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We propose a novel approach to stably immobilize gold nanoparticles (AuNPs) on a plastic substrate and demonstrate that the modified substrate is also capable of immobilizing biomolecules. To immobilize citrate-capped AuNPs, an acrylic substrate was simply dip-coated in a functional polymer solution to decorate the outermost surface with amino groups. Electrostatic interactions between AuNPs and the amino groups immobilized the AuNPs with a high density. The AuNP-modified acrylic substrate was transparent with a red tint. A heat treatment promoted the formation of amide bonds between carboxy groups on the AuNPs and amino groups on the substrate surface. These covalent bonds stabilized the immobilized AuNPs and the resulting substrate was resistant to washing with acid and thiol-containing solutions. The surface density of AuNPs was controlled by the surface density of amino groups on the substrate surface, which was in turn controlled by the dip-coating in the functional polymer solution. We attempted to immobilize functional biomolecules on the AuNPs-functionalized plastic surface by two different approaches. An enzyme (horseradish peroxidase) was successfully immobilized on the AuNPs through amide formation and 5 '-thiolated DNA was also immobilized on the AuNPs through S-Au interactions. These chemistries allow for simultaneous immobilization of two different kinds of biomolecules on a plastic substrate without loss of their functional properties.

α,β-Unsaturated esters were selectively protected in situ in the presence of α,β-unsaturated Weinreb amides using PEt3 and trimethylsilyl trifluoromethanesulfonate (TMSOTf) in toluene under reflux. Diisobutylaluminium hydride (DIBAL-H) reduction of the mixture followed by tetra-n-butylammonium fluoride (TBAF) treatment produced α,β-unsaturated aldehydes in good yields along with the recovered α,β-unsaturated esters.

An organosolv pretreatment consisting of an H2SO4/n-butanol biphasic system was designed to separate lignocellulosic biomass in three distinct phases: a cellulose-rich solid residue, hydrolyzed hem...

Isotactic polypropylene (PP) is one of the most popular plastics. However, the remarkably low surface energy of PP prevents the surface functionalization of PP. We studied the surface functionalization of PP by dip-coating with a maleic anhydride-grafted chlorinated polypropylene (MPO)/methacrylate-based terpolymer mixture. A methacrylate-based terpolymer (PMFP) was synthesized, which contained perfluoroalkyl (Rf)-conjugated monomers and poly(ethylene glycol)-conjugated monomers. Tape-peeling tests revealed that MPO successfully immobilized PMFP on a PP surface, although PMFP was less adhesive to PP. X-ray photoelectron spectroscopy (XPS), contact angle, and protein adsorption measurements revealed that the Rf groups and PEG chains in PMFP were segregated to the outermost surface of the dip-coated layer. The surface segregation of these moieties produced a low-fouling surface on the PP substrate. In addition, we synthesized a terpolymer that contained Rf groups and PEG chains with carboxy groups at their termini (PMFB) and used it to dip-coat a PP substrate. The surface segregation of side chains in PMFB induced the presentation of carboxy groups at the outermost surface, which were used as reactive sites for enzyme immobilization. We studied the surface functionalization of PP by dip-coating with a maleic anhydride-grafted chlorinated polypropylene (MPO) /methacrylate-based terpolymer mixture. A methacrylate-based terpolymer (PMFP) was synthesized, which contained perfluoroalkyl (Rf)-conjugated monomers and poly(ethylene glycol)-conjugated monomers. We found that the presence of MPO aided the adhesion of the terpolymer to the PP surface and that the coated PP surface exhibited low protein-fouling properties. In addition, the dip-coating of a terpolymer containing reactive groups (–COOH) with MPO caused the reactive groups to be presented at an outermost surface.

The biomass-derived source, low-cost and hydrophobicity/oleophilic advantages of activated carbon (AC) were explored for the immobilization of Aspergillus oryzae whole-cells expressing Fusarium heterosporum lipase. The adsorptive influence of AC favored growth of the cells into its porous interfaces with paralleled exterior dense film formation. Increasing AC weights hindered extracellular lipase activity. Cell aggregation of 0.34 ± 0.02 mg/BSP was found to be effective in catalyzing an industrially challenging feedstock (68.77% w/w free fatty acids, 20.48% w/w triglycerides) to 98% fatty acid methyl esters (FAME). In a comparative investigation with polyurethane as matrix, higher trans/esterification facilitation was observed with AC. Benefiting from the oleophilicity of AC; denaturation effect from methanol on the lipases was reduced. Surface characterization with FE-SEM, XPS and FT-IR evidenced effective cell-matrix adhesion and a retention of the AC’s intrinsic properties. The advantageous tribology o...

