JP5148632B2 - Liquid chromatograph - Google Patents

Description

  The present invention is suitable for, for example, high-performance liquid chromatograph (HPLC) recycle separation, and can reduce the external volume (dead volume) of the separation column and suppress the diffusion of the sample by the dead volume by a simple configuration and piping structure. , The flow rate is 0.4 mL / min. In addition to realizing recycling separation in the above-mentioned class of liquid chromatographs, a column switching system using a recycling separation circuit is configured in a simple and inexpensive manner, facilitating analysis, fractionation, extraction and purification, and separation of target components. The present invention relates to a liquid chromatograph that facilitates purification and promotes high-quality purification of multiple components.

As a method for increasing the resolution of the separation column of the liquid chromatograph, there is a recycling separation method in which the eluate from the column is returned to the column again and the separation is repeated. For example, as the recycling separation method, the mobile phase supply unit, the sample introduction unit, The recycle switching valve is connected to the main analysis liquid path in which the separation column and the detector are sequentially connected, and the flow path on the detector outlet side is connected to the inlet side of the liquid feeding pump of the mobile phase supply unit via the switching valve. There is a preparative liquid chromatograph in which a sample is circulated to the same separation column together with a solvent in a state in which switching is possible and a flow path including a sample introduction part is bypassed at the time of recycling separation (for example, see Patent Document 1).

However, since the sample circulates the sample together with the solvent from the outlet side of the liquid pump to the separation column via the pressure gauge at the time of recycling separation, there is an increase in the number of unnecessary passing parts for recycling. There is a concern that the outer volume of the column is increased and the sample is diffused in the outer volume of the separation column, so that the peak of the chromatogram is widened.
In this case, the inner diameter of the preparative column is about 100 mm, and the optimum flow rate for this inner diameter is about 300 mL / min. Thus, the diffusion of the sample in the outer volume of the separation column is diluted and does not significantly affect the resolution of the separation column.

  However, in recycle separation of liquid chromatographs in which a sample is circulated at a low speed with a low flow rate solvent, or in liquid chromatography that does not use a preparative column, the increase in the external volume of the separation column or the sample in the external volume of the separation column The recycling separation method using a pressure gauge as described above cannot be adopted.

Moreover, in the conventional field of recycle separation, the inner diameter of a preparative size column is about 100 mm, and the optimum flow rate for this inner diameter is about 300 mL / min. In contrast, an easy-to-use analytical size column (with an inner diameter of about 4.6 mm and an optimum flow rate of about 1 mL / min.) Was being used.
However, in such conventional recycling separation using a liquid chromatograph, it has been difficult to perform the intended recycling separation due to the column outer volume (dead volume).

  Therefore, as a solution to such a problem, the first and second flow paths connecting the separation column and the detector are provided, and these flow paths are detachably connected via two flow path switching valves. At the time of recycling separation, the two flow path switching valves are alternately switched, and the sample and 1 mL / min. There is one in which the following solvent is introduced from one channel to the other channel and separated by each separation column (see, for example, Patent Document 2).

  However, the apparatus requires two separation columns, a detector, a 6-port flow path switching valve, and a liquid feed pump for recycling separation, and the structure is complicated, large, and expensive. There was a problem that the dead volume increased, the sample was diffused, the precision of recycling separation could not be obtained, and the switching operation of the two flow path switching valves was complicated and troublesome.

Japanese Unexamined Patent Publication No. 7-31188 JP 2006-201039 A

  The present invention solves such a problem, and is suitable for, for example, high-performance liquid chromatograph (HPLC) recycling separation, and reduces the external volume (dead volume) of the separation column by a simple configuration and piping structure, thereby reducing the dead volume. The flow rate is 0.4 mL / min. In addition to realizing recycling separation in the above-mentioned class of liquid chromatographs, a column switching system using a recycling separation circuit is configured in a simple and inexpensive manner, facilitating analysis, fractionation, extraction and purification, and separation of target components. An object is to provide a liquid chromatograph that facilitates purification and promotes high-quality purification of a plurality of components.

  According to the first aspect of the present invention, there is provided a circulation flow path capable of circulating and moving a sample, a liquid feed pump, and a pressure measuring means capable of measuring a pressure downstream of the liquid feed pump. And a liquid chromatograph provided with a damper capable of buffering the pressure downstream of the liquid feed pump, provided with a pressure measuring tube having one end communicating with the circulation channel and the other end being occluded, The pressure measuring means and the damper are inserted into the pressure measuring pipe, and the other end portion of the pressure measuring pipe is closed, whereby the pressure measuring means inserted in the pressure measuring pipe, and the damper provided in the pressure measuring means And the substantial volume outside the separation column is greatly reduced, the diffusion of the sample moving through the circulation channel is greatly suppressed, and the flow rate is 0.4 mL / min. Recycling separation in the above class of liquid chromatographs is realized with a simple configuration and piping structure, and even in chromatograms with peak retention capacity of about 3.0 mL or more, sufficient liquid chromatographs for recycling separation are provided. Is to be realized.

According to a second aspect of the present invention, the liquid feed pump is inserted into the circulation channel, a sample introduction part is provided in the circulation channel between the liquid feed pump and the separation column, and the sample introduction part and the liquid feed are provided. A connector having a plurality of ports is inserted between the pump and one end of the pressure measuring tube is connected to the port of the connector to enable pressure measurement by the pressure measuring means, The pressure measuring tube, the pressure measuring means, and the damper can be separated from the circulation flow path via the sample introduction unit.
In the invention of claim 3, a detector is inserted in the circulation flow path, the inner diameter of the downstream side portion of the circulation flow path communicating with the detector can be reduced, and the back pressure on the downstream pipe line of the detector is increased. In this way, it is possible to reduce the inner diameter of the downstream pipe line and to reduce the external volume of the separation column in the downstream pipe line.

According to a fourth aspect of the present invention, the liquid chromatograph has a flow rate of 0.4 mL / min. As described above, a liquid chromatograph having a remarkable effect of reducing the external volume of the separation column is realized.
According to the invention of claim 5, the liquid chromatograph improves the efficiency of recycle separation of a frequently used liquid chromatograph, wherein the elution volume until the peak of the sample appears is about 3.0 mL or more. I am doing so.

  According to the sixth aspect of the present invention, the recycling valve is attached to the introduction portion of the liquid feeding pump, the recycling valve can be communicated with the introduction portion of the liquid feeding pump, and recycled from the introduction portion of the liquid feeding pump, that is, the suction portion. The piping up to the valve is omitted, the piping of the relevant part is unnecessary, and the integrated configuration of the inlet part of the liquid feed pump and the recycling valve is realized, simplifying the configuration and compacting their installation space In addition, the dead volume of the part is set to a substantially zero value to suppress the dispersion of the sample so that good separation can be obtained.

