CN107589190A - The double SPE high performance liquid chromatography on-line coupling equipment of large volume sample injection - Google Patents
The double SPE high performance liquid chromatography on-line coupling equipment of large volume sample injection Download PDFInfo
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Abstract
The present invention relates to a kind of double SPE high performance liquid chromatography on-line coupling equipment of large volume sample injection.Equipment of the present invention includes：Three chromatogram pumps, one large volume automatic sampler, the automatic sampler of one common sample size, one direction changeover valve of high pressure ten, two solid-phase extraction columns, and column oven, chromatographic column and detector, pass through the combination equipment, connected using simplest hardware device with stream, allow the direct injected of the large volume fluid sample more than 1mL, it is equipped with the SPE extraction column of two parallel processing, it is combined with high performance liquid chromatography, carry out overlapping sample introduction, pre-treatment is with separating, greatly reduce holding time of the single sample in whole batch processing sample, and, this combination equipment can easily switch to common highly effective liquid phase chromatographic system, realize the free switching between on-line system and common separation system.
Description
Technical field
The invention belongs to analytical chemistry sample pre-treatments field, and in particular to a kind of large volume sample injection-bis- SPE-height
The design and analysis of effect liquid phase chromatogram on-line coupling equipment and application, the trace organic substance in fluid sample environment, food
Sample introduction, on-line extraction, enrichment, removal of impurities, desorption, separation and detection.
Background technology
Analysis for trace compound, typically there are two kinds of strategies：Small size samples (such as biological sample), high sensitivity inspection
Survey device detection；Or large volume sampling (such as environmental sample), sample introduction is analyzed after enrichment removal of impurities.
Environmental sample, such as the analysis detection of underwater trace pollutant, sampling volume and sampling volume can be generally improved, so
SPE (Solid Phase Extraction are used afterwards；SPE) it is enriched with, after removal of impurities, sample introduction divides again for concentration, the molten desorbed solution of weight
Analysis.But offline SPE is present, and manual errors are big, cumbersome, the defects of wasting time and energy, it is therefore desirable to develop online " big
Injection volumes (Large Volume Injection；LVI)-SPE-on-line coupling method " is to improve the reproduction of method
Property, automation, shorten analysis time.
High performance liquid chromatography (High Performance Liquid Chromatography, HPLC) is most conventional at present
Analysis means, the technology can to object carry out accurate quantitative analysis, be equipped with the strong detection of the qualitative ability such as diode array, mass spectrum
Device can accomplish accurate Semi-qualitative, and majority of compounds can be efficiently separated and quantified by injecting HPLC after pre-treatment
It is qualitative.Ultra performance liquid chromatography system (Ultra High Performance Liquid Chromatography, UHPLC or
Person Ultra Performance Liquid Chromatography, UPLC) its hardware principle is as common HPLC, together
Sample meets the technical background of this patent.
But the on-line coupling of SPE and high performance liquid chromatography, two limitations be present：When sampling volume compared with
It is small, generally less than 1mL, it is necessary to manual off-line mode sample is concentrated after sample introduction again；Second, SPE loadings and processing sample
Time and the time of HPLC separation objects are all long, cause whole analysis time to extend.Existing SPE is online
Combined system connects single solid-phase extraction column usually using six direction changeover valves, by the switching of six direction changeover valves, realizes that loading extracts
Take the conversion with two states of Desorption separation；In addition, the sample introduction speed of general injector is about 1mL/min, if sampling volume
Up to several milliliters even tens of milliliters, its sample injection time will be difficult to ignore, along with solid-phase extraction column also needs to activate, clean and
Equilibration time, whole analysis time will be very very long, and analysis throughput will substantially reduce.
The content of the invention
Based on this, it is an object of the present invention to provide a kind of on-line coupling equipment, simplest hardware device and stream are used
Road connects, it is allowed to the direct injected of the bulk sample more than 1mL, the SPE extraction column of two parallel processing is equipped with, with
High performance liquid chromatography is combined, and is carried out overlapping sample introduction, pre-treatment and is separated, greatly reduces single sample in whole batch processing sample
Holding time in product, also, this combination equipment can easily switch to common highly effective liquid phase chromatographic system, realize more work(
The purpose of energy system.
