JP5205583B2 - A chemical sensor that detects sensitive and selective taste-free non-ionized substances. - Google Patents

Description

  The present invention relates to a pseudo-biological lipid polymer membrane, a working electrode using the membrane, a membrane potential measuring apparatus having the working electrode, and a measuring method thereof.

  Taste evaluation is an extremely important task in the food and pharmaceutical industries. Such sensory tests have been performed by direct judgment with a conventionally trained human tongue. However, the judgment by the examiner has a problem that it is easily influenced by the physiological state, psychological state, or health state of the person.

  In order to solve the above problems and allow anyone to evaluate the taste objectively and accurately, a taste sensor device as an artificial transducer replacing a human tongue has been developed in recent years. For example, Patent Document 1 is a taste sensor using a lipid polymer membrane by the present inventors, and many taste sensors such as Patent Documents 2 to 5 are known based on this technology. Each of these sensors acquires a change in potential generated between the two substances when the taste substance is adsorbed on the surface of the lipid polymer film, etc., converted into an electrical signal as information on the taste substance. Such a taste substance recognition mechanism mimics a biological taste substance acceptance mechanism. Therefore, a taste sensor device using a lipid polymer film can measure and evaluate the taste with a quality and strength very close to human sensitivity.

  However, with regard to sweet substances, the conventional taste sensor device using a lipid polymer film has a response sensitivity and a response selectivity compared to the other four basic taste substances (salt taste substance, sour taste substance, bitter taste substance, and umami substance). There was a problem that was extremely low. One of the causes is that the device is a potential measuring device using a lipid polymer membrane. That is, non-ionizing substances such as sucrose and glucose that do not ionize are difficult to generate a high membrane potential in the lipid polymer membrane. This problem also affects the accuracy, stability, and reproducibility of the overall measurement results of the taste sensor device. Thus, the present inventors have so far developed a lipid polymer membrane having electrical responsiveness to sweet substances. For example, di-n-hexadecyl ester (2C16) and tetradodecyl ammonium bromide (hereinafter referred to as TDAB), di-n-octylphenyl phosphate (hereinafter referred to as DOPP) described in Non-Patent Document 1, and It has been found that a lipid polymer film composed of polyvinyl chloride (hereinafter referred to as PVC) has a relatively high response to 30 mM, which is the lowest sucrose concentration detectable by humans. However, the lipid polymer membrane has a problem that the specificity of the response to sucrose, that is, the selectivity to the sweet substance is insufficient as compared with other basic taste substances.

From the background described above, the development of lipid polymer membranes with high response sensitivity and selectivity for non-ionizing substances such as sweet ingredients, as well as the development of taste sensor devices that can measure sweet substances with high sensitivity It was desired.
JP-A-3-54446 JP-A-4-238263 JP-A-4-303755 JP-A-4-324351 JP-A-5-34311 JP-A-3-163351 Habara M, Ikezaki H, Toko K, Biosens. Bioelectron. , 2004, 19, 1559. Kiyoshi Toko, Central Analysis Center, Kyushu University: Center News, 1997, 15 (3), 2-8. Schallenberger RS and Acre TE, Nature, 1967, 216, 480. Kier L, Journal of Pharmaceutical Science, 1972, 61, 1394.

  The final subject of this invention is developing and providing the working electrode which can detect a non-ionizing substance electrically highly sensitively, and a membrane potential measuring apparatus which has the said working electrode. In order to solve the problem, it is first necessary to develop a working membrane for a working electrode having high response sensitivity and response selectivity for non-ionized substances. Accordingly, a first object of the present invention is to develop and provide a working membrane for working electrodes for non-ionizing substances, particularly sweet substances, which has been difficult with conventional lipid polymer membranes.

In order to solve the above-mentioned problems, the present inventors have conducted extensive research, and as a result, surface modification of a lipid polymer membrane with a polyhydric phenol makes the membrane highly potential responsive and selective to non-ionizing substances. It was found to have The inventions shown in the following (1) to (12) have been completed based on such findings, and are provided as means for solving the above problems.
(1) The present invention provides a pseudo-biological lipid polymer membrane comprising a membrane body portion comprising a polymer material and a plasticizer thereof, and a modification portion comprising a polyhydric phenol that modifies the surface of the membrane body portion. provide.
(2) The present invention produces a membrane body formed by mixing a polymer material and a plasticizer thereof, and is formed on the surface of the membrane body by immersing the membrane body in a solution containing polyhydric phenol for about 48 hours. Provided is a pseudo-biological lipid polymer membrane comprising a modified portion.
(3) The present invention provides a pseudo-biological lipid comprising a polymer material, a membrane main body made of a plasticizer thereof, and a lipid substance, and a modifying portion made of a polyhydric phenol that modifies the surface of the membrane main body. A polymer membrane is provided.
(4) In the present invention, a membrane body formed by mixing a polymer material, its plasticizer, and a lipid substance, and immersing the membrane body in a solution containing polyhydric phenol for about 48 hours. To provide a pseudo-biological lipid polymer film comprising a modifying part formed on the surface thereof.
(5) The present invention provides a pseudo-biological lipid polymer membrane, wherein the lipid substance is tetradodecyl ammonium bromide (TDAB).
(6) The present invention provides a pseudo-biological lipid polymer membrane, wherein the plasticizer is di-n-octylphenyl phosphate (DOPP) and phenylphosphonic acid monooctyl ester (PPME).
(7) The present invention provides a pseudo-biological lipid polymer membrane, wherein the polyhydric phenol of the modifying part is a polyhydric phenolic acid and / or a hydrolyzed tannin.
(8) The present invention provides a pseudo-biological lipid polymer membrane characterized in that the modifying portion has a function of generating a potential in the membrane main body in response to a foreign non-ionizing substance.
(9) The present invention provides a pseudo-biological lipid polymer membrane characterized in that the modifying part responds to a sweet substance which is a foreign non-ionizing substance.
(10) The present invention provides a support, the pseudo-biological lipid polymer membrane according to any one of (1) to (9), and a non-ionizing substance exogenous on the surface side of the pseudo-biological lipid polymer membrane. Provided is a working electrode having a conductor for acquiring a membrane potential generated in response from the back side thereof.
(11) The present invention provides a membrane potential measuring device having the working electrode.
(12) The present invention provides a non-ionizing substance measuring method in which a reference electrode and the working electrode are brought into contact with a measurement target solution and a difference in potential value generated at each electrode in response to the non-ionizing substance is measured.

  According to the pseudo-biological lipid polymer membrane of the present invention, by capturing a non-ionizing substance on its surface, it is possible to respond sensitively and selectively to the substance. Therefore, electrochemical changes (membrane potential measurement, impedance measurement, surface polarization control method, etc.), refractive index (dielectric constant) change (surface plasmon resonance method, etc.), mass change (quartz crystal, surface elasticity) (Wave measurement, etc.) If used in a sensor unit for measurement, non-ionized substances can be measured with high sensitivity and selectivity. This makes it possible to detect sweet substances with the same sensitivity as other basic taste substances, and solve the problems of conventional taste sensors. Therefore, according to such a taste sensor device, it is possible to objectively evaluate the taste with quality and strength as close as possible to the sensitivity of human taste, and to accurately reproduce the taste.

  The best mode for carrying out each invention described above will be described below with reference to the drawings. However, the present invention is not limited to these embodiments, and various modifications can be made without departing from the scope of the invention. Can be implemented in a manner.

  The first embodiment mainly relates to claims 1 to 2 and 6 to 9. The second embodiment mainly relates to claims 3 to 9. Furthermore, the third embodiment mainly relates to claim 10. The fourth embodiment mainly relates to claims 11 and 12.

  << Embodiment 1 >>

  <Embodiment 1: Overview>

  The present embodiment relates to a pseudo-biological lipid polymer membrane. The pseudo-biological lipid polymer membrane is based on a membrane main body portion and a modifying portion made of a polymer material and its plasticizer. The pseudo-biological lipid polymer membrane can generate a membrane potential that is highly sensitive and selective to the membrane body with respect to non-ionizing substances. As a result, it is possible to selectively measure sweet substances and the like that could be measured only with extremely low sensitivity with the conventional taste sensor device with high sensitivity.

