JPWO2013146142A1 - フッ素アパタイト、吸着装置および分離方法 - Google Patents
フッ素アパタイト、吸着装置および分離方法 Download PDFInfo
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- JPWO2013146142A1 JPWO2013146142A1 JP2014507602A JP2014507602A JPWO2013146142A1 JP WO2013146142 A1 JPWO2013146142 A1 JP WO2013146142A1 JP 2014507602 A JP2014507602 A JP 2014507602A JP 2014507602 A JP2014507602 A JP 2014507602A JP WO2013146142 A1 JPWO2013146142 A1 JP WO2013146142A1
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- Prior art keywords
- fluorapatite
- charged substance
- adsorbent
- substance
- protein
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
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- 238000001179 sorption measurement Methods 0.000 title claims abstract description 73
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- 229940077441 fluorapatite Drugs 0.000 claims abstract description 137
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- XYJRXVWERLGGKC-UHFFFAOYSA-D pentacalcium;hydroxide;triphosphate Chemical compound [OH-].[Ca+2].[Ca+2].[Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O XYJRXVWERLGGKC-UHFFFAOYSA-D 0.000 claims description 52
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Classifications
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- B01D15/26—Selective adsorption, e.g. chromatography characterised by the separation mechanism
- B01D15/38—Selective adsorption, e.g. chromatography characterised by the separation mechanism involving specific interaction not covered by one or more of groups B01D15/265 - B01D15/36
- B01D15/3847—Multimodal interactions
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- B01J20/02—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising inorganic material
- B01J20/04—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising inorganic material comprising compounds of alkali metals, alkaline earth metals or magnesium
- B01J20/048—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising inorganic material comprising compounds of alkali metals, alkaline earth metals or magnesium containing phosphorus, e.g. phosphates, apatites, hydroxyapatites
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- B01J20/28—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof characterised by their form or physical properties
- B01J20/28054—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof characterised by their form or physical properties characterised by their surface properties or porosity
