JP2004224828A - New polymer capable of molecular recognition and its preparation method - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a new polymer capable of molecular recognition which selectively binds to a target molecule, has a photosensitivity inducing its physical/chemical modification when exposed to light and enables its adsorption to and desorption from the target molecule to be controlled by light, and a method for preparing the polymer through molecular imprinting. <P>SOLUTION: The polymer has a functional group which specifically and reversibly binds to the target molecule and a photosensitive moiety which isomerizes when exposed to light. The method for preparing the polymer comprises, e.g. a step wherein an unsaturated monomer having the functional group which specifically forms a reversible bond with the target molecule and the photosensitive moiety is prepared, a step wherein the unsaturated monomer is bound to the target molecule via the reversible bond, a step wherein the unsaturated monomer bound to the target molecule is copolymerized with a crosslinking agent and a step wherein the target molecule is detached from the obtained polymer. <P>COPYRIGHT: (C)2004,JPO&NCIPI

Description

【００３０】
架橋剤としては、付加重合可能な基を複数有するものが好ましい。付加重合可能な基を複数有する架橋剤の例としては、式（４）に示すｐ−ジビニルベンゼン、式（１８）に示すｏ−ジビニルベンゼン、式（１９）に示すｍ−ジビニルベンゼン等のジビニルベンゼン系架橋剤、（メタ）アクリル酸系架橋剤等がある。（メタ）アクリル酸系架橋剤としては、（メタ）アクリロイル基を複数有する化合物、例えば、式（１）に示すエチレングリコールジメタクリレート（ＥＤＭＡ）、式（２）に示すトリメチロールプロパントリメタクリレート（ＴＲＩＭ）、式（３）に示すトリメチロールプロパントリアクリレート、式（６）に示すテトラエチレングリコールジアクリレート、式（１１）に示すトリエチレングリコールジメタクリレート、式（１２）に示すペンタエリスリトールテトラアクリレート、式（１３）に示すペンタエリスリトールトリアクリレート、式（１４）に示すビスフェノールＡジメタクリレート、式（１５）に示すＮ，Ｎ’−メチレンジアクリルアミド、式（１６）に示すＮ，Ｎ’−１，４−フェニレンジアクリルアミド、式（１７）に示す３，５−ビス（アクリロイルアミド）安息香酸、式（２０）に示すＮ，Ｏ−ビスアクリロイル−Ｌ−フェニルアラニノール等が好適である。架橋剤として、ビニル基を１つだけ持つ化合物（式（５）に示す塩化ビニル、式（７）に示すスチレン、式（９）に示すフェニルビニルスルホキシド、式（１０）に示すフェニルビニルスルホン等）や、（メタ）アクリロイル基を１つだけ持つ化合物（例えば式（８）に示すメタクリル酸メチル等）等のような重合可能な基を１つだけ持つ化合物を用いてもよい。架橋剤としては、不飽和単量体が有する重合可能な基と同じ基を持つ架橋剤を使用することが好ましい。また、架橋剤は、結合部位同士のクロストークを避けるために不飽和単量体に対して大過剰で用いることが好ましい。具体的には、不飽和単量体１モルに対して架橋剤を５モル以上用いることが好ましい。
【００３１】
上記脱離工程では、重合工程で得られた重合体における官能基と標的分子との可逆的な結合を分解することにより、標的分子を脱離させる。例えば、可逆的な結合がエステル結合である場合には、エステル結合を酸加水分解あるいはアルカリ加水分解することにより標的分子を脱離させればよい。
【００３２】
〔非共有結合性分子認識ポリマーの製造方法〕
上記非共有結合性分子認識ポリマーは、いわゆる非共有結合型分子インプリンティングにより製造できる。すなわち、上記非共有結合性分子認識ポリマーは、上記非共有結合性官能基および光感応性部位を有する不飽和単量体を調製する調製工程と、上記不飽和単量体を、標的分子の存在下で架橋剤と共重合し、重合体を得る重合工程と、得られた重合体から標的分子を脱離させる脱離工程とを含む方法により製造できる。
【００３３】
上記不飽和単量体は、標的分子の持つ官能基と可逆的な非共有結合（水素結合など）を形成しうる官能基と、重合可能な不飽和結合と、光感応性部位とを持つ化合物である。標的分子の持つ官能基と可逆的な非共有結合を形成しうる官能基としては、標的分子の持つアミノ基と水素結合を形成しうるカルボニル基や、標的分子の持つ含窒素複素環の窒素原子と水素結合を形成しうる水酸基やアミノ基等が挙げられる。
【００３４】
上記調製工程では、例えば、光感応性部位を有する化合物にビニル基やメタクリロイル基等の付加重合可能な不飽和基をつけて付加重合可能とし、さらに標的分子と可逆的に非共有結合を形成しうる非共有結合性官能基（カルボキシル基、アミノ基、フェノール性水酸基等）をつけて付加重合可能な不飽和単量体を調製すればよい。
【００３５】
上記重合工程は、基本的には共有結合性分子認識ポリマーの製造方法と同様であるが、非共有結合型分子インプリンティングでは、標的分子と不飽和単量体との複合体を安定的に存在させるために、不飽和単量体は標的分子に対して過剰に用いることが好ましい。具体的には、標的分子１モルに対して標的分子を２モル以上用いることが好ましい。また、非共有結合が水素結合である場合、使用する溶媒としては、水素結合を妨害しないクロロホルム等のハロゲン系溶媒が好ましい。
【００３６】
上記脱離工程では、標的分子を選択的に溶解しうる溶液（例えばアルコール系溶媒等）で洗浄することにより、標的分子を脱離させる。これにより、非共有結合性分子認識ポリマーが得られる。
【００３７】
上記の非共有結合型分子インプリンティングにおいては、形成される結合部位の不均一性が問題となることがある。通常、標的分子と不飽和単量体（機能性モノマー）との複合体を形成させるために，不飽和単量体（機能性モノマー）は、標的分子に対して過剰に加えて重合する。当然ではあるが、結合部位に関与する不飽和単量体（機能性モノマー）は限られており、加えられた不飽和単量体（機能性モノマー）のうちの過剰分は、特異的な結合部位にはならず、非特異的な結合となる。すなわち、過剰分は、特異的結合部位と無関係に存在するか、あるいは完成度および特異性が低い結合部位として存在する。
【００３８】
そこで、上記の非共有結合型分子インプリンティングにおいては、この問題を解決する方法として、非共有結合型分子インプリンティングの後に、非共有結合型分子インプリンティングにより得られたインプリントポリマーの非特異的結合部位を化学修飾してブロックする処理（以下、「ポストインプリンティング処理」と称する）を行ってもよい。例えば、不飽和単量体（機能性モノマー）として不飽和カルボン酸を用いた非共有結合型分子インプリンティングによって得られたインプリントポリマーに対し、特異的結合部位を保護するために標的分子の共存下で、インプリントポリマー中における不飽和カルボン酸由来のカルボン酸残基で結合部位に関与していないものを、ヨウ化メチルによるメチルエステル化してもよい。このポストインプリンティング処理は、比較的簡便に、標的分子に対する非共有結合性分子認識ポリマーの選択性を向上させ、性能を改善できる可能性がある。
【００３９】
また、上記標的分子が水酸基を有し、かつ、上記非共有結合性分子認識ポリマーが標的分子の水酸基に対して特異的に可逆的な水素結合を形成する水酸基を有する場合には、共有結合型分子インプリンティングにより製造できる。すなわち、この場合、非共有結合性分子認識ポリマーは、上記光感応性部位および水酸基を有する不飽和単量体と、上記水酸基を有する標的分子との炭酸エステルを調製する調製工程と、上記炭酸エステルを架橋剤と共重合し、重合体を得る重合工程と、得られた重合体から、脱炭酸により標的分子を脱離させ、不飽和単量体由来の水酸基を遊離させる脱離工程とを含む方法で製造できる。
【００４０】
上記調製工程では、例えば、標的分子のクロロギ酸エステルを、トリエチルアミン存在下で光感応性部位および水酸基を有する不飽和単量体と反応させればよい。
【００４１】
上記重合工程は、共有結合性分子認識ポリマーの製造方法と同様である。上記脱離工程では、例えば重合工程で得られた重合体を塩基性条件下で加水分解することにより、脱炭酸を伴う標的分子の脱離を行えばよい。
【００４２】
ここで、共有結合型分子インプリンティングおよび非共有結合型分子インプリンティングの各々の利点を挙げる。共有結合型分子インプリンティングの利点は、標的分子と不飽和単量体（機能性モノマー）との複合体が比較的安定であること、結合部位の均一性が比較的高いこと、重合反応に用いる溶媒の選択において、溶媒による複合体の不安定化が起こりにくいために比較的制限が少ないこと等である。一方、非共有結合型分子インプリンティングの利点は、不飽和単量体（機能性モノマー）の重合前に、標的分子と不飽和単量体（機能性モノマー）との複合体を予め合成しておく必要がないこと、標的分子が比較的除去されやすいこと等である。
【００４３】
なお、以上の説明では、架橋剤との共重合を用いた分子インプリンティングによる製造方法について説明したが、分子認識ポリマーは、重縮合反応を利用した分子インプリンティングによっても製造可能である。
【００４４】
また、上記の説明では、分子認識ポリマーを、共有結合性分子認識ポリマーと非共有結合性分子認識ポリマーとの２種類に分類して説明したが、分子認識ポリマーは、標的分子に対して、可逆的な共有結合と可逆的な非共有結合との両方を形成するものであってもよい。このような結合モードの異なる分子認識の組み合わせは、選択性や親和性の増大が期待でき、応用範囲を広げることになる。このような分子認識ポリマーの例としては、標的分子に対して可逆的な配位結合を形成しうる錯体部位と、標的分子の水酸基に対して可逆的な水素結合を形成しうるカルボキシル基とを併せ持つ分子認識ポリマーが挙げられる。この分子認識ポリマーは、例えば、上記錯体部位を有する不飽和単量体と、カルボキシル基および光感応性部位を有する不飽和単量体と、架橋剤との共重合により得ることができる。
【００４５】
（具体例１）アゾベンゼン部位を有する光感応性インプリントポリマー
アゾベンゼンは、光照射により、容易にシス−トランス異性体間を相互変換することが知られている。このアゾベンゼンの二つのベンゼン環に、それぞれ、重合可能な二重結合と標的分子認識のための官能基を導入し、これを機能性モノマーとしてインプリントポリマーを合成すれば、光感応性インプリントポリマー光がえられる（実施例１・２参照）。この光感応性インプリントポリマーは、光照射によるアゾベンゼン部位の異性化にともない結合部位での分子認識能の変化が見られることが期待される。それゆえ、この光感応性インプリントポリマーは、標的分子の光応答性固相抽出（分離）に利用できることが期待される。
