CN1035471C - Preparation-type high voltage electrophoretic separating device of difference speed counter-current - Google Patents
Preparation-type high voltage electrophoretic separating device of difference speed counter-current Download PDFInfo
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- CN1035471C CN1035471C CN92103718A CN92103718A CN1035471C CN 1035471 C CN1035471 C CN 1035471C CN 92103718 A CN92103718 A CN 92103718A CN 92103718 A CN92103718 A CN 92103718A CN 1035471 C CN1035471 C CN 1035471C
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Abstract
The present invention relates to a high-voltage electrophoretic separating device of differential-speed counter current, which is used for separating and purifying biological substances of industrial scale and is mainly composed of an electrophoretic column having a favorable heat exchanging structure, an electrode chamber, electrodes, etc., wherein separating media with different properties are arranged in the electrophoretic column. The present invention has the characteristics of large processing capacity, high resolution, convenient operation, etc.; the present invention can be used for the high-purity and continuous batch production of biological products and biological medicine so as to have considerable economic and social benefit.
Description
The invention belongs to can be for the biological substance separation and purification equipment of industrial-scale production.
Develop rapidly along with bionic, carry out bigger in batches preparation separation and purification at for example various special efficacy pharmaceutical grade proteins of some important biological substance, hormone, interferon etc., be that people urgently expect the new problem that solves, also be to form pressing for of rising high tech, high profit industry.
Electrophoresis is undoubtedly a kind of effective analytical test instrument as the basic isolation technics of analysis and profiling protein matter complex mixture in the biological technical field.Its basic principle is: have the cation (+) of electrostatic charge or anion (-) under the extra electric field effect according to the difference of its " charge-mass ratio ", " differential effect " of moving with different movement velocitys is separated from each other ionic constituent different in the solution.In the Biochemical Research of protein, to compare with other analytical separation means, electrophoretic separation technique has mild condition, resolution ratio height, easy to operate, advantage such as amount of samples is few, therefore, has obtained using widely in clinical medicine and laboratory research.Yet, regrettably, when the production that up to now electrophoretic separation technique is applied to a large amount of protein examples prepares, no matter be its separation accuracy or its practical degree, all fail far away to reach and be used for the such level of protein analysis.Even the electrophoresis method that has also can be used for the preparation of a small amount of protein example, still, because these electrophoresis methods are difficult to realize serialization production in enormous quantities, thereby, be only limited to laboratory scale at present and use and preparation in a small amount, and with high costs.Suitable high voltage is the effective measures that improve electrophoresis rate, increase disposal ability, can bring problems such as heat radiation difficulty, convection current and enlarge-effect increase the weight of but apply high voltage and increase the electrophoretic column size.Address the above problem, and take measures to realize the on-line monitoring of continuous operation and process, then utilize novel electrophoretic separation technique to replace conventional art to realize separation and purification and production in enormous quantities biological and biochemical substances, not only become and may but also can reduce production costs significantly.
The object of the present invention is to provide a kind of plant-scale differential adverse current electrophoresis separator that can be used for, make it to be used for the particularly separation and purification of protein of biological substance.
The present invention compares with existing preparation type electrophoretic separation device, and principal character is:
(1) this device can carry out continuous system fully, but also intermittently operated.The comparable common electrophoretic column of electrophoretic column size is much bigger, so production capacity can improve several times.In the I of this device electrophoretic column, II two zones, respectively with the particular form dress bead polymer gel with different grain size and different cell sizes, the middle part is the purifying enriched chamber of the target product that is made of porous plate.The convection problem that this has not only solved the sample component has improved the resolution ratio of process; And can when guaranteeing target product purity, carry out the serialization lock out operation.In addition, intermittently during lock out operation, target product also can be concentrated in purification chamber.
