JP2950697B2 - Direct connection method and apparatus for high performance liquid chromatograph / mass spectrometer - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for directly connecting a high performance liquid chromatograph (hereinafter, referred to as LC) and a mass spectrometer (hereinafter, referred to as MS) and an apparatus therefor. , LC / MS), and a suitable method for directly connecting to LC / MS, which achieves a plurality of analysis modes in one system, and an apparatus therefor.

In particular, (1) a mode in which the eluted components from the LC analysis column are directly sent to the MS ion source section for analysis, and (2) an eluted component is captured in the LC trap column and desalted and washed. Thereafter, the mode is eluted and introduced into the MS ion source and analyzed, (3) the mode and means for concentrating the analysis target component in the LC trap column, and (4) the analysis is performed from the LC sample inlet. MS sample solution directly without passing through the column
This mode is used to optionally and arbitrarily select a mode for introducing and analyzing into the ion source section of (1) and a mode (5) for removing unnecessary components eluted from the LC analysis column.

[0003]

2. Description of the Related Art MS is an analyzer relating to the molecular weight and structure of an organic compound, and is an essential instrument for instrumental analysis in the field of biochemistry. However, analysis of the mixture was difficult. Therefore, chromatography which is excellent in separation and analysis of a mixture, particularly, if soluble in a solvent, regardless of a nonvolatile substance, a thermally unstable substance, an inorganic compound, an organic compound, a low molecular substance, a high molecular substance, etc. The method used in combination with all the LCs to be analyzed was excellent.

A conventional technique will be described with reference to FIGS. FIG. 29 is a block diagram for defining various solutions used in the LC / MS system, FIG. 30 is a block diagram of a conventional LC / MS using a volatile mobile phase system, and FIG.
1 is a block diagram of a conventional LC / MS using a nonvolatile mobile phase system, and FIG. 32 is a conventional flow injection MS.
It is a block diagram of. 29, 30, 31, and 32, 1 is a mobile phase A, 2 is a pump, 3 is a sample inlet, 4 is an analysis column, 9 is an MS ion source, 12 is a trap column, and 25 is a removal of nonvolatile salts. It is a salt system.

In describing LC / MS techniques, many solutions exist and function differently in analytical systems. These solutions are defined according to FIG. (1) The mobile phase A refers to an eluent flowing through the analytical column 4 to separate the mixture into components. (2) Eluent A refers to a liquid in which mobile phase A eluted through analytical column 4. (3) The mobile phase B is a solution that dilutes the eluate A to make it easier for the analyte component to be trapped in the trap column 12.
A solution for washing the trap column 12. (4) The combined liquid is a liquid in which the eluate A and the mobile phase B are mixed and combined. (5) The eluate B refers to a liquid in which the combined liquid flows into the trap column 12 and elutes. (6) The mobile phase C is an eluent for eluting the analysis target component trapped in the trap column 12. (7) The eluent C refers to the mobile phase C that has passed through the trap column 12 and eluted. (8) The eluent D was prepared using the mobile phase B alone as the trap column 1
2 and washed and eluted.

Conventional LC / MS has three analytical modes: (1) analysis using a volatile mobile phase system, (2) analysis using a non-volatile mobile phase system by desalting, and (3) flow injection analysis. Are roughly divided into (1) Analysis using a volatile mobile phase system will be described. The analysis using the volatile mobile phase system can be performed using a simple system. In FIG. 30, the volatile mobile phase A1 for analysis is sent out by the pump 2. The sample solution is injected from the sample inlet 3 with a micro syringe or the like, and separated by the analysis column 4 for each component. The components eluted from the column 4 are supplied to the MS ion source 9 of the LC / MS interface.
And ionized to give a mass spectrum.

This ionization has, up to now, been
The Mosspray method (Thermo Spray)
TSP method, atmospheric pressure chemical ionization method (Atmosphere)
icPressure Chemical Ioniz
)), ie, APCI method, particle vehicle
Method (Particle Beam), that is, PB
Method, continuous flow FAB (Continuous Flo
w Fast Atom Bombardment),
That is, the CFFAB method and the electrospray method (El
(electrospray), that is, the ESI method
well known.

These TSP method, APCI method, PB method, E
The SI method uses an analytical column 4 under atmospheric pressure or reduced pressure.
Is sprayed and ionized. The generated ions are sampled from pores (not shown), and MS gives a mass spectrum. In the CFFAB method, the eluent A is directly introduced into a MS FAB ion source (not shown) at a flow rate of about several μl / min through a thin tube (inner diameter: about several tens of μm) and ionized.

The general LC analysis has a wide range of analysis fields. For example, (a) analysis using a reversed-phase system that does not contain salts. That is, water, acetonitrile, methano-
Analysis using acetic acid, acetic acid, trifluoroacetic acid, formic acid, etc., and (b) analysis of a reversed phase system using a system containing volatile salts. That is, analysis performed using water, acetonitrile, methanol, tetrahydrofuran, ammonium acetate, ammonium formate, etc., and (c) normal phase system analysis. Examples include analysis performed using hexane, chloroform, isopropanol, and the like, and (d) analysis of a sample containing no nonvolatile ionic substance.

Next, (2) analysis of a nonvolatile mobile phase system by desalting will be described. For LC analysis, mobile phase A
May use non-volatile ionic substances,
In some cases, a sample containing a nonvolatile ionic substance is analyzed. If these analysis conditions are applied to LC / MS analysis as they are, non-volatile components are deposited in and around pores (not shown) and tubules (not shown) for sampling ions. Let me.

As a result, ion sampling becomes extremely unstable, and as a result, stable measurement is hindered. Also,
Sometimes the measurement itself is impossible. If the tubule becomes clogged, these parts need to be cleaned or replaced. Further, it is said that such a non-volatile ionic substance interferes with ionization itself. That is, in general, it has been considered that LC / MS analysis cannot use a nonvolatile salt or a nonvolatile buffer.

In order to solve this problem, an improved method has been proposed from the viewpoint of chromatography. In this method, a target component to be analyzed is trapped in a trap column, and non-volatile ionic substances and the like are removed by washing, so-called desalting is performed. Then, a mobile phase C containing no non-volatile ionic substances is used. The captured components are eluted and sent to MS for analysis.

An analysis procedure by desalination will be described with reference to FIG. The mobile phase A1 containing the non-volatile salt and the buffer is sent out by the pump 2 and sent to the desalination system unit 25 via the analysis column 4. The sample is injected from the sample inlet 3 and separated by the analytical column 4 and flows into the desalting system unit 25 together with the eluate A. The desalination system unit 25 includes a plurality of pumps, a plurality of valves, a trap column, and the like (all not shown). The sample that has flowed into the desalting system 25 together with the eluate A is captured by the trap column 12. The trap column 12 is washed and desalted with the mobile phase B.

The analysis target component trapped in the trap column 12 is eluted with the mobile phase C containing no non-volatile components, sent to the MS ion source unit 9, and analyzed by the above (1) volatile mobile phase system. As in the case of (2), it is ionized and gives a mass spectrum. In this manner, the analyte component was eluted from the system using the nonvolatile salt, the nonvolatile buffer, and the like, and LC / MS became possible. As a method related to these, the techniques described in JP-A-3-175355, JP-A-62-138753, and JP-A-62-19758 are known.

Next, the method of (3) flow injection analysis will be described with reference to FIG. In general, the analysis of a pure compound, that is, a pure product, does not require the separation by the LC analysis column 4 as shown in FIG. In FIG. 32, the flow injection analysis method is based on the flow of the mobile phase A1 sent by the pump 2,
This is an analysis method in which a sample solution is directly sent from the sample inlet 3 to the MS ion source unit 9. The sample solution that has flowed into the MS ion source unit 9 is ionized in the same manner as in (1) the analysis using the volatile mobile phase system and (2) the analysis using the non-volatile mobile phase system by desalting. Give mass spectrum.

Further, this analysis system is a general LC /
Before MS, when checking which of the many ionization modes the sample to be measured fits, what solvent system is optimal, what ionization conditions are suitable, etc. It was very convenient. For this reason, this flow injection analysis method is an analysis mode generally used widely, which is equal to or more than (1) separation analysis using an LC analysis column in analysis using a volatile mobile phase system.

However, such conventional (1),
The three analysis modes (2) and (3) are independent of each other, and when one analysis mode is used and then another analysis mode is to be used, It had to be a separate system, and had the disadvantage that the whole system was complicated and expensive.

[0018]

The above-mentioned conventional analysis is as follows.
The three techniques of (1) analysis using a volatile mobile phase system, (2) analysis of a non-volatile mobile phase system by desalting, and (3) flow injection analysis are widely and equally important in LC / MS. It is used by the way. However, there is a problem in the conventional direct connection method of LC / MS and the apparatus therefor that there is no system which considers all the switching of the analysis mode.

