JPH05339297A - Separation of ovotransferrin - Google Patents
Separation of ovotransferrinInfo
- Publication number
- JPH05339297A JPH05339297A JP4171823A JP17182392A JPH05339297A JP H05339297 A JPH05339297 A JP H05339297A JP 4171823 A JP4171823 A JP 4171823A JP 17182392 A JP17182392 A JP 17182392A JP H05339297 A JPH05339297 A JP H05339297A
- Authority
- JP
- Japan
- Prior art keywords
- solution
- ovotransferrin
- avidin
- protein
- lysozyme
- 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.)
- Pending
Links
- 108010026206 Conalbumin Proteins 0.000 title claims abstract description 28
- 238000000926 separation method Methods 0.000 title description 8
- 108090001008 Avidin Proteins 0.000 claims abstract description 11
- 102000016943 Muramidase Human genes 0.000 claims abstract description 11
- 108010014251 Muramidase Proteins 0.000 claims abstract description 11
- 108010062010 N-Acetylmuramoyl-L-alanine Amidase Proteins 0.000 claims abstract description 11
- 235000010335 lysozyme Nutrition 0.000 claims abstract description 11
- 229960000274 lysozyme Drugs 0.000 claims abstract description 11
- 239000004325 lysozyme Substances 0.000 claims abstract description 11
- 229920002684 Sepharose Polymers 0.000 claims abstract description 8
- 238000001042 affinity chromatography Methods 0.000 claims abstract description 8
- 239000012460 protein solution Substances 0.000 claims abstract description 8
- 238000000034 method Methods 0.000 claims description 11
- 108010000912 Egg Proteins Proteins 0.000 claims description 8
- 102000002322 Egg Proteins Human genes 0.000 claims description 8
- 125000004435 hydrogen atom Chemical group [H]* 0.000 claims description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 abstract description 16
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 abstract description 13
- 239000000243 solution Substances 0.000 abstract description 11
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 abstract description 9
- 239000011780 sodium chloride Substances 0.000 abstract description 8
- 230000001766 physiological effect Effects 0.000 abstract description 7
- 102000004169 proteins and genes Human genes 0.000 abstract description 7
- 108090000623 proteins and genes Proteins 0.000 abstract description 7
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 abstract description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 abstract description 4
- 239000007864 aqueous solution Substances 0.000 abstract description 3
- 229910000147 aluminium phosphate Inorganic materials 0.000 abstract description 2
- 239000007853 buffer solution Substances 0.000 abstract description 2
- 239000003480 eluent Substances 0.000 abstract description 2
- 239000000463 material Substances 0.000 abstract description 2
- 230000002542 deteriorative effect Effects 0.000 abstract 1
- 239000008213 purified water Substances 0.000 abstract 1
- 239000010414 supernatant solution Substances 0.000 abstract 1
- 239000000499 gel Substances 0.000 description 5
- 235000018102 proteins Nutrition 0.000 description 5
- NLXLAEXVIDQMFP-UHFFFAOYSA-N Ammonia chloride Chemical compound [NH4+].[Cl-] NLXLAEXVIDQMFP-UHFFFAOYSA-N 0.000 description 4
- 239000000872 buffer Substances 0.000 description 4
- 239000008363 phosphate buffer Substances 0.000 description 4
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 description 3
- QCVGEOXPDFCNHA-UHFFFAOYSA-N 5,5-dimethyl-2,4-dioxo-1,3-oxazolidine-3-carboxamide Chemical compound CC1(C)OC(=O)N(C(N)=O)C1=O QCVGEOXPDFCNHA-UHFFFAOYSA-N 0.000 description 3
- 108010088751 Albumins Proteins 0.000 description 3
- 102000009027 Albumins Human genes 0.000 description 3
- 235000014103 egg white Nutrition 0.000 description 3
- 210000000969 egg white Anatomy 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 239000002244 precipitate Substances 0.000 description 3
- 238000001556 precipitation Methods 0.000 description 3
- JVIPLYCGEZUBIO-UHFFFAOYSA-N 2-(4-fluorophenyl)-1,3-dioxoisoindole-5-carboxylic acid Chemical compound O=C1C2=CC(C(=O)O)=CC=C2C(=O)N1C1=CC=C(F)C=C1 JVIPLYCGEZUBIO-UHFFFAOYSA-N 0.000 description 2