The present invention is achieved by the following technical solutions：A kind of large volume sample injection-bis- SPE-high performance liquid chromatography
On-line coupling equipment, including large volume sample injection-bis- SPEs on-line coupling device and high performance liquid chromatography separation device；
The large volume sample injection-bis- SPEs on-line coupling device includes being used to push the of solvent needed for SPE
One liquid chromatography pump, the first automatic sampler for being loaded into sample, for stream switching ten direction changeover valves and for richness
Collection object simultaneously removes deimpurity first solid-phase extraction column and second solid phase extraction column；
The high performance liquid chromatography separation device includes being used for the second chromatogram pump and the 3rd chromatogram for pushing separation mobile phase
Pump, the second automatic sampler for being loaded into sample and the chromatographic column for chromatographic isolation；
Ten direction changeover valve is disposed with ten interfaces by clock-wise order：First automatic sampler access the
One interface, the first solid-phase extraction column both ends are respectively connected to second interface and the 9th interface, and the chromatographic column access the 3rd connects
Mouthful, the second solid phase extraction column both ends are respectively connected to the 4th interface and the 7th interface, the 6th interface connection sewer pipe, and described the
Two automatic samplers access the 8th interface, the 5th interface and the connection of the tenth interface.
Prior art is different from, the present invention is using ten direction changeover valves as connection maincenter sample introduction-bis- solid-phase extraction devices
Being connected with high performance liquid chromatography separation device, ten direction changeover valves only have two states, so as to there was only two streams in system,
It is simple to operate；Switch while realizing two SPEs and two streams by the state switching of ten direction changeover valves, carry out weight
Folded sample introduction, pre-treatment and separate, shorten the global analysis time.
Further, the state of ten direction changeover valve be first interface connected with second interface, the 3rd interface and the 4th
Orifice, the 5th interface and the 6th orifice, the 7th interface and the 8th orifice, the 9th interface and the tenth orifice
When, first liquid chromatography pump, the first automatic sampler, the first solid-phase extraction column are linked to be extraction stream, second chromatogram
Pump, the 3rd chromatogram pump, the second automatic sampler, second solid phase extraction column, chromatographic column are linked to be Desorption separation stream.
The state of ten direction changeover valve be second interface and the 3rd orifice, the 4th interface and the 5th orifice,
When 6th interface connects with the 7th orifice, the 8th interface with the 9th orifice, the tenth interface with first interface, described
One liquid chromatography pump, the first automatic sampler, second solid phase extraction column are linked to be extraction stream, second chromatogram pump, the 3rd color
Spectrum pump, the second automatic sampler, the first solid-phase extraction column, chromatographic column are linked to be Desorption separation stream.
So as to be switched by the state of ten direction changeover valves, automatically switch two parallel solid-phase extraction columns, two solid phases
Extraction column is at cleaning, balance, loading or Desorption separation stage at any one time, at one of solid-phase extraction column
While cleaning, balance, loading stage, another solid-phase extraction column is in the Desorption separation stage, makes two solid-phase extraction columns
The overlapping alternating of analysis process.
Further, the extraction stream is opposite with flow direction of the separation flow path in solid-phase extraction column is desorbed.So as to reduce
Migration distance of the object in solid-phase extraction column, improve desorption efficiency and chromatographic isolation performance.
Further, the on-line coupling equipment also includes column oven and the detector for detection signal, and described first
Solid-phase extraction column, second solid phase extraction column, ten direction changeover valves and chromatographic column are located in the column oven, and the detector is connected to
The outlet of chromatographic column.
Further, low pressure gradient proportioning valve built in first chromatogram pump, selectivity push at least three kinds of composition differences
Solvent.Loading or the eluting power of balanced solvent are low, can push loading sample and flow through solid-phase extraction column, but not enrichment
Eluted in the object of solid-phase extraction column；The medium cleaning fluid of eluting power can get off a part of Impurity elution but not
Object can be eluted；The cleaning solution of high eluting power is responsible for cleaning whole extraction pipeline, and handle is remained on solid-phase extraction column
Object and matrix elute, so as to will not cause next time analyze cross pollution.
