JPS5848851A - high performance liquid chromatograph - Google Patents

Abstract

PURPOSE:To send a trace of a constant flow rate accurately with good reproducibility, by carrying out a piston drive by liquid pressure. CONSTITUTION:The area of the rear end face which is in contact with the moving phases A1, B2 of pistons 9, 9' of liquid sending pumps A3, B4, is made smaller than that of the rear end which is in contact with a liquid 30 for driving. Each pump A3, B4 is filled with each phase A1, B2 and suction valves 22, 22' are closed and then, jetting valves 22, 23 are opened. Further, valves 16, 16', 15, 15' are closed and valves 17, 17' are opened. Next, liquid sending of a pump 28 for driving is started and at the same time, a valve 14 or 14' is opened. The phases A1, B2 are sent out at an optional proportion by opening and closing the valves 14, 14' alternately for every prescribed time. Both phases are mixed in a mixer 5 and the mixture is introduced into a column 7 through a sample injector 6 and then, is discharged through a discharge device 8.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a high performance liquid chromatograph, particularly a high performance liquid chromatograph suitable for micro high performance liquid chromatography.

Conventional high performance liquid chromatography generally uses an inner diameter of 4
Using a collar with a length of approximately 100 to 50 cm, the mobile phase was applied at a rate of 1.5 mm per minute. The analysis was conducted at a flow rate of around mt [7]. The number of theoretical plates, which is a measure of the separation capacity, is most often used to be around one column, but recently, attempts have been made to increase the number of theoretical plates of this column to the highest level in history to realize liquid chromatography with even higher resolution. is becoming popular. This is due to the spread of high-performance liquid chromatography, which has broadened the scope of analysis and the need for columns with a higher molecular weight of 11fff.
A method has been announced in which a stainless steel thin tube with an inner diameter of about 0.5 mm is used with a thin tube column filled with conventional column filler X7 agent gold. In case of B, the length of the column is 1m to 1.
With a Q I of about 11, the number of theoretical plates is several tens of stone steps, which is an order of magnitude higher than the conventional one.
It is from 1/00 to 1000 minutes σ month. This low consumption of mobile phase solvent meets the demands of the era of resource saving and energy saving, and this point is also a great advantage. .

However, despite these extremely excellent features and the fact that it is expected to become the mainstream of micro liquid chromatography in the future, the J-7 tograph is , has not yet been commercially available.This is possible because it requires reproducible micro-volume pumping of a few microliters per minute, and an accurate and reproducible gradient, in which the composition of the mobile phase is gradually changed while micro-volume pumping. This seems to be due to the fact that two problems regarding change have not yet been fully resolved.

The purpose of the present invention is to enable accurate and reproducible constant flow minute liquid delivery by driving the piston using hydraulic pressure, and to sequentially drive two or more constant flow minute liquid pumps. The object of the present invention is to provide a high performance liquid chromatograph capable of performing gradient analysis with accuracy and good reproducibility.

The present invention utilizes a large amount of liquid to deliver a small amount of liquid with high precision at the tip surface by making the area of the tip surface of the piston in contact with the mobile phase small and the area of the rear end surface in contact with the driving liquid large. By combining two or more pumps using such pistons and alternately applying pressure to the rear end surface of each piston for a predetermined time period, mixing of each mobile phase can be achieved. This allows the ratio to be controlled accurately.

The T1 embodiment will be described below.

FIG. 1 is a block diagram showing one embodiment of the present invention, and this basic structure is the same as the conventional one. That is, in the same figure, the mobile phase A12 is one digit, and the mobile phase B13 is the liquid pump A1.
4 is a liquid sending pump B, 5 is a mixer, 6 is a sample injector,
7 is a carano, and 8 is a detector.

- In a high-performance liquid chromatograph having the configuration described in
A mobile phase A1 and a mobile phase B2 are sent by a liquid feeding pump A3 and a liquid feeding pot B4, respectively, and mixed in a mixer 5. The mixing ratio in this case can be arbitrarily set by controlling the amount of liquid sent from each pump. The mobile phase leaving the mixer 5 passes through a sample injector 6 and reaches a column 7, where a detector 8 detects each sampled component.

