阅读说明：本技术 液相色谱分析装置及方法、流动相供给装置及方法 (Liquid chromatography device and method, and mobile phase supply device and method ) 是由 家氏淳 于 2019-08-22 设计创作，主要内容包括：本发明提供一种液相色谱分析装置及方法、流动相供给装置及方法。将第一储存部中储存的含有盐的水溶液通过第一配管引导到混合部,将第二储存部中储存的有机溶剂通过第二配管引导到混合部。通过由混合部将水溶液和有机溶剂进行混合来生成流动相。由加热部对第一配管的至少一部分和第二配管的至少一部分进行加热,以使由混合部生成的流动相的温度为水溶液中含有的盐的溶解温度以上。(The invention provides a liquid chromatography device and method, and a mobile phase supply device and method. The salt-containing aqueous solution stored in the first storage unit is guided to the mixing unit through a first pipe, and the organic solvent stored in the second storage unit is guided to the mixing unit through a second pipe. The mobile phase is generated by mixing the aqueous solution and the organic solvent in the mixing section. At least a part of the first pipe and at least a part of the second pipe are heated by the heating unit so that the temperature of the mobile phase generated by the mixing unit is equal to or higher than the dissolution temperature of the salt contained in the aqueous solution.)

1. A liquid chromatography apparatus includes:

a mobile phase supply device for generating a mobile phase;

a syringe to which a sample and the mobile phase generated by the mobile phase supply device are supplied;

a column into which the mobile phase and the sample supplied to the syringe are introduced; and

a detector for detecting the sample passed through the column,

wherein the mobile phase supply device comprises:

a first storage part for storing an aqueous solution containing a salt;

a second storage part for storing an organic solvent;

a mixing part that generates a mobile phase by mixing the aqueous solution stored in the first storage part and the organic solvent stored in the second storage part;

a first pipe for connecting the mixing section and the first storage section;

a second pipe for connecting the mixing section and the second storage section; and

a heating unit configured to heat at least a part of the first pipe and at least a part of the second pipe so that a temperature of the mobile phase generated by the mixing unit is equal to or higher than a dissolution temperature of a salt contained in the aqueous solution.

2. The liquid chromatography apparatus according to claim 1,

further comprising a column thermostatic bath for accommodating the column and adjusting the temperature of the column,

the column thermostatic bath includes the heating unit, and also houses at least a part of the first pipe and at least a part of the second pipe, and heats at least a part of the first pipe and at least a part of the second pipe.

3. A mobile phase supply device for supplying a mobile phase used for liquid chromatography of a sample, the mobile phase supply device comprising:

a first storage part for storing an aqueous solution containing a salt;

a second storage part for storing an organic solvent;

a mixing part that generates a mobile phase by mixing the aqueous solution stored in the first storage part and the organic solvent stored in the second storage part;

a first pipe for connecting the mixing section and the first storage section;

a second pipe for connecting the mixing section and the second storage section; and

a heating unit configured to heat at least a part of the first pipe and at least a part of the second pipe so that a temperature of the mobile phase generated by the mixing unit is equal to or higher than a dissolution temperature of a salt contained in the aqueous solution.

4. A liquid chromatography method comprising the steps of:

generating a mobile phase by a mobile phase supply device;

supplying the sample and the mobile phase generated by the mobile phase supply device to a syringe;

introducing the mobile phase and the sample supplied to the syringe into a column; and

the sample that has passed through the column is detected by a detector,

wherein the step of generating a mobile phase by the mobile phase supply device comprises the steps of:

guiding the salt-containing aqueous solution stored in the first storage unit to the mixing unit through a first pipe;

guiding the organic solvent stored in the second storage unit to the mixing unit through a second pipe;

generating a mobile phase by mixing the aqueous solution and the organic solvent by the mixing section; and

at least a part of the first pipe and at least a part of the second pipe are heated by a heating unit so that the temperature of the mobile phase generated by the mixing unit is equal to or higher than the dissolution temperature of the salt contained in the aqueous solution.

5. The liquid chromatography analysis method according to claim 4,

further comprising the step of adjusting the temperature of the column by a column thermostatic bath for accommodating the column,

the column thermostatic bath includes the heating portion,

the step of heating at least a part of the first pipe and at least a part of the second pipe by a heating unit includes the steps of:

storing at least a part of the first pipe and at least a part of the second pipe in the column thermostat tank; and

at least a part of the first pipe and at least a part of the second pipe are heated by the column thermostat.

