JP2855596B2 - Component separation method - Google Patents
Component separation methodInfo
- Publication number
- JP2855596B2 JP2855596B2 JP19155594A JP19155594A JP2855596B2 JP 2855596 B2 JP2855596 B2 JP 2855596B2 JP 19155594 A JP19155594 A JP 19155594A JP 19155594 A JP19155594 A JP 19155594A JP 2855596 B2 JP2855596 B2 JP 2855596B2
- Authority
- JP
- Japan
- Prior art keywords
- group
- target component
- component group
- charged
- substrate
- Prior art date
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- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Separation Using Semi-Permeable Membranes (AREA)
- Peptides Or Proteins (AREA)
- Apparatus Associated With Microorganisms And Enzymes (AREA)
Description
【０００１】[0001]
【産業上の利用分野】本発明は、水相／有機相の分配係
数と荷電膜による選択透過性を利用した成分の分離方法
に関する。BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for separating components using a partition coefficient between an aqueous phase and an organic phase and selective permeation by a charged membrane.
【０００２】[0002]
【従来の技術】多成分水溶液から目的物質を分離する方
法の１つとして、液−液抽出があり、この液−液抽出を
利用したものとして、具体的にはタンパク質分解酵素の
逆反応によるペプチドの合成がある。2. Description of the Related Art One of the methods for separating a target substance from a multi-component aqueous solution is liquid-liquid extraction, and a method utilizing this liquid-liquid extraction is specifically a peptide obtained by reverse reaction of a protease. There is a synthesis of
【０００３】上記タンパク質分解酵素による合成反応平
衡は低いため、生成物を効率よく反応系外に取り出すこ
とで見掛上の平衡を合成側にシフトさせる方法として、
従来から沈殿法、微水系反応法、二相系反応法が知られ
ている。[0003] Since the synthesis reaction equilibrium by the above protease is low, there is a method for shifting the apparent equilibrium toward the synthesis side by efficiently taking the product out of the reaction system.
Conventionally, a precipitation method, a micro-aqueous reaction method, and a two-phase reaction method are known.
【０００４】沈殿法は、水溶性基質から水不溶性生成物
を合成する方法であり、微水系反応法は、系内の水量を
制限することにより、逆反応である加水分解反応を抑制
する方法であり、二相系反応法は水相で酵素反応を行
い、有機相に生成物を抽出する方法である。沈殿法とし
ては、化学工学会第２０回秋期大会研究発表講演要旨集
（１９８７年、３４３頁）が知られ、微水系反応法とし
ては特開平２−３９８９５号公報に開示されるものが知
られ、二相系反応法としては化学工学論文集１７巻３号
（１９９１年、５８６−５８８頁）に開示されるものが
知られている。[0004] The precipitation method is a method of synthesizing a water-insoluble product from a water-soluble substrate, and the micro-aqueous reaction method is a method of restricting the amount of water in the system to suppress a reverse hydrolysis reaction. There is a two-phase reaction method in which an enzymatic reaction is performed in an aqueous phase and a product is extracted in an organic phase. As the precipitation method, a summary of research presentations of the 20th Fall Meeting of the Chemical Engineering Society (1987, p. 343) is known, and as a micro-water-based reaction method, a method disclosed in JP-A-2-39895 is known. As a two-phase reaction method, a method disclosed in Chemical Engineering Transactions Vol. 17 No. 3 (1991, pp. 586-588) is known.
【０００５】[0005]
【発明が解決しようとする課題】上述の沈殿法は、生成
物が沈殿するため高純度の生成物を得ることができるが
連続化は困難である。また、沈殿物と酵素とが複合体を
形成することにより、酵素が系外へ漏出するため酵素の
再利用も困難である。In the above-mentioned precipitation method, a high-purity product can be obtained because the product precipitates, but it is difficult to make the product continuous. Further, since the enzyme forms a complex with the precipitate and leaks out of the system, it is difficult to reuse the enzyme.
