Thermoresponsive chromatography with regulated electrostatic interactions for effective separation of nucleic acids

具有受控静电相互作用的热响应色谱可有效分离核酸

• 批准号: 13680955
• 负责人: KIKUCHI Akihiko
• 金额: $ 1.22万
• 依托单位: Tokyo Women's Medical University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13680955/
• 关键词: thermoresponsive polymers; poly(N-isopropylacrylamide); cationic groups; chromatography; oligonucleotide; nucleoside base; hydrophobic interaction; electrostatic interaction; 温度応答性ポリマー; 表面荷電密度; 温度応答性クロマトグラフィー; 核酸アナログ; 相転移挙動

This research focused on the design and characterization of novel thermoresponsive chromatography system in which the surface characteristics can change hydrophilic/hydrophobic properties and, at the same time, surface charge densities by temperature as a sole variant in aqueous milieu. Such surfaces was prepared for separation of bioactive nucleic acids efficiently without use of several chromatography columns working with different separation modes. We have already elucidated the surface property alterations of the cationic thermoresponsive poly(N-isopropylacrylamide) (PIPAAm) modified surfaces with temperature in terms of hydrophobicity changes and surface potential changes. Furthermore, separation of adenosine nucleotides, AMP, ADP, and ATP was successfully achieved with this column matrix. In this fiscal year, separation of adenosine oligonucleotides and thymidine oligonucleotides with repeating number of 2-5, were investigated using cationic, thermoresponsive column. Thermorespon … More sive column matrices were prepared by radical polymerization of IPAAm monomer and 3,3-dimethylaminopropylacrylamide (DMAPAA) on the initiator immobilized silica beads. The sample mixture was injected into polymer-modified silica packed column (4.6mmφ, 150mm length), and elution behavior of these nucleotides were monitored. For nonionic PIPAAm homopolymer column, retention times of the oligonucleotides are almost same regardless of the number of repeating units. In contrast, retention of oligonucleotides is increased with increasing number of repeating units on cationic, thermoresponsive surfaces. Polarity of the nucleotide bases is significantly important to alter oligonucleotide retention on cationic thermoresponsive column matrices. Thus the retention times of the thymidine oligonucleotides are slightly longer than those of the adenosine nucleotides with the same amount of the phosphate groups. The interaction of oligonucleotides and cationic surfaces is multi-point interaction of polyelectrolytes, and thus the retention increased with increasing the number of repeating units. In conclusion, the obtained results suggest that the prepared cationic, thermoresponsive surfaces can be used to separate oligonucleotides through the recognition of charge densities and hydrophobicity of sample molecules. Less

本研究的重点是设计和表征一种新型的热响应色谱系统，在这种系统中，表面特性可以改变水环境中材料的亲疏水性，同时表面电荷密度随温度的变化而变化。制备的这种表面可以有效地分离生物活性核酸，而不需要使用几种不同分离模式的色谱柱。我们已经从疏水性变化和表面电位变化两方面阐明了阳离子热敏性聚n -异丙基丙烯酰胺（PIPAAm）修饰表面的表面性质随温度的变化。此外，该柱基质成功地分离了腺苷核苷酸、AMP、ADP和ATP。本会计年度，采用阳离子热响应色谱柱研究了重复数为2-5的腺苷寡核苷酸和胸苷寡核苷酸的分离。用IPAAm单体和3,3-二甲氨基丙基丙烯酰胺（DMAPAA）在引发剂固定的硅胶珠上进行自由基聚合，制备了体积更大的柱基质。将样品混合物注入聚合物修饰的硅胶填充柱（4.6mmφ， 150mm长）中，监测这些核苷酸的洗脱行为。对于非离子型PIPAAm均聚物柱，无论重复单元数多少，寡核苷酸的保留时间几乎相同。相反，寡核苷酸的保留随着在阳离子、热响应表面上重复单元数量的增加而增加。核苷酸碱基的极性对于改变寡核苷酸在阳离子热响应柱基质上的保留是非常重要的。因此，胸腺嘧啶寡核苷酸的保留时间比具有相同数量磷酸基团的腺苷核苷酸的保留时间稍长。寡核苷酸与阳离子表面的相互作用是聚电解质的多点相互作用，因此保留率随着重复单元数目的增加而增加。综上所述，所得结果表明，制备的阳离子热响应表面可以通过识别样品分子的电荷密度和疏水性来分离寡核苷酸。少

项目成果

Chikako SAKAMOTO, Yuji OKADA, Hideko KANAZAWA, Eri AYANO, T.NISHIMURA, M.ANDO, Akihiko KIKUCHI, Teruo OKANO: "Temperature-and pH-responsive aminopropyl silica ion-exchange columns grafted with copolymers of N-isopropylacrylamide"Journal of Chromatography

Chikako SAKAMOTO、Yuji OKADA、Hideko KANAZAWA、Eri AYANO、T.NISHIMURA、M.ANDO、Akihiko KIKUCHI、Teruo OKANO：“接枝有 N-异丙基丙烯酰胺共聚物的温度和 pH 响应型氨丙基二氧化硅离子交换柱”Journal of Chromatography

坂本千賀子, 岡田裕司, 金澤秀子, 菊池明彦, 岡野光夫: "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"分析化学. 52(10). 903-906 (2003)

Chikako Sakamoto、Yuji Okada、Hideko Kanazawa、Akihiko Kikuchi、Mitsuo Okano：“通过温度响应色谱分离儿茶素”52(10) 903-906 (2003)。

C.Sakamoto.Y.Okada.H.Kanazawa, E.Ayano, T.Nishimura, M.Ando, A.Kikuchi, T.Okano: "Temperature- and pH-responsive aminopropyl silica ion-exchange columns grafted with copolymers of N-isopropylacryiamide"Journal of Chromatography A. 1030. 247-253 (2004)

C.Sakamoto.Y.Okada.H.Kanazawa、E.Ayano、T.Nishimura、M.Ando、A.Kikuchi、T.Okano：“接枝有 N 共聚物的温度和 pH 响应型氨丙基二氧化硅离子交换柱

Chikako SAKAMOTO, Yuji OKADA, Hideko KANAZAWA, Akihiko KIKUCHI, Teruo OKANO: "Separation of catechins by temperature-responsive chromatography"BUNSEKI KAGAKU. 52(10). 903-906 (2003)

Chikako SAKAMOTO、Yuji OKADA、Hideko KANAZAWA、Akihiko KIKUCHI、Teruo OKANO：“通过温度响应色谱法分离儿茶素”BUNSEKI KAGAKU。

Akihiko KIKUCHI: "Intelligent interfaces for tissue engineering"Biomaterials-SEITAI ZAIRYO-(Journal of Japanese Society for Biomaterials). 21(3). 214-215 (2003)

Akihiko KIKUCHI：“组织工程的智能界面”生物材料-SEITAI ZAIRYO-（日本生物材料学会杂志）。