JP3217801B2 - Taste sensor and manufacturing method thereof - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an artificial sensor capable of acting on behalf of the five senses of human beings, and more particularly, to a sensor or transducer that can substitute for human sensation, such as taste, which has heretofore been considered impossible by artificial sensors. Electronic device called.

[0002]

2. Description of the Related Art The same applicant has previously filed a patent application for the invention of "taste sensor and its manufacturing method" (Japanese Patent Application No. 1-1908).
No. 19; hereinafter referred to as a prior application of the same applicant), and according to the specification and the drawings, a so-called lipid molecule group housed in a surface matrix of a certain high molecular polymer with a specific molecular arrangement is basically used. It was shown that the sensor could be sensitive to salty, sour, bitter, and sweet tastes. Moreover, it has been shown that this type of sensor can measure taste instead of taste, which is one of the human five senses.

[0003] To explain this more specifically, in the prior application of the same applicant, for example, polyvinyl chloride (PVC) is used as a polymer and a plasticizer such as dioctyl phthalate (DOP) is used. And lipids are roughly 2:
The mixture at a weight ratio of 3: 1 was melted in tetrahydrofuran (THF), transferred to a flat-bottomed container, and kept on a uniformly heated plate at about 30 ° C. for 2 hours to volatilize THF and evaporate lipid. A membrane, ie, a lipidic molecular membrane housed in a PVC surface matrix, has been obtained. Experiments have confirmed that this lipid membrane serves as a taste sensor.

At this time, when the lipid membrane is immersed in an aqueous solution of potassium chloride (KCl), it appears that the hydrophilic groups of the lipid molecules are arranged in such a manner as to appear on the surface of the lipid membrane.
The sensitivity and stability of the sensor (membrane potential) were shown to be improved (Japanese Patent Application No. 1-190819).

However, using a lipid membrane formed in this way as a sensor, although a measurement result can be obtained to a certain extent, the production method of placing lipid molecules in the surface matrix of a high molecular polymer requires a molecular level. Structure is not constant,
In other words, it is not always possible to obtain a constant surface matrix itself, and it is difficult to guarantee that a lipid membrane containing lipid molecules is always of constant quality. The orientation of the reactive molecules is also poor, and therefore, the measurement of the membrane potential and the electrical resistance is likely to cause variations in the data and the sensitivity is inadequate.

[0006] Also, the types of lipid molecules that can be relatively easily accommodated in the surface matrix of a high molecular polymer have been limited. For example, phosphatidylcholine (P
Lipids constituting a biological membrane, such as C), have poor compatibility with a polymer material such as polyvinyl chloride (PVC), and thus it has been difficult to form a lipid membrane used for a taste sensor.

[0007]

SUMMARY OF THE INVENTION It is an object of the present invention to improve a conventional taste sensor using a lipid membrane by focusing on the following three points. That is, the quality of the taste sensor is made constant. Improve the quality of taste sensor. Increase the number of taste sensors. (Diversify)

Accordingly, it is an object of the present invention to realize a structure of a lipid membrane exhibiting a stable and less variable membrane potential with good reproducibility for each basic taste, and to realize a taste sensor using the lipid membrane. I have.

[0009] In addition, lipids that have been difficult to use in the conventional structure of lipid membranes, particularly, lipids that constitute biological membranes and substances that can be used as materials for taste sensors without being limited to lipids, can be used. It is an object of the present invention to realize a structure of a film and a taste sensor using the film.

In particular, a sensor for measuring a physical quantity such as a membrane potential of a substance or an electric conductivity of a surface, that is, a physical quantity where a substance is in contact with the surrounding environment and is slightly affected by the external world as an external interface. The aim is to obtain a product with stable quality and performance.

[0011]

Means for Solving the Problems It is considered that a hydrophilic group having a cationic group, an anionic group, a nonionic group, or an amphoteric group can be used for sensing a taste substance. This is called an amphiphilic substance having a hydrophobic group. Alternatively, a bitter substance such as an alkaloid, for example, quinine or the like is also adsorbed on the membrane and has a charge, so that it can be used as a material for the taste sensor.