【０００６】また微水系反応法については、一般に生成
物の反応平衡度が低いため純度を高めることが困難であ
り、更に有機溶媒中に存在する酵素の安定性にも問題が
ある。上記特開平２−３９８９５号公報においては、カ
ルボキシル基保護の基質Ｂの濃度をアミノ基保護の基質
Ａの濃度よりも高くすることによって基質Ａに関しては
高い転換率を得ているが、反応液中の過剰の基質Ｂにつ
いては再利用ができていない。[0006] In the case of the microaqueous reaction method, it is generally difficult to increase the purity because the reaction equilibrium of the product is low, and there is also a problem in the stability of the enzyme present in the organic solvent. In the above-mentioned JP-A-2-39895, a high conversion rate is obtained for the substrate A by making the concentration of the substrate B for protecting the carboxyl group higher than the concentration of the substrate A for protecting the amino group. The excess substrate B cannot be recycled.
【０００７】更に二相系反応法は、酵素を触媒とした基
質Ａと基質Ｂとの反応を水相で行い、生成したジペプチ
ドを有機相へ抽出させる方法であるため、反応生成物が
除去される水相においては、平衡を保つために更に反応
が進行する理想的なものである。しかしこの方法には、
ジペプチドと共に基質Ｂの一部も有機相へ抽出されると
いう問題があり、このため有機相から高純度のジペプチ
ドを得ることは困難である。また基質Ｂは流失による現
象を考慮して、基質Ａに比較して高濃度で反応系に供給
されるため、前記有機相への流出による損失量は膨大と
なる。更に、浮遊する酵素についてもエマルションの形
で有機相に巻き込まれるために、この有機相の取り出し
に伴って酵素の一部が漏出してしまうという難点もあっ
た。Further, the two-phase reaction method is a method in which a substrate A and a substrate B are catalyzed by an enzyme in an aqueous phase, and the produced dipeptide is extracted into an organic phase. In an aqueous phase, it is ideal that the reaction proceeds further to maintain equilibrium. However, this method
There is a problem that a part of the substrate B is extracted into the organic phase together with the dipeptide, so that it is difficult to obtain a high-purity dipeptide from the organic phase. Further, since the substrate B is supplied to the reaction system at a higher concentration than the substrate A in consideration of the phenomenon caused by the loss, the amount of loss due to the outflow into the organic phase is enormous. Furthermore, since the floating enzyme is involved in the organic phase in the form of an emulsion, there is also a problem that a part of the enzyme leaks with the removal of the organic phase.
【０００８】そこで、本発明者等は先に上記の不利を解
消するジペプチドの製造法を提案している。この方法
は、反応槽において基質Ａ、基質Ｂ、酵素及び生産物で
あるジペプチドが含まれる水相と有機相とを接触せし
め、この有機相からジペプチドを直接取り出さず、ジペ
プチドの他に基質Ｂを含む有機相の溶液を抽出槽へ移
し、ジペプチドを再度水性の抽出相へ移動し基質Ｂと分
離してから取り出すようにしたものである。この方法に
よれば、効率よく高純度のジペプチドを生産することが
できるのであるが、ジペプチドと基質Ｂとが同じ抽出傾
向を示すため２段階の抽出が必要となり、その分だけ装
置も大掛りとなり、抽出に要する時間も長くかかる。Therefore, the present inventors have previously proposed a method for producing a dipeptide which solves the above disadvantages. In this method, an aqueous phase containing a substrate A, a substrate B, an enzyme and a dipeptide which is a product is brought into contact with an organic phase in a reaction vessel, and the dipeptide is not directly taken out from the organic phase. The solution containing the organic phase is transferred to an extraction tank, and the dipeptide is transferred again to the aqueous extraction phase, separated from the substrate B, and then taken out. According to this method, a high-purity dipeptide can be efficiently produced. However, since the dipeptide and the substrate B show the same extraction tendency, two-stage extraction is required, and the apparatus becomes large accordingly. In addition, the time required for extraction is long.