【００４６】
（具体例２）ベンゾスピロピラン基を持つ光感応性インプリントポリマー
スピロピラン類は、アゾベンゼンと並び代表的なフォトクロミック化合物である。互いに直行している二つのヘテロ環のうちの一つが紫外光の照射により開環し、その構造を大きく変える。従って、それぞれのヘテロ環を含む部分に、重合可能な二重結合と分子認識のための官能基を導入することにより、光照射による官能基の配向の変化により分子認識能が変化するインプリントポリマーを合成できると期待される。
【００４７】
ここでは、光異性化可能なベンゾスピロピラン基を導入した機能性モノマー（標的化合物と相互作用可能なモノマー）と、標的分子とを用いて、標的分子を吸脱着可能な光感応性インプリントポリマーを合成する。
【００４８】
まず、重合可能なビニル基やメタクリロイル基等の不飽和基と、標的分子と相互作用可能な官能基とを、ベンゾスピロピランに導入し、スピロピラン系機能性モノマーを合成する（図１（ａ））。例えば、メタクリロイルオキシメチル基を導入したサリチルアルデヒド誘導体を合成し、また、インドリン誘導体のクロル基を分子認識のための官能基（カルボキシル基やボロン酸基）に変換し、その後、これら二つの化合物をカップリングさせてスピロピラン系機能性モノマーを得る。
【００４９】
次いで、スピロピラン系機能性モノマーを標的分子と相互作用させて複合体を形成し、この状態で、架橋剤を加えて重合させる（図１（ｂ））。
【００５０】
最後に、標的分子を脱離させ、光感応性インプリントポリマーを得る（図１（ｃ））。
【００５１】
得られた光感応性インプリントポリマーには、標的分子と相補的な結合部位ができる。この結合部位は、光の入力でベンゾスピロピラン残基の異性化が誘起され、結合特性が変化する。そのため、この光感応性インプリントポリマーは、光のＯＮ−ＯＦＦあるいは波長の異なる光の照射により標的分子を吸着・脱着し、固相抽出が行えることが期待できる。したがって、上記光感応性インプリントポリマーには、固相抽出の際、溶媒を変えることなく、光のＯＮ−ＯＦＦあるいは波長の異なる光の照射で標的分子の濃縮および回収が繰り返し行え、分離科学の広い範囲に適用可能であると考えられる。
【００５２】
この分離手法は、特にトウモロコシ畑やゴルフ場などによく用いられるトリアジン除草剤の一つであるアトラジンを分離するのに有効であると考えられる。また、標的分子がアトラジンである場合、標的分子と相互作用可能な官能基としては、アトラジン１分子に対して複数の水素結合を形成して高い結合能を発現するカルボキシル基が好適である。アトラジンは、次式
【００５３】
【化２】

【００５４】
に示すように、カルボキシル基を持つ化合物と多点（４点）で水素結合を形成して、八員環キレート型の水素結合複合体を形成することが知られている。したがって、標的分子としてアトラジンを用い、カルボキシル基を持つ機能性モノマーを用いて分子インプリンティングを行えば、アトラジンを高選択的に吸着する光非共有結合性分子認識ポリマーが得られる。上記非共有結合性分子認識ポリマーを用いて固相抽出を行えば、アトラジンの回収を効率良く行うことができる。
【００５５】
上記のアトラジンと特異的に結合する非共有結合性分子認識ポリマーは、以下のようにして製造できる。すなわち、まず、図１（ａ）（Ｒ^１、Ｒ^２はそれぞれ、水素原子または炭化水素基を示す）に示すように、メタクリロイルオキシメチル基およびカルボキシル基を有するベンゾスピロピラン系化合物（スピロピラン系機能性モノマー）と、付加重合可能な基を複数有する架橋剤（例えば、エチレングリコールジメタクリレート）とを用意し、これらをクロロホルム等の溶媒中でアトラジンと混合して、スピロピラン系機能性モノマーのカルボキシル基にアトラジンを結合させる。そして、これを２，２’−アゾビス（イソブチロニトリル）等の重合開始剤の存在下で重合した後、アトラジンを溶解しうる酸性溶液（例えばメタノール／酢酸）および溶媒（例えばメタノール）で洗浄することにより、アトラジンが除去され、非共有結合性分子認識ポリマーが得られる。
【００５６】
（具体例３）ペプチドを用いた光感応性分子認識ポリマー
水素結合等の非共有結合による分子認識を可能とする側鎖（標的分子に対して特異的にかつ可逆的に結合する官能基）と光異性化可能な基（光感応性部位）とを導入したペプチドを合成し、このペプチドをドーパミン（３，４−ジヒドロキシエチルアミン；カテコールアミン類の１種）等の標的分子の存在下で縮重合してオリゴペプチドとすることにより、光感応性分子認識ポリマーを得る。この光感応性分子認識ポリマーでは、標的分子と光感応性分子認識ポリマーとの相互作用の補強を図るために、チオール基等をさらに導入してもよい。
【００５７】
この光感応性分子認識ポリマーは、ドーパミン等の標的分子を認識する部位（官能基）近傍に光に応答する光感応性部位が導入されていれば、光により、認識部位の構造変化に伴う認識能の制御が可能となる。この光感応性分子認識ポリマーは、ドーパミン等の薬剤を担持して輸送し、光によるＯＮ−ＯＦＦにより薬剤を吸脱着可能であると考えられるため、ドラッグデリバリーシステムへの応用が期待できる（図６）。また、この光感応性分子認識ポリマーは、オリゴペプチドを主体としていることから、生分解性で生体適合性が高いと考えられる。また、この光感応性分子認識ポリマーは、回収、分解、再利用が可能であり、環境にやさしい材料である。
【００５８】
【実施例】
以下、本発明を実施例により詳細に説明する。なお、本発明は、以下の実施例の記載に限定されるものではなく、本発明の範囲内で種々の変更が可能である。
【００５９】
〔実施例１〕
本実施例では、メタクリロイルオキシ基と水酸基とを有するアゾベンゼンを合成し、これをコレステロールとの炭酸エステルとした機能性モノマーを調製し、この機能性モノマーをエチレングリコールジメタクリレートを架橋剤として重合することによりポリマーを合成し、得られたポリマー中の炭酸エステルを加水分解して、光感応性インプリントポリマーを合成した。この光感応性インプリントポリマーは、コレステロールと水素結合を形成しうるフェノール性水酸基と、光照射によりシス−トランス異性化するアゾベンゼン部位とを有することから、コレステロールを認識する分子認識能を持ち、かつ、光照射によりアゾベンゼン部位の異性化に伴って結合部位での分子認識能が変化することが期待される。
【００６０】
（Ｓ１）３，３’−ジヒドロキシアゾベンゼンの合成
まず、窒素雰囲気下、反応容器に、塩化銅二水和物（分子量１７０．４８）１９．０５ｇ（０．１１ｍｏｌ）、３０重量％アンモニア水３３ｍｌおよび水２０ｍｌを投入した。次に、塩酸ヒドロキシルアミン（分子量６９．４９）２１．０７ｇ（０．３ｍｏｌ）を水６０ｍｌに溶解した溶液を滴下し、一価の銅錯体溶液を調製した。別の反応容器に、３−アミノフェノール（分子量１０９．１３）１０．９３ｇ（０．１ｍｏｌ）と５ｍｏｌ／ｌ塩酸水溶液１００ｍｌとを投入し、氷冷した。ここに、亜硝酸ナトリウム（分子量６９．００）７．０４ｇ（０．１ｍｏｌ）を水２０ｍｌに溶解した溶液を滴下し、ジアゾニウム塩溶液を生成させた。ここで生成したジアゾニウム塩溶液を水１３００ｍｌで希釈し、希釈したジアゾニウム塩溶液を氷冷した上記の銅錯体溶液に約１５分間かけて滴下し、そのまま３時間撹拌した。固体をろ別し、その固体を、３重量％アンモニア水５００ｍｌ、水２００ｍｌ、１ｍｏｌ／ｌ塩酸水溶液３００ｍｌ、および水２００ｍｌで洗浄した。洗浄した固体をカラムクロマトグラフィー（シリカゲル充填カラム、展開溶媒：酢酸エチル／ヘキサン＝２／３）で精製し、３，３’−ジヒドロキシアゾベンゼン（分子量２１４．２２）４．９８ｇ（０．０２３ｍｏｌ）を得た。収率は４６％であった。
【００６１】
（Ｓ２）３’−ヒドロキシ−３−メタクリロイルオキシアゾベンゼンの調製
反応容器に、３，３’−ジヒドロキシアゾベンゼン（分子量２１４．２２）１．２８ｇ（６ｍｍｏｌ）、塩化メチレン１５ｍｌ、およびピリジン３ｍｌを投入し、氷冷した。次いで、反応容器内の混合液を撹拌しながら、３０分かけて、塩化メタクリロイル（分子量１０４．５４）０．３２ｇ（３ｍｍｏｌ）を塩化メチレン５ｍｌに溶解した溶液を滴下した。滴下終了後、０℃で３０分間、室温で３０分間撹拌した。溶媒を留去し、残渣を酢酸エチルに溶解して、水で洗浄した。酢酸エチル層を硫酸マグネシウムで乾燥させた後、溶媒を留去し、残渣をカラムクロマトグラフィー（シリカゲル充填カラム、展開溶媒：酢酸エチル／ヘキサン＝１／２）で精製し、３’−ヒドロキシ−３−メタクリロイルオキシアゾベンゼン（分子量２７０．２９）０．５２ｇ（１．９ｍｍｏｌ）を得た。収率は６３％であった。
【００６２】
（Ｓ３）炭酸コレステリル３−（３−メタクリロイルオキシ）フェニルアゾフェニルの調製
反応容器に対し、３’−ヒドロキシ−３−メタクリロイルオキシアゾベンゼン０．２３ｇ（０．８ｍｍｏｌ）、テトラヒドロフラン１５ｍｌ、およびトリエチルアミン２ｍｌを投入し、氷冷した。次いで、反応容器内の混合液を撹拌しながら、この混合液に対し、氷冷によって液温を０〜５℃に保ちながらクロロギ酸コレステリル（分子量４４９．１２）０．４０ｇ（０．９ｍｍｏｌ）を３０分間かけて滴下した。その後、液温を室温に戻し、室温で２０時間撹拌を行った。
【００６３】
反応混合液を分液ロートに移し、５重量％塩酸水溶液、水、５重量％水酸化ナトリウム水溶液、および水をそれぞれ加えて分液を行い、有機層を分離した。分離した有機層を硫酸マグネシウムで３０分間かけて乾燥させ、硫酸マグネシウムを濾過によって除去した。この濾過の濾液から溶媒を留去して得られた残渣をカラムクロマトグラフィー（シリカゲル充填カラム、展開溶媒：酢酸エチル／クロロホルム／ヘキサン＝１／２／１７）で精製し、炭酸コレステリル３−（３−メタクリロイルオキシ）フェニルアゾフェニル（分子量６８２．９５）０．２１ｇ（０．３ｍｍｏｌ）を得た。収率は３８％であった。
【００６４】
（Ｓ４）ポリマーの調製
炭酸コレステリル３−（３−メタクリロイルオキシ）フェニルアゾフェニル６９．４ｍｇ（０．１ｍｍｏｌ）を、０．６４ｍｌのヘキサンおよび０．１６ｍｌのトルエン（溶媒）に溶解し、得られた溶液に、架橋剤としてのエチレングリコールジメタクリレート（分子量１９８．２２）３７７ｍｇ（１．９ｍｍｏｌ）と、重合開始剤としての２，２’−アゾビス（イソブチロニトリル）（分子量１６４．２１）５．０ｍｇ（０．０２ｍｍｏｌ）とを加えた。
【００６５】
そして、４５℃で１８時間かけて重合反応を行った。粗生成物を乳鉢で破砕し、メタノールで洗浄・乾燥した。これにより、ポリマー４１１ｍｇが得られた。
【００６６】
（Ｓ５）ポリマーの加水分解
得られたポリマーを、０．１ｍｍｏｌ／ｌの水酸化ナトリウムのメタノール溶液に懸濁させて、塩基性条件下で３時間かけて加水分解を行った。
【００６７】
これにより、脱炭酸を伴うコレステロールの脱離が起こり、結合部位にフェノール性水酸基が残った光感応性インプリントポリマー３７０ｍｇが得られた。
【００６８】
（Ｓ６）光感応性インプリントポリマーの分子認識能
クロロギ酸コレステリルに代えてクロロギ酸フェニルを用いる以外は、Ｓ２〜Ｓ４と同様の反応を行い、比較用のインプリントポリマーを合成した。
【００６９】
光感応性インプリントポリマーおよび比較用のインプリントポリマーをｎ−ヘキサン中に懸濁させて、コレステロールとの結合実験を行ったところ、光感応性インプリントポリマーは、比較用のインプリントポリマーに比べ、コレステロールの結合量が有意に多かった。このことから、光感応性インプリントポリマーが、コレステロールを認識する分子認識能を持っていることが確認された。
【００７０】
（Ｓ７）光感応性インプリントポリマーに結合させたコレステロールの紫外照
射による脱離
この光感応性インプリントポリマーをｎ−ヘキサン中に懸濁させ、暗所でコレステロールを結合させた後、高圧水銀灯で波長３３０ｎｍの紫外線を１８分間照射したところ、結合していたコレステロールが遊離することが確認された。