Conventional (2) analysis of a non-volatile mobile phase system by desalting comprises a very complicated system such as a plurality of valves, a plurality of pumps, a trap column, and a plurality of capillaries. Therefore, when changing from one analysis mode to another analysis mode, it is necessary to change the system from that system to another system. Or remove the current system and connect another system to the MS. In the case of the former, extremely complicated procedures, labor and time are required. In the latter case, an LC interface must be prepared for each of the many analysis modes, which is also very complicated. In either case, there is a problem that it is not advantageous from the viewpoint of cost and space.

Further, in a system having a large number of analysis modes, it is necessary to prevent the inside of the system from drying up at the time of mode switching, and to prevent memory and clogging. However, there has been a problem that conventional apparatuses and analytical methods have not been sufficiently studied for these. In addition, LC /
MS differs greatly from general LC analysis in that when a certain component is present at a high concentration, the contamination of the analysis system is reduced.
There has been a problem in that no measures have been taken to block these high-concentration components from entering the MS ion source 9.

The present invention has been made in order to solve the above-mentioned problems of the prior art, and takes into account all the switching of the analysis mode, and switches from one analysis mode to another analysis mode. When changing, complicated procedures, labor and time are unnecessary, and it is advantageous in terms of cost and space, and prevents drying, memory and clogging in the system. It is an object of the present invention to provide a method of directly connecting an LC in which a specific high-concentration component is excluded from the MS ion source section 9 to the MS and an apparatus therefor.

[0022]

In order to achieve the above-mentioned object, a direct connection method of LC / MS according to the present invention comprises an LC / MS comprising a sample inlet, an analytical column, a pump and a detector.
, MS, and a high link connecting the two at the interface unit.
Fast and LC, the direct method of the MS, the interface - the face portion, the sample inlet, by Ri is configured to the MS ion source is a plurality of switching valves and a plurality of pumps and the trap column and piping and ionization means, By switching the valve, the following four analysis modes are independently selected and connected directly as needed. (A) The first analysis mode in which the eluate from the LC analysis column is directly or entirely split into the MS ion source, and the trap column is washed with a washing solution. (B) The second analysis mode in which components eluted from the LC analysis column are captured by a trap column, and the MS ion source is washed with a washing solution. (C) The third analysis mode in which components trapped in the trap column are eluted by the eluent and sent to the MS ion source, and the eluate from the LC analysis column is discharged to the outside. (D) A fourth analysis mode in which the trap column and the MS ion source are washed with a washing solution, and the eluate from the LC analysis column is discharged to the outside.

The LC / MS direct connection apparatus according to the present invention has a liquid chromatograph comprising a sample inlet, an analytical column, a pump and a detector , a mass spectrometer, and an interface for connecting the two. A face portion, the interface portion is constituted by a sample inlet, a plurality of switching valves, a plurality of pumps, a trap column, piping, a mass spectrometer as an ionization means, an ion source portion, and a control device ,
The valve is switched by the control device, and the following four analysis modes are independently selected as needed and connected directly. (A) The first analysis mode in which the eluate from the analytical column of the liquid chromatograph is directly or entirely split into the mass spectrometer ion source, and the trap column is washed with a washing solution. (B) The second analysis mode in which components eluted from the analytical column of the liquid chromatograph are captured by the trap column, and the mass spectrometer ion source is washed with a washing solution. (C) The third analysis mode in which the components trapped in the trap column are eluted by the eluent and sent to the ion source of the mass spectrometer, and the eluate from the analytical column of the liquid chromatograph is discharged to the outside. (D) The fourth analysis mode in which the trap column and the mass spectrometer ion source are washed with a washing solution, and the eluate from the analysis column of the liquid chromatograph is discharged to the outside.

That is, as shown in FIG. 31, (2) an analysis system using a volatile mobile phase system, and (3) a flow injection in a desalination system section 25 in the analysis system for a non-volatile mobile phase system using desalination. A function of switching between the analysis system and the (2) non-volatile mobile phase analysis system by desalination is provided. FIG. 1 is a block diagram showing an outline of the present invention. In FIG.
1 is an analysis mobile phase A, 2 is a pump, 3 is a sample inlet, 4 is an analysis column, 9 is an MS ion source, 21, 22, 23,
24 is a three-way switching valve, 25 is a desalination system for nonvolatile salts, 27 is an LC controller, and 91 and 92 are flow passage systems.

(1) In the case of an analysis system using a volatile mobile phase system, a sample is injected from the sample injection port 3, and the three-way switching valve 21 is moved straight to the analysis column 4 on the right side in the figure, and the analysis column 4 And separated. At this time, the three-way switching valves 22 and 23 are switched to allow the eluate A1 to flow through the channel system
Pour into one. The eluate A containing the analysis target component is
Next, it is sent to the MS ion source 9 through the three-way switching valve 23 and the three-way switching valve 24. As a result, the eluate A containing the analysis target component can be directly sent to the MS ion source section 9 to bypass the desalting system section 25 and be ionized to give a mass spectrum.

(2) In the case of a non-volatile mobile phase based analysis system by desalting, the mobile phase A 1 containing a non-volatile buffer and a non-volatile salt is sent out by the pump 2. The sample injected from the sample injection port 3 moves straight through the three-way switching valve 21 toward the analysis column 4 on the right side in the figure, enters the analysis column 4, and is separated for each component. The analysis target component separated by the analysis column 4 advances the three-way switching valve 22 together with the eluate A to the desalination system 25 on the right side in the figure,
It is sent into the desalination system section 25.

The target component for analysis is the desalination system 25
Once trapped in the trap column 12 (not shown) in the inside, it is washed with a mobile phase B (not shown) and desalted. The captured analyte components are then eluted with a mobile phase C (not shown) free of non-volatile salts. The eluate C containing the analysis target component proceeds through the three-way switching valves 23 and 24 to the right side of the MS ion source unit 9 in the figure, is sent to the MS ion source unit 9, and is ionized to give a mass spectrum.

(3) In the case of the flow injection analysis system, the sample solution injected into the sample injection port 3 is transferred from the three-way switching valve 21 to the flow path system 92 and further to the three-way switching valve 21 together with the carrier solvent, that is, the mobile phase A1. It is sent to the MS ion source unit 9 via the switching valve 24 and is ionized to give a mass spectrum. Thereby, the analysis column 4 and the desalting system unit 25 can be bypassed, and the sample solution is directly sent to the MS ion source unit 9 and ionized to give a mass spectrum.

When the flow path is switched by a plurality of three-way switching valves, the three analysis systems (1) and (2)
(3) can be switched. Switching of these valves can be performed freely and freely by the LC controller 27.

[0030]

The function of each of the above technical means is as follows. According to the configuration of the present invention, (1) in the case of analysis using a volatile mobile phase system, it is also possible to directly send an analysis target component to MS after column separation and analyze it. In the case of (2) a non-volatile mobile phase-based analysis system by desalting, after separating the components in the sample, the analysis target component is captured, only the non-volatile salt is removed, and the analysis target component is subjected to MS. Can be sent to Further, (3) the flow injection analysis system can be configured by switching valves, and can be used together for measurement that does not require separation of columns, search for system optimization conditions, and the like.

More specifically, in LC / MS, the required analysis mode and its means can be achieved in one system by switching valves, increasing the sensitivity by backflushing, and reducing the concentration of low-concentration samples. Concentration and further LC / M
Components that elute into the S system and may contaminate the ion source of the MS can be freely removed. Here, the reason why the term "MS ion source unit" is used is that there are many methods for ionizing LC / MS as described above, and these methods are widely used, and cannot be specified. In all these ionization modes, the present invention works effectively.
Furthermore, during a series of analysis operations, the solvent flows constantly throughout the system without interrupting the LC, the pump, and the like, thereby preventing contamination and clogging of the entire system.

[0032]

Embodiments of the present invention will now be described with reference to FIGS.
This will be described with reference to FIG. [Embodiment 1] FIG. 2 shows an LC / LC according to an embodiment of the present invention.
FIGS. 3, 4, 5, and 6 show an MS analysis system, and FIGS. 3, 4, 5, and 6 show analysis systems of the first, second, third, and fourth analysis modes of the LC / MS according to the embodiment of FIG. FIG. 7 is an explanatory view of analysis mode switching in LC / MS using a nonvolatile mobile phase system, FIG. 8 is an explanatory view of analysis mode switching of front removal by a volatile mobile phase system, FIG. 9 is an explanatory diagram of the analysis mode switching of the unnecessary component removal, and FIG. 10 is an explanatory diagram of the analysis mode switching of the sample concentration by repetition.

In FIGS. 2, 3, 4, 5, and 6, 1 is a mobile phase A, 2 is a pump, 3 is a sample inlet, 4 is an analytical column,
5 is a detector, 6 is a mobile phase B, 7 is a pump, 8 is a tee,
9 is an MS ion source, 10 is a pump, 11 is a mobile phase C,
12 is a trap column, 18 is a tie, 19 is a branch resistor, DR1... DR4 is a drain, V-1 is a multi-way switching valve (hereinafter referred to as a valve), V-2 is a valve, 1
a... 1q are switching ports of the valve V-1, 2a.
2q is the switching port of valve V-2, 30 ... 45
Are the valve V-1, the valve V-2, the pump 7, the tee
8, MS ion source 9, pump 10, mobile phase C11, trap column 12, tee-18, each drain DR1.
A channel capillary system (hereinafter, referred to as DR4) for connecting components such as DR4.
Tubules).