- 229920002134 Carboxymethyl cellulose Polymers 0.000 description 2
- 229920001425 Diethylaminoethyl cellulose Polymers 0.000 description 2
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 2
- 235000019270 ammonium chloride Nutrition 0.000 description 2
- 239000001768 carboxy methyl cellulose Substances 0.000 description 2
- 235000010948 carboxy methyl cellulose Nutrition 0.000 description 2
- 239000008112 carboxymethyl-cellulose Substances 0.000 description 2
- 238000004255 ion exchange chromatography Methods 0.000 description 2
- 229910021645 metal ion Inorganic materials 0.000 description 2
- 239000000843 powder Substances 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 239000007974 sodium acetate buffer Substances 0.000 description 2
- JYEUMXHLPRZUAT-UHFFFAOYSA-N 1,2,3-triazine Chemical compound C1=CN=NN=C1 JYEUMXHLPRZUAT-UHFFFAOYSA-N 0.000 description 1
- 101000766307 Gallus gallus Ovotransferrin Proteins 0.000 description 1
- OWYWGLHRNBIFJP-UHFFFAOYSA-N Ipazine Chemical compound CCN(CC)C1=NC(Cl)=NC(NC(C)C)=N1 OWYWGLHRNBIFJP-UHFFFAOYSA-N 0.000 description 1
- 125000000729 N-terminal amino-acid group Chemical group 0.000 description 1
- 108010058846 Ovalbumin Proteins 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- 102000035195 Peptidases Human genes 0.000 description 1
- JQYMGXZJTCOARG-UHFFFAOYSA-N Reactive blue 2 Chemical compound C1=2C(=O)C3=CC=CC=C3C(=O)C=2C(N)=C(S(O)(=O)=O)C=C1NC(C=C1S(O)(=O)=O)=CC=C1NC(N=1)=NC(Cl)=NC=1NC1=CC=CC(S(O)(=O)=O)=C1 JQYMGXZJTCOARG-UHFFFAOYSA-N 0.000 description 1
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 1
- 239000011543 agarose gel Substances 0.000 description 1
- 235000004279 alanine Nutrition 0.000 description 1
- 125000003295 alanine group Chemical group N[C@@H](C)C(=O)* 0.000 description 1
- 235000001014 amino acid Nutrition 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 239000003638 chemical reducing agent Substances 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 150000004696 coordination complex Chemical class 0.000 description 1
- 238000010168 coupling process Methods 0.000 description 1
- 239000003398 denaturant Substances 0.000 description 1
- 238000000502 dialysis Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000010828 elution Methods 0.000 description 1
- 239000012149 elution buffer Substances 0.000 description 1
- 238000011067 equilibration Methods 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 238000003505 heat denaturation Methods 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 230000001771 impaired effect Effects 0.000 description 1
- 229910052742 iron Inorganic materials 0.000 description 1
- 239000003446 ligand Substances 0.000 description 1
- 229940092253 ovalbumin Drugs 0.000 description 1
- 108090000765 processed proteins & peptides Proteins 0.000 description 1
- 235000004252 protein component Nutrition 0.000 description 1
- 229940024999 proteolytic enzymes for treatment of wounds and ulcers Drugs 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 239000001632 sodium acetate Substances 0.000 description 1
- 235000017281 sodium acetate Nutrition 0.000 description 1
- 239000001488 sodium phosphate Substances 0.000 description 1
- 229910000162 sodium phosphate Inorganic materials 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 238000011282 treatment Methods 0.000 description 1
- RYFMWSXOAZQYPI-UHFFFAOYSA-K trisodium phosphate Chemical compound [Na+].[Na+].[Na+].[O-]P([O-])([O-])=O RYFMWSXOAZQYPI-UHFFFAOYSA-K 0.000 description 1
Landscapes
- Peptides Or Proteins (AREA)
Abstract
Description
【０００１】[0001]
【産業上の利用分野】本発明は、オボトランスフェリン
の分離方法に関する。更に詳しくは、リゾチームおよび
アビジンを除去した卵白たん白質溶液からのオボトラン
スフェリンの分離方法に関する。TECHNICAL FIELD The present invention relates to a method for separating ovotransferrin. More specifically, it relates to a method for separating ovotransferrin from an albumin protein solution from which lysozyme and avidin have been removed.