International audience; Xylitol is a major commodity chemical widely used in both the food and pharmaceutical industries. Although the worldwide demand for xylitol is constantly growing, its industrial production from purified D-xylose involves a costly and polluting catalytic hydrogenation process. Biotechnological production of xylitol from biomass is a promising strategy to establish an environmentally friendly sustainable conversion process. In this study, xylitol was produced from woody Kraft pulp (KP) by using an engineered strain of Saccharomyces cerevisiae (YPH499-XR-BGL-XYL-XYN) expressing cytosolic xylose reductase (XR), along with beta-D-glucosidase (BGL), xylosidase (XYL) and xylanase (XYN) enzymes co-displayed on the cell surface. All these enzymes contributed to the consolidated bioprocessing of KP to xylitol with a yield of 2.3 g L-1 (28% conversion) after 96 hours, along with a significantly reduced amount of commercial enzymes required for pre-treatment (commercial hemicellulase cocktail (CHC), [CHC] = 0.02 g-DW per g). Further improvement of the cell surface display of XYL and XYN was obtained by using a SED1 "SSS" cassette, containing the coding sequences of the SED1 promoter, the SED1 secretion signal, and the SED1 anchoring domain, to generate the improved strain YPH499-XR-BGL-XYLsss-XYNsss. This improved strain showed a significantly enhanced xylitol production capacity reaching a yield of 3.7 g L-1 (44% conversion) after 96 hours. The cellulosic content of KP residues was also significantly increased, from 78% to 87% after 96 hours of fermentation, and nanofibrillation of KP residues was observed by scanning electron microscopy. Pre-treatment and fermentation were successfully performed as a proof of concept to further scale up bio-refinery industrial production of xylitol from lignocellulose.

The development of potentially safe radiosensitizing agents is essential to enhance the treatment outcomes of radioresistant cancers. The titanium peroxide nanoparticle (TiOxNP) was originally produced using the titanium dioxide nanoparticle, and it showed excellent reactive oxygen species (ROS) generation in response to ionizing radiation. Surface coating the TiOxNPs with polyacrylic acid (PAA) showed low toxicity to the living body and excellent radiosensitizing effect on cancer cells. Herein, we evaluated the mechanism of radiosensitization by PAA-TiOxNPs in comparison with gold nanoparticles (AuNPs) which represent high-atomic-number nanoparticles that show a radiosensitizing effect through the emission of secondary electrons. The anticancer effects of both nanoparticles were compared by induction of apoptosis, colony-forming assay, and the inhibition of tumor growth. PAA-TiOxNPs showed a significantly more radiosensitizing effect than that of AuNPs. A comparison of the types and amounts of ROS generated showed that hydrogen peroxide generation by PAA-TiOxNPs was the major factor that contributed to the nanoparticle radiosensitization. Importantly, PAA-TiOxNPs were generally nontoxic to healthy mice and caused no histological abnormalities in the liver, kidney, lung, and heart tissues.

Purpose Titanium dioxide nanoparticles (TiO2 NPs) have been investigated for their role as radiosensitisers for radiation therapy. The study aims to increase the efficiency of these NPs by synthesising them with samarium. Methods Samarium-doped TiO2 NPs (Ti(Sm)O2 NPs) were synthesised using a solvothermal method. Transmission electron microscopy (TEM), X-ray diffraction (XRD), and energy-dispersive X-ray spectroscopy (EDS) were performed for characterising of the Ti(Sm)O2 NPs. The intracellular uptake and cytotoxicity were assessed in vitro using A549 and DU145 cancer cell lines. Furthermore, the effect of dose enhancement and generation of reactive oxygen species (ROS) in response to 6 MV X-rays was evaluated. Additionally, the image contrast properties were investigated using computed tomography (CT) images. Results The synthesised Ti(Sm)O2 NPs were about 13 nm in diameter as determined by TEM. The XRD pattern of Ti(Sm)O2 NPs was consistent with that of anatase-type TiO2. EDS confirmed the presence of samarium in the nanoparticles. At 200 μg/ml concentration, no differences in cellular uptake and cytotoxicity were observed between TiO2 NPs and Ti(Sm)O2 NPs in both A549 and DU145 cells. However, the combination of Ti(Sm)O2 NPs and X-rays elicited higher cytotoxic effect and ROS generation in the cells than that with TiO2 NPs and X-rays. The CT numbers of Ti(Sm)O2 NPs were systematically higher than that of TiO2 NPs. Conclusions The Ti(Sm)O2 NPs increased the dose enhancement of MV X-ray beams than that elicited by TiO2 NPs. Samarium improved the efficiency of TiO2 NPs as potential radiosensitising agent.

