JPS5926058A - Dispensing method and dispensing device - Google Patents

Abstract

PURPOSE:To dispense simply a fraction, by using a vessel providing a means for discharging a liquid when the liquid surface arrives at a prescribed position and varying the capacity of its vessel in accordance with an outflow pattern of the fraction from a column. CONSTITUTION:A fraction sent from a column is received and is held in a vessel 1a and when a liquid surface A arrives at a prescribed position B, the liquid is discharged from a crooked glass tube 4. At this time, the capacity of the vessel 1a is varied by controlling the insertion depth of an insertion rod 2a in accordance with an outflow pattern of the fraction sent from the column. Then, for example, the liquid is discharged from the vessel 1a concurrently with completion of the outflow from the column and this operation is repeated in correspondence to the outflow pattern of the fraction sent from the column and then, the necessary fraction and unnecessary fraction are demarcated and are dispensed.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to the field of liquid chromatography. The present invention provides a method for developing and fractionating a sample containing a plurality of 7-lactones by column chromatography, and a fractionating device used to implement this method.

Hitherto, such methods have been known in which the effluent from the column is introduced into a container having means for draining the liquid when the liquid level reaches a predetermined position. However, in this method, the amount of liquid drained from the container is always constant, and for example, if the amount of liquid effluent from the column is constant, the liquid is drained at regular intervals. Therefore, if the amount of liquid drained from the container is too large, the interval between drains will be long, and there is a risk that two or more 7 lactations may be mixed. Furthermore, if the amount of liquid per draining cycle is too small, the interval between drains becomes short, the same fraction may be discharged multiple times, and efficiency tends to decrease. Furthermore, when a plurality of fractions are separated, it is extremely difficult to introduce the effluent from the column into the container in synchronization with the outflow timing of the 7-lactone.

The present invention is the result of intensive research and has resulted in a new technique that solves these problems.

The preparative separation method of the present invention is a method in which a sample containing a plurality of fractions is developed and fractionated by column chromatography, and the method includes introducing the fractions from the column into a container and holding them; This fractionation method comprises discharging the fraction from the container when it reaches a predetermined amount in the container, and changing the volume of the container in accordance with the outflow pattern of the fraction from the column.

The preparative device of the present invention is a device for developing and fractionating a sample containing a plurality of 7-lactones by column chromatography, and the device includes a container for holding a fraction from the column; A preparative separation device comprising means for discharging the fraction from the container when a predetermined amount is reached, and means for changing the volume of the container in response to the outflow pattern of the fraction from the column.

In the preparative separation method according to the present invention, the fraction from the column/
is introduced, for example, into a container having means for draining when the liquid level reaches a predetermined position, the volume of said container being preferably predetermined, corresponding to the outflow pattern of the fraction from the column. Change according to the program. For example, a fraction is introduced into a container, and when the amount of this fraction is large, the capacity of the container is increased, and when it is small, the capacity is decreased. Then, the volume of the container is changed so that, for example, the container is drained at the same time that the fraction finishes flowing out of the column. The above operations are repeated to fractionate and separate fractions.

, 2, etc., are preferably set by the retention time and outflow pattern of each fraction previously investigated for each sample.

FIG. 1 is a front sectional view of an example of the preparative separation device of the present invention. FIG. 2 is a front sectional view of another example of the preparative separation device of the present invention. FIG. 3 is a front sectional view of an example of a preparative separation apparatus using the preparative separation device of the present invention, and a part thereof is shown in a block diagram. Figure y/l) is an example of a chromatogram obtained by introducing serum into a GPC (gel permeation chromatography) column filled with polyvinyl alcohol gel and developing it. FIG. 9(b) is a graph showing the liquid I in the container corresponding to FIG. 9(,). FIG. 3 is a perspective view of an example of the insertion rod position control device shown in FIGS. 1, 2, and 3. FIG.

In Figure 1, vessel /& receives and holds the fraction from the column. When the liquid level A reaches a predetermined position B, the liquid is drained through the bent glass tube. At this time, the capacity of the container/a is changed by inserting the insertion rod 2a in accordance with the outflow pattern of the 7-lux from the column. For example, the total volume of the container is changed by inserting an insertion rod into the container such that the container is drained just before the required fraction exits the column. When a predetermined amount is reached, the liquid is drained from the glass tube IIa by siphon action. Then, the required fraction is introduced into a container, and for example, when the amount of this 7-lux is large, the capacity is increased, and when it is small, the capacity is decreased. Then, for example, the container is drained at the same time that the required 7 lactations have finished flowing out of the column. This operation is repeated as the fractions flow out/return from the column, and necessary fractions and unnecessary fractions are fractionated.

In FIG. 2, the glass tube a in FIG. 1 is replaced with a valve S and a water level detector t. The fraction from the column is held in container /b, and when the liquid level reaches water level detector B, valve S is opened and the liquid is drained.