In the present invention, in order to improve the orientation and obtain a film for a taste sensor having stable quality and performance while considering the utilization of these substances, the manufacturing procedure is first changed. Thereby, a stable and uniform layer of amphipathic molecules or a layer of molecules of bitter tastants is formed on the substrate.

That is, a predetermined amount of a polymer, a plasticizer, and a lipid substance are mixed, dissolved in a solvent, placed in a flat-bottomed container, and the solvent is evaporated to obtain a lipid membrane. In the invention of the prior application, it is considered that it is difficult to obtain a uniform surface matrix of the polymer, and it is intended to use a substance incompatible with the polymer such as polyvinyl chloride, and To improve the turbidity, avoid turbidity with the high molecular weight polymer, and firstly, dissolve the amphipathic substance or bitter substance in water. For example, use an alkaline aqueous solution such as caustic soda as a solvent, or use various alcohols that are well soluble in water as a solvent to dissolve an amphipathic substance or a bitter substance and make it compatible with water (some turbidity is acceptable). .

Second, as the substrate, at least a part of the surface thereof (the part to which the film of the molecule of the amphipathic molecule or the bitter substance which is a main part of the taste sensor is attached) has the amphipathic substance or the bitter substance. A substance (a substance having properties to be referred to as a hydrophilic-hydrophobic group property) which is close to (well binds to) the hydrophobic group of the substance is attached thereto. For example, a glass plate immersed in stearic acid is a substrate that satisfies the conditions. Further, a rod or plate made of polyvinyl chloride containing a plasticizer is also a substrate satisfying the conditions.

When a substrate having a surface that is familiar with the hydrophobic group of the amphipathic substance or bitter substance is immersed in the aqueous solution of the amphipathic substance or bitter substance formed in the first step, the amphiphilic substance (ie, , A substance having a molecular structure having a hydrophobic part and a hydrophilic part) or a part of the bitter substance having a hydrophobic property adheres to the surface of the substrate prepared in the second stage. Thus, a uniform layer of the amphipathic molecule group or the bitter substance molecule group was formed on the substrate.

[0016]

The amphiphilic molecular layer or the molecular layer of the bitter substance on the substrate prepared in accordance with the procedure described in the preceding paragraph has a property of being friendly to the hydrophobic group of the amphiphilic substance or the bitter substance attached to the surface of the substrate ( (Hereinafter referred to as a hydrophilic-hydrophobic group) substance (for example, stearic acid) and a hydrophobic group of an amphipathic molecule or a bitter substance, so that a group of amphiphilic molecules formed on the substrate is formed. It is considered that the layer or the layer of the molecular group of the bitter substance has its hydrophilic groups exposed on the surface and is uniformly arranged. In other words, it seems to be a neat arrangement like a monolayer of an amphipathic substance or a bitter substance, and observation with an electron microscope suggests that.

[0017]

DESCRIPTION OF THE PREFERRED EMBODIMENTS The amphipathic molecule referred to in the present invention is a molecule having a hydrophobic part and a hydrophilic part, and examples of substances (amphiphilic substances) comprising the molecular groups are as shown in Table 1. is there.

[0018]

[Table 1]

The bitter substance is a substance exhibiting a bitter taste, examples of which are shown in Table 2.

[0020]

[Table 2]