【０００９】[0009]
【課題を解決するための手段】上記課題を解決するため
本発明は、水相と有機相とを接触せしめ、水相中の目的
成分群を有機相中に抽出する成分分離方法において、前
記水相と有機相とは荷電膜を介して接触し、また前記水
相中には目的成分群の他に、水相／有機相の分配係数が
前記目的成分群よりも大きい第１の非目的成分群と、第
２の非目的成分群が含まれ、これら目的成分群、第１の
非目的成分群、第２の非目的成分群及び荷電膜の荷電は
以下の組み合わせのうちのいずれかとした。 In order to solve the above-mentioned problems, the present invention relates to a component separation method for bringing an aqueous phase into contact with an organic phase and extracting a target component group in the aqueous phase into the organic phase. Phase and the organic phase are in contact with each other via a charged membrane, and in the aqueous phase, in addition to the target component group, a first non-target component having a larger aqueous / organic phase partition coefficient than the target component group The group and the second non-target component group were included, and the target component group, the first non-target component group, the second non-target component group, and the charge of the charged membrane were any of the following combinations.
【００１０】[0010]
【００１１】以下の（表１）に本発明方法が適用される
各成分の荷電と荷電膜の荷電との組み合わせを示す。
尚、（表１）において、第１の非目的成分群（Ａ）また
は目的成分群（Ｐ）が無荷電の場合には、ｐＨ以外の要
素によって抽出するが、具体的な要素としては溶媒との
親和性、飽和溶解度等があり、このｐＨ以外の要素によ
って第１の非目的成分群（Ａ）については水相に殆んど
を分配し、目的成分群（Ｐ）については有機相に多く分
配する。The following Table 1 shows combinations of the charge of each component to which the method of the present invention is applied and the charge of the charged film.
In Table 1, when the first non-target component group (A) or the target component group (P) is uncharged, extraction is performed by using an element other than pH. Of the first non-target component group (A), and most of the target component group (P) is distributed in the organic phase due to factors other than the pH. Distribute.
【００１２】[0012]
【表１】 [Table 1]
【００１３】また上記方法としては、基質Ａ及び基質Ｂ
とを酵素触媒の存在下で反応させてジペプチドを生成す
る場合に適用することができ、この場合、基質Ａ及び基
質Ｂは以下の組み合わせのいずれかとする。 組み合わせ 基質Ａ：Ｌｙｓ（リジン）、Ｈｉｓ（ヒスチジン）、Ａ
ｒｇ（アルギニン）、Ｏｒｎ（オルニチン）、Ａｓｐ
（アスパラギン酸）及びＧｌｕ（グルタミン酸）の群
（Ｉ群）から選ばれるアミノ酸であって、そのアミノ基
が保護基によって保護されたアミノ酸 基質Ｂ：Ａｌａ（アラニン）、Ｖａｌ（バリン）、Ｌｅ
ｕ（ロイシン）、Ｉｌｅ（イソロイシン）、Ｍｅｔ（メ
チオニン）、Ｔｒｐ（トリプトファン）、Ｐｈｅ（フェ
ニルアラニン）、Ｐｒｏ（プロリン）、Ｇｌｙ（グリシ
ン）、Ｓｅｒ（セリン）、Ｔｈｒ（トレオニン）、Ｃｙ
ｓ（システイン）、Ｔｙｒ（チロシン）、Ａｓｎ（アス
パラギン）、Ｇｌｎ（グルタミン）、Ｌｙｓ、Ｈｉｓ、
Ａｒｇ、Ｏｒｎ、Ａｓｐ及びＧｌｕの群（ＩＩ群）から
選ばれるアミノ酸であって、そのカルボキシル基がアル
キルエステル化（但し、このアルキルは炭素数１乃至４
である。）されたアミノ酸 組み合わせ 基質Ａ：前記ＩＩ群から選ばれるアミノ酸であって、そ
のアミノ基が保護基によって保護されたアミノ酸 基質Ｂ：前記Ｉ群から選ばれるアミノ酸であって、その
カルボキシル基が前記条件のアルキルエステル化された
アミノ酸[0013] The above-mentioned method includes a substrate A and a substrate B.