Polyacrylic acid (PAA)-modified titanium peroxide nanoparticles (PAA-TiOx NPs) are promising radiosensitizers. PAA-TiOx NPs were synthesized from commercial TiO2 nanoparticles that were modified with PAA and functionalized by H2O2 treatment. To realize practical clinical uses for PAA-TiOx NPs, their tissue distribution and acute toxicity were evaluated using healthy mice and mice bearing tumors derived from xenografted MIAPaCa-2 human pancreatic cancer cells. Healthy mice were injected with PAA-TiOx NPs at 25 mg/kg body weight via the tail vein, and tumor-bearing mice were injected either into the tumor locally or via the tail vein. The concentration of PAA-TiOx NPs in major organs was determined over time using inductively coupled–plasma atomic emission spectrometry. After 1 h, 12% of the PAA-TiOx NP dose had accumulated in the tumor, and 2.8% of the dose remained after 1 week. Such high accumulation could be associated with enhanced permeability and retention effects of the tumor, as PAA-TiOx NPs are composed of inorganic particles and polymers, without tumor-targeting molecules. The liver accumulated the largest proportion of the injected nanoparticles, up to 42% in tumor-bearing mice. Blood biochemical parameters were also investigated after intravenous injection of PAA-TiOx NPs in healthy mice. PAA-TiOx NPs invoked a slight change in various liver-related biochemical parameters, but no liver injury was observed over the practical dose range. In the future, PAA-TiOx NPs should be modified to prevent accumulation in the liver and minimize risk to patients.

We propose a novel approach to prepare affinity membranes using azido-containing surfactants and click chemistry. Porous polymeric membranes were prepared using cellulose acetate via polymer phase separation in the presence of azido-containing surfactants. Thermally induced phase separation and nonsolvent-induced phase separation were used for membrane preparation. The azido groups displayed on the membrane surfaces were conjugated with nitrilotriacetic acid via Huisgen 1,3-dipolar cycloaddition to prepare membranes that displayed affinity toward a hexahistidine-tagged protein. Two types of phase separation successfully produced porous membranes with different microstructures but showed similar separation performances. In place of nitrilotriacetic acid, d -Ala- d -Ala was conjugated to the surface of the azido-functionalized membrane. The membranes functionalized with d -Ala- d -Ala showed high affinity toward vancomycin. The present approach leads to facile surface functionalization of polymeric materials and produces affinity membranes displaying various types of ligands on their surfaces.

Polyacrylic acid-modified titanium peroxide nanoparticles (PAA-TiOx NPs) are promising radiosensitizers that enhance the therapeutic effect of X-ray irradiation after local injection into tumors. However, the mechanism for this reaction has remained unclear with the exception of the involvement of hydrogen peroxide (H2O2), which is released by PAA-TiOx NPs to a liquid phase during dispersion. In the present study, a clonogenic assay was used to compare PAA-TiOx NPs with free H2O2 molecules to investigate the effect exerted on the radiosensitivity of cancer cells in vitro. A cell-free dialysis method revealed that a portion of the H2O2 adsorbed onto the PAA-TiOx NPs during synthesis could be released during a treatment regimen. The H2O2 release lasted for 7 h, which was sufficient for one radiation treatment procedure. For in vitro experiments, cultured human pancreatic cancer cells took up PAA-TiOx NPs in 10 min after administration. Interestingly, when the cells were washed with a buffer after treatment with either a PAA-TiOx NP or H2O2 solution, the intracellular H2O2 levels remained higher with PAA-TiOx NP treatment compared with the H2O2 solution treatment. Furthermore, the effects of subsequent X-ray irradiation corresponded to the intracellular H2O2 levels. These results indicate that PAA-TiOx NPs are efficient carriers of H2O2 into cancer cells and thus enhance the radiosensitivity.