【０００２】[0002]
【従来の技術】卵白中には約40種類のたん白質が含まれ
ており、その内工業的に分離が容易で、有用な生理活性
を有するリゾチームやアビジンが利用されているが、リ
ゾチームやアビジンを分離した後の卵白溶液中にも、有
用な生理活性を有するたん白質であるオボトランスフェ
リンが12%程度存在している。2. Description of the Related Art Egg white contains about 40 kinds of proteins, and lysozyme and avidin, which have industrially easy separation and useful physiological activity, are used. About 12% of ovotransferrin, which is a protein having useful physiological activity, is also present in the egg white solution after the separation of the.
【０００３】オボトランスフェリン(コンアルブミン)
は、鉄を強固に結合させた分子量約７万のたん白質であ
り、N-末端アミノ酸残基はアラニンであり、糖を含んで
いる。アミノ酸組成、ペプチドマップは、血清トランス
フェリンにきわめてよく似ており、免疫学的にも両者は
類似しており、糖鎖部分のみが異なると考えられてい
る。このオボトランスフェリンは、種々の金属イオン
と、たん白質１分子に付き２ヶ所で結合する能力も有し
ており、元来熱変性し易いオボトランスフェリンも、金
属イオンと複合体を形成すると、熱変性温度を上昇させ
たり、たん白分解酵素や種々の変性処理に対する抵抗性
も高められる。また、オボトランスフェリンのS-S結合
は、変性剤なしでも還元剤だけで容易に切断されるが、
複合体を形成した状態では還元に対してきわめて高い抵
抗性を示すようになる。これは、金属複合体となること
によって、分子構造が密になることを示しており、この
ような安定性は卵白の殺菌にとって実用上重要である。Ovotransferrin (conalbumin)
Is a protein having a molecular weight of about 70,000 to which iron is bound tightly, the N-terminal amino acid residue is alanine, and it contains a sugar. The amino acid composition and peptide map are very similar to serum transferrin, immunologically similar to each other, and it is considered that only the sugar chain portion is different. This ovotransferrin also has the ability to bind various metal ions at two positions per protein molecule, and even ovotransferrin, which is naturally susceptible to heat denaturation, forms heat denaturants when it forms a complex with metal ions. It also increases the temperature and resistance to proteolytic enzymes and various denaturing treatments. In addition, the SS bond of ovotransferrin is easily cleaved only by the reducing agent without the denaturing agent,
When the complex is formed, it becomes extremely resistant to reduction. This indicates that the molecular structure becomes dense by forming a metal complex, and such stability is practically important for sterilization of egg white.
【０００４】このような特性を有するオボトランスフェ
リンについては、従来工業的に大量廉価に分離する技術
が確立されてはおらず、以下の如き実験室的レベルの分
離法にとどまっている。With respect to ovotransferrin having such characteristics, a technique for industrially separating a large amount of the ovotransferrin at low cost has not been established, and only the following laboratory-level separation method is available.
【０００５】その代表的な分離法は、沈殿法とイオン交
換クロマトグラフィー法である。Typical separation methods are precipitation method and ion exchange chromatography method.
【０００６】沈殿法では、リゾチーム、アビジン除去済
みの卵白たん白質溶液に酢酸を加えてpHを5.0に調整
し、3時間放置した後、その上層を遠心機にかけ、沈殿
を落して更にその上層を回収し、そこに塩化ナトリウム
を3重量%になるように加え、その溶液のpHを3.5に調整
して一夜放置し、遠心機にかけて上層を捨て、沈殿物と
してオボトランスフェリンを得るという方法がとられて
いる。[0006] In the precipitation method, acetic acid is added to the egg white protein solution from which lysozyme and avidin have been removed to adjust the pH to 5.0, and the mixture is left for 3 hours. The method is to collect, add sodium chloride to 3% by weight, adjust the pH of the solution to 3.5, leave it overnight, centrifuge and discard the upper layer to obtain ovotransferrin as a precipitate. ing.