6. A mobile phase supply method for supplying a mobile phase used in liquid chromatography of a sample, comprising the steps of:

guiding the salt-containing aqueous solution stored in the first storage unit to the mixing unit through a first pipe;

guiding the organic solvent stored in the second storage unit to the mixing unit through a second pipe;

generating a mobile phase by mixing the aqueous solution and the organic solvent by the mixing section; and

at least a part of the first pipe and at least a part of the second pipe are heated by a heating unit so that the temperature of the mobile phase generated by the mixing unit is equal to or higher than the dissolution temperature of the salt contained in the aqueous solution.

Technical Field

The present invention relates to a liquid chromatography apparatus, a mobile phase supply apparatus, a liquid chromatography method, and a mobile phase supply method for supplying a mobile phase.

Background

In a liquid chromatography apparatus, a mixed liquid of an aqueous solution containing a salt (hereinafter, simply referred to as an aqueous solution) and an organic solvent is sometimes used as a mobile phase. For example, the aqueous solution and the organic solvent are mixed in advance in a common bottle to produce a mixed solution. In this case, the salt in the aqueous solution hardly precipitates. Even when the salt is precipitated, the mixed solution is stirred in the bottle to dissolve the salt again in the mixed solution.

On the other hand, the aqueous solution and the organic solvent may be stored in different bottles, and when a sample is analyzed, the aqueous solution and the organic solvent supplied from the bottles are mixed in a common flow path to produce a mixed solution. In this case, at the interface where the aqueous solution and the organic solvent are in contact, the concentration of the organic solvent increases, and therefore precipitation of a salt is liable to occur. In the case where the concentration of the organic solvent is high, the problem is more significant.

Jp 2010-156660 a describes a mobile phase supply device for supplying a mixed solution of a buffer solution (hereinafter, referred to simply as a buffer solution) in which a salt is dissolved and an organic solvent as a mobile phase to an analysis flow path of a liquid chromatograph. In this mobile phase supply device, the buffer solution stored in the first storage tank is supplied to the liquid-feeding pump through the first electromagnetic valve, the first backflow prevention valve, and the mixing flow path. The organic solvent stored in the second storage tank is supplied to the liquid-feeding pump through the second electromagnetic valve, the second backflow prevention valve, and the mixing flow path. The buffer solution and the organic solvent are mixed in a pump chamber of a liquid feeding pump.

Disclosure of Invention

In the mobile phase supply apparatus described in japanese patent application laid-open No. 2010-156660, even when salt is precipitated when a buffer solution and an organic solvent are brought into contact in a mixing flow path, the first and second backflow prevention valves prevent the salt from entering the first and second electromagnetic valves, respectively. However, precipitation itself of the salt cannot be prevented, and the precipitated salt may inhibit stable supply of the mobile phase.

The invention aims to provide a liquid chromatography device, a mobile phase supply device, a liquid chromatography method and a mobile phase supply method which can stably supply mobile phase.

(1) A liquid chromatography apparatus according to an aspect of the present invention includes: a mobile phase supply device for generating a mobile phase; a syringe to which the sample and the mobile phase generated by the mobile phase supply device are supplied; a column into which the mobile phase and the sample supplied to the syringe are introduced; and a detector for detecting the sample having passed through the column, wherein the mobile phase supply device includes: a first storage part for storing an aqueous solution containing a salt; a second storage part for storing an organic solvent; a mixing section that generates a mobile phase by mixing the aqueous solution stored in the first storage section and the organic solvent stored in the second storage section; a first pipe for connecting the mixing section and the first storage section; a second pipe for connecting the mixing section and the second storage section; and a heating unit that heats at least a part of the first pipe and at least a part of the second pipe so that the temperature of the mobile phase generated by the mixing unit is equal to or higher than the dissolution temperature of the salt contained in the aqueous solution.

In this liquid chromatography apparatus, the sample and the mobile phase generated by the mobile phase supply device are supplied to the syringe. The mobile phase and the sample supplied to the syringe are introduced into the column, and the sample having passed through the column is detected by a detector.

In the mobile phase supply apparatus, the salt-containing aqueous solution stored in the first storage unit is guided to the mixing unit through the first pipe, and the organic solvent stored in the second storage unit is guided to the mixing unit through the second pipe. The mobile phase is generated by mixing the aqueous solution and the organic solvent in the mixing section. At least a part of the first pipe and at least a part of the second pipe are heated by the heating unit so that the temperature of the mobile phase generated by the mixing unit is equal to or higher than the dissolution temperature of the salt contained in the aqueous solution.