【００７１】
〔実施例２〕
本実施例では、鋳型分子として４−ニトロフェニル−α−Ｄ−マンノピラノシドを用いて、結合定数の大きい環状ボロン酸エステル結合を持つ機能性モノマーを調製した。この機能性モノマーの合成スキームを以下に示す。
【００７２】
【化３】

【００７３】
そして、架橋剤として分解反応を起こさないジビニルベンゼンおよびスチレンを用いて上記機能性モノマーを重合することによりポリマーを合成し、得られたポリマー中のボロン酸エステルを加水分解して、光感応性インプリントポリマーを合成した。
【００７４】
この光感応性インプリントポリマーは、４−ニトロフェニル−α−Ｄ−マンノピラノシドと環状ボロン酸エステルを形成しうるボロン酸部位と、光照射によりシス−トランス異性化するアゾベンゼン部位とを有することから、４−ニトロフェニル−α−Ｄ−マンノピラノシドを認識する分子認識能を持ち、かつ、光照射によりアゾベンゼン部位の異性化に伴って結合部位での分子認識能が変化することが期待される。
（Ｓ１）アゾカップリング反応
２ｍｏｌ／ｌ塩酸水溶液８５ｍｌ中、０℃、１５分間の条件で、ｐ−ブロモアニリン（分子量１７２．０２）４．３０ｇ（２５ｍｍｏｌ）を亜硝酸ナトリウム（分子量９４．１１）１．９０ｇ（２７．５ｍｍｏｌ）と反応させ、塩化ｐ−ブロモベンゼンジアゾニウムの水溶液を調製した。
【００７５】
続いて、得られた塩化ｐ−ブロモベンゼンジアゾニウム水溶液に対してフェノール（分子量９４．１１）２．３５ｇ（２５ｍｍｏｌ）および水酸化ナトリウム（分子量４０．００）２ｇ（５０ｍｍｏｌ）を加え、０℃、５分間の条件でアゾカップリング反応を行った。
【００７６】
アゾカップリング反応生成物を粗生成物から単離することにより、４−ブロモ−４’−ヒドロキシアゾベンゼン（分子量２７７．１２５）２．７２ｇ（９．８ｍｍｏｌ）を得た。収率は３９％であった。
（Ｓ２）ボロン酸エステルに変換
反応容器に、４−ブロモ−４’−ヒドロキシアゾベンゼン０．５６ｇ（２．０ｍｍｏｌ）、［１，１’−ビス（ジフェニルホスフィノ）フェロセン］パラジウム（ＩＩ）ジクロリド・塩化メチレン（１：１）錯体（分子量８１６．６４）４４ｍｇ（０．０６ｍｍｏｌ）、ビス（ピナコラト）ジボロン（分子量２５３．９４）０．５６ｇ（２．２ｍｍｏｌ）、酢酸カリウム（分子量９８．１５）０．３１ｇ（３．１ｍｍｏｌ）、および溶媒としての乾燥ＤＭＦ（Ｎ，Ｎ−ジメチルホルムアミド）１０ｍｌを投入し、反応混合物を窒素雰囲気下、８０℃で１８時間加熱した。反応の完了を確認した後、反応混合物をジエチルエーテルで希釈し、セライトで濾過し、水および食塩水で洗浄した。洗浄した溶液を硫酸マグネシウムで乾燥し、真空吸引により溶媒を除去した。その後、残渣を酢酸エチル／ヘキサン混合溶媒による再結晶で精製した。これにより、４’−ヒドロキシアゾベンゼン−４−ボロン酸ピナコラト（分子量３２４．１９）０．６０ｇ（１．９ｍｍｏｌ）を得た。収率は９３％であった。
【００７７】
なお、ｄｐｐｆは配位子１，１’−ビス（ジフェニルホスフィノ）フェロセンを指す。
（Ｓ３）メタクリル酸エステル生成
４’−ヒドロキシアゾベンゼン−４−ボロン酸ピナコラト０．６１ｇ（１．９ｍｍｏｌ）を、塩化メチレン１０ｍｌに溶解させ、ピリジン１ｍｌを添加した。この溶液に対し、塩化メタクリロイル（分子量１０４．５４）０．２７ｇ（２．６ｍｍｏｌ）を加えて、０℃で３０分間、室温で１時間撹拌することにより、エステル化反応を行った。
【００７８】
メタクリル酸エステルを粗生成物から単離することにより、４’−メタクリロイルオキシアゾベンゼン−４−ボロン酸ピナコラト（分子量３９２．２６）０．５８ｇ（１．５ｍｍｏｌ）を得た。収率は７９％であった。
（Ｓ４）加水分解
４’−メタクリロイルオキシアゾベンゼン−４−ボロン酸ピナコラト０．５８ｇ（１．５ｍｍｏｌ）を塩酸／アセトン混合液中、室温、２４時間の条件で加水分解させた。
【００７９】
加水分解生成物を粗生成物から単離することにより、分子内にメタクリロイル基とボロン酸基とを有する機能性モノマーとしての４’−メタクリロイルオキシアゾベンゼン−４−ボロン酸（分子量３１０．１２）０．１９ｇを得た。収率は４１％であった。
（Ｓ５）ボロン酸環状エステルモノマー調製
乾燥塩化メチレン２０ｍｌ中で、４’−メタクリロイルオキシアゾベンゼン−４−ボロン酸３１ｍｇ（０．１ｍｍｏｌ）と、鋳型分子としての４−ニトロフェニル−α−Ｄ−マンノピラノシド（分子量３０１．３０）１５ｍｇ（０．０５ｍｍｏｌ）とを混合し、１８時間撹拌した（図２参照）。
【００８０】
溶媒を留去することにより、図２のＡに示す、４−ニトロフェニル−α−Ｄ−マンノピラノシドに対して２分子のボロン酸が環状エステルを形成したモノマー（ボロン酸環状エステルモノマー）（分子量８４９．４１）４２ｍｇを得た。収率は１００％であった。このモノマーは、更なる精製は行わず、そのまま次のポリマー合成に使用した。
（Ｓ６）光感応性インプリントポリマーの調製
ボロン酸環状エステルモノマーを、図２に示すように、重合することで光感応性インプリントポリマーを調製した。
【００８１】
重合は、図３に示す操作により行った。すなわち、まず、ボロン酸環状エステルモノマー４２ｍｇ（０．０５ｍｍｏｌ）をテトラヒドロフラン０．８ｍｌに溶解し、得られた溶液に、架橋剤としてのジビニルベンゼン１９５ｍｇ（１．５ｍｍｏｌ）およびスチレン５２ｍｇ（０．５ｍｍｏｌ）と、重合開始剤としての２，２’−アゾビス（２，４−ジメチルバレロニトリル）５ｍｇ（０．０２ｍｍｏｌ）とを加えた。
【００８２】
そして、４５℃で１８時間かけて重合反応を行った。粗生成物を破砕し、メタノールで洗浄・乾燥することにより、ポリマーを単離した。これにより、ポリマー４１ｍｇが得られた。
（Ｓ７）環状ボロン酸エステルの加水分解
得られたポリマーを、図２および図４に示すように加水分解した。すなわち、得られたポリマーを、５重量％の塩酸を含むアセトン３０ｍｌに懸濁させることにより、酸性条件下で１６時間かけて環状ボロン酸エステルの加水分解を行った。
【００８３】
これにより、鋳型分子４−ニトロフェニル−α−Ｄ−マンノピラノシドがポリマーから脱離し、光感応性インプリントポリマーが得られた。
（Ｓ８）光感応性インプリントポリマーの紫外・可視吸収スペクトルの変化
光感応性インプリントポリマーに対し、波長３３１ｎｍの紫外線を１５分間照射したところ、図５に示すように、波長３３０ｎｍ付近の吸収（トランス体に起因する）が減少し、波長４４０ｎｍ付近の吸収（シス体に起因する）が増加した。これにより、トランス体が減少してシス体が増加すること、すなわちアゾベンゼン部位の異性化が起こっていることが確認された。
【００８４】
なお、図５に示す紫外・可視吸収スペクトルは、光感応性インプリントポリマーをクロロホルム中に懸濁させた状態で測定した。
【００８５】
【発明の効果】
本発明によれば、以上のように、標的分子と選択的に結合すると共に、光照射により物理・化学的変化を誘導できる光感応性を持ち、標的分子に対する吸脱着を光で制御可能な新規分子認識ポリマーを提供できる。
【００８６】
この分子認識ポリマーは、固相抽出の際、溶媒を変える煩雑な操作を行うことなく、光のＯＮ−ＯＦＦあるいは波長変更により標的分子の濃縮および回収が繰り返し行えるので、標的分子を固相抽出するための分離剤として極めて有用であると考えられる。また、固相抽出における溶媒の交換を不要としたことで、自動化が容易な固相抽出系を構築できる。
【００８７】
また、ドーパミン等の薬剤を標的分子とする分子認識ポリマーは、薬剤を担持して輸送し、光によるＯＮ−ＯＦＦにより薬剤を吸脱着可能であると考えられるため、ドラッグデリバリーシステムへの応用が期待できる。
【００８８】
また、本発明の製造方法によれば、上記の新規分子認識ポリマーを製造することができる。
【図面の簡単な説明】
【図１】本発明の一実施例における分子認識ポリマーの製造方法を示す図である。
【図２】本発明の他の実施例における分子認識ポリマーの製造方法を示す図である。
【図３】本発明の他の実施例における分子認識ポリマーの製造方法における、重合工程および脱離工程の操作・条件を示す図である。上記の図である。
【図４】本発明の他の実施例における分子認識ポリマーの製造方法の一工程を示す図である。
【図５】アゾベンゼン部位を持つ光感応性インプリントポリマーに対し、紫外線を照射したときの紫外・可視吸収スペクトルの変化を示す図である。
【図６】本発明の一形態における光感応性インプリントポリマーの分子認識能が外部刺激（光）により変化する様子を示す図である。[0001]
TECHNICAL FIELD OF THE INVENTION
The present invention relates to a molecular recognition polymer that selectively binds to a target molecule, and a method for producing a molecular recognition polymer using a molecular imprinting method.
[0002]
[Prior art]
As a method for producing a molecular recognition polymer that specifically binds to a target molecule, an interesting method called molecular imprinting, in which the design of the functional site of the molecular recognition polymer is left to the molecule itself, is known. The principle of molecular imprinting is very clear. The principle is that when synthesizing a crosslinkable polymer having a specific binding site for the target molecule, the target molecule is mixed with the monomer for polymer synthesis and polymerization is performed. When the target molecule is washed out, a binding site complementary to the target molecule is formed in the polymer. If the target molecule has a characteristic functional group, a monomer (functional monomer) that interacts with the functional group through a non-covalent bond is polymerized together with a cross-linking agent serving as a polymer matrix (hereinafter, referred to as “non-covalent type Molecular imprinting) or synthesize a complex of the target molecule and the functional monomer in advance and polymerize it with a cross-linking agent (hereinafter, this is called “covalent molecular imprinting”). This is a general method. After polymerization, if the polymer is swollen and washed under the condition that the target molecule is detached, the part of the polymer after the disappearance of the target molecule can interact with the target molecule like a target molecule template A functional group remains, and a polymer that specifically binds to the target molecule (hereinafter referred to as “imprint polymer”) is obtained. That is, a functional group derived from a functional monomer is arranged in a matrix of a polymer derived from a cross-linking agent so as to recognize a characteristic functional group according to the shape of a target molecule. The meaning around the meaningfully localized functional group is a site that specifically binds to the target molecule.