  <Embodiment 1: Configuration>

  FIG. 1 shows a configuration example of this embodiment. As shown in this figure, the pseudo-biological lipid polymer membrane (0100) of this embodiment is composed of a membrane body portion (0101) and a modification portion (0102) covering the membrane body portion, and the membrane body portion is made of a polymer material. And a plasticizer. FIG. 1 shows a case where the modifying part is formed on one side, and the surface indicated by the arrow A is in contact with the solution to be measured. Below, each component of this embodiment is demonstrated.

  The “pseudo-biological lipid polymer membrane” (0100) is an artificial membrane having a function imitating the electrical property found in a biological cell membrane, and generates a membrane potential with a substance to be measured. It is configured so that the electric charges generated with the conductor can be exchanged. As shown in Example 6 described later, the pseudo-biological lipid polymer membrane of the present invention can generate a membrane potential higher than that of the conventional lipid polymer membrane described in the background art for non-ionized substances. . Further, as shown in Example 7, when the membrane potential was measured for each of the five basic taste substances, the pseudo-biological lipid polymer membrane of the present invention shows a membrane potential pattern specific to the sweet substance. That is, the pseudo-biological lipid polymer membrane of the present invention can be used as a receptor membrane capable of selectively measuring non-ionizing substances.

  The name of the pseudo-biological lipid polymer membrane is based on imitating the function of the cell membrane of the living body. The lipid mentioned here is only derived from the fact that the cell membrane is mainly composed of protein and lipid, which are macromolecules, and the pseudo-biological lipid polymer membrane does not necessarily need to contain lipid as a component. Absent. There may be a case where the membrane body portion of this embodiment described later also functions as a lipid, as will be described later.

  ((Membrane body))

    (Configuration of membrane body)

  The “membrane main body” (0101) of the present embodiment forms the main body of the pseudo-biological lipid polymer membrane, and is composed of a polymer material and a plasticizer thereof. The membrane main body is configured to be able to adsorb the polyhydric phenol of the modifying part described later on its surface. Although the mechanism of adsorption of the polyhydric phenol to the membrane main body has not been elucidated, it is considered that both are connected through an electrical bond such as a hydrophobic bond and / or a hydrogen bond. The grounds are based on the properties of tannin, which is one of polyhydric phenols, and the results of Example 2 and the like described later. Tannin is known to have a converging action by binding to a protein and aggregating a basic functional group, but the binding of both is performed by a hydrophobic bond and a hydrogen bond. From the results of Example 2, it has been found that the balance of charge density on the film surface and the hydrophobic region due to the components of the film main body are important. Therefore, it is preferable that the membrane main body has a site having a high affinity for the hydrophobic site of the polyhydric phenol on the surface thereof, or a site that easily binds to a hydroxyl group of the polyhydric phenol. That is, at least one of the polymer material or the plasticizer described below only needs to have the above-mentioned properties on the surface when the film body is configured.

  The “polymer material” is configured to function as a support member for the entire membrane.

  As the type of the polymer material, a known polymer material used for a liquid membrane ion selective electrode or the like may be used as long as no charge is required as described later. Specifically, PVC, polyvinylidene chloride (PVDC), polystyrene (PS), polyurethane (PU), polysulfone (PSF), polycarbonate (PC), polyarylate (PAR), polyamide (PA), polyvinyl alcohol (PVA) And the like, natural resins such as lacquer, polysaccharide polymers such as agar, gelled proteins such as gelatin (collagen), and mixtures thereof. In particular, PVC is advantageous in that it is easily available and softening and curing can be easily adjusted by the mixing ratio with a plasticizer described later. Therefore, PVC is a polymer material that is most commonly used for liquid membrane ion-selective electrodes and the like, and is also preferable as the polymer material.

  When the plasticizer has a charge, the polymer material may have either a positive or negative charge corresponding to the charge of the plasticizer. This is because the balance of the charge density on the surface of the film main body is taken into consideration. Charge retention in the polymer material can be achieved by using a charged polymer. The “charged polymer” is a polymer such as plastic having a positive or negative charge. For example, the positively charged polymer includes (poly) vinylamine, and the negatively charged polymer includes (poly) styrene sulfonic acid.

  The “plasticizer” is a substance that can plasticize the polymer material and give it softness. The softness of the polymer material is determined by the amount of plasticizer added. For example, if the plasticizer is an ester compound described later, the molecule has a structure having a polar region and a nonpolar region. Specifically, in the case of DOPP shown in FIG. 2A, the polar region of 0201 is electrically connected to the molecules of the polymer film material, and the nonpolar region of 0202 is between the molecules of the polymer film material. Block access. As a result, the distance between the molecules of the polymer material is kept expanded, so that the softness is maintained.

  The type of plasticizer may be a material that can plasticize it according to the polymer material to be used. When the polymer material is the above-mentioned plastic, generally an ester compound or an ether compound is frequently used. Specifically, DOPP, 2-nitrophenyl octyl ether (POE), 4-nitrophenyl phenyl ether (NPPE), dioctyl phosphate (DOP), dioctyl sebacate (DOS), dibutyl sebacate (DBS), phosphoric acid Examples include tricresyl (TCP) or a combination thereof. Which plasticizer is used may be appropriately selected according to compatibility with the polymer film material, plasticization efficiency, low migration, low volatility, and the like. If the polymer material is a polymer polysaccharide or gelled protein, the plasticizer may be water.

  By the way, none of the plasticizers mentioned as specific examples has a charge. However, when actually used for producing the membrane body, most membrane bodies have a charge. This is considered to be because impurities such as a derivative having a charge are mixed in the plasticizer commercially available. For example, the commercially available DOPP shown by A in FIG. 2 is mixed with 0.6 to 2.5% of phenylphosphonic acid monooctyl ester (PPME), which is a negatively charged derivative shown in B of FIG. (Non-patent document 2). In the production of a general plasticizer, the inclusion of such impurities is inevitable, and it is difficult to purify it purely. Therefore, when a commercially available non-ionizing plasticizer is used, the plasticizer usually has an electric charge due to mixed impurities. Therefore, in the case of this embodiment, it is preferable to use a polymer material having a charge opposite to that of the plasticizer in order to obtain a balance of charge density on the film surface.

    (Function of membrane body)

  The membrane body functions as a body that supports the pseudo-biological lipid polymer membrane and as a scaffold for the polyhydric phenol that constitutes the modification. Furthermore, it functions to acquire a membrane potential generated in response to a non-ionized substance trapped on the membrane surface side on the membrane back surface side as well as lipids of the cell membrane.

    (Production of membrane body)

  The membrane body is prepared by mixing a polymer material and a plasticizer. The method for producing the membrane body may be similar to the method for producing a lipid polymer membrane used in a conventional membrane potential measuring device. For example, the method for producing a lipid polymer membrane described in Patent Document 1 is helpful. However, in this embodiment, no lipid substance is added to the constituent elements of the membrane main body. As an example, the most common method is described below. First, a predetermined amount of a polymer material and a plasticizer are mixed. Since the mixing ratio and the mixing method of the polymer material and the plasticizer differ depending on each type, the mixing ratio and the mixing method may be appropriately adjusted depending on the intended use and usage of the pseudo-biological lipid polymer membrane. For example, when a pseudo-biological lipid polymer membrane is used for the working electrode and the polymer material is polyvinyl chloride and the plasticizer is di-n-octylphenylphosphonate, each is 0.8: What is necessary is just to mix at about 1. Next, the polymer material and the plasticizer are dissolved with a solvent. The type of the solvent is appropriately selected according to the polymer material to be used and the type of the plasticizer. For example, when the polymer material is plastic, tetrahydrofuran (hereinafter referred to as THF), nitrobenzene, cyclohexanone, etc. are generally used. What is necessary is just to adjust the capacity | capacitance of the solvent to add according to the kind of polymer material and a plasticizer, and the weight of a mixture. For example, if the weight of the mixture of polyvinyl chloride and di-n-octylphenylphosphonate is 1800 mg, it may be dissolved in about 10 ml of THF. Subsequently, a film is produced from the liquid mixture. As a method for producing the film, for example, a liquid mixture is spread in a thin layer on a flat plate such as a glass plate, and the solvent is evaporated to resolidify. The thickness of the film is preferably in the range of 10 to 300 μm for functions such as efficient generation of the membrane potential.