Here, the appropriate piston diameter for each pump is about 3 to 58 mm, and if it is smaller than this, troubles such as breakage may occur, which is undesirable for practical use. In this case, in order to transfer a small amount of liquid of several microliters per minute, even if a piston with a diameter of 3+n+n is used, the moving speed will be as low as 0.1 to 1 piston per minute.

Normally, a feed screw or a cam is used to drive the piston, but neither of these methods meet the requirements of feeding the piston evenly at such low speeds.

For this reason, in this example, a piston 9 as shown in FIG. 2 was used as this micro-liquid feeding means. Note that although only the liquid feeding pump A3 for feeding the mobile phase A1 in FIG. 1 is shown in the same figure, the same applies to the liquid feeding pump B4 for feeding the mobile phase B2. In FIG. 2, the rear end surface a of the piston 9 is formed to have a larger area than the front end surface. In addition, a seal material 10 is attached to both ends to maintain airtightness of the pressurizing chamber 11, the return chamber 12, and the pump chamber 13. 14 to 17 are valves, 18.19 are liquid feeding pipes, 20 and 21 are discharge pipes, 22 are absorption valves, 23 are discharge valves, 24 are suction pipes, 25 are discharge pipes, 26 are conduit pipes, 27
28 is a pressure sensor, 28 is a driving liquid, 29 is a liquid reservoir, and 30 is a driving liquid.

In the liquid feeding pot A3 having the above configuration, liquid feeding is performed by pushing out the mobile phase Al' filled in the pump chamber 13 using the piston 9. At this time, by closing the suction valve 22 and keeping the discharge valve 23f open, the mobile phase A1 is discharged from the discharge pipe 25 at a flow rate corresponding to the moving speed of the piston 9. The piston 9 is driven by the pressure of the driving liquid 28 sent to the pressurizing chamber 11.

Here, since the rear end surface I of the piston 9 in contact with the mobile phase A1 has a smaller area than the rear end surface in contact with the driving liquid 28, it is possible to precisely control the liquid feeding of the mobile phase A1. I can do it. That is, if the area ratio between the tip end surface a and the rear end surface 1 of the piston 9 is set to, for example, 1000:1, in order to pump the mobile phase Alk from the pump chamber 13 at a flow rate of 1 μl per minute, pressurization is required. The driving liquid 28 may be sent to the chamber 11 at a flow rate of 1 mt per minute. Therefore, 1000% of the required flow rate
All you have to do is control the flow rate twice as much, and such a large flow rate is
This can be fully realized using a conventional high-speed chromatograph liquid pump. ! , in this case, within this flow rate range ±0.
A high liquid delivery system of 5% or less can be easily achieved. That is, the piston 9 can be moved with an accuracy of 0.5% or less.

During actual operation, valves 15 and 16 are closed and valve 14 is closed.
, 17 are opened, and the driving liquid 30 in the reservoir 29 is sent to the pressurizing chamber 11 using the driving pump 28. As the driving pump 28, a conventional high-performance liquid chromatograph can be used with sufficient flow rate range and accuracy as described above. Piston 9ff: After pressing, valves 14 and 17f: close, and when valves 15 and 16 are opened, 1. The driving solution 30 is sent to the return chamber 12 and the piston 9 returns. At this time, the suction valve 22. Fully open, discharge valve 23
If the pump chamber 13 is closed, the mobile phase A 1 will fill the pump chamber 13 .

Here, the pressure sensor 2 is connected to the pressure chamber 11 by bending the conduit 26.
7 is a provision. Generally, in a micro high-performance liquid chromatograph, it is desirable to reduce the dead volume of the chromato flow path by 40% to feed a small amount of liquid. Therefore, measuring at the end of the discharge pipe 25 to monitor the column pressure is
This is not desirable because the dead volume caused by the pressure cassette/the is large.