[0003]
The present inventors have demonstrated that in a molecular imprinting method, a unique functional monomer is designed and synthesized, and that a molecular recognition polymer capable of selectively adsorbing various target molecules can be tailor-made. Patent Documents 1 to 5).
[0004]
In addition, the present inventors have shown that by imparting some function to this molecular imprinting method, it is possible to synchronize with the binding of a target molecule to create a new molecular recognition polymer that expresses that function. (See Non-Patent Documents 1 and 2).
[0005]
In addition, the present inventors have applied the synthesized imprinted polymer to a separation technique and a sensor, and particularly applied the imprinted polymer to solid-phase extraction to demonstrate its usefulness (Non-Patent Documents 4 to 5). 6).
[0006]
The present inventors have also developed a robot that automatically performs polymer synthesis and evaluation of its binding ability, and have established the concept of combinatorial molecular imprinting (see Non-Patent Document 7).
[0007]
[Non-patent document 1]
Takeuchi, T .; Mukawa, T .; , Matsui, J. et al. Higashi, M .; , Shimizu, K .; D. Molecularly Imprinted Polymers with Metalloporphyrin-Based Molecular Recognition Sites Coassembled with Methacrylic acid, Anal. Chem. 2001, 73, 3869-3874.
[0008]
[Non-patent document 2]
Matsui, J .; Higashi, M .; Takeuchi, T .; Molecularly Imprinted Polymer as 9-Ethyladeneine Receptor Having a Porphyrin-based Recognition Center, J. Mol. Am. Chem. Soc. 2000, 122, 5218-5219.
[0009]
[Non-Patent Document 3]
Takeuchi, T .; , Dobashi, A .; , Kimura, K .; Molecular Imprinting of Biotin Derivatives and Its Application to Competitive Binding Assay Using Non-isotopic Labeled Ligands, Anal. Chem. 2000, 72, 2418-2422.
[0010]
[Non-patent document 4]
Molecularly Imprinted Polymers: Man Made Mimics of Antibodies and Their Application in Analytical Chemistry, Cellularn, B.M. Ed. , Elsevier, 2001.
[0011]
[Non-Patent Document 5]
Takeuchi, T .; , Haginaka, J .; Separation and Sensing Based on Molecular Recognition Using Molecularly Imprinted Polymers, J. Amer. Chromatogr. B 1999, 728, 1-20.
[0012]
[Non-Patent Document 6]
Takeuchi, T .; , Matsui, J. et al. Miniaturized Molecularly Imprinted Continuous Polymer Rods, J. Mol. High Resolution Chromatogr. 2000, 23, 44-46.
[0013]
[Non-Patent Document 7]
Takeuchi, T .; Fukuma, D .; , Matsui, J. et al. Combinatorial Molecular Imprinting: An Approach to Synthetic Polymer Receptors, Anal. Chem. 1999, 71, 285-290.
[0014]
[Problems to be solved by the invention]
However, in solid-phase extraction of target molecules using a molecular recognition polymer manufactured by the conventional molecular imprinting method, solvent conditions under which the target compound is easily adsorbed by the adsorbent are used for enrichment of the target molecules, and target molecules are collected for collecting the target molecules. Use solvent conditions in which the molecules are easily desorbed from the adsorbent. This operation is complicated and hinders automation, and may cause degradation of the target molecule.
[0015]
If a molecular recognition material capable of controlling the adsorption and desorption of a target molecule by an external stimulus can be provided, the enrichment and recovery of the target molecule can be achieved by ON-OFF of light without performing a complicated operation of changing a solvent during solid phase extraction. It is thought that it can be repeated, and it is considered to be extremely useful. However, no attempt has been made to change the molecular recognition polymer in response to an external stimulus.
[0016]
The present invention has been made in view of the above-mentioned conventional problems, and has an object to selectively bind to a target molecule and to have a photosensitivity capable of inducing a physical / chemical change by light irradiation, and to absorb a target molecule. It is an object of the present invention to provide a novel molecular recognition polymer whose desorption can be controlled by light, and a method for producing the molecular recognition polymer using a molecular imprinting method.
[0017]
[Means for Solving the Problems]
In order to solve the above-mentioned problems, the molecular recognition polymer of the present invention is a novel polymer having a functional group that specifically and reversibly binds to a target molecule and a light-sensitive site that isomerizes upon irradiation with light. It is characterized by being a molecular recognition polymer.
