論文 - 金澤　秀子

Temperature-modulated interactions between thermoresponsive strong cationic copolymer-brush-grafted silica beads and biomolecules

Nagase K., Suzuki S., Kanazawa H.

Heliyon 10 （ 15 ） e34668 2024年08月

ISSN  24058440

　概要を見る

Thermoresponsive polymer brushes have attracted considerable research attention owing to their unique properties. Herein, we developed silica beads grafted with poly(N-isopropylacrylamide (NIPAAm)-co-3-acrylamidopropyl trimethylammonium chloride (APTAC)-co-tert-butyl acrylamide (tBAAm) and P(NIPAAm-co-APTAC-co-n-butyl methacrylate(nBMA)) brushes. The carbon, hydrogen, and nitrogen elemental analysis of the copolymer-grated silica beads revealed the presence of a large amount of the grafted copolymer on the silica beads. The electrostatic and hydrophobic interactions between biomolecules and prepared copolymer brushes were analyzed by observing their elution behaviors via high-performance liquid chromatography using the copolymer-brush-modified beads as the stationary phase. Adenosine nucleotides were retained in the bead-packed columns, which was attributed to the electrostatic interaction between the copolymers and adenosine nucleotides. Insulin was adsorbed on the copolymer brushes at high temperatures, which was attributed to its electrostatic and hydrophobic interactions with the copolymer. Similar adsorption behavior was observed in case of albumin. Further, at a low concentration of the phosphate buffer solution, albumin was adsorbed onto the copolymer brushes even at relatively low temperatures owing to its enhanced electrostatic interaction with the copolymer. These results indicated that the developed thermoresponsive strong cationic copolymer brushes can interact with peptides and proteins through a combination of electrostatic and temperature-modulated hydrophobic interactions. Thus, the developed copolymer brushes exhibits substantial potential for application in chromatographic matrices for the analysis and purification of peptides and proteins.

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Thermoresponsive mixed polymer brush to effectively control the adhesion and separation of stem cells by altering temperature

Nagase K., Wakayama H., Matsuda J., Kojima N., Kanazawa H.

Materials Today Bio 20   100627 2023年06月

　概要を見る

During the last few decades, thermoresponsive materials for modulating cell adhesion have been investigated for the application of tissue engineering. In this study, we developed thermoresponsive mixed polymer brushes consisting of poly(N-isopropylacrylamide) (PNIPAAm) and poly(N,N-dimethylaminopropylacrylamide) (PDMAPAAm). The mixed polymer brushes were prepared on a glass substrate via the reversible addition-fragmentation chain transfer polymerization of DMAPAAm and subsequent atom transfer radical polymerization of NIPAAm. The mixed polymer brushes grafted to glass exhibited increased cationic properties by increasing the grafted PDMAPAAm length. The shrinking and extension of PNIPAAm exposed and concealed PDMAPAAm, respectively, indicating that the surface cationic properties can be controlled by changing the temperature. At 37 ​°C, the prepared mixed polymer brushes enhanced cell adhesion through their electrostatic interactions with cells. They also exhibited various thermoresponsive adhesion and detachment properties using various types of cells, such as mesenchymal stem cells. Temperature-controlled cell adhesion and detachment behavior differed between cell types. Using the prepared mixed polymer brush, we separated MSCs from adipocytes and HeLa cells by simply changing the temperature. Thus, the thermoresponsive mixed polymer brushes may be used to separate mesenchymal stem cells from their differentiated or contaminant cells by altering the temperature.

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Real-time in situ X-ray micro-computed tomography study of the effect of impurities on the crystallization of amorphous nifedipine

Amano Y., Misawa T., Miyazaki T., Ando D., Koide T., Izutsu K.i., Kanazawa H., Hanaoka K., Yamamoto E.

Journal of Pharmaceutical and Biomedical Analysis 226   115248 2023年03月

ISSN  07317085

　概要を見る

Controlling the physical stability of noncrystalline active pharmaceutical ingredients remains a major challenge in the development of amorphous formulations such as amorphous solid-dispersion (ASD) formulations. To establish new evaluation and formulation strategies, the spatial distribution of the crystal phase in bulk amorphous nifedipine (NFD) was investigated as a model. The crystallization of amorphous NFD and the effect of a deliberately added impurity were investigated using powder X-ray diffraction (PXRD), differential scanning calorimetry and real-time in situ X-ray micro-computed tomography (X-ray CT). The stability data of amorphous samples, i.e., NFD and a mixture of NFD with an oxidative degradation product of NFD, impurity A (Imp A), at a weight ratio of 90:10, presented as percent amorphous remaining, suggests that Imp A accelerates the bulk crystal growth of NFD. Real-time in situ X-ray CT results showed surface-enhanced crystal growth and cavity formation in solid NFD samples. Moreover, the crystals were heterogeneous in density. These results suggest that Imp A affects the physical stability of the amorphous NFD. X-ray CT equipped with a heating unit can aid in-situ evaluation and assessment of physicochemical properties and physical stability of amorphous samples and formulations.

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Thermoresponsive bio-affinity interfaces for temperature-modulated selective capture and release of targeted exosomes

Nagase K., Yamazaki K., Maekawa Y., Kanazawa H.

Materials Today Bio 18   100521 2023年02月

　概要を見る

The existing methods for exosome isolation, such as ultracentrifugation, size exclusion, and affinity separation, suffer from some limitations. Herein, we aimed to develop temperature-modulated exosome-capturing materials using thermoresponsive polymers and peptides with affinity for exosomes. Poly(2-hydroxyethyl methacrylate-co-propargyl acrylate)-b-poly(N-isopropylacrylamide) (P(HEMA-co-PgA)-b-PNIPAAm) was grafted on silica beads via a two-step process of activator regenerated by electron transfer atom transfer radical polymerization. Peptides with affinity for exosomes were conjugated to the propargyl group of the bottom P(HEMA-co-PgA) segment of the copolymer via a click reaction. The prepared copolymer-grafted beads were characterized by elemental analysis, X-ray photoelectron spectroscopy, scanning electron microscopy, transmission electron microscopy, gel permeation chromatography, and the turbidity of the polymer solution. Results indicated that the copolymer and peptide were successfully modified on the silica beads. Exosomes from SK-BR-3 ​cells, a human breast cancer cell line, were selectively captured on the prepared beads at 37 ​°C, as the upper PNIPAAm segment shrank and the affinity between the peptide and exosome was enhanced. Upon lowering the temperature to 4 ​°C, the captured exosomes were released from the copolymer brush because of the extension of the PNIPAAm segment that reduced the affinity between peptides and exosomes. These findings demonstrated that the prepared copolymer brush-grafted silica beads can capture and release targeted exosomes via temperature modulation. Taken together, the developed copolymer brush-grafted silica beads would be useful for the separation of exosomes using simple procedures such as temperature modulation.

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Two-dimensional temperature-responsive chromatography using a poly(N-isopropylacrylamide) brush-modified stationary phase for effective therapeutic drug monitoring

Nagase K., Inoue S., Inoue M., Kanazawa H.

Scientific Reports 12 （ 1 ） 2653 2022年12月

　概要を見る

Therapeutic drug monitoring (TDM) is an effective pharmacological approach for controlling drug concentration in a patient’s serum. Herein, a new two-dimensional chromatography system was developed using two poly(N-isopropylacrylamide) (PNIPAAm)-modified bead-packed columns for effective and safe drug monitoring. PNIPAAm-modified silica beads were prepared as packing materials using atom transfer radical polymerization of NIPAAm. The increase in the retention times of the drugs requiring TDM with increasing temperature, was attributed to enhanced hydrophobic interactions at elevated temperatures. The drugs and serum proteins were separated on the prepared column at 40 °C using an all-aqueous mobile phase. Differences in the hydrophobic interactions accounted for the elution of the serum proteins and drugs at short and long retention times, respectively, and a primary column was employed to separate the serum proteins and drugs. After eluting the serum proteins from the column, the drug was introduced into the secondary column, leading to a peak of its purified form and enabling determination of the drug concentration. Two-dimensional temperature-responsive chromatography can benefit TDM by allowing the drug concentration in the serum to be measured in all-aqueous mobile phases without sample preparation.

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Temperature-responsive mixed-mode column for the modulation of multiple interactions

Nagase K., Matsumoto K., Kanazawa H.

Scientific Reports 12 （ 1 ） 4434 2022年12月

　概要を見る

In this study, mixed-mode chromatography columns have been investigated using multiple analyte interactions. A mixed-mode chromatography column was developed using poly(N-isopropylacrylamide) (PNIPAAm) brush-modified silica beads and poly(3-acrylamidopropyl trimethylammonium chloride) (PAPTAC) brush-modified silica beads. PNIPAAm brush-modified silica beads and PAPTAC brush-modified silica beads were prepared by atom transfer radical polymerization. The beads were then packed into a stainless-steel column in arbitrary compositions. The elution studies evaluated the column performance on hydrophobic, electrostatic, and therapeutic drug samples using steroids, adenosine nucleotide, and antiepileptic drugs as analytes, respectively. Steroids exhibited an increased retention time when the column temperature was increased. The retention of adenosine nucleotides increased with the increasing composition of the PAPTAC-modified beads in the column. The antiepileptic drugs were separated using the prepared mixed-mode columns. An effective separation of antiepileptic drugs was observed on a 10:1 PNIPAAm:PAPTAC column because the balance between the hydrophobic and electrostatic interactions with antiepileptic drugs was optimized for the bead composition. Oligonucleotides were also separated using mixed-mode columns through multiple hydrophobic and electrostatic interactions. These results demonstrate that the developed mixed-mode column can modulate multiple hydrophobic and electrostatic interactions by changing the column temperature and composition of the packed PNIPAAm and PAPTAC beads.

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Chromatography columns packed with thermoresponsive-cationic-polymer-modified beads for therapeutic drug monitoring

Nagase K., Takagi H., Nakada H., Ishikawa H., Nagata Y., Aomori T., Kanazawa H.

Scientific Reports 12 （ 1 ） 12847 2022年12月

　概要を見る

Therapeutic drug monitoring, which is used to determine appropriate drug doses, is critical in pharmacological therapy. In this study, we developed thermoresponsive chromatography columns with various cationic properties for effective therapeutic drug monitoring. Thermoresponsive cationic copolymer poly(N-isopropylacrylamide-co–n-butyl methacrylate-co–N,N-dimethylaminopropyl acrylamide) (P(NIPAAm-co-BMA-co-DMAPAAm))-modified silica beads, which were used as the chromatographic stationary phase, were prepared by modifying the radical initiator of the silica beads, followed by radical polymerization. Characterization of the prepared silica beads demonstrated that thermoresponsive polymers with various cationic properties successfully modified the beads. The elution behavior of several steroids in the prepared bead-packed columns at various temperatures indicated that the optimal column operating temperature was 30 °C. Appropriate measurement conditions for 13 drugs were investigated by varying the cationic properties of the columns and the pH of the mobile phase. Drug concentrations in serum samples were determined using the developed columns and mobile phases with a suitable pH. Voriconazole concentrations in human serum samples were determined using the developed columns with all-aqueous mobile phases. We anticipate that the developed chromatography columns can be used for therapeutic drug monitoring because drug concentrations can be measured using all-aqueous mobile phases that are suitable in clinical settings.

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Actin cable formation and epidermis–dermis positional relationship during complete skin regeneration

Takaya K., Okabe K., Ishigami A., Imbe Y., Kanazawa H., Sakai S., Aramaki-Hattori N., Kishi K.

Scientific Reports 12 （ 1 ） 15913 2022年12月

　概要を見る

Up to a certain developmental stage, a fetus can completely regenerate wounds in the skin. To clarify the mechanism of fetal skin regeneration, identifying when the skin switches from fetal-type wound regeneration to adult-type wound repair is necessary. We hypothesized that this switch occurs at several time points and that complete skin regeneration requires epidermal–dermal interactions and the formation of actin cables. We compared normal skin and wound morphology at each developmental stage. We examined two parameters: epidermal texture and dermal structure. We found that the three-dimensional structure of the skin was completely regenerated in full-thickness skin incisions made before embryonic day (E) 13. However, the skin texture did not regenerate in wounds made after E14. We also found that the dermal structure regenerates up to E16, but wounds created after E17 heal as scars with dermal fibrosis. By controlling the activity of AMP-activated protein kinase and altering actin cable formation, we could regulate scar formation in utero. These findings may contribute to therapies that allow complete skin regeneration without scarring.

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A thermoresponsive cationic block copolymer brush-grafted silica bead interface for temperature-modulated separation of adipose-derived stem cells

Nagase K., Okada A., Matsuda J., Ichikawa D., Hattori Y., Kanazawa H.

Colloids and Surfaces B: Biointerfaces 220   112928 2022年12月

ISSN  09277765

　概要を見る

Adipose-derived mesenchymal stem cells (ADSCs) have beneficial effects in cell transplantation therapy; these cells are collected from adipose tissue using low-invasive methods. However, to prepare ADSCs for cell therapy, a cell separation method that neither involves modification of the cell surface nor causes loss of cell activity is needed. Here, we aimed to develop ADSC separation columns using thermoresponsive cationic block copolymer brush-grafted beads as packing materials. The block copolymer brush was formed by a bottom cationic segment, poly(N,N-dimethylaminopropylacrylamide) (PDMAPAAm), and an upper thermoresponsive segment, poly(N-isopropylacrylamide) (PNIPAAm), and was grafted in two atom transfer radical polymerization reactions. The copolymer brush-grafted silica beads were packed into a column. An ADSC suspension was introduced into the columns at 37 °C and adsorbed on the copolymer brush-modified beads through electrostatic and hydrophobic interactions with the PDMAPAAm and PNIPAAm segments, respectively. The adsorbed ADSCs eluted from the column by lowering the temperature to 4 °C. In contrast, most Jurkat and vascular endothelial cells eluted at 37 °C, because of the relatively weaker electrostatic interactions with the block copolymer brush compared to ADSCs. Using the prepared column, a mixture of ADSCs and Jurkat cells was separated by changing the column temperature. The recovered ADSCs exhibited cell activity. The developed cell separation column may be useful for isolating ADSCs without cell surface modification, while maintaining cell activity.

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Temperature-modulated antibody drug separation using thermoresponsive mixed polymer brush-modified stationary phase

Nagase K., Ishii S., Takeuchi A., Kanazawa H.