According to this configuration, the aqueous solution and the organic solvent are heated at a position upstream of the mixing section so that the temperature of the mobile phase becomes equal to or higher than the dissolution temperature of the salt contained in the aqueous solution. Therefore, even in the case where the aqueous solution is in contact with the organic solvent, salt precipitation is prevented regardless of the concentration of the organic solvent. Thus, there is no case where salt hinders stable supply of the mobile phase. This enables stable supply of the mobile phase.

(2) The liquid chromatography apparatus may further include a column thermostat which accommodates the column and adjusts the temperature of the column, and the column thermostat may include a heating unit which accommodates at least a part of the first pipe and at least a part of the second pipe and heats at least a part of the first pipe and at least a part of the second pipe. In this case, it is not necessary to provide a heating unit for heating at least a part of the first pipe and at least a part of the second pipe separately from the column thermostatic bath. This can reduce the cost of the mobile phase supply device and can miniaturize the mobile phase supply device.

(3) A mobile phase supply device according to another aspect of the present invention is a mobile phase supply device for supplying a mobile phase used in liquid chromatography of a sample, the mobile phase supply device including: a first storage part for storing an aqueous solution containing a salt; a second storage part for storing an organic solvent; a mixing section that generates a mobile phase by mixing the aqueous solution stored in the first storage section and the organic solvent stored in the second storage section; a first pipe for connecting the mixing section and the first storage section; a second pipe for connecting the mixing section and the second storage section; and a heating unit that heats at least a part of the first pipe and at least a part of the second pipe so that the temperature of the mobile phase generated by the mixing unit is equal to or higher than the dissolution temperature of the salt contained in the aqueous solution.

In the mobile phase supply device, even when the aqueous solution is in contact with the organic solvent, salt precipitation is prevented regardless of the concentration of the organic solvent. Thus, there is no case where salt hinders stable supply of the mobile phase. This enables stable supply of the mobile phase.

(4) The liquid chromatography method according to another other aspect of the present invention comprises the steps of: generating a mobile phase by a mobile phase supply device; supplying the sample and the mobile phase generated by the mobile phase supply device to the syringe; introducing the mobile phase and the sample supplied to the syringe into the column; and detecting the sample having passed through the column by the detector, wherein the step of generating the mobile phase by the mobile phase supply device includes the steps of: guiding the salt-containing aqueous solution stored in the first storage unit to the mixing unit through a first pipe; guiding the organic solvent stored in the second storage unit to the mixing unit through a second pipe; generating a mobile phase by mixing the aqueous solution and the organic solvent by the mixing section; and heating at least a part of the first pipe and at least a part of the second pipe by the heating unit so that the temperature of the mobile phase generated by the mixing unit is equal to or higher than the dissolution temperature of the salt contained in the aqueous solution.

According to this liquid chromatography, even when an aqueous solution is brought into contact with an organic solvent in a mobile phase supply device, salt precipitation is prevented regardless of the concentration of the organic solvent. Thus, there is no case where salt hinders stable supply of the mobile phase. This enables stable supply of the mobile phase.

(5) The liquid chromatography method may further include a step of adjusting the temperature of the column by a column thermostatic chamber that houses the column, the column thermostatic chamber including a heating unit, and the step of heating at least a part of the first pipe and at least a part of the second pipe by the heating unit may include the steps of: storing at least a part of the first pipe and at least a part of the second pipe in a column thermostat; and heating at least a part of the first pipe and at least a part of the second pipe by the column thermostat. In this case, the cost of the mobile phase supply device can be reduced, and the mobile phase supply device can be downsized.

(6) A mobile phase supply method according to another other aspect of the present invention is a method for supplying a mobile phase used in liquid chromatography analysis of a sample, the mobile phase supply method including the steps of: guiding the salt-containing aqueous solution stored in the first storage unit to the mixing unit through a first pipe; guiding the organic solvent stored in the second storage unit to the mixing unit through a second pipe; generating a mobile phase by mixing the aqueous solution and the organic solvent by the mixing section; and heating at least a part of the first pipe and at least a part of the second pipe by the heating unit so that the temperature of the mobile phase generated by the mixing unit is equal to or higher than the dissolution temperature of the salt contained in the aqueous solution.

According to this mobile phase supply method, even when an aqueous solution is brought into contact with an organic solvent, salt precipitation is prevented regardless of the concentration of the organic solvent. Thus, there is no case where salt hinders stable supply of the mobile phase. This enables stable supply of the mobile phase.

