CN103764243A - Method and apparatus for improved resolution chromatography - Google Patents

Abstract

An apparatus and a method are provided for column chromatography which provide improvements in separation resolution and detection sensitivity, comprising a chromatography column, the column having an inlet and an outlet, wherein the outlet is configured to split a flow of eluate as it leaves the column through the outlet into at least two separate portions, wherein the apparatus is configured to separately process the portions, for example to separately detect a portion or separately collect fractions of a portion with improved resolution. A split frit assembly is preferably configured to split the flow of eluate. The portions preferably emanate from different radial regions of the column. An end fitting for the column outlet may be provided having multiple ports to separately convey the portions.

Description

For improvement of the chromatographic method and apparatus of resolution ratio

Invention field

The present invention relates to the field of column chromatography.

background of invention

Chromatographic column has been extensively developed and is conventional for analytic type and preparative scale chromatography.As everyone knows, the separation of the sample (being called again a kind of analyte or solute) of realizing a kind of mixture that comprises a plurality of components by following operation in chromatographic column: make sample dissolution in a kind of eluant, eluent to form fluid mobile phase and to make this mobile phase through being typically filled in the fixedly phase in tubulose post, thereby because different component causes that with the distribution difference (being that these components have different distribution coefficients) between fixing phase this sample separation becomes its component at this mobile phase.Eluant, eluent fluid is a kind of liquid the most commonly, but may be one other fluid as supercritical fluid (SCF), and the present invention relates to the post that together uses with liquid or SCF mobile phase.Yet the present invention does not relate to gas chromatography (being that fluid is not gas).In column chromatography, fixing in post typically in a kind of filler particles bed or a kind of form of porous monoblock.The invention particularly relates to packed column, but it is not limited only to packed column.Often these posts comprise the reusable post with disposable post core (cartridges), and these post cores and post are all columniform conventionally.The present invention can or have together with the post of other cross section profiles with cylindrical pillars (most preferably orbicular cylindrical pillars) and uses.That is to say, post jamb can have numerous cross section profiles, but most preferably the cross section of this post jamb has circular contour.

As described, make mobile phase pass post and along with the variation of time detects leaving the eluent of this post.The signal intensity (chromatogram) detecting in time shows to exist different components in mixture.The separation degree of these different components depends on separative efficiency or the resolution ratio of this post.The resolution ratio of this post depends on many factors.This type of factor comprises mobile phase and the fixing character of phase, these mobile phases and fixingly researched and developed widely mutually.

It is also known that, the shape of the sample being injected into is a factor that affects the resolution ratio of this post.Conventionally make sample substantially traverse the diameter of post and be incorporated in this post.Conventionally in the porch of post, use head joint, as sieve plate (frit).Distributor can be with helping to distribute sample to attempt and to obtain equal one deck that sample traverses the whole diameter of post.United States Patent (USP) 4,999,102 described a kind of for making uniform liquid be assigned to unit of extensive separator system and/or from wherein evenly collecting the manifold system of liquid.As entrance in the situation that, this manifold is divided and is sent away single flow point and be made into a plurality of stream by a row, and to guarantee that interface between this manifold and this unit makes liquid traverse a kind of pattern of roughly distributing equably whole unit the final multiplicity of flow passage device is arranged.With for export in the situation that, oppositely arrange this manifold to collect these streams and divide and send away these streams and join in single pipeline via a row from traverse the uniform distribution of this unit.In a similar fashion, US5,124,133 have described a kind of system of passing the homogeneous flow profile of packed bed for liquid, and this system relates to the top surface that makes this liquid traverse this packed bed and distributes equably.

The entrance system of known other types is often for different objects.For example, EP371648A has disclosed a kind of equipment that is designed for displacement chromatography, wherein has an entrance distributing manifold.With every kind of liquid flow of displacement chromatography order, pass the same radial zone of post.On the other hand, JP62-063857A has described a kind of tabular post having for a plurality of entrances of possible a plurality of samples, wherein can in the direction of inlet streams, apply an electric field so that the two dimensional separation of crossing over post plate to be provided being orthogonal to, this tabular post also has a plurality of outlets.

In JP62-240857A, described a kind ofly for the preparation of the chromatographic chromatographic column of type, the annular release of the stream of drawing from this post is wherein provided, made like this center flow stream to open with the periphery flow point that flows.Compare with the peak shape of elution band from outer shroud, better from the peak shape of the elution band of center ring, this demonstrates obvious band distortion.

EP257582 has described a kind of preparation scale chromatographic column, wherein by the pipe being inserted into from column outlet bed, sample is loaded in this post.This pipe occupy the bed center of column inlet below approximate 20%.That is to say, on this pipe, have packing material.Once sample is loaded on post, mobile phase just carries out wash-out through post (outside of sample inlet tube) to this sample.Then, sample exits from the outlet of post through a series of holes or the line of rabbet joint in exit.This outlet has a plurality of lines of rabbet joint on concentric position, but do not have center, does not exit the line of rabbet joint, because the central segment of inlet tube in this post.

Recently, about high performance liquid chromatography (HPLC), the people such as Xiali gram (Shalliker) flow fluid in post in the later stage nineties can be visual.In use, have in the fixedly phase of identical refractive index and the glass column of mobile phase, they can make fluid flow visual under the help of dye marker thing.Their result demonstrates, and the mode that sample is introduced in the bed of chromatographic column is important.Ideally, filler plug should be a kind of cylindrical-square plug, but various factors finally causes Parabolic sample band, for example, along with sieve plate porosity reduces, the plug flow more parabolic type that becomes.Cloth Roy strangles the visual of solute migration in this (Broyles), Xiali gram (Shalliker) and Guiochon(' chromatographic column. the impact of sieve plate porosity (Visualization of Solute Migration in Chromatographic Columns.Influence of the Frit Porosity) '. chromatography magazine A(J.Chromatography A.), the plug flow more parabolic type that becomes has been described along with entrance sieve plate porosity reduces 917(2001) 1-22).Use a distributor to improve the homogeneity of the flow velocity of the radial section that traverses bed, the increase axial dispersion of still will take is cost, and this has finally caused the significantly reduction of separating property.No matter sieve plate porosity or noly adopted a distributor, it is all inhomogenous that the sample that traverses post radial section distributes.The people (chromatography magazine A(J.Chromatography A) such as Xiali gram (Shalliker), 865(1999) 83-95) demonstrating sieve plate and distributor all can not be used for that sample is distributed equably and traverse post, and the trend of this sample more concentrated in the central area of bed (more dilution in the neighboring area of bed on the contrary) is larger.In addition, the diameter (cloth Roy strangles this (Broyles), Xiali gram (Shalliker) and Guiochon, chromatography magazine A(J.Chromatography A) that sieve plate diameter should matched column internal diameter, 855(1999), 367-382).If not, use the sieve plate with the diameter narrower than the diameter of post can cause very serious parabolic flow.For chromatogram worker, can know intuitively that sieve plate diameter must equal the diameter of column internal diameter, but can not always realize, particularly, by axial compression preparative column in the situation that, wherein head joint is in post itself.For the post of these types, the diameter of sieve plate must be less than post, because this sieve plate must be comprised in a housing unit to prevent leakage and the damage of post jamb.

In another research, the people such as Xiali gram (Shalliker) (' physical proof of two kinds of wall effects in liquid chromatography (Physical Evidence of two Wall Effects in Liquid Chromatography) '. chromatography magazine A(J.Chromatography A) .888(2000) 1-12 and ' in liquid-phase chromatographic column visual (the Visualization of Solute Migration in Liquid Chromatography Columns) of solute migration '. chromatography magazine A(J.Chromatography A) .826(1998) 1-13) proved when sample being incorporated into the bed in the approximate 1cm depth in sieve plate below via central point injection (CPI), in the radial section of the center of this bed, there is the migration of this sample simultaneously.Although when using CPI technology, the people such as Xiali gram (Shalliker) observe excellent separative efficiency, should be understood that a kind of like this sample introducing method is loaded down with trivial details and is easy to destroy post bed.Therefore, this technology is unsuitable for conventional application.In addition, although central point injects the most effective solute method of shipment allowing along post, because injecting, this type of allow the local central area of sample in the most effective fill area of this post concentrated, but modern post has effectively been avoided the use of this type of injection technique, because when sample is entered by the sieve plate in column top and distributor, valve injects and makes sample traverse this post dispersion.Like this, central point method for implanting is abandoned to a great extent.

The existence of post jamb has been discussed and has been approached these walls and filled the impact of this post on sample flow.Nox (Knox), laird (Laird) and thunder literary composition (Raven), (chromatography magazine (J.Chromatography) 122(1976), 129-145) demonstrates the flow velocity of very close wall slightly higher than the flow velocity at Zhu center.The disturbance region that they also demonstrate column filling extends in this post, thereby causes that serious band broadens and peak distortion.Approximately 30 times of particle diameters can be extended in the wall region of filling in good LC post at another in this post.Xiali gram (Shalliker), cloth Roy strangle this (Broyles) and Guiochon, (chromatography magazine A(J.Chromatography A), 888(2000) 1-12) existence of two kinds of wall effects described.These two kinds of wall effects are all caused by the inhomogeneities of column filling.Particles filled chromatographic column Gai Zhu center (radially) section has lower bed density, and this central segment is relative homogeneous, but surpasses this area, and packed density increases gradually towards wall.Therefore, in the central segment of post, flow velocity is the highest and approaches wall to be minimum, and this contributes to Parabolic plug flow.Yet, at the area of next-door neighbour's wall, packed density fast reducing.This factor is the result of the geometric properties of post and particle.The two is all rigidity for particle and post; Neither can be out of shape each other and adapt to.Therefore, the void space at wall place increases, and column permeability is the highest herein just.Therefore, in the region of next-door neighbour's post jamb, flow velocity is the highest.These two kinds of wall effects greatly contribute to reduce the efficiency of migration.

Cloth Roy strangles this (Broyles), Xiali gram (Shalliker) and Guiochon, (chromatography magazine A(J.Chromatography A), 867(2000), 71-92) proved except other things, migration distance after preset time is with radial position marked change, and moves back outlet connection and affect the peak shape detecting after post.

Therefore, be known that the sample that can not describe through particles filled chromatographic column by a cylindrical plugs flow model moves.Many factors (comprising the character of sieve plate, the inhomogeneities of the existence of distributor, injection process itself, wall effect and packed density) causes being sample plug or the band of parabolic type or bowl shape, this sample plug or band have the central area of a higher concentration conventionally, and is moving higher than more approaching under the speed of more diluting region of wall this central area.Also there is a region of the low concentration Rapid Flow of very close wall.The General Parabolic wire shaped of band makes the demand of column efficiency larger.Compare with the area of cylindrical expansion, need more plate to separate the area that parabola expands.Therefore, must realize separately by longer post, this causes the increase of time, also may cause the reduction of potential barrier flow velocity to adapt to longer post.The fixing wall effect that also may be subjected to mutually of integral body in post affects, and is no longer columniform through the plug flow of this type of post.

For this background, obtained the present invention.

summary of the invention

According to an aspect of the present invention, a kind of equipment for column chromatography is provided, this equipment comprises a chromatographic column, this post has an entrance and an outlet, wherein this outlet is configured to make this diverting flow become at least two parts of separating when an eluent stream is left this post through this outlet, and wherein this equipment is configured to process dividually the part that these separate.

According to another aspect of the present invention, provide a kind of method of column chromatography, the method comprises: a mobile phase is provided, and this mobile phase comprises a kind of sample that needs to be separated into a plurality of components; Make this mobile phase to an outlet longitudinal flow of post, pass a liquid-phase chromatographic column from an entrance of post, this mobile phase leaves this post as a kind of eluent by this outlet; When the stream of eluent leaves this post through this outlet, make this diverting flow become at least two parts of separating; And process dividually these at least two parts of separating.

According to an other aspect of the present invention, a kind of equipment for column chromatography is provided, this equipment comprises a chromatographic column, this post has an entrance and an outlet, wherein this outlet is configured to when a part of an eluent stream is left this post, guide this part so that a remainder of itself and this eluent is processed dividually, wherein this part flows out from a restricted radial zone of this post.

The present invention can provide many merits.At least one that separately process in type preferably includes the detection of eluent.The inventive example is improved from the mensuration performance of chromatographic column as made the detection of sample be strengthened and make.In different embodiment, due to the improved detection sensitivity in chromatographic determination and/or improved peak capacity and peak resolution ratio, the present invention may can realize for example by the detection lower limit of the species of chromatography.Have been found that, can obviously increase the number of theoretical tray, and in some embodiment for HPLC, compare with conventional post, the number that has been found that theoretical tray of the present invention increases over 50%, in some cases, do not compare with the similar conventional system of processing that separates of following with there is no the shunting of eluent, have been found that the increase of separative efficiency is higher than 50%.In the situation that the post of other types has been found that column efficiency (column plate counting) surpasses 100% increase.Conventionally, in the situation that column efficiency increases by one * 2 factor, resolution ratio increases by one * 1.44 factors.

Will be appreciated that, as a replacement scheme improving the peak resolution ratio of given column length, compare with a similar conventional system, the present invention can reach with shorter post given peak resolution ratio.A shorter post can make pending chromatographic isolation faster.An additional advantage is for example: only use a part of eluent for detection of meaning that the solvent load being incorporated in detector reduces, this can be highly profitable concerning some detector, and these detectors are as the mass spectrograph operating in vacuum environment and other detectors or biotype reaction detection device (as for finding the antioxidant detector of medicinal compound).Therefore, the present invention can use the post of the routine size with MS detection better.About preparative scale chromatography, the purer fraction (fraction) that the present invention can collect sample due to improved separative efficiency.To describe in more detail these and other advantages, and these and other advantages become clearer by following explanation of the present invention.

This equipment comprises a chromatographic column, this chromatographic column is a liquid-phase chromatographic column typically, but can be a supercritical fluid chromatography post, this post has an entrance and an outlet, thus contain a kind of solvent and need separated sample a mobile phase can by this entrance be incorporated in this post and along this post longitudinally (axially) flow to this outlet.This outlet is configured to make this stream (leaving the mobile phase of this post) shunting when an eluent stream is left this post.Configure this outlet so that this stream is split at least two parts of separating, and each part processed dividually, for example, guided in himself the treating apparatus that is directed to other part (or a plurality of part) that treating apparatus wherein separates.Configuring in this way column outlet can make being separated on post of eluent stream but not carry out after post.Thereby, leave a part of the eluent stream of this post and separate with another part, and process dividually with other part.In some preferred embodiment, leave a part of the eluent stream of this post and separate with another part, and detect dividually with other part.In some preferred embodiment, leave a part of the eluent stream of this post and separate with another part, and collect dividually the fraction of this part with other part.

Preferably, this outlet is configured to make this diverting flow to become at least two parts of separating when an eluent stream is left this post by this outlet: at least one first of this stream and at least one second portion of this stream.Preferably, the Yi Ge first of this stream of eluent is guided in first treating apparatus, and a second portion of this stream of eluent is guided in second treating apparatus that separates and be preferably different from this first treating apparatus with this first treating apparatus.For example, preferably this first treating apparatus comprises that one for detection of the detector that is present in the sample in this eluent, and this second treating apparatus can comprise a refuse receiver, or as the entrance of same or another chromatographic column, so that this second portion stands at least other one chromatography of taking turns.Yet this second treating apparatus can also comprise that one for detection of the detector that is present in the sample in this eluent.Can adopt many other treating apparatus and combinations thereof, and at this, discuss these devices in more detail below.In this way, for example, the invention provides a kind of post with an outlet, this outlet is for example configured to optionally guide, in a part to one first treating apparatus (detector) of eluent stream, and another part of this outlets direct is to another treating apparatus separating with this first treating apparatus simultaneously.

Preferably, at least two parts of separating of eluent flow out from the zones of different of post, and for example this first can flow out from a first area and this second portion can flow out from a second area of post (more specifically packed column bed).More preferably, these at least two parts of separating flow out from the different radial zones of post, and for example this first can flow out from being different from second radial zone of the post of this first radial zone from a first radial zone outflow and this second portion.At this term radial zone, mean cross section in post or a region of plane, perpendicular to the central axis of post or the cross section of longitudinal axis or plane.Thereby, this term radially or radially refer to the direction perpendicular to central axis or the longitudinal axis of post.Preferably, the first area of post is the radial zone being located substantially on away from the wall of this post.Preferably, this post is the packed column therein with a post bed, and a first area of this packed column is a radial zone, and eluent leaves this post from this radial zone through the part of the uniform filling of this post bed.Also more preferably, this first flows out from this post Yi Ge center radial zone, and this second portion flows out from being positioned at this center radial zone radial zone radially outward.Most preferably, Gai Zhu center radial zone is the region being located substantially on a central axis, and this central axis longitudinally runs through this post from entrance to outlet.In this way, a centronucleus of the eluent stream that contains relatively higher sample concentration and the better component of differentiating can be guided in a detector or other treating apparatus as a part, and the remainder of this eluent stream is for example guided to other places as one or more other parts, for example guide in one or more different treating apparatus.Thereby, be preferably different from the remainder of eluent stream and the first that processes dividually eluent stream with the remainder of eluent stream.Never in any form scope of the present invention is limited, it is believed that a part that is due to the fact that the eluent of obtaining from a restricted radial zone has than the less sample axially scattering of eluent of obtaining from traversing the full duration of this post, this is due to due to the parabolic shape of mobile sample band.

In certain embodiments, outlet is arranged to make this diverting flow to become more than two parts when the stream of eluent leaves post.For example, outlet in this type of embodiment can be arranged to make this diverting flow to become three or more parts of separating when the stream of eluent leaves post, each part is directed in the treating apparatus separating with other parts, therefore in this case, there are three or more the treating apparatus that separate.

As mentioned above, the concentration of the mobile example band in the LC post of filling reduces along with the increase of the distance from post central axis, be partly due in column diameter and traverse column diameter inhomogeneous flow and be partly due to fill bed in and fill diffusion that bed transporting be around associated and relevant mass transfer effect, the validity that the density of this filling bed self and uniformity can be filled with coupled columns changes.The sample band that this moves also trends towards having parabolic shape.These phenomenons can cause the shortcoming about sample separation efficiency, detection and resolution ratio, because conventional column outlet makes to assemble from the eluent that traverses the whole diameter of this post.Yet, the present invention can make to draw this post eluent diverting flow or cut apart so that part of this eluent stream is separated and can be processed dividually these parts with another part at least.For example, a part of the eluent better separating in addition that contains relative higher concentration sample optionally can be guided in a detector, so that the detection of enhancing is provided, wherein peak is also differentiated better.That is to say, by the component band in the sample of chromatography, in the centronucleus of post, become and axially scatter still less, so that due to the adjacent component detecting in sample, the peak in chromatogram becomes and separates better or differentiate.In preferred embodiment, detect the part that therefore only concentrates on the stream that the centronucleus (being center radial zone) from post elutes, because the part of this stream has than the radially outer regional concentration of more close post jamb higher sample relatively.In other words, can the application of the invention reduce or eliminate the harmful edge effect being present in eluent stream.The in the situation that of preparative scale chromatography, the part of the eluent better separating that contains relative higher concentration sample can be selected for classification separation to obtain the purer fraction of this sample.

As seen from the above, in a plurality of specific embodiments, this equipment preferably includes a detector, and this detector is arranged to detect dividually with other part or a plurality of part at least a portion of eluent.More preferably, this detector is arranged to separate detection from a part of the eluent of post Yi Ge center radial zone outflow.In other a plurality of specific embodiments, this equipment comprises a fraction collector, and this fraction collector is arranged to collect dividually with other one or more parts the fraction of at least a portion of this eluent.In this type of embodiment, this equipment can be configured to other one or more parts to deliver in a refuse receiver.In certain embodiments, this equipment can be configured to other one or more parts to deliver in the entrance of a post, for the further chromatography of this other one or more parts.Other one or more parts preferably flow out from an outer radial region of the post with respect to center radial zone.

Although can by can for the amount of the fluid of a detector for example of entering be reduced to a part only selecting eluent stream for detection of, the concentration of the sample in detector flow path is compared with regular situation to some extent to be increased.For some detector type (for example, as a mass spectrograph), the amount of wherein allowing the eluant, eluent that enters spectrometer should be low ideally, and some detector (for example, as UV detector) in the situation that, wherein flowing effect is correlated with, and this is a positive benefits.The in the situation that of mass spectrograph (MS) detection system because a part of only getting eluent according to the present invention for detection of, so this can mean that the present invention can use the post with the routine size that MS detects better.For example, a conventional 4.6mm HPLC post can typically have the flow velocity of about 1.5ml/min, yet the part that shunting arrangement of the present invention in certain embodiments can provide eluent for detection of, this part only has the approximately flow velocity of 0.2ml/min, MS system can be processed this part more simply, because evaporate the more solvent of heavy load, requires larger vacuum power.After some biosensors (as DPPH antioxidant detector) require post, reagent is to enter the stream (stream) that flows.Ratio by mobile phase and DPPH reagent carrys out regulation peak response.Can make the diverting flow of drawing post reduce valuably the consumption of DPPH reagent, and there is no loss of sensitivity.Can also there is minimum dead volume and adopt a plurality of checkout gears simultaneously.These detectors can be that sample destroys, because also a part of sample can be collected in fraction for further analysis.

