【０００１】0001
【産業上の利用分野】本発明は、隔膜式過酸化水素電極
型のバイオセンサに関し、更に詳しく言えば、隔膜をシ
リコンゴム等の疎水性材質から構成し、且つこの隔膜に
より電極を被覆した構成により電極面積を大きくするこ
とで、水溶性の妨害物質の影響を小さくし且つ感度の優
れたバイオセンサに関する。本発明は、臨床検査、化学
、食品工業、環境計測等に利用される。[Field of Industrial Application] The present invention relates to a diaphragm-type hydrogen peroxide electrode type biosensor, and more specifically, a diaphragm made of a hydrophobic material such as silicone rubber, and an electrode covered with the diaphragm. The present invention relates to a biosensor that reduces the influence of water-soluble interfering substances and has excellent sensitivity by increasing the electrode area. INDUSTRIAL APPLICATION This invention is utilized for a clinical examination, chemistry, the food industry, environmental measurement, etc.
【０００２】0002
【従来の技術】従来のバイオセンサとしては、電極をガ
ラス、プラスチック等の電極支持筒に、電解液とともに
入れ、電極の端面を親水性の隔膜で被覆した構造の過酸
化水素電極に酵素固定化膜を装着したものが知られてい
る。[Prior art] In conventional biosensors, enzymes are immobilized on hydrogen peroxide electrodes, which have a structure in which an electrode is placed together with an electrolyte in an electrode support tube made of glass or plastic, and the end surface of the electrode is covered with a hydrophilic diaphragm. A type equipped with a membrane is known.
【０００３】0003
【発明が解決しようとする問題点】しかし、従来の上記
バイオセンサでは、親水性の隔膜を使用しているので、
被測定液中の水溶性の妨害物質（アスコルビン酸、尿酸
等）が隔膜内部に浸透し、電極上で誤差電流として検出
され、正確な測定ができなかった。また、電極の端面だ
けを検出部としているので、電極本体の大きさに比べて
小さな電極面積しか得られず、感度が低く、小型化がで
きなかった。[Problems to be solved by the invention] However, since the conventional biosensor uses a hydrophilic diaphragm,
Water-soluble interfering substances (ascorbic acid, uric acid, etc.) in the liquid to be measured penetrated into the diaphragm and were detected as error currents on the electrodes, making accurate measurements impossible. In addition, since only the end face of the electrode is used as the detection part, only a small electrode area can be obtained compared to the size of the electrode body, resulting in low sensitivity and making it impossible to miniaturize.
【０００４】本発明は、上記問題点を解決するものであ
り、水溶性の妨害物質の影響が小さくて正確な測定が可
能で、且つ感度に優れた隔膜式過酸化水素電極型のバイ
オセンサを提供することを目的とする。The present invention solves the above problems and provides a diaphragm-type hydrogen peroxide electrode type biosensor that is less affected by water-soluble interfering substances, enables accurate measurements, and has excellent sensitivity. The purpose is to provide.
【０００５】[0005]
【課題を解決するための手段】本第１発明の、内部電解
液をもつ隔膜式過酸化水素電極型のバイオセンサは、疎
水性の過酸化水素透過性筒体と、該筒体内に保持される
電極と電解液と、該筒体の外側に配設される酵素固定化
膜と、を有することを特徴とする。また、本第３発明の
バイオセンサは、例えば、第６図に示すように、凹部を
有する絶縁基板７と、該凹部内に配設、充填される電極
１、２と電解液（緩衝液）６と、該凹部上に配設され上
記電解液を密封する疎水性の過酸化水素透過性膜８と、
該過酸化水素透過性膜上に積層される酵素固定化膜４と
、を有する積層型構成とすることもできる。[Means for Solving the Problems] A diaphragm-type hydrogen peroxide electrode type biosensor having an internal electrolyte according to the first invention includes a hydrophobic hydrogen peroxide-permeable cylinder and a hydrogen peroxide-permeable cylinder held within the cylinder. The cylindrical body is characterized by having an electrode, an electrolytic solution, and an enzyme-immobilized membrane disposed outside the cylindrical body. Further, the biosensor of the third invention includes, for example, as shown in FIG. 6, an insulating substrate 7 having a recess, electrodes 1 and 2 disposed and filled in the recess, and an electrolyte (buffer). 6, a hydrophobic hydrogen peroxide permeable membrane 8 disposed on the recess and sealing the electrolyte;
It is also possible to have a laminated structure including an enzyme immobilization membrane 4 laminated on the hydrogen peroxide permeable membrane.