Separation and Purification Technology 299 2022年10月

ISSN  13835866

　概要を見る

Recently, antibody drugs have been investigated for medical treatments of intractable diseases. An effective antibody drug separation method is in demand for production of such drugs. We developed an antibody drug separation material using a mixed polymer brush composed of poly(N-isopropylacrylamide) (PNIPAAm), which is a thermoresponsive polymer, and poly(4-vinylpyridine) (P4VP), which is an antibody-affinity polymer, for temperature modulation and adsorption of the antibody drug rituximab that results in its separation. A mixed polymer brush was prepared on silica bead surfaces by radical polymerization to modify P4VP, followed by atom transfer radical polymerization to modify PNIPAAm. The prepared mixed polymer brush was characterized by CHN elemental analysis, chloride analysis, nitrogen adsorption, thermogravimetric analysis, differential thermal analysis, differential scanning calorimetry, Fourier-transform infrared spectroscopy, X-ray photoelectron spectroscopy, scanning electron microscopy, transmission electron microscopy, and zeta potential measurement. Temperature-modulated adsorption of rituximab on the mixed polymer brush was achieved by concealing and exposing P4VP through the extension and shrinkage of PNIPAAm in the brush, and was investigated using mixed polymer brush-modified silica beads as chromatographic packing materials. By utilizing the specific properties of mixed polymer brushes, rituximab and bovine serum albumin could be separated by simply changing the column temperature. Thus, the mixed polymer brush-modified beads prepared in this study may be useful for purifying rituximab by changing the temperature.

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Thermoresponsive block copolymer brush for temperature-modulated hepatocyte separation

Nagase K., Kojima N., Goto M., Akaike T., Kanazawa H.

Journal of Materials Chemistry B 10 （ 42 ） 8629 - 8641 2022年08月

ISSN  2050750X

　概要を見る

Hepatic tissue engineering may be an effective approach for the treatment of liver disease; however, its practical application requires hepatic cell separation technologies that do not involve cell surface modification and maintain cell activity. In this study, we developed hepatocyte cell separation materials using a thermoresponsive polymer and a polymer with high affinity to hepatocytes. A block copolymer of poly(N-p-vinylbenzyl-O-β-d-galactopyranosyl-(1→4)-d-gluconamide) (PVLA) and poly(N-isopropylacrylamide) (PNIPAAm) [PVLA-b-PNIPAAm] was prepared through two steps of atom transfer radical polymerization. On the prepared PVLA-b-PNIPAAm brush, HepG2 cells (model hepatocytes) adhered at 37 °C and detached at 20 °C, attributed to the temperature-modulated affinity between PVLA and HepG2. Cells from the immortalized human hepatic stellate cell line (TWNT-1) did not adhere to the copolymer brush, and RAW264.7 cells (mouse macrophage; model Kupffer cells) adhered to the copolymer brush, regardless of temperature. Using the difference in cell adhesion properties on the copolymer brush, temperature-modulated cell separation was successfully demonstrated. A mixture of HepG2, RAW264.7, and TWNT-1 cells was seeded on the copolymer brush at 37 °C for adherence. By reducing the temperature to 20 °C, adhered HepG2 cells were selectively recovered with a purity of approximately 85% and normal activity. In addition, induced pluripotent stem (iPS) cell-derived hepatocytes adhered on the PVLA-b-PNIPAAm brush at 37 °C and detached from the copolymer brush at 20 °C, whereas the undifferentiated iPS cells did not adhere, indicating that the prepared PVLA-b-PNIPAAm brush could be utilized to separate hepatocyte differentiated and undifferentiated cells. These results indicated that the newly developed PVLA-b-PNIPAAm brush can separate hepatic cells from contaminant cells by temperature modulation, without affecting cell activity or modifying the cell surface. Thus, the copolymer brush is expected to be a useful separation tool for cell therapy and tissue engineering using hepatocytes.

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Role of Wnt Signaling in Mouse Fetal Skin Wound Healing

Takaya K., Sunohara A., Aramaki-hattori N., Sakai S., Okabe K., Kanazawa H., Asou T., Kishi K.

Biomedicines 10 （ 7 ）  2022年07月

ISSN  2227-9059

　概要を見る

Wnt proteins secrete glycoproteins that are involved in various cellular processes to main-tain homeostasis during development and adulthood. However, the expression and role of Wnt in wound healing have not been fully documented. Our previous studies have shown that, in an early-stage mouse fetus, no scarring occurred after cutaneous wounding, and complete regeneration was achieved. In this study, the expression and localization of Wnt proteins in a mouse fetal-wound-healing model and their associations with scar formation were analyzed. Wnt-related molecules were detected by in-situ hybridization, immunostaining, and real-time polymerase chain reaction. The results showed altered expression of Wnt-related molecules during the wound-healing process. Moreover, scar formation was suppressed by Wnt inhibitors, suggesting that Wnt signaling may be involved in wound healing and scar formation. Thus, regulation of Wnt signaling may be a possible mechanism to control scar formation.

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Viral vector purification with thermoresponsive-anionic mixed polymer brush modified beads-packed column

Nagase K., Kitazawa S., Kogure T., Yamada S., Katayama K., Kanazawa H.

Separation and Purification Technology 286 2022年04月

ISSN  13835866

　概要を見る

Viral vectors have attracted attention as a potential new therapeutic modality for gene therapy. In this study, we developed a temperature-modulated viral vector purification column using a mixed polymer brush composed of thermoresponsive and anionic polymers as packing material ligands. The mixed polymer brushes were modified on silica beads using a combination of reversible addition − fragmentation chain transfer (RAFT) polymerization of 2-acrylamido-2-methylpropanesulfonic acid (AMPS) and subsequent atom transfer radical polymerization (ATRP) of N-isopropylacrylamide (NIPAAm). The temperature-modulated zeta potential change in the prepared mixed polymer brush was attributed to the PNIPAAm shrinking and exposing of PAMPS. The prepared mixed polymer brush modified beads were used as packing materials, and the elution behavior of the adeno associated virus type 2 (AAV2) vector was observed. The AAV2 vector was adsorbed on the mixed polymer brush by electrostatic and hydrophobic interactions at 40 °C. By reducing the temperature to 5 °C, adsorbed AAV2 vector on the mixed polymer brush was desorbed and eluted from the column due to lowering the electrostatic and hydrophobic interactions between the AAV2 vector and mixed polymer brush. The AAV2 vector was separated from bovine serum proteins as a contaminant using the column. The ability to infect cells was maintained by the recovered AAV2 vectors from the column. Thus, the developed column would be beneficial for the simple purification of viral vectors.

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Temperature responsive chromatography for therapeutic drug monitoring with an aqueous mobile phase

Nagase K., Nishiyama T., Inoue M., Kanazawa H.

Scientific Reports 11 （ 1 ） 23508 2021年12月

　概要を見る

Therapeutic drug monitoring is a key technology for effective pharmacological treatment. In the present study, a temperature-responsive chromatography column was developed for safe and simple therapeutic drug monitoring without the use of organic solvents. Poly(N-isopropylacrylamide) (PNIPAAm) hydrogel-modified silica beads were prepared via a condensation reaction and radical polymerization. The temperature-dependent elution behavior of the drugs was observed using a PNIPAAm-modified silica-bead packed column and an all-aqueous mobile phase. Sharp peaks with reproducible retention times were observed at temperatures of 30 °C or 40 °C because the PNIPAAm hydrogel on the silica beads shrinks at these temperatures, limiting drug diffusion into the PNIPAAm hydrogel layer. The elution behavior of the sample from the prepared column was examined using a mixture of serum and model drugs. The serum and drugs were separated on the column at 30 °C or 40 °C, and the concentration of the eluted drug was obtained using the calibration curve. The results show that the prepared chromatography column would be useful for therapeutic drug monitoring because the drug concentration in serum can be measured without using organic solvents in the mobile phase and without any need for sample preparation.

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Thermally-modulated cell separation columns using a thermoresponsive block copolymer brush as a packing material for the purification of mesenchymal stem cells

Nagase K., Edatsune G., Nagata Y., Matsuda J., Ichikawa D., Yamada S., Hattori Y., Kanazawa H.

Biomaterials Science 9 （ 21 ） 7054 - 7064 2021年11月

ISSN  20474830

　概要を見る

Cell therapy using mesenchymal stem cells (MSCs) is used as effective regenerative treatment. Cell therapy requires effective cell separation without cell modification and cellular activity reduction. In this study, we developed a temperature-modulated mesenchymal stem cell separation column. A temperature-responsive cationic block copolymer, poly(N,N-dimethylaminopropylacrylamide)-b-poly(N-isopropylacrylamide)(PDMAPAAm-b-PNIPAAm) brush with various cationic copolymer compositions, was grafted onto silica beads via two-step atom transfer radical polymerization. Using the packed beads, the elution behavior of the MSCs was observed. At 37 °C, the MSCs were adsorbed onto the column via both hydrophobic and electrostatic interactions with the PNIPAAm and PDMAPAAm segments of the copolymer brush, respectively. By reducing the temperature to 4 °C, the adsorbed MSCs were eluted from the column by reducing the hydrophobic and electrostatic interactions attributed to the hydration and extension of the PNIPAAm segment of the block copolymer brush. From the temperature-modulated adsorption and elution behavior of MSCs, a suitable DMAPAAm composition of the block copolymer brush was determined. Using the column, a mixture of MSC and BM-CD34+ cells was separated by simply changing the column temperature. The column was used to purify the MSCs, with purities of 78.2%, via a temperature change from 37 °C to 4 °C. Additionally, the cellular activity of the MSCs was retained throughout the column separation step. Overall, the obtained results show that the developed column is useful for MSC separation without cell modification and cellular activity reduction. This journal is

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Anion species-triggered antibody separation system utilizing a thermo-responsive polymer column under optimized constant temperature

Nomoto D., Nagase K., Nakamura Y., Kanazawa H., Citterio D., Hiruta Y.

Colloids and Surfaces B: Biointerfaces 205   111890 2021年09月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  0927-7765

　概要を見る

Although the field of antibody drugs has grown larger, the antibody production still faces several challenges. Effective antibody purification is required, but the conventional purification method for antibodies is cost intensive and often causes aggregation problems, indicating the need for new alternative antibody purification methods. In the present study, a constant temperature antibody purification system for use with a thermo-responsive polymer column was developed based on switching of anion species in eluents. By adjusting the temperature for each antibody, the developed column enabled separation of the therapeutic monoclonal antibodies, rituximab and trastuzumab, from contaminants without changing salt concentration or pH of the eluents. The thermo-responsive hydrogel-modified column packing material was synthesized by introducing n-butyl methacrylate, acrylic acid, N,N′-methylenebisacrylamide and N-isopropylacrylamide to the surface of silica beads with an initiator by a graft-from approach. Elution behavior of antibodies with three types of anions, such as citrate, phosphate, and chloride were tested under three different temperature conditions. It was demonstrated that the thermo-responsive hydrogel grafted column showed a switchable antibody retention behavior at constant temperature and salt concentration, with antibody adsorption by NaCl eluent and desorption by citric acid buffer eluent.

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Temperature-responsive spin column for sample preparation using an all-aqueous eluent

Nagase K., Ishizawa Y., Inoue M., Kokubun M., Yamada S., Kanazawa H.

Analytica Chimica Acta 1179   338806 2021年09月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  0003-2670

　概要を見る

We present a temperature-responsive spin column using an all-aqueous eluent. The method is intended as a simple sample preparation method for protein removal from serum, which is required for serum drug analysis. As packing materials for the spin column, we prepared two types of silica beads via surface-initiated radical polymerization. The large beads (diameter, 40–63 μm) were grafted with a temperature-responsive cationic copolymer, poly(N-isopropylacrylamide-co-N,N-dimethylaminopropyl acrylamide-co-n-butyl methacrylate) (P(NIPAAm-co-DMAPAAm-co-BMA)), and the small beads (diameter, 5 μm) were grafted with a temperature-responsive hydrophobic copolymer, P(NIPAAm-co-BMA). The beads were packed into the spin column as a double layer: P(NIPAAm-co-BMA) silica beads on the bottom and P(NIPAAm-co-DMAPAAm-co-BMA) silica beads on the top. The sample purification efficacy of the prepared spin column was evaluated on a model sample analyte (the antifungal drug voriconazole mixed with blood serum proteins). At 40 °C, the serum proteins and voriconazole were adsorbed on the prepared spin column via hydrophobic and electrostatic interactions. When the temperature was decreased to 4 °C, the adsorbed voriconazole was eluted from the column with the pure water eluent, while the serum proteins remained in the column. This temperature-responsive spin column realizes sample preparation simply by changing the temperature.

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Discrimination of ranitidine hydrochloride crystals using X-ray micro-computed tomography for the evaluation of three-dimensional spatial distribution in solid dosage forms

Yamamoto E., Takeda Y., Ando D., Koide T., Amano Y., Miyazaki S., Miyazaki T., Izutsu K.i., Kanazawa H., Goda Y.

International Journal of Pharmaceutics 605   120834 2021年08月

ISSN  03785173

　概要を見る

A non-destructive discrimination method for crystals in solid dosage drug forms was first developed using a combination of Raman spectroscopy and X-ray micro-computed tomography (X-ray CT). Identification of the crystal form of an active pharmaceutical ingredient (API) at the appropriate pharmaceutical dosage is crucial, as the crystal form is a determinant of the quality and performance of the final formulation. To develop a non-destructive analytical methodology for the discrimination of solid API crystals in a solid dosage form, we utilized a combination of Raman spectroscopy and X-ray CT to differentiate between ranitidine crystal polymorphs (forms 1 and 2) in tablet formulations containing three excipients. The difference in electron density correlated with the true density between ranitidine polymorphs, thereby enabling the discrimination of crystal forms and visualization of their three-dimensional spatial localization inside the tablets through X-ray CT imaging. Furthermore, X-ray CT imaging revealed that the crystal particles were of varying densities, sizes, and shapes within the same batch. These findings suggest that X-ray CT is not only an imaging tool but also a unique method for quantitative physicochemical characterization to study crystal polymorphs and solid dosage forms.

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Effect of pore diameter on the elution behavior of analytes from thermoresponsive polymer grafted beads packed columns

Nagase K., Umemoto Y., Kanazawa H.

Scientific Reports 11 （ 1 ） 9976 2021年05月

研究論文（学術雑誌）, 共著, 査読有り

　概要を見る

Temperature-responsive chromatography using thermoresponsive polymers is innovative and can control analyte retention via column temperature. Analyte elution behavior in this type of chromatography depends on the modified thermoresponsive polymer and the structure of the base materials. In the present study, we examine the effect of the pore diameter of silica beads on analyte elution behavior in temperature-responsive chromatography. Poly(N-isopropylacrylamide-co-n-butyl methacrylate) hydrogel was applied to beads of various pore sizes: 7, 12, and 30 nm. Almost the same amount of copolymer hydrogel was applied to all beads, indicating that the efficiency of copolymer modification was independent of pore size. Analyte retention on prepared beads in a packed column was observed using steroids, benzodiazepines, and barbiturates as analytes. Analyte retention times increased with temperature on packed columns of 12- and 30-nm beads, whereas the column packed with 7-nm beads exhibited decreased retention times with increasing temperature. The difference in analyte elution behavior among the various pore sizes was attributed to analyte diffusion into the bead pores. These results demonstrate that bead pore diameter determines temperature-dependent elution behavior.

