JPH03125955A - Biological gas sensor - Google Patents

Abstract

PURPOSE:To selectively and quantitatively measure a gaseous substance by simple constitution and operation by forming an immobilized membrane of a living body related substance on the responding part of a measuring element and setting a physiologically active stabilizer so as to surround the periphery of the immobilized film. CONSTITUTION:An immobilized membrane 12 having living body related substance immobilized thereon is formed on the responding part of a measuring element 11 detecting physiologically active reaction. A gas permeable membrane 14 is mounted to one opening part of a case 13 setting the element 11 at the central axial part thereof by a ring 15 and a seal material 16 is set to the other opening part of the case 13. The hermetically closed space demarcated by the seal member 16 and the gas permeable membrane 14 in the case 13 is packed with a physiologically active stabilizer 17. When a gas component flows in the case from the outside through the gas permeable membrane 14, specific reaction is generated only in the substance to which the living body related substance in the immobilized membrane 12 shows physiological activity in the gas component and the physical or chemical change due to said reaction is detected by the element 11.

Description

[Detailed Description of the Invention] [Industrial Application Field] The present invention is applicable to, for example, indoor odor measurement, alcohol detection,
The present invention relates to a biogas sensor using biologically related substances that enables the measurement of specific gaseous substances and is used for gas leak detection, disease diagnosis by measuring body odor, etc.

[Prior Art] For example, in the field of gas substance measurement such as environmental analysis, there is a need for qualitative and quantitative gas analysis means that have excellent responsiveness and are easy to operate. Gas chromatography, which is known as a gas analysis method, has high selectivity and is excellent as a qualitative and quantitative analysis method, but it does not have excellent responsiveness and Handling is complicated.

Semiconductor gas sensors are known as ones with excellent responsiveness, and although these semiconductor gas sensors have excellent operability, they have poor selectivity with respect to the type of gas. Although various other measuring means have been developed, there is no simple gas analysis means that can satisfy both high responsiveness and selectivity.

In the medical field, biosensors have been put into practical use in recent years. This biosensor utilizes the physiological activity of biologically related substances and detects it with a chemical/physical sensor, and has excellent responsiveness and selectivity. However, the biological substances used here can only be used in an appropriate environment, and cannot be directly applied to gas analysis.

[Problems to be Solved by the Invention] The present invention has been made in view of the above-mentioned points, and it is possible to perform selective and quantitative gas measurement of gaseous substances with a simple configuration and simple operation. The present invention aims to provide a biogas sensor that can be used as a measuring device for various odor sensors, gas leak detection, alcohol detection, disease diagnosis, and the like.

[Means for Solving the Problems] A biogas sensor according to the present invention includes an immobilized film on which a biologically related substance is immobilized, which is formed on a sensitive part of a measuring element that detects a physiologically active reaction of the immobilized film. configured to set the bioactive stabilizer so as to surround the periphery of the
A measurement gas from the outside is made to act on the immobilization membrane through the gas-permeable membrane.

[Function] In the biogas sensor configured in this way, among the gas components that have flowed in through the gas permeable membrane, only the biologically-related substances in the immobilized membrane exhibiting physiological activity are specific. Begins to show a reaction. The measuring element then detects a physical or chemical change due to the reaction. Therefore, the responsiveness and operability of specific detection of the measurement gas substance can be obtained in accordance with the performance of the measuring element, and excellent responsiveness and easy operability can be obtained. In addition, biologically-related substances that have a large effect on measurement can be reliably maintained in their activity by the use of a physiologically active stabilizer, which is effective in improving reliability.

[Embodiments of the invention] Hereinafter, one embodiment of the present invention will be described with reference to the drawings.

FIGS. 1 and 2 show its configuration, and this biogas sensor is equipped with a measuring element 11 consisting of a Bolaro-type diaphragm-type oxygen electrode. The diaphragm-type oxygen electrode that constitutes this measurement element IT is composed of a platinum cathode, a silver/silver chloride anode, a potassium chloride electrolyte, and a Teflon (registered trademark) diaphragm, and has the function of detecting physiologically active reactions. . An immobilization film 12 on which a biologically related substance is immobilized is formed on the sensitive part of the measurement element II.

The measuring element 11 on which the immobilized film 12 is formed is placed in the solid axis of a cylindrical case 13, and one opening of the case 13 is provided with a gas permeable film 14. is attached by a ring 15. Further, a sealing material 16 is provided on the opposite side of the case 13 from the opening surface where the gas-permeable membrane 14 is set, so as to seal the space between the sealing material 16 and the measuring element 11. A sealed space partitioned by the sealing material 1B and the gas permeable membrane 14 is formed.

Then, this sealed space is filled with a physiologically active stabilizer 17.