As shown in FIG. 2, the mobile phase A1 for analysis stored in the eluent tank is sent out by the pump 2. The sample solution is injected from the sample injection port 3 by a microsyringe or the like, and is continuously analyzed by the mobile phase A1 through the analysis column 4.
Is transferred to The transferred sample is separated and eluted in the analysis column 4 for each component. The eluted components are detected by the detector 5
Is sent to the valve V-2 through the thin tube 31, and reaches the switching port 2i.

As the thin tube, any stainless steel tube having an inner diameter of 0.1 mm to 1.0 mm can be used, and a stainless tube having a diameter of 0.1 mm to 0.5 mm is preferably used. The valve V-1 and the valve V-2 are 8-way and 12-way switching valves, respectively. Switching of the flow path system
The operation is performed by a manual cock or a motor drive controlled by a CPU, an LC controller (neither is shown), or the like. These valves can alternately switch between a flow path shown by a solid line and a flow path shown by a broken line. The state of the solid line is I, and the state of the broken line is II.

Since these flow path systems have two valves in the system, the valve V-1 has two states of I and II, the valve V-2 has two states of I and II, A total of four valve state combinations are possible. As described above, the two valves use 8-way or 12-way switching valves, but use valves having more switching ports (for example, Hitachi K-1600 16-way valves). It is also possible.

Next, each analysis mode will be described. (1) In the case of the first analysis mode In this analysis mode, the valve V-1 is at I and the valve V-2 is at I. In FIG. 3, the connection state of the valve switching port is indicated by a solid line. For example, valve V
-1 of 1a and 1b are connected, but 1
Ports a and 1h are not connected. The eluate A eluted from the analytical column 4 is supplied to the detector 5, the capillary 31, the switching ports 2i and 2j, the capillary 41, and the switching ports 1f and 1
e, through the thin tube 38, the switching ports 2a and 2b, and the thin tube 37, are sent to the MS ion source section 9 and ionized to finally give a mass spectrum.

Mobile phase C11 for elution free of nonvolatile salts
(For example, a 1: 9 solution of water and acetonitrile)
Are sent out by the pump 10 and the thin tube 36, the switching port 2
c, 2d, capillary 35, switching ports 1h, 1g, capillary 4
2 through DR2 to the outside. The role of the mobile phase C11 in the first analysis mode is to clean the channel system.

The mobile phase B6 (for example, water) is sent out by the pump 7 and split into two at the tee-8. One of the branched mobile phases B6 is composed of a thin tube 39 and switching ports 21 and 2.
k, and is discharged from DR3 to the outside via the thin tube 40. The other mobile phase B6 branched at T-8 has a branch resistance of 1
At 9 the flow rate is limited and the tee 18 branches further into two.

The mobile phase B6 of one flow path branched by the tee 18 reaches the thin tube 32 and the switching ports 2h and 2g. However, the switching port 2g is stopped because it is sealed. Therefore, the mobile phase B6 of the other flow path branched at the tee 18 includes the thin tube 33, the switching port 1c,
1d, flows into the trap column 12 from the direction of the arrow shown in FIG. The eluate D from the trap column 12 is discharged from the drain DR1 to the outside via the thin tube 43 and the switching ports 1a and 1b. The branch resistor 19 adjusts the flow rate between the flow path 39 and the flow paths 32 and 33, and may be replaced with a needle valve.

In summary, the analysis column 4
The eluate A from is supplied continuously to the MS ion source unit 9 to give a mass spectrum of the target component. The trap column 12 is washed with the mobile phase B6 in the forward direction of the arrow shown. The elution mobile phase C11 cleans the channel.

(2) In the case of the second analysis mode In this analysis mode, the valve V-1 is I and the valve V-2 is I
In the state II, the analysis target component is the trap column 12
Is captured by the In FIG. 4, the eluate A eluted from the analytical column 4 is sent out to the capillary 32 via the detector 5, the capillary 31, and the switching ports 2i and 2h.

The mobile phase B6 is sent out by the pump 7, and branches off at a tee-8. One branched mobile phase B6
Is the eluate A in the tee 18 through the branch resistance 19
And the eluate A is diluted to form a combined liquid. The combined liquid is sent from the thin tube 33 to the valve V-1, and further flows into the trap column 12 through the switching ports 1c and 1d and the thin tube 45 in the direction of the arrow shown in the figure.

The analysis target component dissolved in the combined liquid is immediately captured by the trap column 12. The combined liquid other than the captured components is almost eluate B, and the capillary 43,
Through the switching ports 1a and 1b, the air is discharged from the drain DR1 to the outside. Mobile phase C for elution without nonvolatile salts
Numeral 11 designates a thin tube 36 and a switching port 2 by the pump 10.
c, 2b, and sent to the MS ion source unit 9 via the thin tube 37 to wash the MS ion source 9 unit and its channel system.

The other mobile phase B6 branched at T-8
Are the thin tube 39, the switching ports 21 and 2a, the thin tube 38, the switching ports 1e and 1f, the thin tube 41, and the switching port 2j,
2k, is discharged from the drain DR3 to the outside through the thin tube 40.

In summary, the analysis column 4
The analysis target component eluted from is trapped in the trap column 12, and the MS ion source unit 9 is washed with the mobile phase C11. The next third analysis mode will be applied.

(3) In the case of the third analysis mode In this analysis mode, the valve V-1 is in the state of II and the valve V-2 is in the state of I, and is captured by the trap column 12 in the second analysis mode. This is a case where the analyzed target component is eluted and introduced into the MS ion source unit 9 after the elution. In FIG. 5, the eluate A of the analytical column 4 is discharged from the drain DR2 to the outside via the switching ports 2i and 2j, the thin tube 41, the switching ports 1f and 1g, and the thin tube 42.

Mobile phase C11 for elution free of nonvolatile salts
Are pumped out by the pump 10, and the thin tube 36, the switching ports 2c, 2d, the thin tube 35, the switching ports 1h, 1a,
It flows into the trap column 12 through the thin tube 43 in the direction indicated by the arrow. Mobile phase C in this third analysis mode
The flow direction of the mobile phase C11 in the first analysis mode and the second analysis mode is opposite to the flow direction of the mobile phase C11.

In the second analysis mode, the trap column 1
The analysis target component captured in step 2 is eluted in the opposite direction to the capture, desalting, and washing. The eluate C containing the eluted and eluted analysis target components is supplied to the capillary tube 45, the switching ports 1d and 1e,
After passing through the thin tube 38, the switching ports 2a and 2b, and the thin tube 37, M
It enters the S ion source unit 9. The analysis target component is ionized by the MS ion source unit 9 to give a mass spectrum.

On the other hand, the mobile phase B6 is sent out by the pump 7 and is branched into two at the tee 8. Among them, the mobile phase B in one of the flow paths comprises the thin tube 39 and the switching ports 21 and 2k. Through the drain DR3. The mobile phase B of the other branched flow path is discharged from the drain DR1 to the outside through the branch resistance 19, the tee 18, the thin tube 33, the switching ports 1c and 1b, and the thin tube 44.

In summary, the eluate A from the analytical column 4 is discharged to the outside. The analysis target component captured in the trap column 12 is eluted by backflushing with the mobile phase C11. And this analysis target component is
It is introduced into the MS ion source unit 9 and gives a mass spectrum. Further, the channel is washed with the mobile phase B6.

(4) Fourth Analysis Mode In this fourth analysis mode, the valve V-1 is II and the valve V-2 is II.
, The trap column 12, the MS ion source 9
Is washed, and if the trap column 12 has captured the analysis target component, it is a case of washing and desalting without eluting the analysis target component. In FIG. 6, the eluate A from the analysis column 4 passes through the capillary 31, the switching ports 2i and 2h, and the capillary 32, and is then sent to the tee 18.

The mobile phase B6 is sent out by the pump 7, and is branched into two at the tee-8. One of them is
The effluent A is sent to the tee 18 via the branch resistor 19 and merges with the eluate A at the tee 18. By this confluence, the eluate A is diluted by the mobile phase B6. Then, the combined liquid is discharged from the drain DR1 to the outside through the thin tube 33, the switching ports 1c and 1b, and the thin tube 44.

The mobile phase B6 in the other flow path branched by the tee 8 is composed of the thin tube 39, the switching ports 21 and 2
a, the thin tube 38, the switching ports 1e and 1d, and the thin tube 45, and flows into the trap column 12 from the direction of the arrow shown in the figure. The eluate D from the trap column 12 is supplied to the capillary 43,
Switching port 1a, 1h, thin tube 35, switching port 2
d, 2e, and are discharged from the drain DR4 to the outside through the thin tube 34. Mobile phase C11 is pumped out by pump 10,
After passing through the thin tube 36, the switching ports 2c and 2b, and the thin tube 37, M
The S ion source unit 9 is cleaned.