In this embodiment, by measuring the pressure in the pressurizing chamber 11, the pressure on the high pressure side can be indirectly measured. That is, the internal pressure of the pressurizing chamber 11 measured by the pressure sensor 27 is
/ I) can be monitored as the pressure peak on the high pressure side, that is, the column pressure. “This”: Uni Chroma!・One of the advantages of the present invention, which uses liquid to drive the liquid pump, is that pressure can be measured without providing a pressure sensor in the flow path system.

The capacity of the pump chamber 13 needs to be set larger than the amount of mobile phase required for one analysis, and is generally desirably 100 μl or more. Assuming that the measurement was made at a flow rate of 1 μt per minute, 1
00 μt allows continuous liquid feeding for 100 minutes. In most analyses, the analysis time per analysis is within 100 minutes, and it is considered sufficient to be able to continuously feed 100 μt.

FIG. 3 is an explanatory diagram showing a mechanism for completely forming a gradient using the two liquid feeding pumps shown in FIG. 2. In FIG. 3, two liquid pumps A3 and +14 are connected to the pipe 31.
Connect in parallel.

The liquid feed pump A3 feeds the mobile phase A1, and the liquid feed pump B4 feeds the mobile phase B2. In the figure, for the system of liquid pump A3, the same parts as in Figure 2 are given the same numbers, and for the system of liquid pump B4, the parts corresponding to the parts of liquid pump A3 are the same numbers with dashes. This is shown using . In FIG. 3, the mixing ratio of both mobile phases is controlled as follows. First, liquid feed pumps A3 and B4
are filled with mobile phases Al and B2, respectively. Next, the suction valve 22.22' is closed and the discharge valve 23.23' is left open. Meanwhile, valves 16, 16' and 15° 15' are closed;
Open valve 17.17'. Next, at the same time as the driving pump 28 starts to feed liquid, the valve 14 or 14'τ is opened. If the valve 14 is opened, the mobile phase A1 is fed by the liquid feeding pump A3, and if the valve 14' is opened, the mobile phase B2 is fed by the liquid feeding pump B4. Therefore, valve 14.14'(z
By alternately opening and closing for a predetermined period of time, the mobile phase A1
and B2 can be fed at any ratio. That is, any mixture of both mobile phases A1 and B2 can be prepared. The two are mixed in a mixer 5, led to a column 7 via a sampling injector 6, and then to an ejector 8'Ir:
It passes through and is discharged. Column 7 is tubule Calano. Such control of the opening/closing time of the valves 14, 14' can be realized very easily by using, for example, a microcomputer.

Note that the drive fluid 30 is that which is discharged from valves 17 and 17', and that which is discharged from valves 16 and 17 when piston 7999' returns.
The liquid discharged from 16' is purple, passing through the entire pipe 31 to the liquid reservoir 29.
By leading to 1 to 2 cycles, it is possible to use the 1 to 3 cycles.

As explained above, according to the present invention, it is possible to perform extremely precise liquid feeding with good reproducibility, and also to achieve an accurate and highly reproducible gradient. In other words, the functions of a high-performance liquid chromatograph can be completely miniaturized.
It is possible to replace a system using a conventional high performance liquid chromatograph with a micro high performance liquid chromatograph according to the present invention. According to the present invention, the amount of mobile phase solvent consumed is as low as about 1/1000 of the conventional method (10), and only several 1111 solvents are consumed in one day of operation, saving 1 and 20 times less time. ′
7'The 3,1, thin, 1t gear pirarikarano, which exhibits remarkable effectiveness, is 1 Fumikawa-(゛), so the number of theoretical plates is significantly higher (-2, the analytical performance is significantly improved, etc.)
This effect has gold.

lYi’i i1’-Description of the drawing

Claims (1)

[Claims] 1 A plurality of mobile phase liquid feeding pumps each having a piston having a small area at the tip end in contact with the mobile phase and a large area at the rear end surface are arranged, and each pump has a piston with a piston having a small area at the tip end in contact with the mobile phase and a large area at the rear end surface. A high-performance liquid chromatograph characterized in that it is driven intermittently by the pressure of one driving liquid that is sequentially supplied.