Further, the sample size of first automatic sampler is more than 1mL.
Further, first automatic sampler is provided with the changeable quantitative loop for leaving stream.After completing to extract,
Need to clean solid-phase extraction column and balanced, now by the quantitative loop of the first automatic sampler with large volume sample injection amount
Stream is switched away from, the time Jing Guo the quantitative loop is cleaned and balance to reduce.
Present invention also offers a kind of highly effective liquid phase chromatographic system, including the second chromatogram pump of the present invention, the 3rd color
Compose pump, the second automatic sampler, chromatographic column and ten direction changeover valves, and duplex fitting；Ten direction changeover valve is by suitable clockwise
Sequence is disposed with ten interfaces：Second automatic sampler accesses the 8th interface, and the duplex fitting both ends are respectively connected to
Second interface and the 9th interface, the chromatographic column access the 3rd interface.
Present invention also offers a kind of highly effective liquid phase chromatographic system, including the second chromatogram pump of the present invention, the 3rd color
Compose pump, the second automatic sampler, chromatographic column and ten direction changeover valves, and duplex fitting；Ten direction changeover valve is by suitable clockwise
Sequence is disposed with ten interfaces：Second automatic sampler accesses the 8th interface, and the duplex fitting both ends are respectively connected to
4th interface and the 7th interface, the chromatographic column access the 3rd interface.
Relative to prior art, the present invention replaces one of them of first solid-phase extraction column and second solid phase extraction column
When changing duplex fitting into, its connected stream is transformed to common highly effective liquid phase chromatographic system, so as to realize on-line analysis
Simple switching between system and common highly effective liquid phase chromatographic system, meet the multifunctional application of system.
In order to more fully understand and implement, the invention will now be described in detail with reference to the accompanying drawings.
Brief description of the drawings
Fig. 1 is the on-line coupling equipment schematic diagram of embodiment 1；
Fig. 2 is the fluid communication schematic diagram of " 0 " number state in embodiment 1；
Fig. 3 is the fluid communication schematic diagram of " 1 " number state in embodiment 1；
Fig. 4 is the fluid communication schematic diagram in embodiment 2；
Fig. 5 is the chromatogram that acid loading analyzes 80 kinds of medicines and personal care articles in experiment 1；
Fig. 6 is the chromatogram that 1 neutral and alkali loading of experiment analyzes 80 kinds of medicines and personal care articles；
Fig. 7 is the chromatogram that 7 kinds of agricultural chemicals are analyzed in experiment 2；
Fig. 8 is the chromatogram of common 25 kinds of medicines of high performance liquid chromatography pattern analysis and personal care articles in experiment 3.
Embodiment
Further to illustrate the present invention to reach technological means and its technique effect that predetermined goal of the invention is taken, with
Under in conjunction with the embodiments and accompanying drawing, large volume sample injection proposed by the present invention-bis- SPEs-high performance liquid chromatography on-line coupling is set
Standby structure, feature and its embodiment illustrates, and describes in detail as follows.
Embodiment 1
Referring to Fig. 1, large volume sample injection-bis- SPEs-high performance liquid chromatography on-line coupling that it is the present embodiment is set
Standby schematic diagram, it includes large volume sample injection-bis- SPEs on-line coupling device and high performance liquid chromatography separation device.
The large volume sample injection-bis- SPEs on-line coupling device includes being used to push different solvents needed for SPE
The first liquid chromatography pump 1, the first automatic sampler 4 with large volume sample injection amount for being loaded into sample, cut for stream
The direction changeover valve 7 of high pressure ten that changes and for being enriched with object and going deimpurity first solid-phase extraction column 8 and second solid phase to extract
Take post 9.
It is provided with specifically, the sample size of first automatic sampler 4 is more than 1mL, in it and changeable leaves determining for stream
Ring is measured, it is necessary to be cleaned and balanced to extraction column after completing to extract, now by first with large volume sample injection amount certainly
The quantitative loop of dynamic injector switches away from stream, and the time Jing Guo the quantitative loop is cleaned and balance to reduce.