In summary, the eluate A from the analytical column 4 is discharged to the outside. The trap column 12 is washed with the mobile phase B6 from the forward direction. The MS ion source unit 9 has been washed with a mobile phase C11 containing no nonvolatile salt. When the analysis target component is trapped in the trap column 12, the analysis target component can be washed and desalted without being eluted by selecting the polarity of the mobile phase B6.

Next, using the present embodiment, the first, second, third, and fourth analysis modes described above are combined and utilized, and FIG.
Various specific analyzes will be described with reference to FIGS. (1) Procedure for introduction into MS after desalting In FIG. 7, the chromatogram shown at the top is the detector 5
Is a liquid chromatogram detected by X, X is a target analysis component, and Y, Z, V, and W are unnecessary components. The analysis mode switching by the valve operation shown at the bottom shows a procedure for introducing the salt into the MS after desalting. It is assumed that the liquid chromatogram shown at the top was obtained by analysis of a mobile phase system containing a nonvolatile ionic substance. The illustrated component X is an analysis target component, and a procedure for desalting this component X for LC / MS measurement and introducing it into MS will be described.

First, prior to the analysis, the trap column 12
The valve is switched to be pre-processed and put into a fourth analysis mode. The sample was then injected until the retention time t ₁ immediately before the X component is eluted fourth analysis mode - to leave the mode. Unwanted components eluted from the analyzer column 4 during this time
V and Y are all discarded to the outside together with the eluate A. During this time, the trap column 12 has been washed with the mobile phase B6.

Next, the holding time t _1, the second analysis mode - switched to de. The analysis target component X is captured by the trap column 12. Analysis time t ₂ that elution of target component X has been completed, the analysis mode - de fourth analysis mode again - is switched to de.

This fourth analysis mode is held for several minutes, actually for about 3 to 5 minutes, and is transferred to the trap column 12.
The mobile phase A1 is extruded by the mobile phase B6, and is desalted and washed. During the desalination washing, the analysis target component X is
It remains trapped in the trap column 12.

Next, at t ₃ , the analysis mode is switched to the third analysis mode, and the analysis target component X trapped in the trap column 12 is backflushed and eluted by the mobile phase C11. You. The eluate C containing the analysis target component X is sent to the MS ion source to give a mass spectrum.

[0061] In t ₄ when infeed of analysis target component to the MS has been completed, the analysis mode - de are quaternary analysis mode again - switched to de, trap column 12 is washed with the mobile phase B6, following LC / MS Prepare for measurement. In this analysis, only the analysis target component X is captured by the trap column 12 and gives a mass spectrum after desalting and elution. On the other hand, unnecessary components V, Y, other than the analysis target component X
Z and W are all discharged outside.

(2) L in a mobile phase system containing no nonvolatile salt
C / MS Procedure In FIG. 8, the chromatogram shown at the top is the detector 5
Is a liquid chromatogram detected in the four components,
V, X, Y, and Z are sequentially eluted from the analytical column 4. X, Y, and Z are the components to be analyzed, and component V is an unnecessary component eluted from the void volume. The analysis mode switching by the valve operation shown at the bottom shows a front removal procedure in LC / MS using a volatile mobile phase system.

In the case of reverse phase chromatography, ionic compounds, salts, compounds having extremely high polarity and the like are eluted without being retained on the column. Generally, these compounds are
When directly introduced into the ion source 9, the ion sampling pores (not shown) may be clogged or the MS ion source 9 may be clogged.
To contaminate.

Even if mass spectra of these components are obtained, in many cases, the mass spectra do not provide significant information because they are mixtures. Therefore, if the components eluted by the void volume are removed and are not introduced into the MS ion source, the MS ion source 9 is kept clean for a long time and there is no loss of information.

In FIG. 9, L is the target component for analysis, O,
P and Q are trace components, M is a main component, V is a component eluted by void volume, and both M and V are unnecessary components. In the case of impurity analysis, when the main component M is introduced into the MS ion source 9, the subsequent trace components O, P, Q, etc. are difficult to identify due to the carryover of the component M. There is. Such a large amount of the component M is repeatedly
When introduced into the ion source 9, contamination of the MS ion source 9 is caused at an early stage, and the cleaning interval is shortened. In order to solve this, a large amount of the non-analytical component M may be discharged to the outside without being introduced into the MS ion source.

The void volume shown in FIGS.
V eluted by the system and non-analytical components M present in large amounts
Can be discharged to the outside by the fourth analysis mode. During this time, the MS ion source 9 is filled with the solvent, that is, the mobile phase C11.
Is prevented from becoming unstable and the washing is performed.

After the unnecessary components M and V elute, the eluate A from the analytical column 4 is introduced into the MS ion source section 9 in the first analysis mode, and a mass spectrum is obtained. If the retention time of the unnecessary component is known in advance, the analysis mode can be switched based on the retention time.

Referring to FIG. 8, the removal of the component V eluted by the void volume will be described. First, the fourth analysis mode is set, and the sample solution is injected from the sample inlet 3. The fourth analysis mode - de is maintained until the holding time t _1. During this time, the eluted unnecessary components V are discharged to the outside.

Next, the holding time t _1, a first analysis mode - to de. The eluate A is directly led to the MS ion source unit 9. The analysis target components X, Y, and Z are ionized one after another to give a mass spectrum. Next, at the holding time t ₂ ,
By entering the fourth analysis mode, the LC / MS is completed, and the column is washed and further prepared for the next analysis.

FIG. 9 shows an example of removal of the component V eluted by the void volume and the non-analytical component M present in a large amount. The analysis mode is set to the fourth analysis mode, and sample injection is performed. Fourth Analysis mode until the retention time t ₁ - to maintain de, discharging undesired components V outside. Next, the holding time t
_{In step 1} , the first analysis mode is switched, and the analysis target component L
Can be measured.

At t ₂ immediately before the elution of the main component M, the mode is switched to the fourth analysis mode, and the main component M is maintained in the fourth analysis mode until a retention time t _{3 at} which the elution of the main component M is completed. Is discharged. In the holding time t _3, the first analysis mode again - to set the de analysis target component O, P, measured Q performed. In holding time t _4, a fourth analysis mode - Set to de,
Prepare for the next measurement. In this way, the first analysis mode and the fourth analysis mode are selectively used, and the components separated on the chromatography can be freely introduced into and removed from the MS ion source.

(3) Flow Injection Analysis Procedure In LC / MS analysis, it may be sufficient to analyze only a pure product or to obtain only a mass spectrum from a mixture. In such a case, separation by an LC column is unnecessary. Also, the type of ionization of the sample, for example, A
The flow injection method is convenient for selecting PCI and ESI, optimizing ionization conditions, and selecting positive and negative ionization modes.

This flow injection system comprises:
In FIG. 2 described above, an injection port 3 'is provided in a part of the flow channel system of the mobile phase C11. As shown in FIG. 6, when the analysis mode is set to the fourth, the analysis column 4 is always washed with the mobile phase A1 for analysis. Further, the trap column 12 is also washed with the mobile phase B6. The MS ion source unit 9
Washed with mobile phase C11. The sample solution is stored in a small tube 36
Is repeatedly sent from the injection port 3 'provided in the middle to the MS ion source unit 9, and a mass spectrum can be obtained simply and in a short time.

In the second analysis mode shown in FIG.
Similarly, analysis by the flow injection method can be performed. However, in the second analysis mode, the analysis column 4
The eluate A from is diluted with the mobile phase B6 and flows into the trap column 12. On the other hand, in the fourth analysis mode,
The analysis column 4 and the trap column 12 are completely independent flow path systems and have no relation to each other. Therefore, the fourth analysis mode is suitable for the flow injection analysis.

(4) Procedure for Concentration Analysis of Analytical Target Component In the LC / MS measurement, the concentration of the analytical target component is low.
Analysis can be difficult. In such a case, it becomes possible to concentrate the analysis target component by the system of this embodiment.

The procedure will be described with reference to FIG. L
In the C / MS, the sample is set in the fourth analysis mode and injected. At time t ₁ immediately before the elution of the analysis target component X, the mode is switched to the second analysis mode, and the trapping column 12 captures the analysis target component X. When the elution of the analysis target component X is completed t
_{At 2} , switch to the fourth analysis mode. After elution of components other than the analysis target component X is completed, the sample is injected again.

As described above, by alternately switching between the fourth analysis mode and the second analysis mode and repeating the injection, the analysis target component X is captured and concentrated in the trap column 12. When the analysis target component X is captured and the concentration is completed, t ₀
Third Analysis mode in - switching to de, analysis target component X is eluted by backflushing, Ru sent to the MS ion source section 9.

By repeating such sample injection, the bandwidth of the analysis target component X trapped in the trap column 12 is wider than that in a single trap, but the type, length, and mobile phase of the trap column 12 are different. By examining the polarity and dilution ratio (ratio) of B, the spread of the bandwidth can be reduced as much as possible. In addition, elution by backflushing has the function of canceling the expansion of the bandwidth.