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Selective capture and non-invasive release of cells using a thermoresponsive polymer brush with affinity peptides

Nagase K., Shimura M., Shimane R., Hanaya K., Yamada S., Akimoto A.M., Sugai T., Kanazawa H.

Biomaterials Science 9 （ 3 ） 663 - 674 2021年01月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  2047-4830

　概要を見る

Tissue engineering and cell transplantation therapy have become promising therapies for intractable diseases. These approaches require cell separation technology without cell modification. Accordingly, in this study, we developed a novel cell separation method using a thermoresponsive block copolymer brush with an affinity peptide. A block copolymer brush with bottom poly(2-hydroxyethyl methacrylate [HEMA]-co-propargyl acrylate) and top poly(N-isopropylacrylamide-co-HEMA) segments was prepared through two steps of atom transfer radical polymerization. Then, cell affinity peptides were conjugated to the bottom segment of the copolymer brush through a click reaction. Using cRGD as a cell-Affinity peptide, enhancement of cell adhesion with rapid adhesion on the copolymer brush was observed at 37 °C, whereas the copolymer brush without cRGD did not exhibit cell adhesion. Temperature-modulated cell adhesion and detachment were performed with a relatively long upper segment because the affinity between peptides and cells was modulated by the swelling and shrinking of the upper thermoresponsive segment. Selective endothelial cell adhesion was performed at 37 °C using GGGREDV as an affinity peptide. Smooth muscle cells and fibroblasts did not adhere to the copolymer brush. Adhered human umbilical vein endothelial cells (HUVECs) were successfully recovered by reducing the temperature to 20 °C. Based on the properties of the copolymer brush, HUVECs could be purified using a mixture of cells simply by changing the temperature. These results demonstrated that the prepared copolymer brush with cell affinity peptides could be a useful cell separation tool because the cells could be separated with specificity and without cell modification using a simple procedure.

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Effective Separation for New Therapeutic Modalities Utilizing Temperature-responsive Chromatography

Maekawa Y., Ayano E., Nagase K., Kanazawa H.

Analytical Sciences 37 （ 5 ） 651 - 660 2021年01月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  09106340

　概要を見る

In recent years, drug discovery and therapeutics trends have shifted from a focus on small-molecule compounds to biopharmaceuticals, genes, cell therapy, and regenerative medicine. Therefore, new approaches and technologies must be developed to respond to these changes in medical care. To achieve this, we applied a temperature-responsive separation system to purify a variety of proteins and cells. We developed a temperature-responsive chromatography technique based on a poly(N-isopropylacrylamide) (PNIPAAm)-grafted stationary phase. This method may be applied to various types of protein and cell separation applications by optimizing the properties of the modified polymers used in this system. Therefore, the developed temperature-responsive HPLC columns and temperature-responsive solid-phase extraction (TRSPE) columns can be an effective separation tool for new therapeutic modalities such as monoclonal antibodies, nucleic acid drugs, and cells.

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Thermoresponsive Cationic Block Copolymer Brushes for Temperature-Modulated Stem Cell Separation

Kenichi Nagase, Ayumu Ota, Tadashi Hirotani, Sota Yamada, Aya Mizutani Akimoto, Hideko Kanazawa

Macromolecular Rapid Communications 2000308 2020年08月

研究論文（学術雑誌）, 共著, 査読有り

Design of two complementary copolymers that work as a glue for cell-laden collagen gels

Taihei Nishimoto, Ko Matsukawa, Kenichi Nagase, Hideko Kanazawa, Aya Mizutani Akimoto, Ryo Yoshida

Chemical Communications 46 （ 10565 ） 10548 2020年08月

研究論文（学術雑誌）, 査読有り

Antibody drug separation using thermoresponsive anionic polymer brush modified beads with optimised electrostatic and hydrophobic interactions

Kenichi Nagase, Saki Ishii, Koji Ikeda, Sota Yamada, Daiju Ichikawa, Aya Akimoto, Yutaka Hattori, Hideko Kanazawa

Scientific Reports 10, 11896, (2020) 10   11896 2020年07月

研究論文（学術雑誌）, 共著, 査読有り

Simultaneous analysis of multiple oligonucleotides by temperature-responsive chromatography using a poly(N-isopropylacrylamide)-based stationary phase

Yutaro Maekawa, Kaichi Yamazaki, Miwa Ihara, Kenichi Nagase, Hideko Kanazawa

Analytical and Bioanalytical Chemistry 412   5341 - 5351 2020年06月

研究論文（学術雑誌）, 共著

Green analytical method for the simultaneous analysis of cytochrome P450 probe substrates by poly(N-isopropylacrylamide)-based temperature-responsive chromatography

Yutaro Maekawa, Naoya Okamoto, Yuji Okada, Kenichi Nagase, Hideko Kanazawa

Scientific Reports10(1), 8828, (2020) 10 （ 1 ） 8828 2020年06月

研究論文（学術雑誌）, 共著, 査読有り

Mixed polymer brush as a functional ligand of silica beads for temperature-modulated hydrophobic and electrostatic interactions

Kenichi Nagase, Sakiko Kitazawa, Sota Yamada, Aya Mizutani Akimoto, Hideko Kanazawa

Analytica Chimica Acta 1095   1 - 13 2020年01月

研究論文（学術雑誌）, 共著, 査読有り

Thermoresponsive anionic copolymer brush-grafted surfaces for cell separation

Nagase K., Uchikawa N., Hirotani T., Akimoto A., Kanazawa H.

Colloids and Surfaces B: Biointerfaces （Colloids and Surfaces B: Biointerfaces）  185   110565 2020年01月

ISSN  09277765

　概要を見る

© 2019 Elsevier B.V. Cell separation methods that do not require surface modification of cells are needed in tissue engineering and regenerative medicine. We developed thermoresponsive anionic polymer brushes for cell separation without modification of the cell surfaces. Copolymer brush poly(N-isopropylacrylamide-co-N-tert-butylacrylamide-co-tert-butyl acrylate, P(NIPAAm-co-tBAAm-co-tBA), was prepared on a cover glass plate through activator regenerated by electron transfer atom transfer radical polymerization (ARGET-ATRP). The tert-butyl group of the copolymer brush was then deprotected and a P(NIPAAm-co-tBAAm-co-acrylic acid (AAc)) brush-modified glass surface was obtained. ARGET-ATRP synthesis achieved polymers with low polydispersity. The negative surface charge of the polymer brush-modified substrates was evaluated using zeta potential measurements and the phase transition temperature of the polymer was modulated between 37–20 °C to perform cell adhesion and detachment, respectively. The adhesion and detachment behavior of cells used in cardiovascular tissue engineering on the thermoresponsive anionic polymer brushes was investigated. Normal human umbilical vein endothelial cells (HUVEC) exhibited prompt detachment from the thermoresponsive anionic polymer brush surfaces. In addition, normal human aortic smooth muscle cells (SMC) exhibited relatively high adhesion on thermoresponsive anionic polymer brush-modified surfaces compared with those modified with thermoresponsive polymer brushes without anionic groups. By utilizing the difference in the cell adhesion and detachment properties of the cell types, a mixture of HUVEC and SMC was separated simply by altering the applied temperature. This result indicated that the prepared thermoresponsive anionic polymer brush-modified glass surface could be used as a tool for the separation of cells in cardiovascular tissue engineering.

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刺激応答性高分子を用いたバイオセパレーション

長瀬健一, 金澤秀子

高分子 69 （ 9 ） 472 - 473 2020年

研究論文（学術雑誌）, 共著, 査読有り

Characteristic differences of cell sheets composed of mesenchymal stem cells with different tissue origins

Nakao M., Inanaga D., Nagase K., Kanazawa H.

Regenerative Therapy （Regenerative Therapy）  11   34 - 40 2019年12月

　概要を見る

© 2019 The Japanese Society for Regenerative Medicine Introduction: Stem cell therapy with mesenchymal stem cells (MSCs)has been widely used in many clinical trials, and therapy with MSC sheets shows promise for patients. However, there are few reports characterizing MSC sheets. In the present study, the properties of MSC sheets derived from bone marrow, adipose tissue, and umbilical cord were evaluated. Methods: Cell sheets were fabricated with MSCs from different tissue origins in temperature-responsive cell culture dishes with and without pre-coating of fetal bovine serum (FBS). MSC adhesion behavior in the culture dish was observed. Secretion of cytokines related to cell proliferation and immune regulation from MSC sheets was investigated by ELISA. The adhesion properties of the MSC sheets were investigated by time-lapse microscopy. Results: Different cell adhesion and proliferation rates in temperature-responsive cell culture dishes were observed among the three types of MSCs. FBS pre-coating of the dishes enhanced cell attachment and proliferation in all cell types. Harvested cell sheets showed high attachment capacity to tissue culture polystyrene dish surfaces. Conclusions: MSC sheets can be fabricated from MSCs from different tissue origins using temperature-responsive cell culture dishes. The fabricated MSC sheets could be useful in cell transplantation therapies by choosing appropriate types of MSCs that secrete therapeutic cytokines for the targeted diseases.

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Phenotypic traits of mesenchymal stem cell sheets fabricated by temperature-responsive cell culture plate: structural characteristics of MSC sheets.

Nakao M, Kim K, Nagase K, Grainger DW, Kanazawa H, Okano T

Stem cell research & therapy （Stem Cell Research and Therapy）  10 （ 1 ） 353 2019年11月

　概要を見る

© 2019 The Author(s). Background: In most stem cell therapy strategies reported to date, stem cells are introduced to damaged tissue sites to repair and regenerate the original tissue structure and function. MSC therapeutic efficacies are inconsistent, largely attributed to transplanted MSC difficulties both in engrafting at tissue sites and in retaining their therapeutic functions from suspension formulations. MSC functional components, including cell adhesion and cell-cell junction proteins, and ECM that contribute to essential cellular therapeutic effects, are damaged or removed by proteolytic enzymes used in stem cell harvesting strategies from culture. To overcome these limitations, methods to harvest and transplant cells without disrupting critical stem cell functions are required. Cell sheet technology, exploiting temperature-responsive cell culture surfaces, permits cell harvest without cell protein damage. This study is focused on phenotypic traits of MSC sheets structurally and functionally to understand therapeutic benefits of cell sheets. Methods/results: This study verified cleaved cellular proteins (vinculin, fibronectin, laminin, integrin β-1, and connexin 43) and increased apoptotic cell death produced under standard trypsin harvesting treatment in a time-dependent manner. However, MSC sheets produced without trypsin using only temperature-controlled sheet harvest from culture plastic exhibited intact cellular structures. Also, MSCs harvested using enzymatic treatment (i.e., chemical disruption) showed higher pYAP expression compared to MSC sheets. Conclusion: Retention of cellular structures such as ECM, cell-cell junctions, and cell-ECM junctions is correlated with human umbilical cord mesenchymal stem cell (hUC-MSC) survival after detachment from cell culture surfaces. Retaining these proteins intact in MSC cultures using cell sheet technology is proposed to enhance stem cell survival and their function in stem cell-based therapy.

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Temperature-modulated cell-separation column using temperature-responsive cationic copolymer hydrogel-modified silica beads

Nagase K., Inanaga D., Ichikawa D., Mizutani Akimoto A., Hattori Y., Kanazawa H.

Colloids and Surfaces B: Biointerfaces （Colloids and Surfaces B: Biointerfaces）  178   253 - 262 2019年06月

ISSN  09277765

　概要を見る

© 2019 Elsevier B.V. There is strong demand for cell separation methods that do not decrease cell activity or modify cell surfaces. Here, new temperature-modulated cell-separation columns not requiring cell-surface premodification are described. The columns were packed with temperature-responsive cationic polymer hydrogel-modified silica beads. Poly(N-isopropylacrylamide-co-n-butyl methacrylate-co-N,N-dimethylaminopropyl acrylamide) hydrogels with various cationic moieties were attached to silica-bead surfaces by radical polymerization using N,Nʹ-methylenebisacrylamide as a crosslinking agent. The beads were packed into solid-phase extraction columns, and temperature-dependent cell elution from the columns was found using HL-60 and Jurkat cells. The retention HL-60 and Jurkat cells in columns containing cationic beads at 37 °C was 95.3% to 99.6% and 95.0% to 98.8%, respectively. By contrast, beads without cationic properties exhibited low cell retention (20.6% for HL-60 and 32.5% for Jurkat cells). The cells were mainly retained through both electrostatic and hydrophobic interactions. The retained HL-60 (4.9%) and Jurkat cells (40%) were eluted at 4 °C from the column with a low composition of cationic monomer (DMAPAAm, 1 mol% in copolymer), because the temperature-responsive hydrogels on the beads became hydrophilic, decreasing the hydrophobic interactions between the cells and the beads. A higher number of Jurkat cells than HL-60 cells were eluted because of differences in their electrostatic properties (Jurkat cells: −2.53 mV; HL-60 cells: −20.7 mV). The results indicated that cell retention by the hydrogel-coated beads packed in a solid phase extraction column could be modulated simply by changing the temperature.

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Adsorption-Desorption Control of Fibronectin in Real Time at the Liquid/Polymer Interface on a Quartz Crystal Microbalance by Thermoresponsivity

Li J., Kaku T., Tokura Y., Matsukawa K., Homma K., Nishimoto T., Hiruta Y., Akimoto A., Nagase K., Kanazawa H., Shiratori S.

Biomacromolecules （Biomacromolecules）  20 （ 4 ） 1748 - 1755 2019年04月

ISSN  15257797

　概要を見る

Copyright © 2019 American Chemical Society. The cell manipulation technique using thermoresponsive polymers is currently attracting much attention for applications in the medical field. To achieve arbitrary and accurate cell control, it is necessary to intensely research fibronectin behavior. A smart surface, which has thermoresponsive wettability and which can adsorb or desorb fibronectin repeatedly without the presence of cells, was fabricated by an electrospinning method. The fabricated coating changed its structure as the temperature was changed, and this transformation could substitute for the pulling force generated by the cytoskeletal contraction of cells. Moreover, a coated quartz crystal microbalance was able to detect the fibronectin behavior as frequency shifts, which could be used in the estimation of the mass shift with the aid of suitable equations. This coating and measurement system can contribute greatly not only to the development in the medical field centered on biomaterial manipulation technologies, but also to the improvement of medical instruments.

