JPS6122895B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention involves selectively measuring spots obtained by simultaneously developing several standard samples of known concentration on the same thin layer chromatography plate, and averaging these measured values to determine the standard sample. The present invention relates to a concentration measurement method that calibrates a known amount of and thereby performs accurate concentration measurement.

Quantitative measurement of thin layer chromatograms is usually performed by simultaneously developing a sample of unknown concentration and a standard sample of known concentration on the same thin layer chromatogram plate and comparing both spots. In other words, if the concentration of the standard sample is C, the measured value of the spot is S, and the measured value of the unknown sample is B, the concentration W of the unknown sample can be found as W=F×B......() .

here, F=C/S……() , and this F is called a concentration conversion factor.

Since the measured value S of the spot of the standard sample includes an error due to non-uniform development depending on the location of the thin layer chromatogram, the concentration conversion factor F also includes the same error. Therefore, if the density is calculated using this value, the calculated density will also include an error.

An object of the present invention is to provide a measurement method that can eliminate such error components and accurately measure concentration.

The gist of the present invention is to spread a standard sample with a known concentration at a plurality of appropriate locations between unknown samples spread at a plurality of locations on a thin layer chromatography plate, and to spread the standard sample at a plurality of appropriate locations where the standard sample is spread on the plate. The known amount of the standard sample is calibrated by the average value of the standard sample at a plurality of locations, and the concentration is thereby measured.

That is, according to the present invention, the concentration conversion factor F is Therefore, the individual errors of the spots of the standard sample are canceled out and removed.
(Hereinafter, F in formula () will be expressed as an average concentration conversion factor to distinguish it from F in formula ().) In carrying out the present invention, for example, the stage on which the thin layer chromatography plate is mounted is - A computer controls the densitometer that moves in the Y direction, and after measuring a spot on one standard sample, the stage is automatically moved to measure the next spot on the standard sample, and this is repeated to measure multiple locations. The values may be averaged and a known amount of standard sample calibrated to make the concentration measurement.

The present invention will be explained in more detail below with reference to FIGS. 1 to 3. However, this is just an example, and the present invention is not limited thereby.

As shown in FIG. 1, an apparatus for carrying out the present invention includes a stage 2 on which a thin layer chromatography plate 1 is placed, a stage driving means 3, a minute light beam irradiation means 4,
It is composed of a characteristic light detection section 5, an arithmetic processing section 6, an A/D converter 7, and an operation console 8. Control console 8
The Y-direction scanning width D, lane interval L, number of lanes N, and known density U can be inputted to the arithmetic processing section 6 from there. (A lane refers to the path along which a spotted sample is developed.) The arithmetic processing unit 6 is a microcomputer equipped with an interface, a memory, and a microprocessor, and drives the stage in the opposite direction by a width D. A procedure for measuring a characteristic optical signal and obtaining a measured value S(j). A procedure for driving the stage to the left in the X direction by a distance L and in the development direction by a width D. Execute the above step (i), and then Execute step (ii) and repeat these steps alternately until the number of executions of step (i) reaches the number of lanes N, and the measured values S(1), S(2), S(3) obtained thereby , ..., S(N), divide this by N to obtain the average value P, and divide the concentration U by P to obtain the average concentration conversion factor F (see Fig. 3). There is.

Next, this operation will be explained.

A standard sample 15 with a known concentration is spotted and spread at several points on the thin layer chromatographic plate 1 along with an unknown sample 17. However, the lane spacing for the standard sample shall be equal to L.

After development, move thin layer chromatography plate 1 to stage 2.
Place it on. By manually moving the stage 2 or by moving the thin layer chromatography plate, the initial position of the minute light beam is set a little before the leftmost spot 16 of the standard sample.

A value slightly larger than the width of the standard sample spot in the development direction is input as the Y-direction scanning width D from the operation console. In addition, the lane interval L of the standard sample and the number N of lanes of the standard sample are input.

Finally, input the concentration U of the standard sample and start the measurement.

When the step (iii) is executed, the spots 16 of the standard sample are continuously measured as shown by the dotted line in FIG.
The average concentration conversion factor F is automatically calculated.

In this embodiment, the average concentration conversion factor obtained in this way is displayed on a display provided on the console and stored in the arithmetic processing section, and is used to calculate the concentration value when measuring an unknown sample.

[Brief explanation of the drawing]

FIG. 1 is a block diagram showing an example of a densitometer used for carrying out the present invention, FIG. 2 is a reference diagram for explaining its operation, and FIG. 3 is a procedure diagram for calculating an average concentration conversion factor. DESCRIPTION OF SYMBOLS 1... Thin layer chromatography plate, 2... Stage, 3... Stage drive unit, 4... Minute light flux irradiation unit, 5... Characteristic light detection unit, 6... Arithmetic processing unit, 7
...A/D converter, 8...Operation console, 16...
Standard sample spot.

Claims (1)

[Claims] 1. Drive the thin-layer chromatography plate in the X-Y direction, sequentially scan unknown samples spread out at several locations on the same plate with monochromatic light, and detect the transmitted light, reflected light, or fluorescent light. In the method of measuring the concentration, a standard sample with a known concentration is spread on a plurality of appropriate locations between the unknown samples spread on the plurality of locations on a thin layer chromatography plate, and the plate is spread on an appropriate number of locations on which the standard sample is spread. The device is selectively driven to scan locations, calibrates the known amount of the standard sample using the average value of the standard sample at multiple locations, and measures the concentration of the unknown sample based on the calibrated known amount. A concentration measurement method characterized by: