JPH0737973B2 - Chromatograph transfer method - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION (Industrial field of application) The present invention relates to thin layer chromatography (TLC) and paper chromatography, and to thin layer chromatography plates and paper chromatography papers (these are collectively referred to as thin layer plates). The present invention relates to a method for transferring a developed sample spot to a medium convenient for analysis by infrared spectroscopy.

(Prior Art) As a plate used in thin layer chromatography, a plate obtained by coating silica gel on a glass substrate or an aluminum sheet is generally used. In paper chromatography, filter paper is used. The sample is spread in the in-plane direction of the silica gel layer or the filter paper by the eluent to form a sample spot.

Attempts have also been made to analyze the components spread on thin plates by Fourier transform infrared spectroscopy (FTIR).
To analyze by FTIR, the developed sample spot is irradiated with the infrared light as it is, and the diffuse reflection spectrum is measured (see Anal. Chem. 1989 , 61, 615-618). , KBr and KCl powder pellets, dried and then measured by FTIR (Anal.Chem.
1987 , 59, 415-418), a method of placing a powder of an infrared non-absorbing material on the edge of a thin plate and transferring a spot component from the thin plate to the powder through a glass fiber (An.
al. Chem. 1986 , 58, 2708-2714)).

(Problems to be Solved by the Invention) For FTIR measurement, a method in which infrared rays are directly applied to a thin layer plate is not practical because the stationary phase silica gel or paper of the thin layer plate has large infrared absorption.

The method of scraping and extracting the sample spot and dropping it on the KBr or KCl powder pellets for FTIR measurement is inaccurate and time-consuming because the sample is lost, decomposed, or contaminated. It is troublesome.

In the method of transferring the fractional component by using the glass fiber at the edge of the thin plate, the resolution in the developing direction is determined by the pitch of the glass fiber provided at the edge, and only the thin plate that is one-dimensionally developed can be used. Not applicable.

When performing infrared spectroscopic analysis of a sample spot, the inventors did not scrape the sample spot, but transferred the sample spot developed on the thin layer plate to the infrared non-absorptive substance layer, thereby ensuring accurate Has already proposed a method for enabling a simple infrared spectroscopic analysis (Japanese Patent Application No. 1-1997).
No. 06). In the proposed method, a thin plate having a sample spot and no support material is placed on a porous material plate,
The infrared non-absorbing material layer is brought into close contact with the thin layer plate, and the eluent is raised by capillary action from the porous material plate to the infrared non-absorbing material layer through the thin layer plate, and the infrared non-absorbing material layer Volatilize from the surface to transfer the sample in the thin layer plate to the infrared non-absorbing material layer.

If the sample spread on the thin plate absorbs visible light, it can be visually confirmed that the thin plate is transferred to the infrared non-absorbing material layer. However, some samples do not have absorption in the visible region, and it may not be possible to visually confirm that they have been transferred. At that time, since the transfer time is decided and the transfer operation is carried out by relying on the intuition, the transfer degree may not be constant.

Therefore, the present invention is a transfer method of the above-mentioned patent application by the present inventors for transferring a sample spot developed on a thin layer plate to an infrared non-absorbing material layer, in which the sample does not have absorption in the visible region. It is an object of the present invention to provide a method capable of performing transfer with good reproducibility even in the case of.

(Means for Solving the Problem) In the present invention, the first embodiment having a sample spot and using no support material
And a second thin layer plate carrying a dye marker are placed side by side on a porous material plate, and an infrared non-absorbing material layer is provided on the first thin layer plate and the second thin layer plate. And the eluent is raised from the porous material plate through each thin layer plate toward the infrared non-absorptive material layer by a capillary phenomenon, and volatilized from the surface of the infrared non-absorptive material layer to form a thin layer plate. The sample is transferred to the infrared non-absorbing material layer.

Further, in the present invention, in order to automatically detect that the transfer has been performed, light irradiation is performed on the infrared non-absorbing material layer at the position of the dye marker, and the reflected light from the light irradiation position is received. , The transfer state is monitored from the received light signal.

As the first thin layer plate and the second thin layer plate,
EmporeTLC sheet (product of Analytichem International, Inc.), glass sintered plate, ceramic plate,
Paper for paper chromatography can be used.

As the porous substance plate, for example, a glass filter material, a ceramic substrate, a polymer material, a material having a solution permeability such as paper or cloth can be used.

As the infrared non-absorbing substance, powdery substances such as KBr and KCl can be used.

Although various solvents can be used as the eluent, those having a low elution force and a low boiling point are preferable. As such a solvent, a strongly polar solvent such as methanol can be used when the thin layer plate is a normal phase system, and a weak polarity solvent such as acetonitrile when the thin layer plate is a reverse phase system. Etc. can be used.

(Operation) A first thin layer plate on which a sample is developed and separated and a second thin layer plate carrying a dye marker are arranged side by side, and an infrared non-absorbing material layer is provided on one surface of both thin layer plates. Close contact,
When the eluent is moved from the other surface of both thin layer plates to the side where the infrared non-absorbing material layer is present by capillary action, the sample components move to the infrared non-absorbing material layer and are transferred in the first thin layer plate. At the same time, the dye of the marker is transferred to and transferred to the infrared non-absorbing material layer in the second thin plate. Even if the sample component does not absorb in the visible range and the transfer cannot be visually recognized only by the sample part, the dye is transferred to the infrared non-absorbing material layer at the same time, so the transfer is visually confirmed. be able to.

If the infrared non-absorbing material layer at the position of the dye marker is irradiated with light and the reflected light is detected by the light receiving section, the transfer can be automatically monitored by the light receiving signal.

(Embodiment) FIG. 1 is an exploded perspective view showing one embodiment, and FIG. 2 is a sectional view taken along line XX of FIG. 1 together with a porous material plate.

Reference numeral 4 is an eluent contained in a container, and methanol is used. The eluent 4 is filled with a porous material plate 6 so that a part thereof is immersed. A ceramic substrate is used as the porous material plate 4.

Reference numeral 8 denotes a transfer frame. A slit-shaped hole 9 is formed in the transfer frame 8, and the infrared non-absorbing material layer 10 is densely formed in the hole 9.
And it is filled uniformly. A KBr layer is used as the infrared non-absorption material layer 10. The hole 9 of the transfer frame 8 has a width a little smaller than the size of the sample spot 18a developed on the first thin layer plate 12a, and the plurality of sample spots 18a and the second sample spot 18a
The thin plate 12b is formed to have a length including the dye spot 18b. Since the size of the sample spot 18a is about 5 to 10 mm, it is appropriate that the width of the hole 9 of the transfer frame 8 be about several mm. Reference numeral 14 denotes a support frame, and the support frame 14 has a hole slightly larger than the hole 9 of the transfer frame 8. Transfer frame 8 and support frame
A first thin layer plate 12a and a second thin layer plate 12b are sandwiched between the two between 14, and the transfer frame 8 and the support frame 14 are fixed to each other by screws 16 and 16 at both ends, thereby forming a thin layer. The plates 12a and 12b are sandwiched and fixed between the transfer frame 8 and the support frame 14, and the infrared non-absorptive material layer 10 and the thin layer plates 12a and 12b filled in the holes 9 of the transfer frame 8 are formed. In close contact.

The thin-layer plates 12a and 12b are normal-phase thin-layer plates for thin-layer chromatography, and are thin plates that do not use a support material, and the EmporeTLC sheet (Analytichem Internation
al., Inc. product) is used. The sample is already developed on the thin layer plate 12a and the sample spot 18a is present.
On the thin layer plate 12b, a dye is carried as a transfer marker to form a dye spot 18b.

In order to bring the porous material plate 6 and the thin layer plates 12a, 12b into contact with the thin layer plates 12a, 12b sandwiched between the transfer frame 8 and the support frame 14, the surface of the porous material plate 6 is supported. A ridge 6a is formed which enters the hole of the frame 14 and comes into contact with the thin plate 12a, 12b.

In order to promote the elution plate 4 to volatilize from the surface of the infrared non-absorption material layer 10, a fan heater may be provided above the infrared non-absorption material layer 10 to blow hot air.

Next, the operation of this embodiment will be described.

A mixture of anhydrous caffeine, phenacetin, and noscapine, which are components of a cold medicine, is used as a sample, and this sample is spread on a thin layer plate 12a by thin layer chromatography. These sample components do not have absorption in the visible region, but have absorption in the ultraviolet region. Therefore, visually check the sample spot
18a cannot be recognized, and the transferred state cannot be confirmed even when transferred. On the other hand, Fast Green FCF is used as the dye of the dye marker. Spots 18b of this dye are formed on the thin layer plate 12b.

As shown in FIG. 1, the transfer layer 8 filled with the infrared non-absorbing material 10 and the support frame 14 with holes are shown in the thin layer plate 12a on which the sample is spread and the explosive layer plate 12b having the dye spots 18b.
Insert it with and fix it. This is placed in a state where the thin plate 12a, 12b is in contact with the surface of the porous material plate 6 as shown in FIG. In this state, as shown by the arrow, the eluent 4 passes through the porous material plate 6 and the thin layer plates 12a, 12b
It rises up to reach the infrared non-absorbing material layer 10 and is volatilized from its surface. At this time, the sample components in the thin layer plate 12a move upward as the eluent 4 moves and are transferred to the infrared non-absorbing material layer 10, and the dye on the thin layer plate 12b also moves as the eluent 4 moves. And transferred to the infrared non-absorbing material layer 10. Since the eluent 4 which is a transfer solvent has a strong elution output, the sample spot 18a and the marker dye spot 18b are transferred at the same time.