As described above, according to [Example 1], desalting analysis of a sample, analysis of a mobile phase system containing no non-volatile salt, removal of unnecessary components, flow injection analysis, and concentration of trace components were also performed. It becomes possible by simple valve operation.

[Embodiment 2] Next, an LC / MS according to another embodiment of the present invention will be described. Figures 11, 12, 13, 1
FIG. 4 is an explanatory view showing an LC / MS analysis system according to another example of the present invention. In the figure, the same reference numerals as those in FIGS. Only new codes will be described. In FIGS. 11, 12, 13, and 14, V-1, V-2, and V-3 are hexagonal switching valves,
.. 3f are switching ports of the hexagonal switching valve V-3, and 80, 81... 88, 89 are thin tubes.

The LC / MS system according to the present embodiment is a system using three six-way valves. The system shown in FIG. 11 corresponds to the first analysis mode of [Embodiment 1]. The mobile phase A1 for analysis is sent out by the pump 2. The sample introduced from the sample inlet 3 is separated for each component in the analytical column 4 and detected by the detector 5. The eluate A containing the analysis target component that has exited the detector 5 is transferred from the small tube 80 to the switching ports 1a and 1f of the valve V-1, the small tube 84, the switching ports 1e and 1d, the small tube 83, and the valve V-2. Through the switching ports 2d and 2c, and further through the thin tube 87.

During this period, the mobile phase B6 sent out by the pump 7 is replaced with the tee-8 and the capillary 8 by the trap column 12.
5. It is sent through the switching ports 2a and 2b and the thin tube 86, and is washed from the direction of the arrow shown in the figure. Note that the valve V-
The third port 3c is sealed. This first analysis mode
In this method, the analysis target component eluted from the analysis column 4 can be directly guided to the MS ion source 9 for analysis.

The analysis system shown in FIG.
Corresponds to the second analysis mode. The eluate A containing the analysis target component eluted from the analysis column 4 is sent out to the thin tube 85 via the switching ports 1a and 1b of the valve V-1 and the thin tubes 81, 3e and 3d. Further, the eluate A merges with the mobile phase B6 at the T-8 provided in the middle of the thin tube 85, and is diluted by the mobile phase B6. The generated combined liquid is supplied to the switching ports 2a and 2b of the valve V-2, the thin tube 8
6 and flow into the trap column 12 in the direction of the arrow. The analysis target component is captured by the trap column 12.

The eluate B not captured by the trap column 12 is supplied to the small tube 88, the switching ports 2e and 2f, and the small tube 89.
Through the drain DR1 to the outside. During this time, the MS ion source 9 is supplied with the mobile phase C11 sent out by the pump 10 and sent through the thin tube 82, the switching ports 1c and 1d, the thin tube 83, the switching ports 2d and 2c, and the thin tube 87. Has been cleaned. In the second analysis mode, the analysis target component eluted from the analysis column 4 can be captured by the trap column 12.

The analysis system shown in FIG. 13 corresponds to the third analysis mode. The mobile phase C11 is sent out by the pump 10 and enters the valve V-1 via the thin tube 82. Next, it enters the valve V-2 via the switching ports 1c and 1d and the thin tube 83, and further flows into the trap column 12 from the switching ports 2d and 2e and the thin tube 88 in the direction of the arrow shown in the figure.

The analyte of interest captured by the mobile phase C11 is eluted by backflushing. The eluate C containing the analysis target component enters the valve V-2 from the capillary 86. Next, it enters the MS ion source section 9 through the switching ports 2b and 2c and the thin tube 87. During this time, the eluate A from the analytical column 4 is supplied to the detector 5, the thin tube 80, the switching ports 1a and 1b, the thin tube 81, and the switching ports 3e and 3f.
Through the drain DR2.

In the third analysis mode, the analysis target component trapped in the trap column 12 is eluted by backflushing and introduced into the MS ion source section 9 to obtain a mass spectrum.

The analysis system shown in FIG. 14 corresponds to the fourth analysis mode. The mobile phase B6 is sent out by the pump 7 and enters the valve V-2 from the tee 8 via the thin tube 85. The switching port 3c of the valve V-3 is sealed. Further, the mobile phase B6 includes the switching ports 2a, 2
b, the liquid flows into the trap column 12 through the thin tube 86 from the direction of the arrow shown in FIG. The eluate D from the trap column 12 is discharged from the drain DR1 to the outside through the small tube 88, the switching ports 2e and 2f, and the small tube 89.

The mobile phase C11 is sent out by the pump 10, and the capillary 82, the switching ports 1c and 1d, the capillary 83,
It is sent to the MS ion source 9 via the switching ports 2d and 2c and the thin tube 87. Thereby, the MS ion source unit 9 is cleaned. In this analysis mode, the trap column 12 can be washed, and the trapped analysis target component can be desalted.

As described above, similarly to [Embodiment 1],
In [Example 2], the second, third, and fourth analysis modes
By switching the mode, it becomes possible to capture and desalinate the target component from the analysis system containing the non-volatile component, and to introduce the target component into the MS. In the analysis system containing no non-volatile components, components eluting in the void volume and a large amount of non-analytical components are removed by using the first analysis mode and the fourth analysis mode.
Only the analyte component can be directly introduced into the MS.
In the fourth injection mode, the flow injection analysis can be easily performed by installing the sample injection port 3 'on the thin tube 82 in the flow path of the eluent C11 and injecting the sample.

[Embodiment 3] An LC / MS according to still another embodiment of the present invention will be described. This embodiment shows an example in which two 16-way valves are used. 15, 16, 17, 18
FIG. 7 is an explanatory diagram showing an LC / MS analysis system according to still another example. In the figure, the same reference numerals as those in FIGS. Only new codes will be described. In FIGS. 15, 16, 17, and 18, V
-1, V-2 are 16-way switching valves, 1a, 1b
1q is the switching port of the 16-way switching valve V-1
, 2a, 2b... 2q are 16-way switching valves V-
The switching ports 2, 50, 51, 72, 73 are thin tubes.

As in [Embodiment 1] and [Embodiment 2],
Four analysis modes are possible: first, second, third and fourth. In the present embodiment, in the fourth analysis mode, the sample injection port 3 is independent of the analysis column 4. By using this sample injection port 3, flow injection becomes possible. That is, the difference from the above-mentioned [Example 1] and [Example 2] is that the sample inlet 3 for LC separation analysis and the sample inlet for flow injection are shared by one sample inlet. It is that you are. this is,
In the case of a system using a large-scale sample injection method such as an autosampler for sample injection, there is an advantage that operability is improved because piping change is not required between LC separation analysis and flow injection analysis. .

FIG. 15 corresponds to the first analysis mode. The mobile phase A1 for analysis is sent out by the pump 2. Thin tube 6
3. Valve V-1 switching ports 1q, 1p, thin tube 5
1 and reaches the sample injection port 3. The sample is injected from the inlet 3 and flows into the analysis column 4 via the thin tube 64, the switching ports 1b and 1a, and the thin tube 50.

The sample is separated for each component and eluted from column 4. After the detection target component contained in the eluate A is detected by the detector 5, the switching port 12n, 2 of the valve V-2 is used.
m, and enters the valve V-1 again through the thin tube 56. further,
Via the switching ports 1j, 1i, the valve V-2 is again entered at the thin tube 57, and the switching ports 2a, 2b, the thin tube 7
After passing through 3, it is introduced into the MS ion source unit 9.

During this time, the mobile phase B6 is sent out by the pump 7, and the tee 8 makes the thin tube 65, the switching port 2
The liquid is branched into the liquid passing through q, 2p and the thin tube 66, and the liquid passing through the branch resistance 19, but merges again at the tee 18. The mobile phase B6 that has joined again passes through the capillary 59, the switching ports 1g and 1h, and the capillary 58, and the trap column 1
2 is washed from the illustrated arrow direction. After washing, eluate D
Is discharged from the drain DR1 to the outside via the thin tube 61, the switching ports 1e and 1f, and the thin tube 60. Mobile phase C1
1 is pumped out by a pump 10 and branched into two at a tee-20.

One of the branched flow paths enters the valve V-1 through the small tube 70, the switching ports 2f and 2e, and the small tube 54, and passes from the drain DR2 through the switching ports 11 and 1k and the small tube 55. It is discharged outside. The other channel is
The tee 20 branches into a thin tube 52, and the switching port
1n, 1m, thin tube 53, switching ports 1d, 1c,
Drain D through thin tube 62 and switching ports 2c and 2d
It is discharged outside from R3. In the first analysis mode, the eluate A from the analysis column 4 is used as it is in the MS ion source section 9.
Can be sent to During this time, the trap column 12, the thin tube, and the like are washed with the solvent.

FIG. 16 corresponds to the second analysis mode. The eluate A eluted from the analytical column 4 flows into the tee 18 via the switching ports 2n and 2p of the valve V-2 and the thin tube 66. The mobile phase B6 is sent by the pump 7 through the tee 8 and the branch resistor 19, and merges with the eluate A at the tee 18, and is diluted to become a merged liquid.

