JPS60259945A - Leading liquid for tubular type uniform speed electrophoretic analysis - Google Patents

Abstract

PURPOSE:To enable the separation and analyzation of components at a high accuracy by adding very small amount of hydroxy propyl methylcellulose to an electrolytic solution for a tubular type uniform speed electrophoretic analysis leading liquid as thickener to inhibit an electric osmotic current even in an alkaline area. CONSTITUTION:A leading liquid containing the ion of the same character with higher mobility than an ion to be analyzed is arranged to ajoin a terminal liquid containing the ion of the same character with lower mobility than the ion being analyzed on the interface in a capillary tube and a sample is injected into the interface while a constant current flows from both ends of the tube to separate and analyze the ion being analyzed in the ascending order of mobility. In this invention, 0.01-0.2% of hydroxy propyl methylcellulose is added to the leading liquid to thicken. This can inhibit the electric osmotic current even in the not only acid to neutral but also alkaline area to prevent components separated from running over a slip to the capillary tube making the boundary between the components unclear thereby enabling separation and analysis at a high accuracy.

Description

DETAILED DESCRIPTION OF THE INVENTION A. Field of Application of the Technology The present invention relates to a leading liquid suitable for a capillary isotachophoresis device.

Conventional technology Capillary isotachophoresis analysis is a type of separation analysis in which leading ions contain ions of the same sign that have a higher mobility than the analyte ions, and ions of the same sign that have a lower mobility than the analyte ions. The terminal liquid contained in the capillary tube is brought into contact with the interface at the interface, the sample is injected into this interface, and a constant current is passed from both ends of the tube to separate the ions to be analyzed in the order of their mobility. .

By the way, sample components separated in this way have the problem that diffusion occurs between adjacent components based on the difference in ion concentration, resulting in a decrease in analysis accuracy. Monoisootatylphenol (C2H40) 9-chain and hydroxymethylcellulose (HEC) are added to the leading liquid as thickeners. These thickeners function in the acidic to neutral range, allowing separated ions to be transported to the detection means with clear boundaries, thereby improving analysis accuracy.

However, in the alkaline region, the electroosmotic flow becomes large, causing slippage between the separated components and the capillary tube, making the boundaries between the components unclear and reducing the accuracy of analysis, which is still a problem.

C. Objectives In view of the above-mentioned problems, an object of the present invention is to provide a novel leading liquid capable of suppressing electroosmotic flow in all hydrogen ion concentration regions.

The structure of the twenty-first invention, that is, the feature of the present invention is that a trace amount of hydroxypropyl methylcellulose is added to the electrolyte.

E. Examples - The details of the present invention will be explained below based on Examples.

[Example 1] (a) Leading liquid mixed with 0.0 IM hydrochloric acid and amediol, pH 8.8
A solution containing 0.1% of the conventionally used hydroxy methylcellulose (HEC).

(b) Terminal liquid with pH 10.8, which is a mixture of 0.01M 6-aminocaproic acid and barium hydroxide.

(C) Capillary tube with inner diameter of 1 mm and length of 40 mm and inner diameter of 0.5 m
m, 150 mm long tubes arranged in series.

(d) Detector Potential gradient detector and ultraviolet absorption photometer with a wavelength of 254 nm A low-molecular sample consisting of a mixture of aspartic acid, cystine, asparagine, tyrosine, glycine, tryptophan, alanine, and β-alanine each at a concentration of 400 PP. When the cells were injected at 1 W and analyzed using a migration current of 100 pA, the results shown in FIG. 1(A) were obtained.

Next, PH containing O, OIM hydrochloric acid and amediol was mixed.
Hydroxy propylmethyl cellulose (HPMC) was added as a thickener to the solution of 8, 8 and used as the leading liquid, and analyzed under the above conditions, as shown in the same figure (B). We were able to obtain analysis results in which the fraction boundaries of each component were much clearer than with conventional leading liquids.

[Example 2] (a) Leading liquid 0.2% monoisooctylphenol was mixed into a solution of PHLl-, which is a mixture of 0.0045M MES and 0.01M 7mediol.

(b) Terminal liquid 0° OIM of 6-aminocaproic acid, barium hydroxide,
and 0.01M 7mediol mixed PH10.8
Of things.

(c) A capillary tube with an inner diameter of 1 mm and a length of 100 mm, and an inner diameter of 0.5 mm.
A series of tubes with a length of 250 mm.

(d) Ultraviolet absorption photometer with detector wavelength of 280 nm 1.8p of a polymer sample made by mixing human control serum and ampholine at a ratio of 1:3 was injected, and the electrophoresis current was 75. When analyzed by A, results as shown in FIG. 2(A) were obtained.

Next, 0.1% hydroxy propyl methylcellulose (HPMC) was added as a thickener to a pH 9.1 solution containing 0.0045M MES and 0.01M 7mediol.
When analysis was conducted under the conditions described above using the sample mixed with
Moreover, what was detected as a single peak in the conventional method could be obtained as a waveform separated into multiple peaks, and it was found that the boundaries of the fractions were sharper than in the conventional method.

Next, hydroxy propyl φ methyl cellulose, (H
PMC) concentration: 10.05%, 0.1
%.

When the protein in the serum was analyzed by changing the concentration to 0.2% and 0.4%, the concentration of hydroxypropyl methylcellulose was 0.05% and 0.1%, as shown in Figure 3.
.

At 0.2z, a sharp signal could be obtained from the ultraviolet absorption photometer, but at 0.4z, the peaks from the ultraviolet absorption photometer became small and unified, and the separation performance deteriorated. From this, it was found that when hydroxypropyl methylcellulose was added in the range of 0.01 to 0.2z, a clear fraction could be obtained.

Furthermore, when we added methylcellulose in hydroxypropyl in the range of 0.01 to 0.2z to the acidic leading solution and analyzed it, we found that it showed clear sample fractionation, similar to the case with the alkaline leading solution. confirmed.

Note that this hydroxy propyl methylcellulose has the effect of suppressing electroosmotic flow and clarifying the fraction when used not only in a capillary type isotachophoresis apparatus but also in a free-flow type electrophoresis apparatus.

As explained above, according to the present invention, hydroxy propyl methylcellulose is used as a thickener for the leading liquid, so that it can be applied to a wide range of materials ranging from low molecular weight to high molecular weight, regardless of the hydrogen ion moisture content of the leading liquid. Samples can be separated and analyzed with clear fractions.

[Brief explanation of the drawing]

Figures 1 (A) and (B') are waveform diagrams showing the analysis results of low-molecular samples using the conventional leading liquid and the leading liquid of the present invention, respectively. FIG. 3 is a waveform diagram showing the analysis results of a polymer sample using the leading liquid and the leading liquid of the present invention. FIG. 3 is a waveform diagram showing the analysis results when the concentration of hydroxy propyl methylcellulose is changed. Applicant: Shimadzu Corporation Representative: Patent attorney: Yoshido Nishikawa Katsuhiko Kimura $72 (A) (B) Figure 2 (8)

Claims (1)

[Claims] A leading solution for capillary isotachophoresis analysis made by adding a minute amount of hydroxy propyl methylcellulose to an electrolytic solution containing ions with the same sign and higher mobility than the ions to be analyzed.