The ratio of the corresponding volume of the different piece of eluent stream can change, thus stream to cut apart can be adjustable.Optimum flow ratio typically depends on carried out specific experiment.Along with the reduction of the ratio of collecting from central segment, separated efficiency system ground increases.This is a positive benefits because can depend on obtain separated associated difficulty or or even how many samples detector (as mass spectrograph) that must flow to sample load dependent form adjust this separation.In addition, can consider the environmental pressure of the method and change from an outer peripheral areas for example to the amount of the stream refuse and the amount of the stream of collecting from a central area of post.The present invention provides a kind of adjustable system that can weigh economic benefits and environmental benefit valuably.

Can change by following various in greater detail means the ratio of eluent stream.Preferably, a part of the eluent of separately processing is 70% or still less or 50% or still less (by volume) of total eluent, more preferably 30% or still less.For example, this part can be total eluent 50% or still less, 45% or still less, 40% or still less, 35% or still less, 30% or still less, 25% or still less, 20% or still less, 15% or still less, 10% or still less or 5% or still less.The part that this part preferably flows out from post Yi Ge center radial zone.The part that this part preferably detects dividually with one or more other parts or collects dividually.

Can by select different sieve section area ratio and/or by selecting to export in greater detail as follows the ratio of the number of port and/or the corresponding volume of the different piece that size changes eluent stream.

Can for example, by adjusting pressure in one or more passages and/or flow velocity (outlet pressure of difference), change the ratio (dividing degree) from the corresponding volume of these parts of the eluent stream of outlet, the one or more passage is connected on outlet port and in the downstream of this outlet port.Can be for example by means of the outlet pressure that is loaded with pressure at least one downstream passage of a part in these parts or flow regulator and changes difference, and/or can be for example by changing along the length of the pipeline of outlet port or by changing the outlet pressure that change difference along the diameter of the pipeline of outlet port.

This outlet is preferably equipped with a sieve plate, this sieve plate is wherein set and makes like this eluent that leaves post flow through this sieve plate.Preferably, within this sieve plate is positioned at the column internal diameter in exit.

Exit sieve plate is preferably in a sieve plate assembly, and this sieve plate assembly is configured to make this diverting flow to become at least two parts of separating when the stream of eluent leaves post.Configure in this way this sieve plate and can make separately carrying out on post but not after post of eluent stream.In order to make this diverting flow, preferably exit sieve plate assembly is a shunting sieve plate assembly (being called again partition type sieve plate assembly at this), this shunting sieve plate assembly comprises at least two sieve section of separating, these at least two sieve section of separating are separated from each other by one or more flow barriers, for example a kind of non-porous (non-porous) body can provide flow barrier, or a kind of non-porous coating can provide flow barrier, for example wherein this kind of coating is provided on one or more surfaces of at least one sieve section of separating, these one or more coated surfaces are in abutting connection with other sieve section or a plurality of sections.Thereby, flow the first of stream can be provided through the eluent of one or more the first sieve section, and flow the second portion of stream can be provided through the eluent of one or more the second sieve section, because the mobile eluent through the one or more the first sieve section separates with the eluent flowing through the one or more the second sieve section by the fluid barriers in non-porous body form.When eluent passes sieve plate assembly, flow barrier has stoped horizontal (radially) of the eluent between these sieve section to flow, thereby separately can make stream be isolated into part separately.Because these sections of shunting sieve plate have occupied the zones of different (especially different radial zones) of post, so this shunting sieve plate has determined that at least two of eluent parts of separating are from the zones of different of post (preferably as described different radial zones) outflow.On the other hand, a kind of single sieve plate sheet of routine is that efficiency is lower, because be not so orderly and therefore flow and may work as its mobile becoming during through this sieve plate through the eluent of the zones of different of post bed and be mixed to undesirable degree through the stream of the eluent of this sieve plate.Use a shunting sieve plate can make the eluent of the mobile zones of different through post bed draw this sieve plate with different parts, these different parts correspond to the zones of different of post.

A kind of preferred configuration of shunting sieve plate assembly comprises: at least one center sieve section, a non-porous body and at least one outside sieve section, this non-porous body surrounds this at least one center sieve section, and this at least one outside sieve section is surrounded this at least one center sieve section but separated therewith by this non-porous body.More preferably, this shunting sieve plate comprises: the center sieve section of a radial center, a non-porous body and an outside sieve section, and this non-porous body annular is surrounded this center sieve section, and this outside sieve section annular is surrounded this non-porous body.Most preferably, the center sieve section of this radial center is located substantially on from entrance to outlet and longitudinally runs through the central axis of this post.This type of shunting sieve plate configuration in exit allows that the one or more center sieve section produces a part of the stream flowing out from post Yi Ge center radial zone, and the one or more outside sieve section produces from being positioned at a part of this center radial zone stream that radially outward a radial zone of (periphery) flows out.

This center sieve section and outside sieve section can have different relative areas, thereby eluent stream is split into from having the central area of different relative areas and a plurality of parts of outer peripheral areas.Thereby the ratio of the area of these sieve section can be for changing a kind of means of ratio of corresponding volume of the splitter section of eluent stream.For example, the ratio of the area of the area of outside sieve section and center sieve section can be from 90%:10% to 50%:50% for example, more typically from 80%:20% to 50%:50%, change, but also can use the ratio outside these scopes.A preferred ratio of the outside area of sieve section and the area of center sieve section be from about 6:1 to about 1:1, more preferably from about 3:1 to about 1:1, also more preferably from about 2.5:1 about 1.5:1 extremely, and be most preferably about 2:1.

These sieve section can have identical or different density.For example, this center sieve section can have the density different from this outside sieve section.In the embodiment of a type, this center sieve section can have the density lower than this outside sieve section.Thereby, can control eluent preferably to flow through the sieve section with less dense for thering is a more highdensity sieve section.

Exit sieve plate assembly typically comprises a non-porous joint in outside (preferably being made by polymer), for example, make the non-porous jiont treatment in this outside to the port of export of post so that this sieve plate assembly forms a sieve plate cap.A kind of like this external lug is preferred, because for example a kind of steel sieve plate will can not seal against steel column wall well.Polymeric joint can for example, be made by different polymer (PTFE, ETFE, PEEK or Kel-F, more preferably PEEK).Conventionally, any non-porous part of this sieve plate assembly can for example, be made by plastics or polymer (PTFE, ETFE, PEEK or Kel-F, more preferably PEEK).

In some other embodiment, exit sieve plate assembly can comprise a single-piece porous sieve plate, is not a plurality of sections of sieve plate.As in other described embodiment, this single-piece porous sieve plate can be accommodated in an outside joint non-porous, preferably polymer, for example, make this jiont treatment to the outlet of post so that sieve plate assembly forms a sieve plate cap.

The non-porous joint in outside of exit sieve plate assembly can have a plurality of openings so that the flow point of eluent is opened.For example, the non-porous joint in this outside can have the opening of a radial center to allow the stream through a part of eluent from sieve plate, the stream of a part of this eluent flows out from a radial center region of post, and can have one or more peripheral openings of radially outward locating at this central opening and with permission, from sieve plate, pass the stream of a part of eluent, the stream of a part of this eluent flows out from a peripheral radial zone (surrounding this radial center region) of post.Can be for example there are one or more peripheral openings in sidewall by non-porous joint externally and the part of assembling the stream flowing out from the outer peripheral areas of post from the outside of sieve plate.

Exit sieve plate assembly preferably has circular external shape to meet circular cross-section post, although can for example depend on that post shapes use has the sieve plate assembly of other shapes.

The material of exit sieve plate can be the conventional sieve plate material using in LC, for example steel.Thereby this sieve plate can be become to make to flow by easy configuration with shunting mode described here and shunt through its eluent stream.For example, sieve plate material, thickness (degree of depth) and porosity can be that routine is used in LC system.For example, can use the sieve plate with 0.25mm to 2mm typical thickness.For example, can use the sieve plate of the 2 μ m porositys with nominal.Yet, can use the sieve plate with other porositys, for example, in the scope of nominal ground 0.1 μ m to 20 μ m.The non-porous body of shunting sieve plate embodiment preferably for example, is made by a kind of plastics or polymer (PTFE, ETFE, PEEK or Kel-F, more preferably PEEK), but can for example, by metal (, stainless steel), be made.This type of non-porous material can also be provided as an one or more lip-deep thin layer or the coating in one or more sieve section, the one or more sieve section in abutting connection with another sieve section so that a flow barrier to be provided.Non-porous flow barrier alternatively can be made by a kind of metal.A kind of like this metallic barrier can an one or more lip-deep thin layer or the coating as the one or more sieve section in abutting connection with another sieve section form by sputter.This type of thin layer or coating flow barrier can have advantages of lower resistance to eluent stream.

Compare with the width of sieve section, the width of flow barrier or non-porous body (in the radial direction measure) is preferably little, is preferably lower than each the width in these sieve section.This width is preferably as far as possible little, is that micron is big or small ideally.Thereby, thereby can in eluent stream, there is flow barrier and any dead time effect to after any possible resistance of the stream of the separative component of tool or flow barrier that causes can be decreased to minimum.Yet, will be appreciated that, this barrier should not be so thin so that can not effectively realize and make stream be isolated into part separately.

In certain embodiments, outlet can be equipped with a plurality of capillaries (outlet capillary) to guide eluent stream, and wherein one or more capillaries are arranged to guide the first of this stream and the second portion that one or more other capillaries are arranged to guide this stream.Thereby this first and second part is to guide in the capillary separating and therefore shunting each other.Outlet capillary preferably for example, is separated from each other by a kind of non-porous material (plastics).In this way, the capillary only separating along these guides this stream.Preferably, sieve plate is provided in these outlet capillaries.In this type of outlet preferred arrangement capillaceous, the first capillary of a radial center can for example, by a plurality of the second capillaries (capillary bundling, the capillary of these bindings can be the capillary of the binding that arranges with a kind of annular) surround, preferably annular is surrounded, the wherein first of the first capillary guiding eluent stream of this radial center and the second portion that these a plurality of the second capillary Yu Gai firsts guide eluent stream dividually.

Preferably, column inlet and outlet are positioned at one end of post separately, are positioned at the end opposite of post.The post using preferably has a flow distributor (rate of discharge distributor) at its port of export place.In the passage that this rate of discharge distributor is preferably configured to separate therein, carry at least the first and second parts of eluent stream, first carries in the one or more passages that separate with second portion etc.Thereby flow distributor is a distributor that is effective to eluent stream.This rate of discharge distributor may be provided in a styletable joint, is in use installed to releasedly a dismountable end connector on column outlet end.Alternately, can manufacture this rate of discharge distributor with the end integration of post.One preferred, and to arrange be to make this rate of discharge distributor as the end connector part separating being in use installed on the end of post.Yet, will be appreciated that, in other embodiments, flow distributor can be integrated and manufacture to carry at least the first and second parts of eluent stream with the post with a plurality of passages that separate is possible.In the embodiment of this type of integration, this flow distributor is not a part separating.At this, although the preferred embodiment of an end connector separating will be mainly used to illustrate this flow distributor, a kind of like this feature of end connector is also generally used for this flow distributor and post end is integrated situation about manufacturing.

The flow distributor in exit is configured to wherein have a plurality of passages that separate separately to carry at least the first and second parts of eluent stream, for example, as make at least the first and second of this eluent stream shunt in the downstream in exit by described shunting sieve plate assembly, or this flow distributor can be configured to make eluent stream to be split into part separately individually, for example, wherein in exit, used (non-shunting) sieve plate of a standard.Thereby this flow distributor has a plurality of passages that separate at least the first and second parts of eluent are flowed into.Therefore, eluent can be split into a plurality of parts of separating, and these parts of separating are assigned in different treating apparatus.As described above, this flow distributor is preferably provided in an end connector, and wherein this end connector comprises a plurality of passages that separate.

The passage separating of rate of discharge distributor (for example end connector) is preferably arranged to carry a plurality of parts of eluent stream, and a plurality of parts of these eluent stream flow out from the zones of different of post (more preferably as described the different radial zones of post).The passage separating of flow distributor can comprise have at least one passage (a preferably first passage) one first group, and this first group is arranged in this distributor so that in use its first area in post, the first radial zone preferably.For example, in the situation that the flow distributor in a kind of end connector form, first group of at least one passage of having of this end connector is arranged in this joint, makes like this when it is installed on the port of export of post this first group first area in post, the first radial zone preferably.This first radial zone is Zhu center radial zone preferably, is more preferably located substantially on a central axis that runs through post, and has in this case first group of at least one passage referred to here as a central passage group.First or central passage group carry the Yi Ge first of eluent stream.First group (for example, central passage group) with at least one passage preferably shunts sieve plate assembly Yi Ge center sieve section radially aligned with one, wherein adopted a shunting sieve plate assembly.The passage separating of rate of discharge distributor can comprise have at least one passage (preferably a plurality of passages) one second group, and this second group is arranged in this distributor so that in use its second area in post, the second radial zone preferably.For example, an in the situation that end in a kind of end connector form, second group of at least one passage (preferably a plurality of passages) of having of this end connector is arranged in this joint, make like this when it is installed on the port of export of post, there is a second area in post of second group of at least one passage, the second radial zone preferably.This second radial zone is preferably positioned at center radial zone radially outward or a radial zone at peripheral place, and has in this case one second group passage group that is referred to herein as an outside or periphery of at least one passage.The second outside or peripheral passage group is carried a second portion of eluent stream.Second group (for example, outside or peripheral passage group) with at least one passage preferably with outside of a shunting sieve plate assembly or peripheral sieve section radially aligned, wherein adopted a shunting sieve plate assembly.In other embodiments, in flow distributor, can comprise and there is one the 3rd group of one or more passages and other group optionally, for example wherein the third part of eluent and optionally other part be separated for processing.In a plurality of preferred embodiments, for guiding the first passage group of the Yi Ge first of eluent to comprise a radial center passage, and second group is included in a plurality of external channels that this central passage is radially outward located.Yet, will be appreciated that, in a plurality of embodiment, this first group can comprise a plurality of central passages (being Yi Ge center radial zone) and at first group with a plurality of passages a plurality of external channels of radially outward locating.In such cases, can with from the flow point of a plurality of external channels, turn up the soil together with the stream aggregation from a plurality of central passages and assign to process as a First, from the stream of a plurality of external channels, can be collected at together and process as a second portion.

Preferably, flow distributor is arranged to very close sieve plate assembly or most preferably contacts with sieve plate assembly, make like this to flow out from the different radial zones of post bed, the group through these parts of the eluents of this sieve plate assembly (preferably shunting sieve plate) through the corresponding passage this distributor, for example through the first and second parts of the eluent of this sieve plate assembly, enter accordingly first group and second group of passage in this flow distributor.By arranging this flow distributor in the direction contacting with this sieve plate assembly, the possibility that makes to introduce space is very little.The flow distributor using can be set level against sieve plate surface.The flow distributor using can contact and place with the one or more non-porous part of sieve plate assembly so that these one or more non-porous parts for example, at sieve plate (sieve section) thus and between flow distributor, provide sealing to seal the neighbouring part of the eluent stream from each other.For example, these parts that non-porous outside sieve plate joint and/or non-porous flow barrier (this non-porous flow barrier make porous sieve plate section separately) thereby can seal against this flow distributor keeps eluent separately.

Through the passage of rate of discharge distributor (preferably end connector) preferably each comfortable its downstream end there is an outlet or outlet port, outlet pipeline can be connected on this outlet or outlet port so that eluent is for example downloaded in treating apparatus.The number of passage can maybe can be not equal to the number of outlet port, and for example in certain embodiments, any two or more passages may be incorporated in flow distributor to share an outlet port.Yet preferably the number of passage equals the number of outlet port.

Preferably, outside or peripheral channel and their port are arranged in around the central axis of post symmetrically.For example, these outsides or peripheral channel and their port can be divided equally spaced apart and/or equidistant spaced apart with central passage and port.Yet these outsides or peripheral channel and their port can be arranged asymmetrically.

Preferably, rate of discharge distributor comprises a central passage and from 2 to 12 external channels (i.e. 2,3,4,5,6,7,8,9,10,11 or 12 external channels), more preferably a central passage and from 3 to 6 external channels.The rate of discharge distributor with 3,4,5 or 6 external channels is good examples.Yet these numbers do not limit the present invention.

As a plurality of preferred examples, in first preferred embodiment, rate of discharge distributor (preferably end connector) comprises a central passage and three external channels (i.e. four-way or a port arrangement).In second preferred embodiment, flow distributor comprises a central passage and six external channels (i.e. the 7th passage or port arrangement).Can change the number of passage and the number of port, for example can use four ports, five ports, six ports, seven ports, eight ports, nine ports, ten ports, 11 ports or 12 port arrangement, or in fact can use and there is the even more configuration of high port number.

About the number of center outlet port and the number of peripheral outlet port, at above mentioned the first preferred embodiment middle outlet flow distributor (preferably end connector), can there are in center Yi Ge center outlet port and its three peripheral outlet ports of encirclement, but should be understood that, the present invention expects the peripheral outlet port of any number, for example one or more peripheral outlet ports.A plurality of preferred examples can have from 3 to 12, more preferably 3 to 10 peripheral outlet ports, particularly 3,4,5,6,7 or 8 peripheral outlet ports.The flow distributor (preferably end connector) with 3,4,5 or 6 peripheral outlet ports is good examples.In addition, the present invention expects the center outlet port (transmitting those ports from an eluent stream of Yi Ge center radial zone) of any number, for example one or more centers outlet port.Preferably, there is Yi Ge center outlet port.These outlet ports can be positioned at end or the side of the body of flow distributor, preferably end conventionally.Outside outlet port or a plurality of port can be positioned at end or the side of flow distributor.Center outlet port or a plurality of port can be positioned at end or the side of flow distributor, but end preferably.In certain embodiments, can assemble the part of the stream flowing out from the outer peripheral areas of post and for example guide through the outside outlet port or a plurality of port that are positioned at these sides of flow distributor from the exterior lateral sides of sieve plate.Can assemble the part of the stream flowing out from the central area of post and for example guide through the end Yi Ge center outlet port or a plurality of port that are positioned at flow distributor from the center of sieve plate.

Selecting the number of the one or more outlet port of each part of guiding eluent and size can be for changing a kind of means of ratio (being dividing degree) of corresponding volume of these parts of eluent stream.Alternately maybe can also change the ratio (dividing degree) from the corresponding volume of these parts of the eluent stream of outlet by adjusting along the pressure in the downstream passage of these outlet ports (being the outlet pressure of difference).For example can be by changing the outlet pressure that changes this difference along the length of pipeline or the diameter of pipeline of these outlet ports.

In use, can seal one or more in (being shutoff) these outlet ports, so that eluent can be from wherein flowing through, but cause on the contrary and flow through the remaining port of opening.

A part of the stream of eluent comprises from outlet and assembles and deliver to all eluents in a concrete treating apparatus by a concrete path.A part of eluent thereby for example can comprise, from one or more ports (a plurality of peripheral ports of flow distributor) and assemble and intensively deliver to an eluent concrete treating apparatus.

Port of export joint can have and the similar external dimensions of conventional end joint.At column outlet place, this end connector can or by manually fastening or be fastened on the end of post by means of an instrument if desired.Port of export joint preferably connects on the port of export that is installed to post by spiral, maybe can be pushed into installation, maybe can connect with the connection of another kind of type.For the end connector of the many general types for analytical column (for example, for HPLC), for a kind of typical connection of the end connector on this post, be included in external screw thread on the port of export of this post and at an internal thread of this end connector inside.In this type of arranges, this end connector thereby be screwed on the end of post and cover outlet.In other embodiments, port of export joint can be arranged in post end by inside, for example for some type from packed column and axial compression column.In this type of embodiment, end connector can be pushed into (friction) and be installed in post end, and optionally can on its outer surface, with a sealing device (as one or more sealing rings or O type ring), with the inner surface against post jamb, seal.Port of export joint can be made by any applicable material.End connector can be made by metal (preferably stainless steel), is especially preferably passing through screw thread or is passing through to use a big Lip river gram of generation ^tM(Swagelok ^tM) type fittings is installed to it for example, in the situation of a metal column (stainless steel column).In other cases, for example, in the situation that post is glass, this end connector can for example, be made by other applicable materials (plastics, such as PEEK).

Will be appreciated that, an important feature of the present invention is on post, to carry out separating of eluent stream, is not as conventional method, after post, to carry out separating of eluent stream.This realizes by feature of the present invention described here (as shunting sieve plate assembly and flow distributor).

Column outlet can make the eluent stream of leaving this post be split into more than two parts, for example three, four or more part.Preferably, this outlet makes eluent stream be split into two parts, and this provides the low cost of the present invention as being described in more detail at this, succinct and favourable embodiment.