The invention according to claim 7 is a pressure measuring means capable of measuring a pressure downstream of the liquid feed pump, a circulation flow path capable of circulating and moving the sample, a separation column and a recycle valve. And a damper capable of buffering the pressure downstream of the liquid feed pump, and a detector, a pressure chromatograph with one end communicating with the circulation channel and the other end being occluded A pipe is provided, and the pressure measuring means and the damper are inserted into the pressure measuring pipe, and a separation column that is the same as or different from the separation column is disposed in series in the circulation channel, and the liquid feeding pump is used. Pressure measurement inserted in the pressure measurement tube by circulating the sample to the circulation channel and enabling it to be introduced into the separation column, enabling recycle separation, and closing the other end of the pressure measurement tube Means and damper Reducing the volume, and significantly reduce the spread of the sample to move through the circulation flow path, together with precisely the separation at the time of recycling separated, together capable of simple and inexpensive construction of the column-switching system utilizing a recycle separation circuit , the liquid feed pump, sets the installation and operating conditions to analyze the conditions of the pump easily and quickly, and the purified or extracted work and analysis or preparative work easily so.
In the invention of claim 8, a detector is arranged inside or outside the circulation flow path, and when it is arranged inside, the state of recycling separation can be confirmed at any time, and precise analysis or extraction or purification work can be performed accurately. In addition, when placed outside, not only high pressure type detectors but also general detectors can be used, and dead volume caused by detectors can be avoided as in the case where they are placed in a circulation channel. , So that accurate analysis or extraction or purification can be performed.

According to the ninth aspect of the present invention, contaminant components other than the target component are removed through the switching operation of the recycle valve, the target component can be extracted or purified, and those operations can be performed rationally and easily. .
The invention according to claim 10 is characterized in that a switching valve is connected in parallel to one separation column disposed in the circulation flow path upstream of the recycling valve, and backflushing can be performed through operation of the switching valve. A backflush function can be obtained easily and inexpensively by a switching valve having a simple configuration.

  According to the eleventh aspect of the present invention, the recycling valve is attached to the introduction portion of the liquid feeding pump, the recycling valve can be communicated with the introduction portion of the liquid feeding pump, and recycled from the introduction portion of the liquid feeding pump, that is, the suction portion. The piping up to the valve is omitted, the piping of the relevant part is unnecessary, and the integrated structure of the inlet part of the pump and the recycling valve is realized, simplifying the configuration and making the installation space compact. At the same time, the dead volume of the part is set to a substantially zero value to suppress the dispersion of the sample so that a good separation can be obtained and the recycling separation can be performed accurately.

  In the first aspect of the present invention, a pressure measurement pipe having one end communicating with the circulation channel and the other end being occluded is provided, and the pressure measurement means and the damper are inserted into the pressure measurement pipe. By closing the other end of the measuring tube, the dead volume between the pressure measuring means inserted in the pressure measuring tube and the damper provided in the pressure measuring means is reduced, and the external volume of the separation column is greatly reduced. Thus, the diffusion of the sample moving through the circulation channel is greatly suppressed, and the flow rate is 0.4 mL / min. Recycling separation in the above class of liquid chromatographs is realized with a simple configuration and piping structure, and even in chromatograms with peak retention capacity of about 3.0 mL or more, sufficient liquid chromatographs for recycling separation are provided. Can be realized.

According to a second aspect of the present invention, the liquid feed pump is inserted into the circulation channel, a sample introduction part is provided in the circulation channel between the liquid feed pump and the separation column, and the sample introduction part and the liquid feed are provided. Since a connector having a plurality of ports is inserted between the pump and one end of the pressure measuring tube is connected to the port of the connector, pressure measurement by the pressure measuring means is enabled, and The pressure measuring tube, the pressure measuring means, and the damper can be separated from the circulation channel via the sample introduction unit.
In the invention of claim 3, since a detector is inserted in the circulation flow path, and the inner diameter of the downstream side portion of the circulation flow path communicating with the detector can be reduced, the back pressure against the downstream pipe line of the detector can be reduced. Can be set high, the inner diameter of the downstream pipe line can be reduced, and the separation column outer volume of the downstream pipe line can be reduced.

According to a fourth aspect of the present invention, the liquid chromatograph has a flow rate of 0.4 mL / min. As described above, it is possible to realize a liquid chromatograph that has a remarkable effect of reducing the external volume of the separation column.
According to the invention of claim 5, since the elution volume until the peak of the sample appears is about 3.0 mL or more, the liquid chromatograph improves the efficiency of recycle separation of the frequently used liquid chromatograph. can do.

  In the invention of claim 6, the recycling valve is attached to the introduction part of the liquid feeding pump, and the recycling valve can communicate with the introduction part of the liquid feeding pump. The piping from the recycle valve to the recycle valve is omitted, and the piping of the relevant part is unnecessary, and an integrated structure of the inlet part of the liquid feed pump and the recycle valve is realized, simplifying the structure and the space for installing them. In addition, the dead volume of the part is made substantially zero and the dispersion of the sample is suppressed, so that an excellent separation can be obtained.

The invention of claim 7 is provided with a pressure measurement pipe having one end connected to the circulation flow path and capable of closing the other end, and the pressure measurement means and the damper are inserted into the pressure measurement pipe, A separation column that is the same as or different from the separation column is arranged in series in the circulation channel, and the sample is circulated and moved to the circulation channel via a liquid feed pump, and can be introduced into the separation column for recycling separation. By closing the other end of the pressure measuring tube, the dead volume of the pressure measuring means and the damper inserted in the pressure measuring tube is reduced, and the sample moving through the circulation channel is diffused. significantly suppressed, together with the correctly perform the separation in time of recycling separation, with a column switching system utilizing a recycle separator may simple and inexpensive construction, the liquid feed pump The installation and operating conditions to analyze the conditions of the pump set easily and quickly, purified or extracted work and analysis or preparative work can be easily performed.
In the invention of claim 8, since the detector is arranged inside or outside the circulation flow path, when it is arranged inside, the state of recycling separation is confirmed at any time, and accurate analysis or extraction or purification work is performed. In addition, when placed outside, not only high pressure resistant detectors but also general detectors can be used, and dead volume caused by detectors can be avoided as in the case of placement in a circulating flow path. Accurate analysis or extraction or purification work

According to the ninth aspect of the present invention, since contaminant components other than the target component are removed and the target component can be extracted or purified through the switching operation of the recycle valve, these operations can be performed reasonably and easily. .
According to the invention of claim 10, a switching valve is connected in parallel to one separation column arranged in the circulation flow path upstream of the recycling valve, and backflushing can be executed through operation of the switching valve. Therefore, there is an effect that the back flush function can be obtained easily and inexpensively by the switching valve having a simple configuration.

  According to the eleventh aspect of the present invention, the recycling valve is attached to the introduction portion of the liquid feeding pump, the recycling valve can be communicated with the introduction portion of the liquid feeding pump, and recycled from the introduction portion of the liquid feeding pump, that is, the suction portion. The piping up to the valve is omitted, the piping of the relevant part is unnecessary, and the integrated structure of the inlet part of the pump and the recycling valve is realized, simplifying the configuration and making the installation space compact. At the same time, the dead volume of the part is made substantially zero to suppress the dispersion of the sample, and good separation can be obtained, and the recycling separation can be performed accurately.