An induction of polyacrylic acid-modified titanium dioxide with hydrogen peroxide nanoparticles (PAA-TiO2/H2O2 NPs) to a tumor exerted a therapeutic enhancement of X-ray irradiation in our previous study. To understand the mechanism of the radiosensitizing effect of PAA-TiO2/H2O2 NPs, analytical observations that included DLS, FE-SEM, FT-IR, XAFS, and Raman spectrometry were performed. In addition, highly reactive oxygen species (hROS) which PAA-TiO2/H2O2 NPs produced with X-ray irradiation were quantified by using a chemiluminescence method and a EPR spin-trapping method. We found that PAA-TiO2/H2O2 NPs have almost the same characteristics as PAA-TiO2. Surprisingly, there were no significant differences in hROS generation. However, the existence of H2O2 was confirmed in PAA-TiO2/H2O2 NPs, because spontaneous hROS production was observed w/o X-ray irradiation. In addition, PAA-TiO2/H2O2 NPs had a curious characteristic whereby they absorbed H2O2 molecules and released them gradually into a liquid phase. Based on these results, the H2O2 was continuously released from PAA-TiO2/H2O2 NPs, and then released H2O2 assumed to be functioned indirectly as a radiosensitizing factor.

Selective one-pot alkylation or allylation of ketones in the presence of α,β-unsaturated ketones (enones) were accomplished by using our in situ protection methodology. Enones selectively react with a combination of PPh3 and silyl triflates in the presence of ketones to produce phosphonium silyl enol ethers, which act as protective groups for the enones during the alkylation or allylation of ketones, and can be easily deprotected to regenerate the parent enones on work-up.

Highly discriminative transformation of α,β-unsaturated esters in the presence of enones using two types of phosphonium salts, and their application to the synthesis of oxacyclic compounds in six steps in one pot have been achieved.

Xylitol is a highly valuable commodity chemical used extensively in the food and pharmaceutical industries. The production of xylitol from d-xylose involves a costly and polluting catalytic hydrogenation process. Biotechnological production from lignocellulosic biomass by micro-organisms like yeasts is a promising option. In this study, xylitol is produced from lignocellulosic biomass by a recombinant strain of Saccharomyces cerevisiae (S. cerevisiae) (YPH499-SsXR-AaBGL) expressing cytosolic xylose reductase (Scheffersomyces stipitis xylose reductase [SsXR]), along with a β-d-glucosidase (Aspergillus aculeatus β-glucosidase 1 [AaBGL]) displayed on the cell surface. The simultaneous cofermentation of cellobiose/xylose by this strain leads to an ≈2.5-fold increase in Yxylitol/xylose (=0.54) compared to the use of a glucose/xylose mixture as a substrate. Further improvement in the xylose uptake by the cell is achieved by a broad evaluation of several homologous and heterologous transporters. Homologous maltose transporter (ScMAL11) shows the best performance in xylose transport and is used to generate the strain YPH499-XR-ScMAL11-BGL with a significantly improved xylitol production capacity from cellobiose/xylose coutilization. This report constitutes a promising proof of concept to further scale up the biorefinery industrial production of xylitol from lignocellulose by combining cell surface and metabolic engineering in S. cerevisiae.