[0018]
The molecular recognition polymer can be selectively bonded to a target molecule by a functional group, and can adsorb and desorb the target molecule. Since the molecular recognition polymer has a light-sensitive portion, isomerization (chemical change, physical change) of the light-sensitive portion occurs due to ON-OFF of light irradiation or irradiation of light having a different wavelength. With this isomerization, the molecular structure near the functional group changes, and the ability of the functional group to bind to the target molecule (molecular recognition ability) changes. Therefore, the molecular recognition polymer can control the molecular recognition ability of the functional group with respect to the target molecule by ON-OFF or wavelength change of external light irradiation, and can adsorb and desorb the target molecule.
[0019]
BEST MODE FOR CARRYING OUT THE INVENTION
The molecular recognition polymer according to the present invention has a functional group that specifically and reversibly binds (interacts) with a target molecule (target compound), and a light-sensitive site that isomerizes by light irradiation.
[0020]
Examples of the molecular recognition polymer include (1) a molecular recognition polymer having a covalent functional group that forms a reversible covalent bond specifically to a target molecule and a light-sensitive site that isomerizes upon irradiation with light (hereinafter referred to as a molecular recognition polymer). , Referred to as a "covalent molecular recognition polymer"), (2) a non-covalent functional group that forms a reversible non-covalent bond specifically to a target molecule, and a photosensitivity that isomerizes by light irradiation There is a molecular recognition polymer having a moiety (hereinafter, referred to as “non-covalent molecular recognition polymer”).
[0021]
In order for the target molecule to form a reversible bond to the covalent functional group of the covalent molecular recognition polymer, it is necessary that the covalent bond is easily formed and the target molecule can be detached. Considering this point, the covalent bond must be not very strong, and the formation and dissociation of the covalent bond must be easy and reversible. Examples of the reversible covalent bond include an ester bond between a boronic acid group and a cis-diol, an acetal bond between an aldehyde and a hydroxyl group, a ketal bond between a ketone and a hydroxyl group, and a Schiff base formed by a bond between an aldehyde and an amino group. A coordination bond using a metal complex (for example, zinc porphyrin); an ester bond between a carboxyl group and a hydroxyl group; an amide bond between a carboxyl group and an amino group. Among these, a reversible ester bond between a boronic acid group and a cis-diol is preferred because of its quick equilibrium.
[0022]
On the other hand, reversible non-covalent bonds that can be used for non-covalent molecular recognition polymers include hydrogen bond-donating groups (hydrogen atoms on amide groups, hydrogen atoms on carboxyl groups, amino groups, hydroxyl groups, phenolic hydroxyl groups, etc.). ) And a hydrogen bond-accepting group (such as a carbonyl portion of an amide group, a carbonyl portion of a carboxyl group, or a nitrogen atom on a nitrogen-containing heterocyclic ring such as a pyridine ring), or an ionic bond (between a cation and an anion). Electrostatic interaction). When the non-covalent molecular recognition polymer forms a hydrogen bond with the target molecule, it must form a multipoint hydrogen bond in order to form a high-selectivity and strong bond with the target molecule. Are preferable, and most preferably form hydrogen bonds at four or more points. Further, in order for the non-covalent molecular recognition polymer to bind to the target molecule in a highly selective and strong manner, the multipoint hydrogen bond may form an eight-membered ring chelate-type hydrogen bond complex. preferable. Examples of the combination of compounds forming an 8-membered ring chelate-type hydrogen bond complex include a compound having at least one amide group and / or carboxyl group, an amino group at a position adjacent to the nitrogen atom on the nitrogen-containing heterocycle, and And / or a compound having at least one carbonylamino group.
[0023]
The combination of the target molecule and the molecular recognition polymer is not particularly limited as long as it can form a reversible bond between them. That is, these combinations are formed between the functional group (boronic acid group, hydroxyl group, carboxyl group, amino group, phenolic hydroxyl group, amide group, etc.) of the molecular recognition polymer and the functional group (hydroxyl group, etc.) of the target molecule. Any combination that forms a reversible bond may be used. Examples of the combination of the target molecule and the molecular recognition polymer include: (1) a compound having a catechol group having a cisdiol as a target molecule or a polyhydric alcohol such as a sugar capable of forming a reversible ester bond; (2) a combination of a compound serving as a hydrogen donor and a compound serving as a hydrogen acceptor (for example, atrazine as a target molecule, and a carboxyl group which forms a hydrogen bond with atrazine) And (3) a combination of a target molecule having a charge and a molecular recognition polymer having a charge opposite to that of the target molecule.
[0024]
The light-sensitive site may be any type as long as a photoisomerization, that is, a chemical or physical change caused by light irradiation occurs reversibly, but the structure (conformation) is formed by reversible isomerization by light irradiation. ) Greatly change, for example, an azobenzene site (azobenzene skeleton) and a spiropyran site (spiropyran skeleton) are preferable.
(Production method of covalent molecular recognition polymer)
The covalent molecular recognition polymer can be produced by so-called covalent molecular imprinting. That is, the covalent molecular recognition polymer is prepared by preparing an unsaturated monomer having a covalent functional group and a light-sensitive site (also referred to as a functional monomer); A bonding step of bonding the molecule with a reversible covalent bond, a polymerization step of copolymerizing an unsaturated monomer bonded to the target molecule with a cross-linking agent to obtain a polymer, and forming a target molecule from the obtained polymer. And a desorption step of desorption.
[0025]
The unsaturated monomer is a functional group capable of forming a reversible covalent bond (the above-described coordination bond, acetal-ketal bond, ester bond between boric acid and cis-diol) with the functional group of the target molecule. And a compound having an addition-polymerizable unsaturated bond and a light-sensitive site. The functional group capable of forming a reversible covalent bond with the functional group of the target molecule includes a metal complex site capable of forming a coordination bond with an oxygen atom of the target molecule, a cis-diol site of the target molecule and a cyclic structure. And a diboronic acid moiety capable of forming an ester bond.
[0026]
In the above-described preparation step, for example, a compound having a light-sensitive site may be subjected to addition polymerization by adding an addition-polymerizable unsaturated group such as a vinyl group or a methacryloyl group, and further may form a reversible covalent bond with a target molecule. An unsaturated monomer may be prepared by adding a covalent functional group (or the like). Azobenzene and benzospiropyran are typical examples of the compound having a photosensitive portion. In addition, as a compound having a light-sensitive moiety, a structure in which the structure does not change so much by light irradiation, but a color is changed, a diarylethene-based compound, a fluidide, a salicylideneaniline-based compound, etc. which can be applied to a rewritable recording material. Of reversible photoresponsive compounds. In addition, triphenylmethane cyanide, in which the carbon surrounded by the three phenyl groups becomes a cation by light irradiation and whose polarity is greatly increased, is also a compound having a light-sensitive site.
[0027]
In the above polymerization step, an unsaturated monomer and a crosslinking agent are dissolved in a solvent, and a copolymerization reaction is performed using a polymerization initiator. The solvent also acts as a pore former to make the polymer porous. The solvent is not particularly limited as long as it can dissolve the target molecule and the unsaturated monomer. Chloroform is preferred because it dissolves each reagent well. Further, as the polymerization initiator, 2,2′-azobis (isobutyronitrile), 2,2′-azobis (2,4-dimethylvaleronitrile) or the like can be used.
[0028]
The crosslinking agent used in the above addition polymerization is a compound having a group copolymerizable with a polymerizable group of the unsaturated monomer, for example, a vinyl group or a (meth) acryloyl group. Examples of the crosslinking agent are shown in the following formulas (1) to (20).
[0029]
Embedded image

[0030]
As the crosslinking agent, those having a plurality of groups capable of addition polymerization are preferable. Examples of the crosslinking agent having a plurality of groups capable of addition polymerization include p-divinylbenzene represented by the formula (4), o-divinylbenzene represented by the formula (18), and divinyl such as m-divinylbenzene represented by the formula (19). There are a benzene-based crosslinking agent and a (meth) acrylic-acid-based crosslinking agent. Examples of the (meth) acrylic acid-based crosslinking agent include compounds having a plurality of (meth) acryloyl groups, for example, ethylene glycol dimethacrylate (EDMA) represented by the formula (1), and trimethylolpropane trimethacrylate (TRIM) represented by the formula (2). ), Trimethylolpropane triacrylate represented by formula (3), tetraethylene glycol diacrylate represented by formula (6), triethylene glycol dimethacrylate represented by formula (11), pentaerythritol tetraacrylate represented by formula (12), Pentaerythritol triacrylate represented by (13), bisphenol A dimethacrylate represented by formula (14), N, N'-methylenediacrylamide represented by formula (15), N, N'-1,4 represented by formula (16) -Phenylenediacrylamide, represented by formula (17) 3,5-bis (acryloylamide) benzoic acid, N shown in Equation (20), O-Bisuakuriroiru -L- phenylalaninol the like. Compounds having only one vinyl group (eg, vinyl chloride represented by formula (5), styrene represented by formula (7), phenylvinyl sulfoxide represented by formula (9), phenyl vinyl sulfone represented by formula (10), etc. ) Or a compound having only one polymerizable group such as a compound having only one (meth) acryloyl group (for example, methyl methacrylate shown in the formula (8)). As the crosslinking agent, it is preferable to use a crosslinking agent having the same group as the polymerizable group of the unsaturated monomer. Further, the crosslinking agent is preferably used in a large excess with respect to the unsaturated monomer in order to avoid crosstalk between binding sites. Specifically, it is preferable to use 5 mol or more of a crosslinking agent per 1 mol of the unsaturated monomer.
[0031]
In the elimination step, the target molecule is eliminated by breaking a reversible bond between the functional group in the polymer obtained in the polymerization step and the target molecule. For example, when the reversible bond is an ester bond, the target molecule may be eliminated by subjecting the ester bond to acid hydrolysis or alkali hydrolysis.
[0032]
(Production method of non-covalent molecular recognition polymer)
The non-covalent molecular recognition polymer can be produced by so-called non-covalent molecular imprinting. That is, the non-covalent molecular recognition polymer is prepared by preparing an unsaturated monomer having the non-covalent functional group and the light-sensitive site, and converting the unsaturated monomer to the presence of the target molecule. The polymer can be produced by a method comprising a polymerization step of copolymerizing with a crosslinking agent below to obtain a polymer, and a elimination step of eliminating a target molecule from the obtained polymer.