It is explanatory drawing which shows the condition at the time of the column equilibrium of the liquid chromatograph to which this invention is applied. The main part of the present invention in a front view of expansion much shows piping condition of liquid feed pump and the connector and the pressure sensor. (A) is explanatory drawing which shows the condition at the time of recycling separation of the liquid chromatograph to which this invention is applied, (b) is an enlarged view of the recycling valve at the time of recycling separation.

It is a chromatogram which shows the condition which isolate | separates a single sample by the recycle separation by this invention. It is a chromatogram which shows the condition which isolate | separates a single sample by the recycle separation by a conventional apparatus. It is a chromatogram which shows the condition which isolate | separates the said single sample with a preparative column.

It is a chromatogram which shows the condition which isolate | separates two samples by the recycle separation by this invention. It is a chromatogram which shows the condition which isolate | separates two samples by the recycle separation by a conventional apparatus. It is a chromatogram which shows the condition which isolate | separates said two samples with a preparative column.

It is explanatory drawing which shows the 2nd Embodiment of this invention, and has shown the condition at the time of column equilibrium. It is a front view which shows the principal part of the said 2nd Embodiment, and has expanded and shown the piping condition of the recycle valve which concerns on this embodiment, a liquid feeding pump, a connector, and a pressure sensor. It is explanatory drawing which shows the condition at the time of the recycling separation of the said 2nd Embodiment. It is a chromatogram which shows the condition where two samples are recycle-separated by 2nd Embodiment using the said recycle valve | bulb.

It is a chromatogram which shows the condition which recycle-separates two samples with the conventional recycle valve | bulb used for 1st Embodiment. In the explanatory view showing the third embodiment of the present invention, a recycling separation / column switching system is configured by connecting two separation columns and a detector to the circulation flow path, and the situation of the system at the time of column equilibrium is shown. Show. It is explanatory drawing which shows the condition at the time of the recycling separation by the said 3rd Embodiment.

FIG. 6 is an explanatory diagram showing a fourth embodiment of the present invention, in which a detector and two separation columns are connected to a circulation flow path, a backflush switching valve is connected to one of the separation columns, and the recycling separation of the system is performed Shows the situation of time. It is a chromatogram which shows the separation condition which applied the back flush after detection of the two target components of vitamin D2 and D3 at the time of the recycle separation by the said 4th Embodiment. In the explanatory view showing the fifth embodiment of the present invention, two separation columns are connected to the circulation flow path, and a detector is connected to the outside of the circulation flow path to constitute a recycling separation / column switching system, It shows the situation when the system is separated for recycling.

It is explanatory drawing which shows the 6th Embodiment of this invention, and connects two separation columns to a circulation flow path, and connects the switching valve for backflushing to one of the separation columns, and is detected outside a circulation flow path. A recycle separation / column switching system is configured by connecting a vessel, and the situation at the time of recycle separation of the system is shown.

Explanation of symbols

2 Solvent 4, 4a Recycling valve 8, 9 Liquid feed pump (plunger pump)
21 Connector 22 Pressure measuring tube 25 Pressure measuring means (pressure sensor)
28 Sample introduction part (injection valve)
30, 35 Separation column 32, 32a Detector 36 Switching valve

Hereinafter, the flow rate of the present invention is 0.4 to 2.0 mL / min. The illustrated embodiment applied to a class of liquid chromatographs will be described. In FIGS. 1 to 3, reference numeral 1 denotes a container containing a solvent 2 therein, and one end of a pumping pipe 3 is accommodated in the container 1, and the other end. Is connected to a predetermined port P4 of the recycle valve 4 which is a high pressure resistant _four- port switching valve.

One end of the solvent conduit 5, one end of the reflux pipe 6, and one end of the drain pipe 7 are connected to the other ports P ₃ , P ₂ , P ₁ of the recycle valve 4. The remaining ports P ₅ and P ₆ have their external connection ports closed by appropriate means or are not formed with openings.
Among these, the other end of the solvent conduit 5 is connected to a T-shaped connector 10 disposed on the side of two plunger pumps 8 and 9 that are liquid feed pumps, and inlet pipes 11 and 12 are connected to the connecting portions on both sides thereof. It is connected.

  Inlet valves 15 and 16 and outlet valves 17 and 18 are attached to both sides of the pump heads 13 and 14 of the plunger pumps 8 and 9, and the other ends of the inlet pipes 11 and 12 are connected to the inlet valves 15 and 16, respectively. The outlet pipes 19 and 20 are connected to the outlet valves 17 and 18, and the other ends of the outlet pipes 19 and 20 are connected to the port of the connector 21.

The connector 21 has four ports that can communicate with each other, of which the outlet pipes 19 and 20 are connected to two ports, and the other port has one end of a pressure measuring tube 22 and a pump outlet. One end of the tube 23 is connected.
The pressure measuring tube 22 is disposed away from a circulation channel at the time of recycling separation, which will be described later, and one end of the measuring tube 22 is connected to the connector 21 and a drain tube 27 that can be closed is connected to the other end. A pressure measuring means and a damper, which will be described later, are inserted in the pressure measuring tube 22.

A pressure sensor 25, which is a pressure measuring means capable of measuring the pressure downstream of the liquid feed pump or the separation column pressure downstream of the liquid feed pump, is disposed facing the upstream side of the closing portion in the drain valve 24, Pressure measurement at column equilibrium and recycle separation is enabled.
In this case, the pressure sensor 25 allows various measuring methods.
In the figure, reference numeral 25a denotes a damper provided in the pressure sensor 25, which communicates with the pressure measuring pipe 22, and can buffer the pressure downstream of the liquid feed pumps 8 and 9 within the drain valve 24. A drain valve knob 27 is a drain pipe having one end connected to the drain valve 24.

The other end of the pump outlet pipe 23 is connected to a port of an injection valve 28 which is a sample introduction part, and a sample is supplied to the sample loop 28a from other ports via an injector (not shown). Injectable.
The sample loop 28 a is connected to one end of the sample introduction tube 29 through the switching operation of the injection valve 28 so as to be disconnected or communicated, and the other end of the tube 29 is connected to the separation column 30.
In the figure, 34 is a column oven that accommodates the injection valve 28, the sample introduction tube 29, the separation column 30, the detector 32, and the like. For convenience, the detector 32 is illustrated outside the column oven 34.

  One end of an elution tube 31 is connected to the elution side of the separation column 30 and the other end is connected to a detector 32. The detector 32 of the embodiment uses a high-pressure resistant UV detector having a UV cell 33, and the other end of the reflux pipe 6 is connected to the detector 32. The UV cell 33 of the embodiment has a pressure resistance of 5 MPa.