FIG. 2 shows an embodiment of a method of manufacturing a taste sensor according to the present invention (an embodiment of the manufacturing method according to claim 2). That is, FIG. 2 shows a flow of the manufacturing method. In the uppermost step 5 on the left side, a solution in which an amphipathic molecule group or a bitter substance molecule group is dissolved in alcohol is dissolved in water to form an amphipathic substance or bitter substance. Prepare an aqueous solution of the substance (may have some turbidity) or prepare a solution of amphiphilic molecules or molecules of bitter substances dissolved in an aqueous solution of a strong alkali (eg, caustic soda). On the other hand, a substrate is made with a film based on a familiar substance that is familiar to the hydrophobic groups of the amphipathic or bitter substance molecules. As a specific example, in step 6, a solution is prepared by dissolving 200 mg of polyvinyl chloride (PVC) and 250 μl of dioctyl phthalate (DOP) as a plasticizer in tetrahydrofuran (THF) as a solvent. In step 7, it is heated at 30 ° C. on a flat bottom petri dish,
The THF is volatilized to form a base film serving as a substrate. Alternatively, stearic acid is prepared in step 6, and the glass substrate is immersed in stearic acid and taken out, substituting step 7 to prepare a glass substrate having a base film of stearic acid on the surface. The base film thus formed has a property that it is easily compatible with the hydrophobic group of the amphiphilic substance or the bitter substance because the hydrophilic / hydrophobic group is exposed on the surface. Step 8
Then, the substrate with a base film obtained in step 7 is immersed in the aqueous solution or suspension of the amphipathic substance or bitter substance obtained in step 5, and left for 3 days. Thereafter, it is taken out, and the amphipathic molecules or molecules of the bitter substance attached to unnecessary portions on the substrate surface are washed away with a weak alkaline solution (step 9).
This manufacturing method is suitable for mass production because the substrate with the base film may be immersed in an aqueous solution or suspension.

FIG. 3 shows another embodiment (an embodiment of the manufacturing method according to claim 3) of the manufacturing method of the taste sensor of the present invention. FIG. 3 shows a flow of the manufacturing method. In the uppermost step 10, an amphipathic substance or a bitter substance is dropped on the water surface to form a monomolecular film of an amphipathic molecule group or a bitter substance molecule group on the water surface. I do. On the other hand, a substrate is made with a film based on a familiar substance that is familiar to the hydrophobic groups of the amphipathic or bitter substance molecules. As a specific example, in step 6, a solution is prepared by dissolving 200 mg of polyvinyl chloride (PVC) and 250 μl of dioctyl phthalate (DOP) as a plasticizer in tetrahydrofuran (THF) as a solvent. In step 7, it is heated at 30 ° C. on a flat-bottomed petri dish,
By evaporating F, a base film serving as a substrate is formed. Alternatively, stearic acid is prepared in step 6, and the glass substrate is immersed in stearic acid and taken out, substituting step 7 to prepare a glass substrate having a base film of stearic acid on the surface. The base film thus formed has a property that it is easily compatible with the hydrophobic group of the amphiphilic substance or the bitter substance because the hydrophilic / hydrophobic group is exposed on the surface. Step 11
Then, as schematically shown in FIG. 4, the substrate with the base film obtained in the step 7 is transferred to the monomolecular film of the amphipathic molecule group or the bitter substance molecule group on the water surface obtained in the step 10 by the water surface. To form a monomolecular film of an amphipathic molecule group or a bitter substance molecule group on the surface of the base film, or, as schematically shown in FIG. By moving the contact point between the monomolecular film formed on the substrate and the substrate, a monomolecular film of an amphipathic molecule group or a bitter substance molecule group is formed on the surface of the base film. Next, in step 9, the amphipathic molecules or the molecules of the bitter substance attached to the unnecessary portions on the substrate surface are washed away with a weak alkaline solution.

The film of the amphipathic molecule or the molecule of the bitter substance on the substrate thus formed, as described in the section of action,
Since the hydrophobic groups (hydrophobic sites) of the molecules constituting the film are laid on the surface of the substrate, the hydrophilic groups (hydrophilic sites) of the amphipathic molecules or the molecules of the bitter substance group are arranged outward,
It can be inferred that a single layer is formed.

According to the expression method used in the design method of chemicals, this pattern is formed by a hydrophobic group extending in the longitudinal direction like a tail and a hydrophilic group indicated by a circle on a part of the atomic arrangement extending in the longitudinal direction. FIG. 1 schematically shows an example of an amphipathic molecule represented as As shown in the figure, since the base film 2 having a hydrophilic / hydrophobic group is formed on the upper surface of the glass substrate 1, the group of the amphiphilic molecules 4 fixed to the base film 2
A hydrophobic group extending in the longitudinal direction like a tail is attracted to the base film 2, and the hydrophilic group indicated by a triangle is aligned with the face on the upper surface as indicated by 3. On the other hand, the glass surface without the hydrophilic / hydrophobic base film 2 (lower surface, side surface)
Then the arrangement of the amphipathic molecules is disturbed.