[0033]
The unsaturated monomer is a compound having a functional group capable of forming a reversible non-covalent bond (such as a hydrogen bond) with a functional group of a target molecule, a polymerizable unsaturated bond, and a light-sensitive site. It is. The functional group capable of forming a reversible non-covalent bond with the functional group of the target molecule includes a carbonyl group capable of forming a hydrogen bond with an amino group of the target molecule and a nitrogen atom of a nitrogen-containing heterocyclic ring of the target molecule. And a hydroxyl group or an amino group capable of forming a hydrogen bond with the amino group.
[0034]
In the above preparation step, for example, a compound having a light-sensitive site is added with an addition-polymerizable unsaturated group such as a vinyl group or a methacryloyl group to enable addition polymerization and further form a non-covalent reversible bond with the target molecule. A non-covalent functional group (carboxyl group, amino group, phenolic hydroxyl group, etc.) may be added to prepare an addition-polymerizable unsaturated monomer.
[0035]
The above polymerization step is basically the same as the method for producing a covalent molecular recognition polymer, but in non-covalent molecular imprinting, a complex of a target molecule and an unsaturated monomer is stably present. For this purpose, it is preferable to use an unsaturated monomer in excess with respect to the target molecule. Specifically, it is preferable to use 2 mol or more of the target molecule per 1 mol of the target molecule. When the non-covalent bond is a hydrogen bond, the solvent to be used is preferably a halogen-based solvent such as chloroform which does not hinder the hydrogen bond.
[0036]
In the desorption step, the target molecule is desorbed by washing with a solution capable of selectively dissolving the target molecule (eg, an alcohol-based solvent). Thereby, a non-covalent molecular recognition polymer is obtained.
[0037]
In the above-described non-covalent molecular imprinting, the heterogeneity of the formed binding site may be a problem. Usually, in order to form a complex between the target molecule and the unsaturated monomer (functional monomer), the unsaturated monomer (functional monomer) is added in excess to the target molecule and polymerized. Naturally, the unsaturated monomer (functional monomer) involved in the binding site is limited, and the excess of the added unsaturated monomer (functional monomer) is limited to specific binding. It is not a site, but non-specific binding. That is, the excess exists independently of the specific binding site or as a less complete and less specific binding site.
[0038]
Therefore, in the above-described non-covalent molecular imprinting, as a method for solving this problem, after the non-covalent molecular imprinting, the non-specific binding of the imprint polymer obtained by the non-covalent molecular imprinting is performed. A process of chemically modifying and blocking the binding site (hereinafter, referred to as “post-imprinting process”) may be performed. For example, coexistence of target molecule to protect specific binding site to imprinted polymer obtained by non-covalent type molecular imprinting using unsaturated carboxylic acid as unsaturated monomer (functional monomer) Below, a carboxylic acid residue derived from an unsaturated carboxylic acid in the imprint polymer that is not involved in the binding site may be subjected to methyl esterification with methyl iodide. This post-imprinting treatment may improve the selectivity of the non-covalent molecular recognition polymer for the target molecule relatively easily and may improve the performance.
[0039]
Further, when the target molecule has a hydroxyl group, and the non-covalent molecular recognition polymer has a hydroxyl group that forms a reversible hydrogen bond specifically to the hydroxyl group of the target molecule, a covalent bond type It can be produced by molecular imprinting. That is, in this case, the non-covalent molecular recognition polymer is a preparation step of preparing a carbonic acid ester of the unsaturated monomer having a light-sensitive site and a hydroxyl group, and a carbonic acid ester of the target molecule having a hydroxyl group, And a cross-linking agent, a polymerization step to obtain a polymer, from the obtained polymer, a target molecule is eliminated by decarboxylation, and a elimination step of releasing a hydroxyl group derived from an unsaturated monomer is included. Can be manufactured by the method.
[0040]
In the preparation step, for example, the chloroformate of the target molecule may be reacted with an unsaturated monomer having a photosensitive site and a hydroxyl group in the presence of triethylamine.
[0041]
The above polymerization step is the same as the method for producing a covalent molecular recognition polymer. In the elimination step, for example, the polymer obtained in the polymerization step may be hydrolyzed under basic conditions to thereby eliminate the target molecule accompanied by decarboxylation.
[0042]
Here, the advantages of each of covalent molecular imprinting and non-covalent molecular imprinting are listed. The advantages of covalent molecular imprinting are that the complex of the target molecule and the unsaturated monomer (functional monomer) is relatively stable, the binding site is relatively uniform, and it is used for the polymerization reaction. In the selection of the solvent, there are relatively few restrictions because the complex is hardly destabilized by the solvent. On the other hand, the advantage of non-covalent molecular imprinting is that before the polymerization of the unsaturated monomer (functional monomer), a complex of the target molecule and the unsaturated monomer (functional monomer) is synthesized in advance. Need not be kept, and the target molecule is relatively easily removed.
[0043]
In the above description, the production method by molecular imprinting using copolymerization with a crosslinking agent has been described. However, the molecular recognition polymer can also be produced by molecular imprinting using a polycondensation reaction.
[0044]
Further, in the above description, the molecular recognition polymer is classified into two types, that is, a covalent molecule recognition polymer and a non-covalent molecule recognition polymer, but the molecule recognition polymer is reversible with respect to the target molecule. It may form both a permanent covalent bond and a reversible non-covalent bond. Such a combination of molecular recognitions having different binding modes can be expected to increase selectivity and affinity, thereby expanding the application range. Examples of such a molecular recognition polymer include a complex site capable of forming a reversible coordination bond with a target molecule and a carboxyl group capable of forming a reversible hydrogen bond with a hydroxyl group of the target molecule. A molecular recognition polymer having the same may be used. This molecular recognition polymer can be obtained, for example, by copolymerizing an unsaturated monomer having the above-described complex site, an unsaturated monomer having a carboxyl group and a light-sensitive site, and a crosslinking agent.
[0045]
(Specific Example 1) Photosensitive imprint polymer having azobenzene moiety
Azobenzene is known to easily convert between cis-trans isomers by light irradiation. By introducing a polymerizable double bond and a functional group for target molecule recognition into each of the two benzene rings of this azobenzene, and synthesizing the imprint polymer with this as a functional monomer, a photosensitive imprint polymer is obtained. Light is obtained (see Examples 1 and 2). This photosensitive imprint polymer is expected to show a change in the molecular recognition ability at the binding site due to the isomerization of the azobenzene site by light irradiation. Therefore, it is expected that this photosensitive imprinted polymer can be used for photoresponsive solid-phase extraction (separation) of target molecules.
[0046]
(Example 2) Photosensitive imprinted polymer having benzospiropyran group
Spiropyrans are typical photochromic compounds along with azobenzene. One of the two heterocycles that are orthogonal to each other is opened by irradiation with ultraviolet light, and significantly changes its structure. Therefore, by introducing a polymerizable double bond and a functional group for molecular recognition into the part containing each heterocycle, the imprint polymer whose molecular recognition ability changes due to a change in the orientation of the functional group due to light irradiation. Is expected to be synthesized.
[0047]
Here, using a functional monomer (a monomer capable of interacting with a target compound) into which a photoisomerizable benzospiropyran group is introduced and a target molecule, a photosensitive imprint polymer capable of adsorbing and desorbing the target molecule is prepared. Combine.
[0048]
First, an unsaturated group such as a polymerizable vinyl group or methacryloyl group and a functional group capable of interacting with a target molecule are introduced into benzospiropyran to synthesize a spiropyran-based functional monomer (FIG. 1 (a)). . For example, a salicylaldehyde derivative into which a methacryloyloxymethyl group has been introduced is synthesized, and a chloro group of the indoline derivative is converted into a functional group (a carboxyl group or a boronic acid group) for molecular recognition. The spiropyran-based functional monomer is obtained by coupling.
[0049]
Next, the spiropyran-based functional monomer interacts with the target molecule to form a complex, and in this state, a crosslinking agent is added to polymerize (FIG. 1 (b)).
[0050]
Finally, the target molecule is eliminated to obtain a photosensitive imprint polymer (FIG. 1 (c)).
[0051]
The resulting photosensitive imprinted polymer has a binding site complementary to the target molecule. At this binding site, light input induces isomerization of the benzospiropyran residue to change the binding characteristics. Therefore, it can be expected that this light-sensitive imprinted polymer can perform solid-phase extraction by adsorbing and desorbing target molecules by ON-OFF of light or irradiation of light having different wavelengths. Therefore, in the above-described photosensitive imprinted polymer, the concentration and recovery of the target molecule can be repeated by ON-OFF of light or irradiation of light of different wavelengths without changing the solvent during solid-phase extraction. It is considered applicable to a wide range.
[0052]
This separation technique is considered to be particularly effective for separating atrazine, which is one of the triazine herbicides frequently used in corn fields and golf courses. When the target molecule is atrazine, the functional group capable of interacting with the target molecule is preferably a carboxyl group that forms a plurality of hydrogen bonds with one molecule of atrazine and exhibits high binding ability. Atrazine has the formula
[0053]
Embedded image

[0054]
It is known that a compound having a carboxyl group forms a hydrogen bond at multiple points (four points) to form an eight-membered ring chelate type hydrogen bond complex as shown in FIG. Therefore, if atrazine is used as a target molecule and molecular imprinting is performed using a functional monomer having a carboxyl group, a photo-noncovalent molecular recognition polymer that adsorbs atrazine with high selectivity can be obtained. If solid phase extraction is performed using the noncovalent molecular recognition polymer, atrazine can be efficiently recovered.
[0055]
The above non-covalent molecular recognition polymer that specifically binds to atrazine can be produced as follows. That is, first, as shown in FIG. ¹ , R ² Each represents a hydrogen atom or a hydrocarbon group), a benzospiropyran-based compound having a methacryloyloxymethyl group and a carboxyl group (a spiropyran-based functional monomer), and a cross-linking agent having a plurality of addition-polymerizable groups ( For example, ethylene glycol dimethacrylate) is prepared, and these are mixed with atrazine in a solvent such as chloroform to bond atrazine to the carboxyl group of the spiropyran-based functional monomer. Then, this is polymerized in the presence of a polymerization initiator such as 2,2′-azobis (isobutyronitrile), and then washed with an acidic solution (eg, methanol / acetic acid) and a solvent (eg, methanol) that can dissolve atrazine. By doing so, atrazine is removed, and a non-covalent molecular recognition polymer is obtained.