The high performance liquid chromatography separation device includes being used for the second chromatogram pump 2 and the 3rd chromatogram for pushing separation mobile phase
Pump 3, the second automatic sampler 5 for being loaded into common amount volume sample, the chromatographic column 10 for chromatographic isolation and for examining
Survey the detector 11 of signal.
The large volume sample injection-bis- SPEs-high performance liquid chromatography on-line coupling equipment also includes being used to load first
Solid-phase extraction column 8, second solid phase extraction column 9, the direction changeover valve 7 of high pressure ten and chromatographic column 10 and the column oven 6 with function of temperature control.
The direction changeover valve 7 of high pressure ten fills the large volume sample injection-bis- SPEs on-line coupling as connection maincenter
Put and connected with the high performance liquid chromatography separation device, it is disposed with first interface a by clock-wise order, second connect
Mouthful b, the 3rd interface c, the 4th interface d, the 5th interface e, the 6th interface f, the 7th interface g, the 8th interface h, the 9th interface i and the
Ten interface j.First automatic sampler 4 accesses first interface a, and the both ends of the first solid-phase extraction column 8 are respectively connected to second interface b and the
Nine interface i, chromatographic column 10 access the 3rd interface c, and the both ends of second solid phase extraction column 9 are respectively connected to the 4th d and the 7th interface g, the
Six interface f connection sewer pipes, the second automatic sampler 5 access the 8th interface h, the 5th interface e and the tenth interface j use one short
Pipe interconnects.
In the present embodiment, the direction changeover valve 7 of high pressure ten includes two kinds of connected states, is respectively " 0 " number state and " 1 " number shape
State.
When the direction changeover valve 7 of high pressure ten is " 0 " number state (as shown in Figure 2), first interface a connects with second interface b,
Three interface c connect with the 4th interface d, the 5th interface e connects with the 6th interface f, the 7th interface g connects with the 8th interface h, the 9th
Interface i connects with the tenth interface j, and the first liquid chromatography pump 1, the first automatic sampler 4, the first solid-phase extraction column 8 are linked to be one
Stream, second chromatogram pump 2, the 3rd chromatogram pump 3, the second automatic sampler 5, second solid phase extraction column 9, chromatographic column 10 are linked to be
Another stream.Specifically, the solvent of the first liquid chromatography pump 1 outflow flows through the first automatic sampler 4 in order, into high pressure
The number one interface a of ten direction changeover valves 7, flowed out from second interface b, connect the first solid-phase extraction column 8, into the 9th interface i, from
Tenth interface j flows out, and enters the 5th interface e by connecting pipeline, flows out waste liquid from the 6th interface f；On the other hand, the second chromatogram
The mobile phase of the chromatogram pump 3 of pump 2 and the 3rd push flows through the second automatic sampler 5, into the 8th interface of the direction changeover valve 7 of high pressure ten
H, flowed out from the 7th interface g, into second solid phase extraction column 9, enter back into the 4th interface d, flowed out from the 3rd interface c, into color
Post 10 is composed, detector 11 is finally flowed into and completes analysis.
When the direction changeover valve 7 of high pressure ten is " 1 " number state (as shown in Figure 3), second interface b connects with the 3rd interface c,
Four interface d connect with the 5th interface e, the 6th interface f connects with the 7th interface g, the 8th interface h connects with the 9th interface i, the tenth
When interface j connects with first interface a, the first liquid chromatography pump 1, the first automatic sampler 4, second solid phase extraction column 9 are linked to be one
Bar stream, the second chromatogram pump 2, the 3rd chromatogram pump 3, the second automatic sampler 5, the first solid-phase extraction column 8, chromatographic column 10 are linked to be separately
One stream.Specifically, the solvent of the first liquid chromatography pump 1 outflow flows through the first automatic sampler 4 in order, into high pressure ten
The number one interface a of direction changeover valve 7, flowed out from the tenth interface j, the 5th interface e is entered by connecting pipeline, from the 4th interface d
Outflow, into second solid phase extraction column 9, enters back into the 7th interface g, flows out waste liquid from the 6th interface f；On the other hand, the second color
The mobile phase that spectrum pump 2 and the 3rd chromatogram pump 3 pushes flows through the second automatic sampler 5, and the 8th into the direction changeover valve 7 of high pressure ten connects
Mouth h, flows out from the 9th interface i, into the first solid-phase extraction column 8, flows into second interface b, then is flowed out from the 3rd interface c, enters
Chromatographic column 10, finally flow into detector 11 and complete analysis.