  ((Modifier))

    (Configuration of modifier)

  The “modifying part” (0102) is configured to modify the surface of the membrane main body part and is made of polyhydric phenol. The polyhydric phenol which comprises a modification part is not restricted to a single kind. For example, it may be a mixture of polyhydric phenolic acid and hydrolyzable tannin such as tannic acid described below. Moreover, the compound which the molecule | numerators of the polyhydric phenol polymerized may be mixed.

  The arrangement of the modifying part may be the entire surface of the membrane main body part or a part thereof. However, in the case of a part of the surface, it is arranged at least in a region in contact with the solution to be measured for the function described later.

  "Polyhydric phenol" is a substance widely distributed in the plant kingdom called polyphenol, and is a general term for compounds having a dihydric or higher phenolic hydroxyl group in an aromatic molecule. For example, polyhydric phenolic acid, flavonoid, tannin and the like are applicable. As the polyhydric phenol of the present invention, polyhydric phenolic acid, hydrolyzed tannin, or a mixture thereof is preferable. This is because these substances are considered to have high affinity with non-ionizing substances such as sugar.

  "Polyhydric phenol" is a polyhydric phenol having a carboxyl group in the molecule, and precisely means a phenolic acid having two or more hydroxyl groups. For example, gallic acid (gallic acid: 3,4,5-trihydroxybenzoic acid) indicated by A in FIG. 3, HHDP (hexahydroxyphenic acid) indicated by A in FIG. 4, which is a dimer of gallic acid, and the like are applicable. In addition, HHDP is lactonized in a free state, and usually immediately becomes ellagic acid shown in FIG. 4B.

  “Hydrolyzed tannin” refers to a tannin that is a compound in which a polyhydric phenol acid and a polyhydric alcohol are ester-bonded and is hydrolyzed by an acid, an alkali, an enzyme, or the like. Specifically, gallotannin, which is a derivative of gallic acid, ellagitannin, which is a derivative of ellagic acid / HHDP, and the like are applicable. As an example of gallotannin, PGG (β-1,2,3,4,6-pentagalloyl-OD-Glucose) indicated by B in FIG. 3 is used, and as an example of ellagitannin, casalictin ( CASARICTIN).

  In the present invention, the “non-ionizing substance” means a substance that is soluble in a polar solvent such as water or ethanol and does not ionize. For example, caffeine, capsaicin, sweet substances and the like are applicable.

The term “sweet substance” as used in the present invention means a natural substance that generally makes a human feel sweet. For example, sugar and stevioside are applicable. However, the sweet substance of the present invention does not necessarily require humans to feel sweetness. Any substance of the same genus that is structurally similar to the substance that makes the sweet taste feel may be used without making the sweet taste. The “sugar” as used herein refers to a relatively low molecular weight polyhydric alcohol obtained by reducing the aldehyde group or keto group of a carbohydrate or sugar represented by the general formula of C _m (H ₂ O) _n. Say things. Specific examples include monosaccharides or oligosaccharides, glycerol, xylitol, sorbitol, mannitol, arabinitol, and the like.

    (Function of modifier)

  The modifying part has a central function in the present invention of interacting with a non-ionizing substance on the film surface. As shown in Example 4 to be described later, the membrane body portion having a modified portion made of a polyhydric phenol on the surface has a drastically increased potential responsiveness to sucrose compared to the case of only the membrane body portion. In addition, as shown in Example 6, the pseudo-biological lipid polymer membrane having the modified portion of the present invention has about four times the sensitivity of a lipid polymer membrane that has been the most sensitive in terms of potential response to sucrose. . Furthermore, as shown in Example 7, it exhibits selective responsiveness to non-ionizing substances such as sweet substances. In this way, by adding a modification part consisting of polyhydric phenol to the membrane body surface, high response sensitivity and selectivity for non-ionizing substances, which was difficult with conventional artificial lipid polymer membranes, can be achieved. Can be generated.

  Here, the chemical acceptance theory of sweet substances will be explained. As a geometric model of a complex composed of a sweetening group (Glucophore) and a sweet substance receptor, models of Non-Patent Document 3 by Schallenberger and Acree and Non-Patent Document 4 by Kier are known. As shown in FIG. 13, they require three underlying sites that are complementary to each other for the interaction of the sweet substance and its receptor. That is, three of a hydroxyl bond donor site (AH: proton donor group) and a hydrogen bond acceptor site (B: proton acceptor group) and a hydrophobic bond site (X) (not shown) that are separated by a certain distance in the molecule. It is. Between the sweet substance and its receptor, it is considered that the AH-B unit of the sweet substance is bound to the AH-B unit on the receptor side, and both X sites are bound by a hydrophobic bond. .

  There is no experimental evidence that the polyphenol constituting the modifying part of the present invention captures a non-ionizing substance by the same mechanism as in the chemoreceptive theory. Therefore, it is unclear how the non-ionized substance is actually captured. However, the fact that non-ionizing substances will be adsorbed to the polyhydric phenol on the membrane surface by some binding force means that the pseudo-biological lipid polymer used for the measurement as described below “Pretreatment for reuse of modified part” It is also inferred from the fact that the membrane does not return to the original potential value unless it is sufficiently washed. In addition, the fact that hydrolyzable catechins such as PGG have sugars such as glucose arranged at the center of their molecular structure indicates that polyphenols have a high affinity for non-ionizing substances such as sugars. It seems to suggest.

(Production of modified part)
The modifying part can be generated on the surface of the membrane body by immersing the membrane body in a solution containing polyhydric phenol. “Immerse” as used herein is not limited to immersing the membrane main body in the mixed liquid, but pouring the mixed liquid over the surface of the membrane main body or applying the mixed liquid using a brush or the like. It means that the mixed liquid is brought into contact with the surface of the membrane main body widely. The composition of the mixed solution may be appropriately selected according to the response sensitivity to the non-ionized substance, the composition of the film main body, and the like, and is not particularly limited. For example, a mixed solution of 0.05% (W / V) tannic acid / 0.3 mM tartaric acid / 30 mM KCl (hereinafter referred to as a tannic acid mixed solution) may be used. Tannic acid is a mixture mainly composed of gallotannin obtained from Urushi family Rhus, Beech family Quercus, etc., and includes gallic acid, PGG and the like. As an example of the production method of the modifying portion, the membrane body may be immersed in the tannic acid mixed solution at 25 ° C. for about 48 hours. Thereby, a modification part can be formed in the surface of a film | membrane main-body part.

(Processing before reuse of modifier)
The modified part after the measurement is considered to be in a state in which a part of the non-ionized substance remains adsorbed on the surface. This can also be inferred from the fact that when the reference liquid is measured again after the pseudo-biological lipid polymer membrane used for the measurement with the measurement object solution is lightly washed, the original reference liquid measurement value is not obtained. Therefore, it is necessary to perform pretreatment when reusing the pseudo-biological lipid polymer membrane. In the case of a membrane having a positive charge, the pretreatment should be sufficiently washed with a 100 mM KCl / 10 mM KOH / 30% ethanol mixed solution, and in the case of a membrane having a negative charge, with a 100 mM HCl / 30% ethanol mixed solution. . By this operation, the modifying part is reset, and the pseudo-biological lipid polymer membrane can be measured a plurality of times, so that the running cost can be suppressed.