【０００６】上記過酸化水素透過性筒体又は過酸化水素
透過性膜としては、被測定水溶液中の水溶性妨害物質（
アルコルビン酸等）を透過させず且つ発生する過酸化水
素を透過させるような疎水性材質からなればよく、例え
ば、シリコンゴム及びその他のテフロン膜等とすること
ができる。また、使用する酵素は被測定物質（例えばグ
ルコース等）との相関において種々選択される。そして
、酵素固定化膜の形態としては、膜状（層状も含む。）
となっておればよい。従って、この酵素を担持、固定す
る基材としては、膜状となるものであればよく、特に限
定されず、例えば、不織布、セルロースアセテート等と
することができるし、更に、直接、筒体等の表面に担持
させることもできる。The hydrogen peroxide-permeable cylinder or hydrogen peroxide-permeable membrane may contain water-soluble interfering substances (
The material may be made of a hydrophobic material that does not allow hydrogen peroxide (such as ascorbic acid) to pass therethrough and allows hydrogen peroxide to pass through, such as silicone rubber or other Teflon membranes. Furthermore, various enzymes are selected based on the correlation with the substance to be measured (for example, glucose, etc.). The form of the enzyme-immobilized membrane is membranous (including layered).
It is sufficient if it becomes . Therefore, the substrate for supporting and immobilizing this enzyme is not particularly limited as long as it is in the form of a membrane. For example, it can be a nonwoven fabric, cellulose acetate, etc. It can also be supported on the surface of.
【０００７】[0007]
【作用】本発明においては、電極が疎水性の過酸化水素
透過性筒体等で覆われているため、発生する過酸化水素
はこの筒体若しくは膜を透過して電極上まで拡散するが
、被測定液中の水溶性の妨害物質が電極上まで拡散でき
ないので、正確な測定ができる。また、この過酸化水素
透過性筒体等及び酵素固定化膜が電極全体を覆うので、
全体を検出部とした構成となり、そのため電極面積が大
きくなり、感度を向上させることができる。更に、この
過酸化水素透過性筒体を用いる場合は、電極支持筒とし
ての機能も同時に有する。[Operation] In the present invention, since the electrode is covered with a hydrophobic hydrogen peroxide permeable cylinder, the generated hydrogen peroxide passes through this cylinder or membrane and diffuses onto the electrode. Accurate measurement is possible because water-soluble interfering substances in the liquid to be measured cannot diffuse onto the electrode. In addition, since this hydrogen peroxide permeable cylinder etc. and the enzyme immobilization membrane cover the entire electrode,
The entire device is configured as a detection section, which increases the electrode area and improves sensitivity. Furthermore, when this hydrogen peroxide permeable cylinder is used, it also functions as an electrode support cylinder at the same time.
【０００８】[0008]
【発明の効果】以上のように、本発明の過酸化水素電極
型のバイオセンサを用いれば、水溶性の妨害物質の影響
が小さな正確な測定ができ、また感度に優れたものとす
ることができる。また、この過酸化水素透過性筒体を用
いる場合は、電極支持筒としての機能も同時に有するの
で、安定して電極を同時に保持できるとともに、全体を
検出部とした構成とすることができるので、センサの小
型化が容易となる。更に、過酸化水素透過性筒体又は過
酸化水素透過性膜の材料としてシリコンゴムを用いる場
合は、柔軟性及び弾力性に優れるので、種々の形状に設
計でき、また、相手材形状に自由に合わせることもでき
る。[Effects of the Invention] As described above, by using the hydrogen peroxide electrode type biosensor of the present invention, accurate measurement can be performed with less influence of water-soluble interfering substances, and it can also be made with excellent sensitivity. can. In addition, when using this hydrogen peroxide permeable cylinder, it also functions as an electrode support cylinder, so it can stably hold the electrodes at the same time, and the entire structure can be used as a detection part. It becomes easy to downsize the sensor. Furthermore, when silicone rubber is used as the material for the hydrogen peroxide permeable cylinder or hydrogen peroxide permeable membrane, it has excellent flexibility and elasticity, so it can be designed into various shapes and can be freely adapted to the shape of the mating material. You can also match them.