Thus, the substrate with the base film was formed,
An example of processing a single layer of amphipathic molecules (or made directly on the base film itself as a substrate) into a taste sensor is shown in FIG. In the figure, 1, 2, 3,
3 'is a single layer of a substrate, a base film, and a group of amphipathic molecules as in the example of FIG. Unnecessary part 1
After removing the amphipathic molecules 2 and 12 ', conductive rods (for example, gold) 14 and 14' are inserted into the holes 13 and 13 'formed in the substrate 1, and one end of the rod is attached to the base film 2. , And solder the lead wires 15 and 15 'to the other end. Thus, the potential of the membrane of the amphipathic molecule can be measured. Alternatively, if the 12 'removed as an unnecessary portion is not removed, the electric conductivity (resistance) between the lead wires 15 and 15' can be measured. Since the membrane potential or the electrical conductivity has a relationship corresponding to the concentration of the substance exhibiting the basic taste, a taste sensor has been realized.

Next, the method of an experiment conducted using the taste sensor manufactured as described above and the data obtained as a result will be described.

FIGS. 7 and 8 show the taste sensor of the present invention and a conventional taste sensor using sodium chloride (NaC).
The relationship between the concentration of 1) and the membrane potential was examined.

The experiment was performed on the taste sensor of 5 samples each according to the following procedures. Immerse the taste sensor in 1 mM potassium chloride solution. After about 4 minutes, the membrane potential is measured, and the value is used as a reference. While the taste sensor is immersed in the solution, sodium chloride is added to make a solution of potassium chloride 1 mM and sodium chloride 1 mM. After about 4 minutes, the membrane potential is measured, and the difference from the reference value is determined. Hereinafter, the concentration of sodium chloride in the procedure was 3 m.
M, 10 mM, 30 mM, 100 mM, 300 mM, 1000 mM
And repeat the procedure. At this time, if the concentration of sodium chloride is changed, the concentration of potassium chloride decreases due to an increase in the amount of the solution. Therefore, potassium chloride is replenished to maintain 1 mM.

As can be seen from FIGS. 7 and 8, the sensitivity of the taste sensor of the present invention is about 5 times higher than that of the conventional taste sensor between 1 mM and 10 mM, and 1 mM to 100 mM.
Approximately 2 fold, and 1.5 fold between 1 mM and 1000 mM. Less variation between taste sensors. Good log linearity. It can be said that. The result is sensitive, consistent quality,
Since the characteristics can be represented by a simple straight line, it indicates that a practical taste sensor has been obtained.

FIGS. 9 and 10 show the relationship between the concentration of hydrochloric acid (HCl) and quinine and the membrane potential of the taste sensor of the present invention and the conventional taste sensor, respectively.
All taste sensors use dioctyl phosphate, a kind of amphiphile, as a material.

The experiment was performed according to the following procedures. Immerse the taste sensor in 1 mM potassium chloride solution. After about 4 minutes, the membrane potential is measured, and the value is used as a reference. With the taste sensor immersed in the solution, hydrochloric acid (quinine in FIG. 10) was added, and potassium chloride 1 mM and hydrochloric acid (FIG. 1) were added.
0 means quinine) 0.001 mM solution. After about 4 minutes, the membrane potential is measured, and the difference from the reference value is determined. Hereinafter, hydrochloric acid in the procedure (quinine in FIG. 10)
Concentration of 0.003 mM, 0.01 mM, 0.03 mM, 0.1 mM, 0.3 mM,
Change to 1 mM, 3 mM, 10 mM, 30 mM, 100 mM,
Repeat the above steps. At this time, if the concentration of hydrochloric acid (quinine in FIG. 10) is changed, the concentration of potassium chloride decreases due to an increase in the amount of the solution. Therefore, potassium chloride is replenished to maintain 1 mM.

FIG. 9 shows that the sensitivity of hydrochloric acid does not change much, but the logarithmic linearity between 0.1 mM and 100 mM is improved. Also, from FIG. 10, it can be seen that the sensitivity of quinine is approximately doubled between 0.01 mM and 3 mM. In FIG. 10, the output of the taste sensor is saturated in a portion where the concentration is higher than 3 mM, which indicates the same phenomenon that the intensity of bitterness is saturated in human taste.