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LAT1-Targeting Thermoresponsive Liposomes for Effective Cellular Uptake by Cancer Cells

Maekawa-Matsuura M., Fujieda K., Maekawa Y., Nishimura T., Nagase K., Kanazawa H.

ACS Omega （ACS Omega）  4 （ 4 ） 6443 - 6451 2019年04月

　概要を見る

© 2019 American Chemical Society. L-type amino acid transporter 1 (LAT1) is a transporter that is more highly expressed in cancer cells compared with normal cells. In the present study, liposomes, composed of egg phosphatidylcholine (EPC) and dioleoyl phosphatidylethanolamine, were modified with LAT1-targeting thermoresponsive polymer, l-tyrosine-conjugated poly(N-isopropylacrylamide-co-N,N-dimethylacrylamide) (P(NIPAAm-co-DMAAm)). The cellular uptake of the prepared LAT1-targeting liposomes was evaluated using HeLa cells as a cancer cell model. At temperatures above the polymer's lower critical solution temperature, uptake of the liposomes into cells was observed because the polymer at the liposome surface became hydrophobic and interacted with the cell membrane. Flow cytometry analysis suggested that l-tyrosine-P(NIPAAm-co-DMAAm)-liposomes exhibited markedly increased cellular uptake by HeLa cells compared with that of liposomes not modified with l-tyrosine. This result indicated that cellular uptake of liposomes can be enhanced by the affinity between l-tyrosine and the LAT1 of HeLa cells. The developed functional liposomes, which exhibit both thermoresponsive and LAT1-targeting properties, would be appropriate for temperature-modulated drug delivery and imaging with good targeting ability.

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Liposomes with temperature-responsive reversible surface properties

Nemoto R., Fujieda K., Hiruta Y., Hishida M., Ayano E., Maitani Y., Nagase K., Kanazawa H.

Colloids and Surfaces B: Biointerfaces （Colloids and Surfaces B: Biointerfaces）  176   309 - 316 2019年04月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  09277765

　概要を見る

© 2019 Elsevier B.V. Liposomes composed of egg phosphatidylcholine and cholesterol were modified with the temperature-responsive polymer poly(N-isopropylacrylamide-co-N, N-dimethylacrylamide) (P(NIPAAm-co-DMAAm)), and exhibited reversible surface properties with temperature. Completely reversible liposome aggregation due to P(NIPAAm-co-DMAAm) hydration/dehydration was demonstrated over four successive cycles of heating and cooling. The P(NIPAAm-co-DMAAm) polymer was hydrated during cooling, which dispersed the liposomes. The rigidity of the liposomal membrane was one of the factors in the reversible aggregation, as was the modification density of the polymer on the liposomes. A low density on relatively rigid liposomes could maintain the polymer property of reversible hydrated layers below critical solution temperature (LCST) boundary. Above the LCST, temperature-responsive polymers could also transport negatively charged liposomes into cells. The reversible behavior of the temperature-responsive polymer-modified liposomes has not been reported previously and could enable new applications for switching deposit forms as alternative drug carriers.

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Effect of polymer phase transition behavior on temperature-responsive polymer-modified liposomes for siRNA transfection

Nagase K., Hasegawa M., Ayano E., Maitani Y., Kanazawa H.

International Journal of Molecular Sciences （International Journal of Molecular Sciences）  20 （ 2 ） 430 2019年01月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  16616596

　概要を見る

© 2019 by the authors. Licensee MDPI, Basel, Switzerland. Small interfering RNAs (siRNAs) have been attracting significant attention owing to their gene silencing properties, which can be utilized to treat intractable diseases. In this study, two temperature-responsive liposomal siRNA carriers were prepared by modifying liposomes with different polymers—poly(N-isopropylacrylamide-co-N,N-dimethylaminopropyl acrylamide) (P(NIPAAm-co-DMAPAAm)) and poly(N-isopropylacrylamide-co-N,N-dimethylacrylamide) P(NIPAAm-co-DMAAm). The phase transition of P(NIPAAm-co-DMAPAAm) was sharper than that of P(NIPAAm-co-DMAAm), which is attributed to the lower co-monomer content. The temperature dependent fixed aqueous layer thickness (FALT) of the prepared liposomes indicated that modifying liposomes with P(NIPAAm-co-DMAPAAm) led to a significant change in the thickness of the fixed aqueous monolayer between 37 ◦ C and 42 ◦ C; while P(NIPAAm-co-DMAAm) modification led to FALT changes over a broader temperature range. The temperature-responsive liposomes exhibited cellular uptake at 42 ◦ C, but were not taken up by cells at 37 ◦ C. This is likely because the thermoresponsive hydrophilic/hydrophobic changes at the liposome surface induced temperature-responsive cellular uptake. Additionally, siRNA transfection of cells for the prevention of luciferase and vascular endothelial growth factor (VEGF) expression was modulated by external temperature changes. P(NIPAAm-co-DMAPAAm) modified liposomes in particular exhibited effective siRNA transfection properties with low cytotoxicity compared with P(NIPAAm-co-DMAAm) modified analogues. These results indicated that the prepared temperature-responsive liposomes could be used as effective siRNA carriers whose transfection properties can be modulated by temperature.

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Mixed polymer brush as a functional ligand of silica beads for temperature-modulated hydrophobic and electrostatic interactions

Nagase K., Kitazawa S., Yamada S., Akimoto A., Kanazawa H.

Analytica Chimica Acta （Analytica Chimica Acta）  1095   1 - 13 2019年

ISSN  00032670

　概要を見る

© 2019 Elsevier B.V. We developed a mixed polymer brush, which consists of temperature-responsive polymer and cationic polymer modified beads as functional chromatographic matrices, for temperature-modulated multiple hydrophobic and electrostatic interactions. The mixed polymer brush was modified on silica beads through the combination of reversible addition−fragmentation chain transfer (RAFT) polymerization of N,N-dimethylaminopropyl acrylamide (DMAPAAm) and surface initiated atom transfer radical polymerization (ATRP) of N-isopropylacrylamide (NIPAAm). Zeta potential measurement of the mixed polymer brush revealed that the zeta potential increased with increasing temperature, which was attributed to the exposed PDMAPAAm on the beads arising from the shrinking of PNIPAAm upon increasing the temperature. The prepared beads were used as the packing material of high performance liquid chromatography (HPLC) columns, and the elution behavior of steroids, adenosine nucleotides, and proteins through the column was observed. The retention time of steroids increased with increasing the column temperature because of the dehydration of PNIPAAm in the mixed polymer brush. Adenosine nucleotides were also retained by the columns, which was attributed to the electrostatic interaction with PDMAPAAm in the mixed polymer brush. Several proteins were adsorbed on the column at elevated temperatures because of the enhanced electrostatic interaction of exposed PDMAPAAm and the enhanced hydrophobic interaction resulting from the dehydration of PNIPAAm. By exploiting this unique property, mixtures of proteins could be separated by simply changing the column temperature. These results indicate that the developed mixed polymer brush modified beads would be useful as functional chromatographic packing matrices for thermally-modulated multiple hydrophobic and electrostatic interactions.

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Temperature-responsive mixed-mode column containing temperature-responsive polymer-modified beads and anionic polymer-modified beads

Nagase K., Watanabe M., Zen F., Kanazawa H.

Analytica Chimica Acta （Analytica Chimica Acta）  1079   220 - 229 2019年

ISSN  00032670

　概要を見る

© 2019 Elsevier B.V. We developed temperature-responsive mixed-mode columns packed with poly(N-isopropylacrylamide) (PNIPAAm)-modified beads and poly(2-acrylamido-2-methylpropane sulfonic acid) (PAMPS)-modified beads in various ratios. The PNIPAAm-modified silica beads and PAMPS-modified silica beads were prepared by surface-initiated atom transfer radical polymerization of N-isopropylacrylamide and 2-acrylamido-2-methylpropane sulfonic acid, respectively. We confirmed polymer modification of the silica beads by CHN elemental analysis, FTIR, zeta-potential measurements, and SEM. To determine the column separation efficiency, we examined the elution behaviors of cold medicine active ingredients and monoamines from each column. Analyte separation occurred on columns with PNIPAAm to PAMPS ratios of 1:20 and 1:10, whereas a column containing only PNIPAAm-modified beads did not retain the analytes. The analytes were retained on the columns through hydrophobic and electrostatic interactions with PNIPAAm and PAMPS, respectively. The separation performance improved with increasing column temperature because of dehydration of PNIPAAm and enhancement of hydrophobic interactions at elevated temperatures. The mixed-mode columns will be useful for separating basic bioactive compounds because the retention of analytes can be modulated by changing the column temperature and the composition.

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Crosslinked Poly(N-Isopropylacrylamide)-Based Microfibers as Cell Manipulation Materials with Prompt Cell Detachment

Konishi T., Mizutani Akimoto A., Nishimoto T., Tokura Y., Tenjimbayashi M., Homma K., Matsukawa K., Kaku T., Hiruta Y., Nagase K., Kanazawa H., Shiratori S.

Macromolecular Rapid Communications （Macromolecular Rapid Communications）  40 （ 23 ） e1900464 2019年

ISSN  10221336

　概要を見る

© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim Stimuli-responsive smart materials are a key to the realization of next-generation medical technologies. Among them, the temperature-responsive polymer poly(N-isopropylacrylamide) (PNIPAAm) is attracting particular attention because it is easy to use in physiological conditions. PNIPAAm-grafted surfaces can undergo temperature-modulated cell adhesion and detachment without proteolytic enzymes, and can be used as cell-separating materials through selective cell adhesion/detachment. However, cell detachment at reduced temperatures is problematic because it takes several hours. A novel thermoresponsive crosslinked microfiber system that can greatly reduce the cell detachment time is introduced in this study. The crosslinked fibers provide temperature-dependent volume change, and enable cell detachment within 10 min of reducing the temperature, which is one-sixth of the time required in previous studies. The prompt cell detachment is thought to arise from a completely new mechanism derived from fiber swelling. This system will make a significant contribution as a novel cell manipulating system for next-generation medical technology.

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CD44-targeting nanocarriers for cancer treatment

Yamanouchi T., Kanazawa H.

Drug Delivery System （Drug Delivery System）  34 （ 1 ） 38 - 45 2019年

ISSN  09135006

　概要を見る

© 2019, Japan Society of Drug Delivery System. All rights reserved. Recently, the theory of cancer stem cell (CSC) is gaining increasing attention from researchers and has become an important focus of cancer research. Since the discovery that the marker molecule is over-expressed in a variety of solid tumors, many studies have focused on methods for targeting the hyaluronan receptor CD44. Hyaluronic acid, the primary CD44 binding molecule, has proved a significant factor in developing nanocarriers that demonstrate preferential tumor accumulation and increased cell uptake. Research is frequently conducted to add further functions to nanocarriers targeting CD44. It is expected that more effective drug delivery will be possible by combining with existing technology. In this review, we introduce research approaches from the current literature that take advantage of CD44-targeting ability and describe the possible advantages for each approach. Finally, we describe our recent progress in hyaluronic acid-modified thermoresponsive liposome aiming for CD44-targeting and combined with oncothermia.

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Protein purification using solid-phase extraction on temperature-responsive hydrogel-modified silica beads

Okubo K., Ikeda K., Oaku A., Hiruta Y., Nagase K., Kanazawa H.

Journal of Chromatography A （Journal of Chromatography A）  1568   38 - 48 2018年09月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  00219673

　概要を見る

© 2018 Elsevier B.V. Recently, the importance of biopharmaceuticals in medical treatments has been increasing, and effective protein purification methods are strongly required for their production. In the present study, a temperature-responsive solid-phase extraction (SPE) column was developed for the purification of proteins without affecting their bioactivity. A temperature-responsive polymer hydrogel-modified stationary phase was prepared by coating aminopropyl silica beads (average diameter, 40–64 μm) with poly(N-isopropylacrylamide) (PNIPAAm)-based thermoresponsive hydrogels. n-Butyl methacrylate and acrylic acid were copolymerized with PNIPAAm as hydrophobic and anionic monomers, respectively. Using these temperature-responsive SPE columns, targeted proteins were retained on the thermoresponsive hydrogel at 40 °C through hydrophobic and electrostatic interactions. After the temperature was reduced from 40 °C to 4 °C, the retained proteins were successfully eluted from the column. Using the temperature-responsive SPE system, lysozyme was successfully separated from ovalbumin without any loss in bioactivity (99.7 ± 0.1%). Rituximab, a monoclonal antibody, was also purified from BSA or hybridoma cell culture medium using the prepared SPE column. Denaturation of rituximab was not observed in the rituximab fraction eluted from the SPE column. These results demonstrate that temperature-responsive polymer-based SPE can be applied in biomedical purifications, while maintaining the biological activity of the proteins.

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Comparison of plasma propofol concentration for apnea, response to mechanical ventilation, and airway device between endotracheal tube and supraglottic airway device in Beagles

Tomoya IIZUKA, Kenichi MASUI, Hideko KANAZAWA, Ryohei NISHIMURA

Journal of Veterinary Medical Science 80 （ 9 ） 1420 - 1423 2018年07月

研究論文（学術雑誌）, 共著, 査読有り

LAT1-targeting thermoresponsive fluorescent polymer probes for cancer cell imaging

Matsuura M., Ohshima M., Hiruta Y., Nishimura T., Nagase K., Kanazawa H.

International Journal of Molecular Sciences （International Journal of Molecular Sciences）  19 （ 6 ） 1646 2018年06月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  16616596

　概要を見る

© 2018 by the authors. Licensee MDPI, Basel, Switzerland. L-type amino acid transporter 1 (LAT1) is more highly expressed in cancer cells compared with normal cells. LAT1 targeting probes would therefore be a promising tool for cancer cell imaging. In this study, LAT1-targeting thermoresponsive fluorescent polymer probes based on poly(N-isopropylacrylamide-co-N,N-dimethylacrylamide) (P(NIPAAm-co-DMAAm)) were synthesized and their affinity for LAT1 was evaluated. The synthesized polymer probes interacted with LAT1 on HeLa cells, and inhibition of L-[3 H]-leucine, one of the substrates for LAT1 uptake, was investigated. L-Tyrosine-conjugated P(NIPAAm-co-DMAAm) inhibited the uptake of L-[3 H]-leucine, while P(NIPAAm-co-DMAAm) and L-phenylalanine-conjugated P(NIPAAm-co-DMAAm) did not. This result indicated that L-tyrosine-conjugated polymer has a high affinity for LAT1. The fluorescent polymer probes were prepared by modification of a terminal polymer group with fluorescein-5-maleimide (FL). Above the polymer transition temperature, cellular uptake of the polymer probes was observed because the polymers became hydrophobic, which enhanced the interaction with the cell membrane. Furthermore, quantitative analysis of the fluorescent probe using flow cytometry indicated that L-tyrosine-conjugated P(NIPAAm-co-DMAAm)-FL shows higher fluorescence intensity earlier than P(NIPAAm-co-DMAAm)-FL. The result suggested that cellular uptake was promoted by the LAT1 affinity site. The developed LAT1-targeting thermoresponsive fluorescent polymer probes are expected to be useful for cancer cell imaging.