The combined liquid flows from the small tube 58 to the trap column 12 through the small tube 59 and the switching ports 1g and 1h of the valve V-1 in the direction of the arrow shown in the figure. After the analysis target component is captured by the trap column 12, the uncaptured eluate B is supplied to the capillary 61, the switching ports 1e, 1
f, It is discharged from the drain DR1 to the outside through the thin tube 60.

The mobile phase C11 is sent out by the pump 10, and is branched into two at a tee-20. The mobile phase C11 in one of the branched flow paths is a thin tube 70, a switching port 2
f, 2g, and discharged from the drain DR4 to the outside via the thin tube 69. The mobile phase C11 of the other flow path branched at the tee 20 reaches the valve V-1 via the thin tube 52, and switches to the switching ports 1n and 1m, the thin tube 53, and the switching port 1.
It enters the MS ion source section 9 via d and 1c, the thin tube 62, the switching ports 2c and 2b, and the thin tube 73.

In the second analysis mode, the analysis target component eluted from the analysis column 4 can be captured in the trap column 12. During this time, the MS ion source unit 9 has been washed with the mobile phase C11.

FIG. 17 corresponds to the third analysis mode. The eluate A eluted from the analytical column 4 is supplied to the switching port 2
n, 2m, the thin tube 56, the switching ports 1j, 1k, and the thin tube 55, and are discharged from the drain DR2 to the outside. The mobile phase B6 is sent out by the pump 7, and is branched at T-8 into a liquid passing through the thin tube 65, the switching ports 2q and 2p, the thin tube 66, and a liquid passing through the branch resistance 19. Then, they merge again at the tee-18. The combined mobile phase B6 is discharged from the drain DR1 to the outside through the thin tube 59 and the switching ports 1g and 1f.

The mobile phase C11 is sent out by the pump 10,
At tee-20, it branches into two. One of the branched mobile phases C11 is composed of a thin tube 70, switching ports 2f and 2
e, thin tube 54, switching ports 11 and 1m, thin tube 53,
It flows into the trap column 12 from the direction of the arrow shown in the figure via the switching ports 1d and 1e and the thin tube 61 in order.

Thus, the analysis target component captured in the trap column 12 is eluted by backflushing. The eluate C containing the analysis target component is supplied to the capillary 5
8, is sent to the MS ion source 9 through the switching ports 1h and 1i, the thin tube 57, the switching ports 2a and 2b, and the thin tube 73 to give a mass spectrum.

The mobile phase C11 of another flow path system branched at the tee 20 is composed of a thin tube 52, switching ports 1n and 1
p, the sample inlet 3 is washed through the thin tube 51,
The water is discharged from the drain DR3 to the outside through the switching ports 1b and 1c, the thin tube 62, and the switching ports 2c and 2d.

In the third analysis mode, the target component to be analyzed captured by the backflushing is eluted to give a mass spectrum. Then, the other channel system is washed, and the eluate A from the analysis column 4 is discharged to the outside.

FIG. 18 corresponds to the fourth analysis mode. The mobile phase A1 for analysis is sent out by the pump 2 and
3. Enter the analysis column 4 via the switching ports 1q and 1a and the capillary 50. Further, the drain D is passed through the detector 5, the thin tube 67, and sequentially through the switching ports 2n and 2p, the thin tube 66, the tee 18, the thin tube 59, and the switching ports 1g and 1f.
It is discharged from R1 to the outside.

The mobile phase C11 is sent out by the pump 10, and is branched into two at a tee-20. The mobile phase C11 in one of the branched flow paths comprises a thin tube 70, a switching port 2f,
2 g is discharged from the drain DR4 to the outside through the thin tube 69. The mobile phase C11 in the other branched flow path enters the valve V-1 from the thin tube 52, and is switched to the switching port 1n, 1.
p, capillary 51, sample inlet 3, capillary 64, switching port
1b, 1c, a thin tube 62, switching ports 2c, 2b,
It enters the MS ion source 9 via the thin tube 73.

The mobile phase B6 is supplied by the pump 7 to the switching ports 2q, 2a, the thin tube 57, and the switching ports 1i, 1.
h, and is sent to the trap column 12 via the thin tube 58 sequentially. The mobile phase B6 cleans the trap column 12 from the direction shown by the arrow. After washing, the eluate D is transferred to the thin tube 6
1, switching ports 1e and 1d, thin tube 53, switching ports 1m and 11l, thin tube 54, switching ports 2e and 2
d, the water is sequentially discharged from the drain DR3 to the outside via the thin tube 72.

[0109] In the fourth analysis mode, the trap column 12 is washed with the mobile phase B6. Also, MS
The ion source 9 is washed with the mobile phase C11. The sample injection port 3 is provided independently of the analysis column 4 in the washing channel system of the mobile phase C11. Therefore, the fourth analysis mode
In this mode, the flow injection analysis of the sample to the MS ion source unit 9 becomes possible. On the other hand, the analytical column 4 is washed with the mobile phase A for analysis.

Also in this embodiment, the first, second, third,
By switching the fourth analysis mode, desalting of the analysis sample and LC
/ MS direct analysis, removal of unnecessary components, and flow injection analysis are achieved.

[Embodiment 4] An LC / MS according to still another embodiment of the present invention will be described. FIG. 19 is a block diagram of an LC / MS according to still another embodiment of the present invention.
FIG. 20 is an operation explanatory diagram of the automatic removal of high concentration components of the LC / MS in FIG. 19, the same reference numerals in FIGS. 2 and 9 denote the same parts, and a description thereof will be omitted. Only new codes will be described. 26 is a comparator, 27 is an LC controller
In FIG. 20, O, P and Q are components to be analyzed, M is a main component, V is a component eluted by void volume, and both M and V are unnecessary components.

As shown in FIG. 19, the detector 5 is disposed at the subsequent stage of the analytical column 4 and monitors components eluted from the analytical column 4, and the comparator 26 always outputs the output voltage Vs of the detector 5. And the comparison voltage Vr. When a high-concentration component is eluted, the output voltage Vs of the detector 5 becomes larger than the comparison voltage Vr, and the LC controller 27 issues an instruction to switch to the fourth analysis mode by the LC / MS2.
8 is output. Elution of high concentration components is completed, and detector 5
Is smaller than the comparison voltage Vr, LC
The controller 27 instructs the LC / MS 28 to restore the fourth analysis mode.

The switching of the analysis mode will be described with reference to FIG. After setting the fourth analysis mode, sample injection is performed. The component V eluted in the void volume is discharged out of the system. In the holding time t _1, a first analysis mode - switched to de analysis target component O, the measurement of P is performed. The high concentration component M elutes, and the output voltage Vs of the detector 5 becomes the comparison voltage Vr.
When exceeded, i.e., at t _2, the analysis mode - de first analysis mode - automatically switched to de - from de fourth analysis mode. Advances the elution of the high concentration of the components M, when the output voltage Vs falls below the comparison voltage Vr, i.e., at t _3, the analysis mode - de first analysis mode - returns to de.

In the first analysis mode, the eluting target component Q can be measured after the main component M is eluted. In holding time t _4, LC / MS measurement is finished, the fourth analysis mode - provided to be switched to de next analysis. According to this embodiment, even if the retention time of the high concentration component is unknown, the high concentration component can be removed, and carry-over due to the high concentration component can be prevented.

(Experimental Example) The results of an experiment conducted using the above-described present invention will be described below. The sample used for the experiment is an azole antifungal agent having a molecular weight of 331. The analysis conditions are as follows. The system used is the system of [Embodiment 2] shown in FIG. Analysis column 4 is Y
MC-Pack A-312 ODS 6 × 150m
m, pump is L-6200 2 manufactured by Hitachi, Ltd.
Table (pumps 5 and 10 shown) and L-60
00 (pump 7 shown), LC / MS device is M-1000 manufactured by Hitachi, Ltd., MS ion source unit 9
Is an atmospheric pressure chemical ionization (APCI), the trap column 12 is a 4 × 30 mm ODS column, the detector 5 is a Hitachi L-4250 UV monitor, and the detection wavelength is 260 n.
m. The sample was dissolved in the mobile phase for analysis and the concentration was 10
It was adjusted to 0 μl / ml. 100 μl of this sample solution (sample injection amount: 10 μg) was injected from the sample injection port 3.

FIG. 21 is a liquid chromatogram of the first experimental example according to the present embodiment, FIG. 22 is a mass chromatogram of the first experimental example according to the present embodiment, and FIG. 23 is a second experimental example according to the present embodiment. Liquid chromatogram, FIG. 24 is a mass chromatogram of the second experimental example according to the present embodiment, FIG. 25 is a liquid chromatogram of the third experimental example according to the present embodiment, FIG.
Is the mass chromatogram of the third experimental example according to the present embodiment,
FIG. 27 is a liquid chromatogram of the fourth experimental example according to the present embodiment, and FIG. 28 is a mass chromatogram of the fourth experimental example according to the present embodiment.