At this time, the capacity of container /b should be adjusted according to the outflow pattern of the fraction from the column. l! , 2b are changed in the insertion order J'+.

In FIG. 3, the eluent in the eluent tank 7 is sent to the column /θ together with the sample injected from the injector 7 by the pump g. The sample is developed using Colors 1 and 10. Corresponding to the outflow pattern of the 7-lux from the column, in the vessel / the insertion rod λ is connected to the insertion rod position control device @-,
The container is changed according to a predetermined program. This insertion rod position control device uses, for example, a motor to move the insertion rod up and down using a cam. Then, the necessary 7 fraction and unnecessary fraction are fractionated and drained, and collected in the chamber //. The chatter moves each time the container/chamber is drained, and each drain is collected in a separate chamber. In this way, the necessary fractions are collected.

FIG. 7 (fl) shows an example of a chromatogram obtained by introducing serum into a GPC column packed with polyvinyl alcohol gel and developing it. The conditions will be described below. 20μ of healthy human serum as a sample! The transport rate of the eluent by the pump was set to /m, and the column temperature was set to 30'.

At this time, transferrin leaked out from 35 to 37 minutes after serum injection, and albumin leaked out from 1I/ to 57 minutes. These fractions were collected using a preparative device such as the one shown in Figure 1.
d was used for fractionation.

Drain 1 oyd three times and 1. After draining S-d seven times, the solution containing transferrin was collected for 2 days.

After further draining the water, the solution containing albumin was collected in sM1 fraction. Transferrin and albumin were fractionated using the apparatus shown in FIG. 3 using the sample, fractionation conditions, and fractionation device as described above. Figure 7(b) shows
This is a graph of the amount of liquid in the container corresponding to the chromatogram in FIG. The reason why the rate of increase in the liquid volume in FIG. 9(b) is always constant is that the 7-rakusion containing the eluent flowing out from the column is constant. Both transferrin and albumin were fractionated by draining each container once. In the example of the insertion rod position control device shown in FIG. 5, a cam/S is rotated by a motor 13 via a gear puffer, and an insertion rod 2c pressed against the cam/S by a spring 2 is moved. The pattern of movement of the insertion rod, 2c, by the cam/S preferably corresponds to the outflow pattern of the fraction from the column.

As described above, the present invention not only allows fractions to be separated very easily, but also has the effect of avoiding mixing of multiple fractions and fractionating the same 7-fraction into multiple fractions. Furthermore, when the amount of effluent from the column is constant, there is an effect that the effluent can be fractionated in accordance with the outflow timing of a plurality of fractions. In addition, there is an effect that the fraction can be classified accurately with a small number of draining times, regardless of the size of the fraction amount. Furthermore, since the number of times the liquid is drained is small, the apparatus can be simplified, and there is an effect that the number of erroneous operations is extremely reduced.

[Brief explanation of the drawing]

FIG. 1 is a front sectional view of an example of the preparative separation device of the present invention. FIG. 2 is a front sectional view of another example of the preparative separation device of the present invention. FIG. 3 is a front sectional view of an example of a preparative separation apparatus using the preparative separation device of the present invention, and a part thereof is shown as a block diaphragm. Figure (a) is an example of a chromatogram obtained by introducing serum into a GPC column packed with a polyvinylalfur gel and developing it. Figure 7 (b
) is a graph showing the amount of liquid in the container corresponding to Figure (a). FIG. S is a perspective view of an example of the insertion rod position control device of FIGS. 1, 2, and 3. FIG. /a, /b, /- container, 2b-, 2c
%Y...Insertion rod 3aJbJ...Insertion position control device @
4'a, Ri... Glass tube S... Valve 6... Water level detector 7-...
...Eluent tank S...---Pump?・・・
...... Injector 10...
・・・Column／／・・・・・・・・・−・Chamber
/, 2...Spring patent applicant Asahi Kasei Industries, Ltd.

Claims (4)

[Claims] (1) A method for developing and fractionating a sample containing multiple fractions by column chromatography, which method includes introducing and holding the fractions from the column into a container, and storing the fractions in the container. A preparative separation method comprising: draining and closing the container when a predetermined amount of the fraction has been reached; and varying the volume of the container in response to the outflow pattern of the fraction from the column. (2) The preparative separation method according to claim 1, wherein the change in the capacity of the container is performed according to a predetermined program. (3) A device for developing and fractionating a sample containing multiple fractions by column chromatography, the device comprising a container for holding the 7 fractions from the column, and a container for holding the 7 fractions from the column; A preparative separation device comprising means for discharging the container when a volume is reached, and means for varying the volume of the container in response to an outflow pattern of 7 radians from a force of 2 mm. (4) The preparative separation device according to claim 3, wherein the volume change of the container is performed according to a predetermined program.