The operation principle of the present embodiment is：When the direction changeover valve 7 of high pressure ten is " 0 " number state, the first liquid chromatography pump 1 is by mesh
Mark thing solution is pushed to the first solid-phase extraction column 8, and after completing extraction process, the switching direction changeover valve 7 of high pressure ten is " 1 " number state,
The first solid-phase extraction column 8 is set to be connected with chromatographic column 10, the mobile phase pushed using the second chromatogram pump 2 and the 3rd chromatogram pump 3 is completed
Desorption separation process, and now, second solid phase extraction column 9 then cleaned in above-mentioned parsing disengaging time section, balance and on
Sample, then switch the direction changeover valve 7 of high pressure ten again, be again introduced into desorption and separation.The front half section of this analysis process, the first solid phase extraction
Cleaning, balance, the loading progress synchronous with the Desorption separation of second solid phase extraction column 9 of post 8 are taken, the direction changeover valve 7 of high pressure ten switches
Enter the second half section afterwards, the Desorption separation of this first solid-phase extraction column of stage 8 and the cleaning of second solid phase extraction column 9, balance, loading
It is synchronous to carry out, realize overlapping analysis.
Embodiment 2
Referring to Fig. 4, it is the fluid communication schematic diagram of the present embodiment.The structure design and connected mode of the present embodiment with
Embodiment 1 is essentially identical, and difference is one of them of the first solid-phase extraction column 8 of embodiment 1 and second solid phase extraction column 9
Zero dead volume bilateral 12 is replaced with, so as to switch to common high performance liquid chromatography separation system.
In the present embodiment, the first solid-phase extraction column 8 of embodiment 1 is replaced with into zero dead volume bilateral 12, so as to switch
The common high performance liquid chromatography separation system arrived includes the second chromatogram pump 2, the 3rd chromatogram pump 3, the second automatic sampler 5, chromatogram
Post 10, the direction changeover valve 7 of high pressure ten and zero dead volume bilateral 12.Second automatic sampler 5 accesses the 8th interface h, and described zero
The both ends of dead volume bilateral 12 are respectively connected to second interface b and the 9th interface i, and the chromatographic column accesses the 3rd interface 7.Specifically,
Second chromatogram pump 2 and the mobile phase of the 3rd chromatogram pump 3 push flow through the second automatic sampler 5, into the direction changeover valve 7 of high pressure ten
8th interface h, flowed out from the 9th interface i, into zero dead volume bilateral 12, flow into second interface b, then flowed out from the 3rd interface c,
Into chromatographic column 10, finally flow into detector 11 and complete analysis.
Relative to prior art, the present invention is connected using simplest hardware device with stream, it is allowed to more than 1mL substantially
The direct injected of product sample, sample introduction again after being concentrated without manual off-line mode to sample；Meanwhile it is equipped with two parallel processing
Solid-phase extraction column, with high performance liquid chromatography be combined, carry out overlapping sample introduction, pre-treatment and separate, shortening the global analysis time,
Greatly reduce holding time of the single sample in whole batch processing sample；Also, this combination equipment can easily switch
For common highly effective liquid phase chromatographic system, the purpose of multifunction system is realized.
The present invention is further described below by way of several experiments.
Experiment 1
This experiment enters the online of 80 kinds of medicines and personal care articles in water-filling using the on-line coupling equipment described in embodiment 1
Analysis.In this experiment, the first solid-phase extraction column 8 and second solid phase extraction column 9 are high pressure resistant reversed phase extraction post, chromatographic column 10
For super-pressure reverse-phase chromatographic column.Object is 80 kinds of medicines and personal care articles, according to the retention characteristic of object, is divided into acidity
It is as follows with alkaline two kinds of loading patterns, specific experiment condition：
Liquid phase chromatogram condition (SPE condition)
First liquid chromatography pump 1 configures low pressure gradient proportioning valve, shares three kinds of optional solution, and wherein A streams are 0.05%
Aqueous formic acid, B streams are acetonitrile/methanol/acetone (the ︰ 1V/V/V of 1 ︰ 1), and C streams are 0.1% ammonia spirit.First liquid phase color
The flow velocity for composing pump 1 is 4mL/min.
