JPS61270658A - Separating and refining device - Google Patents
Separating and refining deviceInfo
- Publication number
- JPS61270658A JPS61270658A JP60111653A JP11165385A JPS61270658A JP S61270658 A JPS61270658 A JP S61270658A JP 60111653 A JP60111653 A JP 60111653A JP 11165385 A JP11165385 A JP 11165385A JP S61270658 A JPS61270658 A JP S61270658A
- Authority
- JP
- Japan
- Prior art keywords
- eluate
- column
- valve
- eluent
- composition
- 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
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/26—Conditioning of the fluid carrier; Flow patterns
- G01N30/38—Flow patterns
- G01N30/44—Flow patterns using recycling of the fraction to be distributed
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Sustainable Development (AREA)
- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Treatment Of Liquids With Adsorbents In General (AREA)
Abstract
Description
【発明の詳細な説明】
定業上の利用−分野
本発明は、工業的規模での混合物の分離精製装置に係り
、更に詳しくは、溶離液再生機能を有する液体クロマト
グラフィー装置に関する。DETAILED DESCRIPTION OF THE INVENTION Field of the Invention The present invention relates to an apparatus for separating and purifying mixtures on an industrial scale, and more particularly to a liquid chromatography apparatus having an eluent regeneration function.
−従沫ρ技−術
液体クロマトグラフィーは、元来、実験室規模での分析
装置として開発された技術であるが、遺伝子組喚えや細
胞融合等のバイオチクノロシイを利用して得られる物質
等を高純度で、しかも短時間に工業的規模で分離精製す
るため利用されるようになってきた。この方法は、耐圧
カラム中にシリカゲル、ODS、多孔性ポリマーゲルを
充填し、カラム上部に混合物を添加した後、/8離液で
カラムから押し出すごとにより混合物の中から目的成分
を分+1iu−t’る方法である。Liquid chromatography is a technology originally developed as a laboratory-scale analysis device, but it can also be obtained by using biotechnology such as genetic recombination and cell fusion. It has come to be used to separate and purify substances with high purity and in a short time on an industrial scale. This method involves filling a pressure-resistant column with silica gel, ODS, and porous polymer gel, adding the mixture to the top of the column, and then extruding it from the column with /8 syneresis to remove the target component from the mixture by +1iu-t. This is the way to do it.
工業的分離精製技術として液体クロマ1〜グラフイーを
適用する場合には、カラムを大型化するごとにより処理
「dを増加さ・するので、それに伴って78月1メ夜量
も増加する。4犀1/&として(31、充虜するゲル及
び目的物質により異なるが、アセトニトリル液がしばし
ば使用される。When applying liquid chroma 1 to graphie as an industrial separation and purification technology, each time the column size increases, the processing amount increases, and the amount of water per month also increases accordingly. As 1/& (31), depending on the gel to be filled and the target substance, acetonitrile solutions are often used.
発明が解決−レ、L−シ)ξず−ろ問題点l容離剤とし
て、特に自tel! /’J媒を使用する場合に(31
、経l斉的観点でY1機溶媒を循環使用する必要がη三
じる。そのため、朶溜により自機溶媒を回収、精製し、
1fi利川する方法が行われているが、二1ス1か高く
なるため、液体り[+7トグラフイーに分離精製純度あ
るいは高速化という14所がありながら、実用化範囲が
制限されることか多い。The invention solves the problems, especially as a release agent. /'When using J medium (31
Therefore, it is necessary to recycle the Y1 organic solvent from a simultaneous point of view. Therefore, we collect and purify the organic solvent by distillation,
A method of 1fi Ikawa is being used, but because it is expensive, the range of practical application is often limited, although there are 14 points in liquid lithography such as separation, purification, purity, or high speed.
−・方、/8離液を循環使用したときに、溶離液の組成
が異なってくると、[−1的物質がカラムから溶11目
゛る時間(保持時間)にずれをη”しろため、隣接する
ピークでは分舗度がイ[(下し、それに伴い、目的物質
の純度も低下する。そのため、溶離液組成のモニタリン
グか重要な課題となってくる。従って、溶離液を循環使
用する場合には、モニタリング装置を(・1加する必要
があV)、il′1i偵1な設備を要する。On the other hand, when the /8 syneresis solution is used repeatedly, if the composition of the eluent changes, the -1 substance will be eluted from the column at the 11th time (retention time) due to a shift of η". , the separation degree of adjacent peaks decreases, and the purity of the target substance decreases accordingly.Therefore, monitoring of the eluent composition becomes an important issue.Therefore, the eluent should be recycled. In some cases, a monitoring device (+1V) is required, and a large amount of equipment is required.