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Mesenchylmal stem cell culture on poly(N-isopropylacrylamide) hydrogel with repeated thermo-stimulation

Akimoto A., Niitsu E., Nagase K., Okano T., Kanazawa H., Yoshida R.

International Journal of Molecular Sciences （International Journal of Molecular Sciences）  19 （ 4 ） 1253 2018年04月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  16616596

　概要を見る

© 2018 by the authors. We prepared thermoresponsive hydrogels by mixing poly(N-isopropylacrylamide) (PNIPAAm) derivatives as the main chain components, octa arm polyethylene glycol (PEG) as a crosslinker, and the Arg-Gly-Asp-Ser (RGDS) peptides as cell adhesion units. Human bone marrow derived mesenchymal stem cells (hbmMSCs) were cultured on the hydrogels. The PNIPAAm gel prepared by the post crosslinking gelation method was revealed to be cytocompatible and showed temperature dependent changes in mechanical properties. Repeated changes in the swelling ratio of the PNIPAAm gel affected the shape of the hbmMSCs. With respect to both cytocompatibility and reversibility of changes in mechanical properties, the PNIPAAm gel system could be potentially useful for the analysis of cell mechanobiology.

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Poly(N-isopropylacrylamide) based thermoresponsive polymer brushes for bioseparation, cellular tissue fabrication, and nano actuators

Nagase K., Okano T., Kanazawa H.

Nano-Structures and Nano-Objects （Nano-Structures and Nano-Objects）  16   9 - 23 2018年04月

研究論文（学術雑誌）, 共著

　概要を見る

© 2018 Elsevier B.V. Many thermoresponsive surfaces for biomedical applications have been developed using various techniques for modifying substrates with thermoresponsive polymers. In addition, surface-initiated living radical polymerization methods such as atom-transfer radical polymerization (ATRP) and reversible addition–fragmentation chain-transfer (RAFT) polymerization have been developed as nano structure controlled surface modification methods for modifying thermoresponsive polymers to substrates. These polymerization techniques allow the accurate control of polymerization and the formation of densely packed polymer brush structures. The present review article summarizes a variety of thermoresponsive polymer brushes, mainly comprising poly(N-isopropylacrylamide) (PIPAAm), prepared by surface-initiated ATRP and RAFT polymerization. Self-oscillating polymer brushes as new nano-actuators are also discussed.

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Design of functional thermoresponsive polymer brushes and their application to bioseparation

Nagase K., Okano T., Kanazawa H.

Kobunshi Ronbunshu （Kobunshi Ronbunshu）  75 （ 2 ） 143 - 154 2018年03月

ISSN  03862186

　概要を見る

©2018, The Society of Polymer Science, Japan. Protein drugs and therapeutic cells are among promising current and future therapies for patients. The importance of these proteins and cells has increased in the clinical field, and protein and cell purification methods, without losing their potency or biological activity, are urgently needed. We prepared thermoresponsive polymer brushes as a new protein and cell separation tool because interactions between the polymer brushes and targeted proteins and cells can be modulated by changing the external solution temperature without using other agents that might contaminate these biologics or disrupt their function. Using these new thermoresponsive polymer brushes, a variety of different bio-separations have been performed by simple temperature changes in aqueous solutions. The present review describes the recent achievement in bioseparation using thermoresponsive polymer brushes.

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Development of nanocarriers functionalized with stimuli-responsive polymer for controlled cellular uptake

Yamanouchi T., Katsuyama N., Hiruta Y., Ayano E., Kanazawa H.

Kobunshi Ronbunshu （Kobunshi Ronbunshu）  75 （ 2 ） 116 - 127 2018年03月

ISSN  03862186

　概要を見る

©2018, The Society of Polymer Science, Japan. Drug delivery system (DDS) is a technology that controls sustained release of medicines in the body and localized delivery to diseased tissues. Therefore, it is possible to keep the concentration of medicine at therapeutically effective concentrations for a long time and to reduce side effects. We have achieved controlled intracellular uptake, and release of medicines by nanocarriers modified with stimuli-responsive polymers based on poly(N-isopropylacrylamide), which undergoes a reversible phase transition in response to external temperature stimulation. In this article, we introduce our recent research on the design of temperature- and pH-responsive polymers, which enables selective intracellular uptake with external stimulation, and applications to DDS of an anti-tumor agent and siRNA by micelle and liposomes modified with those polymers.

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Poly(N-isopropylacrylamide)-based thermoresponsive surfaces provide new types of biomedical applications

Nagase K., Yamato M., Kanazawa H., Okano T.

Biomaterials （Biomaterials）  153   27 - 48 2018年01月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  01429612

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© 2017 The Authors Thermoresponsive surfaces, prepared by grafting of poly(N-isopropylacrylamide) (PIPAAm) or its copolymers, have been investigated for biomedical applications. Thermoresponsive cell culture dishes that show controlled cell adhesion and detachment following external temperature changes, represent a promising application of thermoresponsive surfaces. These dishes can be used to fabricate cell sheets, which are currently used as effective therapies for patients. Thermoresponsive microcarriers for large-scale cell cultivation have also been developed by taking advantage of the thermally modulated cell adhesion and detachment properties of thermoresponsive surfaces. Furthermore, thermoresponsive bioseparation systems using thermoresponsive surfaces for separating and purifying pharmaceutical proteins and therapeutic cells have been developed, with the separation systems able to maintain their activity and biological potency throughout the procedure. These applications of thermoresponsive surfaces have been improved with progress in preparation techniques of thermoresponsive surfaces, such as polymerization methods, and surface modification techniques. In the present review, the various types of PIPAAm-based thermoresponsive surfaces are summarized by describing their preparation methods, properties, and successful biomedical applications.

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Reversible conformational changes in parallel type G-quadruplex structure inside a thermoresponsive hydrogel

Erika Hasuike, Aya Mizutani Akimotoa, Reiko Kurodab, Ko Matsukawa, Yuki Hiruta, Hideko Kanazawa, Ryo Yoshida

Chemical Communications 53   3142 - 3144 2017年02月

研究論文（学術雑誌）, 共著, 査読有り

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Dual temperature- and pH-responsive polymeric micelle for selective and efficient two-step doxorubicin delivery

Yuki Hiruta, Yuki Kanda, Naoya Katsuyama, Hideko Kanazawa.

RSC Adv 7   29540 - 29549 2017年

研究論文（学術雑誌）, 共著, 査読有り

Transcutaneous drug delivery by liposomes using fractional laser technology

Takahiro Fujimoto, Jian Wang, Kazuki Baba, Yuka Oki, Yuki Hiruta, Masayuki Ito, Shinobu Ito, Hideko Kanazawa

Lasers in Surgery & Medicine 49   525 - 532 2017年

研究論文（学術雑誌）, 共著

The use of a temperature-responsive column for the direct analysis of drugs in serum by two-dimensional heart-cutting liquid chromatography

Toshiyasu Mikuma, Ryo Uchida, Mizuki Kajiya, Yuki Hiruta, Hideko Kanazawa

Analytical and Bioanalytical Chemistry 409   1059 - 1065 2017年

研究論文（学術雑誌）, 共著, 査読有り

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Tunable Surface Properties of Temperature-Responsive Polymer Modified Liposomes Induce Faster Cellular Uptake

Jian Wang, Eri Ayano, Yoshie Maitani, Hideko Kanazawa

ACS OMEGA 2 （ 1 ） 316 - 325 2017年

研究論文（学術雑誌）, 共著, 査読有り

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Propofol-fentanyl interaction in Beagles - Apnea, response to mechanical ventilation, endotracheal tube, and tetanic stimulation

T. Iizuka, K. Masui, T. Miyabe-Nishiwaki, H. Kanazawa, R. Nishimura

Research in Veterinary Science 115   34 - 42 2017年

研究論文（学術雑誌）, 共著

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Fractional laser-assisted percutaneous drug delivery via temperature-responsive liposomes

Takahiro Fujimoto, Masayuki Ito, Shinobu Ito, Hideko Kanazaw

Journal of Biomaterials Science, Polymer Edition 28 （ 7 ） 679 - 689 2017年

研究論文（学術雑誌）, 共著, 査読有り

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Enhanced Cellular Uptake and Gene Silencing Activity of siRNA Using Temperature-Responsive Polymer-Modified Liposome.

Wang J, Ayano E, Maitani Y, Kanazawa H

Int J Pharm 523   217 - 228 2017年

研究論文（学術雑誌）, 共著, 査読有り

Design and synthesis of temperature-responsive polymer/silica hybrid nanoparticles and application to thermally controlled cellular uptake

Hiruta Y, Nemoto R, Kanazawa H

Colloids Surf B Biointerfaces 153   2 - 9 2017年

研究論文（学術雑誌）, 共著, 査読有り

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Intracellular localization and delivery of plasmid DNA by biodegradable microsphere-mediated femtosecond laser optoporation

Atsuhiro Ishii, Yuki Hiruta, Dag Heinemann, Alexander Heisterkamp, Hideko Kanazawa, Mitsuhiro Terakawa

Journal of Biophotonics 10   1723 - 1731 2017年

研究論文（学術雑誌）, 共著, 査読有り

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Local Release of VEGF Using Fiber Mats Enables Effective Transplantation of Layered Cardiomyocyte Sheets

Kenichi Nagase, Yuhei Nagumo, Miri Kim, Hee-Jung Kim, Hei-Won Kyung, Hye-Jin Chung, Hidekazu Sekine, Tatsuya Shimizu, Hideko Kanazawa, Teruo Okano, Seung-Jin Lee, Masayuki Yamato

Macromolecular Bioscience 17   1700073 2017年

研究論文（学術雑誌）, 査読有り

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超高速液体クロマトグラフィーを用いた向精神薬の複数同時迅速分析.

芳川満輝、内田亮、黒木巽、三熊敏靖、蛭田勇樹、永田佳子、金澤秀子

分析化学 65 （ 5 ） 173 - 179 2016年

研究論文（学術雑誌）, 共著, 査読有り

Temperature-responsive molecular recognition chromatography using phenylalanine and tryptophan derived polymer modified silica beads.

Hiruta Y, Kanazashi R, Ayano E, Okano T, Kanazawa H

Analyst 141 （ 3 ） 910 - 917 2016年

研究論文（学術雑誌）, 共著

Protein separations via thermally responsive ionic block copolymer brush layers.

Nagase K, Kobayashi J, Kikuchi, A, Akiyama Y, Kanazawa H, Okano T.

RSC Advances 6   26254 - 26263 2016年

研究論文（学術雑誌）, 共著

Thermoresponsive Anionic Block Copolymer Brushes with Strongly Anionic Bottom Segment for Effective Interactions with Biomolecules

Kenichi Nagase, Jun Kobayashi, Akihiko Kikuchi, Yoshikatsu Akiyama, Hideko Kanazawa, Teruo Okano.

RSC Adv. 6   93169 - 93179 2016年

研究論文（学術雑誌）, 共著, 査読有り

Design of tetra-arm PEG-crosslinked thermoresponsive hydrogel for 3D cell culture

Aya Mizutani Akimoto, Erika Hasuike, Hiroto Tada, Kenichi Nagase, Teruo Okano, Hideko Kanazawa, Ryo Yoshida

Anal. Sci. 32   1203 - 1205 2016年

研究論文（学術雑誌）, 共著

Approaching over 10,000-fold sensitivity increase in chiral capillary electrophoresis: cation-selective exhaustive injection and sweeping cyclodextrin-modified micellar electrokinetic chromatography

T Mikuma, YT Iwata, H Miyaguchi, K Kuwayama, K Tsujikawa, T Kanamori, H Kanazawa, H Inoue

ELECTROPHORESIS 37 （ 22 ） 2970 - 2976 2016年

研究論文（学術雑誌）, 共著

Temperature-responsive molecular recognition chromatography using phenylalanine and tryptophan derived polymer modified silica beads

金澤 秀子

Analyst 2015年11月

研究論文（学術雑誌）, 共著, 査読有り

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pH/temperature-responsive fluorescence polymer probe with pH-controlled cellular uptake

Yuki Hiruta, Takaaki Funatsu, Minami Matsuura, Jian Wang, Eri Ayano, Hideko Kanazawa

Sensors and Actuators B 207   724 - 731 2015年

研究論文（学術雑誌）, 共著, 査読有り

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降圧配合錠の製剤学的性質が成分溶出性に与える影響

永田佳子, 阿南ゆりえ, 金澤秀子

分析化学 64 （ 11 ）  2015年

研究論文（学術雑誌）, 共著, 査読有り

Thermoresponsive Hydrophobic Copolymer Brushes Modified Porous Monolithic Silica for High-Resolution Bioseparation

金澤 秀子

RSC Advances 5   66155 - 66167 2015年

研究論文（学術雑誌）, 共著, 査読有り

Effects of Terminal Group and Chain Length on Temperature-Responsive Chromatography Utilizing Poly(N-isopropylacrylamide) Synthesized via RAFT Polymerization

Hiruta Y, Nagumo Y, Miki A, Okano T, Kanazawa H

RSC Advances 5   73217 - 73224 2015年

研究論文（学術雑誌）, 査読有り

Temperature-responsive Solid-phase Extraction Column for Biological Sample Pretreatment

金澤 秀子

Anal. Sci. 31 （ 9 ） 881 - 886 2015年

研究論文（学術雑誌）, 共著, 査読有り

Design of Environmentally Responsive Fluorescent Polymer Probes for Cellular Imaging.

金澤 秀子

Biomacromolecules 16   2356 - 2362 2015年

研究論文（学術雑誌）, 共著, 査読有り

The effects of anionic electrolytes and human serum albumin　on the LCST of poly(N-isopropylacrylamide)-based temperature-responsive copolymers.