【０００７】このような一連の工程として行われる沈殿
法は、試料を一度に大量に処理できるという利点を有す
るものの、オボトランスフェリンを完全に分離する迄に
長時間を要し、非効率的であるばかりではなく、オボト
ランスフェリンをpH3.5という低pH状態にさらすため、
それの生理活性が損なわれるおそれがある。The precipitation method, which is carried out as such a series of steps, has the advantage that a large amount of sample can be treated at one time, but it takes a long time to completely separate ovotransferrin and is inefficient. Not only that, to expose ovotransferrin to a low pH of 3.5,
Its physiological activity may be impaired.
【０００８】一方、イオン交換クロマトグラフィー法で
は、カルボキシルメチルセルロースまたはジエチルアミ
ノエチルセルロースを担体としており、オボトランスフ
ェリンの分離状態や回収率は良いものの、１回の操作に
カルボキシメチルセルロースの場合で約25時間、ジエチ
ルアミノエチルセルロースの場合で約18時間を必要とす
るため、非効率的である。また、溶出液として塩化アン
モニウムを用いているため、分離されたオボトランスフ
ェリンが含まれている溶液中の塩化アンモニウムを透析
などにより除かなければならず、この点でも非効率的で
ある。On the other hand, in the ion exchange chromatography method, carboxymethylcellulose or diethylaminoethylcellulose is used as a carrier, and although the separation state and recovery rate of ovotransferrin are good, diethylaminoethylcellulose takes about 25 hours in the case of carboxymethylcellulose in one operation. In case of about 18 hours, it is inefficient. Further, since ammonium chloride is used as the eluent, ammonium chloride in the separated solution containing ovotransferrin must be removed by dialysis or the like, which is also inefficient.
【０００９】[0009]
【発明が解決しようとする課題】本発明の目的は、リゾ
チームやアビジンを除去した卵白たん白質溶液からオボ
トランスフェリンを分離するに際し、操作に長時間を要
せず、しかもオボトランスフェリンの生理活性を損なわ
ない分離方法を提供することにある。DISCLOSURE OF THE INVENTION An object of the present invention is to eliminate the ovotransferrin from an albumin protein solution from which lysozyme and avidin have been removed, which does not require a long time for operation, and impairs the physiological activity of ovotransferrin. There is no separation method.
【００１０】[0010]
【課題を解決するための手段】かかる本発明の目的は、
リゾチームおよびアビジンを除去した卵白たん白質溶液
について、ブルー・セファローズCL-6Bを担体とするア
フィニティークロマトグラフィーを適用し、オボトラン
スフェリンを分離する方法によって達成される。The object of the present invention is as follows.
This is achieved by a method of separating ovotransferrin by applying affinity chromatography using Blue Sepharose CL-6B as a carrier to an egg protein solution from which lysozyme and avidin have been removed.
【００１１】担体として用いられるブルー・セファロー
ズCL-6BBlue Sepharose CL-6B used as a carrier
【化２】 (ここで、R1，R2は水素原子または−SO2ONa基である)
は、架橋アガロースゲルであるセファローズCL-6Bに、
トリアジンカップリング法によりチバクロン・ブルー3G
A(チバ・ガイギー社製品；上記化学式においてSepharos
e CL-6Bを除いた部分)を共有結合させたものであり、こ
の色素部分がリガンドとして作用する。[Chemical 2] (Here, R 1 and R 2 are a hydrogen atom or a —SO 2 ONa group)
Is a cross-linked agarose gel Sepharose CL-6B,
Cibacron blue 3G by triazine coupling method
A (Ciba Geigy product; Sepharos in the above formula)
(a part excluding CL-6B) is covalently bonded, and this dye part acts as a ligand.
