Method for screening lipid deterioration odor suppressing materials, lipid deterioration odor suppressing agent, method for producing food and beverages containing the agent, and food and beverages containing the agent

Abstract

To provide a method for screening lipid deterioration odor suppressing materials, a lipid deterioration odor suppressing agent, a method for producing food and beverages containing the agent, and food and beverages containing the agent.SOLUTION: Provided is a method for screening lipid deterioration odor suppressing materials, comprising the following steps (1) to (3): (1) bringing a test substance and a lipid deterioration odor causative substance into contact with an olfactory receptor polypeptide that contains an amino acid sequence of OR2 J3 and / or an amino acid sequence that has at least 90% identity with the amino acid sequence of OR2 J3 and is responsive to the lipid deterioration odor causative substance; (2) measuring a response of the olfactory receptor polypeptide to the lipid deterioration odor causative substance; and (3) identifying a test substance that suppresses the response of the olfactory receptor polypeptide based on the measured response.SELECTED DRAWING: Figure 7

Description

[Technical field]

[0001] The present invention relates to a screening method for a lipid deterioration odor suppressing material suitable for reducing or masking off-flavors (unpleasant odors that deviate from the inherent odor of the food or beverage) in foods, beverages, etc., a lipid deterioration odor suppressing agent, a method for producing foods and beverages containing the agent, and foods and beverages containing the agent. [Background technology]

[0002] In mammals such as humans, odors are recognized when odorant molecules bind to olfactory receptors on olfactory nerve cells in the olfactory epithelium in the upper part of the nasal cavity, and the response of the receptor to the molecules is transmitted to the central nervous system. There are approximately 400 types of olfactory receptors in humans, and a single odorant can cause multiple olfactory receptors to respond, making the olfactory receptor response to the actual smell of food and drink complex. In response to such complexity, there is a demand in the food and beverage industry for methods to discover materials that can selectively reduce or mask off-flavors. In addition, OR2J3, one of the above-mentioned human olfactory receptor proteins, is known to detect cis-3-hexen-1-ol, which has a green leaf odor (Non-Patent Document 1).

[0003] Conventionally, screening of agents that reduce or mask off-flavors has been carried out by humans evaluating the off-flavor reduction or masking effect of test substances through sensory testing, but sensory testing has problems such as the need to train experts who can evaluate odors and low throughput.

[0004] In recent years, as an alternative to evaluating odors using the human sense of smell, evaluation techniques have been developed that use evaluation cells in which olfactory receptor (OR) genes have been introduced and the olfactory receptors are expressed on the cell surface. For example, a screening method for odor-suppressing materials has been reported using the olfactory receptor OR2L3, which responds to odor-causing substances (Patent Document 1).

[0005] Incidentally, in foods and beverages containing a large amount of oil, the odor of deteriorated lipids acts as an off-flavor and is one of the factors that impair the palatability of the foods and beverages. Several components are known to be causative of the above-mentioned lipid deterioration odor, including, for example, fat and oil oxides such as linear or branched, saturated or unsaturated aliphatic alcohols or aldehydes, in particular (E,E)-2,4-decadienal (hereinafter simply referred to as "(E,E)-2,4-decadienal" or "decadienal"). [Prior art documents] [Patent documents]

[0006] [Patent Document 1] Patent Publication No. 2022-25044 [Non-patent literature]

[0007] [Non-Patent Document 1] Jeremy F.McRae,et.al.,Chem.Scenses,37,p585-p593,2012 Summary of the Invention [Problem to be solved by the invention]

[0008] The present invention has been made in consideration of the above-mentioned problems in the prior art, and aims to provide a method for screening lipid-deterioration odor suppressing materials, a lipid-deterioration odor suppressing agent, a method for producing food and beverages containing the agent, and food and beverages containing the agent. [Means for solving the problem]

[0009] As a result of intensive research into the above-mentioned problems, the present inventors have newly discovered that the human olfactory receptor OR2J3 and its variants respond to various substances causing lipid deterioration odors. In particular, they have newly discovered that the human olfactory receptor OR2J3 and its variants respond to (E,E)-2,4-decadienal, which is known as a substance causing lipid deterioration odors. They have also newly discovered a substance (a lipid deterioration odor suppressing material) that suppresses the response of OR2J3 to (E,E)-2,4-decadienal. The present invention has been completed based on the above findings. That is, the present invention is as follows.