金澤 秀子

Colloids and Surfaces B: Biointerfaces 132   299 - 304 2015年

研究論文（学術雑誌）, 共著, 査読有り

Measurement of the dynamic behavior of thin poly(N-isopropylacrylamide) hydrogels and their phase transition temperatures measured using reflectometric interference spectroscopy

金澤 秀子

J. Nanopart. Res. 17 （ 148 ）  2015年

研究論文（学術雑誌）, 共著, 査読有り

The mechanism of melanocytes-specific cytotoxicity induced by phenol compound having a pro-oxidant effect, relating to the appearance of leukoderma

Nagata T, Ito S, Itoga K, Kanazawa H, Masaki H

Biomed. Res. Int. 2015年

研究論文（学術雑誌）, 共著, 査読有り

Simultaneous Analysis of Oral Antidiabetic Drug by LC-MS/MS

Kai S, Ishikawa K, Ito H, Ogawa T, Yamashita H, Nagata Y, Kanazawa H

Chromatography 36   19 - 24 2015年

研究論文（学術雑誌）, 共著, 査読有り

ビタミンE含有サプリメントの機能評価および多成分併用効果の検討

永田 佳子，星野 由依，平出 園絵，重田 まりあ，蔭山 大輔，金澤 秀子

ビタミンE研究の進歩XVI 16   86 - 91 2014年12月

研究論文（その他学術会議資料等）, 共著

Liquid Chromatography-Mass Spectrometric Analysis of Dehydroepiandrosterone and Related Steroids Utilizing a Temperature-Responsive Stationary Phase

Ayano E, Suzuki Y, Nishio T, Nagata Y, Kanazawa H, Nagase K, Okano T

Chromatograpy 35   131 - 138 2014年11月

研究論文（学術雑誌）, 共著, 査読有り

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分子認識部位を導入した機能性高分子修飾充填剤による温度応答性クロマトグラフィー

坂田 和貴, 大久保 廣平, 蛭田 勇樹, 綾野 絵理, 金澤 秀子

高分子論文集 71 （ 7 ） 293 - 301 2014年07月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  0386-2186

Temperature-Responsive Fluorescence Polymer Probes with Accurate Thermally Controlled Cellular Uptakes.

Yuki Hiruta, Mirai Shimamura, Minami Matsuura, Yutaro Maekawa, Takaaki Funatsu, Yuichi Suzuki, Eri Ayano, Teruo Okano and Hideko Kanazawa.

ACS Macro Letters 3   281-285 2014年03月

研究論文（学術雑誌）, 共著, 査読有り

Monolithic Silica Rods Grafted with Thermoresponsive Anionic Polymer Brushes for High-Speed Separation of Basic Biomolecules and Peptides

金澤 秀子

Bio Macromolecules 15 （ 4 ） 1204 - 1215 2014年02月

研究論文（学術雑誌）, 共著

High temperature heat source generation with quasi-continuous wave semiconductor lasers at low power levels of 6W for medical use.

Fujimoto T*, Imai Y, Tei K, Fujioka T, Ito S, Kanazawa H, Yamaguchi S.

Journal of Biomedical Optics 19 （ 10 ） 101502 2014年02月

研究論文（学術雑誌）, 共著, 査読有り

Thermoresponsive Copolymer Brushes Possessing Quaternary Amine Groups for Strong Anion-Exchange Chromatographic Matrices.

Nagase K, Geven M, Kimura S, Kobayashi J, Kikuchi A, Akiyama Y, Grijpma D, W, Kanazawa H, Okano T*.

Biomacromolecules 15   1031-1043 2014年02月

研究論文（学術雑誌）, 共著, 査読有り

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Temperature-responsive smart packing materials utilizing multi-functional polymers.

Ayano E, Kanazawa H*.

Analytical Sciences 30   167-173 2014年01月

研究論文（学術雑誌）, 共著, 査読有り

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超高速LCを用いる複合型抗酸化サプリメント成分の迅速定量分析

平出園絵, 星野由依, 重田まりあ, 蔭山大輔, 金澤秀子, 永田佳子.

分析化学 63 （ 8 ） 679 - 685 2014年

研究論文（学術雑誌）, 共著, 査読有り

フォトダイオードアレイ検出器を備えた超高速LCによる降圧配合錠の確認試験.

宮野 弘之, 金澤 秀子, 永田 佳子*.

分析化学 62   743-750 2013年08月

研究論文（学術雑誌）, 共著, 査読有り

フォトダイオードアレイ検出器を備えた超高速LCによる降圧配合錠の確認試験.

宮野弘之, 金澤 秀子, 永田佳子*.

分析化学 62(8)   743-750 2013年08月

研究論文（学術雑誌）, 共著, 査読有り

超高速HPLCを用いる経口糖尿病薬の複数同時迅速分析と配合錠への応用.

柄本佳弘, 石川謙, 伊藤一, 平出園絵, 永田佳子, 金澤 秀子*.

分析化学 62(8)   725-730 2013年08月

研究論文（学術雑誌）, 共著, 査読有り

超高速HPLCを用いる経口糖尿病薬の複数同時迅速分析と配合錠への応用.

柄本 佳弘, 石川 謙, 伊藤 一, 平出 園絵, 永田 佳子, 金澤 秀子*.

分析化学 62   725-730 2013年08月

研究論文（学術雑誌）, 共著, 査読有り

陽イオン交換樹脂を用いる放射性セシウムの除去.

森田裕子, 水村亮介, 橘義貴, 金澤 秀子.

分析化学 62(6)   541-545 2013年06月

研究論文（学術雑誌）, 共著, 査読有り

Thermally modulated cationic copolymer brush on monolithic silica rods for high-speed separation of acidic biomolecules.

Nagase K, Kobayashi J, Kikuchi A, Akiyama Y, Kanazawa H, Okano T*.

ACS Applied Materials & Interfaces 5   1442-1452 2013年02月

研究論文（学術雑誌）, 共著, 査読有り

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Evaluation of the predictive performance of a pharmacokinetic model for propofol in Japanese macaques (Macaca fuscata fuscata).

Miyabe-Nishiwaki T, Masui K, Kaneko A, Nishiwaki K, Shimbo E, Kanazawa H.

J Vet Pharmacol Ther 36 （ 2 ） 169 - 173 2013年

研究論文（学術雑誌）, 共著, 査読有り

Induction of different reactive oxygen species in the skin during various laser therapies and their inhibition by fullerene.

Fujimoto T, Ito S*, Ito M, Kanazawa H, Yamaguchi S.

Lasers in Surgery & Medicine 44(8)   685-94 2012年08月

研究論文（学術雑誌）, 共著, 査読有り

超高速液体クロマトグラフィーを用いるロキソプロフェンナトリウム錠溶出試験.

三浦 唯, 永田佳子, 金澤 秀子.

分析化学 61(8)   713-718 2012年08月

研究論文（学術雑誌）, 共著, 査読有り

Hypnotic effects and pharmacokinetics of a single bolus dose of propofol in Japanese macaques (Macaca fuscata fuscata).

Miyabe-Nishiwaki T, Masui K, Kaneko A, Nishiwaki K, Shimbo E, Kanazawa H.

Vet Anaesth Analg 39(4)   435 2012年07月

研究論文（学術雑誌）, 共著, 査読有り

Thermoresponsive Poly(N-isopropylacrylamide)-Based Block Copolymer Coating for Optimizing Cell Sheet Fabrication.

Nakayama M, Yamada N, Kumashiro Y, Kanazawa H, Yamato M, Okano T.

Macromol Biosci 12(6)   751-60 2012年06月

研究論文（学術雑誌）, 共著, 査読有り

High Stability of Thermoresponsive Polymer-Brush-Grafted Silica Beads as Chromatography Matrices.

Nagase K, Kobayashi J, Kikuchi A, Akiyama Y, Kanazawa H, Okano T.

ACS Appl Mater Interfaces 4(4)   1998-2008 2012年04月

研究論文（学術雑誌）, 共著, 査読有り

Effect of polymer containing a naphthyl-alanine derivative on the separation selectivity for aromatic compounds in temperature-responsive chromatography.

Nishio T, Kanazashi R, Nojima A, Kanazawa H*, Okano T.

J. Chromatogr A 1228:   148-54 2012年03月

研究論文（学術雑誌）, 共著, 査読有り

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pH-Induced Phase Transition Control of Thermoresponsive Nano-micelles Possessing Outermost Surface Sulfonamide Moieties.

Nakayama M, Kawahara Y, Akimoto J, Kanazawa H, Okano T.

Colloids Surf B: Biointerfaces 99   12-19 2012年01月

研究論文（学術雑誌）, 共著, 査読有り

Poly (N-isopropylacrylamide)-PLA and PLA blend nanoparticles for temperature-controllable drug release and intracellular uptake.

Ayano E, Karaki M, Ishihara T, Kanazawa H*, Okano T.

Colloids Surf B: Biointerfaces 99   67-73 2012年01月

研究論文（学術雑誌）, 共著, 査読有り

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Effect of Reaction Solvent on the Preparation of Thermoresponsive Stationary Phase through a Surface Initiated Atom Transfer Radical Polymerization.

Nagase K, Kanazawa H, Okano T*.

J Chromatogr A 1218(48)   8617-28 2011年12月

研究論文（学術雑誌）, 共著, 査読有り

Temperature-responsive chromatography for the separation of biomolecules.

Kanazawa H*, Okano T.

J Chromatogr A 1218(49)   8738-47 2011年12月

研究論文（学術雑誌）, 共著, 査読有り

Thermoresponsive Polymer Brush on Monolithic-Silica-Rod for the High-Speed Separation of Bioactive Compounds.

Nagase K, Kobayashi J, Kikuchi A, Akiyama Y, Kanazawa H, Okano T.

Langmuir 27(17)   10830-9 2011年09月

研究論文（学術雑誌）, 共著, 査読有り

Estimation of the postmortem duration of mouse tissue by electron spin resonance spectroscopy.

Ito S, Mori T, Kanazawa H, Sawaguchi T*.

Journal of Toxicology 2011   973172 2011年07月

研究論文（学術雑誌）, 共著, 査読有り

Reaction monitoring of tocopherols with active nitrogen oxides by ultra high-speed liquid chromatography.

Nagata Y, Nishio T, Kanazawa H.

J Pharm Biomed Anal 55   241-246 2011年05月

研究論文（学術雑誌）, 共著, 査読有り

Thermo-responsive protein adsorbing materials for purifying pharmaceutical protein on exposed charging surface.

Nagase K, Yuk SF, Kobayashi J, Kikuchi A, Akiyama Y, Kanazawa H, Okano T*.

J. Mater Chem 21   2590-2593 2011年04月

研究論文（学術雑誌）, 共著, 査読有り

Separation of phosphorylated peptides utilizing dual pH-and temperature-responsive chromatography.

Nishio T, Ayano E, Suzuki Y, Kanazawa H*, Okano T.

J Chromatogr A 1218(15)   2079-84 2011年04月

研究論文（学術雑誌）, 共著, 査読有り

LC-MS/MSによる畜水産物中のメトリブジンおよび代謝物の分析.

○Kai H, Akaboshi T, Waki M, Fuzimaki T, Kanazawa H.

食品衛生学雑誌 52(1)   28-33 2011年02月

研究論文（学術雑誌）, 共著, 査読有り

LC-MS/MSによる畜水産物中のメトリブジンおよび代謝物の分析.

甲斐茂美, 赤星　猛, 脇　ますみ, 藤巻照久, 金澤 秀子.

食品衛生学雑誌 52 （ 1 ） 28-33 2011年02月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Thermally-modulated on/off-adsorption materials for pharmaceutical protein purification.

Nagase K, Kobayashi J, Kikuchi A, Akiyama Y, Kanazawa H, Okano T.

Biomaterials 32(2)   619-627 2011年01月

研究論文（学術雑誌）, 共著, 査読有り

Thermally-modulated on/off-adsorption materials for pharmaceutical protein purification.

金澤 秀子

Biomaterials 32 （ 2 ） 619-627 2011年01月

研究論文（研究会，シンポジウム資料等）, 単著, 査読有り

超高速液体クロマトグラフィーを用いたビタミンE分析および抗酸化反応モニタリングへの応用.

○永田佳子, 西尾忠, 金澤秀子.

ビタミンE研究の進歩 （ビタミンE研究会）  14   125-129 2010年12月

共著

リポソームに封入したビタミンEの抗酸化効果の検討.

○蕭斐如, 永田佳子, 金澤秀子.

ビタミンE研究の進歩 （ビタミンE研究会）  14   87-91 2010年12月

共著

Preparation of thermo-responsive polymer brushes on hydrophilic polymeric beads by surface-initiated atom transfer radical polymerization for a highly resolutive separation of peptides.

Mizutani A, Nagase K, Kikuchi A, Kanazawa H, Akiyama Y, Kobayashi J, Annaka M, Okano T.

J Chromatogr A 1217(38)   5978-5985 2010年09月

研究論文（学術雑誌）, 共著, 査読有り

Preparation of thermo-responsive polymer brushes on hydrophilic polymeric beads by surface-initiated atom transfer radical polymerization for a highly resolutive separation of peptides.

金澤 秀子

J. Chromatogr. A 1217 （ 38 ） 5978-5985 2010年09月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Effective separation of peptides using highly dense thermo-responsive polymer brush-grafted porous polystyrene beads.

Mizutani A, Nagase K, Kikuchi A, Kanazawa H, Akiyama Y, Kobayashi J, Annaka M, Okano T.

J Chromatogr B 878(24)   2191-2198 2010年08月

研究論文（学術雑誌）, 共著, 査読有り

Effective separation of peptides using highly dense thermo-responsive polymer brush-grafted porous polystyrene beads.

金澤 秀子

J. Chromatogr. B 878 （ 24 ） 2191-2198 2010年08月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Hydration of poly(N-isopropylacrylamide) brushes on micro-silica beads measured by a fluorescent probe.

Hattori Y, Nagase K, Kobayashi J, Kikuchi A, Akiyama Y, Kanazawa H ,Okano T.

Chemical Physics Letters 491   193-198 2010年05月

研究論文（学術雑誌）, 共著, 査読有り

Hydration of poly(N-isopropylacrylamide) brushes on micro-silica beads measured by a fluorescent probe.

金澤 秀子

Chemical Physics Letters 491   193-198 2010年05月

研究論文（学術雑誌）, 共著, 査読有り

Thermo-responsive polymer brush surfaces with hydrophobic groups for all-aqueous chromatography.

Nagase K, Kumazaki M, Kanazawa H, Kobayashi J, Kikuci A, Akiyama Y, Annaka M, Okano T.

ACS Appl Mater Interfaces 2(4)   1247-53 2010年04月

研究論文（学術雑誌）, 共著, 査読有り

Thermo-responsive polymer brush surfaces with hydrophobic groups for all-aqueous chromatography.

