Biosensor containing glucose dehydrogenase

Details Biosensor containing glucose dehydrogenase Biosensor containing glucose dehydrogenase

1999-06-30

2003-12-02

Gitomer, Ralph (Department: 1651)

Chemistry: molecular biology and microbiology

Measuring or testing process involving enzymes or...

Involving oxidoreductase

C205S777500

Reexamination Certificate

active

06656702

ABSTRACT:

This application claims benefit to Japan HEI 10-188799 filed Jul. 3, 1998.
BACKGROUND OF THE INVENTION
The present invention relates to a biosensor which facilitates simple and rapid quantitation of a specific component contained in a sample with high accuracy.
Conventionally, various biosensors have been proposed as the system to enable simple quantitation of a specific component contained in a sample without the need of dilution or agitation of a sample solution. The following is a known example of such biosensor (Japanese Laid-Open Patent Publication Hei 2-062952).
The biosensor disclosed in this prior art is produced by the steps of forming an electrode system including a working electrode, a counter electrode and a reference electrode on an electrically insulating base plate using a screen printing method or the like and subsequently forming immediately above this electrode system an enzyme reaction layer including a hydrophilic polymer, an oxidoreductase and an electron acceptor.
When the biosensor thus produced is added with a drop of a sample solution containing a substrate over the enzyme reaction layer, dissolution of the enzyme reaction layer in the sample solution will occur, which triggers a reaction between the enzyme and the substrate (enzyme reaction). As a result, reduction of the electron acceptor will develop. Upon completion of this enzyme reaction, the reduced electron acceptor is reoxidized electrochemically. Based on the oxidation current value measured during this reoxidizing step, the concentration of the substrate in the sample solution can be quantitated.
The biosensor as described above permits measurements of various materials in principle if a suitable enzyme corresponding to the substrate of a target material is selected.
Enzymes, which contain protein as their main component, are normally present in dry state inside a biosensor. However, water contained in air may come in or out of the enzyme through its surface depending on the condition of temperature and humidity of air. Therefore, long contact of the enzyme with air will result in a change in water content which is present in the enzyme in a slight amount. As a result, the enzyme develops degeneration and loses its enzyme activity.
During preservation of a sensor after its production, when the activity of an enzyme contained in the sensor is impaired with time, the enzyme which should participate in enzyme reaction with a substrate is depleted. This produces a problem of incommensuration of the measured response current value with the substrate concentration.
In order to solve the above-mentioned problem, it is important to secure an environment which can retain enzyme activity for a long time in the vicinity of the enzyme.
It is also necessary to make smooth transfer of the substrate and electrons during enzyme reaction in order to increase sensor response.
BRIEF SUMMARY OF THE INVENTION
In view of the above-mentioned problem, the object of the present invention is to provide a biosensor with high stability against preservation by best reducing impairment of the activity of enzyme contained in the biosensor during its preservation after production.
Particularly, the present invention provides a compact disposable biosensor of low cost.
The present invention provides a biosensor comprising an electrically insulating base plate, an electrode system including at least a working electrode and a counter electrode formed on the base plate, and a reaction layer containing at least an enzyme and a sugar formed on the electrode system, wherein the enzyme is at least one selected from the group consisting of glucose oxidase, glucose dehydrogenase and fructose dehydrogenase and the sugar is at least one selected from the group consisting of trehalose, sucrose, glycerol, mannitol and ribose.
In a preferred mode of the present invention, the enzyme is glucose dehydrogenase whose coenzyme is pyrrolo-quinoline quinone and the sugar is trehalose.
While the novel features of the invention are set forth particularly in the appended claims, the invention, both as to organization and content, will be better understood and appreciated, along with other objects and features thereof, from the following detailed description taken in conjunction with the drawings.


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Derwent abstract of JP 05087767 A (Acc. No. 1993-149757), 1993. Biosensor of long sensor life and prevented deactivation of biological substance.*
Sprules et al. (1996). A disposable reagentless screen-printed amperometric biosensor for the measurement of alcohol in beverages. Analytica Chimica Acta 329(3), pp. 215-221.*
Koji Sode* and Nozomu Yasutake, “Preparation of lyophilized pyrroloquinoline quinone glucose dehydrogenase using trehalose as an additive”, Biotechnology Techniques, vol. 11, No. 8, Aug. 1997, pp. 577-580.
Sprules, S.D. et al.: “A disposable reagentless screen-printed amperometric biosensor for the measurement of alcohol in beverages”, Analytica Chimica Acta, vol. 329, No. 3, pp. 215-221 (Mar., 1996).

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