Is reacted in the presence of an enzyme catalyst to produce a dipeptide. In this case, the substrate A and the substrate B are any of the following combinations. Combination Substrate A: Lys (lysine), His (histidine), A
rg (arginine), Orn (ornithine), Asp
(Aspartic acid) and Glu (glutamic acid), an amino acid selected from the group (Group I), whose amino group is protected by a protecting group. Substrate B: Ala (alanine), Val (valine), Le
u (leucine), Ile (isoleucine), Met (methionine), Trp (tryptophan), Phe (phenylalanine), Pro (proline), Gly (glycine), Ser (serine), Thr (threonine), Cy
s (cysteine), Tyr (tyrosine), Asn (asparagine), Gln (glutamine), Lys, His,
An amino acid selected from the group of Arg, Orn, Asp and Glu (group II), wherein the carboxyl group is alkylesterified (provided that the alkyl has 1 to 4 carbon atoms).
It is. ) Combined amino acids Substrate A: an amino acid selected from the group II, whose amino group is protected by a protecting group Substrate B: an amino acid selected from the group I, wherein the carboxyl group has the above-mentioned condition Alkyl esterified amino acids
【００１４】基質Ａの保護基としては、ペプチド合成反
応に慣用される例えばジフェニルメチル基、トリフェニ
ルメチル基、芳香族基及び脂肪族オキシカルボニル基が
挙げられ、これらはハロゲン、ニトロ基、低級アルキル
基、低級アルコキシ基等によって置換されていてもよ
い。その他、ファルマシアレビューＮｏ，３（日本薬学
会、１９８０年）の第３１頁、表１に記載される保護基
も好適に使用できるが、最も好ましい保護基はベンジル
オキシカルボニル基である。The protecting group for the substrate A includes, for example, diphenylmethyl group, triphenylmethyl group, aromatic group and aliphatic oxycarbonyl group, which are commonly used in peptide synthesis reactions, such as halogen, nitro group and lower alkyl group. Or a lower alkoxy group. In addition, the protecting groups described in Table 1 of Pharmacia Review No. 3 (Japan Pharmaceutical Association, page 1980, page 31) can be suitably used, but the most preferred protecting group is a benzyloxycarbonyl group.
【００１５】また１モルの基質Ａに対する基質Ｂの添加
モル数は１〜５、好ましくは１〜２とする。このモル数
が１未満では基質Ａが蓄積し、また５を超えると基質Ｂ
の系外流出が著しくなるため本発明の目的を達成できな
いことがある。更に触媒として使用する酵素としては、
ペプチド合成一般ではプロテアーゼ全般が対象となる。The number of moles of the substrate B added to 1 mole of the substrate A is 1 to 5, preferably 1 to 2. When the number of moles is less than 1, substrate A accumulates, and when the number exceeds 5, substrate B accumulates.
May be unable to achieve the object of the present invention because of significant outflow from the system. Further, as an enzyme used as a catalyst,
In general, peptide synthesis covers proteases in general.
【００１６】[0016]
【作用】水相／有機相の分配係数の相違によって、基質
Ａ（第１の非目的成分群）と目的成分群とが分離され、
荷電の相違によって基質Ｂ（第２の非目的成分群）と目
的成分群とが分離される。The substrate A (first non-target component group) and the target component group are separated by the difference in the partition coefficient between the aqueous phase and the organic phase.
The substrate B (second non-target component group) and the target component group are separated by the difference in charge.
【００１７】[0017]
【実施例】以下に本発明の実施例を添付図面に基づいて
説明する。ここで、図１は本発明に係る成分分離方法を
実施する装置の概略図、図２は水相／有機相の分配係数
の差を利用した抽出を説明した図、図３は荷電膜を利用
した抽出を説明した図である。Embodiments of the present invention will be described below with reference to the accompanying drawings. Here, FIG. 1 is a schematic diagram of an apparatus for carrying out the component separation method according to the present invention, FIG. 2 is a diagram illustrating extraction using a difference in distribution coefficient between an aqueous phase / organic phase, and FIG. 3 is a diagram using a charged membrane. FIG.