金澤 秀子

ACS Appl Mater Interfaces. 2 （ (4) ） 1247-53 2010年04月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

多機能性ポリマーを用いる環境低負荷型クロマトグラフィーシステムの開発.

Nishio T, Kanazawa H*.

分析化学 59   163-173 2010年03月

研究論文（学術雑誌）, 共著, 査読有り

Thermo-responsive polymer brush-grafted porous polystyrene beads for all-aqueous chromatography.

Mizutani A, Nagase K, Kikuchi A, Kanazawa H, Akiyama Y, Kobayashi J, Annaka M, Okano T.

J Chromatogr A 1217   522-529 2010年01月

研究論文（学術雑誌）, 共著, 査読有り

Preparation of Thermoresponsive Anionic Copolymer Brush Surfaces for Separating Basic Biomolecules.

金澤 秀子

Biomacromolecules 11 （ 1 ） 215–223 2010年01月

研究論文（研究会，シンポジウム資料等）, 単著, 査読有り

Thermo-responsive polymer brush-grafted porous polystyrene beads for all-aqueous chromatography.

金澤 秀子

J. Chromatogr. A 1217 （ 4 ） 522-529 2010年01月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Separation of phosphorylated peptides utilizing dual pH-and temperature-responsive chromatography.

金澤 秀子

J. Chromatogr. A    in press 2010年

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Preparation of Thermoresponsive Anionic Copolymer Brush Surfaces for Separating Basic Biomolecules.

Nagase K, Kobayashi J, Kikuchi A, Akiyama Y, Kanazawa H, Annaka M, Okano T*.

Biomacromolecules 11(1)   215-223 2009年11月

研究論文（学術雑誌）, 共著, 査読有り

Hypnotic effects and pharmacokinetics of a single bolus dose of propofol in Japanese macaques (Macaca fsucata fsucata).

金澤 秀子

Vet Anaesth Analg. 37 （ 6 ） 501-510 2009年11月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Aqueous chromatographic system for the quantification of propofol in biological fluids using a temperature-responsive polymer modified stationary phase.

金澤 秀子

J. Chromatogr. A 1216 （ 44 ） 7427-7432 2009年10月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Dual Temperature- and pH-Responsive Fluorescence Molecular Probe for Cellular Imaging utilizing a PNIPAAm- Fluorescein Copolymer.

金澤 秀子

Anal. Sci. 25 （ 8 ） 1043-1047 2009年08月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Polymeric nanoparticles encapsulating betamethasone phosphate with different release profiles and stealthiness.

Ishihara T, Kubota T, Choia T, Takahashi M, Ayano E, Kanazawa H, Higaki M*.

Int J Pharm 375(1-2)   148-154 2009年06月

研究論文（学術雑誌）, 共著, 査読有り

Polymeric nanoparticles encapsulating betamethasone phosphate with different release profiles and stealthiness.

金澤 秀子

Int J Pharm. 375 （ 1-2 ） 148-154 2009年06月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Dual temperature- and pH-responsive fluorescence molecular probe for cellular imaging utilizing a PNIPAAm-fluorescein copolymer.

Kobayashi H, Nishikawa M, Sakamoto C, Nishio T, Kanazawa H*, Okano T.

Anal Sci 25   1043-1047 2009年06月

研究論文（学術雑誌）, 共著, 査読有り

Aqueous chromatographic system for the quantification of propofol in biological fluids using a temperature-responsive polymer modified stationary phase.

Nishio T, Suzuki R, Tsukada Y, Kanazawa H*, Okano T, Miyabe-nishiwaki T.

J Chromatogr A 1216   7427-7432 2009年04月

研究論文（学術雑誌）, 共著, 査読有り

Intracellular delivery of siRNA by cell-penetrating peptides modified with cationic oligopeptides.

Ishihara T, Goto M, Kodera K, Kanazawa H, Murakami Y, Mizushima Y, Higaki M.

Drug Delivery 16(3)   153-159 2009年04月

研究論文（学術雑誌）, 共著, 査読有り

分子内に塩基性部位を有するビタミンE誘導体の合成とラジカル消去活性.

中西郁夫, 川島知憲, 薬丸晴子, 乳井美奈子, 大久保敬, 金澤秀子, 福原潔, 奥田晴宏, 福住俊一, 小澤俊彦, 伊古田暢夫, 安西和紀.

ビタミンE研究の進歩 （ビタミンE研究会）  13   7-10 2009年03月

共著

Efficient entrapment of poorly water-soluble pharmaceuticals in hybrid nanoparticles.

Ishihara T, Goto M, Kanazawa H, Higaki M, Mizushima Y.

J. Pharm. Sci 98(7)   2357-2363 2009年03月

研究論文（学術雑誌）, 共著, 査読有り

ビタミンE異性体の併用効果の検討.

永田佳子, 藤岡孝允, 金澤秀子.

ビタミンE研究の進歩 （ビタミンE研究会）  13   97-101 2009年03月

共著

Influence of graft interface polarity on hydration/dehydration of grafted thermoresponsive polymer brushes and steroid separation using all-aqueous chromatography.

Nagase K, Kobayashi J, Kikuchi A, Akiyama Y, Annaka M, Kanazawa H, Okano T.

Langmuir 24(19)   10981-10987 2008年08月

研究論文（学術雑誌）, 共著, 査読有り

Antioxidant Activity of Tocopherols against Nitrogen Oxides by the Simultaneous Use of Toc Derivatives.

'Nagata Y*, Fujioka T, Kanazawa H.'

J Clin Biochem Nutr 43(Suppl.1)   436-439 2008年07月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Preparation of Thermoresponsive Polymer Brush Surfaces and Their Interaction with Cells.

'Mizutani A, Kikuchi A, Yamato M, Kanazawa H, Okano T.'

Biomaterials 29 （ 13 ） 2073-2081 2008年05月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Aqueous Chromatographic system for Separation of Biomolecules using Thermoresponsive Polymer Modified Stationary Phase.

'Kanazawa H*, Nishikawa H, Mizutani A, Sakamoto C, Morita-Murase Y, Nagata Y, Kikuchi A, OkanoT.'

J. Chromatogr. A 1191 （ 1-2 ） 157-161 2008年05月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Preparation of Thermoresponsive Cationic Copolymer Brush Surfaces and Application of the Surface to Separation of Biomolecules.

Nagase K, Kobayashi J, Kikuchi A, Akiyama Y, Kanazawa H, Okano T.

Biomacromolecules 9(4)   1340-1347 2008年04月

研究論文（学術雑誌）, 共著, 査読有り

Cu(II) Chelates of an Intermediate Species Absorbing in the 500-nm Region in Nonenzymatic Pyridoxal Catalysis.

'Nagata Y, Matsushima Y, Kanazawa H.'

J Kyoritsu Univ Pharm Vol.4   9-16 2008年03月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Effects of graft densities and chain lengths on separation of bioactive compounds by nanolayered thermoresponsive polymer brush surfaces.

'Nagase K, Kobayashi J, Kikuchi A, Akiyama Y, Kanazawa H, Okano T.'

Langmuir 24 （ 2 ） 511-517 2008年01月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

畜水産物中動物用医薬品スクリーニング分析法としての高速液体クロマトグラフィー/タンデム質量分析法の検討.

'甲斐茂美,赤星猛,藤巻照久,伊藤伸一,金澤秀子.'

分析化学 56(12)   56(12) 2007年12月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Differential effects of the ascorbyl and tocopheryl derivative on the methamphetamine-induced toxic behavior and toxicity.

'Ito S, Mori T, Kanazawa H, Sawaguchi T.'

Toxicology 240(1-2)   96-110 2007年10月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Interfacial property modulation of thermoresponsive polymer brush surfaces and their interaction with biomolecules.

'Nagase K, Kobayashi J, Kikuchi A, Akiyama Y, Kanazawa H, Okano T.'

Langmuir 23(18)   9409-15 2007年08月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Thermally responsive chromatographic materials using functional polymers.

Kanazawa H.

J Sep Sci （/Wiley-VCH）  30(11)   1646-1656 2007年07月

研究論文（学術雑誌）, 単著, 査読有り

環境応答性高分子を用いた新規分離システム.

'金澤将史, 南部杏子, 鈴木祐輔, 西川真由美, 金澤秀子*.'

分析化学(Bunseki Kagaku) （/日本分析化学会）  56(6)   397-407 2007年06月

研究論文（学術雑誌）, 共著, 査読有り

Analysis of Melatonin using a pH- and Temperature-responsive Aqueous Chromatographic System.

'Ayano E, Suzuki Y, Kanezawa M, Sakamoto C, Morita-Murase Y, Nagata Y, Kanazawa H*, Kikuchi A, Okano T.'

J Chromatogr A 1156   213-219 2007年06月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

機能性高分子の分子設計と分離及び薬物送達システムへの応用.

金澤秀子.

ファルマシア （/(社)日本薬学会）  43   307-309 2007年04月

研究論文（学術雑誌）, 単著

A developed determination of midazolam and 1'-hydroxymidazolam in plasma by liquid chromatography-mass spectrometry: Application to human pharmacokinetic study for measurement of CYP3A activity.

'Shimizu M, Uno T*, Tamura HO, Kanazawa H, Murakami I, Tateishi T.'

J Chromatogr B 847 (2)   275-281 2007年03月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

薬学教育年限延長(6年制)に伴う共立薬科大学生涯学習の動向調査研究.

'藤本 和子*, 荒井 範子, 柏崎 あや, 金澤 秀子, 望月 正隆.'

共立薬科大学雑誌 2   2 2007年03月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

'Metabolism of bisphenol A in the rat syncytiotrophoblast cell line, TR-TBT 18d-1.'

'Ikeda Y, Nakashima E, Kanazawa H, Tamura H*.'

J Health Sci 53   146-150 2007年02月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

ビタミンE類縁体におけるラジカル消去反応に対する塩基触媒作用.

'中西郁夫, 川島知憲, 大久保敬, 金澤秀子, 稲見圭子, 望月正隆, 福原潔, 奥田晴宏, 小澤俊彦, 伊東忍, 福住俊一, 伊古田暢夫.'

ビタミンE研究の進歩 （/ビタミンE研究会）  ⅩⅡ   11-16 2006年12月

研究論文（学術雑誌）, 共著

Study of a Capillary LC System using Temperature-Responsive Polymer-Modified Packing Materials.

'Kanezawa M, Ayano E, Kanazawa H*, Akiyama Y, Okano T.'

CHROMATOGRAPHY 27   111-115 2006年12月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

窒素酸化物との反応におけるビタミンE異性体併用における効果の「検討.

'藤岡孝允, 石川奈都子, 笠井愛子, 永田佳子, 金澤秀子.'

ビタミンE研究の進歩 （/ビタミンE研究会）  ⅩⅡ   17-21 2006年12月

研究論文（学術雑誌）, 共著

ペットボトルのお茶でアルツハイマー病を予防する.

'○村上勲, 小西茉美, 渡邉まどか, 永田佳子, 森田裕子, 金澤秀子.'

日本分析化学会第５５年会　展望とトピックス （/社団法人　日本分析化学会）  　   8 2006年09月

研究論文（学術雑誌）, 共著

Pharmacokinetics of fentanyl after single intravenous injection and constant rate infusion in dogs.

'Sano T, Nishimura R, Kanazawa H, Igarashi E, Nagata Y, Mochizuki M, Sasaki N.'

Vet Anaesth Analg 33(4)   266-273 2006年07月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Aqueous chromatography system using temperature-responsive polymer modified stationary phase.

'Ayano E, Kanazawa H*.'

J Sep Sci （/WILEY-VCH）  29   738-749 2006年06月

研究論文（学術雑誌）, 共著, 査読有り

Study of temperature-responsibility on the surfaces of a thermo-responsive polymer modified stationary phase.

'Ayano E, Okada Y, Sakamoto C, Kanazawa H*, Kikuchi A, Okano T.'

J Chromatogr A 1119/1-2   51-57 2006年06月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Aqueous chromatography system using pH- and temperature-responsive stationary phase with ion-exchange groups.

'Ayano E, Nambu K, Sakamoto C, Kanazawa H*, Kikuchi A, Okano T.'

J Chromatogr A 1119(1-2)   58-65 2006年06月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

環境応答性ポリマーを用いた水系クロマトグラフィー.

'綾野絵理, 金澤秀子*.'

化学工業 （/化学工業社）  57（5）   13-19（341-347） 2006年05月

研究論文（学術雑誌）, 共著

'Increased F2-Isoprostane Levels in the Rat Brain and Plasma Caused by Oxidative Stress and Aging, and Inhibitory Effect of Vitamin E.'

'Nishio T, Miyadera R, Sakai R, Abe K, Kanazawa H, Fukui K, Urano S.'

J Clin Biochem Nutr 38   161-166 2006年05月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Separation of Nucleotides with an Aqueous Mobile Phase Using pH- and Temperature-Responsive Polymer Modified Packing Materials.

'Ayano E, Sakamoto C, Kanazawa H*, Kikuchi A, Okano T.'

Anal Sci 22   539-543 2006年04月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Temperature-responsive stationary phase utilizing a polymer of prorine derivative for hydrophobic interaction chromatography using an aqueous mobile phase.

'Kanazawa H*, Ayano E, Sakamoto C,Yoda R, Kikuchi A, OkanoT.'

Journal of chromatography A 1106   152-158 2006年03月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Simultaneous determination of catechins and procyanidins in bottled tea drinks by LC/MS.

'Murakami I,, Nakamura T, Ishibashi Y, Shibuya R,, Ayano E,, Morita-Murase Y, Nagata Y, Kanazawa H*.'

Chromatography 27   19-25 2006年03月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

環境センサーを備えた高分子による環境にやさしいクロマトグラフィー.

'綾野絵理,南部杏子,飯塚元気,坂本千賀子,金澤秀子,菊池明彦,岡野光夫.'

日本薬学会第126年会　講演ハイライト （/（社）日本薬学会）  　   ｐ25 2006年03月

研究論文（学術雑誌）, 共著

Temperature-responsive stationary phase utilizing a polymer of proline derivative for hydrophobic interaction chromatography using an aqueous mobile phase.

'Kanazawa H*, Ayano E, Sakamoto C, Yoda R, Kikuchi A, Okano T.'

J Chromatogr A 1106   152-158 2006年02月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

カテキンと新カテキン.

金澤秀子.

'Clinical Engineering,' （/秀潤者）  16(12)   1241 2005年12月

研究論文（学術雑誌）, 単著

温度応答性充填剤を利用した新たな手の平サイズの分離システムの開発.

'飯塚元気,金澤将史,綾野絵理,金澤秀子,小林純,菊池明彦,岡野光夫.'

