Savoriness sensor film, method for producing savoriness sensor film, and method for detecting "savoriness" using savoriness sensor film

Abstract

The present invention provides a savoriness sensor film which makes it possible to evaluate the degree of "savoriness".　The savoriness sensor film is produced by mixing a polymer material (PVC), a lipid (TDAB) having a positive charge, and a plasticizer (DOPP), forming these into the shape of a film, and modifying, on the film surface, a negatively charged lipid, which is an aromatic polycarboxylic acid having at least one hydroxyl group and in which, on a benzene ring, a carboxyl group and a hydroxyl group are adjacent or carboxyl groups are adjacent. In this sensor film, film potential is changed by a flavor substance in an aqueous solution to be measured, and the degree of "savoriness" is detected. In such a savoriness sensor film, one lipid having a negative charge is selected from the group of 2,6-dihydroxyterephthalic acid (2,6-DHTA), 2-hydroxyterephthalic acid (2-HTA), 2,3-dihydroxyterephthalic acid (2,3-DHTA), 2,5-dihydroxyterephthalic acid (2,5-DHTA), 2-hydroxyisophthalic acid (2-HIPA), 4-hydroxyisophthalic acid (4-HIPA), 3-hydroxyphthalic acid (3-HPA), and 4-hydroxyphthalic acid (4-HPA).

Description

Umami sensor film, method for manufacturing umami sensor film, and method for detecting "umami" using umami sensor film

[0001] The embodiments relate to an umami sensor film for evaluating "umami", a method for manufacturing this umami sensor film, and a method for detecting "umami" using this umami sensor film.

[0002] Umami is recognized worldwide by academic societies as the fifth basic taste. In recent years, as Japanese food has become the most popular cuisine in the world, umami as a taste has also become recognized worldwide, and people all over the world are now enjoying umami.

[0003] Meanwhile, taste testing systems have been developed in recent years that can test the taste of foods, beverages, etc. without relying on human sensory testing. These systems use taste sensor membranes (also called taste sensors or lipid membrane sensors) made by mixing a polymeric material, lipids, and plasticizers in a predetermined ratio. This taste sensor membrane changes its membrane potential in response to substances in the test liquid, allowing it to quantify taste and be used for taste evaluation.

[0004] The inventors have already proposed taste testing methods using molecular membranes made of lipids or the like as sensors in Patent Documents 1 and 2, etc. For example, in the "Taste Sensor and Manufacturing Method Thereof" section of Patent Document 1, they point out the importance of lipid membranes as taste sensors. Furthermore, in the "Lipid Membrane" section of Patent Document 2, they propose a lipid membrane highly selective for bitterness and a lipid membrane highly selective for astringency. Furthermore, in the "Molecular Membrane for Taste Testing" section of Patent Document 3, they propose a lipid membrane highly selective for sourness, a lipid membrane highly selective for saltiness, and a lipid membrane highly selective for "umami." Furthermore, while Non-Patent Document 1 does not contain any disclosure directly related to taste sensors, it does propose a lipid membrane highly sensitive to non-ions such as caffeine, which are different from taste.

[0005] Patent No. 2578370 Patent No. 4395236 Patent No. 4520577

[0006] Yoshimatsu, J.; Toko, K.; Tahara, Y.; Ishida, M.; Habara, M.; Ikezaki, H.; Kojima, H.; Ikegami, S.; Yoshida, M.; Uchida, T. Development of a Taste Sensor to Detect Non-Charged Bitter Substances. Sensors 2020, 20, 3455, doi: 10.3390 / s20123455.

[0007] The umami substance with the strongest umami flavor is monosodium glutamate (MSG), found in kelp broth. Another typical umami substance is monosodium aspartate (MSA), an amino acid with a similar structure found in many foods, such as bonito broth. The molecular structures of monosodium glutamate (MSG) and monosodium aspartate (MSA) are shown in Figures 1A and 1B, respectively.

[0008] An umami sensor has already been put to practical use based on the disclosure of Patent Document 3. This conventional umami sensor has already been put to practical use by many food manufacturers, including seasoning manufacturers. However, the conventional umami sensor that has been put to practical use is said to have the following problems.

[0009] The first problem is that, as shown in Figure 2, it is affected by pH, has high sensitivity to sourness, and has low selectivity for "umami."

[0010] In Figure 2, the vertical axis represents the output (mV) of a conventional umami sensor, and the horizontal axis represents the basic taste components (from left to right: salt, acid, monosodium glutamate (MSG), monosodium aspartate (MSA), bitterness +, bitterness -, and astringency) as shown in Figure 2, and the graphs show the output (mV) of the conventional umami sensor versus the basic taste components. The basic taste components are the same as the components of the samples shown in Table 1 below.

[0011]

[0012] The second problem is that with conventional umami sensors, when the concentration of an umami substance becomes high, the sensor output reverses, as shown in Figure 3. In other words, conventional umami sensors have a problem in that the measurement range of "umami" is narrow.

[0013] In FIG. 3, the vertical axis represents the output (mV) of the conventional umami sensor, and the horizontal axis represents the concentrations of monosodium glutamate (MSG) and monosodium aspartate (MSA).

[0014] The causes of the above-mentioned problems are believed to be as follows: Most foods have a pH of 4 to 8, and in this pH range, umami substances such as MSG are negatively ionized. Therefore, conventional umami sensors use a positively charged lipid that responds to negative ions, i.e., a positively charged lipid (e.g., trioctylmethylammonium chloride (TOMA) as shown in Figure 4B), as the sensor material. This positively charged lipid membrane has the property of responding negatively to negatively ionized umami substances. However, a problem with positively charged lipid membranes is that they also respond to negative Cl ions in salt (NaCl) (a so-called negative response). On the other hand, sensor membranes using negatively charged lipids (for example, bis(2-ethylhexyl)phosphate (Phosphoric Acid di(2-ethylhexyl) Ester: 2C8) as shown in Figure 4A) have the property of responding positively to the positive Na ions of salt (NaCl). Therefore, by utilizing the negative response of such positively charged lipid membranes and the negative response of positively charged lipid membranes, the positive and negative responses in both membranes are canceled out, thereby reducing the sensitivity to salt (NaCl) to zero. This method improves selectivity for "umami."