  <Embodiment 1: Effect>

  According to the pseudo-biological lipid polymer membrane of the present embodiment, a non-ionizing substance such as a sweet substance can be captured on the surface of the substance and can respond to the substance with high sensitivity and selectivity. Therefore, electrochemical changes (membrane potential measurement, impedance measurement, surface polarization control method, etc.), refractive index (dielectric constant) change (surface plasmon resonance method, etc.), mass change (quartz crystal oscillator, surface acoustic wave) Measurement, etc.) If used in a sensor unit for measurement, non-ionized substances can be selectively measured with high sensitivity.

  << Embodiment 2 >>

  <Embodiment 2: Overview>

  The present embodiment relates to a pseudo-biological lipid polymer membrane. The basic configuration is the same as that of the pseudo-biological lipid polymer membrane of the first embodiment, but the first embodiment is different from the first embodiment in that the membrane body includes a lipid substance in addition to the polymer material and the plasticizer. Different. Since the lipid substance has a charge with a hydrophobic site, it has a function of facilitating adsorption of polyhydric phenol to the membrane body and transfer of charge through the membrane body. As a result, it is possible to obtain a stronger membrane potential response to non-ionizing substances.

  <Embodiment 2: Configuration>

  FIG. 5 shows a configuration example of this embodiment. As shown in this figure, the basic configuration of the pseudo-biological lipid polymer membrane (0500) of this embodiment is composed of a membrane body (0501) and a modification (0502) as in the first embodiment. However, in addition to the components of the membrane main body of Embodiment 1, the membrane main body of the present embodiment further has a lipid substance (0503). For each component, the description of the same parts as in Embodiment 1 is omitted, and here, the characteristic points of the present embodiment and the lipid substance that is a component unique to the present embodiment will be described below. To do.

  The “membrane main body” (0501) of the present embodiment mainly includes a polymer material, a plasticizer, and a lipid substance. The constituent elements of the membrane body of this embodiment are the same as those of the first embodiment. However, since the lipid substance has a positive or negative charge, the polymer material does not necessarily have a charge. A lipid substance having a charge corresponding to the plasticizer described in Embodiment 1 (more precisely, impurities contained therein) may be used.

  The “lipidic substance” (0503) of the present embodiment refers to an amphipathic lipid or an amphiphilic substance having a similar structure. In either case, the molecular structure has a polar hydrophilic portion and a hydrophobic portion composed of a long carbon chain. Examples of the hydrophilic site include a phosphate group, an amino group, a carboxyl group, and a hydroxyl group, as well as ionic ammonium. The type of lipidic substance to be used is not particularly limited, but it is preferable that the lipidic substance exposed on the membrane surface when adsorbing the membrane main body has a property of adsorbing polyhydric phenol. For example, as described above, the molecular structure has a region having a charge such as a hydroxyl group and / or a hydrophobic site. Specific examples include TDAB shown in FIG. 6 and quaternary ammonium which is a derivative thereof. As shown in Example 3 to be described later, TDAB is very preferable as a lipid substance of the present embodiment because it has a very high membrane potential responsiveness to sugar.

  <Embodiment 2: Effect>

  According to the pseudo-biological lipid polymer membrane of this embodiment, a higher membrane potential can be obtained for non-ionizing substances such as sweet substances by adding a lipid substance to the membrane body.

  << Embodiment 3 >>

<Embodiment 3: Overview>
This embodiment relates to a working electrode. The working electrode has the pseudo-biological lipid polymer membrane of Embodiment 1 or 2 as a non-ionizing substance receiving membrane, and generates a membrane potential selectively with high sensitivity to the non-ionizing substance, and Can get it. As a result, it is possible to provide a membrane electron measuring apparatus that measures a non-ionizing substance such as a sweet substance with high potential in terms of potential.

<Embodiment 3: Configuration>
FIG. 7 shows a configuration example of this embodiment. As shown in this figure, the working electrode (0700) of the present embodiment includes the support (0701), the pseudo-biological lipid polymer membrane (0702) of the first or second embodiment, and the conductor (0703) as essential components. ). A buffer layer (0704) is selectively provided. Among these, since the pseudo-biological lipid polymer membrane (0702) has been described in detail in the first or second embodiment, the description thereof will be omitted. Here, a support (characteristic component of the present embodiment) 0701), the conductor (0703), and the buffer layer (0704) will be described below.

  ((Support))

    (Structure of support)

  The “support” (0701) is a base of the working electrode and is configured to dispose a pseudo-biological lipid polymer membrane on a part thereof. Therefore, the shape of the support determines the main shape of the working electrode. The shape of the support is not particularly limited as long as it can acquire the membrane potential generated in the pseudo-biological lipid polymer membrane in response to the non-ionizing substance. What is necessary is just to determine suitably according to the use application etc. of the sensor part which has a working electrode and a reference electrode in a membrane potential measuring apparatus. For example, in the case of a working electrode used in a desktop sensor unit, it may be a closed tubular support (0701) as shown in FIG. 7, or a working electrode in a chip type sensor as shown in FIG. In the case of (0802), it may be a plate-shaped support (0801).

  The material of the support is not particularly limited as long as it is an insulator and is water resistant. Specific examples include glass, plastic, ceramics, synthetic rubber, water-resistant paper, wood, and the like. However, as long as the outermost part is covered with a water-resistant insulating material, the support may have a conductive material inside. For example, a multilayer substrate having a laminated structure composed of an insulator material and a conductor material and having the outermost portion coated with a water-resistant insulator material is applicable.

  The position of the pseudo-biological lipid polymer membrane on the support is not particularly limited as long as it is disposed at a position where it can directly contact the solution to be measured.

  ((conductor))

    (Construction of conductor)

  The “conductor” (0703) is disposed in contact with the solution to be measured on the back side of the pseudo-biological lipid polymer membrane, and acquires a membrane potential generated in response to a non-ionizing substance on the front side. It is configured as follows. The term “front side / back side” as used herein is not limited to the front / back relationship of a planar substance, but means the single side and the other side of a substance as wide as the outer side and inner side of a hollow substance.

  The material of the conductor may be any of an electronic conductor, an ionic conductor, or an electron / ion mixed conductor. In consideration of processing points, an electronic conductor is preferable. Specific examples of the electronic conductor include metals such as aluminum (Al), chromium (Cr), copper (Cu), silver (hereinafter referred to as Ag), platinum (Pt), gold (Au), and carbon (C). Etc. Above all, Ag is used as the most common wiring material in the working electrode of the liquid membrane type ion selective electrode device, and is preferable as the material of the conductor because its performance has been confirmed and is easily available. Further, Ag / AgCl in which silver chloride (hereinafter referred to as AgCl) is generated or adhered on the Ag surface is particularly preferable because it can suppress a polarization phenomenon that occurs in a conductor made of only Ag.

  The shape and size of the conductor may be appropriately determined according to the shape of the working electrode and the shape and size of the electrode part and the pseudo-biological lipid polymer membrane, and is not particularly limited. For example, it may be rod-like like the conductor (0703) shown in FIG. 7 and larger than the pseudo-biological lipid polymer membrane (0702), or like the conductor (0804) shown in FIG. It may be planar and substantially the same size as the pseudo-biological lipid polymer membrane (0803).

    (Function of conductor)

  In the membrane potential measuring apparatus according to the fourth embodiment, which will be described later, this conductor is present at the terminal portion of the apparatus wiring. It has a function to transmit to.

  ((Buffer layer))

    (Composition of buffer layer)

  The “buffer layer” (0704) is a layer existing between the pseudo-biological lipid polymer membrane and the conductor. The buffer layer is not an essential component. What is necessary is just to select suitably as needed depending on the structure of a working electrode. For example, when the pseudo-biological lipid polymer membrane (0803) and the conductor (0804) are in direct contact like the working electrode (0802) shown in FIG. 8, the buffer layer is not necessary.

  The buffer layer is composed of an electrolytic solution containing an electrolyte having a buffering action, or a wetting material that holds the electrolytic solution and has a structure as a matrix such as a polymer. Here, the “electrolytic solution” is a liquid having conductivity by dissolving an electrolyte in a solvent such as water.