【０００９】[0009]
【実施例】以下実施例により本発明を具体的に説明する
。実施例１本実施例は、妨害物質に対する選択性を比較例１と比較
しつつ検討したものである。本実施例１の隔膜式過酸化
水素電極型グルコースセンサは、図１（縦断面図）及び
図２（横断面図）に示すように、シリコンゴムチューブ
３と、このチューブ１の両端を閉じるシリコン栓５と、
この中に保持される電極（作用極１と対極２）と電解液
６と、このチューブ３の外側に配設される酵素固定化不
織布（膜）４とからなる。このセンサは、以下のように
して作製された。[Examples] The present invention will be explained in detail with reference to Examples below. Example 1 In this example, selectivity to interfering substances was investigated in comparison with Comparative Example 1. As shown in FIG. 1 (longitudinal cross-sectional view) and FIG. 2 (cross-sectional view), the diaphragm-type hydrogen peroxide electrode type glucose sensor of Example 1 includes a silicone rubber tube 3 and a silicone rubber tube 3 that closes both ends of the tube 1. Stopper 5 and
It consists of electrodes (working electrode 1 and counter electrode 2) held therein, an electrolyte 6, and an enzyme-immobilized nonwoven fabric (membrane) 4 disposed outside the tube 3. This sensor was manufactured as follows.
【００１０】まず、シリコンゴムチューブ（直径；２ｍ
ｍφ、長さ；ｌ１５ｍｍ、肉厚；５０μｍ）に緩衝液（
リン酸緩衝液、ｐＨ＝７．０）を入れ、一端をシリコン
栓５で封止する。次に、一対の電極として１ｍｍφの白
金線１、２を２本用意し、１本の白金線２にナイロンネ
ット９を巻き、白金線１、２が互いに直接接触しない様
にしてシリコンゴムチューブ３内に挿入する。シリコン
ゴムチューブ３のもう一端からリード１１、２１を取り
出し、他方の開口部を同様にシリコン栓５で封止する。その後、所定の不織布を５％グルコースオキシダーゼ溶
液に浸漬し、その後グルタルアルデヒドで固定化処理を
行った、そして、この酵素が固定化された不織布４をシ
リコンゴムチューブ３の外側全体に巻きつけて、グルコ
ースセンサを構成した。First, a silicone rubber tube (diameter: 2 m)
mφ, length: 15 mm, wall thickness: 50 μm) and buffer solution (
A phosphate buffer solution (pH=7.0) is added thereto, and one end is sealed with a silicone stopper 5. Next, prepare two platinum wires 1 and 2 with a diameter of 1 mm as a pair of electrodes, wrap a nylon net 9 around one platinum wire 2, and keep the platinum wires 1 and 2 from coming into direct contact with each other in a silicone rubber tube 3. Insert inside. Leads 11 and 21 are taken out from the other end of silicone rubber tube 3, and the other opening is similarly sealed with silicone plug 5. After that, a predetermined nonwoven fabric was immersed in a 5% glucose oxidase solution, and then immobilized with glutaraldehyde, and this enzyme-immobilized nonwoven fabric 4 was wrapped around the entire outside of the silicone rubber tube 3. A glucose sensor was constructed.