[0056]
(Specific Example 3) Photosensitive molecule recognition polymer using peptide
Introduction of a side chain (functional group that binds specifically and reversibly to the target molecule) and a photoisomerizable group (photosensitive site) that enable molecular recognition by non-covalent bonds such as hydrogen bonds Of the photo-sensitive molecule-recognition polymer by synthesizing the synthesized peptide and subjecting the peptide to condensation polymerization in the presence of a target molecule such as dopamine (3,4-dihydroxyethylamine; one of catecholamines) to form an oligopeptide. obtain. In this photosensitive molecule recognizing polymer, a thiol group or the like may be further introduced in order to reinforce the interaction between the target molecule and the photosensitive molecule recognizing polymer.
[0057]
This light-sensitive molecule-recognition polymer has a light-sensitive light-sensitive site introduced in the vicinity of a site (functional group) for recognizing a target molecule such as dopamine. Function can be controlled. It is considered that this photosensitive molecule-recognizing polymer can carry and transport a drug such as dopamine, and can absorb and desorb the drug by ON-OFF by light, so that application to a drug delivery system can be expected (FIG. 6). ). In addition, since this photosensitive molecule-recognition polymer is mainly composed of oligopeptides, it is considered that it is biodegradable and has high biocompatibility. In addition, this photosensitive molecule-recognizing polymer can be recovered, decomposed, and reused, and is an environmentally friendly material.
[0058]
【Example】
Hereinafter, the present invention will be described in detail with reference to examples. Note that the present invention is not limited to the description of the following embodiments, and various modifications can be made within the scope of the present invention.
[0059]
[Example 1]
In this example, azobenzene having a methacryloyloxy group and a hydroxyl group was synthesized, a functional monomer was prepared by converting it to a carbonate ester with cholesterol, and the functional monomer was polymerized using ethylene glycol dimethacrylate as a crosslinking agent. And a carbonate in the obtained polymer was hydrolyzed to synthesize a photosensitive imprinted polymer. Since the photosensitive imprint polymer has a phenolic hydroxyl group capable of forming a hydrogen bond with cholesterol and an azobenzene moiety which is cis-trans isomerized by light irradiation, it has a molecular recognition ability for recognizing cholesterol, and In addition, it is expected that the molecular recognition ability at the binding site changes with the isomerization of the azobenzene site by light irradiation.
[0060]
(S1) Synthesis of 3,3′-dihydroxyazobenzene
First, under a nitrogen atmosphere, 19.05 g (0.11 mol) of copper chloride dihydrate (molecular weight 170.48), 33 ml of 30% by weight aqueous ammonia and 20 ml of water were charged into a reaction vessel. Next, a solution prepared by dissolving 21.07 g (0.3 mol) of hydroxylamine hydrochloride (molecular weight: 69.49) in 60 ml of water was added dropwise to prepare a monovalent copper complex solution. In a separate reaction vessel, 10.93 g (0.1 mol) of 3-aminophenol (molecular weight 109.13) and 100 ml of a 5 mol / l hydrochloric acid aqueous solution were charged, and the mixture was cooled with ice. A solution of 7.04 g (0.1 mol) of sodium nitrite (molecular weight 69.00) dissolved in 20 ml of water was added dropwise to form a diazonium salt solution. The resulting diazonium salt solution was diluted with 1300 ml of water, and the diluted diazonium salt solution was added dropwise over about 15 minutes to the ice-cooled copper complex solution, and the mixture was stirred for 3 hours. The solid was separated by filtration, and the solid was washed with 500 ml of 3% by weight aqueous ammonia, 200 ml of water, 300 ml of a 1 mol / l aqueous hydrochloric acid solution, and 200 ml of water. The washed solid is purified by column chromatography (silica gel packed column, developing solvent: ethyl acetate / hexane = 2/3), and 4.98 g (0.023 mol) of 3,3′-dihydroxyazobenzene (molecular weight 214.22) is obtained. Obtained. The yield was 46%.
[0061]
(S2) Preparation of 3′-hydroxy-3-methacryloyloxyazobenzene
1.28 g (6 mmol) of 3,3′-dihydroxyazobenzene (molecular weight 214.22), 15 ml of methylene chloride, and 3 ml of pyridine were charged into a reaction vessel, and the mixture was cooled with ice. Next, a solution prepared by dissolving 0.32 g (3 mmol) of methacryloyl chloride (molecular weight: 104.54) in 5 ml of methylene chloride was added dropwise over 30 minutes while stirring the mixture in the reaction vessel. After completion of the dropwise addition, the mixture was stirred at 0 ° C. for 30 minutes and at room temperature for 30 minutes. The solvent was distilled off, and the residue was dissolved in ethyl acetate and washed with water. After the ethyl acetate layer was dried over magnesium sulfate, the solvent was distilled off, and the residue was purified by column chromatography (silica gel packed column, developing solvent: ethyl acetate / hexane = 1/2) to give 3′-hydroxy-3 0.52 g (1.9 mmol) of methacryloyloxyazobenzene (molecular weight 270.29) was obtained. The yield was 63%.
[0062]
(S3) Preparation of cholesteryl carbonate 3- (3-methacryloyloxy) phenylazophenyl
0.23 g (0.8 mmol) of 3'-hydroxy-3-methacryloyloxyazobenzene, 15 ml of tetrahydrofuran, and 2 ml of triethylamine were added to the reaction vessel, and the mixture was cooled with ice. Next, 0.40 g (0.9 mmol) of cholesteryl chloroformate (molecular weight 449.12) was added to the mixture while stirring the mixture in the reaction vessel while maintaining the temperature of the mixture at 0 to 5 ° C. by ice cooling. It was added dropwise over 30 minutes. Thereafter, the liquid temperature was returned to room temperature, and the mixture was stirred at room temperature for 20 hours.
[0063]
The reaction mixture was transferred to a separating funnel, and a 5% by weight aqueous solution of hydrochloric acid, water, a 5% by weight aqueous solution of sodium hydroxide, and water were added thereto to carry out liquid separation, and the organic layer was separated. The separated organic layer was dried over magnesium sulfate for 30 minutes, and magnesium sulfate was removed by filtration. The residue obtained by evaporating the solvent from the filtrate obtained by this filtration was purified by column chromatography (silica gel packed column, developing solvent: ethyl acetate / chloroform / hexane = 1/2/17), and cholesteryl carbonate 3- (3 (Methacryloyloxy) phenylazophenyl (molecular weight 682.95) 0.21 g (0.3 mmol) was obtained. The yield was 38%.
[0064]
(S4) Preparation of polymer
Cholesteryl carbonate 3- (3-methacryloyloxy) phenylazophenyl 69.4 mg (0.1 mmol) was dissolved in 0.64 ml of hexane and 0.16 ml of toluene (solvent), and the resulting solution was added as a crosslinking agent. 377 mg (1.9 mmol) of ethylene glycol dimethacrylate (molecular weight 198.22) and 5.0 mg (0.02 mmol) of 2,2′-azobis (isobutyronitrile) (molecular weight 164.21) as a polymerization initiator And added.
[0065]
Then, the polymerization reaction was performed at 45 ° C. for 18 hours. The crude product was crushed in a mortar, washed with methanol and dried. This resulted in 411 mg of polymer.
[0066]
(S5) Hydrolysis of polymer
The obtained polymer was suspended in a methanol solution of 0.1 mmol / l sodium hydroxide, and hydrolyzed under basic conditions for 3 hours.
[0067]
Thereby, elimination of cholesterol accompanied by decarboxylation occurred, and 370 mg of a photosensitive imprinted polymer having a phenolic hydroxyl group remaining at the binding site was obtained.
[0068]
(S6) Molecular recognition ability of photosensitive imprinted polymer
The same reaction as in S2 to S4 was performed except that phenyl chloroformate was used in place of cholesteryl chloroformate, to synthesize an imprint polymer for comparison.
[0069]
When the photosensitive imprint polymer and the comparative imprint polymer were suspended in n-hexane and the binding experiment with cholesterol was performed, the photosensitive imprint polymer was compared with the comparative imprint polymer. Cholesterol binding was significantly higher. From this, it was confirmed that the photosensitive imprint polymer had a molecular recognition ability for recognizing cholesterol.
[0070]
(S7) Ultraviolet illumination of cholesterol bound to photosensitive imprinted polymer
Desorption by shooting
This photosensitive imprint polymer is suspended in n-hexane, cholesterol is bound in a dark place, and then irradiated with ultraviolet light having a wavelength of 330 nm for 18 minutes using a high-pressure mercury lamp, the bound cholesterol is released. Was confirmed.
[0071]
[Example 2]
In this example, a functional monomer having a cyclic boronic ester bond having a large binding constant was prepared using 4-nitrophenyl-α-D-mannopyranoside as a template molecule. The synthesis scheme of this functional monomer is shown below.
[0072]
Embedded image

[0073]
Then, a polymer is synthesized by polymerizing the above functional monomer using divinylbenzene and styrene which do not cause a decomposition reaction as a cross-linking agent, and a boronic ester in the obtained polymer is hydrolyzed to form a photosensitive photosensitive polymer. Printed polymers were synthesized.
[0074]
Since the photosensitive imprint polymer has a boronic acid moiety capable of forming a cyclic boronic ester with 4-nitrophenyl-α-D-mannopyranoside and an azobenzene moiety which undergoes cis-trans isomerization by light irradiation, It is expected that it has a molecular recognition ability to recognize 4-nitrophenyl-α-D-mannopyranoside, and that the molecular recognition ability at the binding site changes with the isomerization of the azobenzene site by light irradiation.
(S1) Azo coupling reaction
In 85 ml of a 2 mol / l hydrochloric acid aqueous solution, 4.30 g (25 mmol) of p-bromoaniline (molecular weight: 172.02) was added to 1.90 g (27.5 mmol) of sodium nitrite (molecular weight: 94.11) under the conditions of 0 ° C. and 15 minutes. ) To prepare an aqueous solution of p-bromobenzenediazonium chloride.