【００１８】本発明に係る成分分離方法を実施する装置
は装置本体１内を荷電膜２によって水相３と有機相４に
分け、水相３は反応性を良好にするために循環されてお
り、この循環系には基質Ａ及び基質Ｂを供給する手段が
設けられ、また触媒となる酵素Ｃは水相３中に予め分散
させるか又は基質Ａ、Ｂと共に供給される。一方、有機
相４からは生成物Ｐを含んだ溶液を取り出し、必要に応
じて酸分解等によって保護基をはずし、再結晶等で精製
して高純度の結晶として析出させる。In the apparatus for carrying out the component separation method according to the present invention, the interior of the apparatus main body 1 is divided into an aqueous phase 3 and an organic phase 4 by a charged membrane 2, and the aqueous phase 3 is circulated to improve the reactivity. The circulating system is provided with means for supplying a substrate A and a substrate B, and the enzyme C serving as a catalyst is dispersed in the aqueous phase 3 in advance or supplied together with the substrates A and B. On the other hand, the solution containing the product P is taken out of the organic phase 4, the protective group is removed by acid decomposition or the like, if necessary, and purified by recrystallization or the like to precipitate as high-purity crystals.
【００１９】前記荷電膜２としては例えばイオン交換膜
を用い、また荷電膜２の荷電は生成物Ｐと異付号で基質
Ｂとは同付号のものを選定する。即ち、荷電膜２の荷電
は（＋）、基質Ａの荷電は（−）、基質Ｂの荷電は
（＋）、生成物Ｐの荷電は（−）となる。As the charged membrane 2, for example, an ion exchange membrane is used, and the charge of the charged membrane 2 is selected to be different from the product P and the same as the substrate B. That is, the charge of the charged film 2 is (+), the charge of the substrate A is (-), the charge of the substrate B is (+), and the charge of the product P is (-).
【００２０】ところで、上記の装置内では同時に２つの
分離がなされている。即ち、第１の分離は図２に示すよ
うに水相／有機相の分配係数の相違を利用した分離であ
り、基質Ａは有機相に殆ど溶解せず、逆に生成物Ｐと基
質Ｂは有機相により多く溶解するため、基質Ａと生成物
Ｐ及び基質Ｂとが分離される。また第２の分離は図３に
示すように電気的な性質を利用した分離であり、基質Ａ
と生成物Ｐは荷電膜２の荷電と異付号であるため荷電膜
２を透過するが、基質Ｂは荷電膜２の荷電と同付号であ
るため反発して透過できない。尚、酵素Ｃについても荷
電膜２を透過することができない。Incidentally, two separations are made simultaneously in the above-mentioned apparatus. That is, the first separation is a separation utilizing the difference in the partition coefficient between the aqueous phase and the organic phase as shown in FIG. 2, and the substrate A hardly dissolves in the organic phase, while the product P and the substrate B Substrate A is separated from product P and substrate B because it is more soluble in the organic phase. The second separation is a separation utilizing electric properties as shown in FIG.
And the product P have a different sign from the charge of the charged membrane 2 and pass through the charged membrane 2, but the substrate B has the same sign as the charge on the charged membrane 2 and repels and cannot pass therethrough. Note that the enzyme C cannot pass through the charged membrane 2.
【００２１】以下に具体的な実施例を挙げる。基質Ａと
してベンジルオキシカルボニルアスパラギン酸が１０ミ
リモル、基質Ｂとしてフェニルアラニンメチルエステル
が１０ミリモル、及びこれらのペプチド（生成物Ｐ）で
あるベンジルオキシカルボニルアスパラギルフェニルア
ラニンメチルエステルが５ミリモル含まれる水溶液を作
成し、この水溶液をｐＨ６に調整した。Specific examples will be described below. An aqueous solution containing 10 mmol of benzyloxycarbonyl aspartic acid as the substrate A, 10 mmol of phenylalanine methyl ester as the substrate B, and 5 mmol of benzyloxycarbonyl asparagyl phenylalanine methyl ester as the peptide (product P) was prepared. This aqueous solution was adjusted to pH 6.
【００２２】更に触媒として使用する酵素としては、メ
タロプロティナーゼの一種であるサーモライシンが好適
に使用できるが、その他の酵素の例を挙げれば、Staphy
lococcalプロティナーゼ（EC 3.4.21）、パパイン（EC
3.4.22.2）がある。Further, as an enzyme used as a catalyst, thermolysin, which is a kind of metalloproteinase, can be suitably used.
lococcal proteinase (EC 3.4.21), papain (EC
3.4.22.2).
