CN1366180A - Efficient liquid-phase conic chromatographic column - Google Patents
Efficient liquid-phase conic chromatographic column Download PDFInfo
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- CN1366180A CN1366180A CN 02106928 CN02106928A CN1366180A CN 1366180 A CN1366180 A CN 1366180A CN 02106928 CN02106928 CN 02106928 CN 02106928 A CN02106928 A CN 02106928A CN 1366180 A CN1366180 A CN 1366180A
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- cone
- tapered pole
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- 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
- G01N30/60—Construction of the column
- G01N30/6052—Construction of the column body
- G01N30/6065—Construction of the column body with varying cross section
Abstract
A high-efficiency cone-shaped preparation column for liquid chromatography is composed of head body and tail of column with inside diameter of head inlet larger than inside diameter of tail outlet. Cone-shaped liquid flow guide channel, distribution disk and sieve plate constitute the liquid flow distributing system located on the head of the column. The distribution disk consists of multiple radial liquid flow channels concentric liquid collecting channel and liquid exuding holes. Inside diameter of the inlet is 2R=10mm-2000mm and inside diameter of the outlet is 2r=3mm-1800mm, the length of the column is L=5cm-100cm. Angle of cone is is in range of 1-20 degrees. Various chromatographic separation media can be filled into the cone. The ivnented cone-shaped preparation column can increase column efficiency and specimen loading and its diluting effect of separated components is lower than that of cylindrical chromatographic column.
Description
Technical field
The invention belongs to a kind of performance liquid chromatographic column that is used to prepare separation, be applicable to that petrochemical complex, fine chemistry industry, biotechnology, pharmacy etc. need obtain the field of net product, the preparation that is particularly useful for the highly purified complex samples of needs such as biomacromolecule, active skull cap components separates.
Background technology
Chromatogram originated from 1906, and the botanist M.S.Tswett of Russia has separated plant pigment with chromatography first.Early stage preparative liquid chromatography is mainly used in the purification that separates of organic compound and petroleum products, purity requirement to product is not high, therefore, the viewpoint in past thinks that imitating at preparative column chromatogram center pillar is not subject matter, people's real concern be yield of unit time.But, since the eighties in 20th century, pharmacy and development of biology require samples such as medicine, protein, nucleic acid to have more than 95% even 99.9% purity, its preparation amount not only will be satisfied with milligram such as structure, activity, physical property measurement and toxicological experiment to the requirement of gram level, and to reach restrain to the kilogram levels suitability for industrialized production to satisfy the demand of clinical practice.Applied widely, quick, the efficient and gentle condition of high performance liquid chromatography makes it become first-selected separation means.To the requirement of high separating efficiency, the research of new packing and new column packing technique development have been promoted.In recent years, people are more prone to the small article size fillers of operational analysis level in preparative chromatography, and its purpose is exactly to imitate in order to improve post.Because the separation and purification difficulty of some gene engineering product and natural products is bigger, need repeatedly cycling could obtain a certain amount of pure product, make preparation cost become the controlling factor of product price.Therefore, the post that how to improve preparative liquid chromatography is imitated and selectivity, reduces separating cycle number of times and the detachment process dilution to target components, and reducing production costs becomes the most significant problems of research.
In preparative chromatography was separated, people once considered a wide internal diameter post is connected with a long narrow internal diameter post, because the wide internal diameter post of front has bigger capacity, can avoid the pillar overload, and the narrow internal diameter post of back can provide effective separation.Nineteen sixty-eight, [Joel Kwok such as Joel Kwok, L R.Snyder, J.C.Sternberg.Anal.Chem., 1968,40 (1): 118-122] studied the problem that the different inner diameters post is connected, find that series connection is not to improve the good method that post is imitated, because the plate height of the wide internal diameter post of front is very big in the columns in series, the narrow internal diameter post of back is compared sometimes with it and can be ignored, and therefore can not play effectively and improve the effect that post is imitated.But, 1981, Said [A.S.Said.Journal of High Resolution Chromatography ﹠amp; ChromatographyCommunications, 1979, Febuary:63-67] draw tapered pole [100 (column length) * 3 (inlet internal diameter) → 1 (outlet internal diameter) by Theoretical Calculation, unit: cm, down with], series connection cylindrical pillars [41.7 (thick column length) * 3 (thick column internal diameter)+58.3 (buttress shaft length) * 1 (buttress shaft internal diameter) that total length identical more identical in quality than stationary phase, unit: cm] higher efficient arranged, the tapered pole post is imitated and is almost 2 times of columns in series post effects, and the tapered pole that therefore has the even transition of wide portion internal diameter is hopeful to be used for preparative chromatography.
1966, [H.M.Stahr, R.M.Ikeda, the E.T.Oakley such as H.M.Stahr in U.S. Philip Marris research centre, B.M.Carter.Anal.Chem., 1966,38:19741 makes low pressure taper liquid-phase chromatographic column first, and they adopt vinyon as column material, silica gel is stationary phase, the homogenate method is filled, and sample is an azobenzene, and eluant, eluent is a normal hexane.Be to investigate the characteristics of tapered pole, having made two in addition has the chromatographic column of equal volume to compare with tapered pole, and one is the cylindrical glass post of routine, and another is three rank cylindrical glass posts of internal diameter varies.Relatively the bands of a spectrum volume of three pillars to be weighing column efficiency, and the result shows that tapered pole compares with three rank cylindrical pillars, and post is imitated basically identical, but the tapered pole post imitates and obviously be better than the single order cylindrical pillars, and its bands of a spectrum volume is half of single order cylindrical pillars.Nineteen ninety, the Moerker Theophile of Switzerland has invented a kind of tapered pole [Switzerland Patent CH675212,1990] of internal diameter less than the outlet internal diameter that enter the mouth.Can be when sample flows in post gradually to the tube wall diffusion, in the tapered pole that this internal diameter enlarges gradually, because column type has met the trend of sample diffusion, solute band can not be diffused into tube wall, thereby presents " infinite diameter effect ".It is high that the great advantage of this tapered pole is that post is imitated, but because the inlet internal diameter is little, thus the ability of load sample is less than same volume, with the column type post of length.In addition, the research of relevant taper liquid-phase chromatographic analysis post also has some reports, tapered pole (150 * 4 → 2.5mm, 0.6 ° of cone angle) [A.Pecavar, I.Vovk as Slovenia country chemistry institute report, J.Marsel, M.Prosek.Analytical Sciences.1999,15:233-240], the tapered pole (25mm * 2 → 0.5mm of the Nugent Kerry D. invention of the U.S., 3.4 ° of cone angles) [World Patent WO00/74808,2000].But, do not have report so far about efficient liquid-phase conic chromatographic column.
Summary of the invention
At above-mentioned background, the purpose of this invention is to provide the liquid chromatography preparative column that a kind of post effect is high, volume containing the sample is big, sample is had lower dilution effect, be applicable to that petrochemical complex, fine chemistry industry, biotechnology, pharmacy etc. need obtain the field of net product, the preparation that is particularly useful for the highly purified complex samples of needs such as biomacromolecule, active skull cap components separates.
A kind of efficient liquid-phase conic chromatographic column of the present invention mainly constitutes (Fig. 1, Fig. 2) by the column cap that is positioned at cylinder top, cylinder and three major parts of post tail of being positioned at the cylinder bottom.Be the taper cylinder of column cap inlet internal diameter (2R) greater than post tail outlet internal diameter (2r).The taper cylinder is the cone of the even transition of inlet internal diameter 2R=10mm～2000mm, outlet internal diameter 2r=3mm～1800mm, column length L=5cm～100cm, and the cone cone angle is in 1 °～20 ° scopes, and the angle of optimum taper angle is 3 °-15 °.Can fill various types of chromatograph packing materials in the cone, as positive, reversed-phase liquid chromatography filler, or size exclusion chromatograph filler and affinity chromatography filler etc.
For the tapered pole of inlet internal diameter 2R＜40mm, column cap comprises nut, screw thread, taper gathering sill, O shape circle, distribution plate and porous sieve plate (Fig. 1).The tapered pole of inlet internal diameter 2R＞40mm adopts flange seal, and column cap comprises ring flange, bolt, taper gathering sill, packing ring, distribution plate and porous sieve plate (Fig. 2).
For the tapered pole of inlet internal diameter 2R＜40mm, the post tail comprises sieve plate, taper gathering sill, screw thread, O shape circle, nut (Fig. 1).The tapered pole of inlet internal diameter 2R＞40mm adopts flange seal, and the post tail comprises ring flange, bolt, taper gathering sill, packing ring and porous sieve plate (Fig. 2).The post tail is the pyramidal structure of gradual change, and sectional area changes the stream dead angle of causing suddenly when being connected with connecting pipe owing to post with minimizing.