Among the pipes, the solvent conduit 5 and the reflux pipe 6, the inlet pipes 11 and 12 and the outlet pipes 19 and 20, the pump outlet pipe 23, the sample introduction pipe 29, and the elution pipe 31 provide a solvent circulation channel during the recycling separation. It is possible to form.
In this case, the drain valve 24, the pressure measuring tube 22, the pressure sensor 25, the damper 25a, and the like constitute an external volume of the separation column, but are spaced apart from the circulation flow path as described above, and at the time of recycling separation Since the drain valve 24 is closed and one end of the pressure measuring tube 22 is closed, it cannot be a flow path or circulation path for the sample and the solvent, and does not form a substantial circulation path.

  Among these circulation channels, the length of each tube is the longest in the elution tube 31, and then the reflux tube 6, the pump outlet tube 23, the solvent conduit 5, the sample introduction tube 29, the outlet pipes 19 and 20, and the inlet pipe. 11 and 12, the solvent conduit 5 is about 1/3 of the elution tube 31, the sample introduction tube 29 is about 1/4 of the elution tube 31, and the outlet pipes 19 and 20 and the inlet pipes 11 and 12 are elution tubes. About 1/8 of 31.

  Among the circulation channels, the inner diameter of the solvent conduit 5 and the inlet pipes 11 and 12 are the largest, and all others are formed to have the same diameter. For example, the reflux pipe 6 on the smaller diameter side is approximately the size of the solvent conduit 5. It is formed in 1/3.

  And the separation column outer volume of the circulation channel is the largest at the damper 25a, the pump heads 13 and 14 parts (for two groups), then the solvent conduit 5 parts, the inlet pipes 11 and 12 parts (for two pipes), 31 parts of elution pipes, 6 parts of reflux pipes, 23 parts of pump outlet pipes, 19 and 20 outlet pipes (for two pipes), and 29 parts of sample introduction pipes are arranged in this order.

Among these, the volume of the pump heads 13 and 14 is about 7% of the total volume outside the separation column, the volume of the solvent conduit 5 is about 6% of the total volume outside the separation column, and the inlet pipes 11 and 12 are outside the separation column. About 5% of the total volume, 31 parts of the elution tube is about 2% of the total volume outside the separation column, about 1 / 3.5 of the volume of the pump heads 13 and 14 parts, and 6 parts of the reflux pipe is the total volume outside the separation column. It is about 1.7%, and is set to about 1/4 of the volume of the pump heads 13 and 14 part.
In this case, the drain valve 24, the pressure measuring pipe 22, and the pressure measuring means 25 and the damper 25a inserted in the pressure measuring pipe 22 are about 76% of the external volume of the separation column and occupy most of the volume. Since the circulation channel is not formed during the recycling separation, there is no influence of the dead volume during the recycling separation.

The liquid chromatograph of the present invention configured as described above includes the solvent container 1, the recycle valve 4, the liquid feed pumps 8 and 9, the pressure sensor 25, the injection valve 28, the separation column 30, and the detector 32. .
The configuration is substantially the same as that of the existing one, but the detector 32 equipped with a high-pressure resistant UV cell 33 is used, and the inner diameter of the adjacent pipe is reduced as described later, so that the outer volume of the separation column is reduced. We are trying to reduce it.

  The piping structure of the liquid chromatograph is as shown in FIGS. 1 and 2, the solvent container 1 and the recycle valve 4 are connected by the pumping pipe 3, and the recycle valve 4 and the liquid feed pumps 8 and 9 are connected to the solvent conduit 5. The pumps 8 and 9 and the injection valve 28 are connected by a pump outlet pipe 23, the injection valve 28 and the separation column 30 are connected by a sample introduction pipe 29, and the separation column 30 and the detector 32 are eluted. The detector 31 and the recycling valve 4 are connected by a reflux pipe 6.

The piping structure is substantially the same as that of the existing one. However, as described above, the detector 32 having the high-pressure resistant UV cell 33 is used, and the back of the reflux pipe 6 of the circulation flow path, which is an adjacent pipe. The pressure can be increased, and the inner diameter of the reflux pipe 6 is reduced.
In the embodiment, by using the detector 32, the inner diameter of the reflux pipe 6 is reduced from 0.33 mm to 0.13 mm.

  Thus, by reducing the inner diameter of the reflux pipe 6 of the circulation flow path, the outer volume of the separation column in the portion is reduced, and the diffusion of the sample in the outer volume of the column during the recycle separation can be suppressed.

  On the other hand, in the piping structure, the drain valve 24 and the pressure measuring pipe 22 are arranged apart from the circulation flow path as described above, and the drain valve 24 is closed during recycling separation, and one end of the pressure measuring pipe 22 is disposed. Therefore, the part cannot be a flow path or a circulation path for the sample and the solvent at the time of recycling separation, and cannot be separated from the circulation path to form a substantial circulation path.

  The drain valve 24 and the pressure measuring tube 22 occupy about 76% of the separation column outer volume, but do not form a circulation flow path at the time of recycling separation, so that the separation column outer volume at the time of recycling separation is greatly reduced. Diffusion of the sample in the column external volume can be greatly suppressed.

  The effect of suppressing the diffusion of the sample in the outer volume of the separation column at the time of such recycling separation can be obtained relatively easily by the above-described piping structure, and the desired effect can be obtained with a simple configuration.

Next, when recycle separation is performed by the liquid chromatograph, the separation column 30 is equilibrated in advance.
At that time, the drain valve 24 is opened in advance, the drain side end of the pressure measuring tube 22 is opened, the plunger pumps 8 and 9 are started, the solvent 2 is pumped up from the container 1, and this is connected to the solvent conduit 5 and the connector 10. Then, it moves to the connector 21 via the pump heads 13 and 14 and the outlet pipes 19 and 20, and further moves to the drain valve 24 and the drain pipe 27 via the pressure measuring pipe 22.

Thereafter, the drain valve 24 is closed, the drain side end of the pressure measuring tube 22 is closed, and the solvent 2 is introduced from the pump outlet tube 23 into the separation column 30 through the sample introduction tube 29, and a part thereof. Is led from the connector 21 to the pressure measuring tube 22, and the measuring tube 22 and the drain valve 24 are filled.
A part of the solvent 2 is moved from the elution tube 31 to the detector 32, led from the reflux tube 6 to the recycle valve 4, and discharged from the drain tube 7.

  In this case, one end of the pressure measurement pipe 22 communicates with the pump outlet pipe 23 via the connector 21 and the pressure sensor 25 is disposed facing the pressure measurement pipe 22. The pressure in 22 or the pressure downstream of the plunger pumps 8 and 9 can be measured.

Thereafter, the injection valve 28 is switched, the sample loop 28a is disconnected from the sample introduction tube 29, and the sample is injected into the sample loop 28a.
Next, the injection valve 28 is switched, the sample loop 28 a is communicated with the sample introduction tube 29, the sample is sent together with the solvent 2 to the sample introduction tube 29, and these are introduced into the separation column 30.
The sample is separated while moving through the separation column 30, and a part of the sample is eluted. The sample moves along the elution tube 31 together with the solvent and is guided to the detector 32.

