JP3063321B2 - Electrochemical detector for sugars - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an electrochemical detector for detecting saccharides with high sensitivity by utilizing an electrode reaction, that is, a charge transfer occurring at an interface between an electrode and a solution, for example, flow injection analysis, capillary electrophoresis. Alternatively, the present invention relates to an electrochemical detector that can be used in an analysis system such as high performance liquid chromatography.

[0002]

2. Description of the Related Art Conventionally, a differential refractometer or a detector utilizing ultraviolet absorption at a short wavelength of 200 nm or less has been generally used for detecting saccharides, but these methods have low sensitivity and high sensitivity analysis. Not suitable for use.

For example, as an approach to high-sensitivity analysis of saccharides in the pharmaceutical and medical fields, a fluorescence detection method utilizing a post-column reaction has been employed, focusing on the fact that sugar molecules have a large number of hydroxyl groups. .

In recent years, a method using a pulse amperometric detector (PAD), that is, a pulse amperometry, has attracted attention as a method for detecting sugars with high sensitivity by an electrochemical technique. In this method, gold (A
By applying a pulsed potential to the u) or platinum (Pt) electrode, the amount of charge transfer based on the electrode reaction is detected while washing / regenerating the electrode surface.

[0005]

Among the conventional methods for detecting saccharides as described above, the method using a refractometer and ultraviolet absorption has the disadvantage of low sensitivity as described above. In the fluorescence detection method using the column reaction, the sugar itself does not have a fluorophore, so it is necessary to label the sugar. In many cases, the reaction temperature in the post-column reaction must be increased, and the reaction coil (column) And the heated solution must be cooled before entering the detector, which complicates the apparatus configuration.

[0006] On the other hand, pulse amperometry has a feature that detection with higher sensitivity is possible as compared with other detection methods. However, since the sugar oxidation reaction intermediate is easily adsorbed on the electrode surface, the pulse amperometry is used. On the other hand, it is necessary to apply a triple pulse having a total of three potentials of a washing potential and a regeneration potential in addition to a detection potential in a short period of time, which makes the operation difficult and reproducible.

An object of the present invention is to provide a detector which has a simpler apparatus or system configuration than conventional detection methods, is easy to operate, and can detect sugars with high sensitivity and high reproducibility. To provide.

[0008]

Means for Solving the Problems In order to achieve the above object, the present invention provides a method for electrochemically detecting a saccharide by utilizing an electrode reaction, wherein rhodium is used as a working electrode of the electrochemical detector. (Rh) electrode.

[0009]

Effect It was confirmed that rhodium functions as a catalyst in an alkaline solution, and that oxidization of saccharides easily occurs on the electrode by using this rhodium as a working electrode.

Therefore, by using the detector of the present invention in an alkaline solution atmosphere and measuring the oxidation current of the saccharide, the saccharide can be detected with high sensitivity. That is, in the present invention, since the electron transfer reaction is monitored, the sensitivity can be easily increased as compared with other methods.

Further, in the rhodium electrode, there is no adsorption of sugar on the electrode surface, and therefore, it is only necessary to apply a constant voltage of, for example, about 500 mV without pulsing the voltage applied to the electrode.

[0012]

FIG. 1 is a longitudinal sectional view showing a configuration of a cell using an electrochemical detector (ECD) to which the present invention is applied.
The structure is the same as that of a normal ECD cell. The working electrode 1 is embedded in an insulator block 2 made of an electrically inactive fluorine resin or the like.
For example, an auxiliary electrode block 3 made of stainless steel is provided. The auxiliary electrode block 3 is provided with a solution inlet tube 4 and an outlet tube 5, and the outlet tube 5 is guided to a reference electrode component 7 holding a reference electrode 6 made of, for example, silver-silver chloride. .

A feature of the embodiment of the present invention is that the material of the working electrode 1 is rhodium in the ECD cell having the above structure. That is, in this example, a rhodium wire having a diameter of 1 mm is used as a working electrode 1 and pressed into an insulating block 2 made of a fluorine-based resin (for example, trade name Kel-F).

An example in which a saccharide is actually detected using the above-described embodiment of the present invention will be described below. In this example,
As shown in FIG.
0, sample injector 11, LC column 12, and EC
Standard high performance liquid chromatography (HPLC) consisting of D cell 13 was used. ECD cell 1 of this HPLC
As No. 3, the ECD cell of the embodiment of the present invention described above is used.

The analysis conditions are as shown in Table 1 below.

[0016]

[Table 1]

FIG. 3 shows a chromatogram obtained under the above conditions. The sample was obtained by detecting 100 pmol (10 μM, 10 μl) of each with sorbitol, glucose, and lactose.

FIG. 4 is a graph showing the reproducibility of the response under the above conditions. This graph was obtained by injecting the sample 20 times every 30 minutes and plotting the transition of the response peak to glucose and lactose in percentage at each time, and it was confirmed that good reproducibility was obtained.

Furthermore, the detection limit is 1.6 pmol (S / N =
In 3), the comparison of the detection limit with other conventional methods was the highest in sensitivity as shown in Table 2 using glucose as an example.

[0020]

[Table 2]

As the rhodium electrode in the electrochemical detector according to the present invention, in addition to the rhodium wire press-fitted into the insulator as in the above-described embodiment, a rhodium electrode coated with rhodium on the surface of various materials is used. be able to.

The electrochemical detector of the present invention can of course be used for flow injection analysis, capillary electrophoresis analysis and the like in addition to the above-described high performance liquid chromatography.

[0023]

As described above, according to the present invention, by forming a working electrode of an electrochemical detector with rhodium, the rhodium acts as a catalyst for accelerating a saccharide oxidation reaction in an alkaline solution, By measuring the oxidation current, it is possible to detect saccharides with high sensitivity and high reproducibility.

Moreover, since no sugar is adsorbed on the rhodium electrode surface, there is no need to apply a complicated pulse potential unlike the PAD, and the system configuration is simplified as compared with the PAD, and the operation is simplified. It will be easier.

[Brief description of the drawings]

FIG. 1 is a longitudinal sectional view showing a configuration of an embodiment of the present invention.

FIG. 2 is a block diagram showing a standard configuration of a high performance liquid chromatography to which an embodiment of the present invention is applied.

FIG. 3 is an example of a chromatogram when the present invention is mounted on the apparatus of FIG. 2 and saccharides are actually detected.

FIG. 4 is a graph showing the reproducibility by transition of detection peaks of glucose and lactose.

[Explanation of symbols]

 1 working electrode (rhodium electrode) 2 insulator block 3 auxiliary electrode block 6 reference electrode 6 10 liquid pump 11 sample Injector 12 ... LC column 13 ... ECD cell

────────────────────────────────────────────────── ─── Continued on the front page (58) Fields surveyed (Int. Cl. ⁷ , DB name) G01N 27/416 G01N 27/28 331 G01N 27/30 G01N 30/64 BIOSIS (DIALOG) JICST file (JOIS) WPI (DIALOG)

Claims (1)

(57) [Claims] 1. An electrochemical detector for detecting saccharides with high sensitivity using an electrode reaction, wherein a rhodium electrode is used as a working electrode.