The used distribution plate of column cap of the present invention is characterized in that the inlet internal diameter of its diameter and tapered pole mates.During for cylinder inlet internal diameter 2R＜40mm, adopt distribution plate as shown in Figure 3, promptly on distribution plate top, 3/5 of radius has concentric circles liquid stream collection channel, in the circle 12 flow passages is radially arranged, and every the center of a passage liquid-leadage hole is arranged; The passage of no liquid-leadage hole and above-mentioned concentric circles joining respectively have a liquid-leadage hole.Have the passage of liquid-leadage hole outwards to tell two passages (the passage angle is 120 °) at above-mentioned concentric circles joining place (no liquid-leadage hole), the central point of two passages has a liquid-leadage hole respectively.All passages are of a size of 0.1～0.5mm, and the liquid-leadage hole aperture is 0.5～0.8mm.During for cylinder inlet internal diameter 2R＞40mm, can increase liquid-leadage hole and concentrically ringed number as required, or adopt other any type of distribution plate.The distribution plate effect is: sample average is distributed on the whole column section, makes full use of the column cap cross section, prevent the local overrich of sample; There is more hole in zone away from post jamb, helps liquid and flows into.
Taper liquid stream gathering sill structure of the present invention, adopting cone angle is 120 °～180 °.Its role is to: when a large amount of samples enter on the column cap, can be distributed to rapidly on the xsect of whole chromatographic column, in time be washed away, avoid causing local overload of post and band broadening because of the local overrich of sample by moving phase; The destruction of avoiding sample in narrow injection port, to form injection stream and bed being caused; Get rid of the column cap bubble, prevent that the existence of bubble from destroying the distribution of sample at column cap.
Essential characteristic of the present invention is that this taper preparative column can improve post significantly and imitate and the sample charge capacity, and the dilution effect that target components is separated significantly reduces than cylindrical chromatographic column.The present invention mainly utilizes the tapered pole internal diameter to reduce gradually, the amplitude that causes the moving phase linear velocity increase of tube wall district is greater than center in the post, and the moving phase linear velocity from post inner tubal wall district to post in the center progressively reduce, thereby make chromatogram bands of a spectrum flow pattern be parabolic shape now.The inlet internal diameter 2R of the tapered pole by designing certain even transition, outlet internal diameter 2r, column length L or cone angle can make tapered pole under optimum flow rate, and the parabolic curvature of bands of a spectrum flow pattern reduces, and obtains near stopper shape flow pattern, imitates thereby improve post; Compare with the column type post of same volume, equal length, adopt the long-pending structure of cone entrance section, the sample load capacity of tapered pole is increased, thereby improve productive rate greater than discharge area; The cone internal diameter of tapered pole reduces to reduce to dilution of sample degree (Fig. 4, Fig. 5) gradually, and this is significant to preparative chromatography, because the solvent that only need evaporate still less just can obtain reducing solvent consumption with the product of measuring, reduces production costs.
Description of drawings
Fig. 1 is the efficient liquid-phase conic chromatographic column synoptic diagram of thread seal;
Fig. 2 is the efficient liquid-phase conic chromatographic column synoptic diagram of flange seal;
Fig. 3 is the distribution plate synoptic diagram;
Fig. 4 is the chromatogram of one-component sample taper preparative column when flow velocity is 4mL/min;
Fig. 5 is the chromatogram of one-component sample column type preparative column when flow velocity is 4mL/min.
Among the figure: the 1-column cap; The 2-cylinder; 3-post tail; 4-taper gathering sill; The 5-distribution plate; The 6-porous sieve plate; The 7-nut; The 8-screw thread; 9-O shape circle; The 10-ring flange; The 11-bolt; The 12-packing ring; The 13-flow passage; The 14-liquid-leadage hole.
Embodiment
The column cap of taper liquid chromatography preparative column links to each other with sampling valve, sample and solvent enter tapered pole through high pressure pump, sampling valve, by the acting in conjunction of taper gathering sill, distribution plate and sieve plate with the sample uniform distribution at the chromatographic column column cap, sample passes through repeated dispensing in the taper chromatographic column after, go out chromatographic column through post tail sheet plate current, enter detecting device.
Embodiment 1: taper liquid chromatography preparative column inlet internal diameter is 20mm, and the outlet internal diameter is 7mm, long 5cm.By cylinder shown in Figure 1 two is screw bolt-type column cap 1 and post tail 3, and they are compatible with the upper/lower terminal of cylinder 2 respectively with screw thread 8 and nut 7; The inner face of column cap 1 and post tail 3 is recessed into taper gathering sill 4; There are O shape circle 9, distribution plate 5, porous sieve plate 6 in column cap 1 below; Post tail 3 top O shape circles 9, porous sieve plate 6.By shown in Figure 3, distribution plate 5 tops, there is concentric circles liquid stream collection channel at 3/5 place of radius, there are 12 to be the radiation-like flow passage in the circle, a liquid-leadage hole, the passage of no liquid-leadage hole arranged every the center of a passage, punish out two passages with above-mentioned concentric circles joining, its angle is 120 °, and the central point of two passages has a liquid-leadage hole respectively, and all channel sizes are 0.1-0.8mm.