【００２３】そして、上記の水溶液を正荷電膜を介し
て、tert-アミルアルコール（有機相）に接触させた。
尚、分配係数（水相濃度／有機相濃度）は、基質Ａ＝４
５５８、基質Ｂ＝０．４７０７、生成物Ｐ＝０．２６５
８である。以下の（表２）は正荷電膜を介在させない場
合と介在させた場合の１６８時間経過後の水相と有機相
内における各物質の濃度である。この（表２）から明ら
かなように本発明方法による場合には、有機相に生成物
Ｐのみが検出される。Then, the above aqueous solution was brought into contact with tert-amyl alcohol (organic phase) via a positively charged membrane.
Incidentally, the partition coefficient (aqueous phase concentration / organic phase concentration) was calculated as follows: substrate A = 4
558, substrate B = 0.4707, product P = 0.265
8 The following (Table 2) shows the concentration of each substance in the aqueous phase and the organic phase after 168 hours when the positively charged film is not interposed and when it is interposed. As is apparent from Table 2, only the product P is detected in the organic phase in the case of the method of the present invention.
【００２４】[0024]
【表２】 [Table 2]
【００２５】尚、本発明に係る方法は連続式、回分式の
どちらにも適用可能であり、水相のｐＨは生成するジペ
プチドの有する側鎖解離基の種類等によって適宜調整す
る。また、実施例としてはジペプチドの製造について本
発明方法を適用した例を示したが、本発明に係る成分の
分離方法は、水相中に３成分以上の成分が含まれている
場合に広く適用できる。The method according to the present invention can be applied to both a continuous system and a batch system, and the pH of the aqueous phase is appropriately adjusted depending on the type of the side chain dissociating group of the dipeptide to be formed. Further, as an example, an example in which the method of the present invention is applied to the production of a dipeptide is shown. However, the method of separating components according to the present invention is widely applied when three or more components are contained in an aqueous phase. it can.
【００２６】[0026]
【発明の効果】以上に説明したように本発明によれば、
分配係数の差を利用した分離と荷電を利用した分離とを
巧みに組み合わせることで、３以上の成分を含む水相か
ら１回の操作で目的成分群を抽出することができる。According to the present invention as described above,
By skillfully combining separation using the difference in partition coefficient and separation using charge, the target component group can be extracted from the aqueous phase containing three or more components in one operation.
【００２７】例えば、基質Ａと基質Ｂを酵素触媒を用い
て反応させてジペプチドを生産する場合に装置も１つ、
抽出も１回で済み、しかも基質の使用も無駄なく高純度
のジペプチドを生産することができる。For example, when a substrate A and a substrate B are reacted using an enzyme catalyst to produce a dipeptide, one apparatus is used.
Only one extraction is required, and a high-purity dipeptide can be produced without wasting substrate.
【図１】本発明に係る成分分離方法を実施する装置の概
略図FIG. 1 is a schematic diagram of an apparatus for performing a component separation method according to the present invention.
【図２】水相／有機相の分配係数の差を利用した抽出を
説明した図FIG. 2 is a diagram illustrating extraction using a difference in partition coefficient between an aqueous phase and an organic phase.
【図３】荷電膜を利用した抽出を説明した図FIG. 3 is a diagram illustrating extraction using a charged membrane.
１…装置本体、２…荷電膜、３…水相、４…有機相、
Ａ，Ｂ…基質、Ｃ…酵素、Ｐ…生成物。DESCRIPTION OF SYMBOLS 1 ... Device main body, 2 ... Charge membrane, 3 ... Water phase, 4 ... Organic phase,
A, B: substrate, C: enzyme, P: product.