Can by eluent guiding device to treating apparatus further guiding leave these splitter sections of the eluent of this outlet, in the downstream of flow distributor.Preferably, for these parts of guiding eluent to the eluent guiding device for the treatment of apparatus, at the stream of eluent, in exit, for example, by (after flow distributor) after shunting, received the stream of this eluent, and this eluent guiding device typically comprises a plurality of conduits, for example pipeline.In use eluent flow through each outlet port of flow distributor preferably there is a conduit attached with it.Preferably, in the passage separating or conduit, guide at least two parts of separating of eluent to the treating apparatus separating of their correspondences.Applicable pipeline for this purpose for example can comprise, for any standard of liquid chromatography or conventional pipeline, plastic lined piping (preferably PEEK pipeline) or metal pipe line (preferably steel pipeline).Each part in eluent stream preferably guides in a corresponding treating apparatus through the group with one or more conduits of himself.

Preferably, entrance and exit is positioned at one end of post separately, and entrance and exit is positioned at the opposite end of post.This entrance is configured to a stream of mobile phase to be incorporated in the post that is loaded with a kind of sample, thereby when this sample longitudinally advances through post by this mobile phase from entrance to outlet, this sample separation becomes a plurality of components.This entrance can be a conventional entrance for column chromatography.In other words, can only by the port of export place at post, carry out modification as the described herein and implement the present invention.Can be for example or via single ingress port or via a plurality of ingress ports (as an entrance or there is the head flow distributor of a plurality of ingress ports, these ingress ports are spaced apart on the width of post to distribute more fifty-fifty the flow that traverses post width) and traverse post width porch or for example by central point, inject (CPI) and at a restricted radial zone place of post, or use a kind of heavy curtain stream method at a restricted radial zone place of post, sample to be incorporated in post, this heavy curtain stream method realizes by adopting a shunting at column inlet place or cutting apart streaming joint (being similar to port of export joint).Therefore in such cases, can preferably introduce sample (with relatively larger amount) via one or more ports, wherein other ports are by a small amount of sample or do not have sample to be incorporated in mobile phase.Therefore the part that, is introduced into the mobile phase that preferably is contained in sample wherein can flow through post to be introduced into a kind of radially restricted mode of the part restriction of the mobile phase that contains a small amount of sample or there is no sample.

About being permitted eurypalynous conventional LC, can in post, use in porch a joint.

In some preferred embodiment, this entrance is configured to independent at least two controlled parts of separating, the stream of mobile phase is incorporated in post, and these parts are incorporated in the different radial zones of post, make like this these parts longitudinal flow in different radial zones pass this post.Preferably, the flow velocity of the sample concentration in each part or each part or the two are independent controlled.The composition of mobile phase part can be controlled.Therefore, these mobile phase parts can have identical or different composition, for example identical or different solvent.A part (a for example radial outer periphery part) can or even a kind of non-solvent or at least there is more a kind of solvent of low solubility (for example water) for sample to be separated, thereby promote sample for example, comprising in other (radial center) parts.

Preferably, the sample concentration in each part is independent controlled, thereby will have sample to be separated to be included in of these parts with the concentration than higher in other one or more parts.

Preferably configuration outlet, makes like this by means of partition type entrance to flow in zones of different and through these mobile phase parts of post, shunts each other to provide in exit the part of separating of eluent.Thereby, the eluent that is separated into two or more parts in exit can comprise a kind of eluent part, this eluent partly comprise contain the sample of a large amount relatively more (or substantially all sample) flow through in the mobile phase part of post at least some or all.Thereby, the eluent that is separated into two or more parts in exit can comprise a kind of eluent part, this eluent partly comprise the relative lower amount of containing sample (or there is no sample) flow through in the mobile phase part of post at least some or all.For example can recycle this part.

Preferably, entrance is configured to at least two parts of separating, a stream of mobile phase is incorporated in post: a second portion of the Yi Ge first of this stream and this stream at least.Preferably, first mobile phase partly comprises the sample higher than this second portion concentration.Thereby in such cases, the sample concentration of each part is independent controlled.In simple situation, this can be by arranging sample to inject valve sample is injected into a part (Yi Ge first) rather than other the one or more parts with controlled amounts, for example by before column inlet only have a part flow through pipeline on arrange this sample valve to realize.Other one or more parts can not have separately sample and inject valve on their pipeline, before column inlet, this part flows through their pipeline, make like this other parts only contain solvent, or can on their pipeline, there is a sample injection valve separating for the sample of difference amount (comprising the situation that there is no sample) is incorporated into the sample that is injected into this first.

Therefore, method provided by the invention can comprise: for an entrance of chromatographic column provides at least two of mobile phase parts of separating, at least one in these at least two parts of separating comprises the sample that is separated into a plurality of components than higher the needing of other one or more part concentration; These at least two parts of separating of mobile phase are incorporated in the different radial zones of this post; An outlet in these different radial zones, from the entrance of this post to this post makes these parts of mobile phase longitudinally through this post, so that sample separation becomes a plurality of components.Preferably, the method comprises by the stream of other one or more parts and controlling having the sample horizontal proliferation of the part of higher sample concentration.

Have sample to be separated preferably at least therein these parts be introduced in porch in post to be included in than higher concentration in other one or more parts in of these parts.Be included in than the sample in a higher mobile phase part of other one or more part concentration and comprise preferred situation, wherein preferably at least in these parts, be introduced in porch in post and more preferably also in exit, the sample of all (substantially all) is comprised in a mobile phase part and other one or more parts do not comprise (substantially not comprising) any sample (being that other one or more parts are only solvents).Can to thering is the sample horizontal proliferation of the part of higher sample concentration, limit by other one or more parts.

Thereby in this type of preferred embodiment, the present invention allows sample to be introduced in the entrance of post, be preferably included in part of mobile phase and have in to be introduced and the part through the restricted radial zone (preferably central area) of post.The such part that contains higher sample concentration is called again sample stream at this.Then can pass through one or more other parts of mobile phase, preferably the mobile wall through post or outer peripheral areas (annular is surrounded central area) provide a so-called heavy curtain stream, this heavy curtain flow restriction the lateral transfer of sample (i.e. diffusion), for example to wall, move.In some preferred embodiment, this heavy curtain stream is to be in the sample stream form of an endless belt of mobile mobile phase around, and this sample stream is the form in mobile phase Yi Ge center band.When in exit together with shunting or when cutting apart stream and using, draw the sample of post thereby be concentrated and draw in a concrete radial zone (preferably central area).Therefore, can by sample is comprised and is concentrated in post a restricted region (central area preferably because this central area typically in packed column the region of uniform filling and wherein sample separation efficiency is also the highest typically) in strengthen the detection of sample.Therefore, this type of embodiment plays and through the sample of chromatographic column, is included in the interior effect in a localized area (preferably central cylindrical area) of post bed by flowing in itself, to strengthen the detection of the component eluting from this post.Thereby these embodiment play the effect that reduces the sample horizontal proliferation in this post when sample longitudinally advances through post.Thereby can be by sample being included in to the improvement that realizes the signal to noise ratio (S/N) of chromatographic peak in the eluent of small size more.The sample being advantageously used in a stream that prevents the mobile phase in the radial zone of post Yi Ge center at the wall of post or a stream of the mobile phase in peripheral radial zone moves on this post jamb.In this way, the collection of the core of mobile phase eluent and/or detection can cause the sample up to 100% effectively to use (only 0% sample is wasted in periphery) individually, wherein substantially can limit this sample by function of the present invention.In this type of embodiment, one more the post of narrow diameter be effectively used for separating of and analyze, but the flow velocity of the pressure drop of wider boring post, this wider boring bed is higher and the extra dead volume advantage with more volume bed.

In an additional advantage of this type of embodiment, can control the linear flow rate of different mobile phase parts.Can or by using pump separately (using a pump separating for each part) for these parts or by other devices, for example, by flow restrictor being placed in the pipeline that is loaded with each part, (so that being loaded with between the pipeline of each part, for example, between a plurality of ambient fluid pipelines and a core flow pipeline, provide different pressure) carrys out coutroi velocity.

Flow velocity can be arranged to substantially the same.In having the conventional post of a single ingress port, sample stream concentrates on the central area of post bed, in any case wherein flowing velocity is higher naturally.This causes a bowl-shape sample band, and wherein sample moves faster and sample in periphery or wall region is moved slower in central area.The embodiment of these descriptions of the present invention can overcome or reduce this defect by the stream arrangement of separating for different mobile phase parts (as a center flow part and a mobile part in other periphery), wherein the flow velocity of these streams can be arranged to substantially the same, thereby a kind of flow velocity profile is provided on cross section, and this flow velocity profile is smooth or at least than obviously more smooth under regular situation substantially.Smooth flow profile can make peak be delivered to outlet and thereby improve resolution ratio in narrower time bands of a spectrum (be peak width narrower) in chromatogram.

Have been found that sample may not contact wall when application is only this heavy curtain stream of solvent.Have been found that the sample up to 100% can be arranged to elute from the central area of post, and be low to moderate the from then on peripheral wash-out in region of 0% sample.Thereby, use heavy curtain stream, can exist from central area and be low to moderate and there is no sample loss, and can exist to the solvent for peripheral heavy curtain stream up to 100% recovery substantially.Therefore, sample can be arranged to as a plug through Zhu center wash-out.This provides one of major advantage of heavy curtain stream, this advantage is that sample concentration increases, and the treating apparatus by concentrated sample being separated from center regional guidance to via a central outlet with lower volume flow (preferably a sample collection device or detector) utilizes this advantage.Can recycle for the solvent of the heavy curtain stream part of mobile phase up to 100% recovery, thereby produce numerous environmental benefit.

Mobile phase is in a plurality of embodiment in the different radial zones that two or more parts of separating are introduced in post therein, entrance is preferably configured to a stream of mobile phase to be incorporated in post and to make like this this mobile phase be provided at least two parts of separating, and wherein each part preferably for example has the group of one or more introduction passages and separately introduces (this entrance is referred to here as partition type entrance) via it.In some preferred embodiment, a part of mobile phase flows to and through this post from another part in a zones of different of post, and more preferably with other part separate detection or collection.

Preferably, partition type entrance is configured to at least two parts of separating, a stream of mobile phase is incorporated in post: a second portion of the Yi Ge first of this stream and this stream at least.Preferably, first mobile phase partly comprises the sample higher than this second portion concentration.The first and second parts of eluent stream preferably flow through the effluent of correspondence first and second parts of the mobile phase of post.Similarly, from wherein flow out the first and second regions of the first and second parts of eluent stream be accordingly preferably the first and second parts of mobile phase flow through the first and second regions of post.

Preferably, use partition type entrance, at least two parts of separating of mobile phase are introduced in the zones of different of post, and for example first be directed in a first area and second portion be directed in a second area of post (post bed more specifically).More preferably, these at least two parts of separating be directed in the different radial zones of post, and for example this first be directed in first radial zone and this second portion be directed in second radial zone of the post that is different from this first radial zone.Preferably, the first area of post is the radial zone being located substantially on away from the wall of this post.Preferably, this post is the packed column therein with a post bed, and a first area of this packed column is a radial zone, and through this radial zone, mobile phase is longitudinally through the part of the uniform filling of this post bed.Also more preferably, the first of mobile phase introduces and flows through post Yi Ge center radial zone, and a second portion radial zone introducing and flow and radially outward locate through being positioned at this center radial zone, i.e. an outer peripheral areas.Most preferably, Zhu center radial zone is the region being located substantially on a central axis, and this central axis longitudinally runs through this post from entrance to outlet.In this way, can a centronucleus that preferably contains the mobile phase stream of higher sample concentration and the better component of differentiating be guided in a detector or other treating apparatus as a part via outlet, and via outlet, for example remainder as this stream of one or more other parts is guided to other places, for example guide in one or more different treating apparatus.Thereby, be preferably different from the remainder of stream and separate and the first that processes this stream with the remainder of stream.In certain embodiments, entrance is arranged to more than two parts, the stream of mobile phase to be incorporated in post.For example, in this type of embodiment, this entrance can be arranged to three or more parts of separating, this stream is incorporated in post.

As mentioned above, fill the concentration of the mobile example band in LC post along with the distance from post central axis increases and reduces, part due in column diameter and traverse column diameter inhomogeneous flow and part due to fill bed in and fill diffusion that bed transporting be around associated and relevant mass transfer effect, the validity that the density of this filling bed self and uniformity can be filled with coupled columns changes.The sample band that this moves also trends towards having parabolic shape.These phenomenons can cause the shortcoming about sample separation efficiency, detection and resolution ratio, because conventional column outlet makes to assemble from the eluent that traverses the whole diameter of this post.Yet the stream that partition type entrance can make to enter the mobile phase of post is cut apart so that part of this stream is separated with another part at least, and these parts can flow along post substantially in a parallel manner.For example, a part of the mobile phase of the sample that contains relative higher concentration can be selected and flows through the central area of post and advantageously separately guided at a detector place, so that a kind of detection of enhancing is provided, wherein peak is also differentiated better.That is to say, by the component band in the sample of chromatography, in the centronucleus of post, become and axially scatter still less, so that due to the adjacent component detecting in sample, the peak in chromatogram becomes and separates better or differentiate.In preferred embodiment, detect the part that therefore only concentrates on the stream that the centronucleus (being center radial zone) from post elutes, because the part of this stream has than the radially outer regional concentration of more close post jamb higher sample relatively.Thereby the outlet of post is preferably configured to make the diverting flow that leaves the eluent of post to become different parts, these different parts flow out from the zones of different of post, and these different parts are separately processed.

Therefore, partition type entrance preferably provides a stream of mobile phase (more preferably only having solvent), referred to here as heavy curtain stream, thereby the part that this heavy curtain stream is flowed through the mobile phase of post by the region that is preferably introduced in a radially outer of post in a radially outer region of post provides.This heavy curtain stream preferably with for example contain core (referred to here as sample stream) relative to the mobile phase of higher concentration sample (more preferably containing all samples) comparable, more preferably under substantially the same linear speed, move, thereby this sample stream is preferably introduced in a radial center region of post, in a radial center region of post, flow through post.Thereby this speeds match provides the tolerance of the sample horizontal proliferation of sample stream in coupled columns.In this way, the mobile mobile phase in the mobile phase part that maintains a center flow and periphery partly separates.

Therefore, in this type of embodiment, this equipment is preferably arranged for the stream separating of different mobile phase parts (as a center flow part and the peripheral part that flows of another one), wherein their flow velocity can be arranged to substantially the same, thereby a kind of flow velocity profile is provided on cross section, and this flow velocity profile is smooth or than much smooth under regular situation substantially.Smooth flow profile also makes peak be delivered to outlet in narrower time bands of a spectrum (be peak width narrower) in chromatogram.In certain embodiments, with respect to outer peripheral areas, in central area, there is lower flow velocity.

Along with mobile phase flows to the outlet that reaches post, eluent is preferably to avoid a large amount of a kind of like this modes of mixing of these parts of mobile phase stream separated.In the exit of post, eluent is preferably shunted, and makes like this heavy curtain stream and sample stream keep separating.In this way, for example, can guide in a detector containing relatively higher sample concentration Yi Ge center stream, and a circumferential flux (heavy curtain stream) is guided to other places.This can strengthen by the detection lower limit of the component of chromatography and improve the peak capacity in chromatographic determination, has wherein realized and has improved the benefit of measuring performance.

Can use change the to flow ratio of corresponding volume of different piece of stream of partition type entrance, thus stream to cut apart can be adjustable.Optimum flow ratio typically depends on carried out specific experiment.In addition, can consider the environmental pressure of the method and change through the amount of stream of an outer peripheral areas and the amount of the stream of a central area through the post that contains sample that contain seldom or do not have sample.The present invention provides a kind of adjustable system that can weigh economic benefits and environmental benefit valuably.

Can change the flow-rate ratio of using partition type entrance by following various in greater detail means.Preferably, mobile phase part be total mobile phase 70% or still less (by volume) 50% or still less, more preferably 30% or still less.For example, this part can be total mobile phase 50% or still less, 45% or still less, 40% or still less, 35% or still less, 30% or still less, 25% or still less, 20% or still less, 15% or still less, 10% or still less or 5% or still less.This part preferably flows through a part of post Yi Ge center radial zone.

Can by select different sieve section area ratio and/or by selecting as follows, at this, the number of ingress port and/or size change the ratio through the corresponding volume of the different piece of the mobile phase stream of partition type entrance in greater detail.

This partition type entrance is preferably equipped with an entrance sieve plate, this sieve plate is wherein set and makes like this mobile phase that enters post flow through this sieve plate.Preferably, within this entrance sieve plate is positioned at the column internal diameter of porch.

This entrance sieve plate preferably in an entrance sieve plate assembly, this entrance sieve plate assembly be configured to when mobile phase flow to into post time be separated at least two parts of separating.In order to make this diverting flow, preferably this entrance sieve plate assembly is a shunting sieve plate assembly (being called again partition type sieve plate assembly at this), this shunting sieve plate assembly comprises at least two sieve section of separating, these at least two sieve section of separating are separated from each other by one or more flow barriers, for example a kind of non-porous body can provide flow barrier, or a kind of non-porous coating can provide flow barrier, for example wherein this kind of coating is provided on one or more surfaces of at least one sieve section of separating, these one or more coated surfaces are in abutting connection with other sieve section or a plurality of sections.Thereby, flow the first of mobile phase stream can be provided through the mobile phase of one or more the first sieve section, and flow the second portion of mobile phase stream can be provided through the mobile phase of one or more the second sieve section, because the mobile mobile phase through the one or more the first sieve section separates with the mobile phase flowing through the one or more the second sieve section by the fluid barriers in non-porous body form.When mobile phase passes sieve plate assembly, flow barrier has stoped horizontal (radially) of the mobile phase between these sieve section to flow, thereby can make stream be isolated into part separately.Because these sections of shunting sieve plate have occupied the zones of different (especially different radial zones) of post, so having determined at least two of mobile phase parts of separating, this shunting sieve plate is introduced in the zones of different of post (preferably as described different radial zones).On the other hand, a kind of single sieve plate sheet of routine is that efficiency is lower, because be not so orderly and therefore when mobile phase is mobile, it may become therein and is mixed to undesirable degree during through this sieve plate through the stream of the mobile phase of this sieve plate.

A kind of preferred configuration of entrance shunting sieve plate assembly comprises: at least one center sieve section, a non-porous body and at least one outside sieve section, this non-porous body surrounds this at least one center sieve section, and this at least one outside sieve section is surrounded this at least one center sieve section but separated therewith by this non-porous body.More preferably, this shunting sieve plate comprises: the center sieve section of a radial center, a non-porous body and an outside sieve section, and this non-porous body surrounds this center sieve section annularly, and this outside sieve section is surrounded this non-porous body annularly.Most preferably, the center sieve section of this radial center is located substantially on from entrance to outlet and longitudinally runs through the central axis of post.This type of shunting sieve plate configuration allows that the one or more center sieve section produces a part of the stream of post Yi Ge center radial zone, and the one or more outside sieve section produces a part of the stream that is positioned at this center radial zone radial zone that radially outward (periphery) located.

This center sieve section and outside sieve section can have different relative areas, thereby mobile phase diverting flow is become from having the central area of different relative areas and a plurality of parts of outer peripheral areas.The ratio of the area of these sieve section can because of but for changing a kind of means of ratio of corresponding volume of these parts of mobile phase stream.For example, the ratio of the area of the area of outside sieve section and center sieve section can be from 90%:10% to 50%:50% for example, more typically from 80%:20% to 50%:50%, change, but also can use the ratio outside these scopes.A preferred ratio of the outside area of sieve section and the area of center sieve section be from about 6:1 to about 1:1, more preferably from about 3:1 to about 1:1, also more preferably from about 2.5:1 about 1.5:1 extremely, and be most preferably about 2:1.

These sieve section can have identical or different density.For example, this center sieve section can have the density different from this outside sieve section.In the embodiment of a type, this center sieve section can have the density lower than this outside sieve section.Thereby, can control mobile phase preferably to flow through the sieve section with less dense for thering is a more highdensity sieve section.

Entrance sieve plate assembly typically comprises a non-porous joint in outside (preferably being made by polymer), for example, make the non-porous jiont treatment in this outside to the arrival end of post so that this sieve plate assembly forms a sieve plate cap.A kind of like this external lug is preferred, because for example a kind of steel sieve plate will can not seal against steel column wall well.Polymeric joint can for example, be made by different polymer (PTFE, ETFE, PEEK or Kel-F, more preferably PEEK).Conventionally, any non-porous part of this sieve plate assembly can for example, be made by plastics or polymer (PTFE, ETFE, PEEK or Kel-F, more preferably PEEK), or can for example, by metal (stainless steel), be made.

In some other embodiment, entrance sieve plate assembly can comprise a single-piece porous sieve plate, is not a plurality of sections of sieve plate.As in other described embodiment, this single-piece porous sieve plate can be accommodated in an outside joint non-porous, preferably polymer, for example, make this jiont treatment to the outlet of post so that sieve plate assembly forms a sieve plate cap.

The non-porous joint in this outside can have a plurality of openings to allow separately to introduce the stream of mobile phase.For example, the opening that the non-porous joint in this outside can have a radial center is worn to a radial center region of post through the stream of a part of the mobile phase of sieve plate allowing, and can have one or more peripheral openings of radially outward locating at this central opening and through the stream of a part of the mobile phase of sieve plate, wear in the peripheral radial zone (surrounding this radial center region) to post allowing.Can for example by thering are one or more peripheral openings in the sidewall at this non-porous joint in outside, the part of mobile phase stream be incorporated into the outer peripheral areas of post from the outside of sieve plate.