(Experimental Example 1) FIGS. 21 and 22 show an analytical experimental example in which the entire amount of the eluate in the volatile mobile phase analysis system is directly injected into MS. Mobile phase A1 comprises
This is a solution in which acetonitrile and a 0.01 M aqueous solution of ammonium acetate are mixed at a ratio of 5:12. The flow rate of the mobile phase A1 was 1.2 ml / min, and the analysis was performed by Isocratec.

FIG. 21 is an output of the detector 5, ie, a liquid chromatogram, in which the target component is eluted at 8.78 minutes. FIG. 22 shows m / C detected by LC / MS.
It is a mass chromatogram of the pseudo molecular ion (M + H) of z332. This analysis was determined by the first analysis mode. That is, all of the LC eluate was sent to the MS ion source unit 9 and analyzed. In FIG. 22, the numeral 4611 on the right shoulder of the mass chromatogram indicates the peak height, that is, m /.
This corresponds to the maximum detected current value (unit: mA) of the pseudo molecular ion of z332.

(Experimental Example 2) FIGS. 23 and 24 correspond to FIGS.
22 shows an example of an analysis experiment in which the analysis is the same as that of Example 22, but the components eluted by the void volume are removed and the analysis target component is introduced into MS. FIG. 23 is a liquid chromatogram monitored by the detector 5, and the analysis target component elutes at 8.72 minutes. On the other hand, the unnecessary components are eluted by the void volume at 3.46 minutes.

In this analysis, after setting the fourth analysis mode, a sample is injected and LC analysis is started. The fourth analysis mode is maintained for up to 7 minutes to remove void volume components appearing around 3 minutes. After a lapse of 7 minutes, the mode is switched to the first analysis mode, and the eluate A of the LC is introduced into the MS ion source unit 9.

In this manner, the salt and the like from which the void volume elutes are discharged to the outside, and the analysis target component can be introduced into the MS ion source unit 9. FIG. 24 shows m / z
332 is a mass chromatogram by the pseudo-molecular ions of 332, and the retention time of the analysis target component does not change even if the valve switching operation is performed.

(Experimental Example 3) FIGS. 25 and 26 show the same analysis as in (Experimental Example 1) and (Experimental Example 2), except that components eluted by the void volume were removed, and a trap column for the target component was used. The example of measurement which capture | acquires and introduced into MS after desalting is shown. FIG. 25 is a liquid chromatogram obtained by the detector 5. The mobile phase A1 is an ammonium acetate solution, and there is no need for desalting in LC / MS.

This experiment was conducted to verify the function of the system. Up to 7 minutes after the sample injection, the eluent A is maintained in the fourth analysis mode, the eluate A is discarded outside, and at the same time, the pretreatment of the trap column 12 is performed. From 7 minutes to 9.8 minutes, the second analysis mode is maintained and analysis column 4
The analysis target component eluted from is captured. 9.8 minutes to 1
Up to 5 minutes, the fourth analysis mode is maintained, desalted, and the eluate A is discarded outside. After 15 minutes, the system enters the third analysis mode, in which the target component trapped in the trap column 12 is eluted by backflushing.

It should be noted here that the LC / LC of FIG. 22 in (Experimental Example 1) and FIG.
The peak height values detected in the MS measurement were 4611 and 3925, respectively, whereas the peak height value obtained in the present experimental example was 7970. The peak eluted by the backflushing becomes sharper than when the peak was eluted from the analytical column 4, meaning that the height increased by that much. This is extremely advantageous for measuring a sample at a concentration just below the detection limit. Therefore, this increase in sensitivity is another advantage of the present system. This suggests that even in a volatile mobile phase system, high sensitivity analysis can be performed by using this system.

(Experimental Example 4) FIGS. 27 and 28 show an analysis example using a nonvolatile mobile phase. The mobile phase A1 was used with a ratio of methanol to a 0.05 M aqueous solution of potassium dihydrogen phosphate of 35:65. The flow rate is 1.2 ml /
min. The mobile phase C11 that elutes the analysis target component captured in the trap column 12 is a 90% acetonitrile aqueous solution. The mobile phase B6 for dilution is water 10
0%. The sample was dissolved in the mobile phase A1 for analysis, adjusted to a concentration of 100 μg / ml, and 100 μl thereof was injected. The column is YMC A-312 ODS 6 × 15
0 mm was used. Trap column is 4 × 30mm
It is an ODS column. The analysis result by this LC is shown in FIG.
Shown in Under these conditions, a peak appears at 17.38 minutes.

FIG. 28 shows an experimental measurement example using the desalination system of this experimental example. From the time immediately after injection of the sample to the time of 16 minutes, the fourth analysis mode is set, and the eluate A from the analysis column 4 is discharged to the outside. During this time, trap column 1
2 has been washed with mobile phase B6, ie with water. 16
After a lapse of minutes, the mode is switched to the second analysis mode, and the analysis target component is captured by the trap column 12.

At 18.4 minutes when the elution of the analysis target component was completed, the mode was switched to the fourth analysis mode. In the trap column 12, the mobile phase B6, that is, water was flown, and desalting was performed. At a retention time of 23 minutes, in the third analysis mode, the components trapped in the trap column 12 are eluted by backflushing with the mobile phase C11,
Sent to MS.

In this case, the peak height value of the mass chromatogram is 9894, and the peak becomes a sharp as in the case of the analysis in the ammonium acetate solution system of (Experimental Example 3).
This is more than twice the peak height value 4611 in the case of direct MS introduction by the ammonium acetate solution system of (Experimental Example 1). According to this system, LC / MS can be performed in a system that shares a nonvolatile buffer and a nonvolatile salt.
Further, higher sensitivity can also be achieved.

Each of the above-described embodiments and experimental examples is described in the following.
By switching the valve, the first analysis mode and the second analysis mode
The direct connection method of LC / MS, which selects and connects the third analysis mode and the fourth analysis mode, has been described.
As shown in FIG. 1, each embodiment and each experimental example in the MS direct connection apparatus were controlled by the valves V-1 and V-1 by the LC controller 27.
-2, V-3 and the flow path systems 91 and 92 are freely switched to be the same as the respective examples and experimental examples of the direct connection method of LC / MS. Description is omitted.

[0130]

As described above in detail, according to the present invention, when all of the analysis modes are switched, one analysis mode is changed to another analysis mode. It does not require complicated procedures, labor and time, and is advantageous in terms of cost and space.
It is possible to provide a method of directly connecting an LC to an MS in which a memory or clogging is prevented and a specific high-concentration component is excluded from the MS ion source section 9, and an apparatus therefor. That is, the following three analysis modes required in LC / MS can be achieved in one system by switching valves. (1) In the case of analysis using a volatile mobile phase system, the eluate A from the analysis column 4 is directly introduced into the MS ion source 9 for analysis. Unnecessary components are removed to prevent the MS ion source 9 from being contaminated. (2) The target component separated in the mobile phase system containing the non-volatile components is desalted and introduced into the MS ion source 9 for analysis. (3) The target component to be analyzed is directly introduced into the MS ion source section 9 for analysis by flow injection analysis that does not require LC separation. This eliminates the need to change the piping and equipment for each analysis mode, and can significantly improve operability.

[Brief description of the drawings]

FIG. 1 is a block diagram showing an outline of the present invention.

FIG. 2 is an explanatory diagram showing an LC / MS analysis system according to one example of the present invention.

FIG. 3 is a first analysis mode of LC / MS according to the embodiment of FIG. 2;
FIG. 2 is an explanatory diagram showing an analysis system of a sword.

4 is a second analysis mode of LC / MS according to the embodiment of FIG.
FIG. 2 is an explanatory diagram showing an analysis system of a sword.

5 is a third analysis mode of LC / MS according to the embodiment of FIG.
FIG. 2 is an explanatory diagram showing an analysis system of a sword.

6 is a fourth analysis mode of LC / MS according to the embodiment of FIG. 2;
FIG. 2 is an explanatory diagram showing an analysis system of a sword.

FIG. 7 is an explanatory diagram of analysis mode switching in LC / MS using a nonvolatile mobile phase system.

FIG. 8 is an analysis mode of front removal by a volatile mobile phase system.
FIG. 9 is an explanatory diagram of the mode switching.

FIG. 9 is an explanatory diagram of an analysis mode switching of unnecessary component removal.

FIG. 10 is an explanatory diagram of switching of an analysis mode of sample concentration by repetition.

FIG. 11 is an explanatory view showing an analysis system of a first analysis mode of LC / MS according to another embodiment of the present invention.

FIG. 12 is an explanatory diagram showing an analysis system of a second analysis mode of LC / MS according to another embodiment of the present invention.

FIG. 13 is an explanatory view showing an analysis system of a third analysis mode of LC / MS according to another embodiment of the present invention.

FIG. 14 is an explanatory view showing an analysis system of a fourth analysis mode of LC / MS according to another embodiment of the present invention.

FIG. 15 shows an LC / MS according to still another embodiment of the present invention.
FIG. 4 is an explanatory diagram showing an analysis system of a first analysis mode.

FIG. 16 shows an LC / MS according to still another embodiment of the present invention.
FIG. 4 is an explanatory diagram showing an analysis system of a second analysis mode.