Drawings

Fig. 1 is a diagram showing a configuration of a liquid chromatography apparatus according to an embodiment of the present invention.

Fig. 2 is a diagram showing a configuration of a liquid chromatography apparatus according to a first modification.

Fig. 3 is a diagram showing a configuration of a liquid chromatography apparatus according to a second modification.

Fig. 4 is a graph showing a change over time in the pressure instruction value of the liquid feeding portion in example 1.

Fig. 5 is a graph showing a change with time of the pressure instruction value of the liquid feeding portion in comparative example 1.

FIG. 6 is a graph showing the results of analysis of the sample in example 2,

fig. 7 (a) to (c) are graphs showing the results of gradient analysis of the sample in example 3.

Detailed Description

[ description of preferred embodiments ]

(1) Structure of liquid chromatography device

Hereinafter, a mobile phase supply device, a liquid chromatography device, a mobile phase supply method, and a liquid chromatography method according to embodiments of the present invention will be described in detail with reference to the drawings. Fig. 1 is a diagram showing a configuration of a liquid chromatography apparatus according to an embodiment of the present invention. The liquid chromatography apparatus 100 of fig. 1 is an HPLC (high performance liquid chromatography) apparatus.

As shown in fig. 1, the liquid chromatography apparatus 100 includes a mobile phase supply apparatus 10, an injector 20, a column thermostat 30, and a detector 40. The column thermostat 30 includes a heating portion. The column 31 is accommodated in the column oven 30, and the inside of the column oven 30 is adjusted to a predetermined fixed temperature.

The mobile phase supply apparatus 10 includes pipes 1, 2, and 3, storage units 11 and 12, a mixing unit 13, a degasifier 14, and a liquid feed unit 15. In the present embodiment, the mobile phase supply apparatus 10 includes a part (heating part) of the column thermostat tank 30. The storage unit 11 is a liquid medicine bottle for storing an aqueous solution containing salt (hereinafter, simply referred to as an aqueous solution). The storage unit 12 is a liquid medicine bottle similar to the storage unit 11 and stores an organic solvent. The storage parts 11, 12 are examples of a first storage part and a second storage part, respectively.

The mixing section 13 is, for example, a low pressure gradient unit, including port A, B, C. The storage section 11 and the port a of the mixing section 13 are connected by the pipe 1. The storage section 12 and the port B of the mixing section 13 are connected by the pipe 2. The pipes 1 and 2 are examples of a first pipe and a second pipe, respectively. The port C of the mixing section 13 is connected to the pipe 3. The mixing section 13 mixes the aqueous solution supplied from the reservoir section 11 to the port a through the pipe 1 and the organic solvent supplied from the reservoir section 12 to the port B through the pipe 2, thereby generating a mobile phase, and outputs the generated mobile phase from the port C.

In the following description, the upstream and downstream of the liquid chromatography apparatus 100 are defined based on the flow of an aqueous solution or an organic solvent. At least a part of the pipes 1 and 2 located upstream of the mixing section 13 is heated so that the temperature of the mobile phase after mixing is equal to or higher than the dissolution temperature of the salt contained in the aqueous solution. In the present embodiment, a part of each of the pipes 1 and 2 is disposed in the column thermostat 30. This enables heat exchange between the aqueous solution and the organic solvent in the column thermostat 30 to bring the temperature of the mixed mobile phase to a temperature equal to or higher than the dissolution temperature of the salt.

The portions of the pipes 1 and 2 disposed in the column thermostat 30 may be formed in a ring shape. In this case, the portions of the pipes 1 and 2 disposed in the column thermostat 30 can be made sufficiently long while maintaining the miniaturization. As a result, the heat exchange can be performed more easily.

The degasifier 14 is inserted into each of the pipes 1 and 2 to remove gas contained in the aqueous solution flowing through the pipe 1 and to remove gas contained in the organic solvent flowing through the pipe 2. The liquid feeding portion 15 is, for example, a pump unit, and is inserted into the pipe 3. The liquid feeding portion 15 pressure-feeds the mobile phase output from the port C of the mixing portion 13 to the downstream.

The syringe 20, the column 31, and the detector 40 are inserted into the pipe 3 downstream of the liquid feeding portion 15 in this order. The sample to be measured is supplied to the syringe 20, and the sample to be measured is introduced into the column 31 together with the mobile phase that is pressure-fed by the liquid feeding unit 15. The sample introduced into the column 31 is separated for each component and eluted at different times. The detector 40 detects the sample eluted from the column 31.