Entrance sieve plate assembly preferably has circular outer shape with the circular cross-section post that coincide, although can for example depend on that post shapes use has the sieve plate assembly of other shapes.

The material of entrance sieve plate can be the conventional sieve plate material using in LC, for example steel.Thereby this sieve plate can be become to make to flow by easy configuration with shunting mode described here and shunt through its eluent stream.For example, sieve plate material, thickness (degree of depth) and porosity can be that routine is used in LC system.For example, can use the sieve plate with 0.25mm to 2mm typical thickness.For example, can use the sieve plate with 2 μ m porositys.The non-porous body of shunting sieve plate embodiment preferably for example, is made by a kind of plastics or polymer (PTFE, ETFE, PEEK or Kel-F, more preferably PEEK), or can for example, by metal (, stainless steel), be made.This type of non-porous material can also be provided as an one or more lip-deep thin layer or the coating in one or more sieve section, the one or more sieve section in abutting connection with another sieve section so that a flow barrier to be provided.Non-porous flow barrier alternatively can be made by a kind of metal.A kind of like this metallic barrier can an one or more lip-deep thin layer or the coating as the one or more sieve section in abutting connection with another sieve section form by sputter.This type of thin layer or coating flow barrier can convection current have advantages of lower resistance.

Compare with the width of sieve section, the width of flow barrier or non-porous body (in the radial direction measure) is preferably little, is preferably lower than the width of these sieve section.Thereby, thereby can in stream, there is flow barrier and any dead time effect to after any possible resistance of the stream of the separative component of tool or flow barrier that causes can be decreased to minimum.Yet, will be appreciated that, this barrier should not be so thin so that can not effectively realize and make stream be isolated into part separately.

The partition type entrance using preferably has an inlet flow rate distributor at its arrival end, for example, so that Yi Ge center stream produces heavy curtain fluidised form around.The structure of this inlet flow rate distributor is preferably similar to rate of discharge distributor described above.At least the first and second parts that in the passage that this inlet flow rate distributor is preferably configured to separate therein, transport flow flows mutually, first carries in the one or more passages that separate with second portion etc.Thereby this inlet flow rate distributor is a distributor that is effective to mobile phase stream.This inlet flow rate distributor may be provided in a styletable joint, is in use installed to releasedly a dismountable end connector on column outlet end.Alternately, can manufacture this inlet flow rate distributor with the arrival end integration of post.One preferred, and to arrange be to make this inlet flow rate distributor as the end connector part separating being in use installed on the end of post.Yet, will be appreciated that, in other embodiments, flow distributor can be integrated and manufacture at least the first and second parts that flow mutually with transport flow with the post with a plurality of passages that separate is possible.In the embodiment of this type of integration, this flow distributor is not a part separating.At this, although the preferred embodiment of an end connector separating will be mainly used to illustrate inlet flow rate distributor, a kind of like this feature of end connector is also generally used for flow distributor and post end is integrated situation about manufacturing.

This inlet flow rate distributor is configured to have therein at least the first and second parts that a plurality of passages that separate flow mutually to separate transport flow.The part that this inlet flow rate distributor can be configured to separate is individually introduced this mobile phase stream, for example, wherein in porch, used (non-shunting) sieve plate of a standard.Thereby this inlet flow rate distributor has a plurality of passages that separate at least the first and second parts of mobile phase are flowed into.Therefore, these parts of separating of mobile phase can be assigned in the zones of different of post.As described above, this flow distributor is preferably provided in an end connector, and wherein this end connector comprises a plurality of passages that separate.

The passage separating of inlet flow rate distributor (for example end connector) is preferably arranged to a plurality of parts that transport flow flows mutually, and a plurality of parts of these mobile phases stream are introduced in the zones of different of post (more preferably as described the different radial zones of post).The passage separating of flow distributor can comprise have at least one passage (a preferably first passage) one first group, and this first group is arranged in this distributor so that in use its first area in post, the first radial zone preferably.For example, an in the situation that inlet flow rate distributor in a kind of end connector form, first group of at least one passage of having of this end connector is arranged in this joint, make like this when it is installed on the arrival end of post this first group first area in post, the first radial zone preferably.This first radial zone is Zhu center radial zone preferably, is more preferably located substantially on a central axis that runs through post, and has in this case first group of at least one passage referred to here as a central inlet passage group.First or the central passage group transport flow Yi Ge first of flowing mutually.First group (for example, central passage group) with at least one passage preferably with a minute inflow entrance sieve plate assembly Yi Ge center sieve section radially aligned, wherein adopted a shunting sieve plate assembly.The passage separating of inlet flow rate distributor can comprise have at least one passage (preferably a plurality of passages) one second group, and this second group is arranged in this distributor so that in use its second area in post, the second radial zone preferably.For example, an in the situation that flow distributor in a kind of end connector form, second group of at least one passage (preferably a plurality of passages) of having of this end connector is arranged in this joint, make like this when it is installed on the arrival end of post, there is a second area in post of second group of at least one passage, the second radial zone preferably.This second radial zone is preferably positioned at center radial zone radially outward or a radial zone at peripheral place, and has in this case one the second group access road group referred to here as an outside or periphery of at least one passage.The second portion that the second outside or peripheral passage group transport flow flows mutually.Second group (for example, outside or peripheral access road group) with at least one passage preferably with outside of a minute inflow entrance sieve plate assembly or peripheral sieve section radially aligned, wherein adopted a shunting sieve plate assembly.In other embodiments, in flow distributor, can comprise and there is one the 3rd group of one or more passages and other group optionally, for example wherein the third part of mobile phase and optionally other the part of separating be introduced in post.In a plurality of preferred embodiments, for introducing the first passage group of the Yi Ge first of mobile phase, comprise a radial center passage, and second group is included in a plurality of external channels that this central passage is radially outward located.Yet, will be appreciated that, in a plurality of embodiment, this first group can comprise a plurality of central passages (being Yi Ge center radial zone) and at first group with a plurality of passages a plurality of external channels of radially outward locating.In other embodiments, can not there is not central passage.In such cases, can with from the flow point of a plurality of external channels, turn up the soil will be through assigning to process as a First together with the mobile phase stream aggregation of a plurality of central passages and finally, from the stream of a plurality of external channels, can be collected at together and process as a second portion separating.

Preferably, the passage of inlet flow rate distributor is substantially in the identical radial position of the passage with rate of discharge distributor.

Preferably, this inlet flow rate distributor is arranged to very near entrance sieve plate assembly or most preferably contacts with entrance sieve plate assembly, makes like this through these parts of the mobile phase of the group of the corresponding passage in this inlet flow rate distributor as the first and second parts of the mobile phase in the different radial zones of post accordingly through sieve plate assembly.By arranging this flow distributor in the direction contacting with this sieve plate assembly, the possibility that makes to introduce space is very little.The inlet flow rate distributor using can be set level against sieve plate surface.The flow distributor using can contact and place with the one or more non-porous part of sieve plate assembly so that these one or more non-porous parts for example, at sieve plate (sieve section) thus and sealing is provided between flow distributor the neighbouring part from mobile phase stream each other seals to porch.For example, non-porous outside sieve plate joint and/or non-porous flow barrier (this non-porous flow barrier make porous sieve plate section separately) thus can when entrance flow distributor seal these parts when mobile phase through entrance sieve plate, keep separating.

Through the passage of inlet flow rate distributor (preferably end connector), preferably locate at its upstream end separately to have an inlet port or ingress port, can by enter pipeline be connected to this enter or ingress port on so that mobile phase be downloaded in post.The number of passage can maybe can be not equal to the number of inlet port port, and for example in certain embodiments, any two or more passages in inlet flow rate distributor can be shared an inlet port port.Yet preferably the number of passage equals the number of inlet port port.

Preferably, the outside in inlet flow rate distributor or peripheral channel and their port are arranged in around the central axis of post symmetrically.For example, these outsides or peripheral channel and their port can be divided equally spaced apart and/or equidistant spaced apart with central axis/central passage and port.Yet these outsides or peripheral channel and their port can be arranged asymmetrically.

Preferably, inlet flow rate distributor comprises a central passage and from 2 to 12 external channels (i.e. 2,3,4,5,6,7,8,9,10,11 or 12 external channels), more preferably a central passage and from 3 to 6 external channels.The inlet flow rate distributor with 3,4,5 or 6 external channels is good examples.Yet these numbers do not limit the present invention.

Inlet flow rate distributor preferably has the number of the central passage identical with rate of discharge distributor described here (an especially central passage) and the number of identical external channel.

As a plurality of preferred examples, in first preferred embodiment, inlet flow rate distributor (preferably end connector) comprises a central passage and three external channels (i.e. four-way or a port arrangement).In second preferred embodiment, inlet flow rate distributor comprises a central passage and six external channels (i.e. the 7th passage or port arrangement).Can change the number of passage and the number of port, for example can use four ports, five ports, six ports, seven ports, eight ports, nine ports, ten ports, 11 ports or 12 port arrangement, or in fact can use and there is the even more configuration of high port number.

About the number of center inlet port port and the number of peripheral inlet port port, inlet flow rate distributor in above mentioned the first preferred embodiment (preferably end connector) can have in center Yi Ge center inlet port port and surround its three peripheral inlet port ports, but should be understood that, the present invention expects the peripheral inlet port port of any number, for example one or more peripheral inlet port ports.A plurality of preferred examples can have from 3 to 12, more preferably 3 to 10 peripheral inlet port ports, particularly 3,4,5,6,7 or 8 peripheral inlet port ports.The inlet flow rate distributor (preferably end connector) with 3,4,5 or 6 peripheral outlet ports is good examples.In addition, the present invention expects the center inlet port port (being about to those ports of a flow transmission Zhi Yige center radial zone) of any number, for example one or more centers inlet port port.Preferably, there is Yi Ge center inlet port port.These inlet port ports can be positioned at end or the side of the body of flow distributor, preferably end conventionally.Outside inlet port port or a plurality of port can be positioned at end or the side of flow distributor.Center inlet port port or a plurality of port can be positioned at end or the side of flow distributor, but end preferably.

Selecting the number of the one or more inlet port port of each part of guiding mobile phase and size can be for changing a kind of means of ratio (being entrance dividing degree) of the corresponding volume of these parts that mobile phase flows.Alternately maybe can also change by adjusting pressure (being the inlet pressure of difference) in access road the ratio (dividing degree) of corresponding volume of these parts of the mobile phase stream of porch.

In use, can seal one or more in (being shutoff) these inlet port ports, so that mobile phase can be from wherein flowing through, but cause on the contrary and flow through the remaining port of opening.

Arrival end joint can have and the similar external dimensions of conventional end joint.At column inlet place, this arrival end joint can or by manually fastening or be fastened on the end of post by means of an instrument if desired.End connector preferably connects on the arrival end that is installed to post by spiral, maybe can be pushed into installation, maybe can connect with the connection of another kind of type.For the end connector of the many general types for analytical column (for example, for HPLC), for a kind of typical connection of the end connector on this post, be included in external screw thread on the arrival end of this post and at an internal thread of this end connector inside.In this type of arranges, this end connector thereby be screwed on the end of post and cover entrance.In other embodiments, this end connector can be arranged in post end by inside, for example for some type from packed column and compression post axially or radially.In this type of embodiment, this end connector can be pushed into (friction) and be installed in post end, and optionally can on its outer surface, with a sealing device (as one or more sealing rings or O type ring), with the inner surface against post jamb, seal.This end connector can be made by any applicable material.This end connector can be made by metal (preferably stainless steel), is especially preferably passing through screw thread or is passing through to use a big Lip river gram of generation ^tM(Swagelok ^tM) type fittings is installed to it for example, in the situation of a metal column (stainless steel column).In other cases, for example, in the situation that post is glass, this end connector can for example, be made by other applicable materials (plastics, such as PEEK).

At this, term post means for sample being carried out to chromatographic any tubular structure.Therefore, post can be the post of a straight post or a coiling, preferably a straight post.Preferably, this post is can be with a kind of post of applicable Filled Dielectrics.It can be for example for the stake of technical grade preparative scale chromatography or for the small-sized post of the preparative scale chromatography of a small amount of sample and/or laboratory environment.It can be for the chromatographic post of analytic type.Typically, this post is the post for liquid chromatography, but can be for example for the chromatographic post of supercritical fluid (SCF).In this case, the flow path for making SCF run through equipment is suitably pressurizeed.This post can be for example for high performance liquid chromatography (HPLC), Ultra Performance Liquid Chromatography method (UHPLC), flash column chromatography, fast protein liquid chromatography (FPLC) and other forms of chromatographic post.This post can comprise a kind of capillary (as used in MECC).Advantageously, in certain embodiments, this post can be a standard (conventional) HPLC post, thereby allow user on standard column, to adopt the present invention, wherein only need to column outlet and optionally column inlet place modify, for example use sieve plate assembly and/or the flow distributor of a modification as the described herein.

Applicable post can be made by various materials as is known, comprises for example metal (preferably stainless steel), glass, pottery, polymer etc.This post can be made into fluid chip structure (the chip post of integration) micro-manufacture or that integrate.This post can be any applicable length; Preferably post has the length of in 5mm to 1000mm scope (may be longer), 50mm to 200mm for example, and for example about 100mm, especially for example, applies for analytic type (HPLC).This post can have any applicable diameter; Preferably this column internal diameter is between 300 μ m and 1000mm, and for example, for HPLC, normal inner diameter is as 4.6mm diameter.This post preferably has circular cross-section (being cross section), although can use the post of other shaped cross.

In different embodiment, the present invention can be analytic type chromatography to its useful chromatography, for example high performance liquid chromatography (HPLC), Ultra Performance Liquid Chromatography method (UHPLC), multidimensional or two-dimensional highly effective liquid phase chromatographic method (MDHPLC or 2DHPLC), flash column chromatography, fast protein liquid chromatography (FPLC), parallel detection chromatography, SCF chromatography and other chromatographys, especially HPLC.

In a plurality of embodiment, the present invention can be preparative scale chromatography to its useful chromatography, for example preparative high performance liquid chromatography (PHPLC), process chromatography, protein purification, enzyme purification, antibody purification, little molecule purifying, medical purifying or natural products purifying.

In a plurality of embodiment, the present invention can be analytic type and preparative scale chromatography to its useful liquid chromatography, for example, wherein not only for analysis purpose, detected eluent but also eluent be collected in the fraction of purifying.

About having mobile phase to be used and the fixing type of phase, can use the mobile phase of any applicable type and fixing phase (for example any applicable and/or known phase), the mobile phase of these types with fixing be to be suitable for the chromatographic type of just carrying out, for example any known HPLC mobile phase when carrying out HPLC and fixedly phase mutually.Type about operable separation method, can use any applicable conventional method, such as or etc. degree or gradient elution or displacing elution, or positive is anti-phase or hydrophilic or ion-exchange or ion exclusion or compatibility or chirality or size exclusion LC etc.

In the situation that chromatography is supercritical fluid chromatography, mobile phase can be a kind of conventional SCF, as carbon dioxide, but is not limited to this.

Use the equipment of the present invention and method of the present invention for various application, to comprise for example isolated or purified of purity analysis, component analysis, attributional analysis, quantitative analysis and AG, pilot-scale or technical grade.Market application comprises for example drug discovery, clinical analysis, environmental analysis and for example diagnostic flag research in protein-based (glycoprotein, phosphoprotein), metabolin and nucleic acid field.Therefore, the present invention for example can be applicable in medicine, chemistry, biotechnology, biological medicine and process industry.

The present invention can be applicable to packed column.At this packed column, mean to comprise the fixedly post of any applicable bed of phase.Depend on the chromatographic type of just carrying out, can using the bed Filled Dielectrics of any routine in post as post bed.Bed medium can comprise for example particle or porous whole block material (for example polymer or ceramic monolithic bed), preferably particle.Alternately, this bed medium can comprise a film bed or any other bed.The present invention is particularly useful to the bed of uneven distribution.The granular size of preferred graininess medium is for example in the scope between 1 μ m and 150 μ m, but the lower limit of granular size or the upper limit do not limit the present invention.Depend on chromatographic type, for the porous media using in the present invention, can use the bore dia of wide scope; Preferably hole size exists with

in scope between (3nm and 300nm).Can use the porosity of wide scope; Preferably porosity is in 0% to 80% scope.Depend on chromatographic type, filled media can comprise different chemical property, for example phase and the chiral stationary phase of alkyl linked phase, typical polar binding.Integral chip post can be used particle or porous monoblock bed.

Can comprise the detector for any routine of column chromatography for the treatment of the type of the detector installing, for example ultraviolet or visible (UV/Vis), mass spectrum (MS), fluorescence (FL), chemiluminescence (CL), refractive index (RI), electrical conductivity (CD), evaporative light-scattering (ELSD) detector etc.In addition, for any unconventional detector of column chromatography, can be used to treating apparatus, for example nuclear magnetic resonance (NMR) or infrared (IR) or antioxidant detector or any other biotype detector.In some cases, can use in post and detect, wherein a detector is placed in post bed so that for example this detector only detects the mobile phase from a specific radial region (especially central area).This type of detector can comprise conductivity detector.

Preferably, the Yi Ge first of the stream of eluent is guided in first treating apparatus, and a second portion of the stream of eluent is guided in second treating apparatus.Be separated from each other and process these parts, this first and second treating apparatus separates.In this way, for example, can detect dividually and there is the Yi Ge first of higher isolation resolution relatively with a second portion with relative lower isolation resolution.Yet, even if having the part of lower isolation resolution relatively also can show than routine arrangement and differentiate better peak, this routine arrangement is to detecting from traverse the eluent that the whole width of post assembles via a single central outlet port, or in other mode to focusing on from the eluent of assembling through the whole width of post.Therefore, the present invention can provide for example chromatogram of this first and/or the chromatogram of this second portion, and each chromatogram has the resolution ratio that together detects the peak that these parts are higher such as fruit.In another example, can dividually the Yi Ge first with higher isolation resolution be collected in fraction with a second portion with lower isolation resolution.Therefore, this can provide the fraction of collecting in Cong Gai first, and these are collected fraction and have such as fruit fraction is higher purity in the situation of collecting together from these parts.

Therefore, to be clear that, in yet another aspect, the present invention further provides a kind of for carrying out multiplexing chromatographic equipment, this equipment comprises: a chromatographic column, this post has an entrance and an outlet, and wherein this outlet is configured to make this diverting flow become at least two parts of separating when an eluent stream is left this post through this outlet, and wherein this equipment is configured to separately process these parts.Similarly, the present invention also provides a kind of multiplexing chromatographic method, and the method comprises: a kind of mobile phase is provided, and this mobile phase includes a kind of sample of a plurality of components of one-tenth to be separated; Make this mobile phase to an outlet longitudinal flow of post, pass a chromatographic column from an entrance of post, this mobile phase that leaves this post through this outlet is as a kind of eluent; When the stream of eluent leaves this post through this outlet, make this diverting flow become at least two parts of separating; And separately process these at least two parts of separating.These parts of separating have experienced separately the chromatographic isolation of component in post.Separately process and can comprise these parts of separate detection, for example, use different detectors to detect each part.In this way, can use different detector (each detector has the detection advantage of himself uniqueness) to carry out a plurality of parts of the identical eluent of parallel parsing.Multiplexing advantage can be to increase quantity of sample handling.Together with other aspects of the present invention, at this, described for carrying out the equipment of multiplexing liquid chromatography and the preferred feature of method.

In thering is the embodiment of partition type entrance, thereby sample is preferably all included in substantially, in the first that introduces the mobile phase in post, provide a sample stream part, and the second portion that is incorporated into the mobile phase in post do not basically contain sample (only being formed by mobile phase solvent) thus a heavy curtain stream part containing sample is provided not.In this way, for example, can detect dividually first or the sample stream part with sample with second or the heavy curtain stream part substantially therein without sample, this sample has been separated into a plurality of components along post.This first or sample stream part preferably flow through the part of the central area of post, wherein separative efficiency is the highest.This heavy curtain stream plays lateral transfer or the diffusion of restriction sample and/or plays the smooth effect of flow velocity profile making on cross section.The heavy curtain stream substantially therein without sample can be processed as a part leaving the eluent of post by reuse it in other one takes turns chromatography.In this way, realize the cost savings of solvent consumption, wherein followed environmental benefit.Therefore,, in this type of embodiment, this equipment is preferably configured to reuse part or the heavy curtain stream of the eluent flowing out from peripheral radial zone in other chromatography.