FIG. 17 shows an LC / MS according to still another embodiment of the present invention.
FIG. 7 is an explanatory diagram showing an analysis system of a third analysis mode.

FIG. 18 is an LC / MS according to still another embodiment of the present invention.
FIG. 10 is an explanatory diagram showing an analysis system of a fourth analysis mode.

FIG. 19: LC / MS according to still another embodiment of the present invention.
It is a block diagram of.

20 is an operation explanatory diagram of the automatic removal of high concentration components of the LC / MS in FIG. 18;

FIG. 21 is a liquid chromatogram of the first experimental example.

FIG. 22 is a mass chromatogram of the first experimental example.

FIG. 23 is a liquid chromatogram of a second experimental example.

FIG. 24 is a mass chromatogram of a second experimental example.

FIG. 25 is a liquid chromatogram of a third experimental example.

FIG. 26 is a mass chromatogram of a third experimental example.

FIG. 27 is a liquid chromatogram of a fourth experimental example.

FIG. 28 is a mass chromatogram of a fourth experimental example.

FIG. 29 is a block diagram for defining various solutions.

FIG. 30 is a block diagram of LC / MS using a conventional volatile mobile phase system.

FIG. 31 is a block diagram of LC / MS using a conventional nonvolatile mobile phase system.

FIG. 32 is a block diagram of a conventional flow injection MS.

[Explanation of symbols]

 Reference Signs List 1 mobile phase A 2 pump 3 sample inlet 4 analysis column 5 detector 6 mobile phase B 7 pump 8 TAY 9 MS ion source unit 10 pump 11 mobile phase C 12 trap column 18 TAY 19 branch resistance DR1 ... DR4 Drain V-1, V-2, V-3 Valve 1a..1q Switching port of valve V-1 2a..2q Switching port of valve V-2 3a..3f Switching port of valve V-3 30.・ 89 thin tube 21,22,23,24 three-way switching valve 25 non-volatile salt desalination system part 26 comparator 27 LC controller 28 LC / MS 30 ・ ・ 89 thin tube 91,92 channel system

────────────────────────────────────────────────── ─── Continuation of the front page (72) Inventor Yoshiaki Kato 882, Momo, Katsuta-shi, Ibaraki Pref. In the measuring instrument division of Hitachi, Ltd. Hei 3-175355 (JP, A) (58) Fields investigated (Int. Cl. ⁶ , DB name) G01N 27/62-27/70 G01N 30/72 H01J 49/00-49/48

Claims (14)

(57) [Claims] 1. Sample inlet, analytical column, pump and detection
Liquid chromatograph composed of a vessel and the mass spectrometer Tooi
In a method for directly connecting a high-performance liquid chromatograph and a mass spectrometer connected by an interface, the interface is a sample inlet, a plurality of switching valves, a plurality of pumps, a trap column, piping, and ionization means. A liquid chromatograph and a mass spectrometer comprising a mass spectrometer and an ion source, wherein the following four analysis modes are connected so as to be independently selected as needed by switching the valves. (A) The eluate from the analytical column of the liquid chromatograph is directly or entirely split or introduced into the ion source of the mass spectrometer, and the trap column is washed in the first analytical mode ( b) Components eluted from the analytical column of the liquid chromatograph are trapped in the trap column, and the mass spectrometer ion source is washed with a washing solution. Two analysis mode - de. (C) The components trapped in the trap column are eluted by the eluent and sent to the ion source of the mass spectrometer, and the eluate from the analytical column of the liquid chromatograph is discharged to the outside in the third analysis mode. (D) A fourth analysis mode in which the trap column and the mass spectrometer ion source are washed with a washing solution, and the eluate from the analysis column of the liquid chromatograph is discharged to the outside. 2. In the third analysis mode, the flow direction of the washing solution into the trap column is opposite to that of the first analysis mode, the second analysis mode, and the fourth analysis mode. The method according to claim 1, wherein the liquid chromatograph is connected to a mass spectrometer. 3. In the first analysis mode and the fourth analysis mode, both modes are switched by a plurality of valves to remove analysis-interfering components and unnecessary components eluted from the analytical column of the liquid chromatograph. 3. The liquid chromatograph and mass spectrometer according to claim 1, wherein the liquid chromatograph and the mass spectrometer are connected so that introduction and analysis of the analysis target component into the ion source of the mass spectrometer can be freely selected. Direct connection with. 4. In the second analysis mode, the analysis target component is captured by the trap column, in the fourth analysis mode, the trap column is washed, and in the third analysis mode, the analysis target component is captured by the trap column. Components are eluted and MS
4. The method for directly connecting a liquid chromatograph to a mass spectrometer according to claim 1, wherein the connection is performed such that the analysis target component is sent to the ion source. 5. In each sample injection analysis, a fourth analysis mode and a second analysis mode are combined, and a target component for analysis is concentrated in a trap column. third analysis mode - direct connection with any of the liquid chromatograph and mass spectrometer of claims 1 to 4, wherein the connecting to more analysis target component to the mass spectrometer ion source is fed to the de Method. 6. In the fourth analysis mode, the sample solution is directly sent from the sample inlet to the mass spectrometer ion source through a path for introducing the sample solution directly from the sample inlet to the mass spectrometer ion source. 6. The method according to claim 1, wherein the liquid chromatograph is connected to a mass spectrometer. 7. A detector which can monitor an elution component is provided between a flow channel between an analysis column of a liquid chromatograph and an interface part of a mass spectrometer directly connected to the liquid chromatograph, and an elution set with the output value of the detector is provided. 2. The method according to claim 1, further comprising the step of: comparing the component values with each other and, when the output value of the detector exceeds a set value, switching the connection of the analysis mode so that the liquid chromatograph eluate is discharged to the outside. A method for directly connecting a liquid chromatograph to a mass spectrometer according to any one of claims 5 to 5. 8. A liquid chromatograph comprising a sample inlet, an analytical column, a pump and a detector, a mass spectrometer,
An interface unit that couples the two, and the interface unit includes a sample inlet, a plurality of switching valves, a plurality of pumps, a trap column, piping, a mass spectrometer serving as ionization means, and an ion source unit. A liquid chromatograph and a mass spectrometer, comprising a control device, wherein the valve is switched by the control device, and the following four analysis modes are connected so as to be independently selected as needed. Direct connection device. (A) The first analysis mode in which the eluate from the analytical column of the liquid chromatograph is directly or entirely split into the mass spectrometer ion source, and the trap column is washed with a washing solution. (B) The second analysis mode in which components eluted from the analytical column of the liquid chromatograph are captured by the trap column, and the mass spectrometer ion source is washed with a washing solution. (C) The third analysis mode in which the components trapped in the trap column are eluted by the eluent and sent to the ion source of the mass spectrometer, and the eluate from the analytical column of the liquid chromatograph is discharged to the outside. (D) The fourth analysis mode in which the trap column and the mass spectrometer ion source are washed with a washing solution, and the eluate from the analysis column of the liquid chromatograph is discharged to the outside. 9. In the third analysis mode, the flow direction of the washing solution into the trap column is opposite to the direction of the first, second, and fourth analysis modes. 9. The apparatus according to claim 8, wherein the apparatus is connected to a liquid chromatograph and a mass spectrometer. 10. In the first analysis mode and the fourth analysis mode, both modes are switched by a plurality of valves to remove an analysis interfering component and an unnecessary component eluted from an analytical column of a liquid chromatograph. 10. The liquid chromatograph and the mass spectrometer according to claim 8, wherein the liquid chromatograph and the mass spectrometer are connected so that introduction and analysis of a component to be analyzed into the ion source of the mass spectrometer can be freely selected. Direct connection device with. 11. In the second analysis mode, the analysis target component is captured in the trap column. In the fourth analysis mode, the trap column is washed. In the third analysis mode, the analysis target component is captured by the trap column. Components are eluted and MS
11. The apparatus for directly connecting a liquid chromatograph to a mass spectrometer according to any one of claims 8 to 10, wherein the apparatus is connected so that the analysis target component is sent to the ion source. 12. In each sample injection analysis, a fourth analysis mode and a second analysis mode are combined, and a target component for analysis is concentrated in a trap column. 12. The apparatus for directly connecting a liquid chromatograph to a mass spectrometer according to any one of claims 8 to 11, wherein the third analysis mode is connected so that the analysis target component is sent to the mass spectrometer ion source. . 13. In the fourth analysis mode, the sample solution is directly sent from the sample inlet to the mass spectrometer ion source through a path for introducing the sample solution directly from the sample inlet to the mass spectrometer ion source. 13. The apparatus according to claim 8, wherein the liquid chromatograph is connected to the mass spectrometer. 14. A detector capable of monitoring an eluted component between flow paths between an analytical column of a liquid chromatograph and an interface part of a mass spectrometer directly connected to the liquid chromatograph, and an output value of the detector and a set value of the eluted component. And a comparator for comparing the analysis mode so that the liquid chromatograph eluate is discharged to the outside when the output value of the detector exceeds a set value. A device for directly connecting the liquid chromatograph according to any one of claims 8 to 12 to a mass spectrometer.