【００１１】本発明の妨害物質（アスコルビン酸及び尿
酸）に対する選択性を調べるため、作用極に＋６００ｍ
Ｖの定電位を印加し、電流値を測定した。この場合、グ
ルコース（１００ｍｇ／ｄｌ）だけの場合の検出電流値
（実施例１の場合は０．２０μＡ、比較例１の場合は０
．１８μＡ）を１００とし、これにアスコルビン酸１０
ｍｇ／ｄｌを添加した場合、尿酸１０ｍｇ／ｄｌを添加
した場合の各々を百分率で示した。尚、比較例１として
は、隔膜として親水性の酢酸セルロース膜を用いたグル
コースセンサを用いた。これらの結果を表１に示す。In order to investigate the selectivity of the present invention for interfering substances (ascorbic acid and uric acid), +600 m was applied to the working electrode.
A constant potential of V was applied and the current value was measured. In this case, the detected current value in the case of only glucose (100 mg/dl) (0.20 μA in Example 1, 0 in Comparative Example 1)
．． 18 μA) as 100, and ascorbic acid 10
When uric acid was added at 10 mg/dl, and when 10 mg/dl of uric acid was added, each was expressed as a percentage. In addition, as Comparative Example 1, a glucose sensor using a hydrophilic cellulose acetate membrane as a diaphragm was used. These results are shown in Table 1.
【００１２】0012
【表１】[Table 1]
【００１３】表１の結果に示すように、実施例１のセン
サにおいては、アスコルビン酸及び尿酸の影響が全くな
かったが、比較例１においては、各々４０％及び４５％
も影響を示した。As shown in the results in Table 1, the sensor of Example 1 was not affected by ascorbic acid and uric acid at all, but in Comparative Example 1, the effects of ascorbic acid and uric acid were 40% and 45%, respectively.
also showed an impact.
【００１４】実施例２本実施例は、電極端面だけを検出部としたセンサ（比較
例２）と本発明のセンサとの感度の比較を行ったもので
ある。本実施例で用いたセンサは実施例１と同じグルコ
ースセンサである。比較例２のセンサとしては、図３に
示すように、、実施例１で用いた２本の白金線（φ１ｍ
ｍ、ｌ１０ｍｍ）１、２をガラス管（直径；３ｍｍφ、
長さ；ｌ１５ｍｍ）１０内に挿入し、白金線１の端面に
シリコンゴム膜８を装着し、この膜８の外側に酵素固定
化膜（実施例１で用いた酵素固定化不織布）４を配置し
た構成のものを用いた。尚、電極本体の大きさは、実施
例２、比較例２ともにほとんど同じである。上記両セン
サを用いて、グルコース１００ｍｇ／ｄｌ及び３００ｍ
ｇ／ｄｌに対する応答電流値を測定して、感度を比較し
た結果を表２に示す。Example 2 In this example, the sensitivity was compared between a sensor in which only the electrode end face was used as a detection part (Comparative Example 2) and the sensor of the present invention. The sensor used in this example is the same glucose sensor as in Example 1. As shown in FIG. 3, the sensor of Comparative Example 2 was made of two platinum wires (φ1 m
m, l10mm) 1 and 2 are glass tubes (diameter; 3mmφ,
length: 15 mm) 10, a silicone rubber membrane 8 is attached to the end face of the platinum wire 1, and an enzyme immobilization membrane (enzyme immobilization nonwoven fabric used in Example 1) 4 is placed on the outside of this membrane 8. A device with a similar configuration was used. Note that the size of the electrode body is almost the same in both Example 2 and Comparative Example 2. Using both the above sensors, glucose 100mg/dl and 300m
Table 2 shows the results of measuring the response current values to g/dl and comparing the sensitivities.
【００１５】[0015]
【表２】[Table 2]
【００１６】表２の結果によれば、本実施例２では、シ
リコンゴムチューブで電極全体を被覆し、且つこのチュ
ーブの全表面上に酵素固定膜を被覆したので、電極全体
を検出部とすることができ、広い電極面積が確保できた
。一方、比較例２では、電極の端面だけを検出部とする
ものである。従って、本実施例２のセンサは、比較例２
のセンサと比べて感度が５〜６倍向上した。また、この
シリコンゴムチューブが電極支持筒の役割をも兼ねてい
る。According to the results in Table 2, in this Example 2, the entire electrode was covered with a silicone rubber tube, and the entire surface of this tube was covered with an enzyme-immobilized membrane, so the entire electrode was used as a detection section. This enabled us to secure a large electrode area. On the other hand, in Comparative Example 2, only the end face of the electrode is used as a detection part. Therefore, the sensor of Example 2 is the same as that of Comparative Example 2.