本発明(、31、前記の従来技術の欠点を解/l!i
(−、、冷帛1[液の11牛及び組成のモニタリングを
簡CY(にfiうごとかできる分litll稍製V2置
を1にOj−J’イ〕こ、とを目的とす4〕。The present invention (,31, solves the drawbacks of the above-mentioned prior art/l!i
(-,, cold cloth 1 [11 cows and composition of the liquid to be easily monitored in CY (to make a small V2 as much as possible to 1 Oj-J'i]), and the purpose is 4) .
問題点を解決すで)ムーン)の手段
本発明は、膜分離により/”6 i’ill /&を再
/L1’ると共に、溶離液組成のモニタリングをII的
酸成分溶出パターンで検出し、制御することに31−、
り前記の問題点をIW決したものである。Means for Solving Problems) The present invention re-re/L1'/"6i'ill/& by membrane separation, and monitors the eluent composition by detecting the acid component elution pattern, 31- to control;
The above-mentioned problems have been resolved by IW.
即し、本発明による分離精製装W U91、り1丁17
1グラフイーカラフ・の流入「1側に溶離lfk貯槽、
溶離液流人ハルゾルひラインミキサーを接続し、該ライ
ンミキサーの前にII史モジュールからの+IC 11
+ i’a岡11液の循環1/3を合流さ・口、りLJ
71グラフィーカラムの流出口側に成分検出器及び検出
パターンの記録itを設置し、成分検出器からの流出管
4;Z i;Iバルブを介してフラクション:ルクター
及び股上シフ。Therefore, the separation and purification equipment W U91, RI 1-17 according to the present invention
1 Graph E carafe inflow '1 side elution lfk storage tank,
Connect the eluent flow line mixer to the +IC 11 from II history module before the line mixer.
+ 1/3 of the circulation of i'a Oka 11 liquid is merged with the mouth, ri LJ
71 A component detector and a recording of the detection pattern are installed on the outlet side of the graphics column, and the fractions are passed through the outflow pipe 4;Zi;I valve from the component detector: Lucter and Rise Schiff.
−ルを接続し、該バルブは=1ンピュータに予め入力し
ζおいた情報に基づいて開閉されて目的物質ノヒークを
示す溶出液をフラクションコレクターへ、他の/8別液
は股モジュールへ流出させることを特徴とする。The valve is opened and closed based on the information entered into the computer in advance, allowing the eluate showing the target substance to flow out to the fraction collector, and the other /8 separate fluid flowing out to the crotch module. It is characterized by
不発明番J,熔離液として使用する液体がアルコール、
アセトニトリル等低分子の物質であり、逆に混合物は、
液体クロマトグラフィーがT業的な分離精製プロセスの
中でも最終段階で使用されることから、夾雑物が少ない
こと、また、目的物質が糖や蛋白質等、高分子物質を対
象にすることが多いこと、及び目的物質の保持時間は別
途、紫外線吸収計、示差屈折11等で既にモニタリング
しているごとに着目し、この特徴を有効に活用するため
に、膜分離で溶離液を再生した後、目的物質の保持時間
を検出し、その変化から溶11i液絹成を一定に保つよ
うに構成したものである。Non-inventive number J, the liquid used as a dissolution liquid is alcohol,
It is a low-molecular substance such as acetonitrile, and conversely, a mixture is
Since liquid chromatography is used in the final stage of industrial separation and purification processes, there are few impurities, and the target substances are often high molecular substances such as sugars and proteins. We focused on the fact that the retention time of the target substance and the retention time of the target substance were already monitored separately using an ultraviolet absorption meter, differential refraction 11, etc., and in order to make effective use of this feature, we regenerated the eluent through membrane separation and then detected the target substance. The system is designed to detect the retention time of 11i and keep the solution silk composition constant based on the change.
作用
本発明による分IiiIt精製装置においては、クロマ
トグラフィーカラムの流出口側に設けられた検出器は、
カラムから流出するフラクションの成分パターンを検出
し、検出結果を二1ンビ1−夕に出力する。この、:l
ンピュータは、予め入力しておいた情IIに基づいて、
バルブを開閉してフラクションをフラクションコレクタ
ー又は股モジュール・・・流入させ、また、他力では、
新しい〆容離液の流入バルブを開閉する。Function: In the fraction III purification device according to the present invention, the detector provided on the outlet side of the chromatography column is
The component pattern of the fraction flowing out from the column is detected, and the detection results are output to the 21st column. This, :l
Based on the information entered in advance, the computer
Open and close the valve to allow the fraction to flow into the fraction collector or crotch module, or by external force,
Open and close the inlet valve for the new synergic liquid.
膜モジュールは、カラJえから流出したフラクションか
ら共存物質を除去し、溶離液を内生ずる。The membrane module removes coexisting substances from the fraction flowing out from the membrane and generates an eluent internally.