[0075]
Subsequently, 2.35 g (25 mmol) of phenol (molecular weight 94.11) and 2 g (50 mmol) of sodium hydroxide (molecular weight 40.00) were added to the obtained aqueous solution of p-bromobenzenediazonium chloride. The azo coupling reaction was performed under the conditions of minutes.
[0076]
By isolating the azo coupling reaction product from the crude product, 2.72 g (9.8 mmol) of 4-bromo-4′-hydroxyazobenzene (molecular weight 277.125) was obtained. The yield was 39%.
(S2) Convert to boronic ester
In a reaction vessel, 4-bromo-4'-hydroxyazobenzene 0.56 g (2.0 mmol), [1,1'-bis (diphenylphosphino) ferrocene] palladium (II) dichloride / methylene chloride (1: 1) complex (Molecular weight 816.64) 44 mg (0.06 mmol), bis (pinacolato) diboron (molecular weight 253.94) 0.56 g (2.2 mmol), potassium acetate (molecular weight 98.15) 0.31 g (3.1 mmol), Then, 10 ml of dry DMF (N, N-dimethylformamide) as a solvent was added, and the reaction mixture was heated at 80 ° C. for 18 hours under a nitrogen atmosphere. After confirming the completion of the reaction, the reaction mixture was diluted with diethyl ether, filtered through celite, and washed with water and brine. The washed solution was dried with magnesium sulfate, and the solvent was removed by vacuum suction. Thereafter, the residue was purified by recrystallization using a mixed solvent of ethyl acetate / hexane. Thereby, 0.60 g (1.9 mmol) of 4'-hydroxyazobenzene-4-boronic acid pinacolato (molecular weight 324.19) was obtained. The yield was 93%.
[0077]
Note that dppf indicates the ligand 1,1′-bis (diphenylphosphino) ferrocene.
(S3) Methacrylate formation
0.61 g (1.9 mmol) of 4'-hydroxyazobenzene-4-boronic acid pinacolato was dissolved in 10 ml of methylene chloride, and 1 ml of pyridine was added. To this solution, 0.27 g (2.6 mmol) of methacryloyl chloride (molecular weight 104.54) was added, and the mixture was stirred at 0 ° C. for 30 minutes and at room temperature for 1 hour to perform an esterification reaction.
[0078]
By isolating the methacrylic acid ester from the crude product, 0.58 g (1.5 mmol) of 4'-methacryloyloxyazobenzene-4-boronic acid pinacolato (molecular weight 392.26) was obtained. The yield was 79%.
(S4) Hydrolysis
0.58 g (1.5 mmol) of 4'-methacryloyloxyazobenzene-4-boronic acid pinacolato was hydrolyzed in a hydrochloric acid / acetone mixture at room temperature for 24 hours.
[0079]
By isolating the hydrolysis product from the crude product, 4'-methacryloyloxyazobenzene-4-boronic acid (molecular weight 310.12) as a functional monomer having a methacryloyl group and a boronic acid group in the molecule is obtained. .19 g were obtained. The yield was 41%.
(S5) Preparation of boronic acid cyclic ester monomer
In 20 ml of dry methylene chloride, 31 mg (0.1 mmol) of 4′-methacryloyloxyazobenzene-4-boronic acid and 15 mg of 4-nitrophenyl-α-D-mannopyranoside (molecular weight 301.30) (0.1 wt. 05 mmol) and stirred for 18 hours (see FIG. 2).
[0080]
By distilling off the solvent, a monomer in which two molecules of boronic acid formed a cyclic ester with 4-nitrophenyl-α-D-mannopyranoside (boronic acid cyclic ester monomer) (molecular weight: 849) shown in FIG. .41) 42 mg were obtained. The yield was 100%. This monomer was used for the next polymer synthesis without further purification.
(S6) Preparation of photosensitive imprint polymer
As shown in FIG. 2, the boronic acid cyclic ester monomer was polymerized to prepare a photosensitive imprint polymer.
[0081]
The polymerization was performed by the operation shown in FIG. That is, first, 42 mg (0.05 mmol) of a boronic acid cyclic ester monomer is dissolved in 0.8 ml of tetrahydrofuran, and 195 mg (1.5 mmol) of divinylbenzene as a crosslinking agent and 52 mg (0.5 mmol) of styrene are added to the obtained solution. And 5 mg (0.02 mmol) of 2,2′-azobis (2,4-dimethylvaleronitrile) as a polymerization initiator were added.
[0082]
Then, the polymerization reaction was performed at 45 ° C. for 18 hours. The polymer was isolated by crushing the crude product, washing and drying with methanol. This resulted in 41 mg of polymer.
(S7) Hydrolysis of cyclic boronic ester
The obtained polymer was hydrolyzed as shown in FIG. 2 and FIG. That is, the obtained polymer was suspended in 30 ml of acetone containing 5% by weight of hydrochloric acid to hydrolyze the cyclic boronic ester under acidic conditions for 16 hours.
[0083]
Thereby, the template molecule 4-nitrophenyl-α-D-mannopyranoside was eliminated from the polymer, and a photosensitive imprinted polymer was obtained.
(S8) Change in ultraviolet / visible absorption spectrum of photosensitive imprinted polymer
When the photosensitive imprint polymer was irradiated with ultraviolet light having a wavelength of 331 nm for 15 minutes, the absorption near the wavelength of 330 nm (attributable to the trans-form) was reduced and the absorption near the wavelength of 440 nm (cis) was observed, as shown in FIG. Due to the body) increased. As a result, it was confirmed that the trans form decreased and the cis form increased, that is, the isomerization of the azobenzene site occurred.
[0084]
The ultraviolet / visible absorption spectrum shown in FIG. 5 was measured in a state where the photosensitive imprint polymer was suspended in chloroform.
[0085]
【The invention's effect】
According to the present invention, as described above, a novel compound capable of selectively binding to a target molecule, having photosensitivity capable of inducing a physical / chemical change by light irradiation, and controlling the adsorption and desorption of a target molecule by light. A molecular recognition polymer can be provided.
[0086]
The solid phase extraction of the target molecule can be performed by solid-phase extraction of the molecular recognition polymer because the concentration and collection of the target molecule can be repeated by ON-OFF or wavelength change of light without performing a complicated operation of changing the solvent during solid phase extraction. Is considered to be extremely useful as a separating agent for In addition, by eliminating the need for solvent exchange in solid phase extraction, a solid phase extraction system that can be easily automated can be constructed.
[0087]
In addition, a molecular recognition polymer targeting a drug such as dopamine or the like is considered to be capable of carrying and transporting the drug, and being capable of adsorbing and desorbing the drug by ON-OFF by light, and is expected to be applied to a drug delivery system. it can.
[0088]
Further, according to the production method of the present invention, the above-mentioned novel molecular recognition polymer can be produced.
[Brief description of the drawings]
FIG. 1 is a diagram showing a method for producing a molecular recognition polymer according to one embodiment of the present invention.
FIG. 2 is a diagram illustrating a method for producing a molecular recognition polymer according to another embodiment of the present invention.
FIG. 3 is a view showing operations and conditions of a polymerization step and a desorption step in a method for producing a molecular recognition polymer according to another embodiment of the present invention. FIG.
FIG. 4 is a view showing one step of a method for producing a molecular recognition polymer according to another embodiment of the present invention.
FIG. 5 is a diagram showing a change in an ultraviolet / visible absorption spectrum when a photosensitive imprint polymer having an azobenzene moiety is irradiated with ultraviolet light.
FIG. 6 is a diagram showing a state in which the molecular recognition ability of the photosensitive imprint polymer in one embodiment of the present invention is changed by an external stimulus (light).

Claims (10)

A molecular recognition polymer having a functional group that specifically and reversibly binds to a target molecule and a light-sensitive site that isomerizes upon irradiation with light. The molecular recognition polymer according to claim 1, wherein the functional group includes a covalent functional group that forms a reversible covalent bond specifically with a target molecule. The target molecule has a plurality of hydroxyl groups,
The molecular recognition polymer according to claim 2, wherein the covalent functional group includes a boronic acid group that forms a cyclic boronic ester by bonding to a plurality of hydroxyl groups of the target molecule. The molecular recognition polymer according to claim 1, wherein the functional group includes a non-covalent functional group that forms a reversible non-covalent bond specifically with a target molecule. The target molecule has a hydroxyl group,
5. The molecular recognition polymer according to claim 4, wherein the non-covalent functional group includes a hydroxyl group that forms a reversible hydrogen bond specifically with a hydroxyl group of the target molecule. The molecular recognition polymer according to any one of claims 1 to 5, wherein the light-sensitive moiety includes an azobenzene moiety. The molecular recognition polymer according to any one of claims 1 to 5, wherein the light-sensitive site includes a spiropyran site. It is a manufacturing method of the molecular recognition polymer of Claim 2, Comprising:
A preparing step of preparing an unsaturated monomer having the covalent functional group and the photosensitive moiety,
A binding step of binding the unsaturated monomer and the target molecule by a reversible covalent bond,
A polymerization step of copolymerizing an unsaturated monomer bonded to a target molecule with a crosslinking agent to obtain a polymer,
A elimination step of elimination of a target molecule from the obtained polymer. It is a manufacturing method of the molecular recognition polymer of Claim 4, Comprising:
A preparation step of preparing an unsaturated monomer having a non-covalent functional group and a light-sensitive site,
A polymerization step of copolymerizing the unsaturated monomer with a crosslinking agent in the presence of a target molecule to obtain a polymer,
A elimination step of elimination of a target molecule from the obtained polymer. It is a manufacturing method of the molecular recognition polymer of Claim 5, Comprising:
A step of preparing a carbonate ester of the photosensitive monomer and the unsaturated monomer having a hydroxyl group, and a target molecule having the hydroxyl group,
A polymerization step of copolymerizing the carbonate with a crosslinking agent to obtain a polymer,
A step of removing a target molecule from the obtained polymer by decarboxylation to release a hydroxyl group derived from an unsaturated monomer.

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