(2) whole electrophoretic column has special heat exchange structure (seeing accompanying drawing 2,3).Fig. 2 is the A type, is applicable to the situation that general preparation amount or multiplication factor are not high, and the outer sleeve of pillar is communicated with inner radial heat exchange arm, the cooling medium cooling that circulates therein simultaneously; Fig. 3 is a Type B, is applicable to the preparation requirement that treating capacity is bigger, and pillar is made shell and tube, load the bead polymer gel in the pipe on request, shell side passes to cooling medium, and this can realize good heat exchange on the one hand, can alleviate even eliminate the influence of convection current on the other hand greatly.In addition, add continuous feeding and discharging, thereby thoroughly solved the queueing problem of electrophoresis heat, guaranteed the permanent cryogenic conditions in the commercial scale electrophoretic separation post reliably.
(3) the indoor buffer solution of electrophoretic column two end electrodes is made the unique design that circulates respectively, has in time taken away the gas that electrode generates, and has solved in the free solution electrophoresis because of producing the influence of bubble to separation process; Simultaneously, the buffer solution diffusion that provides by perforated membrane can also guarantee the stable of PH environment in the whole electrophoretic separation post.
(4) target product can carry out on-line monitoring by the optical fiber testing system that this device is specially joined in the concentration process and the concentration of purifying enriched chamber.
Accompanying drawing 1 is a separation principle schematic diagram of the present invention.
Wherein, P represents target protein, and α, β represent two kinds of separating mediums (as the gel of different-grain diameter or different cell sizes) with different physical properties respectively.This two media will cause the flowing velocity of solution in I, II zone different.In the electrophoretic separation post, various ionic constituent all have two kinds of motion modes: electrophoresis motion and material flow flow.In the I district, has the carrier fluid flowing velocity V of target protein P
r, surpassed should the zone electrophoretic migration speed V
eThereby target protein will be washed away by logistics, and the direction of two kinds of clean differences of speed (VI) is by I → II; Otherwise in the II district, the target protein P with higher electrophoretic velocity will not submit to has moving and " sailing against the current " of relatively low carrier fluid flow velocity, and the direction of two kinds of clean differences of speed (VII) is by II → I.Corollary is: under suitable operating condition, target protein P cancels out each other and obtains " enrichment " in the intersection speed in I, II two zones, finishes and other separate impurities purifying.This separation process is set up according to biological substance bulk of molecule and two physical factors of charged character, and therefore, more simple electrophoresis and simple chromatographic separation technology have higher resolution ratio.
Accompanying drawing the 2, the 3rd, apparatus structure schematic diagram of the present invention.
Accompanying drawing 2 is A type device of the present invention.Wherein, 1 is cooling jacket, and 2 for being communicated with 1 column internal diameter to the cooling arm, and cooling medium circulates in chuck and arm, thereby realizes the heat exchange to electrophoretic column; 3 is electrode chamber, circulates for cushioning liquid, takes away the gas that electrode produces; 4 is electrophoretic separation post main body, and the bead polymer gel of different physical properties is equipped with at two ends respectively; 5 is platinum electrode, produces the uniform DC electric field at 4 two ends; 6 is separating medium (bead polymer gel), both can play the effect that prevents the sample convection current, simultaneously, participates in separation process again, improves resolution ratio; 7 is porous plate, in order to separate different separating mediums, constitutes the purifying enriched chamber of target protein simultaneously; 8 is fibre-optical probe, in order to the concentration of tracking and monitoring target protein; 9 is target protein purifying enriched chamber; 10 is stainless (steel) wire, as the supporter of separating medium 6 and perforated membrane 11; 11 is perforated membrane, is communicated with electrode chamber and main body splitter, constant in order to the pH value of guaranteeing whole splitter, prevents protein composition to be separated diffusion in electrode chamber simultaneously; 12 is "O, seals.
Explanation to import and export in the accompanying drawing 2: on behalf of negative pole circular buffering liquid, A, A ' import and export, B, the anodal circular buffering liquid of B ' representative are imported and exported, C, C ' represent the carrier fluid import and export of (containing the biological substance for the treatment of branch), and D, D ' represent the import and export of circulating cooling medium, the outlet of P representative products.