【００１２】このブルー・セファローズCL-6Bは、凍結
乾燥粉末25g包装で市販されており、これは膨潤すると
約90mlのゲルに相当し、カップリングしている色素の濃
度は膨潤ゲル１mlに対し約2μMである。それの使用に際
しては、必要量の凍結乾燥品を蒸留水で15分間膨潤し、
グラスフィルタ上で洗浄する。水は、1gの乾燥粉末当り
約200mlが数回に分けて加えられ、1gの凍結乾燥物質は
最終的には3.5mlのゲル体積になる。このようなゲルを
カラムに充填し、開始緩衝液で洗浄して、アフィニティ
ークロマトグラフィーに供する。This Blue Sepharose CL-6B is commercially available in a package of 25 g of freeze-dried powder, which when swollen corresponds to about 90 ml of gel, and the concentration of the coupled dye is 1 ml of swollen gel. It is about 2 μM. When using it, swell the required amount of lyophilized product with distilled water for 15 minutes,
Wash on glass filter. Water is added in several divided doses, about 200 ml per gram of dry powder, so that 1 g of lyophilized material results in a final gel volume of 3.5 ml. Such gel is packed in a column, washed with starting buffer and subjected to affinity chromatography.
【００１３】ゲルの開始緩衝液および溶出バッファーと
しては、トリス-酢酸、酢酸ナトリウム、リン酸、リン
酸ナトリウム、トリス-塩酸などが好んで用いられる。As the gel starting buffer and elution buffer, Tris-acetic acid, sodium acetate, phosphoric acid, sodium phosphate, Tris-hydrochloric acid and the like are preferably used.
【００１４】このようにして調製された担体を用いての
アフィニティークロマトグラフィーは、次のようにして
行われる。 (1)リゾチーム、アビジン除去済みの卵白たん白質溶液
(pH7.0)に酢酸を加え、pHを5.0に調整後3時間放置す
る。 (2)遠心機で沈殿を落し、上層を回収する。 (3)0.1N水酸化ナトリウム水溶液で上層のpHを7.0に調整
する。 (4)10mMリン酸緩衝液(pH7.0)などをバッファーとするア
フィニティークロマトグラフィーを行い、オボアルブミ
ン、オボトランスフェリンなどの未吸着成分を除いた
後、1M NaClを含む10mMリン酸緩衝液(PH7.0)などに用い
て、吸着成分としてのオボトランスフェリンを分離す
る。 (5)使用済みカラムは、0.5M NaClを含む10mM酢酸ナトリ
ウム緩衝液(PH4.5)、0.5M NaClを含むトリス−塩酸緩衝
液(PH8.5)などで洗浄する。Affinity chromatography using the carrier thus prepared is carried out as follows. (1) Egg protein protein solution with lysozyme and avidin removed
Acetic acid is added to (pH 7.0) to adjust the pH to 5.0, and the mixture is left for 3 hours. (2) Drop the precipitate with a centrifuge and collect the upper layer. (3) Adjust the pH of the upper layer to 7.0 with 0.1N sodium hydroxide aqueous solution. (4) Affinity chromatography using 10 mM phosphate buffer (pH 7.0) as a buffer to remove unadsorbed components such as ovalbumin and ovotransferrin, and then 10 mM phosphate buffer containing 1 M NaCl (PH7 .0) etc. to separate ovotransferrin as an adsorbing component. (5) The used column is washed with 10 mM sodium acetate buffer (PH4.5) containing 0.5 M NaCl, Tris-hydrochloric acid buffer (PH8.5) containing 0.5 M NaCl, and the like.
【００１５】[0015]
【発明の効果】リゾチームおよびアビジン除去済みの卵
白たん白質溶液から選択的にオボトランスフェリンを分
離するに際し、コンピュータなどで制御可能なアフィニ
ティークロマトグラフィーを、ブルー・セファローズCL
-6Bを担体として行うことにより、約8時間程度で分離操
作を行うことができ、従来法よりは著しく短時間でしか
も生理活性を損なわせないでオボトランスフェリンを得
ることができる。[Effect of the Invention] When selectively separating ovotransferrin from an albumin protein solution from which lysozyme and avidin have been removed, affinity chromatography that can be controlled by a computer or the like is applied to Blue Sepharose CL.
By using -6B as a carrier, the separation operation can be performed in about 8 hours, and ovotransferrin can be obtained in a significantly shorter time than the conventional method without impairing physiological activity.
【００１６】[0016]
【実施例】次に、実施例について本発明を説明する。EXAMPLES The present invention will now be described with reference to examples.