Preferably, the first treating apparatus is the treating apparatus dissimilar with the first treating apparatus.Yet, in certain embodiments, will be appreciated that, the treating apparatus that this first and second treating apparatus can be same type (for example they can comprise the detector of same type separately), a device is processed Yi Ge first and another device processing second portion, as long as these parts are separately processed.Yet preferably, this second treating apparatus is dissimilar with this first treating apparatus, because typically having different separation degrees and wish, these parts process them with diverse ways.As Multi-instance, this first and second treating apparatus (and the optionally other treating apparatus in the situation that eluent is separated into three or four parts) can comprise independently of one another with lower one or more: detector, refuse reservoir, fraction receiver or collector and a column inlet.Thereby, for the processing of each part preferably (independently) be with lower one or more: detect, collect fraction, deliver in refuse and deliver in a column inlet.Preferably, in certain embodiments, the processing of a part (for example first) comprises with other one or more parts and detects dividually this part, for example, for analytic type chromatography.Preferably, in certain embodiments, the processing of a part (for example first) comprises with other one or more parts collects fraction from this part dividually, for example, for preparative scale chromatography or for multiplexing HPLC, or a comprehensive analysis or in a heart cutting analysis.Preferably, in certain embodiments, the processing of a part (for example second portion) comprises with other one or more parts to be delivered to this part in a column inlet dividually.In certain embodiments, for provide different optionally objects in minute analysis of variance, a plurality of parts are delivered in a plurality of posts that separate.Preferably, in certain embodiments, the processing of a part (for example second portion) comprises with other one or more parts to be delivered to this part in refuse dividually.

Preferably, for example, detect dividually Yi Ge first (for example typically thering is a part of higher isolation resolution relatively) with a second portion (part typically with relative lower isolation resolution).More preferably, in this type of embodiment, this first is the part flowing out from post Yi Ge center radial zone.

In certain embodiments, can wish both to have detected the part flowing out from center radial zone, and also detect a part from outer radial region, because these two regions can show than the routine arrangement with a single central outlet port, differentiate better peak, when cutting apart stream, can use for the object of analyzing and/or prepare the stream of these two eluents.Never in any form scope of the present invention is limited, this is considered to be due to the fact that a part of the eluent of obtaining from a restricted radial zone has than the less sample axially scattering of eluent of obtaining from traversing the full duration of this post.

Preferably, the first treating apparatus comprises first detector.As an example, the first treating apparatus can comprise that a detector is present in the sample in eluent for detecting, and the second treating apparatus can comprise a refuse receiver, typically this refuse receiver does not arrive through the detector of the first treating apparatus.As another example, this first treating apparatus can comprise a detector as previously described, and this second treating apparatus can comprise entrance identical or another chromatographic column, so that second portion experience at least other is taken turns chromatography, for example, relate to the recirculation of mobile phase.This second treating apparatus can comprise a valve, this valve can be controlled and this valve is switchable between primary importance and the second place by a control system, this primary importance allows second portion to flow in entrance identical or another chromatographic column, and this second place allows second portion to flow to other places, for example, flow in a detector, fraction collector, another post or a refuse.As another example, this first treating apparatus can comprise that as in the previous a detector (first detector) and this second treating apparatus can comprise a detector (second detector), optionally wherein this second detector can provide a kind of measurement, can use this measurement (for example passing through control system) to determine whether second portion flows in entrance identical or another chromatographic column, so that second portion experience at least other is taken turns chromatography.Control system is preferably provided, and this control system preferably receives the signal from being included in the one or more detectors in the first and/or second treating apparatus.For one or more treating apparatus, this control system can be based on from treating apparatus a detector signal and the operation of one or more valves is controlled so that in a part of eluent through guiding this part extremely for example in refuse or in a column inlet after this detector to one desirable destination.Thereby, the form that this equipment can executing data dependent form be processed.

In the situation that treating apparatus comprises the entrance of a chromatographic column, this treating apparatus may further include a container or flow circuits, has wherein assembled to need the eluent of further chromatography and preferably concentrated again before further chromatography.

In these different modes, for example, the invention provides a kind of post with an outlet, this outlet is for example configured to optionally guide, in a part to one first treating apparatus (detector) of eluent stream, and another part of this outlets direct is to another treating apparatus different from this first treating apparatus simultaneously.Preferably, for the object of analyzing, this first treating apparatus comprises a detector, arranges this detector so that detect dividually first with second portion.Preferably, object for preparation, this first treating apparatus comprises a fraction receiver, arrange this fraction receiver so that receive dividually the fraction (in the situation that this first treating apparatus comprises a fraction receiver, it can also comprise a detector) of first with second portion.

Will be appreciated that, can adopt many other treating apparatus and combinations thereof, and at this, discuss some in these devices in more detail below.At this, treating apparatus comprises by a part of eluent and leaves after post and the path of obtaining, so that for example in certain embodiments these parts can be directed in identical detector, but by different paths so that these parts separately when different (for example in this case) detect (processing).Yet, in most of the cases, preferably these parts of separating after column outlet place is shunted according to distinct path.

In certain embodiments, can to the different part of the eluent of exit shunting, recombinate in the downstream of outlet.Can after having experienced processing separately, these parts recombinate to these parts.For example, can after detecting dividually with other one or more parts, a part (preferably core) recombinate to these parts.Then for example can make the one or more other chromatographic isolation of part experience of restructuring, optionally after these parts are concentrated again.

A plurality of equipment according to the present invention can be joined together, and that is to say that one (or a plurality of) part of the eluent separating in the exit of post can be fed in the entrance of an other post (according to an other equipment of the present invention).Thereby, according to the post of series connection of the present invention, can be joined together.Alternately, this other post can be a conventional post or not be another post according to post of the present invention.Therefore, generally the present invention can comprise by eluent one (or a plurality of) part flow in an other post for further chromatographic isolation.In this way, a plurality of components of sample wherein can be fed in an other post by a part (or more than one part) of the eluent of fully separated (differentiating), and experience chromatography therein so that further separated these components.The number of the post that the mode of connecting with this can connect is not concrete restriction.For example, in this series connection, two, three or more post can be linked together.Can before entering each other post or a plurality of post, injection concentrate again eluent.Guide to the part of eluent of other chromatographic isolation preferably from the outside of post or part that peripheral radial zone flows out, because a plurality of components are therein typically not as they separated must getting well the part flowing out from central area, but the part of the eluent flowing out from core can be optionally directed for further chromatographic isolation, as being situation in multidimensional or two-dimentional HPLC.

As one of above-described embodiment other variation, can be via conduit recycling elution liquid, lead back in the entrance of same column for one or more other through same column (taking turns or take turns more chromatography).One or more parts of the eluent that specifically, the exit of post can be separated are fed in the entrance of getting back to post.In this way, a plurality of components of sample wherein can be fed in post again by a part (or more than one part) of the eluent of fully separated (differentiating), and experience therein other chromatography so that further separated these components.Preferably, refill get back to post in for one or more other through concentrate eluant before post.The part of the eluent of recirculation is preferably from the outside of post or part that peripheral radial zone flows out, because a plurality of components are therein typically not as their separated must getting well the part flowing out from central area in this way.Eluent does not partly basically contain sample (being only solvent) therein, for example one of them is only that the curtain stream part of solvent flows the in the situation that of shunting with other parts through post and in exit, be only that the part of solvent can be used as mobile phase and is reused in other chromatography, thereby the overall flow that reduces equipment consume mutually.

In different embodiment, the present invention can comprise the other known tip assemblies in a chromatographic equipment.For example, the present invention may further include at least one mobile phase reservoir with the entrance supply mobile phase to post.The present invention may further include at least one pump to pass mutually post from this at least one mobile phase reservoir pumping flow.In certain embodiments, for example wherein two or more parts of separating of mobile phase are introduced into through a multiport entrance, can use two or more pumps (for example using a pump for each part).Yet, even if introduce the situation of a plurality of mobile phase parts in porch for regulating by use traffic, it is also possible using a single pump, as there is a single pump but in pipeline, there is a kind of arrangement of pressure limit, these pipelines are loaded with mobile phase to change independently the degree of flow restriction in every pipeline.The present invention may further include at least one sample injector (for example is injected valve) sample is injected into the mobile phase of the upstream of column inlet.The present invention may further include one or more pressure regulators or flow restrictor so that from the mobile equilibrium of numerous outlet ports in desirable level, these one or more pressure regulators are preferably located in the downstream of column outlet.This equipment is preferably under the control of a control and data gathering system, and this control and data gathering system for example comprise a computer and a plurality of control electronic device.This control and data gathering system be following for controlling (existing in every kind of situation of this assembly) therein preferably: for pumping flow one or more pressure regulators through the stream of outlet port through one or more pumps of post, the sample injector injecting for sample and balance mutually.This control and data gathering system are preferably for receiving and optionally process the data from one or more detectors for the treatment of apparatus.When requiring, this control and data gathering system can also provide a kind of output of data (for example have or do not have these data are processed).This control and data gathering system be other assemblies of control appliance extraly.

Accompanying drawing explanation

Fig. 1 is with a kind of conventional configuration of the schematically illustrated HPLC system of flow chart form.

The schematically illustrated axial cross section side view that passes a conventional chromatographic column of filling of Fig. 2, wherein has a kind of simple sample component wash-out.

Fig. 3 is shown with the view of Fig. 2 of four kinds of different sample component wash-outs.

Fig. 4 is schematically illustrated to be passed according to the axial cross section side view of a chromatography column of the present invention, and this view is illustrated in the principle of a flow distributor in exit.

Fig. 5 A illustrates according to the perspective view of a sieve plate assembly of the present invention preferred embodiment; Fig. 5 B illustrates the bottom view of this embodiment observing in the direction of arrow A; And Fig. 5 C illustrates the side sectional view of this embodiment that B-B along the line obtains.

Fig. 5 D illustrates according to the perspective view of another preferred embodiment of a sieve plate assembly of the present invention; And Fig. 5 E illustrates the bottom view of this embodiment.

Fig. 5 F and 5G are schematically illustrated according to the corresponding other embodiment of a plurality of sieve plate assemblies of the present invention.

Fig. 5 H and 5I are schematically illustrated according to a plurality of embodiment that arrange at a plurality of outlet capillaries at this column outlet place of the present invention.

Fig. 6 A illustrates the one embodiment of the present of invention with a shunting section sieve plate and four port end joints with decomposition view; And Fig. 6 B illustrate with Fig. 6 category-A like but there is an embodiment of a single-piece sieve plate.

Fig. 7 illustrates an other embodiment of the present invention with a shunting section sieve plate and four port end joints with decomposition view.

Fig. 8 A illustrates another the other embodiment of the present invention with a shunting section sieve plate and seven port end joints with decomposition view.

Fig. 8 B illustrates the embodiment of the assembling of Fig. 8 A that is broken away view.

Fig. 9 illustrates the cutaway side sectional view of the embodiment shown in Fig. 7, and this embodiment illustrates and be attached with outlet pipeline with the form of assembling.

Figure 10 is schematically illustrated according to an embodiment of an analytic type LC system of the present invention in a flowchart.

Figure 11 A is schematically illustrated according to a LC system of the present invention other embodiment in a flowchart, and this embodiment adopts the recirculation of a part for eluent.

Figure 11 B schematically illustrated for by mobile phase from a devices in series according to the present invention to according to a kind of system of another equipment of the present invention.

Figure 12 is illustrated in the comparison that different eluents is cut apart the single butyl benzene eluting peak reaching in a HPLC system under condition.

Figure 13 illustrates three kinds of test solute toluene, propylbenzene and butyl benzenes and at different eluents, cut apart the elution curve under condition in a HPLC system.

Figure 14 illustrates from the overlapping of Figure 13 and highly normalized butyl benzene peak.

The elution curve of the sample of toluene, propylbenzene and butyl benzene that Figure 15 illustrates central area (solid-line trace) from this post and wall region (dotted line trace) wash-out a HPLC system.

Figure 16 A is illustrated in and the sample of situation Toluene, ethylbenzene, propylbenzene and the butyl benzene cut apart without the eluent elution curve in the FPLC system of using a glass column.

Figure 16 B illustrates the zoomed-in view in a region of these elution curves shown in Figure 16 A.

Figure 17 A is illustrated in the elution curve of cutting apart a kind of four component sample of the chromatographic column of using a bad filling in situation without stream; And Figure 17 B is illustrated in stream and cuts apart the elution curve that uses the same sample of same post in situation.

Figure 18 is schematically illustrated to be passed according to the axial cross section side view of a chromatography column of the present invention, and this view illustrates the principle of an inlet flow rate distributor and a rate of discharge distributor.

Figure 19 is schematically illustrated according to another embodiment of an analytic type LC system of the present invention in a flowchart, and this embodiment has an inflow entrance of cutting apart.

Figure 20 is schematically illustrated according to another embodiment of a LC system of the present invention in a flowchart, and this embodiment has an inflow entrance of cutting apart and adopts the recirculation of a part for eluent.

Figure 21 illustrates and uses an inlet flow rate distributor at different mobile phases, to cut apart the comparison at a plurality of peaks from three kinds of solute toluene, propylbenzene and butyl benzenes that reach under condition in a HPLC system.

Figure 22 illustrates by using an inlet flow rate distributor to cut apart at different mobile phases the elution curve that under condition, the elution volume at propylbenzene peak compares in this HPLC system.

detailed description of the invention

In order further to understand the present invention, but in the situation that not limiting its scope, with reference now to accompanying drawing, different exemplary embodiments and experiment are described.

In the routine configuration (as schematically illustrated in Fig. 1 in a flowchart) of a HPLC system, one or many bottle of mobile phase solvent 2 is delivered to a solvent delivery system 6 via pipeline, and this solvent delivery system adopts a pump of pumping under high pressure or low pressure; Or by gravity, send (only low pressure) via pipeline.Solvent delivery system 6 sends desirable mobile phase solvent or its mixture (being called simply solvent at this) by a sample injection port or valve 8, wherein a kind of sample is introduced in this solvent streams and then in a chromatographic column 15, this chromatographic column is filled with a kind of fixedly phase or fixed bed or is equipped with fixedly phase of a kind of integral body.This post is a circular cross-section cylindrical pillars typically.Stream by post 15 is by a head or entrance sieve plate 9 and by post bed itself, be radially dispersed on whole width in cross section of this post bed, and then at sample, by mobile phase solvent, is carried length along this post subsequently chromatographic isolation occurs when downward.Outlet or exit at a conventional H PLC post, outwards mobile mobile phase or eluent are by one second or exit sieve plate 11 gatherings, this second or exit sieve plate typically by the end connector being installed on the port of export of this post, be retained on appropriate location so that the whole cross section of this stream is delivered to a little outlet port 40 of the center in the cross section that is positioned at this post.That is, near the material in the outer radial region of post wall, be inwardly forced to radially in case with through together with the material of Gai Zhu center radial zone through single center outlet port 40.Then the separation component of sample is carried by applicable connecting line to detector 16 by the mobile stream of eluent, and this detector produces a chromatogram trace.In analytic type chromatography, these separated components are sent to refuse 18 or destroyed in testing process after detection.In preparative scale chromatography, the part and this part that detect eluent stream are used as in order to use a reservoir or fraction gathering-device 19 to collect the basis of desirable component from this stream that flows.This system for example, in a control and data gathering system 4, under the control of computer and relevant control electronic device, this control and data gathering system are controlled particularly solvent delivery system 6 and injector 8 and are controlled and receive the data of self-detector 16, and control miscellaneous part.Control and data gathering system 4 can also deal with data for example, be exported as a chromatogram for ().

As schematically illustrated pass through a conventional chromatography column 15 axially or institute's diagram in Fig. 2 of longitudinal cross-section side view, the shape that a kind of sample component that is applied to the head of this post from this injector via a single central inlet port 41 is a kind of Flat belt 10 of relative thin is accumulated in that.In three-dimensional, with 10, be similar to the thin flat disc that the internal diameter by this post housing or wall limits.On this post in separation process and along with sample be with 10 by mobile phase, to be carried along this post downwards, this band starts to change shape, as by as shown in the band at 20 places and as description in more detail in above introduction.Briefly, be positioned on Gai Zhu center or longitudinal axis 17 and around Dai center than the periphery of the band of more close wall, move faster, thereby make the band of material form the shape of a kind of bowl or cup, as the band by 25 places is clearly shown that.In addition, the sample near post jamb starts to scatter (broadening) and becomes dilution more.This phenomenon is progressive, so that it is the most remarkable when this post is left in the outlet end of post or exit at fluid.In the end of post 30, " bowl " of material is being positioned at central axis 17Shang center and drawing oral pore or port 40 through starting to draw after having the thin sieve plate layer 11 of minimum lateral flow impedance.Right side in Fig. 2 has the end-view of end 30 of the post of central port 40 in the middle of being illustrated in.The object of this routine arrangement is that the gross section of this sample " band " is focused to central port 40, and it is guided to detector with sharp-pointed as far as possible peak.

When various ingredients is applied to a packed column, they for the mobile phase in this post and fixedly the different chemical affinity of phase make them with the speed with differing from one another, move through this post.This is basis separated in a LC post.Illustrate in the length of post 15 from partly differentiating each other the different sample component 105,110,115,120 of (i.e. separation) four kinds from the similar Fig. 3 of Fig. 2.Should understand in the illustrative examples shown in Fig. 2 B, in post, in the heart, on Ji center center line 17, a plurality of peaks are differentiated completely, but periphery, more close post jamb place broaden and dilution makes these components overlapping and can not be resolved.Along with this group component 105,110,115,120 is drawn single centre bore or the port 40 in the center of this chromatographic column outlet, wherein from the liquid that strides across whole width, be forced to by this port these display sections, peak in resulting chromatogram Figure 135 or not exclusively resolution.That is, there is discernible a plurality of peak but they at the early part at each peak and more late part place not for example, completely from separated from one another, in middle crest region 140.

Compare with described conventional arrangement, in a preferred embodiment of the invention, the single outlet port 40 arranging with routine is contrary, and a plurality of outlet ports are disposed in the outlet of post.As hereinafter described in more detail, a plurality of preferred embodiments can be used an end connector or outlet joint (being also called end cap) on the port of export of this post, and this end connector or outlet joint have been modified so that different from a conventional LC end connector.A kind of preferred modification is that exit sieve plate and/or end connector are designed to when this sieve plate and/or joint location are discharged by being positioned at a plurality of passages at the difference place in horizontal (radially) cross section of this post mobile phase during in this exit from this post.The mobile phase of discharging by these a plurality of passages in this way, is from the zones of different of this post, more particularly the different radial zones of this post flow out.Allow like this to traverse the isolation of different fluid component of the diameter of this post.From the stream of these a plurality of passages, can be used as a plurality of parts of separating processes and differently processes.In a plurality of preferred embodiments, from the more favourable region of this post, can process respectively with so not favourable mobile phase as the mobile phase of center radial zone, to for example improve the resolution ratio of a resulting chromatogram or the purity of collected fraction.Together with eluent from striding across whole post is collected at and the regular situation detecting as a stream compare, by a plurality of parts of eluent being limited in to the limited radial zone of this post, each part can show the improvement in separative efficiency.

Different exemplary embodiments of the present invention are described now.There are three main preferred aspects in these embodiment.The sieve plate in a first main preferred aspect Shi exit is modified to the mobile phase in the mobile phase at the center from this column section and region (from neighboring area) from around this center is separated and separates.Therefore, eluent stream is split into a part that moves through the part at Gai Zhu center and moved through neighboring area by this sieve plate.Second main preferred aspect be then by center stream and the disengaging of periphery stream for example, in different outlet passages and port (steel end connector or cap) in a flow distributor, this flow distributor is typically installed (being for example screwed to) on the end of this post.In certain embodiments likely in the situation that use this distributor without the shunting sieve plate of shunting this eluent, because having the flow distributor of a plurality of ports can carry out eluent stream to be split into different piece individually, for example wherein towards the surface of the flow distributor of this sieve plate, be positioned at the near surface of this sieve plate, preferably contact the surface of this sieve plate.The 3rd main preferred aspect is with a stream joint of cutting apart and carry out thus a kind of stream of cutting apart and inject so that the heavy curtain stream producing by this post further improves efficiency at column inlet place.

Fig. 4 is schematically illustrated in the principle of a flow distributor in the exit of post 15, and this post is for example for a packed column of HPLC.Fig. 4 illustrates the schematic longitudinal cross-section side view with Fig. 2 and the similar post of Fig. 3.The end-view (i.e. the end-view of the flow distributor 172 on the end at this post) of this column outlet is shown on the right side of this longitudinal cross-section side view.This flow distributor comprises a central outlet port one 75 of locating similarly with the single central port 40 of routine arrangement, this central outlet port receive and transmission from the mobile eluent of the center radial zone in the cross section of this post (being positioned at a region the central axis 17 of this post), this eluent comprises the component higher with resolution ratio that concentration is higher.This flow distributor further comprises and symmetrically six peripheral ports 180 of spaced apart location equidistant around central port 175, these peripheral ports receive and transmission mobile eluent in the neighboring area of the madial wall of more close this post, the more dilution that this eluent comprises this sample, circulation subsequently and the component of good discrimination so not typically.