(2) Modification example

Fig. 2 is a diagram showing the configuration of the liquid chromatography apparatus 100 according to the first modification. As shown in fig. 2, the liquid chromatography apparatus 100 according to the first modification includes heating portions 16 and 17. The liquid chromatography apparatus 100 according to the first modification does not include the degasser 14, but may include the degasser 14. The same applies to the liquid chromatography apparatus 100 according to the second modification example described later.

The heating units 16 and 17 may be any of a warm water tank, an electric heater, a peltier element, and the like, and heat at least a part of the pipes 1 and 2, respectively, so that the temperature of the mixed mobile phase is equal to or higher than the dissolution temperature of the salt contained in the aqueous solution. In this case, a part of each of the pipes 1 and 2 may not be disposed in the column thermostat tank 30. The liquid chromatography apparatus 100 according to the first modification may include a common heating unit for heating the mixing unit 13, instead of the heating units 16 and 17.

Fig. 3 is a diagram showing the configuration of a liquid chromatography apparatus 100 according to a second modification. As shown in fig. 3, the liquid chromatography apparatus 100 according to the second modification includes liquid feeding portions 15a and 15b similar to the liquid feeding portion 15, instead of the liquid feeding portion 15. The mixing section 13 is not a low-pressure gradient unit, and is, for example, a mixer unit.

The liquid feeding unit 15a is inserted into the pipe 1 and is used to pressure-feed the aqueous solution stored in the storage unit 11 toward the mixing unit 13. The liquid feeding unit 15b is inserted into the pipe 2 and is used to pressure-feed the organic solvent stored in the storage unit 12 toward the mixing unit 13. In this case, the aqueous solution and the organic solvent are transported by the liquid transport portion 15a and the liquid transport portion 15b provided separately. This enables the sample to be analyzed by a high-pressure gradient method.

The liquid chromatography apparatus 100 according to the second modification may include the same heating portions 16 and 17 as those of the first modification or a common heating portion for heating the mixing portion 13. In this case, a part of each of the pipes 1 and 2 may not be disposed in the column thermostat tank 30.

(3) Effect

In the liquid chromatography apparatus 100 according to the present embodiment, the sample and the mobile phase generated by the mobile phase supply apparatus 10 are supplied to the syringe 20. The mobile phase and the sample supplied to the syringe 20 are introduced into the column 31 housed in the column thermostat 30, and the detector 40 detects the sample that has passed through the column 31.

In the mobile phase supply apparatus 10, the aqueous solution containing the salt stored in the storage unit 11 is guided to the mixing unit 13 through the pipe 1, and the organic solvent stored in the storage unit 12 is guided to the mixing unit 13 through the pipe 2. The mobile phase is generated by mixing the aqueous solution and the organic solvent in the mixing section 13. Here, at least a part of the pipe 1 and at least a part of the pipe 2 are stored in the column thermostat tank 30. At least a part of the pipe 1 and at least a part of the pipe 2 are heated by the column thermostat 30 so that the temperature of the mobile phase generated in the mixing section 13 is equal to or higher than the dissolution temperature of the salt contained in the aqueous solution.

According to this configuration, the aqueous solution and the organic solvent are heated at a position upstream of the mixing section 13 so that the temperature of the mobile phase becomes equal to or higher than the dissolution temperature of the salt contained in the aqueous solution. Therefore, even in the case where the aqueous solution is in contact with the organic solvent, salt precipitation is prevented regardless of the concentration of the organic solvent. Thus, there is no case where salt hinders stable supply of the mobile phase. This enables stable supply of the mobile phase.

In the present embodiment, at least a part of the pipe 1 and at least a part of the pipe 2 are heated by the column thermostat 30, and therefore, it is not necessary to provide a heating unit separately from the column thermostat 30. This can reduce the cost of the mobile phase supply device 10 and also reduce the size of the mobile phase supply device 10.

(4) Example 1 and comparative example

In example 1, a previously cooled potassium phosphate buffer and a previously cooled acetonitrile were used as an aqueous solution and an organic solvent, respectively, and a mobile phase was supplied using the mobile phase supply apparatus 10 of fig. 1. Hereinafter, potassium phosphate buffer is referred to as solution A, and acetonitrile is referred to as solution B. The same applies to comparative example 1, example 2, and example 3 described later.