Sensitivity has improved by 5 to 6 times compared to previous sensors. This silicone rubber tube also serves as an electrode support tube.
【００１７】尚、本発明においては、上記具体的実施例
に示すものに限られず、目的、用途に応じて本発明の範
囲内で種々変更した実施例とすることができる。即ち、
上記実施例では白金電極を用いたが、金、カーボン、金
属酸化物等の電極でもよい。また、上記のように、電極
支持筒としてシリコンゴムチューブを使用する場合は、
柔軟性、弾力性があるので、バイオセンサの形状はどの
様にでも設計可能である。例えば、図５に示すように長
尺形状としこれを螺旋状に巻いたり、また図５に示すよ
うにＵ字形状としたり、目的、用途に応じて種々の曲管
状その他の形状とすることができる。更に、酵素固定膜
を配設する位置、場所も、筒体等の全表面上であっても
よいが、図４に示すようにその所望部分であってもよい
。It should be noted that the present invention is not limited to those shown in the above-mentioned specific embodiments, but can be modified in various ways within the scope of the present invention depending on the purpose and use. That is,
Although platinum electrodes were used in the above embodiments, electrodes made of gold, carbon, metal oxides, etc. may also be used. Also, as mentioned above, when using a silicone rubber tube as an electrode support tube,
Due to its flexibility and elasticity, the biosensor can be designed in any shape. For example, it can be made into a long shape and wound into a spiral as shown in Figure 5, or it can be made into a U-shape as shown in Figure 5, or it can be made into various curved tube shapes or other shapes depending on the purpose and use. can. Further, the enzyme-immobilized membrane may be disposed on the entire surface of the cylindrical body, or may be placed on a desired portion thereof as shown in FIG. 4.
【図１】実施例１のグルコースセンサの縦断面図である
。FIG. 1 is a longitudinal cross-sectional view of a glucose sensor of Example 1.
【図２】実施例１のグルコースセンサの横断面図である
。FIG. 2 is a cross-sectional view of the glucose sensor of Example 1.
【図３】比較例２のグルコースセンサの縦断面図である
。FIG. 3 is a longitudinal cross-sectional view of a glucose sensor of Comparative Example 2.
【図４】長尺曲管形状のバイオセンサの説明図である。FIG. 4 is an explanatory diagram of a biosensor in the shape of a long curved tube.
【図５】Ｕ字型管形状のバイオセンサの縦断面図である
。FIG. 5 is a longitudinal cross-sectional view of a U-shaped tube-shaped biosensor.
【図６】絶縁基板上に電極、内部電解液、過酸化水素透
過性膜及び酵素固定化膜を積層したバイオセンサの説明
断面図である。FIG. 6 is an explanatory cross-sectional view of a biosensor in which electrodes, an internal electrolyte, a hydrogen peroxide permeable membrane, and an enzyme immobilization membrane are laminated on an insulating substrate.
１ 作用極２ 対極３ 過酸化水素透過性筒体４ 酵素固定化膜５ シリコン栓６ 緩衝液７ 絶縁基板８ 過酸化水素透過性膜1 Working electrode2. Opposite3 Hydrogen peroxide permeable cylinder4 Enzyme immobilization membrane5 Silicone stopper6 Buffer solution7 Insulating substrate8 Hydrogen peroxide permeable membrane
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP3142445AJPH04343065A (en) | 1991-05-17 | 1991-05-17 | Biosensor |
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP3142445AJPH04343065A (en) | 1991-05-17 | 1991-05-17 | Biosensor |
| Publication Number | Publication Date |
|---|---|
| JPH04343065Atrue JPH04343065A (en) | 1992-11-30 |
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP3142445APendingJPH04343065A (en) | 1991-05-17 | 1991-05-17 | Biosensor |
| Country | Link |
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| JP (1) | JPH04343065A (en) |
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