再生された溶離液は、その組成に応じて新しい溶離液と
ラインミキサーにより混合され、常に適切な一定組成の
溶離液としてカラムに流される。The regenerated eluent is mixed with fresh eluent according to its composition by a line mixer, and is always passed through the column as an eluent with a constant composition.
実施例 次に、図面に基づいて本発明を詳述する。Example Next, the present invention will be explained in detail based on the drawings.
第1図Let、本発明の一実施態様を示す分離粘剖装置
のフローシートである。この装置で分離精をJを実施す
る場合、まず、カラノ・5の1一部に精製したい物質を
含む混合液を添加する。溶離液を/8冊1液貯槽1から
バルブ2を経て高圧ポンプ4でラインミキサ3に送り、
ここで充分混合した後、カラムに送液する。カラム5の
中で、物質により移動速度の差か生し、甲くカラムから
溶出するものから順次検出器〔iで検出され、止め入力
しておいた情+Uに基づいてバルブ7を開け、精製した
6寸1的物質のめを含むフラクションをフラクションコ
レクタ8に集める。FIG. 1 is a flow sheet of a separation viscodissection device showing one embodiment of the present invention. When carrying out separation and purification using this apparatus, first, a liquid mixture containing the substance to be purified is added to a portion of Karano-5. The eluent is sent from the liquid storage tank 1 through the valve 2 to the line mixer 3 using the high pressure pump 4.
After mixing thoroughly, the solution is sent to the column. In the column 5, there are differences in movement speed depending on the substance, and the substances eluted from the column are detected by the detector [i], and the valve 7 is opened based on the input information +U, and the purification is carried out. The fraction containing the six-dimensional substance obtained is collected in a fraction collector 8.
検出器6で検出された検出パターン4J、記録剖10に
記tiされる。The detection pattern 4J detected by the detector 6 is recorded in the record 10.
カラ人中で物質の移動i重度を決定する因子としてLe
t、カラムに充填するゲルの種類、溶離液組成及び混合
物中の各成分の物理化学的特性があり、これらは精製し
たい[1的物質の特性に応じて決定される。Le as a factor determining the severity of substance movement in humans
t, the type of gel packed in the column, the composition of the eluent, and the physicochemical characteristics of each component in the mixture, which are determined depending on the characteristics of the substance to be purified.
目的とする物質以外の物質を含む溶離液は、膜モジュー
ル9で共存物質を除去され、初期の/8離l& i11
成に調整された(多、再度、高圧ポンプ4でカラムに送
られ、循環を繰り返す。ごの際、循環を繰り返ずことに
よりン容離液絹成が変化することがY想される。そごで
、1」的とする物質がカラムから溶出している時間(保
持時間)を予め二2ンビュータに入力しておき、保持時
間にずれが生しだ場合に、バルブ2を開けて溶離液の組
成を1ントシスールする。The eluent containing substances other than the target substance has coexisting substances removed in the membrane module 9, and the initial /8 separation l&i11
The mixture is then sent to the column again using the high-pressure pump 4 and the circulation is repeated.It is thought that repeating the circulation will change the liquid composition. 1) Enter the time for the target substance to elute from the column (retention time) into the 22-meter monitor in advance, and if there is a discrepancy in the retention time, open valve 2 and elute. The composition of the liquid is 1%.
−L業的規模で使用されることが多いと嵩えられる条件
(ゲル: 01) S、溶離液:水とメタノールとの混
合液)をもとに、更に詳述すると、第2図に示ずl−J
的成分の保持時間が、第3図に示すように、メタノール
1度に依存して変化することkl知られている。従って
、この関係を目的成分に応じて予め把握しζおき、保持
時間の変化の許容範囲(他の除去したい物質の保持時間
を考慮して設定する)をコンピュータに入力しておくご
とによりバルブの自動制御を可能にする。Based on the conditions that are often used on an industrial scale (gel: 01S, eluent: a mixture of water and methanol), the following conditions are shown in Figure 2. Zl-J
It is known that the retention time of the target components changes depending on the degree of methanol, as shown in FIG. Therefore, by understanding this relationship in advance according to the target component and inputting the allowable range of change in retention time (set by taking into account the retention time of other substances to be removed) into the computer, it is possible to adjust the valve accordingly. Allow automatic control.
なお、膜モジュールとしては、l容M?夜にアルコール
、アセトニ(・リル等の有機溶媒を使用することが多い
ので、ポリアミ[系等の耐溶剤性の膜モジュールが好ま
しい。In addition, as a membrane module, 1 volume M? Since organic solvents such as alcohol and acetonyl are often used at night, solvent-resistant membrane modules such as polyamide-based membrane modules are preferred.