[0010] <1> A method for screening a lipid deterioration odor suppressing material, comprising the following steps (1) to (3): (1) contacting a test substance and the causative substance of a lipid deterioration odor with an olfactory receptor polypeptide that contains the amino acid sequence of OR2J3 and / or an amino acid sequence having at least 90% identity to the amino acid sequence of OR2J3 and is responsive to the causative substance of a lipid deterioration odor; (2) measuring the response of the olfactory receptor polypeptide to the lipid deterioration odor causative substance; and (3) identifying a test substance that inhibits the response of the olfactory receptor polypeptide based on the measured response; the lipid deterioration odor causative substance is (E,E)-2,4-decadienal, heptanal, octanal, nonanal, decanal, undecanal, dodecanal, (E,E)-2,4-hexadienal ((E,E)-2,4-hexadienal), (E,E)-2,4-heptadienal ((E,E)-2,4-heptadienal), (E,E)-2,4-oct the compound is at least one selected from the group consisting of (E,E)-2,4-octadienal, (E,E)-2,4-nonadienal ((E,E)-2,4-nonadienal), (E,Z)-2,6-nonadienal ((E,Z)-2,6-nonadienal), (E,E)-2,4-undecadienal ((E,E)-2,4-undecadienal), (E)-2-nonenal ((E)-2-nonenal), (E)-2-decenal ((E)-2-decenal), and (E,E)-2,4-decadienol ((E,E)-2,4-decadienol). <2> The lipid deterioration odor causative substance is (E,E)-2,4-decadienal; <1> The method described above. <3> The amino acid sequence of the OR2J3 is an amino acid sequence set forth in any one of SEQ ID NOs: 1 to 5 in the sequence listing. <1> The method described above. <4> A lipid deterioration odor inhibitor containing at least one lipid deterioration odor inhibitor material selected from the group consisting of allyl isothiocyanate, dimethylbenzylcarbonyl butyrate, sec-butyl isothiocyanate, indole, and 3,4-dimethyl-1,2-cyclopentanedione. <5> the above <1> A lipid deterioration odor inhibitor produced by a method comprising the method according to claim 2, or <4> A method for producing a food or drink containing the lipid deterioration odor inhibitor described in claim 1. <6> the above <4> A food or drink containing the lipid deterioration odor inhibitor described in claim 1. Effect of the Invention

[0011] According to the present invention, it is possible to provide a method for screening a lipid deterioration odor suppressing material suitable for reducing or masking off-flavors, a lipid deterioration odor suppressing agent, a method for producing a food or beverage containing the agent, and a food or beverage containing the agent. [Brief description of the drawings]

[0012] [Figure 1] (a) and (b) show the response strength of receptors OR2J3 and OR51E1, respectively, to various concentrations of (E,E)-2,4-decadienal. [Diagram 2] 1(a) to (f) are diagrams showing the response strength of the receptor OR2J3 to various substances causing lipid deterioration odors at various concentrations. [Diagram 3] 1(a) to 1(i) are diagrams showing the response strength of the receptor OR2J3 to various substances causing lipid deterioration odors at various concentrations. [Figure 4] FIG. 1 shows the response strength of receptor OR2J3 to various concentrations of conventional masking agents. [Diagram 5] FIG. 1 shows the effect of attenuating the response intensity of receptor OR2J3 by various concentrations of a conventional masking agent. [Figure 6] This figure shows the OR2J3 response (AUC value) when (E,E)-2,4-decadienal was further added to the above test substances (220 flavoring raw materials that can be used in food), and the relative AUC value % is shown when the positive control (containing only (E,E)-2,4-decadienal) is indicated as 100%. [Figure 7] FIG. 13 is a diagram showing the results of a sensory evaluation test (cotton ball) of the lipid deterioration odor suppression effect of a new lipid deterioration odor suppression material against (E,E)-2,4-decadienal. [Figure 8] FIG. 13 is a diagram showing the results of a sensory evaluation test (with ion-exchanged water) of the lipid deterioration odor suppression effect of a new lipid deterioration odor suppression material against (E,E)-2,4-decadienal. [Figure 9] FIG. 13 shows the results of a sensory evaluation test (lemon carbonated water) of the lipid deterioration odor suppression effect of a new lipid deterioration odor suppression material against (E,E)-2,4-decadienal. [Figure 10] FIG. 1 shows the results of measuring the response strength of various haplotypes (Hap1 to Hap5) of OR2J3 to (E,E)-2,4-decadienal using a luciferase assay method. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0013] The following describes in detail the embodiments of the present invention. However, the present invention is not limited to the following embodiments and can be implemented with appropriate modifications within the scope of the object of the present invention.