Leave the eluent representative at Gai Zhu center and make us the material of wishing most, because it is concentration component the highest and that have the most clear resolution in the various ingredients of this sample.Therefore, this part of making us most wishing allowing this eluent of separately using of central port 175 optionally guides to a detector and/or fraction collector (not shown).The peak of the better resolution in resulting chromatogram of right side diagram (trace 195) of Fig. 4, this chromatogram produces from the detection from the eluent of central port 175 only.Wherein all eluents from striding across the gross section of this post are flocked together and are detected with the chromatogram 135(that uses as shown in Figure 3 the arrangement of conventional chromatogram post to obtain) compare, chromatogram Figure 195 illustrates the improvement in resolution capability.Eluent from six peripheral ports 180 is flocked together and jointly form not the part with the eluent of processing together with eluent from central port 175.For example, in an embodiment who detects with a detector from the eluent of central port 175, from the eluent of peripheral port 180, can use on the contrary another detector separating to detect or separate collection maybe can be sent to refuse, or the entrance that it can be sent to same post or another post for further chromatographic isolation to differentiate better these components, after optionally again being concentrated before this further chromatographic isolation.This periphery eluent typically makes us wishing not as center eluent so because it be more dilute and resolution ratio lower.It has the region of stream not too uniformly, and this not too uniform conductance causes the overall dilution of solute, because this solute scatters in larger volume with respect to center flow state.In a plurality of preferred embodiments, (more axial diffusion) solute that the present invention's (axially comprising more closely) solute that therefore the separated centre concentration eluting is the earliest higher and the peripheral concentration eluting are after a while lower.Yet this periphery eluent may still show than better differentiating peak in conventional an arrangement as shown in Figure 3.Never in any form scope of the present invention is limited, this is considered to be due to the fact that a part of eluent of obtaining from a restricted radial zone has than the less sample that axially scatters of eluent of obtaining from traversing the full duration of this post.

Exist different possible mode designs to be suitable for separating an exit sieve plate 11 from the object of the stream of the mobile phase of the outlet of this post.Fig. 5 A illustrates the perspective view of a preferred embodiment of a sieve plate assembly 220, and wherein Fig. 5 B illustrates in the direction of arrow A the side sectional view that the bottom view of this embodiment observing and Fig. 5 C illustrate this embodiment that B-B along the line obtains.In an illustrated embodiment, this sieve plate assembly is assembled from multistage sieve plate, i.e. central circular sieve plate dish 235 and concentric sieve plate ring 245 around, both for example, by the porous material (steel) that is conventionally used as sieve plate material, made, they are for example, by being the non-porous flow barrier of solid (being made as PEEK by polymer) of concentric rings 40 forms separated from one another.Non-porous adapter ring 240 prevents the lateral flow of eluent between two sieve section 235 and 245, thereby maintenance center is separated with peripheral eluent stream.The width of non-porous ring 240 is less than the width of these sieve section in this case, thereby reduces the resistance to the separated component in this eluent stream.These dishes and ring 235,240,245 are fixed together and are retained on is also external lug 250 inside of a circular contour for example, being made by the non-porous material of solid (PEEK), and this external lug is as the joint of one end of this post as described above.Therefore assemble above-mentioned part 235,240,245,250 so that they are combined together to form assembly 220, this assembly is as a sieve plate cap, wherein external rings 250 is set size and is set the peripheral edge 224 that profile becomes to have an extension, this edge mate on one end of this post so that bottom surface 222 sucking fit on one end of this post (being frictional fit) of this sieve plate cap.Once this end connector is screwed on one end of this post, in use the side of sieve plate cap 225 contacts with this steel end connector (below describing in more detail).For a plurality of sieve plate assemblies (as assembly 220) that use together with the present invention, can be typically as directed sucking fit or be screwed to this styletable, but preferably by sucking fit or frictional fit, apply.Fig. 5 D and Fig. 5 E illustrate one and construct with the similar sieve plate assembly shown in Fig. 5 A, Fig. 5 B and Fig. 5 C, except sieve plate dish 235 ' and ring 240 ' and 245 ' have different relative areas, thereby eluent stream is split into from having the different central areas of horizontal area and a plurality of parts of outer peripheral areas relatively.As Multi-instance, outside sieve section 245(245 ') area and center sieve plate dish 235(235 ') the ratio of area can be from 90%:10% to 50%:50% for example, still typically from 80%:20% to 50%:50%, change, wherein outside this scope ratio be also possible.As Multi-instance, above-mentioned ratio can be about 70%:30% or 75%:25%.Preferably, described ratio is about 2:1.The ratio of the area of these sieve section can be for changing a kind of means of ratio of corresponding volume of a plurality of splitter sections of eluent stream.In certain embodiments, may likely use Yi Ge center sieve plate dish 235(235 '), this center sieve plate dish has and sieve section 245(245 ') compare different (for example lower) density.

Fig. 5 F and Fig. 5 G illustrate can be in the present invention for the other sieve plate design in column outlet punishment stream eluent stream, one of them center porous sieve plate dish 535 by a plurality of porous sieve plate dishes 550 around, these sieve plate dishes 535 and 550 are arranged in a non-porous body 540 separately, thereby this body prevents the lateral flow of eluent between two sieve section 535 and 550.With in Fig. 5 F and 5G those similarly other arrangements can have greater or less than shown in six peripheral sieve plate dishes 550.Can change the relative area of center sieve plate dish 535 and peripheral sieve plate dish 550, wherein at the embodiment of different relative areas shown in Fig. 5 F and Fig. 5 G.

In some other embodiment, as shown in Fig. 5 H and Fig. 5 I, column outlet 580 can be equipped with a plurality of outlet capillaries 585,590 to the eluent stream in a plurality of separate sections is transferred out to the end of this post, and one of them radial center capillary 585 is arranged to transport from the first of this stream of the central area of this post and around peripheral ten capillaries 590 that arrange of center capillary 585, is provided to guide the second portion of this stream.Therefore, this first and second part is in the capillary separating, to carry and therefore shunting each other.Preferably, in this class embodiment, as shown in Fig. 5 I, in these ends capillaceous, provide sieve plate 592.Total number capillaceous can change, and for example 5,6,7,8,9,10,11 or 12 or more capillary.

Fig. 6 illustrates one embodiment of the present of invention with decomposition view, and this embodiment can be for example used together with having a standard analysis HPLC steel column 270 of 4.6mm internal diameter.Embodiment in Fig. 6 A is used a shunting sieve plate assembly 278,279 and is a flow distributor of end connector 280 forms, and this end connector has a plurality of flow channels therein, and these flow channels have corresponding outlet port 281,282.A plurality of parts of shunting section sieve plate 278 (illustration 285) (corresponding with the part 235,240 and 245 in Fig. 5 A to Fig. 5 E) are assembled and be placed on this column outlet place.Sieve plate part 285 comprises and is retained on Yi Ge center sieve plate dish section 255 in a non-porous middle PE EK ring 260, wherein encircles 260 and then be retained in an outer peripheral ring sieve section 265.On sieve plate part 285, there is a joint 279, an outer cap of being made by PEEK, this joint is as flow adaptation device and for the flow path of the correspondence separation by shunting sieve plate 278 is alignd with the corresponding outlet port 281,282 of a steel end connector 280 as described later.Joint 279 comprises a main body 271, and this main body closely coordinates (passing through frictional fit) and is enclosed in the sieve plate part 285 of the shunting section sieve plate 278 at this styletable place on the end 268 of this post.In one substitute to arrange, replace frictional fit, joint 279 can coordinate (at an internal thread of main body 271 inside of joint 279, may be screwed on an external screw thread on the end 268 of this post) by a kind of being threaded with one end of this post.Joint 279 has a radial center opening 272, and this radial center opening aligns with the center sieve section 255 of this shunting sieve plate and through eluent this center sieve section, that Cong Gaizhu center radial zone flows out, passes opening 272 to allow thus.Joint 279 also has a plurality of (preferably equidistant) peripheral openings 273(is 5 openings in this case), these peripheral openings peripherally locate and are arranged in an annular concave seat of the end face 277 of this adapter around central opening 272.Should understand these openings 273 and can be circular or they can be slit or there are some other shapes.Peripheral openings 273 is radially to align with the peripheral sieve section 265 of this shunting sieve plate in this case.Thereby these openings 273 allow the eluent that the peripheral radial zone from this post through peripheral sieve section 265 flows out to pass.The peripheral eluent of collecting by a plurality of openings 273 is in this way transferred in this annular concave seat, and wherein these openings 273 are arranged in the end face 277 of joint 279.

Steel end connector or end cap 280 are screwed onto on the external screw thread 275 on the outside of this styletable, manually fastening or fastening by means of a kind of instrument as required.End connector 280 provides one or more outlet ports (three outlet ports 281 in this case) from the peripheral radial zone (via peripheral sieve section 265 and a plurality of peripheral openings 273) of this post, to draw this post He Yige center outlet port 282 for mobile phase and draws this post for mobile phase Cong Gaizhu center radial zone (via center sieve section 255 and opening 272).

When assembling, the end face of the bottom surface of fastening end connector 280 (invisible in the figure) contact joint 279.Contain a plurality of openings 273, at the annular peripheral dimple described in one end of joint 279, align with these peripheral ports 281 and the circumferential flux of the eluent in fluid communication so that by these peripheral openings 273 transmits by these peripheral ports 281 in end connector 280 thus.The annular essence of this dimple means these peripheral ports 281 and aligns optional with the rotation of these peripheral openings 273.The eluent receiving from these peripheral ports 281 flows out and therefore preferably assembles and process together as a part for eluent from same peripheral radial zone, and the entrance that for example detects or guide to refuse or deliver to a chromatographic column is for further chromatographic isolation.More typically, the stream from these peripheral ports 281 will be grouped together and process as a part.Yet, should understand from the stream of one or more peripheral ports and can process respectively.Really, from the stream of each peripheral port, can process respectively.

The central port 282 of the central opening in joint 279 272 and this end connector is in fluid communication on the other hand.Eluent so Cong Gaizhu center radial zone from the central port 282 of this end connector flow out and differently process with the eluent from these peripheral ports.Therefore the part that comprises mobile phase from the eluent of central port 282, in this part, these components of sample have been carried out better resolution and can the in the situation that of analytic type chromatogram, with the resolution ratio increasing, have been detected or with the form of purer fraction, collect at preparative scale chromatography in the situation that.Outlet port 281,282 has interior threaded surface (negative thread) to accept as described below a nipple for outlet pipeline.Alternately, likely these outlet ports are arranged to and have a pin thread, the pin thread on a ledge for example, is screwed onto this pin thread for a joint of outlet pipeline.

Embodiment in Fig. 6 B is identical with shown in Fig. 6 A to a great extent, but replace shunting section sieve plate 278, it uses has a sieve plate assembly of a single sieve plate sheet 278 ' in joint 279 inside, its center tap 279 separates this stream by its opening 272 and place, 273 downstreams at this sieve plate sheet.

The decomposition view of an example of a post 293 of Fig. 7 diagram, this post has a male end part 293 ' in its exit, and end connector 289 is screwed onto this end and divides, and this end connector has an interior threaded surface for this connection.There is a shunting sieve plate assembly 290(of as directed a plurality of sieve plate parts referring to decomposition view illustration) be pushed in a part 299 that is engaged in this styletable, this part has the external diameter less than threaded portion 293 '.Once end connector 289 is screwed onto on the end of this post, shunting sieve plate assembly 290 is retained on appropriate location.Shunting in this embodiment sieve plate assembly 290 comprises by the non-porous PEEK as a flow barrier and encircles 296 Yi Ge separated from one another center sieve plate dish 295 and peripheral sieve plate rings 294.These shunting sieve section are retained in outside PEEK ring joint 297, and this outside PEEK ring joint is used as a sucking fit cap to coordinate on the end 299 of this post.Joint 297 is further fixed on this post with ring steel belt 298, the circumference of this ring steel belt clamping PEEK joint 297.In Fig. 7 embodiment, joint 297 is different from the joint 279 that is used as flow adaptation device in Fig. 6.In this example, these non-porous parts of the sealing bottom surface sieve plate assembly 290 of end connector 289 (being part 296,297) are to keep thus separated from these segment fluid flows of sieve section 295 eluent mobile with 294.End connector 289Ta center has Yi Ge center outlet port 292 and around its three peripheral outlet ports 291, but should be appreciated that the present invention expects the peripheral outlet port 291 of any number, for example one or more peripheral outlet ports 291.A plurality of preferred examples can have from 3 to 10 peripheral outlet ports, particularly 3,4,5,6,7 or 8 peripheral outlet ports.A plurality of end connectors with 3 and 6 peripheral outlet ports are good embodiment.In addition, the present invention expects the center outlet port (transmitting those ports from an eluent stream of Yi Ge center radial zone) of any number, for example one or more centers outlet port.Yet, preferably, there is Yi Ge center outlet port in as directed embodiment.End connector center outlet port 292 is radially arranged in the central area of this post and radially aligns with center sieve section 295, and transmits by the part of the eluent stream of this center sieve section thus.Three peripheral outlet ports 291 are radially arranged in the outer peripheral areas of this post and radially align with peripheral sieve plate ring 294, and the part of the eluent of transport flow by this periphery sieve plate ring thus.Outlet port 291,292 has interior threaded surface to accept as described below a nipple for outlet pipeline.

Fig. 8 A diagram generally with that the similar alternative sieve plate assembly 303 shown in Fig. 7 and the decomposition view of end connector 305.But in this example, end connector 305 has six peripheral outlet ports 302 around a central outlet port 301.At the sieve plate assembly 303 of assembling shown in Fig. 8 B and end connector 305, the sieve plate assembly of this assembling and end connector have been broken away to the relation of the part of these inner assemblings is shown.

The cutaway side sectional view of same four the port end joints shown in Fig. 9 diagram Fig. 7, but illustrate together with attached outlet Tube formation on post 293.From this view, can more clearly find out that outlet port 291 and 292 is through the respective channel 291 ' of this end connector and 292 ' outlet at the inner surface from end connector 289, this end connector it outer surface and sieve plate assembly 290 in fluid communication, outlet port 291,292 is arranged in this outer surface.

The outlet pipeline pipe 331 having for the type of HPLC is installed to center outlet port 292 by male joint 341.A plurality of outlet pipeline pipes 332 also with type are installed on these peripheral outlet ports 291 by having with a plurality of nipples 342 of nipple 341 same types.Nipple 341 and 342 is screwed onto in the passage 292 ' and 291 ' of port 292 and 291 accordingly, these passages are for this purpose with (the moon) screw thread in, and pipeline pipe 331,332 from but compressed fit.Should understand in multiple other designs, this outlet pipeline can be installed on these outlet ports by the female coupling being installed on a convex outlet port on this end connector.End connector 289 is arranged on one end of post 310 by screw thread closely, thus so that the end face of the inner surface of end connector 289 or bottom surface contact sieve plate assembly 290 and making through the corresponding core of the eluent of this sieve plate and corresponding central passage 292 ' and the peripheral channel 291 ' that periphery enters this end connector.These non-porous parts of the sealing bottom surface sieve plate assembly 290 of end connector 289 (being part 296,297) sealing is to keep thus separated from these segment fluid flows of sieve section 295 eluent mobile with 294.

Should understand and can design many other embodiment of the present invention to provide a stream of cutting apart at this column outlet place.

The present invention has following very big advantage as can be seen from the above description: it can be put into practice with existing conventional chromatography column in the following manner: eluent stream is cut apart, for example, by an end connector with a plurality of ports is provided, so that differently process from the eluent of the zones of different of this post.Advantageously, cutting apart of this eluent stream is to carry out on post, is not after post.The present invention therefore (for example) puts into practice by a conventional H PLC system, and wherein a part for this eluent stream is directed to this detector and another part is sent to refuse or recovery or processes in another way.

Schematically illustrated according to an embodiment who analyzes LC system of the present invention in a flowchart in Figure 10.This system is similar with the conventional system shown in Fig. 1 to a great extent, and therefore equal reference numbers is used in reference to same parts.Main difference is the exit at this post, wherein installs by shunting sieve plate assembly of the common indication of reference number 411 and multiport end connector so that outwards mobile eluent is split at least two parts as described above.The Yi Ge little center outlet port 440 of center that is arranged in the cross section of this post flows to detector 16 eluent flowing out from the central area of this post.Thereby the component You Gai center eluent of multiple separation with the sample of improved separating degree flows, stream is carried in a detector 16, and this detector produces an improved chromatogram trace.In analytic type chromatogram, these separated components are sent to refuse receiver 18 after detection, or can be destroyed in testing process.A plurality of little peripheral outlet port 442 in end connector 411 flows to refuse receiver 18 eluent flowing out from the outer peripheral areas of this post.For example, yet in other embodiments, the eluent itself flowing out from this outer peripheral areas via a plurality of peripheral outlet ports 442 can be for analysis purpose and therefore can also () be detected by a detector separating.Owing to having than the higher peak resolution ratio of eluent of assembling from striding across the whole width of this post, this periphery eluent also has higher break-up value.Should understand can be for the preparation of type chromatogram with the system of that the similar a kind of improved resolution ratio shown in Figure 10, wherein the sample from central port 440 is collected in a plurality of fractions, preferably after detecting at least a portion of this sample, similar with the gathering system of Fig. 1.In the improved system shown in Figure 10, preferably connect a kind of device so as to control and or centre of equilibrium outlet port 440 and peripheral outlet port 442 between mobile phase pressure so that the condensed matter of maintenance maximum moves down along the central area of this post.For this purpose, therefore flow or pressure regulator 450 can be connected on the one or more outlet pipelines from these peripheral ports 442 and/or central port 440, preferably as shown on the outlet pipeline from these peripheral ports 442.Control particularly solvent delivery system 6 and injection port or valve 8 and control and receive control and the data gathering system 4 of the data of self-detector 16 also to control flow or pressure regulator 450, for example, in response to the signal feedback from this detector and/or a plurality of flow sensors.By managing on one's own initiative the resistance in these flowline, likely drive more cross or still less fluid by this center or peripheral channel.

The present invention can also make to substitute LC system can be in preparative application.In an embodiment as shown in illustrative diagram 11A, the eluent of assembling from center outlet port 440 is delivered by 16 to fraction gathering-devices 455 of a detector.Yet, the eluent of assembling from these peripheral outlet ports 442 is not delivered by a detector or to a fraction gathering-device 455, but be stored in one or more these reservoirs of reservoir 445(via valve 446, it can be any applicable reservoir, for example trap or in addition post) in, these one or more reservoirs with sample injection valve 8 for the first post 15 in UNICOM for recirculation.Once the first post 15 is completed to chromatogram circulation, this post of load again then, now the material from the peripheral wash-out of previous one or more operations is collected in reservoir 445, and this material is injected in solvent streams by sample injection valve 8.The eluent of center wash-out again detects and is collected in fraction gathering-device 455.Can use chromatogram processing a plurality of other circulation until total material of the sufficiently high and desirable ratio in primary sample under the highest level of obtainable chromatogram, to carry out purifying separated with classification, by each, obtain dividually and collect the material from center outlet port 440.

Figure 11 B is schematically illustrated for mobile phase is connected serially to according to a kind of system of another equipment of the present invention (by that analogy) from a device according to the present invention on by the direction shown in these arrows.In an illustrated embodiment, three LC posts 615 are connected in series, and each their porch of leisure of these three LC posts has their fresh mobile phase supply 606.Should understand the number of LC post in other embodiments can be greater or less than three.Each post 615 has a flow distributor to the diverting flow that leaves the eluent of this post is become to two parts in its exit: Yi Ge center stream part 600 and a circumferential flux part 602.In an illustrated embodiment, stream part 600 in center detects and/or is collected in a plurality of fractions in the exit of each post.On the other hand, circumferential flux part 602 is supplied in the entrance in next post downstream, except rear pillar can be delivered to its circumferential flux part 602 refuse or its recirculation is got back to the first post in series connection, or it is delivered to other places.

Example (1)

Below provide the details of different experiments and result to further illustrate the present invention by other example.

Studied HPLV and FPLC experiment post the two.

Separated for HPLC, to use Ru Yousaimo Science and Technology Ltd. (Thermo Scientific) to supply standard stainless steel chromatogram post (100 * 20mm).In this case, adopt a kind of anti-phase pentafluorophenyl group phase.Yet, should understand in other cases, can adopt the fixedly phase (for example, as C18 or HILIC phase or without a kind of silica of Bonded Phase) of any type.For the stream multiport end connector Shi Yousaimo Science and Technology Ltd. that the customization of column outlet is cut apart as shown in Figure 9, it is this operation making.Cut apart the end connector that flows and therefore there are 4 ports (central port and three outside ports).This end connector replaces a conventional joint to be closely screwed on this styletable.For contrast operation, use a conventional end joint with a single central port to replace this multiport end connector.Adopt the exit sieve plate of a ring-shaped P EEK sealing.

Separated for fast protein liquid chromatogram (FPLC), adopt a glass column (OMNI) with 70mm length * 17mm diameter.Fixing in this post is C18 silica mutually.At this column outlet place, use a stream multiport end connector of cutting apart with five outlet ports (central port and four outside ports).In this situation, this end connector is arranged in this post by inside, because this post is an axial compression column.For contrast operation, use a conventional end joint with a single central port to replace this multiport end connector.For whole post, adopt the exit sieve plate of a ring-shaped P EEK sealing.

All mobile phases are from preparing purchased from the HPLC level solvent of Merck & Co., Inc. (Merck) (this (Kilsyth) matched in Kiel, Victoria (Victoria), Australia).Toluene, ethylbenzene, propylbenzene and butyl benzene as test solute are purchased from Sigma's aldrich (Sigma Aldrich) (castle mountain (Castle Hill), New South Wales (New South Wales)).Milli-Q water (18.2M Ω) is prepared by self-control and filters by a 0.2 μ m filter.