  The component to be recycled in the sample is detected by the detector 32 to form a peak in the chromatogram, and the recycling valve 4 is switched before a known time for forming the peak, and the drain pipe 7 is turned on. In addition, the injection valve 28 is switched and the sample loop 28a is separated from the sample introduction pipe 29 to form a circulation flow path, thereby performing substantial recycle separation.

  In the recycle separation, the recycle target component led to the detector 32 is led to the recirculation pipe 6 together with the solvent 2 and moved to the recycle valve 4, from which the solvent conduit 5, connector 10, pump head 13, 14. Move to connector 21 via outlet pipes 19 and 20.

  In this case, one end of the pressure measurement pipe 22 communicates with the pump outlet pipe 23 via the connector 21 and the pressure sensor 25 is disposed facing the pressure measurement pipe 22. The pressure in 22 or the pressure downstream of the plunger pumps 8 and 9 can be measured.

  At the time of such recycle separation, the pressure measuring tube 22 is filled with the solvent 2 as described above, and one end thereof is closed by the drain valve 24 to maintain a constant pressure state. It cannot be a flow path or a circulation path for the sample and the solvent 2, and does not form a substantial circulation path.

  For this reason, the sample and the solvent 2 move to the pump outlet pipe 23 side and are guided to the injection valve 28, move the sample introduction pipe 29 while avoiding the sample loop 28a, and separate the separation column from the introduction pipe 29. 30 is moved, and during the movement, the sample is eluted and the first recycle separation is performed.

Thereafter, the separated eluate moves along the elution tube 31 together with the solvent 2 and is guided to the UV cell 33 of the detector 32, where the elution component is detected by the UV cell 33 and moves through the reflux tube 6. Then, it is guided to the recycle valve 4.
In this way, the first recycle separation is performed, and thereafter, the recycle separation is performed a desired number of times.

When such recycle separation according to the present invention was performed on a single sample (vanillic acid), a chromatogram as shown in FIG. 4 was obtained.
In the figure, the first peak is a liquid chromatographic peak before the transition to recycle separation, and the second to sixth peaks are recycle separation peaks.

The second to sixth peaks are generally sharp and good, and the peak height gradually decreases with each recycle, the peak width gradually increases, and the sample diffuses slowly. I'm angry
The second to sixth peaks have substantially the same shape except that the peak height is slightly lower than the chromatogram obtained by the preparative column shown in FIG. It was done.
In the above experiment, the holding capacity at the first peak is about 10 mL. However, with the apparatus of the present invention, the elution volume of the peak, which has not been sufficient for recycle separation in the past, is 4-30. It was confirmed by the inventors' experiment that the desired separation effect can be obtained even in the recycle separation using mL liquid chromatograph. That is, it was confirmed that separation with a small elution volume was possible.

On the other hand, in FIG. 5, the drain valve 24 and the pressure measurement pipe 22 in which the pressure sensor 25 is arranged are inserted between the outlet pipes 19 and 20 and the pump outlet pipe 23, and pass through the pressure measurement pipe 22 during recycling separation. FIG. 2 shows a chromatogram obtained by recycling and separating the same sample under the same conditions for a conventional apparatus that circulates the sample.
In the figure, the first peak is a liquid chromatographic peak before the transition to recycle separation, and the second to sixth peaks are recycle separation peaks.

  The second to sixth peaks are lower in height and wider than the peaks of the present invention shown in FIG. In particular, the 3rd to 6th peaks have a low peak height and become round, the width of the peak is considerably widened, and the collapse of the peak becomes remarkable, and the sample is diffused rapidly. Panicked.

  FIG. 6 shows a chromatogram separated by a preparative column with the sample injection amount and moving flow rate greatly increased for the above-mentioned sample, and the peaks due to recycle separation appearing in the 2nd to 5th are all sharp. The peak height is gradually lower, but the peak height is generally high and the spread of the peak width is suppressed.

Next, two types of samples (caffeic acid and vanillic acid) were subjected to recycling separation according to the present invention, and a chromatogram as shown in FIG. 7 was obtained.
In the figure, the first peak is a liquid chromatographic peak before the transition to recycle separation, and the second to sixth peaks are recycle separation peaks.

In the second to sixth peaks, two peaks corresponding to the two samples appear, and they are gradually separated from each other. -The height of the peak gradually decreases with each recycle, but it can be seen that the peak width widens and the sample diffuses slowly.
The second to sixth peaks have substantially the same shape except that the peak height is slightly lower than the chromatogram obtained by the preparative column shown in FIG. It was done.

  In the above experiment, the elution volume at the first peak was about 10 mL, but the elution volume of the peak, which was not sufficient for recycle separation by the apparatus of the present invention, was 4-30. The inventors' experiments confirmed that the desired effect can be obtained even in recycle separation using mL liquid chromatography. Even in this case, it was confirmed that separation with a small elution volume was possible.

On the other hand, FIG. 8 shows a chromatogram obtained by recycling and separating the same two types of samples as those described above under the same conditions for the conventional apparatus used in FIG.
In the figure, the first peak is a liquid chromatographic peak before the transition to recycle separation, and the second to sixth peaks are recycle separation peaks.

In the second to sixth peaks, two peaks corresponding to the two samples appear, respectively. These peaks are different from those of the present invention in FIG. Its height is very low and its peak width is wide.
In particular, the 3rd to 6th peaks have a low peak height and become round, the width of the peak is considerably widened, and the collapse of the peak becomes remarkable, and the sample is diffused rapidly. Panicked.

  FIG. 9 shows chromatograms separated by a preparative column for the above-mentioned two types of samples, with the injection amount and transfer flow rate of the above-mentioned samples being greatly increased. All of the two peaks are sharp and good, and the peak height is gradually lower. However, the peak height is generally high and the spread of the peak width is suppressed.

  Based on the result of such recycle separation according to the present invention, the number of theoretical plates is calculated, and when this is plotted on the vertical axis and the number of passes through the separation column 30 on the horizontal axis, Formed a straight line going up to the right approximately proportional to the number of times, and it was confirmed that it passed through the zero point, and it was confirmed that it had good separation characteristics.

  On the other hand, when the above correlation diagram is created based on the result of the recycling separation by the conventional apparatus, a straight line with a downward slope in which the theoretical plate number is approximately inversely proportional to the number of times or a straight line substantially parallel to the horizontal axis is formed. It was confirmed that it has insufficient separation characteristics.

As described above, the liquid chromatograph of the present invention has a simple configuration and piping structure to reduce the external volume of the separation column, suppress the diffusion of the sample in the external volume of the separation column, and 0.4 to 2.0 mL / min. Recycling separation of a liquid chromatograph with a class flow rate can be realized.
Of course, the present invention is useful not only for recycle separation at a low flow rate but also for recycle separation at a large flow rate, for example, in a large volume fractionation system.