[0015] However, the POOH group of the negatively charged lipid (2C8) has the property that its dissociation state changes depending on the pH. Therefore, when negatively charged lipids are used as membrane materials, they are greatly affected by pH, which causes a decrease in selectivity for "umami." This is why conventional sensor membranes have the output characteristics shown in Figures 2 and 3.

[0016] In addition, umami substances are weakly alkaline, and as the concentration of umami substances increases, the pH also increases, and the negatively charged POOH group of the lipid (2C8) dissociates, i.e., POOOH － The negatively charged lipids ionize, resulting in a more negative response in the conventional umami sensor membrane. In particular, at high concentrations of umami substances, all negatively charged lipids dissociate, preventing further negative responses. On the other hand, the conventional umami sensor membrane is affected by the sodium ions contained in the umami substance, resulting in a positive response, resulting in a stronger positive response. Although the conventional umami sensor membrane responds negatively to umami substances, as the umami substance concentration increases, the positive response of the negative lipid membrane becomes stronger, resulting in a reversal of the sensor output. As is clear from Figure 3, which shows the MSG concentration characteristics of the negative lipid membrane, a negative response occurs at low MSG concentrations, but a positive response occurs at higher MSG concentrations. This leads to the second problem mentioned above. Thus, using negative lipids as membrane materials in taste sensor membranes poses a risk of reversal of the umami substance's charge at high concentrations.

[0017] According to an embodiment, an umami sensor membrane is provided, which is characterized in that the surface of a membrane formed by mixing a polymer material, a positively charged amphipathic substance, and a plasticizer contains an aromatic polycarboxylic acid having one or more hydroxyl groups, a negatively charged amphipathic substance in which a carboxyl group and a hydroxyl group are adjacent on the benzene ring, or a negatively charged amphipathic substance in which carboxyl groups are adjacent to each other, and the membrane potential is changed by the umami substance in the aqueous solution to be measured, thereby detecting the intensity of the ``umami'' taste.

[0018] In this embodiment, there is provided an umami sensor membrane of the above-described form, characterized in that the negatively charged amphiphilic substance is one selected from the group consisting of 2,6-dihydroxyterephthalic acid (2,6-DHTA), 2-hydroxyterephthalic acid (2-HTA), 2,3-dihydroxyterephthalic acid (2,3-DHTA), 2,5-dihydroxyterephthalic acid (2,5-DHTA), 2-hydroxyisophthalic acid (2-HIPA), 4-hydroxyisophthalic acid (4-HIPA), 3-hydroxyphthalic acid (3-HPA), and 4-hydroxyphthalic acid (4-HPA).

[0019] Furthermore, according to an embodiment, there is provided a method for producing an umami sensor membrane, which comprises the steps of: mixing a polymer material and a positively charged amphipathic substance in an organic solvent to produce a mixed solution; drying the mixed solution to form a membrane; immersing the membrane in a negatively charged amphipathic substance dissolved in water for a predetermined period of time; and drying the membrane to form a membrane structure containing the negatively charged amphipathic substance on the surface of the membrane.

[0020] In this embodiment, there is provided the above-mentioned method for producing an umami sensor membrane, wherein the negatively charged amphiphilic substance is one selected from the group consisting of 2,6-dihydroxyterephthalic acid (2,6-DHTA), 2-hydroxyterephthalic acid (2-HTA), 2,3-dihydroxyterephthalic acid (2,3-DHTA), 2,5-dihydroxyterephthalic acid (2,5-DHTA), 2-hydroxyisophthalic acid (2-HIPA), 4-hydroxyisophthalic acid (4-HIPA), 3-hydroxyphthalic acid (3-HPA), and 4-hydroxyphthalic acid (4-HPA).

[0021] According to an embodiment, a method for detecting umami using an umami sensor membrane is provided, the method comprising: measuring the potential of the umami sensor membrane relative to a reference electrode in a reference solution to detect a first response voltage Vr between the reference electrode and the umami sensor membrane; measuring the potential of the umami sensor membrane relative to the reference electrode in a sample solution; measuring the potential of the umami sensor membrane relative to the reference electrode in a reference solution; measuring the potential of the umami sensor membrane relative to the reference electrode in a sample solution; and determining a second response voltage Vr' between the reference electrode and the umami sensor membrane. The method determines the difference voltage (Vr'-Vr) between the first and second response voltages Vr and Vr', and outputs this difference voltage (Vr'-Vr) to detect the umami intensity.

[0022] In this embodiment, a method for detecting "umami" using an umami sensor membrane is provided, wherein the negatively charged amphiphilic substance is one selected from the group consisting of 2,6-dihydroxyterephthalic acid (2,6-DHTA), 2-hydroxyterephthalic acid (2-HTA), 2,3-dihydroxyterephthalic acid (2,3-DHTA), 2,5-dihydroxyterephthalic acid (2,5-DHTA), 2-hydroxyisophthalic acid (2-HIPA), 4-hydroxyisophthalic acid (4-HIPA), 3-hydroxyphthalic acid (3-HPA), and 4-hydroxyphthalic acid (4-HPA).

[0023] The sensor using the umami sensor film according to the embodiment can evaluate the degree of umami.