That is, this physiologically active stabilizing material 17 is set to surround the immobilization film 12 formed on the measurement element 11 and is allowed to flow into the inside of this film 12. This prevents the physiologically active stabilizer 17 from leaking to the outside, and also prevents the inflow of substances from the outside. At least a portion of the bioactive stabilizer (7) is in contact with the gas permeable JII 114, and this gas permeable membrane 14 allows gas to pass through from the outside. however,
This gas permeable membrane 14 may be configured to be in contact with the immobilized membrane 2 containing the physiologically active stabilizer I7.

In the gas sensor configured in this way, a gas component flows in from the outside through the gas permeable membrane I4, and the biologically related substances in the immobilized membrane 12 become physiologically active in the gas component that flows in. A specific reaction occurs only with the indicated substance, and a physical or chemical change due to the reaction is detected by the measuring element 11.

In this case, the responsiveness and operability of specific detection of this gaseous substance come to conform to the basic performance of the measuring element 11,
Good responsiveness and easy operability can be obtained. Furthermore, the bioactive substance is maintained in its active state by the physiologically active stabilizer 17.

This gas sensor is constructed using, for example, alcohol oxidase as a biologically related substance, and thus constitutes a biogas sensor for alcohol.

Specifically, the immobilized membrane 12 is constituted by an immobilized enzyme membrane in which alcohol oxidase is immobilized using acrylamide gel. Alcohol oxidase specifically oxidizes primary alcohols and during this oxidation reduces oxygen as a hydrogen acceptor.

In other words, oxygen is consumed according to the amount of reaction alcohol due to the catalytic action.

As the measuring element 11, a polaro-type diaphragm-type oxygen electrode is used, and an immobilized membrane 12 is attached to the sensitive part.Enzymes are a type of protein, and form a polypeptide chain made of 20 types of amino acids. ing. The physiological activity of enzymes is based on the unique three-dimensional structure of this chain, which is made up of hydrogen bonds, hydrophobic bonds, etc.

This bond is easily broken by the surrounding environment, resulting in denaturation of the three-dimensional structure and loss of biological activity.

Therefore, in order to maintain this physiological activity, the immobilized membrane 12 is held in a physiologically active stabilizer 17, and as this physiologically active stabilizer 17, a phosphate buffer is used. This phosphate buffer solution is held in a tank partitioned by an O-ring 15 of a case 13 made of a Teflon vibrator, a gas permeable membrane, and a sealing material 1B, and leakage of the solution to the outside is prevented. This prevents substances from entering from outside. Then, the opening surface of the case 13 is closed by a gas permeable membrane 14 made of a Teflon membrane.

In this case, the immobilized enzyme membrane (immobilized membrane 12) impregnated with a phosphate buffer is in contact with the Teflon gas-permeable membrane 14, allowing the inflow of gas substances from this gas-permeable membrane 14. .

In the biogas sensor for alcohol configured in this way, when gas containing alcohol exists outside the sensor, this alcohol gas passes through the gas permeable membrane 14 and contains the phosphate buffer. It reaches the immobilization membrane 12. In this case, the immobilized enzyme membrane of the immobilized membrane 12 is adjusted to an appropriate pH by the phosphate buffer constituting the physiologically active stabilizer 17.
is maintained and stabilized.

The alcohol that has reached the immobilization membrane (2) is oxidized by enzymes and oxygen is consumed. In this case, the amount of oxygen consumed corresponds to the concentration of permeated alcohol gas from the outside, and oxygen consumption occurs in accordance with the external alcohol gas concentration.

This amount of oxygen consumption is then displayed as a current value by the oxygen electrode of the measuring element 11.

However, if a gas in which alcohol oxidase does not exhibit physiological activity exists outside, such as acetone or benzene, and passes through the gas permeable membrane 14, these gases will not cause an equivalent catalytic reaction, and therefore will not release oxygen. It is not consumed and is not displayed by the measuring element 11.

Figure 3 shows the relationship between the output current of the biogas sensor configured according to this embodiment and the external gas concentration at that time.The horizontal axis is a logarithmic display, and the current output corresponding to the alcohol gas concentration is obtained. There is. However, there is no response output for other gases, and the effectiveness of this gas sensor for alcohol gas measurement as well as its substance specificity can be confirmed. Furthermore, the response of this biogas sensor reached a saturation value within a few minutes, which indicates good responsiveness.

The alcohol biosensor constructed in this way maintains its activity with a sufficiently simple construction, and by using a measuring element with a diaphragm-type oxygen electrode, it can be easily operated.
It is possible to specifically measure primary alcohol, and its constituent materials and sensor conditions will be further described below.

The measuring element 11 composed of a diaphragm-type oxygen electrode is an electrochemical sensor using a Polaro-type Teflon (registered trademark) membrane as a diaphragm, and includes a platinum cathode, a silver/silver chloride anode,
Using potassium chloride electrolyte, cathode potential is -0.7V
vs, Ag/Ag C1l. Furthermore, the immobilized membrane 12 was immersed in a photocrosslinkable acrylamide gel solution at 1280° C.
units/ml of alcohol oxidase (E,
C, 1, 1, 3, 13) were mixed at a volume ratio of 4:1, and 2
A 54 nm ultraviolet ray (15 W) was irradiated for several minutes to form a film with a thickness of 150 μm. Furthermore, the phosphate buffer solution which is considered to be the bioactive stabilizer 17 is 67m
mol/fl, pH 7.0 is used, and the Teflon vibrator making up case 13 has an outer diameter of 40 m.
A tss with an inner diameter of 2011% and a length of 20I was used. The Teflon membrane used as the gas permeable membrane 14 had a pore diameter of 1 to 2 μm and a membrane thickness of 0.25 m5.