[0014] <Screening method for lipid deterioration odor suppressing materials> A first aspect of the present invention is a method for screening a lipid-deterioration odor suppressing material, comprising the following steps (1) to (3): (1) contacting a test substance and the causative substance of a lipid deterioration odor with an olfactory receptor polypeptide that contains the amino acid sequence of OR2J3 and / or an amino acid sequence having at least 90% identity to the amino acid sequence of OR2J3 and is responsive to the causative substance of a lipid deterioration odor; (2) measuring the response of the olfactory receptor polypeptide to the lipid deterioration odor causative substance; and (3) identifying a test substance that inhibits the response of the olfactory receptor polypeptide based on the measured response. In the first embodiment, screening means at least narrowing down the population of test substances, and it is preferable to select candidate lipid-deterioration odor-inhibiting materials. There are no particular limitations on the lipid deterioration odor suppression material as long as it can reduce the off-flavor lipid deterioration odor, but lipid deterioration odor suppression materials that can inhibit the binding of lipid deterioration odor-causing substances with the olfactory receptor polypeptide are preferred, and lipid deterioration odor suppression materials that are antagonists of the olfactory receptor polypeptide and have a masking effect due to the antagonist are more preferred.

[0015] (Lipid deterioration odor causing substance) The above-mentioned lipid deterioration odor causative substances are substances that cause lipid deterioration odors, and specifically, (E,E)-2,4-decadienal, heptanal, octanal, nonanal, decanal, undecanal, dodecanal, (E,E)-2,4-hexadienal ((E,E)-2,4-hexadienal), (E,E)-2,4-heptadienal ((E,E)-2,4-heptadienal), (E,E)-2,4-octadienal ((E,E)-2,4-octadienal), (E,E)-2,4-octadienal ...), (E,E)-2,4-octadienal ((E,E)-2,4-octadienal), (E,E)-2,4-octadienal), (E,E)-2,4-octadienal, (E,E)-2,4-octadienal), (E,E)-2,4-octadienal, (E,E)-2,4-octadienal), (E,E)-2,4-octadienal, (E,E)-2,4-octadienal), (E,E)-2,4-octadienal, (E,E)-2,4-octadienal), At least one selected from the group consisting of (E,E)-2,4-nonadienal, (E,Z)-2,6-nonadienal, (E,E)-2,4-undecadienal, (E)-2-nonenal, (E)-2-decenal, and (E,E)-2,4-decadienol is preferred, with (E,E)-2,4-decadienal being more preferred.

[0016] (Test substance) The test substance applied to the method of the present invention as a candidate material for inhibiting the odor of lipid deterioration may be either a gaseous or liquid substance. The test substance is not particularly limited. The test substance may be composed of a single component (i.e., a pure substance) or may be composed of a combination of two or more components (i.e., a mixture). When the test substance is a mixture, the number of types and composition ratio of the components constituting the mixture are not particularly limited. The test substance may be a known substance or a new substance. The test substance may be a natural product or an artificial product. The test substance may be, for example, a compound library prepared using combinatorial chemistry technology. Examples of the test substance include alcohols, ketones, aldehydes, ethers, esters, hydrocarbons, sugars, organic acids, nucleic acids, amino acids, peptides, lipids, and various other organic or inorganic components. Test substances may include various kinds of flavoring raw materials, synthetic flavors, natural flavors, natural essential oils, plant extracts, and the like, such as the natural essential oils, natural flavors, flavoring raw materials, and synthetic flavors described in "Japan Patent Office Gazette, Collection of Well-Known and Commonly Used Technology (Flavors) Part II Food Flavors, pp. 88-131, published January 14, 2000." Test substances also include existing food additives. "Existing food additives" refers to substances that are approved for use as food additives. The test substance may be one type of test substance, or two or more types of test substances may be used in combination. By bringing two or more components into contact with a receptor protein together and carrying out the method of the present invention, it is possible to identify whether the combination of components as a whole is a lipid-deterioration odor suppressing material. Examples of "contacting two or more components together with an olfactory receptor protein" include contacting a test substance that is a mixture with an olfactory receptor protein and contacting two or more test substances together with an olfactory receptor protein.