[0024] 1 is a chemical formula showing the molecular structure of monosodium glutamate (MSG), one of the representative umami substances. 2 is a chemical formula showing the molecular structure of sodium aspartate (MSA), another representative umami substance. 3 is a graph showing the selectivity of taste components in a conventional umami sensor, shown as the output (mV) of the taste sensor on the vertical axis against the basic taste components on the horizontal axis. 4 is a graph showing the concentration characteristics of monosodium glutamate (MSG) and sodium aspartate (MSA) in a conventional umami sensor, shown as the output (mV) of the sensor on the vertical axis against the concentration (mM) of the umami components on the horizontal axis. 5 is a chemical formula showing the molecular structure of bis(2-ethylhexyl)phosphate (2C8), a negatively charged lipid material used in conventional umami sensors. 6 is a chemical formula showing the molecular structure of trioctylmethylammonium chloride (TOMA) (R=C), a positively charged lipid material used in conventional umami sensors. ６ ~C １０1 is a chemical formula showing the molecular structure of 2,6-dihydroxyterephthalic acid (2,6-DHTA) as a negatively charged lipid used in an umami taste sensor membrane according to an embodiment of the present invention. 2,6-dihydroxybenzoic acid (2,6-DHBA) as a negatively charged lipid used in a caffeine sensor membrane for detecting caffeine, as disclosed in Non-Patent Document 1, is shown in (a) a chemical formula showing the molecular structure in an undissociated state at a pH of 1 or less, and (b) a chemical formula showing the molecular structure in an dissociated state at a pH of 3 or more. 6(a) is a schematic diagram of the interaction between caffeine and negatively charged 2,6-dihydroxybenzoic acid (2,6-DHBA) used in the caffeine sensor membrane shown in FIG. 6(a), showing (a) negatively charged 2,6-DHBA dissociated in aqueous solution, and (b) stacking interaction with caffeine (protons approach each other as shown by the dashed lines), resulting in either a stacking state of non-dissociated 2,6-DHBA as indicated by the arrows. This graph shows how the sensor output (vertical axis) changes with pH (horizontal axis) in the presence of caffeine (100 mM) in the caffeine detection sensor membrane using 2,6-DHBA as the negatively charged lipid shown in FIG. 6(a). This graph shows how the sensor output (mV) (vertical axis) changes with pH (horizontal axis) in the absence of caffeine in the caffeine detection sensor membrane using 2,6-DHBA as the negatively charged lipid shown in FIG. 6(a). 1 is a graph showing the concentration characteristics (sensor output (mV) on the vertical axis versus the concentration (mM) of the umami component on the horizontal axis) for monosodium glutamate (MSG) and monosodium aspartate (MSA) of an umami sensor employing 2,6-dihydroxyterephthalic acid (2,6-DHTA) as the lipid according to an embodiment. 2 is a graph showing the selectivity of taste components (sensor output (mV) on the vertical axis versus the basic tastes on the horizontal axis) of an umami sensor using 2,6-dihydroxyterephthalic acid (2,6-DHTA) according to an embodiment.1A and 1B are schematic diagrams showing the molecular structure of 2,6-dihydroxyterephthalic acid (2,6-DHTA) as a negatively charged lipid used in an umami sensor membrane according to an embodiment, in a state where it is not dissociated at a pH of 1 or less, and in a state where it is dissociated at a pH of 3 or more. 2,6-Dihydroxyterephthalic acid (2,6-DHTA) as a negatively charged lipid used in an umami sensor membrane according to an embodiment and monosodium glutamate (MSG) are shown in (a) a schematic diagram showing how 2,6-DHTA dissociates in an aqueous solution, becomes negatively charged, and interacts with MSG, and (b) a schematic diagram showing how the interaction results in the formation of non-dissociated 2,6-DHTA. 1 is a schematic diagram showing the molecular structure of 2-hydroxyterephthalic acid (2-HTA) in an undissociated state as a negatively charged lipid used in an umami sensor membrane according to an embodiment, with the acid dissociation constant (pka) indicated by a number in the diagram; FIG. 2 is a schematic diagram showing the molecular structure of 2,3-dihydroxyterephthalic acid (2,3-DHTA) in an undissociated state as a negatively charged lipid used in an umami sensor membrane according to an embodiment, with the acid dissociation constant (pka) indicated by a number in the diagram; and FIG. 3 is a schematic diagram showing the molecular structure of 2,5-dihydroxyterephthalic acid (2,5-DHTA) in an undissociated state as a negatively charged lipid used in an umami sensor membrane according to an embodiment, with the acid dissociation constant (pka) indicated by a number in the diagram. 1 is a schematic diagram showing the molecular structure of terephthalic acid (TPA) as a lipid according to a comparative example in an undissociated state, with the acid dissociation constant (pka) indicated by a number in the diagram. 2 is a schematic diagram showing the molecular structure of 2-hydroxyisophthalic acid (2-HIPA) as a negatively charged lipid used in an umami sensor membrane according to an embodiment in an undissociated state, with the acid dissociation constant (pka) indicated by a number in the diagram.FIG. 1 is a schematic diagram showing the molecular structure of 4-hydroxyisophthalic acid (4-HIPA) as a negatively charged lipid used in an umami sensor membrane according to an embodiment in an undissociated state, with the acid dissociation constant (pka) indicated by a number in the diagram. FIG. 2 is a schematic diagram showing the molecular structure of isophthalic acid (IPA) as a lipid in a comparative example in an undissociated state, with the acid dissociation constant (pka) indicated by a number in the diagram. FIG. 3 is a schematic diagram showing the molecular structure of 3-hydroxyphthalic acid (3-HPA) as a negatively charged lipid used in an umami sensor membrane according to an embodiment in an undissociated state, with the acid dissociation constant (pka) indicated by a number in the diagram. 1 is a schematic diagram showing the undissociated molecular structure of 4-hydroxyphthalic acid (4-HPA) as a negatively charged lipid used in an umami sensor membrane according to an embodiment, with the acid dissociation constant (pka) indicated by a number in the diagram. 2 is a schematic diagram showing the undissociated molecular structure of phthalic acid (PA) as a lipid according to a comparative example, with the acid dissociation constant (pka) indicated by a number in the diagram. 3 is a graph showing the MSG concentration characteristics of umami sensor membrane lipids in which the positional relationship of the carboxyl group on the benzene ring is para-isomer, showing the relationship between the sensor output and the MSG concentration (mM) in an umami sensor membrane containing lipids (2,6-DHTA, 2-HTA, 2,3-DHTA, 2,5-DHTA) according to an embodiment and an umami sensor membrane containing a lipid (TPA) according to a comparative example. 1 is a graph showing the relationship between the sensor output and the MSG concentration (mM) in an umami sensor membrane containing lipids (2-HIPA, 4-HIPA) according to an embodiment and a comparative example containing lipid (IPA), showing the MSG concentration characteristics in an umami sensor membrane lipid in which the positional relationship of the carboxyl group on the benzene ring is meta-type. 2 is a graph showing the relationship between the sensor output and the MSG concentration (mM) in an umami sensor membrane containing lipids (3-HPA, 4-HPA) according to an embodiment and a comparative example containing lipid (PA), showing the MSG concentration characteristics in an umami sensor membrane lipid in which the positional relationship of the carboxyl group on the benzene ring is ortho-type.29 is a schematic diagram showing a taste sensor system using an umami sensor membrane according to this embodiment. 30 is a front view showing a sensor probe incorporating a sensor according to this embodiment in the taste sensor system shown in FIG. 27, with a portion thereof seen through. 31 is a front view showing a reference electrode probe in the taste sensor system shown in FIG. 27, with a portion thereof seen through. 32 is a schematic diagram for explaining the composition and chemical structure of an umami sensor membrane according to this embodiment used in the sensor system shown in FIG. 28. 33 is a chemical formula showing the molecular structure of tetradodecyl ammonium bromide (TDAB) as a positively charged lipid mixed into the polymer material of the umami sensor membrane shown in FIG. 34. 34 is a chemical formula showing the molecular structure of di-n-octylphenyl phosphonate (DOPP) as a plasticizer mixed into the polymer material of the umami sensor membrane shown in FIG. 34. 35 is a flowchart showing a manufacturing process of the umami sensor membrane shown in FIG. 35. 28 is a flowchart showing the process of measuring the degree of "umami" in the taste sensor system shown in FIG. 28.