  The type of electrolyte constituting the electrolytic solution is not particularly limited as long as it has a buffering action. A potassium chloride (hereinafter referred to as KCl) solution is preferable. This is because the transport number of the cation (K ion) and the anion (Cl ion) constituting KCl is almost equal, so that it is difficult to generate a local battery in the electrolytic solution or the wetting material. Moreover, when a part of said conductor is comprised with a metal, it is more preferable that it is a mixed solution of KCl and the metal salt of the said metal. For example, the electrolyte solution when the conductor is composed of Ag / AgCl is preferably a mixed solution of KCl and AgCl. This is to prevent the electrode from being consumed due to the ionization of Ag on the Ag / AgCl surface and to extend the life. The concentration of the electrolyte in the electrolytic solution is not particularly limited. What is necessary is just to adjust suitably according to the solubility of electrolyte, or the material of the wetting material holding the said electrolyte solution.

(Function of buffer layer)
The function of the buffer layer is to keep the ion concentration around the conductor constant. Furthermore, by suppressing fluctuations in the potential between the conductor and the pseudo-biological lipid polymer membrane that occur when ions present in the solution used for measurement reach the conductor via the pseudo-biological lipid polymer membrane. is there.

  <Embodiment 3: Function>

  The function of the working electrode of this embodiment is the same as that of a conventional liquid membrane ion selective working electrode. In other words, the receptor membrane responds to the target substance to be measured in the solution, and the generated membrane potential is acquired and transmitted to other parts. In particular, the working electrode of this embodiment has a function of selectively acquiring and transmitting a membrane potential generated in response to a non-ionizing substance.

  <Embodiment 3: Effect>

  According to the working electrode of this embodiment, it is possible to provide a working electrode that can selectively and highly sensitively measure a membrane potential generated in response to a non-ionizing substance such as a sweet substance.

  << Embodiment 4 >>

<Embodiment 4: Overview>
The present embodiment relates to a membrane potential measuring apparatus and a measuring method thereof. FIG. 9 shows an example of the membrane potential measuring apparatus of this embodiment. As shown in this figure, this apparatus acquires and analyzes a sensor part (0903) composed of a reference electrode (0901) and a working electrode (0902), and a membrane potential generated by bringing each electrode into contact with a measurement target solution. Analysis device (0904). Therefore, the basic configuration is the same as that of the conventional membrane potential device except that the working electrode (0905) of Embodiment 3 is provided.

  However, the apparatus of this embodiment has the working electrode of Embodiment 3 so that it has high sensitivity and selectivity for all five basic taste substances (sweet substance, sour substance, salty substance, bitter substance, umami substance). It is possible to provide a taste sensor device capable of measuring by the above.

<Embodiment 4: Configuration>
As shown in FIG. 9, a characteristic point in the configuration of the membrane potential measuring apparatus according to the present embodiment is that the apparatus has the working electrode (0905) according to the third embodiment. Other configurations are based on the configuration of a known membrane potential measuring device. For example, the apparatus comprised by the measurement circuit shown in FIG. 10 based on the taste sensor described in patent document 1 to 6 grade | etc., May be sufficient. The measurement circuit shown in FIG. 10 has a reference electrode (1001), a working electrode (1002), a differential amplifier (1003), an A / D converter (1004), and a computer (PC, etc .: 1005) as basic components. . FIG. 10 shows a case where a plurality of working electrodes (multi-channel) are provided, but at least one of them is composed of the working electrode of the third embodiment. Therefore, the description of the same configuration as that of the known technique is omitted, and only the configuration characteristic to the present embodiment will be described below.

  The apparatus is characterized in that the sensor unit (0903) includes the working electrode (0905) of the third embodiment. It is sufficient that at least one working electrode is included in one membrane potential measuring device. Further, when one apparatus has a plurality of working electrodes as shown in FIG. 9, the other working electrode can be an electrode having a response sensitivity different from that of the working electrode of the third embodiment. For example, if the membrane potential measuring device of the present embodiment is a taste sensor device, the other working electrode may be a working electrode that exhibits selective response sensitivity with respect to basic taste substances other than sweet substances.

  The configuration requirement of the present embodiment is that the working electrode of the third embodiment is included, and the potential value acquired from the electrode can be measured and analyzed. Therefore, the basic components of the membrane potential measuring device itself may be the same as those of the conventional membrane potential measuring device. For example, the structure which adds the working electrode of Embodiment 3 to a commercially available taste sensor apparatus may be sufficient.

  <Embodiment 4: Measurement method>

  Methods for measuring non-ionized substances using the working electrode of Embodiment 3 can be broadly divided into the following A and B.

    A. When standard solution treatment is not performed

FIG. 11 shows an example of a processing flow when each electrode is not previously treated with a reference solution to be described later when the non-ionized substance is measured. The measuring method shown in this figure is performed based on the membrane potential measuring apparatus of this embodiment. Hereinafter, measurement of non-ionized substances when the reference solution treatment is not performed will be described along the flow of the treatment.
(1) Measurement object solution contact step (S1101): A reference electrode and the working electrode described in the third embodiment are brought into contact with a measurement object solution containing a non-ionizing substance. The contact may be performed, for example, by immersing the electrode in a solution.
(2) Reference electrode potential value acquisition step (S1102): In S1101, a potential value generated from the reference electrode is acquired. The potential value acquired in this step is defined as Vc.
(3) Working electrode potential value acquisition step (S1103): In S1101, a potential value corresponding to the amount of non-ionizing substance in the measurement target solution generated from the working electrode is acquired. The potential value acquired in this step is defined as Vs. If the acquired potential value is low, the potential value may be amplified as necessary. For amplification, a high input impedance amplifier or the like may be used. Note that the order of processing in S1102 and this step is not limited. Both processes may be performed simultaneously.
(4) Difference potential value acquisition step (S1104): A difference between potential values generated in S1102 and S1103, that is, Vs−Vc is acquired.
(5) Differential potential value output step (S1105): The differential potential value Vs−Vc obtained in S1104 is output. The output pattern of the difference potential value may be appropriately selected according to the purpose. For example, the difference potential value may be output as it is, or may be output as a graph in which the difference potential value for each concentration obtained from different concentrations of the same non-ionized substance is plotted. Examples of the output destination include a monitor, a printer, and a speaker. The membrane potential generated for non-ionized substances can be measured by the above processing flow.