[0017] (Olfactory Receptor Polypeptides) In the present invention, at least one olfactory receptor polypeptide selected from the group consisting of olfactory receptor polypeptides comprising the amino acid sequence of OR2J3 and olfactory receptor polypeptides comprising an amino acid sequence having at least 90% identity with the amino acid sequence of OR2J3 and having responsiveness to substances causing lipid deterioration odor is used, and it is preferable to use an olfactory receptor polypeptide comprising the amino acid sequence of OR2J3. "OR2J3 protein" refers to a human olfactory receptor protein classified as Olfactory Receptor Family 2 Subfamily J Member 3. OR2J3 protein is also referred to simply as "OR2J3." The gene encoding OR2J3 protein is also referred to as the "OR2J3 gene." An "olfactory receptor polypeptide that contains an amino acid sequence that is at least 90% identical to the amino acid sequence of OR2J3 and is responsive to substances that cause lipid deterioration odor" preferably has a sequence identity of 93% or more, more preferably 95% or more, even more preferably 97% or more, and particularly preferably 98% or more. In this specification, the sequence identity of the amino acid sequence can be calculated using the BLAST search algorithm (publicly available from NCBI).

[0018] It is known that various variants (variants; for example, haplotypes) exist in the human olfactory receptor OR2J3 (for example, Non-Patent Document 1). The main haplotypes of OR2J3 are summarized in Table 1 below.

[0019] [Table 1]

[0020] In the present invention, the amino acid sequence of OR2J3 is preferably an amino acid sequence set forth in any one of SEQ ID NOs: 1 to 5 in the sequence listing. In the present invention, the olfactory receptor polypeptide may be one type of the olfactory receptor polypeptide, or two or more types of the olfactory receptor polypeptides may be used in combination.

[0021] As used herein, a gene encoding an olfactory receptor polypeptide is also referred to as an "olfactory receptor polypeptide gene," and a gene encoding an olfactory receptor protein is also referred to as an "olfactory receptor gene." In this specification, the term "gene" is not limited to DNA and may include any polynucleotide as long as it codes for a protein of interest. The olfactory receptor polypeptide gene (preferably, the olfactory receptor gene) may be DNA, RNA, or a combination thereof. The olfactory receptor polypeptide gene (preferably, the olfactory receptor gene) may be single-stranded DNA or single-stranded RNA. The olfactory receptor gene may be double-stranded DNA, double-stranded RNA, or a hybrid chain consisting of a DNA chain and an RNA chain. The olfactory receptor polypeptide gene (preferably, the olfactory receptor gene) may contain both DNA residues and RNA residues in a single polynucleotide chain. When the olfactory receptor polypeptide gene (preferably, the olfactory receptor gene) contains RNA, the description of DNA such as the exemplified base sequence above may be appropriately interpreted according to RNA. The olfactory receptor polypeptide gene (preferably, the olfactory receptor gene) may or may not contain an intron. The form of the olfactory receptor polypeptide gene (preferably, the olfactory receptor gene) can be appropriately selected depending on various conditions such as the form of use.