[0025] Hereinafter, an umami sensor film according to an embodiment, a method for manufacturing an umami sensor film, and a method for detecting "umami" using an umami sensor film will be described with reference to the drawings.

[0026] First, we will explain the process by which the inventors arrived at the idea that umami can be detected using an umami sensor membrane using 2,6-dihydroxyterephthalic acid (2,6-DHTA) according to the embodiment, based on the knowledge they gained from Non-Patent Document 1. In this explanation, we will provide an overview of the response principle of caffeine detection presumed in Non-Patent Document 1, and compare it with the response principle of umami detection using an umami sensor membrane presumed in the same way.

[0027] Figure 5 shows the molecular structure of 2,6-dihydroxyterephthalic acid (2,6-DHTA), a lipid amphiphilic substance used in the umami sensor membrane according to the embodiment. 2,6-dihydroxyterephthalic acid (2,6-DHTA) is a type of carboxylic acid, but it has an extremely low acid dissociation constant (pKa) of 1 to 2. This is believed to be due to the formation of hydrogen bonds between the OH group in the 2,6 portion and the carboxylic acid group, which releases H+ ions even at extremely low pH levels, resulting in a dissociated state. However, the inventors discovered that when 2,6-dihydroxyterephthalic acid (2,6-DHTA) interacts with umami substances, monosodium glutamate (MSG) or monosodium aspartate (MSA), this dissociation does not occur and the dissociated state does not occur. The inventors focused on this phenomenon of interaction and came up with the idea that by utilizing this phenomenon, it should be possible to fabricate a sensor film that detects "umami" with desirable characteristics.

[0028] The inventors first inferred that the response principle of Non-Patent Document 1 is based on the following process, and from this inference, they focused on the phenomenon of no discrepancy described above. Non-Patent Document 1 does not describe the detection of "umami" but describes a sensor membrane for detecting caffeine. This caffeine detection sensor membrane uses 2,6-dihydroxybenzoic acid (2,6-DHBA) as the negative lipid, which exhibits a strong positive response to non-ionic caffeine. Although 2,6-DHBA has a carboxylic acid group, its acid dissociation constant (pKa) is very low, at approximately 1.6. In other words, as shown in Figure 6(a), 2,6-DHBA does not dissociate at pH 1.0 or below, but dissociates by 50% even at a strongly acidic pH of 1.6. As shown in Figure 6(b), it dissociates by nearly 100% at pH 3 or above. Since most foods have a pH between 3 and 8, 2,6-DHBA is always in a dissociated state in the pH range of food samples. Generally, organic acids with a carboxylic acid group, such as acetic acid, are weak acids, and their acid dissociation constants (hereinafter simply referred to as pka) are high, at 4 to 5. In other words, organic acids with a carboxylic acid group, such as acetic acid, do not dissociate unless the pH is 4 to 5 or higher. However, 2,6-dihydroxybenzoic acid (simply referred to as 2,6-DHBA) is prone to forming a stable form due to intramolecular hydrogen bonds, as shown in Figure 6(b), and therefore dissociates even in an acidic pH of 1.6.

[0029] Regarding this dissociation phenomenon, the inventors have inferred that, in water, 2,6-DHBA (shown in Figure 7(a)) and caffeine (shown in Figure 7(b)) undergo stacking interactions as indicated by the arrows in Figure 7(a) or 7(b), resulting in protons approaching each other as indicated by the dashed circle, and that 2,6-DHBA does not dissociate even at high pH levels. In other words, 2,6-DHBA alone dissociates to become negatively charged ions at pH 2 or higher, but the presence of caffeine prevents dissociation and eliminates the negative charge. By utilizing this effect, as shown in Figure 8, a sensor membrane using 2,6-DHBA can achieve a high sensitivity of approximately 50 mV to caffeine (100 mM) at pH levels of 2 to 10. Furthermore, in the absence of caffeine, the sensor membrane exhibits a sensor output of approximately 0 mV at pH 4 to pH 12, meaning that it is not affected by pH, as shown in Figure 9. In other words, a sensor membrane using 2,6-DHBA responds only to caffeine without being affected by pH. Non-Patent Document 1 also describes that even in the case of dihydroxybenzoic acid, those in which the OH groups are not at positions 2 and 6 have a low response to caffeine.

[0030] The caffeine response is based on a completely different response principle from that of conventional lipid membrane sensors. In other words, with conventional lipid membranes, charged taste substances adsorb to and interact with the sensor membrane, causing a change in the potential of the sensor membrane, which is then used as the output. However, with a sensor membrane that detects caffeine using 2,6-DHBA, uncharged caffeine interacts with 2,6-DHBA, causing the negatively charged 2,6-DHBA to dissociate and become zero-charged, resulting in a change in the potential of the sensor membrane from negative to zero, i.e., a positive response.