Specifically, in example 1, 50mmol/L of the solution A was stored in the storage unit 11, and 50mmol/L of the solution B was stored in the storage unit 12. The mixing section 13 is controlled so that the ratio of the pressure in these storage sections 11, 12 is 45: 55 to deliver liquid a and liquid B. Here, the mixing section 13 is a low-pressure gradient unit.

About 5mL of the respective pipes 1 and 2 are placed in the column thermostat 30 so that the liquids a and B flowing through the pipes 1 and 2 stay in the column thermostat 30 for about 10 minutes and are heated. The liquid a and the liquid B are heated in the column thermostat bath 30 and mixed in the mixing section 13, and then sent downstream by the liquid sending section 15.

Fig. 4 is a graph showing the change with time of the pressure instruction value of the liquid feeding portion 15 in example 1. As shown in fig. 4, in example 1, the pressure of the liquid feeding portion 15 was substantially constant, and the pressure did not fluctuate. From this, it was confirmed that in example 1, even when the solution a and the solution B were mixed, no salt was precipitated, and thus the mobile phase could be stably supplied.

On the other hand, in comparative example 1, the same mobile phase supply as in example 1 was performed using a liquid chromatography apparatus having the same configuration as the mobile phase supply apparatus 10 in fig. 1 except that a configuration for heating the pipes 1 and 2 was not provided. Therefore, the liquid a and the liquid B are mixed by the mixing unit 13 without being heated, and then are sent downstream by the liquid sending unit 15.

Fig. 5 is a graph showing the change over time in the pressure instruction value of the liquid feeding portion 15 in comparative example 1. As shown in fig. 5, in comparative example 1, the pressure of the liquid feeding portion 15 was substantially constant and did not fluctuate within about 7 minutes after the start of liquid feeding. This is considered to be because the temperature of the liquid-feeding portion 15 at the start of liquid feeding was high and no salt was precipitated. However, if the time exceeds 7 minutes after the start of liquid feeding, the pressure in the liquid feeding portion 15 fluctuates greatly at short intervals, and the mobile phase cannot be supplied stably. This is considered to be because the salt is precipitated due to the decrease in temperature of the liquid feeding portion 15.

(5) Example 2

In example 2, a sample was analyzed using the liquid chromatography apparatus 100 of fig. 1. The samples were thiuram standard solutions of the pesticide. Specifically, the thiuram standard solutions were supplied to the syringe 20 at concentrations of 0.1mg/L, 0.2mg/L, 0.5mg/L, and 1.0mg/L in this order, and introduced into the column 31 through the mobile phase produced under the same conditions as in example 1. Here, the column 31 is an ODS (Octadecylsilyl: octadecyl) column, and the temperature of the column 31 is 40 ℃. The flow rate and supply amount of the mobile phase were 1.0mL/min and 20. mu.L, respectively.

Fig. 6 is a graph showing the results of analysis of the sample in example 2. In fig. 6, the horizontal axis represents time, and the vertical axis represents the detection intensity of the sample. The results of analyzing samples at concentrations of 0.1mg/L, 0.2mg/L, 0.5mg/L, and 1.0mg/L are shown by a solid line, a broken line, a one-dot chain line, and a two-dot chain line, respectively. As shown in fig. 6, in example 2, the baseline of the detection intensity did not change regardless of the concentration of the sample. This confirmed that the mobile phase was stably supplied and that the sample could be stably detected and analyzed.

(6) Example 3

Gradient analysis of the samples was performed in example 3. Specifically, a thiuram standard solution having a concentration of 1.0mg/L was used as a sample, and the concentrations of the solutions A and B were set in the range of 70: 30-30: 70 is continuously varied. Other analysis conditions were the same as those in example 2.

Fig. 7 (a) to (c) are graphs showing the results of gradient analysis of the sample in example 3. The horizontal axes of fig. 7 (a) to (c) indicate common time, the vertical axis of fig. 7 (a) indicates the ratio (concentration) of the B liquid, the vertical axis of fig. 7 (B) indicates the pressure indication value of the liquid sending unit 15, and the vertical axis of fig. 7 (c) indicates the detection intensity of the sample.

As shown in fig. 7 (a) and (B), even when the concentration of the liquid B is increased from 30% to 70%, the pressure of the liquid feeding portion 15 does not fluctuate at short intervals. From this, it was confirmed that the mobile phase could be stably supplied because no salt precipitated even when the solution a was mixed with the solution B having a high concentration. As shown in fig. 7 (c), the baseline of the detection intensity did not change. This confirmed that the detection of the sample and the gradient analysis could be stably performed.