The present invention can be widely used in the ion that has different electric charges or have the large biological molecule of ionogen such as the separation and the purification of amino acid, polypeptide, protein, isoenzymes, various hormone and nucleic acid etc.Adjust operating parameter and can isolate wherein a kind of pure component from the mixing system of complexity, the way of also available increase segregation section and purifying enriched chamber is told several pure components simultaneously.
The invention solves the high purifying of biological products and biochemical drug, quantity-produced technology in enormous quantities and plant issue, wide prospect in industrial application is arranged, will play a significant role aspect bioengineering and separation of other macromolecular compound and the preparation.For example, this device has been successfully used to the separation preparation of γ-Gu aminoacyl transferring enzymes, and has obtained good economic benefit and social benefit.
Claims (1)
1. the preparation type high voltage electrophoretic separating device of difference speed counter-current of a separation and purification biological substance comprises the electrophoretic column that separating medium is housed, and purifying enriched chamber and the electrode chamber with circular buffering liquid and separation membrane is characterized in that:
A. electrophoretic column is divided into two zones, the gel separation media in dress different-grain diameter or different apertures in two zones, pillar has by the cooling jacket of radially arm connection or shell and tube cooling shell side, between two area segments, be provided with the concentrated and purified chamber that constitutes by porous plate, be provided with the optical fiber detection probe of monitoring concentration process and product purity in this chamber, and external test macro;
B. be provided with perforated membrane and supporter thereof between electrode chamber and the pillar, be respectively equipped with the import and export that circulate for buffer solution in the positive and negative electrode chamber again.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN92103718A CN1035471C (en) | 1992-05-20 | 1992-05-20 | Preparation-type high voltage electrophoretic separating device of difference speed counter-current |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN92103718A CN1035471C (en) | 1992-05-20 | 1992-05-20 | Preparation-type high voltage electrophoretic separating device of difference speed counter-current |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1066405A CN1066405A (en) | 1992-11-25 |
| CN1035471C true CN1035471C (en) | 1997-07-23 |
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| Application Number | Title | Priority Date | Filing Date |
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| CN92103718A Expired - Fee Related CN1035471C (en) | 1992-05-20 | 1992-05-20 | Preparation-type high voltage electrophoretic separating device of difference speed counter-current |
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Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107412718A (en) * | 2017-08-07 | 2017-12-01 | 安徽宏业药业有限公司 | A kind of biology extraction preparation method of high-purity medicinal bone peptide solution |
| CN111560063B (en) * | 2020-05-12 | 2022-11-01 | 蚌埠医学院 | Raw material medicine purification device for animal pancreas-derived insulin and use method |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4323439A (en) * | 1979-12-31 | 1982-04-06 | The Regents Of The University Of California | Method and apparatus for dynamic equilibrium electrophoresis |
| US4983267A (en) * | 1988-10-18 | 1991-01-08 | Innova/Pure Water, Inc. | Water deionization and contaminants removal or degradation |
| US5032247A (en) * | 1989-09-14 | 1991-07-16 | Separations Technology, Inc. | Method and apparatus for electrophoretic separations |
-
1992
- 1992-05-20 CN CN92103718A patent/CN1035471C/en not_active Expired - Fee Related
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4323439A (en) * | 1979-12-31 | 1982-04-06 | The Regents Of The University Of California | Method and apparatus for dynamic equilibrium electrophoresis |
| US4983267A (en) * | 1988-10-18 | 1991-01-08 | Innova/Pure Water, Inc. | Water deionization and contaminants removal or degradation |
| US5032247A (en) * | 1989-09-14 | 1991-07-16 | Separations Technology, Inc. | Method and apparatus for electrophoretic separations |
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| Publication number | Publication date |
|---|---|
| CN1066405A (en) | 1992-11-25 |
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