日本分析化学会第54年会　展望とトピックス （/(社）日本分析化学会）  　　   p14 2005年09月

研究論文（学術雑誌）, 共著

LC/MS/MSによる畜産物中のベンズイミダゾール系寄生虫駆虫剤の分析.

'甲斐茂美, 赤星猛, 岸美智子, 金澤秀子, 小林静子.'

分析化学 54（9） 2005年09月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

機能性高分子を用いた温度応答性クロマトグラフィーの開発.

'綾野絵理, 金澤秀子*.'

分析化学 （/（社）日本分析化学会）  54（7）   593-603 2005年07月

研究論文（学術雑誌）, 共著, 査読有り

薬剤師のための生涯学習支援システム.

金澤秀子.

薬剤学 （/日本薬剤学会）  65（6）   334-337 2005年06月

研究論文（学術雑誌）, 単著

環境応答性高分子を用いた新規分離システムの開発.

金澤 秀子.

薬学研究の進歩 （/薬学研究奨励財団）  21   19-24 2005年04月

研究論文（学術雑誌）, 単著, 査読有り

Electron-transfer mechanism in radical-scavenging reactions by a vitamin E model in a protic medium.

'Nakanishi I, Kawashima T, Ohkubo K, Kanazawa H, Inami K, Mochizuki M, Fukuhara K, Okuda H, Ozawa T, Itoh S, Fukuzumi S, Ikota N.'

Org Biomol Chem 3(4)   626-629 2005年04月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Analysis of herbicides in water using temperature-responsive chromatography and an aqueous mobile phase.

'Ayano E, Okada Y, Sakamoto C, Kanazawa H*, Okano T, Ando M, Nishimura T.'

J Chromatogr A 1069   281-285 2005年04月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

未来を拓くナノ医療.

金澤秀子.

日本薬学会第125年会講演ハイライト （/日本薬学会）  　   48 2005年03月

研究論文（学術雑誌）, 単著

機能性食品としてペットボトル緑茶飲料を考える.

'村上勲, 石橋悠紀子, 永田佳子, 森田裕子, 金澤秀子.'

日本薬学会　第125年会　講演ハイライト （/社団法人　日本薬学会　広報委員会）  　   27 2005年03月

研究論文（学術雑誌）, 単著

環境応答性高分子を用いた新規分離システムの開発.

金澤秀子.

薬学研究の進歩 （/(財）　薬学研究奨励財団）  21   19-24 2005年03月

研究論文（学術雑誌）, 単著

未来を拓くナノ医療.

金澤秀子.

薬事日報 （/薬事日報社）  10045   10 2005年03月

研究論文（学術雑誌）, 単著

新刊紹介 ゲノムと疾患.

金澤秀子.

ファルマシア （/(社）日本薬学会）  41（1）   93 2005年01月

研究論文（学術雑誌）, 単著

α-Tocopherol とγ-Tocopherolの共存、非共存下での窒素酸化物との反応における反応場および活性種の影響.

'永田佳子, 松本洋太, 須原幸恵, 寺岡奈那恵, 金澤秀子.'

ビタミンE研究の進歩 （/びたみんＥ研究会）  XI   144-149 2004年12月

研究論文（学術雑誌）, 単著

Water-Accelerated Radical-Scavenging Reaction of (+)-Catechin in an Aprotic Medium.

'Nakanishi I*, Kawashima T, Fuikuhara K, Kanazawa H, Okuda H, Fukuzumi S, Ozawa T, Ikota N*.'

ITE Lett. Batt. New Tech. Med. 5 (6)   585-588 2004年12月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Radical-Scavenging Activities of Lipophilic Planar Catechin Derivatives.

'Nakanishi I, Ikota N, Kawashima T, Yakumaru H, Kanazawa H, Okuda H, Anzai K, Ozawa T, Fukuhara K.'

Free Radical Biol. Med. （/the Society for Free Radical Biology and Medicine）  36 (S1)   ｐS40 2004年11月

研究論文（学術雑誌）, 共著

Solvent Effect on the Mechanism of Radical-Scavenging Reactions of a Vitamin E Analogue.

'Nakanishi I,Kawashima T,Ohkubo K, Kanazawa H, Okuda H, Ozawa T, Fukuzumi S, Fukuhara K, Ikota N.'

Free Radical Biol. Med. （/the Society for Free Radical Biology and Medicine）  36 (S1)   ｐ S39-S40 2004年11月

研究論文（学術雑誌）, 共著

薬物相互作用の予測のためのin vitro 代謝阻害実験.

金澤秀子.

都薬雑誌 （/(社）東京都薬剤師会）  26（10）   12-18 2004年10月

研究論文（学術雑誌）, 単著

酸化ストレスに対するビタミンE異性体の併用効果の検討.

永田佳子 石倉彩恵子 前夷香織 松下奈々恵 金澤秀子.

日本分析化学会第53年会・展望とトピックス （/(社）日本分析化学会）  第53年会版   ｐ11 2004年09月

研究論文（学術雑誌）, 共著

大学における分析専攻学生の育成.

金澤秀子.

日本分析化学会関東支部ニュース （/（社）日本分析化学会関東支部）  第15号   6 2004年09月

研究論文（学術雑誌）, 単著

分けるということ.

金澤秀子.

LC talk （/島津製作所）  55   2 2004年07月

研究論文（学術雑誌）, 単著

ビタミンＥモデルのラジカル消去メカニズムに対する溶媒の影響.

'川島知憲,中西郁夫,金澤秀子,小澤俊彦,伊古田暢夫.'

第４回AOB(Antioxidant Biofactor)研究会プロシーディング （/AOB(Antioxidant Biofactor)研究会）  4   p 2004年06月

研究論文（学術雑誌）, 共著

温度応答性クロマトグラフィー.

金澤秀子.

ぶんせき （/（社）日本分析化学会）  6   303-308 2004年06月

研究論文（学術雑誌）, 単著

Radical-Scavenging Reactions of 4-Hydroxycinnamic Acid Derivatives in the Presence of Metal Ions.

'Nakanishi I, Uto Y, Kawashima T,Yakumaru H, Kanazawa H, Ozawa H, Nagasawa H, Hori H, Ikota N.'

J. Pharm. Soc. Jpn. （/日本薬学会）  124 (Suppl. 1)   94 2004年06月

研究論文（学術雑誌）, 共著

Effects of Metal Ions on the Stability of Oxygen and Nitrogen Radicals.

'Nakanishi I, Kawashima T, Kanazawa K, Ohkubo K, Fukuhara K, Ozawa T, Fukuzumi S, Ikota N.'

J. Pharm. Soc. Jpn. （/日本薬学会）  124 (Suppl. 1)   p95 2004年06月

研究論文（学術雑誌）, 共著

Products of the reaction between a- or g-tocopherol and nitrogen oxides analyzed by high-performance liquid chromatography with UV-Vis and atmospheric pressure chemical ionization mass spectrometric detection.

'Nagata Y, Matsumoto Y, Kanazawa H*.'

Journal of Chromatography A 1036   177-182 2004年04月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

機能性高分子を用いた温度応答性クロマトグラフィーによるタンパク分離.

金澤秀子.

Hitachi Scientific Instrument News （/（株）日立ハイテクノロジーズ）  46   7-11 2004年03月

研究論文（学術雑誌）, 単著

Determination of midazolam and it's metabolite as a probe for CYP3A4 phenotype by liquid chromatography-mass spectrometry.

'Kanazawa H*, Okada A, Igarashi E, Higaki M, Miyabe T, Sano T, Nishimura R.'

J Chromatogr A 1031   213-218 2004年02月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Determination and quantitation of sulfonylurea and urea herbicudes in water samples using liquid chromatography with electrospray ionization mass spectrometric detection.

'Ayano E, Kanazawa H, Ando M, Nishimura T.'

Analytica Chimica Acta 507   215-222 2004年01月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Temperature-responsive polymers for liquid-phase separations.

Kanazawa H*.

Anal Bioanal Chem 378   46-48 2004年01月

研究論文（研究会，シンポジウム資料等）, 単著, 査読有り

Temperature- and pH-responnsive aminopropylsilica ion-exchange columns grafted with copolymers of N-isopropylacrylamide.

'Sakamoto C, Okada Y,Kanazawa H*, Ayano E, Nishimura T, Ando M,Kikuchi A, Okano T.'

J Chromatogr A 1030   247-253 2004年01月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

温度応答性クロマトグラフィーによるカテキン類の分離.

'坂本千賀子,岡田裕司,金澤秀子*,菊池明彦,岡野光夫.'

分析化学 52   52 2003年10月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

緑茶飲料中カテキン類のCYP3A4による代謝系への阻害効果の解析.

'中村拓己,浅田絵美,永田佳子,金澤秀子*.'

分析化学 52   52 2003年09月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Stereospecific analysis of omeprazole in human plasma as a probe for CYP2C19 phenotype.

'Kanazawa H*, Okada A, Higaki H, Yokota H, Mashige F, Nakahara K.'

J Pharm Biomed Anal 30(6)   195-202 2003年01月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

α-TocopherolとNOの酸素存在下での反応における溶媒効果について.

'永田佳子, 金澤秀子, 望月正隆, 松島美一, 松本茂信.'

ビタミンE研究と進歩Ⅹ （/ビタミンE研究会）  10   1-6 2002年12月

研究論文（学術雑誌）, 共著

Stereospecific Analysis of Loxoprofen in Plasma by Chiral Column Liquid Chromatography with A Circular Dichroism -Based Detector.

'Kanazawa H*, Tsubayashi H, Nagata Y, Matsushima Y, Mori C, Kizu J, Higaki H.'

J Chromatogr A 948   303-308 2002年03月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

'Determination of omeprazole and its metabolites in human plasma by liquid chromatography-mass spectrometry,.'

'Kanazawa H*, Okada A, Matsushima Y, Mashige F, Okubo S, Yokota H, Nakahara K.'

J Chromatogr A 949   1-9 2002年03月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Temperature-Responsive Chromatography Using Poly(N-isopropylacrylamide) Hydrogel-Modified Silica.

'Kanazawa H*, Sunamoto T, Ayano E, Matsushima Y, Kikuchi A, OKANO T.'

Anal Sci 18   45-48 2002年01月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Environmental Responsive Chromatography.

'Ayano E, Kanazawa H*, Kikuchi A, OKANO T.'

Anal Sci 17   I873-I874 2001年12月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Determination of Omeprazole and its Metabolites in Human Plasma as a Probe For CYP2C19 Phenotype.

'Okada A, Kanazawa H*, Mashige F, Okubo S, Yokota H.'

Anal Sci 17   871-873 2001年09月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Temperature-Responsive Chromatography-Effect of Temperature Gradient-.

'Kanazawa H*, Ayano E, Chiba K, Kikuchi A, OKANO T.'

Anal Sci 17   875-876 2001年09月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Enantiometric determination of L- and D-lactatic acid in human cerebrospinal fluid by chiral ligand exchange high-performance liquid chromatography.

'Okubo S, Mashige F, Omori M, Hashimoto Y, Nakahara K, Kanazawa H, Matsushima Y.'

Biomed. Chromatogr. 14   474-477 2000年12月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Temperature-Responsive Chromatographic Separation of Amino Acid Phenylthiohydantoins using Aqueous Media as the Mobile Phase.

'Kanazawa H*, Sunamoto T, Matsushima Y, Kikuchi A, Okano T.'

Anal.Chem. 72   5961-5966 2000年12月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

'Reaction of 2,2,5,7,8-pentamethyl-6-chromanol, an α-tocopherol analogue, with NO in the presence of oxygen.'

'Nagata Y*, Nishio T, Matsumoto S, Kanazawa K, Mochizuki M, Matsushima Y.'

Bioorg Med Chem Lett 10   2709-2712 2000年10月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Determination of a-Tocopherol and a-Tocopherylquinone in Rat Tissues and Plasma by High-Performance Liquid Chromatography with Electrochemical Detection.

'Kanazawa H*, Miyata C, Nagata Y, Urano S, Matsushima Y.'

Chem.Pharm.Bull. 48(10)   1462-1466 2000年09月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

ＨＰＬＣによるテオフィリンとその代謝物の同時分析.

'金澤秀子,木津純子,松島美一.'

YAKUGAKU ZASSHI （/日本薬学会）  120(10)   1051-1060 2000年08月

研究論文（学術雑誌）, 共著, 査読有り

Preparation of Packing Materials for Temperature-Responsive Chromatography. Modification of Silica Surface with Poly(N-isopropylacrylamide).

'Yamamoto K, Kanazawa H*, Matsushima Y, Takai N, Kikuchi A, Okano T.'

Chromaotgraphy 21(3)   209-215 2000年07月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

D-Serine Concentration in Blood Serum of Liver Transplanted Patients.

'Okubo S, Mashige F, Hashimoto Y, Nakahara K, Kanazawa H, Matsushima Y,Fukushima T, Huang Y, Imai K.'

Chromaotgraphy 21(1)   43-47 2000年04月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Determination of theophylline and its metabolites in biological samples by liquid chromatography-mass spectrometry.

'Kanazawa H*, Atsumi R, Matsushima Y, Kizu J.'

J. Chromatography A 870   87-96 2000年03月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

カチオン性温度応答表面による酸性生理活性物質の分離.

'小林純,中山幸子,菊池明彦,金澤秀子,松島美一,酒井清孝,岡野光夫.'

医工学治療 12(1)   12(1) 2000年03月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Spectrophotometric analysis of lorazepam in plasma by chiral chromatography-circular dichroism (CD) based detector.

'Kanazawa H*, Kunito Y, Matsushima Y, Okubo S, Mashige F.'

J. Chromatography A 871   181-188 2000年03月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Temperature-Responsive Chromatographic Separation of Bisphenol A with Water as a Sole Mobile Phase.

'K,Yamamoto, H,Kanazawa*, Y,Matsushima, K, Oikawa, A, Kikuchi, T, Okano.'

Environmental Sciences 7(1)   47-56 2000年01月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り

Species Absorbing in the 500-nm Region in the Reactions of Pyridoxamine with Pyrroloquinoline quinone and Phenathroline quinones.

'Nagata Y, Matsushima Y, Fujii N, Fukuda S, Kanazawa H, Yoda R, Matsumoto S.'

Chem Pharm Bull 47(12)   1774-1777 1999年12月

研究論文（研究会，シンポジウム資料等）, 単著, 査読有り

Stereospecific Analysis Of Chiral Drugs In Plasma By Chromatography-Chiroptical Detector.

'Kanazawa H*, Kunito Y, Matsushima Y, Okubo S, Mashige F.'

Chromatography 21   81-87 1999年01月

研究論文（研究会，シンポジウム資料等）, 共著, 査読有り