FIGS. 11 and 12 show that the taste sensor of the present invention and the conventional taste sensor are immersed in a 1M sodium chloride solution.
It is a result of examining a temporal change of a membrane potential. The measurement of the membrane potential was performed at intervals of about 20 seconds.

From these results, as described above, the taste sensor of the present invention has a small change in the membrane potential in spite of the fact that the sensitivity is about 1.5 times as compared with the conventional taste sensor. In particular, the change between 0 and 10 minutes is 以下 or less. The changes (curves) are aligned. It can be said that. This result indicates that the taste sensor of the present invention is stable, and has three times the stability when used for short-time measurement, and has a constant quality.

FIG. 13 shows the response of the taste sensor of the present invention to five basic tastes. In the figure, ch1
-Ch8 are taste sensors using different amphiphilic substances or bitter substances, respectively. Table 3 shows the amphiphilic substances or bitter substances used.

[0036]

[Table 3]

The experiment was carried out by using five taste sensors of 8 channels and using the following procedure for a solution of 0.1 mM hydrochloric acid, 100 mM sodium chloride, 0.3 mM quinine, 1 M sucrose and 30 mM MSG for each measurement object. Immerse the taste sensor in 1 mM potassium chloride solution. After about 4 minutes, the membrane potential is measured, and the value is used as a reference. While the taste sensor is immersed in the solution, the concentration of the measurement target is set to the target concentration. After about 4 minutes, the membrane potential of each channel is measured, and the difference from the reference value is determined. At this time, if the concentration of the measurement object is changed, the concentration of potassium chloride decreases due to an increase in the amount of the solution. Therefore, potassium chloride is replenished to maintain 1 mM.

It should be noted that the response of the taste sensor of ch5 to sucrose is large. This is because phosphatidylcholine (PC), which is a kind of lipid constituting the biological membrane, was available and the orientation was improved. Ch3 uses quinine, one of the bitter substances, as a material for the taste sensor.
When viewed from the whole of h1 to ch8, a taste sensor which shows a strong response to hydrochloric acid, a moderate response to sodium chloride and MSG, and a weak response to quinine and sucrose to obtain taste information It can be seen that In the present invention,
Not only phosphatidylcholine (PC) but also the range of substances that can be used as taste sensors has been expanded, so that the amount of information obtained about taste has increased.

FIGS. 14 and 15 show a taste sensor using phosphatidylcholine (PC) as a material, and examined the relationship between the concentrations of sodium chloride and sucrose and the membrane potential.

The experiment was performed according to the following procedures. Immerse the taste sensor in 1 mM potassium chloride solution. After about 4 minutes, the membrane potential is measured, and the value is used as a reference. While the taste sensor is immersed in the solution, sodium chloride (sucrose in FIG. 15) is added to obtain a solution of 1 mM potassium chloride and 1 mM sodium chloride (sucrose in FIG. 15). After about 4 minutes, the membrane potential is measured, and the difference from the reference value is determined. Hereinafter, the concentration of sodium chloride (sucrose in FIG. 15) in the procedure was 3 mM, 10 mM, 30 mM, 100 mM, and 3 mM.
The procedure is repeated, changing to 00 mM and 1000 mM. At this time, if the concentration of sodium chloride is changed, the concentration of potassium chloride decreases due to an increase in the amount of the solution. Therefore, potassium chloride is replenished to maintain 1 mM.

From FIG. 14, it can be said that the concentration of sodium chloride can be determined by measuring the membrane potential. FIG.
From the results, it can be seen that the concentration of sucrose cannot be determined from the membrane potential unless the concentration is higher than 300 mM. In view of the fact that humans do not feel sweet unless the concentration is considerably high, it can be said that the taste sensor of the present invention is close to human taste.

[0042]

According to the present invention, as a taste sensor,
We discovered a new method of forming a single layer of the amphipathic molecule group and the bitter substance molecule group and realized a single layer based on the formation method.
Sensitivity was improved, measurement stability was obtained, and there was little variation in measured values among the sensors. In addition, a sensor having logarithmic linearity was obtained, so that a taste sensor approaching human taste could be realized.