For HPLC post, the standard liquid of toluene, propylbenzene and butyl benzene is under the concentration of about 4mmol, to prepare in mobile phase.The toluene, ethylbenzene, propylbenzene and the butyl benzene that for the standard liquid of FPLC post, at mobile phase, comprise about 2mmol.

The all chromatography experiments that use this stainless steel HPLC post are to use to be equipped with a water generation 600E multi-solvent of water generation (Waters) 717+ automatic injector, a water generation 600E pump, two serial UV/VIS detectors of water generation 2487 and two water generation 600E system controllers to send LC system to carry out.Separation is under isocratic condition, to use a kind of mobile phase of 80/20 or 70/30 methanol/water to carry out under the flow velocity of 18mL/ minute.These inject volume is 200 μ L.It is to regulate difference outlet pressure to change by the PEEK pipeline via adopting different length that outlet stream is cut apart.Not only on the mobile stream of cutting apart, (on the part of stream of drawing this post from the central area of this packed bed) but also the separate section of drawing the stream of this post in the wall region from this carried out UV-absorbance detection (254nm).

Chromatographic isolation on glass FPLC post is used a kind of Shimadzu (Shimadzu) LC-10APvp system to carry out, and this system comprises a Shimadzu LC-10APvp pump, Shimadzu SIL-10ADvp automatic injector, Shimadzu SPD-10Avp UV detector and the online degas module of a Féraud door (Phenomenex) Degassex model DG-440.Separation is to use isocratic condition to use a kind of mobile phase of 80/20 methanol/water to carry out under the flow velocity of 2mL/ minute.Injecting volume is 250 μ L.It is with being that the port of export joint that flows cutting apart of specific purpose manufacture is realized that stream is cut apart, yet in this case, only the eluent from the central area of this post is detected under 254nm and monitored by UV.

The diagram of the parabola essence of the piston flow by a chromatographic bed shown in Figure 2 and being described above.In these experiments described herein, design this port of export joint so that separated near the region of stream of this post jamb and the region of the stream in the central segment of this packed bed.The design of the head joint of testing for these HPLC is carried out diagram at Fig. 7 and Fig. 9.In this end connector, from the eluent stream in this wall region, via a UV detector, be passed in refuse, and this central area is to use a UV detector separating to analyze individually with a kind of usual manner.

Figure 12 is illustrated in the comparison of the single butyl benzene eluting peak reaching in this HPLC system (standard steel column (100mm length * 20mm diameter)) under different eluent splitting ratios.Therefore the difference the chromatographic performance between the sample that has compared the sample of Cong Gaizhu center discharge and drawn from the outer peripheral areas of this post.Figure 12 is illustrated in the peak shape of the butyl benzene of wash-out under various flows splitting ratio: peak 1(is without cutting apart, via Yi Ge center outlet 100%, detect), Feng2(center 53% is detected, peripheral 47% in refuse), Feng3(center 42% is detected, peripheral 58% in refuse) and 27% detection of Feng4(center, peripheral 73% in refuse).For draw the sample record elution curve of this post via this center outlet port.When a stream part over 70% is sent in these peripheral outlets, to compare with non-situation about cutting apart, the increase of efficiency (N value) is up to 43%.These plate counts (N) are: peak 1(2047); Peak 2(2487); Peak 3(2799); And peak 4(2905).Peak width is: peak 1(43 second); Peak 2(41 second); Peak 3(37 second); And peak 4(35 second).

Improved other information in relevant efficiency can be found out from the elution curve of all three kinds of test solutes.Figure 13---curve a(solid line) illustrate when a conventional end joint (single outlet port) is not during for separating of (have utilize cut apart stream), these three kinds test solutes are the elution curve a kind of 80/20 methanol/water mobile phase from this HPLC post.By contrast, Figure 13---curve b(dotted line) in the graphic chromatogram of institute illustrate same sample, but at the elution curve that uses this multiport end connector stream to realize after cutting apart, wherein the 55%(volume % of this eluent stream) stream of port around these (from) be transferred in refuse and the residue 45%(of this stream from the stream of this central port) be detected and for generation of this chromatogram.Similarly, at Figure 13---in curve C (dotted line), identical separation is shown again, but the 77%(of current wherein this sample is from these stream of port around) be transferred in refuse and only remaining 23%(from central port also) be detected for this chromatogram.

The disintegrate-quality of the stream arrangement that use is cut apart and the improvement in detection efficiency are apparent with the form at adjacent peak-to-peak still less overlapping, narrower peak width, peak hangover still less and more strong peak from Figure 13.This further illustrates by more carefully checking normalized peak shape and these overlapping regions at this butyl benzene peak.Figure 14 illustrates the overlapping and highly normalized butyl benzene peak of these chromatograms that the separation since this toluene, propylbenzene and butyl benzene standard obtains, and this separation is under flow, to use the detection stream from the central port of this multiport end connector to carry out in cutting apart of the different ratios as shown in Figure 13 curve a to c.Be clearly shown that on the one end at overlapping, this peak of more low degree of peak and adjacent peak the more hangover of low degree and the peak width of minimizing.These factors are greatly improved this separated resolution capability.For ease of visual inspection, elution time shown in Figure 14 be normalized to they peak maximum in case compensation along with the ratio of the stream in refuse increases, the minimizing of the flow velocity that passes through this detector producing due to the difference by the flow velocity of dead volume after other post.Peak heights is also normalized to compensate the difference time of staying and the amount that is lost to the sample in refuse in this detector flow pond.Most clearly, the degree of cutting apart along with stream increases (along with the ratio being sent in refuse increases), and the separative efficiency in wash-out increases.Yet, also the obvious vision essence after the band broadening post of consideration sample of wash-out in slower fluid flow stream is not taken into account.If also this point is considered in this analysis, after the cutting apart of this stream, the improvement in peak shape will be even more obvious so.

Even if an other advantage of the stream arrangement of cutting apart is the amount of substance guiding in refuse, increase, also observing sensitivity increases (as shown in the peak strength by increasing in not normalized Figure 13).This is considered to the result of two factors, i.e. the solute concentration increasing in the less region of the dilution of this solute piston flow and the minimizing of flow velocity after stream is cut apart; Therefore the solute time of staying in this detector flow pond increases.This is an additional advantage of the detector (for example, as UV detector) of flow-sensitive.

From Figure 13 and Figure 14 most clearly, with the sample belt profile of bulk flow by the non-head joint of cutting apart (curve a) is compared, from this cut apart sample belt profile (curve b and c) that the central port wash-out stream produces for separated be more effective.In table 1, provide these three kinds of analytes asymmetry value at resulting plane-table operation value and peak separately in this HPLC system.Data in table 1 are to obtain under a kind of mobile phase composition of 70/30 methanol/water, because under these conditions, the baseline of realizing between all three kinds of components is differentiated.This allows to use the method for second variance to carry out the accurate measurement of theoretical cam curve N.At this, using under the stream condition of cutting apart, the gain in efficiency (increase of N) is up to 57%.Under the stream condition of cutting apart at these, the asymmetry at peak also greatly reduces.

The curve that chromatogram diagram in Figure 15 is compared with the sample (dotted line trace) from wall region wash-out from the sample (solid-line trace) of the standard liquid of toluene, propylbenzene and the butyl benzene of the central segment wash-out of this post, for this situation, wherein by 70% of total flow, be guided through these peripheral waste port to produce this dotted line trace.From this diagram, can most clearly find out the detrimental effect of this wall.Band tailing is for significant many sample of wash-out in this wall region, and these result pages in this wall region provide in table 1.

table 1

Because these improvement of using stream to cut apart in the elution efficiency described above of generation are observed on the 100mm of marketing quality HPLC post.Even so, observe in theoretical cam curve up to the improvement in 57% efficiency.Use the further improvement of this equipment can realize other gain.Yet, when use and can in FPLC system, adopt those similar glass columns time, still observe larger gain.For example,, at Figure 16 A(solid line) shown in standard four component solutions of toluene, ethylbenzene, propylbenzene and butyl benzene the elution curve of after separating on filling (with C18 spherical (nucleosil) silica) FPLC post of a standard (port) end connector and conventional monoblock type sieve plate is being installed.When outlet stream (multiport) end connector that uses this to cut apart introduces at this column outlet place that stream is cut apart (dotted line) so that stream while drawing by central port and a plurality of peripheral port, when detecting dividually this core, theoretical cam curve increases up to twice, at table 1, describes in detail.That is, depend on every kind of component, as the efficiency of measuring by N has increased (measurement of N is subject to the impact of the peak hangover in separating degree and this non-flow separation of cutting apart) between 51% and 106%.When using stream to cut apart, observe resolution ratio and be increased to 1.03 from 0.81.Figure 16 B illustrates the zoomed-in view in a region of chromatogram of Figure 16 A so that the separation of two more clearly in these tight elution fractions of diagram.

Another advantage of this flow point cut node joint is that the chromatographic column of bad filling can convert the functional column with good separation performance to.For example, the separation of simple four component mixtures of the chromatographic column of a bad filling of use shown in Figure 17.The head wilful damage that this post is passed through to destroy this post is so that when with a kind of usual manner operation, and it provides than desired lower column efficiency by the post for a good filling.Even the chromatogram diagram in Figure 17 A when via a single outlet port, draw 100% mobile phase of this post be sent to detector with there is a single center outlet port a conventional end joint time this four component mixture elution curve.Shown in Figure 17 B when using with identical mobile phase for Figure 17 A at same post but use a multiport end connector, chromatographic isolation in these peripheral outlet open-ended in the situation that during separated these four kinds of identical components.Therefore, liquid flow is drawn this post from a central port and four peripheral ports, but only this centre bore is connected to the detector that produces the chromatogram in Figure 17 B.In Figure 17 A and these the separated important differences of relatively announcement shown in 17B.In Figure 17 A, similar with a kind of conventional outlet arrangement, the wide hangover 355,360,365 of material is observed after each relatively sharp-pointed main peak 350,352,357,362 and departs from this post.Notice that these wide hangovers cause some in these peaks to lack and their adjacent peak completely separated.By contrast, in Figure 17 B, use the eluent stream cut apart, the peak 370,373,375 and 378 of the stream of obtaining since Gai Zhu center outlet port be narrow, do not have and trail and differentiate completely.This data acknowledgement is cut apart stream end connector and not only by improving separative efficiency, is improved column performance, and improves peak symmetry significantly.On good stainless steel column of filling, asymmetry value is reduced to and approaches perfect symmetrical peak from 1.32, and wherein dissymmetry factor is low to moderate 1.04.This improvement is more remarkable on glass FPLC post, and effectively the peak of shunting converts the uniform peak of rule with only appropriate hangover to thus.

For some application, the preferred embodiments of the present invention provide, and the single ingress port 41 arranging with the routine shown in Fig. 2 and Fig. 3 is contrary, and the entrance and exit of this post disposes a plurality of ingress ports.As hereinafter described in more detail, a plurality of preferred embodiments can be used an end connector or inlet attack (being also called end cap) on the arrival end of this post, and this end connector or inlet attack have been modified so that different from a conventional LC end connector.Preferred modification is that entrance sieve plate and/or arrival end joint are designed to when by this sieve plate and/or joint location during in this porch, and mobile phase is incorporated into this post by being positioned at a plurality of passages at the difference place in horizontal (radially) cross section of this post.In this way, the mobile phase of introducing by these a plurality of passages arrives the different radial zones of this post.In a plurality of preferred embodiments, contain more favourable region that a part for the mobile phase of Multi-example is more introduced in this post as radial center region and contain still less or containing another part of the mobile phase of sample, be not introduced in the so not favourable Ru Bi region, region of this post.This entrance arrangement allows the sample concentration of diameter that coutroi velocity and control strides across this post to produce more concentrated, better differentiate and narrower component band.

Figure 18 illustrates the schematic longitudinal cross-section side view with Fig. 2, Fig. 3 and the similar post of Fig. 4.Figure 18 is schematically illustrated in the principle of an inlet flow rate distributor 182 in the exit of post 15, and this post is for example for a packed column of HPLC.The end-view (i.e. the end-view of the flow distributor 182 on the arrival end at this post) of this column inlet is shown in the left side of this longitudinal cross-section side view.This inlet flow rate distributor comprises a central inlet port one 85 of the single central inlet port 41 similar location that arrange with routine, and this central inlet port transfers to (being that central outlet port one 85 is positioned on the central axis 17 of this post) in this post by mobile phase.Preferably, central inlet port one 85 transmission of inlet flow rate distributor 182 are more enriched with sample and more preferably comprise the mobile phase of all samples substantially.Inlet flow rate distributor 182 further comprises and symmetrically six peripheral ingress ports 190 of spaced apart location equidistant around central port 185, these peripheral ports transfer to mobile phase the neighboring area of the madial wall of more close this post, this mobile phase comprise still less or preferably containing sample (being that it is only wherein not have the solvent of sample).Mobile phase periphery stream provides a kind of heavy curtain stream flowing around this center sample circlewise.Should understand at all samples is be introduced into via central port in the situation that, only for a flowline that should central inlet port, need on it, have a sample injection valve, these flowline of supplying these six peripheral ingress ports 190 do not need to have sample injection valve.

A rate of discharge distributor 172 provides the exit at post 15.The end-view (i.e. the end-view of the rate of discharge distributor 172 on the one end at this post) of this column outlet is shown on the right side of this longitudinal cross-section side view.This rate of discharge distributor comprises a central outlet port one 75 of locating similarly with the single central port 40 with conventional arrangement, this central outlet port receive and transmission from the mobile eluent of the center radial zone in the cross section of this post (being positioned at a region the central axis 17 of this post), this eluent comprises the component higher with resolution ratio that concentration is higher.This rate of discharge distributor further comprises six peripheral ports 180 opening location around central port 175 equi-spaced apart, these peripheral ports receive and transmission mobile eluent in the neighboring area of the madial wall of more close this post, and this eluent comprises more dilute sample or containing sample.

The bowl shape producing with conventional entrance arrangement in Fig. 3, part is differentiated is with 105,110,115,120 to compare, entrance arrangement of the present invention shown in Fig. 4 provides more flat sample band 150,155 and 160, these sample bands may not contact these post jambs, and wherein this sample mainly or is all included in the center of this porch sample stream.The flow velocity of the mobile phase in the outer peripheral areas of this post of the heavy curtain of separately introducing stream permission providing by these peripheral ingress ports 190 closer mates the flow velocity in this central area, thereby makes this sample wait to flatten.Can be for this purpose dividually these corresponding parts of pump flow (for example, by a plurality of pump pumpings that separate) or, alternately, a single pump can drive two parts of stream, and in these flowline, a plurality of flow restrictors for each part can be controlled corresponding flow velocity.The sample that this heavy curtain stream has also reduced in this central area laterally moves or spreads the tendency of (outwards towards this wall), thereby concentrates the sample in this central area.Therefore, the sample band 150,155 and 160 that arrives the outlet of this post is more concentrated, more flat and narrower, thereby causes having higher S/N and more sharp-pointed and the better detected peaks of differentiating.These mobile phase parts can have identical or different composition, for example identical or different solvent.A part (a for example radial outer periphery part) can or even a kind of non-solvent or at least there is more a kind of solvent of low solubility (for example water) for thering is sample to be separated, thereby further promote sample for example, comprising in another (radial center) part.

Can more effectively utilize this improvement in the sample band curve of realizing by this post by rate of discharge distributor 172.Leave the eluent representative at Gai Zhu center and make us the material of wishing most, because it is concentration component the highest and that have the most clear resolution in the various ingredients of this sample.Therefore, this part of making us most wishing allowing this eluent of separately using of the central port 175 of the non-orchestration 172 of rate of discharge optionally guides to a detector and/or fraction collector (not shown).Eluent from these six peripheral ports 180 is flocked together and jointly form not the part with the eluent of processing together with eluent from central port 175.For example, in an embodiment who comprises some samples at this heavy curtain stream, wherein with a detector, detect the eluent from central port 175, from the heavy curtain stream eluent of these peripheral ports 180, can use on the contrary another detector separating to detect or separate collection maybe can be sent to refuse, or the entrance that it can be sent to same post or another post for further chromatographic isolation to differentiate better these components, after optionally again being concentrated before this further chromatographic isolation.This periphery eluent typically makes us wishing not as center eluent so because it be more dilute and resolution ratio lower.If this periphery heavy curtain stream eluent comprises seldom or there is no a sample, so it can from these peripheral outlet port one 80Yu Gai center sample flow points turn up the soil assemble and subsequently one take turns chromatography again as mobile phase, thereby the consumption of saving mobile phase.

Should understand this inlet flow rate distributor can be and the design identical with any these rate of discharge distributors described above shown in these accompanying drawings, and wherein this outlet stream refers to that this class is with reference to substituting the reference for separating of the inlet streams of mobile phase as passed through this rate of discharge distributor separation.Should understand this inlet flow rate distributor can use together with an entrance sieve plate assembly, this entrance sieve plate assembly can be shown in these accompanying drawings and the identical design of any these exit sieve plate assemblies described above equally, and wherein this outlet stream refers to that this class is with reference to substituting the reference for separating of the inlet streams of mobile phase as passed through this exit sieve plate components apart.

Should understand the present invention can provide remarkable improvement, and these improvement are to be the sample detection of enhancing and from the form of the improved mensuration performance of a liquid-phase chromatographic column.In a plurality of different embodiment, due to the improved detection sensitivity in a chromatographic determination and/or improved peak capacity and peak resolution ratio, the present invention may can realize for example by the detection lower limit of the species of chromatography.Should be understood that as improve a replacement scheme of peak resolution ratio for a given column length, compare with a similar conventional system, the present invention may can use a plurality of shorter posts to reach a given peak resolution ratio.Shorter post can carry out chromatographic isolation faster.An additional advantage is for example: use an only part for eluent to be introduced in detector for detection of the solvent load that may mean minimizing, this can be highly profitable for some detector, other detectors that these detectors operate in a kind of vacuum environment as mass spectrograph lotus.Therefore the present invention may realize the use of the stock size post with MS detection better.About preparative scale chromatography, due to improved separative efficiency, the present invention may can collect purer sample fraction.The present invention can for example, implement and design in a simple manner from cheap material (sieve plate material and steel end connector) easily, for example, be the form of the end connector with multiport rather than a single-port.

Figure 19 in a flowchart an entrance system of cutting apart of schematically illustrated employing according to one of the present invention another embodiment that analyzes LC system.This system to a great extent with the system class shown in Figure 10 seemingly, therefore and equal reference numbers is used in reference to same parts.In the porch of this post, difference is that this kind of or multi-solvents 2 is delivered to a solvent delivery system 6 via pipeline, and this solvent delivery system adopts two pumps in this example.One or more solvents that enter, if not yet provided to system 6 with the form of two parts of separating, are divided into two parts so.The first of this solvent is directed to and is directed to and by a second pump pumping in system 6 by the second portion of a first pump pumping and this solvent.These independently pump enable to control independently the flow velocity of these parts.These solvents are pumped to post 15 via the flowline that is pipeline 6a, 6b and 6c form.The first of solvent is pumped through pipeline 6a via a sample injection valve 8, and wherein sample is introduced in this partial solvent stream.Load has the first of the solvent of sample to be supplied in post 15 via central inlet port 440 ', and this central inlet port is positioned on the central axis of this post.The second portion that does not contain the solvent of sample is supplied in post 15 by pipeline 6b via a plurality of outsides or peripheral ingress port 442 ', and these outsides or peripheral ingress port are circlewise around central inlet port 440 '.This entrance also has a shunting sieve plate assembly 9 ', so that the first of solvent flows through central segment of sieve plate and the second portion of solvent, flows through circlewise around a peripheral members of the sieve plate of this central segment.The version of embodiment shown in can providing should be provided, and for example pipeline 6b can also have a sample injection valve 8 and be supplied with as required thus a certain amount of sample.

In another embodiment as shown in schematically Figure 20, the eluent of assembling from center outlet port 440 is delivered by 16 to fraction gathering-devices 455 of a detector.Yet, the eluent of assembling from these peripheral outlet ports 442 is not delivered by a detector or to a fraction gathering-device 455, but may be delivered into solvent delivery system 6 via valve 446, for one or more these reservoirs of reservoir 445(that re-use or be stored in of this solvent, can be any applicable reservoir, for example trap or in addition post) in, these one or more reservoirs with sample injection valve 8 UNICOMs for the first post 15 for recirculation.At a reasonable time, then the first post 15 can carry out load again, now the material (comprising therein in the situation of sample) from the peripheral wash-out of previous one or more operations is collected in reservoir 445, and this material is injected in solvent streams by sample injection valve 8.Center wash-out eluent again detects and is collected in fraction gathering-device 455.Can use chromatogram processing a plurality of other circulation until in primary sample sufficiently high and total material of desirable ratio under the highest level of obtainable chromatogram, to carry out purifying separated with classification, by each, obtain dividually and collect the material from center outlet port 440.

Example (2)

Carry out a plurality of other experiments to confirm the effect of an entrance of cutting apart.