【００１７】実施例 リゾチーム、アビジン除去済みの卵白たん白質に純水を
加え、約5倍に希釈した後、0.2%酢酸水溶液を加えてpH
5.0とし、約3時間放置して上層を回収した。この上層を
遠心機にかけて沈殿を落し、更にその上層を回収して、
そのpHを0.1N水酸化ナトリウム水溶液で7.0に調整し
た。Example Pure water was added to the egg white protein from which lysozyme and avidin had been removed, diluted to about 5 times, and a 0.2% acetic acid aqueous solution was added to adjust the pH.
It was set to 5.0 and left for about 3 hours to recover the upper layer. The upper layer is centrifuged to remove the precipitate, and the upper layer is recovered,
The pH was adjusted to 7.0 with 0.1N aqueous sodium hydroxide solution.
【００１８】ブルー・セファローズCL-6Bが充填されて
いるカラム(容量500ml、内径30mm、長さ180mm)を、10mM
リン酸緩衝液(pH7.0)で平衡化した後、上記上層液(pH7.
0)100mlをカラムに流入させる。約１時間放置後、溶出
液からたん白質成分が検出されなくなる迄(紫外線モニ
タで管理)、2M NaClを含む10mMリン酸緩衝液を流速10ml
/分で流入させ、溶出液を回収した。この回収液を濃縮
し、オボトランスフェリン(純度95.7%;HPLCによる)44mg
を得た。A column (capacity 500 ml, inner diameter 30 mm, length 180 mm) packed with Blue Sepharose CL-6B was adjusted to 10 mM.
After equilibration with phosphate buffer (pH 7.0), the above upper layer solution (pH 7.
0) Pour 100 ml into the column. After leaving for about 1 hour, 10 mM phosphate buffer containing 2M NaCl was added until the protein component was no longer detected in the eluate (controlled by UV monitor) at a flow rate of 10 ml.
The eluate was collected by flowing in at a flow rate of / min. The collected solution was concentrated to give 44 mg of ovotransferrin (purity 95.7%; by HPLC).
Got
【００１９】その後、0.5M NaClを含むトリス−塩酸緩
衝液(PH8.5)および0.5M NaClを含む10mM酢酸ナトリウム
緩衝液(PH4.5)を、カラム中の残留たん白質が溶出しな
くなる迄(紫外線モニタで管理)流入、溶出させて、洗浄
した。Thereafter, Tris-hydrochloric acid buffer solution (PH8.5) containing 0.5M NaCl and 10 mM sodium acetate buffer solution (PH4.5) containing 0.5M NaCl were added until the residual protein in the column was no longer eluted ( (Controlled by UV monitor) Inflow, elution, and cleaning.
Claims (1)
白たん白質溶液について、ブルー・セファローズCL-6B 【化１】 (ここで、R1，R2は水素原子または−SO2ONa基である)を
担体とするアフィニティークロマトグラフィーを適用す
ることを特徴とするオボトランスフェリンの分離方法。1. A blue-sepharose CL-6B for an egg white protein solution from which lysozyme and avidin have been removed. A method for separating ovotransferrin, which comprises applying affinity chromatography using (wherein R 1 and R 2 are hydrogen atoms or —SO 2 ONa groups) as a carrier.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP4171823A JPH05339297A (en) | 1992-06-05 | 1992-06-05 | Separation of ovotransferrin |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP4171823A JPH05339297A (en) | 1992-06-05 | 1992-06-05 | Separation of ovotransferrin |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH05339297A true JPH05339297A (en) | 1993-12-21 |
Family
ID=15930406
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP4171823A Pending JPH05339297A (en) | 1992-06-05 | 1992-06-05 | Separation of ovotransferrin |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH05339297A (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2013527473A (en) * | 2010-06-03 | 2013-06-27 | ジーイー・ヘルスケア・バイオサイエンス・アクチボラグ | Chromatographic column module parallel assembly |
| US9950277B2 (en) | 2010-06-03 | 2018-04-24 | Ge Healthcare Bioprocess R&D Ab | Parallel assembly of chromatography column modules |
| CN110204609A (en) * | 2019-05-31 | 2019-09-06 | 华中农业大学 | The method and its albumen iron product of ovotransferrins are extracted in a kind of industrialization |
-
1992
- 1992-06-05 JP JP4171823A patent/JPH05339297A/en active Pending
Cited By (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2013527473A (en) * | 2010-06-03 | 2013-06-27 | ジーイー・ヘルスケア・バイオサイエンス・アクチボラグ | Chromatographic column module parallel assembly |