Since the infrared non-absorptive substance layer 10 is filled in the slit-shaped holes 9 of the transfer frame 8, it is possible to simultaneously transfer a plurality of sample spots 18a arranged in the developing direction to the infrared non-absorptive substance layer 10. A chromatogram can be obtained by performing FTIR measurement by the diffuse reflection method using the infrared non-absorbing material layer 10 after the transfer as a sample. At this time, since the width of the slit-shaped hole 9 corresponds to the size of the sample spot 18a, the sample is prevented from spreading in the slit width direction during transfer.

FIG. 3 shows another embodiment.

The transfer frame 8 is provided with a slit-shaped hole 9a in a region for transferring the sample spot 18a and a hole 9b in a region for transferring the dye spot 18b of the marker separately. The KBr layer, which is the absorption material layers 10a and 10b, is filled. In this way, by making the transfer holes of the sample spot 18a portion and the dye spot 18b portion separate, it is possible to more easily recognize the transfer.

FIG. 4 shows a method for automatically detecting transcription.

A light source 20 for irradiating the infrared non-absorbing material layer 10b at the position where the dye of the dye spot 18b is transferred is provided, and for receiving the reflected light from the infrared non-absorbing material layer 10b by the light from the light source. Is provided with a photodetector 22. Reference numeral 24 denotes a data processing unit that determines from the detection signal of the photodetector 22 that the dye has appeared on the surface of the infrared non-absorbing material layer 10b, and a signal of the data processing unit 24 notifies the end of transfer. A lamp 26 and a buzzer 28 are provided.

According to the present embodiment shown in FIG. 4, when the thin layer plates 12a and 12b are sandwiched between the transfer frame 8 and the support frame 14 and placed on the porous material plate 6, and the elution plate 4 is raised to perform the transfer, There is a difference between the detected intensity of the reflected light from the infrared non-absorbing substance layer 10b before being transferred and the detected intensity of the reflected light when the dye appears on the infrared non-absorbing substance layer 10b after being transferred. Occurs, the data processing unit
This difference is detected by 24.

Thin layer plate 12b with constant concentration and amount of marker dye
By carrying it on, it is possible to transfer with good reproducibility.

(Effect of the Invention) In the present invention, a thin layer plate having a sample spot to be transferred and a thin layer plate carrying a dye as a marker are arranged side by side, and the dye is transferred simultaneously to the sample to the infrared non-absorbing material layer. By doing so, even if the sample does not have absorption in the visible region, it can be easily confirmed that the sample has been transferred.

If a monitoring device that optically detects the dye is provided on the infrared non-absorbing material layer at the position of the dye marker, the transfer can be automatically detected, and the reproducibility of the transfer operation can be improved.

[Brief description of drawings]

FIG. 1 is an exploded perspective view showing an embodiment, FIG. 2 is a sectional view showing a sectional view taken along the line XX of FIG. 1 together with a porous material plate, and FIG. 3 is another embodiment. FIG. 4 is an exploded perspective view, and FIG. 4 is a sectional view showing the sectional view taken along the line YY of FIG. 3 together with the porous material plate and the optical monitoring device. 4 ... Elution plate, 6 ... Porous material plate, 8 ... Transfer frame, 9,
9a, 9b …… Transfer frame hole, 10,10a, 10b …… Infrared non-absorbing material layer, 12a …… First thin layer plate, 12b …… Second thin layer plate, 18a …… Sample spot, 18b …… marker dye spot, 20 …… light source, 22 …… photodetector, 24 …… data processing unit.

Claims (2)

[Claims] 1. A first sample spot having no support material.
And a second thin layer plate carrying a dye marker are placed side by side on a porous material plate, and an infrared non-absorbing material layer is provided on the first thin layer plate and the second thin layer plate. And the eluent is increased from the porous material plate through the respective thin layer plates toward the infrared non-absorptive material layer by capillary development and volatilized from the surface of the infrared non-absorptive material layer to form a thin layer plate. A transfer method in which a sample is transferred to an infrared non-absorbing material layer. 2. The method according to claim 1, wherein the infrared non-absorbing material layer at the position of the dye marker is irradiated with light, the reflected light from the light irradiation position is received, and the transfer state is monitored from the received light signal. Chromatograph transfer method.