    B. When performing standard solution processing

FIG. 12 shows an example of the flow of processing when processing with a reference solution when measuring non-ionizing substances. In this method, a potential value obtained by subtracting the reference potential difference value obtained from the reference solution from the potential difference value obtained from the measurement target solution is set as a target difference potential value. Therefore, it is generally possible to measure with higher accuracy than A. The measurement method shown in FIG. 12 is performed based on the membrane potential measurement apparatus of this embodiment. Hereinafter, measurement of non-ionized substances in the case of performing the reference solution treatment will be described along the flow of the treatment.
(1) Standard solution contact step (S1201): The reference electrode and the working electrode described in Embodiment 3 are brought into contact with the standard solution. The reference solution is not particularly limited as long as it is a solution that stably maintains the state of each electrode. A mixed solution of 30 mM KCl / 0.3 mM tartaric acid is preferred.
(2) Reference electrode reference potential value acquisition step (S1202): The reference electrode reference potential value generated from the reference electrode in S1201 is acquired. The reference potential value acquired in this step is defined as bVc.
(3) Working electrode reference potential value acquisition step (S1203): The reference potential value generated from the working electrode in S1201 is acquired. The working electrode reference potential value acquired in this step is defined as bVs. The order of processing in S1202 and this process is not limited. Both processes may be performed simultaneously.
(4) Measurement object solution contact step (S1204): Similar to S1101, the reference electrode and the working electrode described in Embodiment 3 are brought into contact with the measurement object solution containing a non-ionizing substance.
(5) Reference electrode potential value acquisition step (S1205): Similar to S1102, the measured potential value generated from the reference electrode in S1204 is acquired. The measured potential value acquired in this step is defined as Vc.
(6) Working electrode potential value acquisition step (S1206): The measured potential value generated from the working electrode in S1204 is acquired as in S1103. The measured potential value acquired in this step is defined as Vs. The order of processing in S1205 and this step is not limited. Both processes may be performed simultaneously. When the acquired potential value is low, the potential value may be amplified as necessary.
(7) Reference difference potential value acquisition step (S1207): The difference between the reference potential values generated in S1202 and S1203, that is, bVs−bVc is acquired.
(8) Measurement difference potential value acquisition step (S1208): The difference between the measurement potential values generated in S1205 and S1206, that is, Vs−Vc is acquired.
(9) Target difference potential value acquisition step (S1209): difference between both potential values of the reference difference potential value acquired in S1207 and the measured difference potential value acquired in S1208 (Vs−Vc) − (bVs−bVc) Is obtained as a target differential potential value. According to this measurement method, the potential change based on the adsorption of the non-ionizing substance to the pseudo-biological lipid polymer membrane can be continuously measured. The measurement method will be described below.
(10) Washing step (S1210): After obtaining the potential value in S1205 and S1206, the solution to be measured attached to each electrode is gently washed using the reference solution.
(11) Standard solution recontacting step (S1211): The cleaned reference electrode and working electrode are brought into contact with the standard solution again.
(12) Reference electrode re-reference potential value acquisition step (S1212): The reference potential value obtained at the reference electrode in S1211 is acquired. The re-reference potential value acquired in this step is defined as bVc ′.
(13) Working electrode age reference potential value acquisition step (S1213): The reference potential value obtained at the working electrode in S1211 is acquired. The re-reference potential value acquired in this step is defined as bVs ′. The order of processing in S1212 and this step is not limited. You can go at the same time.
(14) Re-reference difference potential value acquisition step (S1214): A difference between re-reference potential values generated in S1212 and S1213, that is, bVs′−bVc ′ is acquired.
(15) Re-difference potential value acquisition step (S1215): difference between both potential values of the reference difference potential value acquired in S1207 and the re-reference difference potential value acquired in S1214 (bVs′−bVc ′) − (bVs) -BVc) is acquired as the re-difference potential value. Here, if the non-ionizing substance is not adsorbed on the pseudo-biological lipid polymer membrane, the re-difference potential value is 0. When the non-ionizing substance is a sweet substance, the aftertaste remaining on the tongue after eating the sweet substance can be measured by acquiring the re-differential potential value.
(16) Differential potential value output step (S1216): target differential potential value (Vs−Vc) − (bVs−bVc) obtained in S1209, redifferential potential value (bVs′−bVc ′) − (obtained in S1215) bVs−bVc) and the like are output. The output pattern of the difference potential value may be appropriately selected according to the purpose.

  The above processing can be executed by a program for causing a computer to execute. Moreover, this program can be recorded on a recording medium readable by a computer.

<Embodiment 4: Effect>
According to the membrane potential measuring apparatus of the present embodiment, it is possible to measure a non-ionized substance with high potential sensitivity. In the conventional taste sensor device, the remarkably low response sensitivity to sweet substances is the biggest problem. However, according to the membrane potential measuring apparatus of the present embodiment, the above problem can be solved by having the working electrode that responds to each of the other basic taste substances. That is, it becomes possible to selectively measure all of the basic tastes with high sensitivity. In addition, it is possible to provide a taste sensor device that is as close as possible to the sensitivity of human taste, such as enabling accurate measurement of synergistic effects and suppression effects of each taste substance. In addition, by using the apparatus, it is possible to reproduce the taste with high accuracy, create a taste database, and the like.

  << Example >>

  The present invention will be specifically described with reference to the following Examples 1 to 7. However, the following examples are merely illustrative, and the present invention is not limited to these examples.

<Preparation of pseudo-biological lipid polymer membrane>
Specific examples of the method for producing the pseudo-biological lipid polymer membrane of the present invention are given below. Here, the pseudo-biological lipid polymer membrane of Embodiment 2 will be described. Note that the description of the modified part of the pseudo-biological lipid polymer membrane is partly the same as the description of the production of the working electrode of Example 2 because of the production relationship.

(Membrane body production method)
1. A stir bar was placed in the screw-mouth bottle (SV-20), and then 5 ml of THF (Sigma-Aldrich: the same applies below) was placed with a micropipette. Since THF is a volatile substance, the lid was immediately covered with a fluororesin sheet (100 μm thick) immediately after the operation.
2. After adding 2.5 ml of TDAB (Fluka) as a lipid substance and 1.5 ml of DOPP (Dojin Chemical: containing 0.3% PPME) as a plasticizer, each was stirred for 30 minutes.
3. THF (5 ml) was added (total 10 ml), and the mixture was further stirred for 30 minutes.
4). As a polymer material, 800 mg of PVC (Wako Pure Chemical: the same applies hereinafter) was added and stirred for 1 hour.
5. The mixed solution dissolved in THF was taken out into a petri dish and spread thinly.
6. Drying for 72 hours or more in a fume hood with a petri dish covered. In this step, since the THF is volatilized, the mixed solution is solidified again. The membrane main body was produced by the above process. In addition, the thickness of the film | membrane main-body part obtained by this method was about 200 micrometers.

(Method for producing the modified part)
1. As a tannic acid solution, 35 ml of a mixed solution of 30 mM KCl / 0.3 mM tartaric acid was prepared.
2. After adding 17.5 mg (0.05% W / V) of tannic acid (Kanto Chemical) as a polyhydric phenol, the mixture was stirred with a stir bar for 1 minute. Thereby, a tannic acid mixed solution was prepared.
3. The membrane main body in the petri dish prepared above was cut into 21 sections with a cutter washed with THF or ethanol.
4). The membrane body was attached to the support. The method for attaching the film to the support was described in Example 2.
5. The support on which the one-compartment membrane body part was attached to the tannic acid solution was completely immersed for 48 hours. The membrane body was handled with tweezers washed with THF (or ethanol).
Through the above steps, a pseudo-biological lipid polymer membrane was produced.

Moreover, the following method was also used as a production method of the modified part other than the above.
1. As a tannic acid solution, 35 ml of a mixed solution of 30 mM KCl / 0.3 mM tartaric acid was prepared.
2. After adding 17.5 mg (0.05% W / V) of gallic acid (Tokyo Kasei Kogyo) as polyhydric phenolic acid, the mixture was stirred with a stirrer for 1 minute. Thereby, a tannic acid mixed solution was prepared.
3. The membrane main body in the petri dish prepared above was cut into 21 sections with a cutter washed with THF or ethanol.
4). The membrane body was attached to the support.
5. After being immersed in a 100 mM KCl / 10 mM KOH / 30% ethanol mixed solution for 90 seconds, it was immersed in a mixed solution of 30 mM KCl / 0.3 mM tartaric acid for 270 seconds. Then, after being immersed in a tannic acid mixed solution for 30 seconds, it was again immersed in a mixed solution of 30 mM KCl / 0.3 mM tartaric acid for 40 seconds.
6). This procedure 5 was repeated about 140 times.
Through the above steps, a pseudo-biological lipid polymer membrane was produced.

<Production of working electrode>
A working electrode is produced using the pseudo-biological lipid polymer membrane produced in Example 1.

(Production of adhesive)
1. A stir bar was placed in a screw-mouth bottle (SV-20), and 10 ml of THF was added using a micropipette. Immediately after the operation, a lid was placed with a fluororesin sheet (100 μm thick) in between.
2. 800 mg of PVC was added, and the mixture was stirred with a stirrer at a rotational speed of about 1500 rpm for 1 hour. The solution obtained by the above steps was used as an adhesive for film coating.