10 to 20 show another embodiment of the present invention, and the same reference numerals are given to the portions corresponding to the above-described configuration.
Of these, FIGS. 10 to 14 show a second embodiment of the present invention, and propose a recycle valve 4a which is an improvement over the conventional recycle valve 4 of the above embodiment.
That is, in the above-described embodiment, the suction pressure between the suction side of the plunger pumps 8 and 9 and the recycle valve 4 is the suction pressure, so the inner diameter of the pipe between them cannot be made constant or less, and the dead volume of the part concerned There is a limit to the reduction of

This embodiment solves the above-mentioned problem, and the improved recycle valve 4a has ports P ₅ , P in which the external connection port of each port is closed by an appropriate means or no opening is formed. one port of ₆ - opens the door P _5,該Po - - DOO, ports in the embodiment allows connecting the door P ₅ to the outside, the other port - DOO P ₆ is closed the external connection port as it is, this A new recycling valve 4a is integrally mounted at the inlets of the plunger pumps 8 and 9.

That is, the port P ₅ is connected to one end of the pumping pipe 3, the port P ₄ is connected to one end of the inlet pipe 11, and the port P ₃ is connected to one end of the inlet pipe 12. The port P ₂ is connected to one end of the reflux pipe 6, the port P ₁ is connected to one end of the drain pipe 7, and the ports P ₃ and P ₄ are connected to each other. It is always in communication.

Therefore, the port - DOO _P 3, _{P 4} are always communicates with the inlet pipe 11, and because communicating with recycling valve 4a, the port - DOO _P 3, _{P 4} are equivalent to the connector 10 of the embodiment The recycle valve 4a can be regarded as having the connector 10 built therein.
For this reason, the connector 10 and the solvent conduit 5 connecting the connector 10 and the recycle valve 4a can be omitted, the number of parts can be reduced, the configuration can be simplified, and the entire dead volume can be reduced. By reducing the dead volume corresponding to the solvent conduit 5 occupying about 6%, it is possible to suppress the diffusion of the sample and to expect a good recycle separation.

  Therefore, when the recycling separation is performed using the recycling valve 4a, the piping structure is as shown in FIG. 12, and the recycling valves 4a can be arranged on the inlet pipes 11 and 12 side. Compared with the case where it does, rational installation can be aimed at, the installation space can be made compact, and the piping space of the solvent conduit 5 can be made compact, and the piping space can be made compact.

Furthermore, the recycling valve 4a is port - Remove stopcock of bets P ₅ (not shown), or can be connected to the outside perforated, in the case of existing devices remove the solvent conduit 5,該Po - DOO P with connecting one end portion of the pumping pipe 3 to _5, communication slit is formed, and port - DOO P _4, can piping easy because the P ₃ may be connected to one end of the inlet pipe 11 and 12, existing recycling The modification of the valve 4 can be easily accommodated.

Next, when recycle separation is performed by the liquid chromatograph, the separation column 30 is equilibrated in advance. This equilibration operation is performed substantially in the same manner as in the above-described embodiment. However, the solvent 2 can quickly move on the above-described equilibration path and the equilibration processing time can be shortened by the omission of the piping of the solvent conduit 5. .
That is, in the equilibration operation, the drain valve 24 is opened, the drain side end of the pressure measuring tube 22 is opened, the plunger pumps 8 and 9 are started, the solvent 2 is pumped up from the container 1, and this is recycled valve. 4a port of - leads to the door _{P 5.}

Then, the solvent 2 is diverted to the inlet pipes 11 and 12 from the ports P ₄ and P ₃ communicating with the port P ₅ , and moved from the pump heads 13 and 14 and the outlet pipes 19 and 20 to the connector 21. Then, it moves to the drain valve 24 and the drain pipe 27 through the pressure measuring pipe 22.

Thereafter, the drain valve 24 is closed, the drain side end of the pressure measuring tube 22 is closed, the pump outlet tube 23 is moved to the injection valve 28, and the sample introduction tube 29 is moved to the first column 30. Then, a part thereof is led from the connector 21 to the pressure measuring tube 22 and the measuring tube 22 and the drain valve 24 are filled.
Meanwhile, the part of the solvent 2 to move from the elution tube 31 to the detector 32, it moves to P ₂ recycling valve 4a via a return pipe 6 from the detector 32,該Po - port communicating with preparative P ₂ - discharged by moving from door P ₁ to the drain pipe 7. This situation is as shown in FIGS.

Next, the injection valve 28 is switched, the sample loop 28a is disconnected from the sample introduction tube 29, and the sample is injected into the sample loop 28a.
Thereafter, the injection valve 28 is switched, the sample loop 28a is communicated with the sample introduction tube 29, the sample is sent together with the solvent 2 to the sample introduction tube 29, and these are introduced into the separation column 30.
The sample is separated while moving through the separation column 30, and a part of the sample is eluted. The sample moves along the elution tube 31 together with the solvent and is guided to the detector 32.

The component to be recycled in the sample is detected by the detector 32 to form a peak in the chromatogram, and the recycling valve 4a is switched before a known time for forming the peak. with closing the door P _1, switching the injection valve 28, the sample Le - the flop 28a separately from the sample inlet tube 29 to form a circulation flow path, performing a substantial recycling separation.

  In the recycle separation, the recycle target component led to the detector 32 is guided to the recirculation pipe 6 together with the solvent 2 and moved to the recycle valve 4a, from which the solvent conduit 5, the connector 10, the pump head 13, 14. Move to connector 21 via outlet pipes 19 and 20.

  In this case, one end of the pressure measurement pipe 22 communicates with the pump outlet pipe 23 via the connector 21 and the pressure sensor 25 is disposed facing the pressure measurement pipe 22. The pressure in the pipe 22 or the pressure downstream of the plunger pumps 8 and 9 can be measured.

  At the time of such recycle separation, the pressure measuring tube 22 is filled with the solvent 2 as described above, and one end thereof is closed by the drain valve 24 to maintain a constant pressure state. It cannot be a circulation path for the sample and the solvent 2 and is separated from the substantial circulation path.

  For this reason, the sample and the solvent 2 move to the pump outlet pipe 23 side and are guided to the injection valve 28, move the sample introduction pipe 29 while avoiding the sample loop 28a, and separate the separation column from the introduction pipe 29. 30 is moved, and during the movement, the sample is eluted and the first recycle separation is performed.

Thereafter, the separated eluate moves along the elution tube 31 together with the solvent 2 and is guided to the UV cell 33 of the detector 32, and the elution component is detected by the UV cell 33, as shown in FIG. Then, the recirculation pipe 6 is moved and led to the recycle valve 4a.
In this way, the first recycle separation is performed, and thereafter, the recycle separation is performed a desired number of times.
The recycle separation is performed in substantially the same manner as in the above-described embodiment. However, the sample and the solvent can quickly move through the recycle path, and the recycle separation time can be shortened, because the piping of the solvent conduit 5 is omitted.