That is, in the Polaro diaphragm type oxygen electrode constituting the measurement element 11, by applying a predetermined potential to the cathode side using a constant potential setting device (potentiostat),
Since a reduction reaction occurs on the cathode surface and a reduction current flows, a current corresponding to the oxygen concentration outside the diaphragm can be obtained. In the immobilization membrane 12, the primary alcohol is specifically oxidized by the alcohol oxidase immobilized inside the porous interior, and oxygen is reduced and consumed as a hydrogen acceptor, that is, the catalytic reaction corresponds to the amount of alcohol. Oxygen consumption occurs.

The phosphate buffer serving as the bioactive stabilizer 17 functions to stabilize the activity of the enzyme in the immobilized enzyme membrane constituting the immobilized membrane 12 . Enzymes exert their activity due to their unique three-dimensional structure, but this structure can easily change depending on the environment around the enzyme, resulting in loss of activity. However, the phosphate buffer permeates into the interior of the immobilized enzyme membrane, maintaining its structure and acting to stabilize and maintain the physiological activity of the enzyme itself.

In the biogas sensor configured in this manner, oxygen molecules pass through the gas-permeable membrane 14 and flow into the interior together with the atmosphere when present in the general atmosphere. The oxygen molecules that have passed through the gas-permeable membrane 14 are then dissolved in the buffer solution, which is the physiologically active stabilizer 17, and also pass through the immobilization membrane 14 to reach the sensitive part of the measurement element 11. In this sensitive part, a reduction reaction occurs due to the reduction voltage, a reduction current flows according to the dissolved oxygen molecular weight, and this reduction current is measured.

Here, if alcohol scum molecules exist outside the gas-permeable membrane 14, the alcohol molecules also pass through the gas-permeable membrane 14 and reach the immobilization membrane 12 in the same way as oxygen molecules. The alcohol molecules that have reached the immobilized film 12 are oxidized by alcohol oxidase using oxygen as a hydrogen acceptor. Through this catalytic reaction, alcohol is converted into an oxidant, and oxygen corresponding to the amount of alcohol that has flowed in, that is, the concentration of alcohol gas present outside, is consumed.

Due to such oxygen consumption, the amount of oxygen molecules reaching the electrode sensitive part of the measuring element 11 is reduced, and the photocurrent is reduced.

Through such an action, it is possible to obtain an output current corresponding to the alcohol gas concentration outside the gas permeable membrane of this biogas sensor. If the substance permeates through the gas permeable membrane 14, no equivalent catalytic reaction occurs and no oxygen consumption and associated reduction in output current occurs.

[Effects of the Invention] As described above, in the biogas sensor according to the present invention, an immobilized film on which a biologically related substance is immobilized is formed on the sensitive part of the measurement element capable of detecting the physiologically active reaction thereof, It was configured to be surrounded by a bioactive stabilizer. Among the gas components that have passed through the gas-permeable membrane from the outside, a specific reaction occurs only with biologically-related substances that exhibit physiological activity, and physical or chemical changes due to this reaction are detected by the measurement element. It becomes like this. In other words, selective and quantitative measurement of gaseous substances can be performed with simple operations, and the response in this case can be obtained according to the measuring element used, making it suitable for odor measurement, alcohol detection, gas leak detection, etc. A biogas sensor can be obtained that can be effectively applied to disease diagnosis through detection and body odor measurement.

[Brief Description of the Drawings] Fig. 1 is a perspective view showing a biogas sensor according to an embodiment of the present invention, Fig. 2 is a cross-sectional configuration diagram corresponding to the line ■-■ in Fig. 1, and Fig. 3 is the above-mentioned FIG. 3 is a diagram showing the operating characteristics of the gas sensor shown in the example. 1■...11 constant element, 12...immobilization film, 13...
- Case, 14... Gas permeable membrane, 1B... Seal material, ■7... Bioactive stabilizer.

Claims (1)

[Scope of Claims] A measurement element that detects a physiologically active reaction, an immobilization film that immobilizes a biological substance formed on a sensitive part of the measurement element, and a case set around the measurement element. and a gas permeable membrane set to cover an opening formed in correspondence with a portion of the case where the immobilization membrane is formed, and a gas permeable membrane partitioned by the gas permeable membrane and formed inside the case. a physiologically active stabilizer filled in the sealed space, and among the gas components introduced through the gas permeable membrane,
A biogas sensor in which only components reacted on the immobilized membrane are detected by the measurement element.