Further, according to the present invention, it is possible to use a lipid, an amphipathic substance other than lipid, and a bitter substance, which are difficult to use in the conventional method, as a taste sensor, so that taste can be sensed. The increased amount of information and, in particular, the availability of lipids that make up biological membranes have enabled the realization of a taste sensor that is closer to human taste.

[0044]

[Brief description of the drawings]

FIG. 1 is a diagram schematically showing a taste sensor of the present invention.

FIG. 2 is a diagram showing a flow of an embodiment of the manufacturing method of the present invention.

FIG. 3 is a diagram showing a flow of another embodiment of the manufacturing method of the present invention.

FIG. 4 is a view schematically showing one method for forming a monomolecular film of an amphipathic molecule group or a bitter substance molecule group on a substrate with a base film.

FIG. 5 is a diagram schematically showing another method for forming a monomolecular film of an amphipathic molecule group or a bitter substance molecule group on a substrate with a base film.

FIG. 6 is a diagram showing a processing example for extracting a membrane potential from the taste sensor of the present invention.

FIG. 7 is a graph showing concentration characteristics of the taste sensor of the present invention with respect to sodium chloride.

FIG. 8 is a graph showing a concentration characteristic of a conventional taste sensor with respect to sodium chloride.

FIG. 9 is a diagram showing concentration characteristics of hydrochloric acid of the taste sensor of the present invention and a conventional taste sensor.

FIG. 10 is a graph showing density characteristics of a taste sensor of the present invention and a conventional taste sensor with respect to quinine.

FIG. 11 is a diagram showing a time characteristic of the taste sensor of the present invention with respect to sodium chloride.

FIG. 12 is a diagram showing a time characteristic of a conventional taste sensor with respect to sodium chloride.

FIG. 13 is a diagram showing responses of the taste sensor of the present invention to five basic tastes.

FIG. 14 is a graph showing concentration characteristics of the taste sensor of the present invention with respect to sodium chloride.

FIG. 15 is a graph showing the concentration characteristics of the taste sensor of the present invention with respect to sucrose.

[Explanation of symbols]

 DESCRIPTION OF SYMBOLS 1 Substrate (glass substrate) 2 Base film 3 Amphipathic molecule group or bitter substance molecule group 4 Amphiphilic molecule or bitter substance molecule 5 Step 5 6 Step 6 7 Step 7 8 Step 8 9 Step 9 10 Step 10 11 Step 11

Continued on the front page (72) Inventor Kaoru Yamafuji 1-6-21 Kusakae, Chuo-ku, Fukuoka City, Fukuoka Prefecture (72) Inventor Kiyoshi Toko 2-83-2, Miwadai, Higashi-ku, Fukuoka City, Fukuoka Prefecture (72) Inventor Kenji Hayashi 2-14-18-407 Takatori, Sawara-ku, Fukuoka City, Fukuoka Prefecture (72) Inventor Hidekazu Ikezaki 5-10-27 Minamiazabu, Minato-ku, Tokyo Inside Anritsu Corporation (72) Inventor Rieko Higashikubo Tokyo 5-10-27 Minamiazabu, Miyako-ku, Japan An Examiner at Anritsu Corporation Jun Koriyama (56) References JP-A-62-294085 (JP, A) Kenji Hayashi, Kiyoshi Toko and Kaoru Yamafuji, BIO INDUSTRY, No. Vol. 7, No. 4, 1990 (pp. 268-274) Satoru Iiyama, Kiyoshi Toko and Kaoru Yamafuji, MEMBRANE, Vol. 12, No. 4, (1987), pp. 231-237 (58) (Int.Cl. ⁷ , DB name) G01N 27/327 G01N 27/02 G01N 27/416

Claims (1)

(57) [Claims] (1) using a solvent that dissolves in water to remove bitter substances;
Amphiphiles having molecules or hydrophobic and hydrophilic sites
The step of preparing an aqueous solution or suspension of a soluble molecule group (5)
And a substance whose surface is at least partially hydrophobic.
Providing a covered substrate (6, 7);
Immersing the board in said aqueous or suspended water (8)
Manufacturing method of taste sensor.