In order to verify the performance of the improved recycle valve 4a, the conventional recycle valve 4 was analyzed under the same conditions as the improved recycle valve 4a, and a chromatogram as shown in FIG. 14 was obtained.
Comparing these chromatograms, in the chromatogram of the new recycling valve 4a, each peak after the start of recycling separation is a peak compared to the corresponding peak of the chromatogram by the old recycling valve 4. It was confirmed that the sample was high, the peak width was narrow, and the diffusion of the sample was suppressed. This is considered to be due to the effect of suppressing the sample diffusion accompanying the reduction of the dead volume due to the omission of the solvent conduit 5 described above.

  Further, in the chromatograms of FIGS. 13 and 14, the first peak appears 7 to 8 minutes after the start of analysis, and the elution amount in this case is considered to be 2.8 to 3.2 mL. It was confirmed that separation was possible with an elution volume of 8 mL or more.

15 to 20 show a liquid chromatograph in which the improved recycle valve 4a is applied to a column switching system, and the flow rate of the liquid chromatograph in this case is 0.4 to 2.0 mL / min. It is not limited to.
Of these, FIGS. 15 and 16 show a third embodiment of the present invention. In this embodiment, a detector 32 is provided in a recycle separation circuit composed of a liquid feed pump and an injection valve, a separation column and a recycle valve. Applies to intervening systems.

  That is, the recycling separation circuit includes a set of two plunger pumps 8 and 9 that can prevent pulsating flow, and thus substantially equivalent to one liquid feed pump, an injection valve 28 that can introduce a sample, A separation column 30 (hereinafter referred to as a first column) capable of roughly separating a sample and a recycle valve 4a, the detector 32 being inserted in series in the circuit, and the liquid feed pumps 8, 9 Between the first column 30 and the first column 30, a second column 35 capable of separating components difficult to separate by the first column 30 is inserted in series.

Various detectors 32 are used depending on the analysis conditions, but a high pressure resistant type is desirable for insertion in the recycling separation circuit.
Further, the first and second columns 30 and 35 are the same or different from each other, and in the embodiment, different ones that can separate different components are used. (Registered trademark) ODS-P, Inertsil (registered trademark) ODS-EP having a stronger polar interaction than the first column 30 is used for the second column 35.

When the recycle separation is performed by the liquid chromatograph, the first and second columns 30 and 35 are equilibrated in advance.
The balancing operation, except that movement of the solvent 2 to the pump outlet pipe 23 and the second shift and dynamic to the column 35, the first column 30 through the sample introduction pipe 29 from the injection valve 28, a second embodiment In this case, the solvent 2 can be moved quickly through the equilibration path and the equilibration time can be reduced.

  After the equilibration operation, the injection valve 28 is switched, the sample loop 28a is disconnected from the sample introduction tube 29, the sample is injected into the sample loop 28a, and after the sample is introduced, the injection valve 28 is switched and the sample loop is switched. 28 a is communicated with the sample introduction tube 29, the sample is sent together with the solvent 2 to the sample introduction tube 29, and these are introduced into the first column 30.

While the sample is moving through the first column 30, a plurality of components including the target component and the contaminating component are roughly separated and a part is eluted, and these are moved along the elution tube 31 together with the solvent to detect the detector 32. In the UV cell 33, a contaminant component that is eluted earlier than the target component is detected and appears in the chromatogram.
Thereafter, when the solvent containing the elution component is guided to the reflux pipe 6 and moves to the recycle valve 4a, the precipitating contaminant component is discharged from the recycle valve 4a to the drain pipe 7, and elution is performed earlier than the target component. Remove impurities

After discharging the contaminated components, the recycle valve 4 a is switched, and the sample and solvent containing the target components are sent to the pump outlet pipe 23 via the liquid feed pumps 8, 9 and moved to the second column 35.
This situation is as shown in FIG.

  Therefore, the recycle valve 4 a is switched to allow the contaminants eluted from the first column 30 to flow into the drain pipe 7, and the sample loop 28 a is separated from the sample introduction pipe 29. This situation is as shown in FIG.

The recycle separation is performed by circulating a plurality of components including the target component together with the solvent 2 in the circulation channel and repeating the separation by the first and second columns 30 and 35 inserted in the circulation channel. .
At that time, the target component or the plurality of components elutes while moving through the first and second columns 30 and 35, and the eluted components are detected by the UV cell 33 and appear in the chromatogram, which are eluted earlier than the target components. In the case of other contaminant components, the recycle valve 4a is switched and discharged from the drain pipe 7.
Thereafter, the recycle valve 4a is switched, and a plurality of components including the target component are circulated through the circulation channel together with the solvent 2 to sequentially remove the contaminated components and purify the target components.

The target component elutes while moving through the first and second columns 30 and 35, and is detected by the UV cell 33 and appears in the chromatogram. When the target component is completely separated, the recycle valve 4a is switched to drain the target component. Lead to tube 7 and dispense.
That is, in the recycle separation, column switching is performed by the first and second columns 30 and 35, and the target component can be purified or concentrated and extracted.

  As described above, in the third embodiment, column switching is performed at the time of recycle separation, and the sample and the solvent 2 are substantially circulated through the circulation flow path by one liquid feed pump 8 and 9. Compared to those requiring multiple circulation pumps, the configuration is simple and inexpensive, and compared to those requiring multiple circulation pumps, setting of pump installation and operating conditions and mobile phase analysis conditions are set. Etc., and analysis or fractionation can be easily performed.

In addition, the circulation channel is equipped with the above-mentioned new type of recycling valve 4a, and by reducing the dead volume, it is possible to suppress the diffusion of the sample and obtain a reliable separation. Can be purified or concentrated and extracted or separated.
Furthermore, since the detector 32 is inserted in the circulation channel, the elution state and the separation state of the sample circulating in the circulation channel can be confirmed at any time, and the target component can be purified or concentrated to accurately extract or sort. Yes.

  FIG. 17 shows a fourth embodiment of the present invention. This embodiment relates to an application form of the third embodiment, and a backflush switching valve 36 is arranged in parallel with the first column 30 in the third embodiment. After the detection or extraction of the target component, the first column 30 is washed by backflushing.

At that time, a four-way valve is used as the switching valve 36, and one end portions of the sample introduction tube 29a, the elution tube 31, the elution tube 31a, and the sample introduction tube 29 are connected to the ports a to d.
The switching valve 36 communicates the port a and the port d as shown by a solid line at the time of recycling separation, and communicates the solvent through the port b and the port c as shown by a solid line. The first column 30 is movable in the forward direction.
When backflushing, the switching valve 36 is switched as indicated by a broken line, port d and port c are communicated, port a and port b are communicated, and the solvent is indicated as indicated by a broken arrow. In addition, the first column 30 is movable in the reverse direction. This situation is as shown in FIG.

In the fourth embodiment, after the equilibration operation as described above, the recycling valve 4a is switched to separate and separate.
In this embodiment, as in the third embodiment, column switching is performed under recycle separation, contaminant components eluting before and after the target component are removed, and the target component is extracted or purified and backflushed.