Background Biological applications of nanoparticles are rapidly increasing, which introduces new possibilities to improve the efficacy of radiotherapy. Here, we synthesized titanium peroxide nanoparticles (TiOxNPs) and investigated their efficacy as novel agents that can potently enhance the effects of radiation in the treatment of pancreatic cancer. Methods TiOxNPs and polyacrylic acid-modified TiOxNPs (PAA-TiOxNPs) were synthesized from anatase-type titanium dioxide nanoparticles (TiO2NPs). The size and morphology of the PAA-TiOxNPs was evaluated using transmission electron microscopy and dynamic light scattering. The crystalline structures of the TiO2NPs and PAA-TiOxNPs with and without X-ray irradiation were analyzed using X-ray absorption. The ability of TiOxNPs and PAA-TiOxNPs to produce reactive oxygen species in response to X-ray irradiation was evaluated in a cell-free system and confirmed by flow cytometric analysis in vitro. DNA damage after X-ray exposure with or without PAA-TiOxNPs was assessed by immunohistochemical analysis of γ-H2AX foci formation in vitro and in vivo. Cytotoxicity was evaluated by a colony forming assay in vitro. Xenografts were prepared using human pancreatic cancer MIAPaCa-2 cells and used to evaluate the inhibition of tumor growth caused by X-ray exposure, PAA-TiOxNPs, and the combination of the two. Results The core structures of the PAA-TiOxNPs were found to be of the anatase type. The TiOxNPs and PAA-TiOxNPs showed a distinct ability to produce hydroxyl radicals in response to X-ray irradiation in a dose- and concentration-dependent manner, whereas the TiO2NPs did not. At the highest concentration of TiOxNPs, the amount of hydroxyl radicals increased by >8.5-fold following treatment with 30 Gy of radiation. The absorption of PAA-TiOxNPs enhanced DNA damage and resulted in higher cytotoxicity in response to X-ray irradiation in vitro. The combination of the PAA-TiOxNPs and X-ray irradiation induced significantly stronger tumor growth inhibition compared to treatment with either PAA-TiOxNPs or X-ray alone (p

Purpose: Pancreatic cancer is a highly lethal disease and notoriously resistant to many types of cytotoxic chemotherapy and radiotherapy. A novel strategy should be explored for these radioresistant tumors. We originally synthesized a titanium peroxide nanoparticle (TiOxNP) which had a distinct ability to produce reactive oxygen species (ROS) upon X-ray irradiation (IR). Here, the characterization and efficacy of TiOxNP as radiosensitizer for pancreatic cancer therapy were investigated. Methods: TiOxNPs were synthesized from anatase titanium oxide nanoparticles (TiO2NPs) by H2O2 processing and coated by polyacrylic acid. The ability of TiOxNPs to enhance ROS generation upon X-ray irradiation (IR) was tested by using 3′-(p-Aminophenyl) fluorescein and several antioxidants. A xenograft mouse model using the human pancreatic cancer cell line MIAPaCa-2 was used to evaluate the cytotoxic effects of a combination of TiOxNPs and IR in vivo. MIAPaCa-2 cells were injected subcutaneously into the hind legs of immunodeficient BALB/c mice. Seven days after injection, the tumor was treated with a TiOxNP suspension injected directly into the tumor and application of 5 Gy of IR. Tumor size and health of the mice were monitored for 43 days after the treatment. Induction of apoptosis was evaluated by immunohistochemistry using terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling staining. We also characterized TiOxNPs by ESR and XAFS analyses. Results: ROS generation was enhanced dramatically by TiOxNPs in a concentration- and radiation dose- dependent manner compared to enhancement by TiO2NPs. At the highest concentration of TiOxNPs, ROS generation upon an irradiation dose of 30 Gy was enhanced by a factor of more than 10. Also, ROS generated by TiOxNPs upon IR were scavenged by vitamin C or glutathione. Growth inhibition of the tumor in the group receiving both TiOxNP and IR was significantly greater than single treatment subgroups (P < 0.05). The number of apoptotic cells was also significantly higher in the combination group (P < 0.05). Histologically, large pools of TiOxNPs around tumor cells and small amounts inside the tumor cells were observed. No mice died within a 43-day observation period or showed any apparent body weight loss. In the ESR spectra, some signals were observed for TiOxNPs. The spectral shapes of Ti K-edge X-ray absorption near edge structure, spectra were the same among the mineral anatase, TiO2NPs and TiOxNPs. Conclusions: TiOxNPs induced remarkable ROS production upon IR. They were safe and effective in a xenograft mouse model using engrafted human pancreatic cancer cells. Further studies would be necessary to elucidate a mechanism of radical generation and long-term toxicity, however our study shows that TiOxNPs are promising radiosensitizers for application in pancreatic cancer therapy. Citation Format: Masao Nakayama, Ryohei Sasaki, Toru Mukohara, Chiaki Ogino, Kenta Morita, Mitsuo Umetsu, Satoshi Ohara, Kazuyoshi Sato, Chiya Numako, Seiichi Takami, Akihiko Kondo. Titanium peroxide nanoparticles enhance antitumor efficacy through reactive oxygen species in pancreatic cancer radiation therapy. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3337. doi:10.1158/1538-7445.AM2015-3337