When loading condition for it is acid when, balance solution is A stream solution, and the direction changeover valve 7 of high pressure ten is " 0 " number shape during loading
Object solution is pushed to the first solid-phase extraction column 8 with 4mL/min flow velocitys, completed after 2.6min by state, the first liquid chromatography pump 1
Extraction process, while it is " 1 " number state to switch the direction changeover valve 7 of high pressure ten, the first solid-phase extraction column 8 is connected with chromatographic column 10,
The first liquid chromatography pump 1 switches to B streams simultaneously, with 4mL/min flow velocitys cleaning second solid phase extraction column 9, the first liquid after 1min
Phase chromatogram pump 1 switches to A streams, with 4mL/min flow velocitys balance second solid phase extraction column 9.On the other hand, high pressure ten is logical during loading
Switching valve 7 is " 1 " number state, and object solution is pushed to second solid phase extraction by the first liquid chromatography pump 1 with 4mL/min flow velocitys
Post 9, extraction process is completed after 2.6min, while it is " 0 " number state to switch the direction changeover valve 7 of high pressure ten, makes second solid phase extraction column 9
Connected with chromatographic column 10, while the first liquid chromatography pump 1 switches to B streams, and the first solid-phase extraction column is cleaned with 4mL/min flow velocitys
The first liquid chromatography pump 1 switches to A streams after 8,1min, and the first solid-phase extraction column 8 is balanced with 4mL/min flow velocitys.If upper batten
Part is alkalescence, then cleaning solution is C stream solution, and loading process is same as previously described.
Liquid phase chromatogram condition (separation condition)
Chromatographic column：InertSustain AQ-C18(2.10mm I.D.×50mm L.,1.9μm)
Flow velocity：0.4mL/min
Column temperature：40℃
Sample size：5mL
After the switching position of ten direction changeover valve of high pressure 7, the mobile phase of the second chromatogram pump 2 and the push of the 3rd chromatogram pump 3 is opened
Begin desorption and separation process, organic phase initial proportion is 5%, starts linear graded after keeping 0.2min, organic after 8min
Organic Phase Proportion is 100% and keeps 2.1min Phase Proportion after being 80%, 0.1min, and organic Phase Proportion is changed within 0.1min
5% and keep it is constant until analysis terminate.
The chromatogram that the acid loading of this experiment and alkaline loading obtain is distinguished as shown in Figure 5 and Figure 6, the peak of all compounds
Type is symmetrical, sharp, and for the rate of recovery between 60~120%, the global analysis time shortens half than single SPE on-line systems, leads to
Amount doubles.
Experiment 2
This experiment is used for the on-line analysis of 7 kinds of agricultural chemicals in water using the on-line coupling equipment described in embodiment 1.In this experiment
In, the first solid-phase extraction column 8 and second solid phase extraction column 9 are high pressure resistant reversed phase extraction post, and chromatographic column 10 is that super-pressure is anti-phase
Chromatographic column.Object is 7 kinds of agricultural chemicals, and specific experiment condition is as follows：
Liquid phase chromatogram condition (SPE condition)
Cleaning fluid：A-20mM ammonium acetate solutions；B- acetonitriles
Flow velocity：2mL/min
The direction changeover valve 7 of high pressure ten is " 0 " number state during loading, and the first liquid chromatography pump 1 is with 2mL/min flow velocitys by object
Solution is pushed to the first solid-phase extraction column 8, and extraction process is completed after 4min, while it is " 1 " number shape to switch the direction changeover valve 7 of high pressure ten
State, the first solid-phase extraction column 8 is set to be connected with chromatographic column 10, while the first liquid chromatography pump 1 switches to B streams, is flowed with 2mL/min
Speed cleans second solid phase extraction column 9, and the first liquid chromatography pump 1 switches to A streams after 3min, and second is balanced with 2mL/min flow velocitys
Solid-phase extraction column 9.On the other hand, the direction changeover valve 7 of high pressure ten is " 1 " number state during loading, and the first liquid chromatography pump 1 is with 2mL/
Object solution is pushed to second solid phase extraction column 9 by min flow velocitys, and extraction process is completed after 4min, while it is logical to switch high pressure ten
Switching valve 7 is " 0 " number state, second solid phase extraction column 9 is connected with chromatographic column 10, while the first liquid chromatography pump 1 switches to B
Stream, the first solid-phase extraction column 8 is cleaned with 2mL/min flow velocitys, the first liquid chromatography pump 1 switches to A streams after 3min, with 2mL/
Min flow velocitys balance the first solid-phase extraction column 8.