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- B01J20/281—Sorbents specially adapted for preparative, analytical or investigative chromatography
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- C01B—NON-METALLIC ELEMENTS; COMPOUNDS THEREOF; METALLOIDS OR COMPOUNDS THEREOF NOT COVERED BY SUBCLASS C01C
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Abstract
Description
前記フッ素アパタイトは、下記一般式で表され、
Ca10(PO4)6(OH)2−2xF2X
前記一般式中、xは0.05≦x≦0.76の関係を満足することを特徴とするフッ素アパタイト。
Ca10(PO4)6(OH)2−2xF2Xの一般式で表され、前記一般式中、xは0.05≦x≦0.76であるフッ素アパタイトで構成された充填剤を充填空間の少なくとも一部に充填してなる吸着装置内に、前記試料液を供給する供給工程と、
前記吸着装置内に、緩衝液を供給して、前記吸着装置内から流出する流出液を得る工程と、
前記流出液を所定量ずつ分画することにより、この分画された各画分の流出液中に、前記試料液から前記帯電物質を分離する工程とを有することを特徴とする分離方法。
図1は、本発明で用いる吸着装置の一例を示す縦断面図である。なお、以下の説明では、図1中の上側を「流入側」、下側を「流出側」と言う。
フッ素アパタイトは、ハイドロキシアパタイトが有する水酸基の少なくとも一部がフッ素原子で置換されていることに起因して、ハイドロキシアパタイトと比較して、耐酸性に優れるものとなる。その結果、吸着剤3に吸着した酸性タンパク質を溶出液で溶出させる際に、用いる溶出液の選択の幅が広がる。すなわち、溶出液のpH値を広範囲に設定することができるため、高精度で酸性タンパク質が精製される条件に、溶出液をより確実に設定することができるようになる。
フッ素アパタイトは、ハイドロキシアパタイトが有する水酸基の少なくとも一部がフッ素原子で置換されていることに起因して、ハイドロキシアパタイトと比較して、耐酸性に優れるものとなる。その結果、吸着剤3に吸着した塩基性タンパク質を溶出液で溶出させる際に、用いる溶出液の選択の幅が広がる。すなわち、溶出液のpH値を広範囲に設定することができるため、高精度で塩基性タンパク質が精製される条件に、溶出液をより確実に設定することができるようになる。さらに、塩基性タンパク質の精製の過程で、一時的に溶出液のpH値が酸性側に変化したとしても、吸着剤3の変質(溶解等)を好適に防止することができる。このように、水酸基をフッ素原子で置換することにより、フッ素アパタイトの耐酸性が向上する。
次に、このような吸着装置1を用いた酸性タンパク質の分離方法(本発明の分離方法)について説明する。
まず、精製すべき酸性タンパク質(抗体等)と、酸性タンパク質以外の混入物(物質)とを含有する試料液を用意する。
次に、得られた試料液を、流入管24およびフィルタ部材4を介して吸着剤3に供給して、カラム2(吸着装置1)内を通過させて、吸着剤3に接触させる。
次に、流入管24からカラム2内に、酸性タンパク質を溶出させるための溶出液として例えばリン酸系緩衝液を供給する。そして、カラム2内から流出管25を介して流出する流出液を、所定量ずつ分画(採取)する。これにより、吸着剤3に吸着している酸性タンパク質および混入物は、それぞれ、それぞれが有する吸着剤3に対する吸着力の差に応じて、各分画内に溶解した状態で回収(分離)される。
次に、このような吸着装置1を用いた塩基性タンパク質の分離方法(本発明の分離方法)について説明する。
まず、精製すべき塩基性タンパク質(抗体等)と、塩基性タンパク質以外の混入物(物質)とを含有する試料液を用意する。
次に、得られた試料液を、流入管24およびフィルタ部材4を介して吸着剤3に供給して、カラム2(吸着装置1)内を通過させて、吸着剤3に接触させる。
次に、流入管24からカラム2内に、塩基性タンパク質を溶出させるための溶出液として例えばリン酸系緩衝液を供給する。そして、カラム2内から流出管25を介して流出する流出液を、所定量ずつ分画(採取)する。これにより、吸着剤3に吸着している塩基性タンパク質および混入物は、それぞれ、それぞれが有する吸着剤3に対する吸着力の差に応じて、各分画内に溶解した状態で回収(分離)される。
次に、前述したようなフッ素アパタイトは、いかなる製造方法を用いて製造してもよいが、例えば、以下のようなIまたはIIの方法により製造することができる。
A1:まず、ハイドロキシアパタイトを含むスラリーを調製する。
Ca10(PO4)6(OH)2−2xF2X
[ただし、式中、xは0.05≦x≦0.76である。]
[1]まず、水酸化カルシウムを純水に懸濁させ、その中へ、リン酸水溶液を滴下していき、かつ十分に撹拌した。これにより、10wt%のハイドロキシアパタイト一次粒子を含むスラリー200Lを得た。
2−1.Ca溶出による耐酸性評価
まず、各焼結粒子1〜5を、それぞれ、ステンレスカラム(内径4.0mm×長さ100mm)の充填空間にほぼ満量となるように充填して、焼結粒子1〜5が充填されたサンプルNo.1〜5のカラムを製造した。
まず、各焼結粒子1〜5を、それぞれ、ステンレスカラム(内径4.6mm×長さ35mm)の充填空間にほぼ満量となるように充填して、焼結粒子1〜5が充填されたサンプルNo.1〜5のカラムを製造した。