(Membrane on support)
1. A fluororesin sheet (500 μm thick) was laid on a non-woven fabric (Bencot: Asahi Kasei Fiber), and the pseudo-biological lipid polymer membrane prepared in Example 1 was placed.
2. Using a pipette, 20 μl of the adhesive was applied to the support where the film was applied.
3. After pressing the film pasting portion of the support to the film on the fluororesin sheet, the whole fluororesin sheet was turned over and pressed further closely from above the sheet so as to remove air.
4). The fluororesin sheet was carefully removed from the support and dried for 1 hour.
5. In order to ensure the adhesion between the support and the membrane body, THF was applied to the gap between the support and the membrane body using a pipette, and then dried for one day.
6). 3.3M KCl / saturated AgCl was placed inside the support as a buffer layer, and an electrode rod made of AgCl / Cl as a conductor was placed inside the support. All of the above operations were performed in a draft with tweezers and wearing gloves. Further, the tweezers were washed with THF or ethanol, and the film handling and the miscellaneous use were separately used. The working electrode was produced through the above steps.

(Precondition processing)
In the measurement using the working electrode, a preconditioning process was performed as a process before actual measurement in order to obtain a stable measurement potential. The treatment is an operation of preliminarily dipping the electrode in a reference solution to adsorb a taste substance to the electrode film and stabilize the measurement potential. Usually, one of the measurement target solutions is used as the reference solution. Here, the prepared working electrode was immersed in the solution for 2 days using 30 mM KCl / 0.3 mM tartaric acid as a reference solution.

<Verification of optimum concentration conditions for non-ionizing substance response sensitivity of membrane body components>

  (Purpose) To verify the concentrations of lipid substances and plasticizers that respond to non-ionizing substances with the highest sensitivity.

(Method) Pseudo-biological lipid polymer membranes having different concentrations of lipid substance and plasticizer were prepared, and the response potential of the solution to be measured for each membrane was measured.
1. As the lipid substance and the plasticizer, TDAB and DOPP were used in the concentration ranges shown below.
Lipid substance: TDAB ... 1.3mg-3.0mg
Plasticizer: DOPP (containing 0.3% PPME) .. 1.0 ml to 1.7 ml
2. Pseudo-biological lipid polymer membranes in which the concentrations of TDAB and DOPP were changed within the above range were prepared. The method for producing the membrane body of each pseudo-biological lipid polymer membrane was performed according to Example 1.
3. The response potential of the solution to be measured for each membrane was measured. A mixed solution of 1M sucrose / 30 mM KCl / 0.3 mM tartaric acid was used as the measurement target solution.

  Note that a commercially available taste sensor (SA402B: Incent Co., Ltd.) was used for each of the membrane potential measuring devices for measuring potentials in and after this example. Therefore, the working electrode of Example 2 (hereinafter referred to as the working electrode of the present invention) was made of the same type as the working electrode support of the sensor. And the working electrode of this invention was arrange | positioned to the said sensor by the method similar to another working electrode, and it used for the measurement by the same operation. The processing of the potential acquired by the electrode was performed in the same manner as the conventional working electrode according to the analysis program installed in the computer of the sensor.

  (Results) The measurement results are shown in FIG. In the figure, the X-axis represents DOPP concentration (ml), the Y-axis represents TDAB concentration, and the Z-axis represents response potential (mV) obtained from the pseudo-biological lipid polymer membrane.

  As shown in this figure, the potential response to sucrose showed a charge peak when DOPP was between 2.25 and 2.5 mg and TDAB was 1.5 ml as indicated by the broken line. That is, the potential responsiveness to sucrose does not increase linearly as the concentration of TDAB or DOPP increases, but positively charged TDAB (lipid substance) and negatively charged DOPP (plasticizer: It was suggested that the balance of the charge density on the film surface of the film (impurities such as PPME that actually have a negative charge) is important. Therefore, the pseudo-biological lipid polymer membrane of Example 1 is produced under the concentration conditions of TDAB and DOPP having high response sensitivity to non-ionizing substances.

<Verification of responsiveness of non-ionized substances in the modified part>

  (Purpose) The function of the modified part composed of polyhydric phenol was verified.

(Method) The responsiveness to the non-ionizing substance of only the membrane main body and the pseudo-biological lipid polymer membrane having the modifying portion was verified.
1. Using only the membrane body of Example 1, a working electrode was produced according to the method of Example 2. This working electrode is referred to as a working electrode C (Control).
2. Using the pseudo-biological lipid polymer membrane having the modified portion of Example 1, a working electrode was produced according to the method of Example 2. This working electrode is referred to as a working electrode T (Test).
3. Each working electrode was attached to taste sensor A402B.
4). Sucrose was used as the non-ionizing substance. The following sucrose solutions were prepared as measurement target solutions.
(A) 1 mM sucrose / 30 mM KCl / 0.3 mM tartaric acid (b) 3 mM sucrose / 30 mM KCl / 0.3 mM tartaric acid (c) 10 mM sucrose / 30 mM KCl / 0.3 mM tartaric acid (d) 30 mM sucrose / 30 mM KCl / 0.3 mM tartaric acid (e) 100 mM sucrose / 30 mM KCl / 0.3 mM tartaric acid (f) 300 mM sucrose / 30 mM KCl / 0.3 mM tartaric acid (g) 1 M sucrose / 30 mM KCl / 0.3 mM tartaric acid The response potentials of the sucrose solutions at the respective concentrations (a) to (g) at each working electrode were measured according to the method of Example 3.

  (Results) The measurement results are shown in FIG. At each concentration, the potential value obtained at the working electrode C was a black square, the potential value obtained at the working electrode T was spotted as a black circle, and an approximate line based on them was marked.

  As shown in this figure, with the working electrode C, almost no response was observed within the above measurement concentration range, and a response potential was obtained to some extent in the measurement of the 1M sucrose aqueous solution. On the other hand, a response potential was obtained from 30 mV with the working electrode T, and a particularly significant response potential was confirmed at a concentration of 100 mM or more. From the above results, it was proved that the modified portion of the present invention performs an important function in responding to the non-ionized substance by the pseudo-biological lipid polymer membrane.

<Verification of responsiveness of pseudo-biological lipid polymer to non-ionizing substances>

  (Purpose) We examined the responsiveness of pseudo-biological lipid polymers to non-ionizing substances other than sucrose.

(Method) Using the working electrode T of Example 4, the responsiveness of caffeine, which is a non-ionizing substance other than a sweet substance, was verified. Moreover, it measured also about 1 mM sucrose in which the responsiveness was seen in Example 4 for the comparison. The measuring apparatus and measuring method were the same as in Examples 3 and 4. The solution to be measured was prepared as follows.
(A) 0.1 mM caffeine / 30 mM KCl / 0.3 mM tartaric acid (b) 0.3 mM caffeine / 30 mM KCl / 0.3 mM tartaric acid (c) 1 mM caffeine / 30 mM KCl / 0.3 mM tartaric acid (d) 10 mM Caffeine / 30 mM KCl / 0.3 mM tartaric acid (e) 1M sucrose / 30 mM KCl / 0.3 mM tartaric acid

  (Results) The measurement results are shown in FIG. The measurement was performed five times at each concentration, and the average value is shown by a bar graph. The error bar in the figure indicates the error range of the measurement result.

  As shown in this figure, the response potential of caffeine is 10 mV or less even at 10 mM, which is lower than that of sucrose. However, with conventional lipid polymer membranes, no response potential could be obtained from caffeine even at high concentrations (not shown). In view of the fact, it can be understood that the pseudo-biological lipid polymer of the present invention has higher responsiveness to caffeine than the conventional membrane. This also suggests that the pseudo-biological lipid polymer membrane of the present invention has higher responsiveness to non-ionizing substances other than sweet substances than conventional membranes.

<Verification of response sensitivity to non-ionizing substances>

  (Objective) The response sensitivity of the pseudo-biological lipid polymer membrane of the present invention to non-ionizing substances was examined.