This situation is as shown in FIG. 18. Two samples (vitamin D2 and vitamin D3) having different elution times of components and having contaminating components between the elution times are column-switched under recycle separation.
That is, after the sample is introduced, the injection valve 28 is switched to “Load” at the point of “2” and moved to the sample introduction tube 29 together with the solvent 2, and one vitamin D2 component is eluted and completely separated. At the point of “3” when starting to perform, the recycle valve 4a is turned on, and the recycle separation is performed by switching to the recycle separation.

Then, when the vitamin D2 component is completely eluted, the recycle valve 4a is turned off at the time point "4", and the recycle valve 4a is switched to guide the contaminated component between the vitamin D2 and the vitamin D3 to the drain pipe 7.
Thereafter, when the other vitamin D3 component elutes and begins to be completely separated, the recycling valve 4a is turned on to switch to recycling separation, and when the vitamin D3 component is completely eluted, the time point "6" is reached. Then, the recycle valve 4a is turned off, and the recycle valve 4a is switched to guide the contaminant components after vitamin D3 to the drain pipe 7.

Then, after separating or extracting the target ingredients vitamin D2 and D3, the backflush is turned on at the time of “7”, the switching valve 36 is switched as shown by a broken line, and the solvent is changed to the first as shown by the broken line. The column 30 is moved in the reverse direction, and the contaminated components remaining in the first column 30 are washed efficiently and quickly, and this is discharged from the drain pipe 7 together with the solvent or the like by switching the recycle valve 4a.

  As described above, the fourth embodiment shares the advantages of the third embodiment described above, and uses a simple switching valve 36 in a system capable of performing recycle separation and column switching, thereby providing a backflush function. Is realized easily and inexpensively.

Figure 19 shows a fifth embodiment of the present invention, this embodiment relates to another aspect of the third embodiment, port of recycling Li valves 4a - connecting one end of the connection pipe 37 to the preparative P ₁ A detector 32a is connected to the other end, and a drain tube 7 is connected to the detector 32a.
That is, in this embodiment, the detector 32a is arranged outside the recycling separation circuit, and a general detector can be adopted as the detector 32a.

  Further, by arranging the detector 32a as described above, the dead volume by the detector 32a is reduced and the sample is prevented from being diffused and separated as compared with the case where the detector 32a is arranged inside the recycling separation circuit. The accuracy can be improved. The operation of the fifth embodiment is substantially the same as that of the third embodiment, and shares the advantages described above.

  In the embodiment, when the component eluted in the first column 30 or the second column 35 is detected by the detector 32a, the eluted component is moved to the recycle valve 4a together with the solvent, and the recycle valve 4a is switched, and the elution component is sent to the detector 32a through the connecting pipe 37 and detected by the detector 32a. After the detection, the eluted component is discharged from the drain pipe 7 or collected.

Figure 20 shows a sixth embodiment of the present invention, this embodiment relates to a modified embodiment of the fifth embodiment, the switching valve 36 for backflushing the first column 30 of definitive to the fifth embodiment The first column 30 is connected in parallel, and after the detection or extraction of the target component, the first column 30 is washed by backflushing. The configuration and operation of the switching valve 36 are substantially the same as those in the fourth embodiment.

  In addition to sharing the advantages of the fifth embodiment described above, the sixth embodiment uses a simple switching valve 36 in a system capable of performing recycle separation and column switching, thereby making the backflush function easy and inexpensive. It has been realized.

  As described above, the liquid chromatograph of the present invention has a simple configuration and a piping structure that reduces the external volume (dead volume) of the separation column and suppresses the diffusion of the sample by the dead volume. min. In addition to realizing recycling separation in the above-mentioned class of liquid chromatographs, a column switching system using a recycling separation circuit is configured in a simple and inexpensive manner, facilitating analysis, fractionation, extraction and purification, and separation of target components. -It is suitable for recycle separation of, for example, a high performance liquid chromatograph (HPLC) because it facilitates purification and promotes high quality purification of a plurality of components.

Claims (11)

  A circulation channel that is interposed between a separation column and a recycle valve and can circulate a sample, a liquid feed pump, a pressure measuring means that can measure a pressure downstream of the liquid feed pump, and a downstream of the liquid feed pump In a liquid chromatograph equipped with a damper capable of buffering the pressure on the side, a pressure measuring tube having one end communicating with the circulation channel and the other end being occluded is provided, and the pressure measuring tube is provided with the pressure A liquid chromatograph characterized by interposing a measuring means and a damper.   The liquid supply pump is inserted in the circulation flow path, a sample introduction part is provided in the circulation flow path between the liquid supply pump and the separation column, and a plurality of pieces are provided between the sample introduction part and the liquid supply pump. 2. The liquid chromatograph according to claim 1, wherein a connector having a port is inserted, and one end of the pressure measuring tube is connected to the port of the connector.   The liquid chromatograph according to claim 1, wherein a detector is inserted in the circulation channel, and the inner diameter of the downstream side portion of the circulation channel communicating with the detector can be reduced.   The liquid chromatograph has a flow rate of 0.4 mL / min. The liquid chromatograph according to any one of claims 1 to 3, which is as described above.   The liquid chromatograph according to any one of claims 1 to 4, wherein the liquid chromatograph has an elution volume of about 3.0 mL or more until a peak of the sample appears.   The liquid chromatograph according to claim 1, wherein the recycling valve is attached to the introduction part of the liquid feeding pump, and the recycling valve can communicate with the introduction part of the liquid feeding pump. A circulation channel that is interposed between a separation column and a recycle valve and can circulate a sample, a liquid feed pump, a pressure measuring means that can measure a pressure downstream of the liquid feed pump, and a downstream of the liquid feed pump In a liquid chromatograph equipped with a damper capable of buffering the pressure on the side and a detector, a pressure measuring tube having one end communicating with the circulation channel and the other end being occluded is provided, and the pressure measurement with interposing said pressure measuring means and the damper tube, the separation column is the same as or dissimilar separation column to the circulation flow path arranged in series, the specimen via the liquid feed pump to the circulation flow path A liquid chromatograph characterized in that it is circulated and provided so as to be able to be introduced into the separation column so that recycle separation can be performed.   The liquid chromatograph according to claim 7, wherein a detector is disposed inside or outside the circulation channel.   8. The liquid chromatograph according to claim 7, wherein contaminant components other than the target component are removed through the switching operation of the recycle valve so that the target component can be analyzed and extracted or purified.   8. The liquid chromatograph according to claim 7, wherein a switching valve is connected in parallel to one separation column disposed in a circulation flow path upstream of the recycling valve, and backflushing can be performed through operation of the switching valve. Graph.   The liquid chromatograph according to claim 7, wherein the recycling valve is attached to the introduction portion of the liquid feeding pump, and the recycling valve can communicate with the introduction portion of the liquid feeding pump.

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