[0031] The inventors focused on this new response principle and believed that an umami sensor could be developed based on this response principle. In this embodiment, by employing 2,6-dihydroxyterephthalic acid (2,6-DHTA) as the negative lipid shown in Figure 5 in the umami sensor of the embodiment, it has been found that the umami sensor of this embodiment can be given higher sensitivity to umami substances (MSG and MSA) as shown in Figures 10 and 11, compared to salt, acid, bitter +, bitter -, and astringent tastes as shown in Table 1. Furthermore, like the 2,6-DHBA used in the caffeine sensor membrane, 2,6-dihydroxyterephthalic acid (2,6-DHTA) is not in a dissociated state at pH 1 or below, as shown in Figure 12(a), but is always in a dissociated state at pH 3 or above, and is not affected by pH, as shown in Figure 12(b). Therefore, as is clear from the sensor detection of the sample components in Table 1 above, it was found that the sensor has low sensitivity to sourness and high selectivity for the sensor detection of "umami" as shown in Figure 11. Furthermore, as shown in Figure 11, the effect of Na positive ions is constant at pH 1 or higher, so the effect of Na positive ions is constant and it is clear that no reversal occurs as shown in Figure 10.

[0032] As shown in Figure 12(b), 2,6-DHTA according to the embodiment is easily stabilized by intramolecular hydrogen bonding, allowing it to remain dissociated even at an acidic pH of 1.6. The inventors confirmed that 2,6-DHTA and MSG interact in water, as shown in Figure 13(a), and that 2,6-DHTA does not dissociate even at high pH levels, as shown in Figure 13(b). Specifically, 2,6-DHTA according to the embodiment dissociates to form negatively charged ions at pH levels above 2, but the presence of MSG, an umami component, prevents dissociation and eliminates the negative charge. Incidentally, it has also been experimentally confirmed that 2,6-DHTA, used in caffeine detection, does not respond at all to the umami substances MSG or MSA. It is presumed that the umami component MSG or MSA molecules are larger than 2,6-DHBA and have weaker intermolecular interactions, which is why they did not respond. It is clear that the contribution of 2,6-DHTA according to this embodiment to umami detection is a major discovery.

[0033] As already explained, conventional umami sensors exhibit a phenomenon in which the output is inverted at high concentrations of umami components, as shown in Figure 3. Furthermore, it is known that in actual foods, this inversion occurs at lower concentrations. This is thought to occur because Ca ions and Mg ions in the food strongly bind to the negative lipids. In contrast, it has been found that the umami sensor employing 2,6-DHTA as the lipid according to this embodiment has low reactivity with Ca ions and Mg ions in the stable state shown in Figure 12(a), and thus no inversion occurs. Thus, the umami sensor employing 2,6-DHTA as the lipid according to this embodiment exhibits higher food adaptability than conventional umami sensors, and is expected to dramatically expand its versatility for use with foods.

[0034] In addition to 2,6-DHTA, there are other molecular structures that are optimal for use as lipids in umami sensors based on their interactions. These molecular structures are aromatic polycarboxylic acids with one or more hydroxyl groups, such as benzene rings with adjacent carboxyl groups and hydroxyl groups on the benzene ring, or benzene rings with adjacent carboxyl groups. When a carboxyl group and a hydroxyl group on the benzene ring are adjacent, even if there is only one hydroxyl group, the acid dissociation constant (pka) of the carboxyl group decreases due to intramolecular hydrogen bonding with the carboxyl group. When the length between the carboxyl groups on the benzene ring, or the length between the carboxyl group and the hydroxyl group on the benzene ring, becomes the same as the length between the two carboxylic acid groups in MSG, an intermolecular hydrogen bond with MSG is formed. This intermolecular hydrogen bond with MSG increases the acid dissociation constant (pka) of the carboxyl group. As a result, it has been found that the sensor has the same sensitivity to MSG as 2,6-DHTA. If the two carboxyl groups on the benzene ring are in a meta or para configuration, and if there is a hydroxyl group on the carboxyl group, the carboxyl group and the hydroxyl group will be adjacent. The adjacent carboxyl groups will form hydrogen bonds with the hydroxyl group, lowering the acid dissociation constant (pka). Even at low pH, they will dissociate and assume a negative charge (Figures 5, 14, 15, 16, 18, and 19). If the two carboxyl groups on the benzene ring and the two carboxyl groups on MSG are at the same distance from each other, as shown in Figure 13(b), intermolecular hydrogen bonding will occur with MSG, increasing the lowered acid dissociation constant (pka). The negative charge will disappear, becoming zero, and the sensor membrane potential will respond positively. If the two carboxyl groups on the benzene ring are in an ortho configuration, the hydrogen bonding between the two carboxyl groups will lower the acid dissociation constant (pka), and they will dissociate and assume a negative charge even at low pH (Figures 22 and 23). The distance between the carboxyl group and the OH group on the benzene ring then becomes the same as the distance between the two carboxyl groups of MSG, and as shown in Figure 13 (b), an intermolecular hydrogen bond occurs with MSG, the acid dissociation constant (pka) that had decreased increases, the molecule no longer dissociates, the negative charge disappears, the negative charge becomes zero, and the membrane potential of the sensor responds positively.

[0035] Examples of specific molecular structures include 2-hydroxyterephthalic acid (2-HTA) shown in Figure 14, 2,3-dihydroxyterephthalic acid (2,3-DHTA) shown in Figure 15, 2,5-dihydroxyterephthalic acid (2,5-DHTA) shown in Figure 16, 2-hydroxyisophthalic acid (2-HIPA) shown in Figure 18, 4-hydroxyisophthalic acid (4-HIPA) shown in Figure 19, 3-hydroxyphthalic acid (3-HPA) shown in Figure 21, and 4-hydroxyphthalic acid (4-HPA) shown in Figure 22. The numbers in these figures indicate acid dissociation constants (pka).

[0036] On the other hand, as a comparison example to the molecular structure described above, if the two carboxyl groups on the benzene ring are in a meta or para positional relationship, and there is no hydroxyl group, the pKa of the carboxyl groups does not decrease, and the molecule does not respond to MSG. Examples of this are terephthalic acid (TPA) shown in Figure 17 and isophthalic acid (IPA) shown in Figure 20. If the two carboxyl groups on the benzene ring are ortho, the two carboxyl groups are adjacent and indeed hydrogen bond with each other, lowering the pKa. However, this does not match the distance between the two carboxyl groups of MSG, preventing intermolecular hydrogen bonding with MSG, and resulting in a response to MSG. An example of this is phthalic acid (PA) shown in Figure 23.

[0037] As already mentioned, the molecular structures of the above-mentioned negatively charged amphiphilic substances are shown in FIGS. 14 to 23, and in these figures, the acid dissociation constants (pka) are shown numerically.