(Method) The sensitivity of non-ionized substances was compared between the lipid polymer film (hereinafter referred to as conventional film) having the highest response to conventional non-ionized substances and the film of the present invention.
1. As the conventional membrane, a lipid polymer membrane composed of PVC, DOPP, 2C16, and TDAB described in Non-Patent Document 1 described in the background art was used. The membrane consists only of the membrane body and has no modifier. Using this membrane, a working electrode was produced according to the method shown in Example 2 above. The working electrode is referred to as a working electrode F (Former).
2. The working electrode T was the same as the electrode prepared in Example 5.
3. Each working electrode was attached to the taste sensor A402B and then measured under the same measurement conditions as in Examples 3-5.

  (Results) FIG. 17 shows the results of this example. At each concentration, the potential value obtained at the working electrode F was a black triangle, the potential value obtained at the working electrode T was spotted as a black circle, and an approximate line based on them was marked. In this example, the measurement was performed five times under the same conditions, and the average value was shown by a plot. The error bar in the figure indicates the error range of the measurement result.

  As shown in this figure, the response potentials were obtained from the working electrodes F and T from a sucrose solution of about 30 mM. However, the subsequent response sensitivity was remarkable at the working electrode T, and a response potential of about 3 times that of the working electrode F was obtained at 300 mM and about 4 times that at 1M. Thus, it has been clarified that the pseudo-biological lipid polymer membrane of the present invention has a higher response sensitivity to non-ionizing substances than the conventional membrane.

  Moreover, the absolute value of the response potential obtained from the sucrose concentration in the range of 300 mM to 1 M at the working electrode T was about 20 mV to about 80 mV. The response potential for each basic taste substance measured with other receptor membranes is usually in the range of 0 mV to 10 mV. In view of this fact, it can be said that the measured value is sufficient as response sensitivity to non-ionized substances.

<Verification of selectivity for non-ionized substances>

  (Objective) It was verified whether the response sensitivity of the pseudo-biological lipid polymer membrane of the present invention has selectivity for non-ionizing substances.

(Method) The composition of the solution to be measured for each basic taste substance used in this example is as follows.
(1) Salty taste: 300 mM KCl / 0.3 mM Tartaric acid (2) Acidity: 30 mM KCl / 3 mM Tartaric acid (3) Umami: 10 mM MSG (Sodium glutamate) / 30 mM KCl / 0.3 mM Tartaric acid (4) Bitter taste: 0.1 mM Quinine hydrochloride / 30 mM KCl / 0.3 mM tartaric acid (5) Sweetness: 1 M sucrose / 30 mM KCl / 0.3 mM tartaric acid The response potential of each measurement target solution was measured with the working electrode T and the working electrode F. In addition, about an assumption apparatus, a measuring method, etc., it applied to the said Examples 3-6.

  (Results) FIG. 18 shows the results of this example. The measurement target solution was measured five times under the same conditions, and the average value was shown as a bar graph. The error bar in the figure is the error range of each measurement result. In the figure, the black bar represents the measured value of the working electrode T, and the white bar represents the measured value of the working electrode F.

  As shown in this figure, when the basic taste is measured with the working electrode F having a conventional membrane, the sour and bitter substances have a response potential of 20 mV to 30 mV, and the umami substance and the sweet substance all have about − It showed 40 mV to -20 mV. Thus, the conventional membrane showed a response sensitivity relatively close to umami substances other than sweet substances. Therefore, even if the conventional membrane shows the responsiveness to the uncharged substance shown in Example 6, it is difficult to say that the conventional membrane has a sufficient function as a receiving film for the uncharged substance because of its low selectivity.

  On the other hand, when the basic taste was measured with the working electrode T having the pseudo-biological lipid polymer membrane of the present invention, a response potential of about −70 mV was obtained from the sweet substance. This can be clearly distinguished from the response potential obtained from any other basic taste substance. Since the response potential to the salty substance is slightly high at about −30 mV, there may be a concern about erroneous measurement due to the salty substance. However, this can be easily solved by using a working electrode having a receptor film for salty substances. This is because a non-ionizing substance can hardly obtain a response potential in a salty substance receiving membrane. Therefore, it was proved that the pseudo-biological lipid polymer membrane of the present invention has high selectivity for non-ionizing substances.

  (Conclusion) From the results of Examples 3 to 7, it was proved that the pseudo-biological lipid polymer membrane of the present invention has high response sensitivity and selectivity for non-ionized substances. Therefore, the pseudo-biological lipid polymer membrane of the present invention, a working electrode using the same, and a membrane potential measuring device having the electrode are useful for measuring non-ionizing substances.

The figure for demonstrating the structure of the pseudo | simulation biological lipid polymer membrane of Embodiment 1. FIG. The figure for demonstrating the structure of plasticizer DOPP and the impurity PPME contained in it Diagram for explaining the structure of gallic acid and PGG Diagram for explaining the structures of HHDP, ellagic acid, and casalictin Enlarged conceptual diagram of the membrane for explaining the configuration of the pseudo-biological lipid polymer membrane of Embodiment 2 Diagram for explaining structure of lipidic substance TDAB Sectional drawing of the working electrode for demonstrating the structure of Embodiment 3. FIG. FIG. 6 is a perspective view for explaining a working electrode in the chip-type sensor according to the third embodiment. The figure for demonstrating the membrane potential apparatus of Embodiment 4. The figure for demonstrating the measurement circuit of a taste sensor Flow chart of measurement method when reference solution treatment is not performed in Embodiment 4 Flow chart of measurement method when performing reference solution treatment in Embodiment 4 Illustration for explaining the chemical acceptance theory of sweet substances The figure for demonstrating the result of Example 3 The figure for demonstrating the result of Example 4 The figure for demonstrating the result of Example 5 The figure for demonstrating the result of Example 6 The figure for demonstrating the result of Example 7

Explanation of symbols

0101, 0501: Membrane body parts 0102, 0502: Modification part 0503: Lipid substances 0700, 0806, 0905, 1006: Working electrodes 0702, 0803 having pseudo-biological lipid polymer membranes: Pseudo-biological lipid polymer membranes 0802, 0902, 1002: Working electrode 0703, 0804: Conductor 0805, 0901, 1001: Reference electrode

Claims (11)

A membrane body formed by mixing a polymer material and its plasticizer;
A modified part generated on the surface of the membrane main body by immersing it in a solution containing polyphenol for about 48 hours ,
A pseudo-biological lipid polymer membrane comprising: A membrane body formed by mixing a polymer material, its plasticizer, and a lipid substance;
A modified part generated on the surface of the membrane main body by immersing it in a solution containing polyphenol for about 48 hours ,
A pseudo-biological lipid polymer membrane comprising: The pseudo-biological lipid polymer membrane according to claim 2 , wherein the lipid substance is tetradodecyl ammonium bromide (TDAB).   The pseudo-biological lipid polymer membrane according to any one of claims 1 to 3, wherein the plasticizer is di-n-octylphenyl phosphate (DOPP) and phenylphosphonic acid monooctyl ester (PPME).   The pseudo-biological lipid polymer membrane according to any one of claims 1 to 4, wherein the polyhydric phenol in the modifying portion is polyhydric phenolic acid and / or hydrolyzable tannin.   The pseudo-biological lipid polymer membrane according to any one of claims 1 to 5, wherein the modifying unit has a function of generating a potential in the membrane body in response to an external non-ionizing substance.   The pseudo-biological lipid polymer membrane according to claim 6, wherein the modifying unit responds to a sweet substance that is a foreign non-ionizing substance.   The pseudo-biological lipid polymer film according to any one of claims 1 to 7, wherein the modifying part is formed on the surface of the membrane main body part by being immersed in a solution containing tannic acid. A support;
The pseudo-biological lipid polymer membrane according to any one of claims 1 to 8,
A conductor for acquiring a membrane potential generated in response to a foreign non-ionizing substance on the surface side of the pseudo-biological lipid polymer membrane from the back side;
Working electrode.   A membrane potential measuring device comprising the working electrode according to claim 9.   A non-ionizing substance measuring method in which a reference electrode and a working electrode according to claim 9 are brought into contact with a measurement target solution, and a difference in potential value generated at each electrode in response to the non-ionizing substance is measured.