実施例1
第1図に示した装置において、水系G P C用ゲル(
排除限界分子量300000)を直径5cm、高さ60
cmのカラムに充填し、50mMのリン@緩衝ン夜(p
H7,5’)及び0.2MのNaC1を含む水l容7夜
を〆容部1液として用い、流速50m1/分で流して、
卵白アルブミン及び牛血清アルブミンの混合物を分&1
1精製した結果、効率よく分離精製された。Example 1 In the apparatus shown in FIG. 1, an aqueous GPC gel (
exclusion limit molecular weight 300,000), diameter 5 cm, height 60
Packed into a 50mM phosphorus buffered column (p
1 volume of water containing H7,5') and 0.2 M NaCl was used as the first liquid in the final container, and was flowed at a flow rate of 50 m1/min.
Mixture of ovalbumin and bovine serum albumin for 1 minute & 1
As a result of 1 purification, efficient separation and purification was achieved.
発明−Φ−効果
本発明によれば、省エネルギー装置である膜モジュール
で?g !ttII液を再生し、再利用することができ
る。また、カラムからの溶出物質を検出し、フラクショ
ン用バルブを開閉するためのコンピュータは、従来も設
けられCおり、新たに設備を付加することなく、これら
を効果的に組み合わセるごとにより簡111な設備で、
効率よく分1iiIIネNMを行・うごとができる。Invention - Φ - Effect According to the present invention, the membrane module which is an energy saving device? G! The ttII fluid can be regenerated and reused. In addition, computers for detecting eluted substances from columns and opening and closing valves for fractions have been installed in the past, and these can be effectively combined without adding new equipment. With facilities,
Able to perform and move NM in minutes efficiently.
第1図(J、本発明の一実施態様を示す分離精製装置の
)【コーシート、第2図は溶出成分のピークパターンの
模式図、第3I♀1ば/8離液組成が成分の溶出時間に
及ばず影響を示す概念図である。Figure 1 (J, of a separation and purification device showing one embodiment of the present invention) [Cosheet, Figure 2 is a schematic diagram of the peak pattern of eluted components, and Figure 3 I♀1ba/8 synergic composition is the elution time of the component FIG.
Claims (2)
する装置において、クロマトグラフィーカラムの流入口
側に溶離液貯槽、溶離液流入バルブ及びラインミキサー
を接続し、該ラインミキサーの前に膜モジュールからの
再生溶離液の循環路を合流させ、クロマトグラフィーカ
ラムの流出口側に成分検出器及び検出パターンの記録計
を設置し、成分検出器からの流出管にはバルブを介して
フラクションコレクター及び膜モジュールを接続し、該
バルブはコンピュータに予め入力しておいた情報に基づ
いて開閉されて目的物質のピークを示す溶出液をフラク
ションコレクターへ、他の溶離液は膜モジュールへ流出
させることを特徴とする分離精製装置。(1) In an apparatus for separating and purifying mixtures by liquid chromatography, an eluent storage tank, an eluent inlet valve, and a line mixer are connected to the inlet side of the chromatography column, and the regenerated eluent from the membrane module is connected before the line mixer. The liquid circulation paths are merged, a component detector and a detection pattern recorder are installed on the outflow side of the chromatography column, and a fraction collector and membrane module are connected to the outflow pipe from the component detector via a valve. , the separation and purification device is characterized in that the valve is opened and closed based on information input into a computer in advance to allow the eluate showing the peak of the target substance to flow out to a fraction collector, and the other eluent to flow out to the membrane module. .
き、保持時間の変化に応じて溶離液流入バルブが開閉さ
れる特許請求の範囲第1項記載の分離精製装置。(2) The separation and purification apparatus according to claim 1, wherein the retention time of the target substance is input into a computer, and the eluent inflow valve is opened and closed according to changes in the retention time.
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP60111653A JPS61270658A (en) | 1985-05-25 | 1985-05-25 | Separating and refining device |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP60111653A JPS61270658A (en) | 1985-05-25 | 1985-05-25 | Separating and refining device |
Publications (1)
Publication Number | Publication Date |
---|---|
JPS61270658A true JPS61270658A (en) | 1986-11-29 |
Family
ID=14566777
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
JP60111653A Pending JPS61270658A (en) | 1985-05-25 | 1985-05-25 | Separating and refining device |
Country Status (1)
Country | Link |
---|---|
JP (1) | JPS61270658A (en) |
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20200331960A1 (en) * | 2017-07-17 | 2020-10-22 | UCB Biopharma SRL | Chromatography |
-
1985
- 1985-05-25 JP JP60111653A patent/JPS61270658A/en active Pending
Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20200331960A1 (en) * | 2017-07-17 | 2020-10-22 | UCB Biopharma SRL | Chromatography |
US11498941B2 (en) | 2017-07-17 | 2022-11-15 | UCB Biopharma SRL | Chromatography |
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