[0038] Figures 24, 25, and 26 show the concentration characteristics of MSG versus sensor output for each positional relationship of the carboxyl groups (para, meta, ortho). In the para and meta forms, the molecular length is the same as that of MSG due to the positional relationship of the two carboxyl groups. When the acid dissociation constant (pka) is reduced due to intramolecular hydrogen bonding caused by the hydroxyl group adjacent to the carboxyl group, as is clear from Figures 24 and 25, the sensor sensitivity to MSG is achieved due to intermolecular hydrogen bonding.

[0039] 24, it has been found that the para forms 2,3-dihydroxyterephthalic acid (2,3-DHTA), 2-hydroxyterephthalic acid (2-HTA), 2,3-dihydroxyterephthalic acid (2,3-DHTA), and 2,5-dihydroxyterephthalic acid (2,5-DHTA) provide sufficient sensor output in response to changes in MSG concentration. In contrast, it has also been found that terephthalic acid (TPA), as a comparative example, does not provide sufficient sensor output in response to changes in MSG concentration.

[0040] Similarly, as shown in Figure 25, it has been found that in the meta-forms 2-hydroxyisophthalic acid (2-HIPA) and 4-hydroxyisophthalic acid (4-HIPA), sufficient sensor output can be obtained in response to changes in MSG concentration. In contrast, it has also been found that in the comparative example, isophthalic acid (IPA), sufficient sensor output cannot be obtained in response to changes in MSG concentration.

[0041] On the other hand, the ortho isomer has two adjacent carboxyl groups, which reduces the acid dissociation constant (pKa) due to intermolecular hydrogen bonding. The molecular length matches that of MSG, which increases the acid dissociation constant (pKa) due to intermolecular hydrogen bonding with MSG. This is thought to be why the ortho isomer responded to MSG, as shown in Figure 26. More specifically, as shown in Figure 26, it has been found that, for the ortho isomers, 3-hydroxyphthalic acid (3-HPA) and 4-hydroxyphthalic acid (4-HPA), sufficient sensor output was obtained in response to changes in MSG concentration. In contrast, it has also been found that phthalic acid (PA), used as a comparative example, did not provide sufficient sensor output in response to changes in MSG concentration.

[0042] Furthermore, while the above examples illustrate negatively charged amphiphilic substances, the moiety responds to MSG, and negatively charged amphiphilic substances containing this moiety are included in the materials for the umami sensor in the examples of this application. For example, to prevent elution from the sensor membrane into water, benzene rings or carbon chains can be added to the molecules in the examples to increase the durability of the sensor membrane while maintaining MSG responsiveness. Furthermore, while the examples illustrate examples with two carboxyl groups on the benzene ring, if there are three or more hydroxyl groups, the carboxyl groups may be adjacent to each other or adjacent to a hydroxyl group, reducing the acid dissociation constant (pka) of the carboxyl group. Furthermore, the distance between the carboxyl groups or between the carboxyl group and the hydroxyl group becomes identical to the two carboxyl groups of MSG, forming intermolecular hydrogen bonds with MSG, increasing the lowered acid dissociation constant (pka), resulting in a response to MSG.

[0043] The data shown in FIGS. 24, 25 and 26 are measured using a sensor system shown in FIG. 27, which will be described below.

[0044] FIG. 27 shows a taste sensor system that uses an umami sensor membrane according to an embodiment.

[0045] In the taste sensor system shown in FIG. 27 , a container 10 is prepared for separately storing a reference solution, a sample solution, a cleaning solution, etc., and a reference electrode probe 11 and one or more taste sensor probes 15 shown in FIGS. 28A and 28B are supported and fixed by an arm mechanism (not shown) so that they can be inserted and removed from the container 10. The arm mechanism (not shown) moves the probes 11 and 15 up and down. This taste sensor system employs the CPA (Change of Membrane Potential Caused by Adsorption) measurement method. A sensor membrane for detecting each taste quality is prepared, and each sensor membrane is attached to the taste sensor probe 15. Here, the taste qualities are "umami," sourness, saltiness, sweetness, astringency, and bitterness. Therefore, a taste sensor probe 15 equipped with a sensor membrane corresponding to each taste quality is prepared. Naturally, a taste sensor probe 15 equipped with a sensor membrane for detecting "umami" is prepared.

[0046] As shown in Fig. 28A, this taste sensor probe 15 has a sensor membrane 17 for detecting a corresponding taste quality fixed around a through-hole in a tube 16. The surface of the sensor membrane 17 is exposed to the interior of the container 11, and the opposite surface of the sensor membrane 17 is exposed to a conductive internal liquid contained in the tube 16. One example of this internal liquid is a 3.3 M KCl saturated AgCl solution. A sensor electrode 19 is inserted into the tube 16, and the tip of the sensor electrode 19 is immersed in the conductive internal liquid so as to face the sensor membrane 17. The electrode 19 is connected to a voltage detector 20 via a lead wire 19A.

[0047] 28B, the reference electrode probe 11 is composed of a tubular glass tube 13 whose tip opening is sealed with a liquid-tight porous ceramic 18. The glass tube 13 is filled with a conductive internal liquid, similar to the sensor, and the reference electrode 12 is immersed in this conductive internal liquid. The reference electrode 12 is connected to a voltage detector 20 via a lead wire 12A.

[0048] In the CPA measurement method, changes in membrane potential due to electrostatic / hydrophobic interactions between the sensor membrane and the taste substance are output as voltage signals. A voltage detector 20 detects the voltage between the reference electrode 12 and the sensor electrode 19 for the reference and sample solutions, detecting changes over time after the probes 12 and 15 are immersed in the reference and sample solutions. The detected voltage signals are converted into digital voltage signals by an A / D converter 22, which are then sent to a computing device 23 and stored in memory 23A. The computing device 23 calculates the temporal changes in the sensor voltage Vr in the reference solution and the response voltage Vs in the sample solution as response values ​​(relative values: Vs-Vr) and stores them in memory 23A. The memory 23A stores the response values ​​(relative values: Vs-Vr) as first taste response value data. The computing device 23 compares these first taste response values ​​(relative values: Vs-Vr) with known data to quantify the taste quality of the sample solution, which is then output to an output device 24 for use in taste assessment.

[0049] As will be explained later, in an embodiment for measuring umami substances, after a response value (relative value: Vs-Vr) is detected as the first taste, the probes 12 and 15 are immersed in a reference solution and washed with the reference solution. Then, the voltage Vr' between the reference electrode 12 and the sensor electrode 19 relative to the reference solution is detected, and the differential voltage (CPA value: Vr'-Vr) is calculated as the aftertaste response value and stored in memory 23A. Thereafter, the probes 12 and 15 are washed and used again for taste testing.

[0050] Next, the structure of the umami sensor membrane according to the embodiment and the method for manufacturing the same will be described below.

[0051] The umami sensor membrane has a structure as shown in FIG. 29, in which polyvinyl chloride (PVC) is used as the polymer material for the base membrane, tetradodecyl ammonium bromide (TDAB) shown in FIG. 30A is used as the positively charged lipid mixed into the polymer material, 2,6-dihydroxyterephthalic acid (2,6-DHTA) shown in FIG. 5 is used as the negatively charged lipid provided on the surface of the base membrane, and di-n-octylphenyl phosphonate (DOPP) shown in FIG. 30B is used as the plasticizer mixed into the polymer material.

[0052] The umami sensor membrane shown in Figure 29 is manufactured according to the process shown in Figure 31. First, as shown in step S11, PVC (800 mg) as a polymer material, TDAB (0.01 mmol) as a positively charged lipid, and DOPP (1.5 mL) as a plasticizer are dissolved and mixed in tetrahydrofuran (THF) (10 mL) as an organic solvent. Next, as shown in step S12, the mixture is spread on a clean petri dish (90 mm diameter), and a TDAB lipid / polymer membrane is formed by evaporation of the THF organic solvent. Examples of plasticizers are listed in Tables 2 and 3 below. Examples of positively charged lipids are listed in Table 4 below.

[0053]

[0054]

[0055]

[0056] The resulting lipid / polymer membrane is cut to prepare an untreated sensor membrane, which is then adhered to the sensor electrode. Next, as shown in step S13, a negatively charged lipid, 2,6-DHTA, is dissolved in a solvent such as water to produce a 0.03 wt% aqueous solution of the negatively charged lipid, 2,6-DHTA, as a predetermined amphiphilic substance. Then, as shown in step S14, the sensor membrane is immersed in the 0.03 wt% aqueous solution of the negatively charged lipid, 2,6-DHTA, for 72 hours to modify the sensor surface. Then, as shown in step S13, the umami sensor electrode is immersed in a reference solution containing 30 mM KCl and 0.3 mM tartaric acid for 72 hours to produce a sensor membrane.

[0057] As already explained, the completed umami sensor membrane 17 will be composed of a lipid-polymer membrane as shown in Fig. 29. In the umami sensor membrane 17 according to the embodiment, the inside of the sensor membrane is composed of PVC (polyvinyl chloride) 17A as a polymer material, and is composed of TDAB 17B as a positively charged lipid and a plasticizer 17C. The surface of the sensor membrane is modified with the negatively charged lipid 2,6-DHTA.

[0058] Taste measurement of umami substances using the umami sensor membrane 17 fabricated as described above is specifically performed in the sensor system shown in Fig. 27 according to the procedure shown in Fig. 33, and the sensor output (voltage change) shown in Fig. 32(a) is obtained. The sensor output (voltage change) shown in Fig. 32(a) corresponds to the action of negative charges on the sensor membrane 17 in the reference solution 30 and the aqueous solution to be measured (so-called sample solution 28) in Fig. 32(b), (c), (d), and (e).

[0059] As shown in Table 1, an example of a reference solution 30 prepared for taste measurement is a solution of 30 mM KCl and 0.3 mM tartaric acid (30 mM KCl + 0.3 mM tartaric acid). As shown in Table 1, an example of a test solution (so-called sample solution 28) prepared for umami taste measurement is a solution of monosodium glutamate (MSG) and monosodium aspartate (MSA). The reference solution and the test solution of the umami substance (so-called sample solution 28) are stored in separate containers 10. First, as shown in FIG. 33 , the reference electrode probe 11 and the taste sensor probe 15 are immersed in the reference solution 30, and the reference voltage Vr between the reference electrode 12 and the sensor electrode 19 is measured (S1). Because the reference solution 30 does not contain any charge of the substance to be measured, the potential of the sensor membrane 17 is maintained constant, and a baseline voltage is output from the reference electrode 12 and the sensor electrode 19.

[0060] It is preferable that this reference voltage Vr be compared with the previously measured reference voltage to confirm whether it is within the reference range. If the difference between the previously measured reference voltage Vr and the currently measured reference voltage Vr is within a predetermined value, the process proceeds to the next step S2. If the difference between the previously measured reference voltage Vr and the currently measured reference voltage Vr is not within the predetermined value, step S1 is executed again to measure the reference solution again. When measuring the reference solution 30 for the first time, step S1 is repeated a predetermined number of times and averaged to determine the reference value Vr, which is then stored in memory 23A (S1).

[0061] Next, the reference electrode probe 11 and the taste sensor probe 15 are immersed in the solution to be measured (sample solution) 28, and the sample voltage Vs between the reference electrode 12 and the sensor electrode 19 is measured (S2).

[0062] Thereafter, the reference electrode probe 11 and the taste sensor probe 15 are again immersed in the reference solution 30, and the sensor membrane 17 is washed with the reference solution 30. As the sensor membrane 17 is washed with the reference solution 30 from time t2, sour and salty substances 36 are moved (dispersed) into the reference solution 30 so as to be separated from the sensor membrane 17, as shown in FIG. 19(d). Therefore, the potential of the sensor membrane 17 is slightly increased to a potential where only bitter or astringent substances (hydrophobic substances) and umami substances 34 remain. The output voltage between the reference electrode 12 and the sensor electrode 19 is changed to a sensor output Vr' corresponding to a potential where only an aftertaste remains. This reference value voltage Vr' is measured and stored in memory 23A (S4). Thereafter, the differential voltage between the reference voltage Vr' and the reference voltage Vr (CPA value: ΔV'=Vr'-Vr) is calculated by the calculation device 23 as the aftertaste response value (CPA value) for the solution to be measured (sample solution) 28, and is stored in the memory 23A (S5).

[0063] At time t3, the reference electrode probe 11 and the taste sensor probe 15 are immersed in a cleaning solution (alcoholic cleaning solution) prepared in another container 10 and washed (S6). When the sensor membrane 17 is washed with the cleaning solution (alcoholic cleaning solution), bitter or astringent substances (hydrophobic substances) and umami substances 34 are separated from the sensor membrane 17 and diffused into the cleaning solution (alcoholic cleaning solution). Therefore, the potential of the sensor membrane 17 is changed so as to rise over time to the baseline reference voltage Vr.

[0064] Thereafter, steps S1 to S4 are repeated as necessary. When the taste measurement of the aqueous solution to be measured (the so-called sample solution 28) is completed, which is the process of steps S1 to S6, if there is a new aqueous solution to be measured (the so-called sample solution 28), the process of steps S1 to S6 is performed again. If there is no new aqueous solution to be measured (the so-called sample solution 28), the process ends in step S5.

[0065] Furthermore, in step S1, if the measurement value falls outside the specified range even after repeated measurements of the reference liquid, it is determined that the taste sensor probe 15 has not been sufficiently cleaned or that the sensor membrane of the taste sensor probe 15 has not recovered even after cleaning, and the taste sensor probe 15 is subject to replacement.

[0066] In a conventional umami sensor, the umami concentration characteristics are reversed in the high concentration region of umami substances as shown in Figure 3, but in the umami sensor according to the embodiment, this is improved as shown in Figure 10. Furthermore, in a conventional umami sensor, the selectivity is as shown in Figure 2, but in the umami sensor according to the embodiment, this is improved as shown in Figure 11.

[0067] Although embodiments of the present invention have been described, these embodiments are presented as examples and are not intended to limit the scope of the invention. These novel embodiments can be embodied in various other forms, and various omissions, substitutions, and modifications can be made without departing from the spirit of the invention. These embodiments and their modifications are included within the scope and spirit of the invention, and are also included in the scope of the invention and its equivalents as defined in the claims.

[0068] 10...container, 11...reference electrode probe, 12...reference electrode, 13...glass tube, 15...taste sensor probe, 16...tube, 17...sensor membrane, 17A...PVC (polyvinyl chloride), 17B...lipid, 17C...plasticizer, 18...porous ceramic, 19...sensor electrode, 12A, 19A...lead wire, 20...voltage detector, 22...A / D converter, 23...arithmetic unit, 23A...memory, 24...output device, 28...aqueous solution, 30...reference solution, 32...alcohol cleaning solution, 34...bitter or astringent substance (hydrophobic substance), 36...sour, salty or umami substance (hydrophilic substance)

Claims

1. On the surface of a film formed in a film shape by mixing a polymer material, an amphiphilic substance having a positive charge, and a plasticizer, there is an aromatic polycarboxylic acid having one or more hydroxyl groups, in which a carboxyl group and a hydroxyl group are adjacent on the benzene ring, or an amphiphilic substance having a negative charge in which carboxyl groups are adjacent to each other, or an amphiphilic substance having a negative charge in which carboxyl groups are adjacent to each other, and the membrane potential is changed by the umami substance in the aqueous solution to be measured to detect the intensity of "umami". This is a umami sensor film characterized by this.

2. The amphiphilic substance having a negative charge is selected from the group of 2,6-dihydroxyterephthalic acid (2,6-DHTA), 2-hydroxyterephthalic acid (2-HTA), 2,3-dihydroxyterephthalic acid (2,3-DHTA), 2,5-dihydroxyterephthalic acid (2,5-DHTA), 2-hydroxyisophthalic acid (2-HIPA), 4-hydroxyisophthalic acid (4-HIPA), 3-hydroxyphthalic acid (3-HPA), and 4-hydroxyphthalic acid (4-HPA). The umami sensor film according to claim 1, characterized by this.

3. A step of mixing a polymer material and an amphiphilic substance having a positive charge with an organic solvent to generate a mixed solution, a step of drying the mixed solution to form a film, a step of immersing the film in an amphiphilic substance having a negative charge dissolved in water for a predetermined time, and a step of drying this film to form a film structure containing an amphiphilic substance having a negative charge on the surface of the film. A method for manufacturing a umami sensor film, characterized by comprising these steps.

4. The amphiphilic substance having a negative charge is selected from the group of 2,6-dihydroxyterephthalic acid (2,6-DHTA), 2-hydroxyterephthalic acid (2-HTA), 2,3-dihydroxyterephthalic acid (2,3-DHTA), 2,5-dihydroxyterephthalic acid (2,5-DHTA), 2-hydroxyisophthalic acid (2-HIPA), 4-hydroxyisophthalic acid (4-HIPA), 3-hydroxyphthalic acid (3-HPA), and 4-hydroxyphthalic acid (4-HPA). The method for manufacturing a umami sensor film according to claim 3, characterized by this.

5. A umami sensor in which a polymer material, an amphiphilic substance having a positive charge, and a plasticizer are mixed and formed into a film, and a negatively charged amphiphilic substance is contained on the surface of the film. In a method for detecting the intensity of "umami" by changing the membrane potential with umami substances in the aqueous solution to be measured, in a reference solution, the potential of the umami sensor membrane with respect to the reference electrode is measured to detect a first response voltage Vr between the reference electrode and the umami sensor membrane. Next, in a sample solution, the potential of the umami sensor membrane with respect to the reference electrode is measured. Then, again in the reference solution, the potential of the umami sensor membrane with respect to the reference electrode is measured to detect a second response voltage Vr' between the reference electrode and the umami sensor membrane. A differential voltage (Vr' - Vr) between the first and second response voltages Vr and Vr' is obtained, and this differential voltage (Vr` - Vr) is output to detect the intensity of "umami". A method for detecting "umami" using the umami sensor membrane, characterized in that.

6. The method for detecting "umami" using the above-mentioned umami sensor membrane, characterized in that the amphiphilic substance having a negative charge is selected from the group consisting of 2,6-dihydroxyterephthalic acid (2,6-DHTA), 2-hydroxyterephthalic acid (2-HTA), 2,3-dihydroxyterephthalic acid (2,3-DHTA), 2,5-dihydroxyterephthalic acid (2,5-DHTA), 2-hydroxyisophthalic acid (2-HIPA), 4-hydroxyisophthalic acid (4-HIPA), 3-hydroxyphthalic acid (3-HPA), and 4-hydroxyphthalic acid (4-HPA).

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