Column type column internal diameter 14mm, long 5cm.
Two kinds of chromatographic columns have identical volume, the amorphous C of dry-packing 40-63 μ m in the post
18The employing cyclohexane is a moving phase, and flow velocity is 2-14mL/min, and sample is the cyclohexane saturated solution of iodine.With as seen-UV-detector is that the 470nm place is detected at wavelength.Because sample solution is a colored solutions, can clearly observe the bands of a spectrum flow pattern of sample.The result shows: the parabolic curvature of tapered pole bands of a spectrum flow pattern is significantly less than the column type post; The post of tapered pole is imitated and is higher than the column type post; The elution curve peak area of tapered pole is greater than column type post (Fig. 4,5).
Embodiment 2: taper liquid chromatography preparative column inlet internal diameter is 800mm, and the outlet internal diameter is 280mm, long 25cm.By cylinder shown in Figure 22 two column cap 1 and the post tail 3 that ring flange 10 is formed is housed respectively; Ring flange 10 is fastening by bolt 11, in the middle of it packing washer 12 is arranged, and the inner face of column cap 1 and post tail 3 is recessed into taper gathering sill 4; There is distribution plate 5 column cap 1 below, porous sieve plate 6; There is porous sieve plate 6 post tail 3 tops.By shown in Figure 3, when big, can increase liquid-leadage hole and concentrically ringed number as required, or adopt other any type of distribution plates for cylinder inlet internal diameter 2R.The effect of distribution plate is, sample average is distributed on the whole column section.
Fill the spherical C of 10 μ m in the post
18The chromatogram carrier.The employing cyclohexane is a moving phase, and flow velocity is 3-22L/min, and sample is the cyclohexane saturated solution of iodine.With as seen-UV-detector is that the 440nm place is detected at wavelength.The result shows: the post of tapered pole is imitated and is higher than the column type post; The elution curve peak area of tapered pole proves dilution of sample than column type post little (Fig. 4, Fig. 5) greater than the column type post.
Claims (7)
1. an efficient liquid-phase conic chromatographic column is characterized in that, the chromatogram preparative column mainly is made up of column cap, cylinder and post tail three parts, is the taper cylinder of column cap inlet internal diameter greater than post tail outlet internal diameter.
2. liquid chromatography preparative column according to claim 1, it is characterized in that, its cylinder is the cone of the even transition of inlet internal diameter 2R=10mm～2000mm, outlet internal diameter 2r=3mm～1800mm, column length L=5cm～100cm, and the cone cone angle is in 1 °～20 ° scopes.
3, liquid chromatography preparative column according to claim 2 is characterized in that, described cone De Cone angle is at 3 °～15 °.
4. liquid chromatography preparative column according to claim 1 is characterized in that,
(a) for the tapered pole of inlet internal diameter 2R＜40mm, column cap comprises nut, screw thread, taper gathering sill, O shape circle, distribution plate and porous sieve plate;
(b) for the tapered pole of inlet internal diameter 2R＜40mm, the post tail comprises sieve plate, taper gathering sill, screw thread, O shape circle and nut.
5. liquid chromatography preparative column according to claim 1 is characterized in that
(a) for the tapered pole of inlet internal diameter 2R＞40mm, adopt flange seal, column cap comprises ring flange, bolt, taper gathering sill, packing ring, distribution plate and porous sieve plate;
(b) for the tapered pole of inlet internal diameter 2R＞40mm, adopt flange seal, the post tail comprises ring flange, bolt, taper gathering sill, packing ring and porous sieve plate.
6. according to claim 4,5 described liquid chromatography preparative columns, it is characterized in that the inlet internal diameter coupling of distribution plate diameter and tapered pole comprises a plurality of radial flow passages, concentric circles liquid stream collection channel and liquid-leadage hole:
(a) for the tapered pole of cylinder inlet internal diameter 2R＜40mm, on distribution plate top, 3/5 of radius has concentric circles liquid stream collection channel, in the circle 12 flow passages is radially arranged, and every the center of a passage liquid-leadage hole is arranged; The passage of no liquid-leadage hole and above-mentioned concentric circles joining respectively have a liquid-leadage hole; Have the passage of liquid-leadage hole outwards to tell two passages at above-mentioned concentric circles joining place, the passage angle is 120 °, and the central point of two passages has a liquid-leadage hole respectively; All passages are of a size of 0.1～0.5mm, and the liquid-leadage hole aperture is 0.5～0.8mm;
(b) for the tapered pole of cylinder inlet internal diameter 2R＞40mm, can increase liquid-leadage hole and concentrically ringed number as required.