───────────────────────────────────────────────────── フロントページの続き (51)Int.Cl.6 識別記号 ＦＩ // Ｃ１２Ｍ 1/40 Ｃ１２Ｍ 1/40 Ａ (72)発明者 礒野 康幸 茨城県稲敷郡茎崎町若葉１−８ コンド レアＢ−Ａ (72)発明者 星野 信夫 東京都練馬区西大泉５−８−４ (72)発明者 星野 明 埼玉県越谷市越谷本町３−４ (72)発明者 福島 健司 東京都北区志茂４−１−９−201 (56)参考文献 特開 昭61−205490（ＪＰ，Ａ) (58)調査した分野(Int.Cl.6，ＤＢ名) C12P 21/00 - 21/02 B01D 61/42 C07K 1/02 C07K 1/14 C07K 5/06 C12M 1/40 ＢＩＯＳＩＳ（ＤＩＡＬＯＧ) ＷＰＩ（ＤＩＡＬＯＧ)──────────────────────────────────────────────────の Continued on the front page (51) Int.Cl. 6 Identification symbol FI // C12M 1/40 C12M 1/40 A (72) Inventor Yasuyuki Isono 1-8 Wakaba Kusazaki-cho, Inashiki-gun, Ibaraki Pref. A (72) Inventor Nobuo Hoshino 5-8-4 Nishi-Oizumi, Nerima-ku, Tokyo (72) Inventor Akira Hoshino 3-4, Koshigaya-Honmachi, Koshigaya-shi, Saitama (72) Inventor Kenji Fukushima 4-1 Shimo, Kita-ku, Tokyo -9-201 (56) References JP-A-61-205490 (JP, A) (58) Fields investigated (Int. Cl. 6 , DB name) C12P 21/00-21/02 B01D 61/42 C07K 1 / 02 C07K 1/14 C07K 5/06 C12M 1/40 BIOSIS (DIALOG) WPI (DIALOG)
Claims (2)
目的成分群を有機相中に抽出する成分分離方法におい
て、前記水相と有機相とは荷電膜を介して接触し、また
前記水相中には目的成分群の他に、水相／有機相の分配
係数が前記目的成分群よりも大きい第１の非目的成分群
と、第２の非目的成分群が含まれ、これら目的成分群、
第１の非目的成分群、第２の非目的成分群及び荷電膜の
荷電は以下の組み合わせのうちのいずれかであることを
特徴とする成分分離方法。 組み合わせ：第１の非目的成分群は正荷電 第２の非目的成分群は負荷電 目的成分群は正荷電 荷電膜は負荷電 組み合わせ：第１の非目的成分群は負荷電 第２の非目的成分群は正荷電 目的成分群は負荷電 荷電膜は正荷電 組み合わせ：第１の非目的成分群は正荷電 第２の非目的成分群は負荷電 目的成分群は無荷電 荷電膜は負荷電 組み合わせ：第１の非目的成分群は負荷電 第２の非目的成分群は正荷電 目的成分群は無荷電 荷電膜は正荷電 組み合わせ：第１の非目的成分群は無荷電 第２の非目的成分群は負荷電 目的成分群は正荷電 荷電膜は負荷電 組み合わせ：第１の非目的成分群は無荷電 第２の非目的成分群は正荷電 目的成分群は負荷電 荷電膜は正荷電 組み合わせ：第１の非目的成分群は無荷電 第２の非目的成分群は負荷電 目的成分群は無荷電 荷電膜は負荷電 組み合わせ：第１の非目的成分群は無荷電 第２の非目的成分群は正荷電 目的成分群は無荷電 荷電膜は正荷電1. A component separation method for bringing an aqueous phase and an organic phase into contact with each other and extracting a target component group in the aqueous phase into the organic phase, wherein the aqueous phase and the organic phase are contacted via a charged membrane, Further, in the aqueous phase, in addition to the target component group, a first non-target component group and a second non-target component group in which the partition coefficient of the aqueous phase / organic phase is larger than the target component group are included, These target components,
A component separation method, wherein the first non-target component group, the second non-target component group, and the charge of the charged film are any one of the following combinations. Combination: The first non-target component group is positively charged. The second non-target component group is negatively charged. The target component group is positively charged. The charged film is negatively charged. Combination: The first non-target component group is negatively charged. The component group is positively charged. The target component group is negatively charged. The first non-target component group is positively charged. The second non-target component group is negatively charged. The target component group is uncharged. : The first non-target component group is negatively charged. The second non-target component group is positively charged. The target component group is uncharged. The charged film is positively charged. Combination: The first non-target component group