These experiments are to carry out on the 100 * 21mm steel column that is filled with 12 μ m pentafluorophenyl group-silica stationary-phase particle size.On same post, test the different experiments of different post hardware configuration, wherein depend on the circumstances and exchange head joint.These experiments of carrying out are used: (1) conventional post (single ingress port and single outlet port); (2) there is the conventional column inlet joint (single ingress port) of the flow export joint (4 ports: 1 central port and 3 peripheral ports) of cutting apart; And (3) have the heavy curtain fluidization tower inlet attack (4 ports: 1 central port and 3 peripheral ports) of the flow export joint (4 ports: 1 central port and 3 peripheral ports) of cutting apart.Toluene, propylbenzene and butyl benzene are infused in a kind of 30/70 water/methanol as mobile phase (250 μ L) for testing the performance of these three kinds of operator schemes.Result is being described below.

The diagram of the parabola essence of the piston flow by a conventional post shown in Figure 2 and being described above.In these experiments (2) and (3) described herein, design this port of export joint so that separated near the region of stream of this post jamb and the region of the stream in the central segment of this packed bed.Design this arrival end joint to a plurality of separated flows of mobile phase are incorporated into accordingly near in the region in the region of this post jamb and the central segment of this packed bed.For the design of the joint of these experiments be Fig. 7 and Fig. 9 graphic that.Use this end connector, from the eluent stream in this wall region, via a UV detector, be passed in refuse, and this central area is to use a UV detector separating to analyze individually with a kind of usual manner.

The chromatographic isolation of the ternary mixture that Figure 21 diagram obtains for every kind of operator scheme (1), (2) and (3).Eluting order is toluene, propylbenzene and butyl benzene then.Conventional chromatogram pattern (1) is to illustrate by a dotted line, and the outlet stream mode (2) of cutting apart is to illustrate and heavy curtain stream mode (3) is to illustrate by solid line by dotted line.The shunting ratio of the outlet stream of cutting apart is that 54% of mobile phase is eluted in refuse and remainder detects or collects via this central port via these peripheral ports.Heavy curtain stream ratio is 1:3.5(center: heavy curtain district).All overall flow rates at column inlet place are 18mL minute.What in these separation, have main conspicuousness is to compare with a kind of normal manipulation mode (1), and the heavy curtain stream for experiment in (3) injects 155% gain of observing sensitivity, below in table 2, describes in detail.

table 2

This gain in sensitivity is considered to a kind of result of double effect:

(i) all samples in heavy curtain stream mode (3) directly loads to the central port of this post center and the outlet connection that this sample (its 100%) wash-out is cut apart by this subsequently via this central inlet port, wherein at detectable limit place, does not observe sample component via these peripheral outlet ports wash-out in waste stream.Therefore sample is used the mass loading identical with a kind of normal manipulation mode (1) but carries out wash-out with substantially less elution volume, as following further description.

(ii) flow through the volume of this detector lower, because only 46% solvent is delivered by this detector.That is, with at normal manipulation mode, (18mL/ in 1 minute compares, and the flow velocity by this detector in operator scheme (3) is 8.3mL/ minute.Therefore the increase of detector in the time of staying produces larger sensitivity.

Also in Figure 21, be clear that the export model (2) cut apart produces substantially than the two narrower chromatographic curve of normal manipulation mode (1) and heavy curtain flow operation pattern (3).Plate count (N) and other quality factor of describing efficiency are also included in table 2.Compare with normal manipulation mode, exist the operator scheme of cutting apart for this 16% of efficiency to increase, but separative efficiency does not change for this heavy curtain stream mode.Yet, should note (referring to hereinafter) as further, discussed, for these used specific designs, these tolerance of N are not best tolerance.Yet contrary with viewed slight hangover in normal manipulation mode (1), the separation in heavy curtain stream mode (3) produces a peak of full symmetric.Data from table 2 also noteworthy is that, the export model of cutting apart of using operation (2) via the sample efficiency of these peripheral ports wash-out in waste stream higher than a kind of normal manipulation mode about 12.5%.

The limit of the degree such as the measurement requirement of plate count N.A critical aspects that limits the described experiment of Performance Ratio between different operator schemes be flow velocity by detector this normal manipulation mode from relate between any other operator scheme of the outlet of the cutting apart stream by a detector different.In all operations pattern---normal (1), the outlet (2) of cutting apart or the heavy curtain with the outlet of cutting apart flow in (3), volume flow by bed keeps constant, yet, for the operator scheme of cutting apart, the mobile conductance flowing in column outlet punishment causes only 46% solvent and passes detector.Therefore, the apparent peak width of pattern (2) and (3) is in fact than the wide twice of normal manipulation mode (1).The apparent tolerance of theoretical cam curve is reduced in this artificially.Therefore in the operator scheme of cutting apart, the better reflection of separating property can obtain by measuring the amount (being the concentration of solute) of the solute of wash-out in the collected volume of sample.In order to measure this, therefore carry out a research in addition, wherein a plurality of sample bands are that the elution volume that strides across them with five seconds intervals carries out classification separation.Then analyze each fraction to measure the amount of the solute of wash-out.Then the amount of collected solute is mapped as the function of the elution volume from this post, its result is shown in Figure 22.Trace A obtains from normal mode (1), trace B is used from the sample of central port wash-out to obtain from cutting apart stream mode (2), trace C is used the sample of (a plurality of peripheral port) wash-out from wall region to obtain from cutting apart stream mode (2), and trace D obtains from heavy curtain stream mode (3).

Figure 22 diagram normal manipulation mode (1) and for the comparison (wash-out based on propylbenzene band) of cutting apart the separating property between each of flow operation pattern (2) and heavy curtain stream injection way (3) of this classification Separation Research.These separation clearly diagram these cut apart each the level of raising of separating property in flow operation pattern and heavy curtain stream.The sample collection volume of cutting apart flow operation pattern (2) of the sample (trace B) of collecting from center flow stream be about 4mL and from wall region (trace C) 7mL.The sample collection volume with the heavy curtain stream injection (trace D) of the outlet stream of cutting apart is about 5.5mL.In routine operation pattern, sample collection volume is 11mL.Therefore, the most effective pattern (at least with respect to peak volume) of sample component extraction is to cut apart flow operation pattern, and wherein sample is collected by central outlet section.Yet in this operator scheme, about 50% sample is sent to waste stream, although then this sample is included in the solvent of about 7mL subsequently.Because 100% sample in heavy curtain flow operation pattern flows wash-out via center flow, this operator scheme produces more effective extracting process, its efficiency higher than normal mode 100%(therefore at least with respect to sample collection concentration (and detection sensitivity)), and present improved separative efficiency with respect to peak volume.Cost effectiveness can obtain about solvent recycled in addition, because use heavy curtain flow regime not observe sample from the wall region wash-out of post.Therefore this solvent can recycle and without any ability need, in the operation described pattern, it will account for 54% of whole solvent consumptions.

As used herein, comprise claim, unless context illustrate in addition, otherwise should be understood to comprise plural form at the singulative of this term, and vice versa.For example, unless context illustrate in addition, singular reference is as " one (a) " or " one (an) " expression " one or more ".

The description and the claim that run through this description, word " comprises (comprise) ", the version of " comprising (including) ", " having (having) " and " containing (contain) " and these words (for example " comprise (comprising) " and " comprising (comprises) " etc.) represents " including but not limited to " and be not intended to (and not can) and get rid of miscellaneous part.

Should be understood that, can make the version of above embodiment of the present invention, still fall within the scope of the present invention simultaneously.Except as otherwise noted, otherwise each feature disclosing in this description can be identical by serving, be equal to or the alternative features of similar object replaces.Therefore, except as otherwise noted, otherwise each disclosed feature is an example in being equal to of a general series or similar characteristics.

Any one and whole examples or exemplary language (" for example (for instance) ", " as (such as) ", " for example (for example) ", " for example (e.g.) " and similar language) that use provides at this, only be intended to illustrate better the present invention and do not indicate scope of the present invention is limited, unless requirement in addition.Any language in this description be not appreciated that be indication: any element that does not propose claim be to of the present invention realize vital.

Any step of describing in this description can be carried out or carry out simultaneously according to random order, except as otherwise noted or context requirement.

The all features that disclose in this description can combine with any combination, except the combining form of at least some the phase mutual repulsions in this category feature and/or step.Specifically, preferred feature of the present invention is applicable to all aspects of the present invention and can uses with any combination.Equally, the feature of describing in nonessential combining form can be used separately (not combining).

Claims (65)

1. the equipment for column chromatography, this equipment comprises a chromatographic column, this post has an entrance and an outlet, wherein this outlet is equipped with a sieve plate assembly, this sieve plate assembly is configured to make this eluent stream be split at least two parts of separating when an eluent stream is left this post by this outlet, and wherein this equipment is configured to process dividually these parts.

2. equipment as claimed in claim 1, wherein this outlet is configured to the Yi Ge first of this stream to guide in first treating apparatus and by a second portion of this stream and guide in second treating apparatus separating with this first treating apparatus.

3. equipment as claimed in claim 1 or 2, wherein these at least two parts of separating flow out from the different radial zones of this post.

4. equipment as claimed in claim 3, one of them first flows out from this post Yi Ge center radial zone, and a second portion flows out from being positioned at a radial zone radially outward of this center radial zone.

5. equipment as claimed in claim 4, wherein Gai Zhugai center radial zone is to be located substantially on from this entrance to this outlet to run through a region a central axis of this post.

6. the equipment as described in any above claim, wherein a part is to flow out from a region of this post, from the eluent stream in this region through the part of the uniform filling of this post.

7. the equipment as described in any above claim, wherein this outlet is arranged to when this eluent stream is left this post, this eluent stream is split into three or more parts of separating.

8. the equipment as described in any above claim, wherein a part for eluent be by volume total eluent 50% or still less, 45% or still less, 40% or still less, 35% or still less, 30% or still less, 25% or still less, 20% or still less, 15% or still less, 10% or still less or 5% or still less.

9. the equipment as described in any above claim, wherein this sieve plate assembly comprises at least two sieve section of separating, these sieve section are by a flow barrier separation.

10. equipment as claimed in claim 9, wherein this sieve plate comprise at least one radial center sieve section, around flow barrier of this at least one center sieve section and around this at least one center sieve section and by this flow barrier at least one radially outer sieve section separated with it.

11. equipment as claimed in claim 10, wherein the ratio of the area of the area Yu Gai center sieve section of this outside sieve section is from about 2.5:1 to about 1.5:1.

12. equipment as described in claim 10 or 11, wherein the width of this flow barrier is less than the width of each sieve section..

13. equipment as described in any one in claim 9 to 12, wherein this sieve plate assembly comprises that the non-porous joint in outside with at least two openings is to be separated at least two parts of separating by this eluent stream.

14. equipment as described in any above claim, wherein this post has a flow distributor in this exit to carry these at least two parts of separating of this eluent stream in the passage separating therein.

15. equipment as claimed in claim 14, wherein this flow distributor is the part separating being in use installed on the end of this post.

16. equipment as described in claims 14 or 15, wherein this flow distributor comprises one second group that has first group of at least one passage and have at least one passage, this first group be arranged so that in use this first group be arranged in first radial zone of this post to carry the Yi Ge first of eluent, this second group be arranged so that in use this second group second radial zone that is arranged in this post to carry a second portion of eluent.

17. equipment as claimed in claim 16, wherein this first radial zone Shi Yige center radial zone of this post and this second radial zone of this post are to be positioned at the region that this center radial zone is radially outward located.

18. equipment as described in claim 16 or 17, wherein this flow distributor is included in central passage and from three to ten external channels in this second group in this first group.

19. as being subordinated to the equipment as described in any one in the claim 16 to 18 of any one in claim 9 to 13, wherein this flow distributor in use contacts with the one or more non-porous part of this sieve plate assembly, so that the one or more non-porous part of this sieve plate assembly provides a kind of sealing between this sieve plate and flow distributor, thereby the adjacent part of eluent stream is sealed each other.

20. equipment as described in any above claim, wherein this equipment comprises a detector, this detector is arranged to detect dividually with other part or a plurality of part at least a portion of this eluent.

21. equipment as described in any above claim, wherein this equipment comprises a fraction collector, this collector is arranged to collect dividually with this other part or a plurality of part the fraction of at least a portion of this eluent.

22. equipment as described in claim 20 or 21, wherein this detector or fraction collector are arranged to detect dividually or collect the fraction of a part for the eluent having flowed out from this post Yi Ge center radial zone.

23. equipment as described in any above claim, wherein this equipment is configured to that one or more parts are delivered to a refuse receiver or to the entrance of a LC post.

24. equipment as described in any above claim, wherein this equipment comprises for processing dividually at least the first and second treating apparatus of this at least the first and second part of eluent, and wherein this first and second treating apparatus comprises one or more in the following independently of one another: detector, refuse reservoir, fraction collector and column inlet.

25. equipment as described in any above claim, wherein this post is for being selected from the chromatographic post of analytic type of the following: high performance liquid chromatography (HPLC), Ultra Performance Liquid Chromatography method (UHPLC), multidimensional or two-dimensional highly effective liquid phase chromatographic method (MDHPLC or 2DHPLC), flash column chromatography, fast protein liquid chromatography (FPLC) and supercritical fluid (SCF) chromatography.

26. equipment as described in any above claim, wherein this post is for the preparation of the chromatographic post of type.

27. equipment as described in any above claim, wherein this entrance is configured to the stream of mobile phase to be incorporated in this post with independent at least two controlled parts of separating, and these parts are incorporated in the different radial zones of this post, so that these parts longitudinal flow in different radial zones is passed this post.

28. equipment as claimed in claim 27, wherein the Yi Ge first of mobile phase is incorporated in the radial zone of this post Yi Ge center, and a second portion of mobile phase is incorporated into and is arranged in the peripheral radial zone that this center radial zone is radially outward located.

29. equipment as claimed in claim 28, wherein a kind of to have sample to be separated be to be contained in this first of mobile phase with the concentration than higher in this second portion.

30. equipment as described in any one in claim 27 to 29, wherein these mobile phases are partly controlled to have substantially the same flow velocity.

31. equipment as described in any one in claim 27 to 30, wherein this entrance is equipped with an entrance sieve plate assembly, and this entrance sieve plate assembly is configured to this mobile phase to be divided at least two parts of separating.

32. equipment as described in any one in claim 27 to 31, wherein this post has an inlet flow rate distributor in this porch to carry at least two parts of separating of this mobile phase in the passage separating therein.

The method of 33. 1 kinds of column chromatographies, the method comprises: provide a mobile phase, a kind of sample that this mobile phase comprises a plurality of components of one-tenth to be separated; Make this mobile phase to an outlet longitudinal flow of this post, pass this post from an entrance of a chromatographic column, this mobile phase leaves this post as a kind of eluent by this outlet; A sieve plate assembly is provided in this exit, and this sieve plate assembly is arranged to when eluent stream is left this post by this outlet and makes this eluent stream be split at least two parts of separating; And process dividually these at least two parts of separating.

34. methods as claimed in claim 33, the method comprises the Yi Ge first of this stream is guided in first treating apparatus and by a second portion of this stream and guided in second treating apparatus separating with this first treating apparatus.

35. methods as described in claim 33 or 34, wherein these at least two parts of separating flow out from the different radial zones of this post.

36. methods as claimed in claim 35, one of them first flows out from this post Yi Ge center radial zone, and a second portion flows out from being positioned at this center radial zone radial zone radially outward.

37. methods as claimed in claim 36, wherein Gai Zhugai center radial zone is to be located substantially on from this entrance to this outlet to run through a region a central axis of this post.

38. methods as described in any one in claim 33 to 37, wherein a part flows out from a region of this post, from the eluent stream in this region through the part of the uniform filling of this post.

39. methods as described in any one in claim 33 to 38, wherein this outlet is arranged to when this eluent stream is left this post, this eluent stream is split into three or more parts of separating.

40. methods as described in any one in claim 33 to 39, wherein a part for eluent be by volume total eluent 50% or still less, 45% or still less, 40% or still less, 35% or still less, 30% or still less, 25% or still less, 20% or still less, 15% or still less, 10% or still less or 5% or still less.

41. methods as described in any one in claim 33 to 40, wherein this sieve plate assembly comprises at least two sieve section of separating, these sieve section are by a flow barrier separately.

42. methods as claimed in claim 41, wherein this sieve plate comprise at least one radial center sieve section, around flow barrier of this at least one center sieve section and around this at least one center sieve section and by this flow barrier at least one radially outer sieve section separated with it.

43. methods as claimed in claim 42, wherein the ratio of the area of the area Yu Gai center sieve section of this outside sieve section is from about 2.5:1 to about 1.5:1.

44. methods as described in any one in claim 42 to 43, wherein this sieve plate assembly comprises that the non-porous joint in outside with a plurality of openings is to be separated at least two parts of separating by this eluent stream.

45. methods as described in any one in claim 33 to 44, are included in a flow distributor that this exit offers this post to carry these at least two parts of separating of this eluent stream in the passage separating therein.

46. methods as claimed in claim 45, wherein this flow distributor is the part separating being in use installed on one end of this post.

47. methods as described in claim 45 or 46, wherein this flow distributor comprises one second group that has first group of at least one passage and have at least one passage, this first group be arranged so that in use this first group be arranged in first radial zone of this post to carry the Yi Ge first of eluent, this second group be arranged so that in use this second group second radial zone that is arranged in this post to carry a second portion of eluent.

48. methods as claimed in claim 47, wherein this first radial zone Shi Yige center radial zone of this post and this second radial zone of this post are to be positioned at the region that this center radial zone is radially outward located.

49. methods as claimed in claim 48, wherein this flow distributor is included in central passage and from three to ten external channels in this second group in this first group.

50. as being subordinated to the method as described in any one in the claim 45 to 49 of any one in claim 41 to 44, wherein this flow distributor is in use to contact with the one or more non-porous part of this sieve plate assembly, so that the one or more non-porous part of this sieve plate assembly provides a kind of sealing between this sieve plate and flow distributor, thereby the adjacent part of eluent stream is sealed each other.

51. methods as described in any one in claim 33 to 50, comprise at least a portion that detects dividually this eluent with other part or a plurality of part.

52. methods as described in any one in claim 33 to 51, comprise the fraction of collecting dividually at least a portion of this eluent with this other part or a plurality of part.

53. methods as described in claim 51 or 52, comprise the fraction that detects or collect dividually a part for the eluent having flowed out from this post Yi Ge center radial zone.

54. methods as described in any one in claim 51 to 53, comprise this other part or a plurality of part are delivered in a refuse receiver or to the entrance of a post.

55. methods as described in any one in claim 33 to 54, wherein this chromatography is the analytic type chromatography that is selected from the following: high performance liquid chromatography (HPLC), Ultra Performance Liquid Chromatography method (UHPLC), flash column chromatography, fast protein liquid chromatography (FPLC) and supercritical fluid (SCF) chromatography; Or this chromatography is preparative scale chromatography.

56. methods as described in any one in claim 33 to 55, comprise at least a portion of this eluent from this outlet for giving back to the entrance of this post, and this at least a portion is optionally for again concentrated before in giving back to this entrance; Or be supplied in the entrance of another post, this at least a portion is again concentrated before optionally in being supplied to the entrance of these other posts.

57. methods as described in any one in claim 33 to 56, further comprise mobile phase are provided to this entrance with independent at least two controlled parts of separating, at least one in these at least two parts of separating includes sample to be separated; And these at least two parts of separating of mobile phase are incorporated in the different radial zones of this post.

58. methods as claimed in claim 57, comprise the Yi Ge first of mobile phase is incorporated in the radial zone of this post Yi Ge center, and a second portion of mobile phase is incorporated into and is arranged in the peripheral radial zone that this center radial zone is radially outward located.

59. methods as claimed in claim 58, wherein having sample to be separated is to be contained in this first with the concentration than higher in this second portion.

60. methods as described in claim 58 or 59, wherein this second portion restriction sample Cong Gai first of mobile phase is towards the migration of a wall of this post.

61. methods as described in any one in claim 57 to 60, wherein these mobile phases are partly controlled to have substantially the same flow velocity.

62. methods as described in any one in claim 57 to 61, further comprise to this entrance an entrance sieve plate assembly are provided, this entrance sieve plate assembly is configured to this mobile phase to be divided at least two parts of separating.

63. methods as described in any one in claim 57 to 62, further comprise to this post and provide an inlet flow rate distributor to carry these at least two parts of separating of this mobile phase in the passage separating therein.

64. 1 kinds of equipment for column chromatography, this equipment comprises a chromatographic column and a detector, this post has an entrance and an outlet, wherein this outlet is configured to make this eluent stream to be split at least two parts of separating that flow out from the different radial zones of this post when an eluent stream is left this post by this outlet, this detector is arranged to detect dividually with one or more other parts the part having flowed out from a radial center region of this post, these one or more other parts from this central area a region radially outward or a plurality of region flow out.

65. 1 kinds of equipment for column chromatography, this device comprises a chromatographic column, this post has an entrance and an outlet, wherein this outlet is configured to guide this part when a part for an eluent stream is left this post so that a remainder of itself and this eluent is processed dividually, and wherein this part flows out from a restricted radial zone of this post.

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