7, ask 4 and 5 described liquid chromatography preparative columns according to right, it is characterized in that, described taper gathering sill, its cone angle are 120 °～180 °.
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| CN 02106928 CN1204402C (en) | 2002-03-07 | 2002-03-07 | Efficient liquid-phase conic chromatographic column |
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Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2007025467A1 (en) * | 2005-08-27 | 2007-03-08 | Ke Wang | A liquid-phase chromatographic separating column |
| CN101537343A (en) * | 2009-04-10 | 2009-09-23 | 朱靖博 | Conical dynamic axial compression high-pressure chromatographic column |
| CN102160937A (en) * | 2011-03-23 | 2011-08-24 | 江苏迪沃特仪器设备科技有限公司 | Dynamic axial compression column fluid distribution device |
| CN102628841A (en) * | 2011-03-24 | 2012-08-08 | 苏州汇通色谱分离纯化有限公司 | Parabolic high-performance liquid chromatographic preparative column |
| CN103285656A (en) * | 2013-06-21 | 2013-09-11 | 苏州新区枫桥净化设备厂 | A purifying column |
| CN102062765B (en) * | 2009-11-16 | 2013-10-09 | 中国科学院大连化学物理研究所 | Column head structure of conical preparative liquid chromatographic column |
| CN104849378A (en) * | 2015-04-29 | 2015-08-19 | 华东理工大学 | Capillary column system capable of increasing open tubular column sample loading quantity by variable-diameter packing column |
| CN106062551A (en) * | 2013-09-18 | 2016-10-26 | 安捷伦科技有限公司 | Liquid chromatography columns with structured walls |
| TWI635894B (en) * | 2017-01-26 | 2018-09-21 | 義守大學 | Fluid distributor and design method thereof |
| CN111939595A (en) * | 2020-08-07 | 2020-11-17 | 青岛理工大学 | Chromatographic column device, equipment and method |
| WO2023080868A1 (en) * | 2021-11-05 | 2023-05-11 | Eczacibaşi Monrol Nükleer Ürünler Sanayi̇ Ve Ti̇caret Anoni̇m Şi̇rketi̇ | Separation column with variable geometry |
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2002
- 2002-03-07 CN CN 02106928 patent/CN1204402C/en not_active Expired - Fee Related
Cited By (13)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2007025467A1 (en) * | 2005-08-27 | 2007-03-08 | Ke Wang | A liquid-phase chromatographic separating column |
| CN101537343A (en) * | 2009-04-10 | 2009-09-23 | 朱靖博 | Conical dynamic axial compression high-pressure chromatographic column |
| CN102062765B (en) * | 2009-11-16 | 2013-10-09 | 中国科学院大连化学物理研究所 | Column head structure of conical preparative liquid chromatographic column |
| CN102160937A (en) * | 2011-03-23 | 2011-08-24 | 江苏迪沃特仪器设备科技有限公司 | Dynamic axial compression column fluid distribution device |
| CN102628841A (en) * | 2011-03-24 | 2012-08-08 | 苏州汇通色谱分离纯化有限公司 | Parabolic high-performance liquid chromatographic preparative column |
| CN103285656A (en) * | 2013-06-21 | 2013-09-11 | 苏州新区枫桥净化设备厂 | A purifying column |
| US10718743B2 (en) | 2013-09-18 | 2020-07-21 | Agilent Technologies, Inc. | Liquid chromatography columns with structured walls |
| CN106062551A (en) * | 2013-09-18 | 2016-10-26 | 安捷伦科技有限公司 | Liquid chromatography columns with structured walls |
| US10073068B2 (en) | 2013-09-18 | 2018-09-11 | Agilent Technologies, Inc. | Liquid chromatography columns with structured walls |
| CN104849378A (en) * | 2015-04-29 | 2015-08-19 | 华东理工大学 | Capillary column system capable of increasing open tubular column sample loading quantity by variable-diameter packing column |
| TWI635894B (en) * | 2017-01-26 | 2018-09-21 | 義守大學 | Fluid distributor and design method thereof |
| CN111939595A (en) * | 2020-08-07 | 2020-11-17 | 青岛理工大学 | Chromatographic column device, equipment and method |
| WO2023080868A1 (en) * | 2021-11-05 | 2023-05-11 | Eczacibaşi Monrol Nükleer Ürünler Sanayi̇ Ve Ti̇caret Anoni̇m Şi̇rketi̇ | Separation column with variable geometry |
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