Liquid phase chromatogram condition (separation condition)
Chromatographic column：InertSustain AQ-C18(2.10mm I.D.×50mm L.,3μm)
Mobile phase：A phase -20mM ammonium acetate solutions；B phases-acetonitrile
Flow velocity：0.4mL/min
Column temperature：40℃
Sample size：2mL
After the switching position of ten direction changeover valve of high pressure 7, the mobile phase of the second chromatogram pump 2 and the push of the 3rd chromatogram pump 3 is opened
Begin desorption and separation process, organic phase initial proportion is 10%, starts linear graded after keeping 0.5min, organic after 1min
Organic Phase Proportion is 95% and keeps 0.5min Phase Proportion after being 60%, 3.6min, and organic Phase Proportion is changed into 5% within 0.1min
And keep constant until analysis terminates.
Chromatogram that this experiment obtains as shown in fig. 7, the peak type of all compounds is symmetrical, sharp, the rate of recovery between 60~
Between 120%, the global analysis time shortens half than single SPE on-line systems, and flux doubles.
Experiment 3
This experiment carries out 21 kinds of medicines using the common high performance liquid chromatography function of the on-line coupling equipment described in embodiment 2
With the on-line analysis of personal care articles.In this experiment, using stream shown in Fig. 4, it is dead that the first solid-phase extraction column 8 is replaced by zero
Volume bilateral 12, chromatographic column 10 are super-pressure reverse-phase chromatographic column.Object is 21 kinds of medicines, the μ of direct injected 10 after concentrating sample
L, specific experiment condition are as follows：
Liquid-phase condition
Chromatographic column：Shim-pack XR-ODS III 2.0mm I.D.×75mm L.,1.6μm
Flow velocity：0.3mL/min
Column temperature：40℃
Sample size：10μL
Mobile phase A：0.1% aqueous formic acid
Mobile phase B：Methanol
After the completion of sample introduction, the mobile phase of the second chromatogram pump 2 and the push of the 3rd chromatogram pump 3 starts desorption and separation process, has
After being 35%, 1min there is organic Phase Proportion after organic Phase Proportion is 25%, 4.5min after machine phase initial proportion is 5%, 0.5min
Machine Phase Proportion is 95% and keeps 3min, and organic Phase Proportion is changed into 5% and keeps constant until analysis terminates within 0.1min.
Chromatogram that this experiment obtains as shown in figure 8, its obtained chromatographic isolation result and method index with it is common super
Highly effective liquid phase chromatographic system does not have marked difference, it was demonstrated that the system can simply switch to common ultra performance liquid chromatography system
System.
Embodiment described above only expresses the several embodiments of the present invention, and its description is more specific and detailed, but simultaneously
Can not therefore it be construed as limiting the scope of the patent.It should be pointed out that come for one of ordinary skill in the art
Say, without departing from the inventive concept of the premise, various modifications and improvements can be made, these belong to the protection of the present invention
Scope.
