly, the surface layer markedly thickened at temperatures at which the molecular motion of PS is on a relatively small scale, namely, at the gamma and beta relaxation temperatures.