is uncharged. The group is negatively charged. The target component group is positively charged. The charged membrane is negatively charged. Combination: The first non-targeted component group is uncharged. The second non-targeted component group is positively charged. The target component group is negatively charged. The first non-target component group is uncharged The second non-target component group is negative charge The uncharged charged membrane is negatively charged combination: a first non-target component groups uncharged second non-target group of components positively charged target component groups uncharged charged membrane is positively charged
て、前記第１の非目的成分群と第２の非目的成分群は水
相中で酵素触媒により反応して目的成分群であるジペプ
チドを生産する基質Ａ及び基質Ｂであり、基質Ａ及び基
質Ｂは以下の組み合わせのいずれかであることを特徴と
する成分分離方法。 組み合わせ 基質Ａ：Ｌｙｓ、Ｈｉｓ、Ａｒｇ、Ｏｒｎ、Ａｓｐ及び
Ｇｌｕの群（Ｉ群）から選ばれるアミノ酸であって、そ
のアミノ基が保護基によって保護されたアミノ酸 基質Ｂ：Ａｌａ、Ｖａｌ、Ｌｅｕ、Ｉｌｅ、Ｍｅｔ、Ｔ
ｒｐ、Ｐｈｅ、Ｐｒｏ、Ｇｌｙ、Ｓｅｒ、Ｔｈｒ、Ｃｙ
ｓ、Ｔｙｒ、Ａｓｎ、Ｇｌｎ、Ｌｙｓ、Ｈｉｓ、Ａｒ
ｇ、Ｏｒｎ、Ａｓｐ及びＧｌｕの群（ＩＩ群）から選ば
れるアミノ酸であって、そのカルボキシル基がアルキル
エステル化（但し、このアルキルは炭素数１乃至４であ
る。）されたアミノ酸 組み合わせ 基質Ａ：前記ＩＩ群から選ばれるアミノ酸であって、そ
のアミノ基が保護基によって保護されたアミノ酸 基質Ｂ：前記Ｉ群から選ばれるアミノ酸であって、その
カルボキシル基が前記条件のアルキルエステル化された
アミノ酸2. The component separation method according to claim 1, wherein the first non-target component group and the second non-target component group react in an aqueous phase with an enzyme catalyst to form a dipeptide as the target component group. A method for separating components, which is a substrate A and a substrate B to be produced, wherein the substrate A and the substrate B are any of the following combinations. Combination Substrate A: an amino acid selected from the group (Group I) of Lys, His, Arg, Orn, Asp and Glu, whose amino group is protected by a protecting group. Substrate B: Ala, Val, Leu, Ile , Met, T
rp, Phe, Pro, Gly, Ser, Thr, Cy
s, Tyr, Asn, Gln, Lys, His, Ar
g, an amino acid selected from the group consisting of Orn, Asp and Glu (group II), wherein the carboxyl group is alkyl-esterified (however, this alkyl has 1 to 4 carbon atoms). An amino acid selected from the group II, wherein the amino group is protected by a protecting group. Substrate B: an amino acid selected from the group I, wherein the carboxyl group is alkylesterified under the above conditions.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP19155594A JP2855596B2 (en) | 1994-08-15 | 1994-08-15 | Component separation method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP19155594A JP2855596B2 (en) | 1994-08-15 | 1994-08-15 | Component separation method |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPH0851994A JPH0851994A (en) | 1996-02-27 |
| JP2855596B2 true JP2855596B2 (en) | 1999-02-10 |
Family
ID=16276627
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| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP19155594A Expired - Fee Related JP2855596B2 (en) | 1994-08-15 | 1994-08-15 | Component separation method |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JP2855596B2 (en) |
-
1994
- 1994-08-15 JP JP19155594A patent/JP2855596B2/en not_active Expired - Fee Related
Also Published As
| Publication number | Publication date |
|---|---|
| JPH0851994A (en) | 1996-02-27 |
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