Claims (10)
- A kind of 1. large volume sample injection-bis- SPEs-high performance liquid chromatography on-line coupling equipment, it is characterised in that：Including substantially Product sample introduction-bis- SPEs on-line coupling device and high performance liquid chromatography separation device；The large volume sample injection-bis- SPEs on-line coupling device includes being used for the first liquid for pushing solvent needed for SPE Phase chromatogram pump, the first automatic sampler for being loaded into sample, for stream switching ten direction changeover valves and for being enriched with mesh Mark thing simultaneously removes deimpurity first solid-phase extraction column and second solid phase extraction column；The high performance liquid chromatography separation device includes being used for the second chromatogram pump and the 3rd chromatogram pump, the use for pushing separation mobile phase In the second automatic sampler and chromatographic column for chromatographic isolation that are loaded into sample；Ten direction changeover valve is disposed with ten interfaces by clock-wise order：The first automatic sampler access first connects Mouthful, the first solid-phase extraction column both ends are respectively connected to second interface and the 9th interface, and the chromatographic column accesses the 3rd interface, institute State second solid phase extraction column both ends and be respectively connected to the 4th interface and the 7th interface, the 6th interface connection sewer pipe, described second certainly Dynamic injector accesses the 8th interface, the 5th interface and the connection of the tenth interface.
- 2. large volume sample injection according to claim 1-bis- SPEs-high performance liquid chromatography on-line coupling equipment, it is special Sign is：The state of ten direction changeover valve be first interface connected with second interface, the 3rd interface and the 4th orifice, the When five interfaces and the 6th orifice, the 7th interface and the 8th orifice, the 9th interface and ten orifices, described first Liquid chromatography pump, the first automatic sampler, the first solid-phase extraction column are linked to be extraction stream, second chromatogram pump, the 3rd chromatogram Pump, the second automatic sampler, second solid phase extraction column, chromatographic column are linked to be Desorption separation stream.
- 3. large volume sample injection according to claim 2-bis- SPEs-high performance liquid chromatography on-line coupling equipment, it is special Sign is：The state of ten direction changeover valve is second interface and the 3rd orifice, the 4th interface and the 5th orifice, the When six interfaces connect with the 7th orifice, the 8th interface with the 9th orifice, the tenth interface with first interface, described first Liquid chromatography pump, the first automatic sampler, second solid phase extraction column are linked to be extraction stream, second chromatogram pump, the 3rd chromatogram Pump, the second automatic sampler, the first solid-phase extraction column, chromatographic column are linked to be Desorption separation stream.
- 4. large volume sample injection according to claim 3-bis- SPEs-high performance liquid chromatography on-line coupling equipment, it is special Sign is：The extraction stream is opposite with flow direction of the separation flow path in solid-phase extraction column is desorbed.
- 5. large volume sample injection according to claim 1-bis- SPEs-high performance liquid chromatography on-line coupling equipment, it is special Sign is：The on-line coupling equipment also includes column oven and the detector for detection signal, first solid-phase extraction column, Second solid phase extraction column, ten direction changeover valves and chromatographic column are located in the column oven, and the detector is connected to going out for chromatographic column Mouthful.
- 6. large volume sample injection according to claim 1-bis- SPEs-high performance liquid chromatography on-line coupling equipment, it is special Sign is：Low pressure gradient proportioning valve built in first chromatogram pump, selectivity push the different solvent of at least three kinds of compositions.
- 7. large volume sample injection according to claim 1-bis- SPEs-high performance liquid chromatography on-line coupling equipment, it is special Sign is：The sample size of first automatic sampler is more than 1mL.
- 8. large volume sample injection according to claim 1-bis- SPEs-high performance liquid chromatography on-line coupling equipment, it is special Sign is：First automatic sampler is provided with the changeable quantitative loop for leaving stream.
- A kind of 9. highly effective liquid phase chromatographic system, it is characterised in that：Including the second chromatogram pump described in claim 1, the 3rd chromatogram Pump, the second automatic sampler, chromatographic column and ten direction changeover valves, and duplex fitting；Ten direction changeover valve presses clock-wise order It is disposed with ten interfaces：Second automatic sampler accesses the 8th interface, and the duplex fitting both ends are respectively connected to the Two interfaces and the 9th interface, the chromatographic column access the 3rd interface.
- A kind of 10. highly effective liquid phase chromatographic system, it is characterised in that：Including the second chromatogram pump described in claim 1, the 3rd chromatogram Pump, the second automatic sampler, chromatographic column and ten direction changeover valves, and duplex fitting；Ten direction changeover valve presses clock-wise order It is disposed with ten interfaces：Second automatic sampler accesses the 8th interface, and the duplex fitting both ends are respectively connected to the Four interfaces and the 7th interface, the chromatographic column access the 3rd interface.
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