まず、各焼結粒子1〜5を、それぞれ、ステンレスカラム(内径4.6mm×長さ35mm)の充填空間にほぼ満量となるように充填して、焼結粒子1〜5が充填されたサンプルNo.1〜5のカラムを製造した。
以上のことから、焼結粒子におけるフッ素原子の置換率を5%以上に設定することで、焼結粒子(フッ素アパタイト)を優れた耐酸性を有するものとすることができる。さらに、置換率を6%以上、76%以下に設定することで、焼結粒子(フッ素アパタイト)を優れた塩基性タンパク質吸着能を有するものとすることができる。そのため、焼結粒子におけるフッ素原子の置換率を6%以上、76%以下に設定することで、結果的に、焼結粒子(フッ素アパタイト)は、耐酸性および塩基性タンパク質吸着能の双方に優れた特性を発揮するものとなる。
Claims (12)
- 帯電物質と、該帯電物質以外からなる物質とを含有する試料液中から、前記帯電物質を分離するために用いられるフッ素アパタイトであって、
前記フッ素アパタイトは、下記一般式で表され、
Ca10(PO4)6(OH)2−2xF2X
前記一般式中、xは0.05≦x≦0.76の関係を満足することを特徴とするフッ素アパタイト。 - 前記帯電物質は、負帯電物質であり、
前記一般式中のxは、0.05≦x≦0.55の関係を満足する請求項1に記載のフッ素アパタイト。 - 前記負帯電物質は、酸性タンパク質である請求項2に記載のフッ素アパタイト。
- 前記フッ素アパタイトは、粒状をなし、その比表面積が30m2/g以上、50m2/g以下である請求項3に記載のフッ素アパタイト。
- 前記帯電物質は、正帯電物質であり、
前記一般式中のxは、0.06≦x≦0.76の関係を満足する請求項1に記載のフッ素アパタイト。 - 前記正帯電物質は、塩基性タンパク質である請求項5に記載のフッ素アパタイト。
- 前記フッ素アパタイトは、粒状をなし、その比表面積が25m2/g以上、50m2/g以下である請求項6に記載のフッ素アパタイト。
- 前記フッ素アパタイトは、ハイドロキシアパタイトを含むスラリーと、フッ化水素を含有するフッ化水素含有液とを混合した反応液中において、前記ハイドロキシアパタイトと前記フッ化水素とを反応させることにより、前記水酸基の少なくとも一部を、フッ素原子で置換して得られたものである請求項1に記載のフッ素アパタイト。
- 請求項1に記載のフッ素アパタイトを吸着剤として備えることを特徴とする吸着装置。
- 帯電物質と、該帯電物質以外からなる物質とを含有する試料液中から、前記帯電物質を分離する分離方法であって、
Ca10(PO4)6(OH)2−2xF2Xの一般式で表され、前記一般式中、xは0.05≦x≦0.76の関係を満足する、フッ素アパタイトで構成された充填剤を充填空間の少なくとも一部に充填してなる吸着装置内に、前記試料液を供給する工程と、
前記吸着装置内に、緩衝液を供給して、前記吸着装置内から流出する流出液を得る工程と、
前記流出液を所定量ずつ分画することにより、この分画された各画分の流出液中に、前記試料液から前記帯電物質を分離する工程とを有することを特徴とする分離方法。 - 前記帯電物質は、負帯電物質であり、
前記一般式中のxは、0.05≦x≦0.55の関係を満足する請求項10に記載のフッ素アパタイト。 - 前記帯電物質は、正帯電物質であり、
前記一般式中のxは、0.06≦x≦0.76の関係を満足する請求項10に記載のフッ素アパタイト。
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JPH01264915A (ja) * | 1988-04-15 | 1989-10-23 | Asahi Optical Co Ltd | フッ素アパタイトの製造方法 |
JPH0247551A (ja) * | 1988-08-09 | 1990-02-16 | Asahi Optical Co Ltd | 液体クロマトグラフィー用カラム |
JP2009035464A (ja) * | 2007-08-03 | 2009-02-19 | Hoya Corp | フッ素アパタイトの製造方法、フッ素アパタイトおよび吸着装置 |
JP2009084119A (ja) * | 2007-10-01 | 2009-04-23 | Hoya Corp | フッ素アパタイトの乾燥粒子および吸着装置 |
JP2010083713A (ja) * | 2008-09-30 | 2010-04-15 | Hoya Corp | フッ素アパタイト粉体の製造方法、フッ素アパタイト粉体および吸着装置 |
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JPS6413453A (en) * | 1987-07-08 | 1989-01-18 | Asahi Optical Co Ltd | Packing agent for liquid chromatography |
JPH01264915A (ja) * | 1988-04-15 | 1989-10-23 | Asahi Optical Co Ltd | フッ素アパタイトの製造方法 |
JPH0247551A (ja) * | 1988-08-09 | 1990-02-16 | Asahi Optical Co Ltd | 液体クロマトグラフィー用カラム |
JP2009035464A (ja) * | 2007-08-03 | 2009-02-19 | Hoya Corp | フッ素アパタイトの製造方法、フッ素アパタイトおよび吸着装置 |
JP2009084119A (ja) * | 2007-10-01 | 2009-04-23 | Hoya Corp | フッ素アパタイトの乾燥粒子および吸着装置 |
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