| US9943781B2 (en) | 2010-06-03 | 2018-04-17 | Ge Healthcare Bioprocess R&D Ab | Parallel assembly of chromatography column modules |
| US9950277B2 (en) | 2010-06-03 | 2018-04-24 | Ge Healthcare Bioprocess R&D Ab | Parallel assembly of chromatography column modules |
| US10092856B2 (en) | 2010-06-03 | 2018-10-09 | Ge Healthcare Bioprocess R&D Ab | Parallel assembly of chromatography column modules |
| US10933350B2 (en) | 2010-06-03 | 2021-03-02 | Cytiva Bioprocess R&D Ab | Parallel assembly of chromatography column modules |
| US10940403B2 (en) | 2010-06-03 | 2021-03-09 | Cytiva Bioprocess R&D Ab | Parallel assembly of chromatography column modules |
| US11491417B2 (en) | 2010-06-03 | 2022-11-08 | Cytiva Bioprocess R&D Ab | Parallel assembly of chromatography column modules |
| US11529570B2 (en) | 2010-06-03 | 2022-12-20 | Cytiva Bioprocess R&D Ab | Parallel assembly of chromatography column modules |
| US11911711B2 (en) | 2010-06-03 | 2024-02-27 | Cytiva Bioprocess R&D Ab | Parallel assembly of chromatography column modules |
| US12370467B2 (en) | 2010-06-03 | 2025-07-29 | Cytiva Bioprocess R&D Ab | Parallel assembly of chromatography column modules |
| CN110204609A (en) * | 2019-05-31 | 2019-09-06 | 华中农业大学 | The method and its albumen iron product of ovotransferrins are extracted in a kind of industrialization |
| CN110204609B (en) * | 2019-05-31 | 2022-01-07 | 华中农业大学 | Industrial extraction method of ovotransferrin and protein iron product thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| JP4218980B2 (en) | Immunoglobulin G concentrate for treatment and method for producing the concentrate | |
| US5169936A (en) | Protein purification on immobilized metal affinity resins effected by elution using a weak ligand | |
| JP2562192B2 (en) | Purification method of serum albumin | |
| JPS6037996A (en) | Recovery of cell wall protein | |
| JP2005500265A (en) | Method for preparing human immunoglobulin concentrates for therapeutic use | |
| JP2003530326A (en) | Method for producing IgG | |
| JPS6361318B2 (en) | ||
| JP2003096092A (en) | Purification method of pharmacologically active protein through cation exchange chromatography | |
| US4181713A (en) | Isolation of HBs Ag | |
| US4541952A (en) | Purification method of human interferon | |
| US5252217A (en) | Blood coagulation factor XI concentrate having high specific activity, suitable for therapeutic use, and process for preparing same | |
| JPH0580455B2 (en) | ||
| US5665868A (en) | Chromatographic agent and its use for the separation or proteins, polypeptides of metals | |
| KR100436857B1 (en) | Methods for Producing Factor Nine from Biological Sources | |
| US4990597A (en) | Process for the purification of placental tissue protein PP4 | |
| US4065445A (en) | Pregnancy-specific β1 -glycoprotein and process for isolating it | |
| JP2573467B2 (en) | Pharmaceutical compositions suitable for therapeutic use containing factor IX protein | |
| JPH05339297A (en) | Separation of ovotransferrin | |
| JPWO2021262963A5 (en) | ||
| JPH08225599A (en) | Process for the preparation of C1-esterase inhibitor concentrate (C1-INH), and the resulting concentrate for therapeutic use | |
| WO1995004077A1 (en) | Method of purifying plasminogen | |
| US4694074A (en) | Process for the purification of HBsAg | |
| JPS62259596A (en) | Purification of hybrid protein | |
| JPH05339296A (en) | Separation of ovomucoid | |
| JP2000103800A (en) | Method for purifying immunoglobulin A |