[0022] The amino acid sequences of the human olfactory receptor proteins and the nucleotide sequences of the human olfactory receptor genes can be obtained from public databases such as NCBI and Ensembl. Furthermore, the olfactory receptor polypeptide may contain other amino acid sequences in addition to the amino acid sequence of the human olfactory receptor protein. That is, the olfactory receptor polypeptide may be a fusion protein of the amino acid sequence of the human olfactory receptor protein and other amino acid sequences. The other amino acid sequences are not particularly limited as long as they do not impair the responsiveness of the human receptor protein to substances causing lipid deterioration odors. Examples of the other amino acid sequences include tag sequences such as His tag and V5 epitope tag. The other amino acid sequences may be linked to, for example, the N-terminus or C-terminus, or both, of the human olfactory receptor protein.

[0023] The responsiveness of the olfactory receptor polypeptide to substances causing lipid deterioration odors can be confirmed, for example, by measuring the response of the olfactory receptor polypeptide when the olfactory receptor polypeptide is contacted with substances causing lipid deterioration odors. Furthermore, the inhibition of the lipid deterioration odor by the test substance can be confirmed by measuring the reduction in the response of the above-mentioned olfactory receptor polypeptide to substances that cause the lipid deterioration odor.

[0024] The olfactory receptor polypeptide can be used in any form that can be used for screening lipid deterioration odor suppressing materials. Specifically, the olfactory receptor polypeptide can be used in any form as long as it can contact lipid deterioration odor causative substances and test substances and does not lose responsiveness to lipid deterioration odor causative substances. The form of use of the olfactory receptor polypeptide can be appropriately set according to various conditions such as the embodiment of the first aspect.

[0025] The olfactory receptor polypeptide may be used in a form isolated to a desired degree, such as a purified product or a crude product, or may be used in a form contained in any material. Specifically, the olfactory receptor polypeptide may be used in a form supported by a structure. Examples of the structure include cells, cell membranes, artificial lipid bilayer vesicles, and artificial lipid bilayer membranes. Examples of the structure include cells. In other words, the olfactory receptor polypeptide may be used in the form of a structure having (supporting) an olfactory receptor protein, such as a cell having the olfactory receptor polypeptide, a cell membrane having the olfactory receptor polypeptide, an artificial lipid bilayer vesicle having the olfactory receptor polypeptide, or an artificial lipid bilayer membrane having the olfactory receptor polypeptide.

[0026] The olfactory receptor polypeptide can be produced, for example, by expressing the olfactory receptor polypeptide gene. The expression of the olfactory receptor polypeptide gene may be carried out, for example, using cells or a cell-free protein synthesis system. For the expression of the olfactory receptor polypeptide gene using cells, the following description of cells having an olfactory receptor polypeptide can be referred to. The expressed olfactory receptor polypeptide can be obtained appropriately in the form described above and used in the first embodiment.

[0027] The cells having the above-mentioned olfactory receptor polypeptide may be cells that inherently have an olfactory receptor gene, or may be cells that have been modified to have the above-mentioned olfactory receptor polypeptide gene.

[0028] Cells that inherently have an olfactory receptor gene include human cells from which the above-mentioned olfactory receptor gene is derived, such as human olfactory cells, etc. Cells that inherently have an olfactory receptor gene can be obtained, for example, from an organism or tissue that contains the cell. Cells modified to have the olfactory receptor polypeptide gene include cells into which the olfactory receptor polypeptide gene has been introduced.

[0029] In addition, cells having the above-mentioned olfactory receptor polypeptide and cells used to obtain them (e.g., cells into which the above-mentioned olfactory receptor polypeptide gene is introduced or has been introduced) are collectively referred to below simply as "host cells".

[0030] The host cell is not particularly limited as long as it can express the functional olfactory receptor polypeptide and can be used for screening of lipid-degraded odor suppressing materials. Examples of the host cell include bacterial cells, fungal cells, plant cells, insect cells, and animal cells. Examples of the preferred host cell include eukaryotic cells such as fungal cells, plant cells, insect cells, and animal cells. Examples of the more preferred host cell include animal cells. Examples of the animal include mammals, birds, and amphibians. Examples of the mammal include rodents and primates. Examples of the rodents include Chinese hamsters, hamsters, mice, rats, and guinea pigs. Examples of the primates include humans, monkeys, and chimpanzees. Examples of the birds include chickens. Examples of the amphibians include Xenopus laevis. In addition, the tissue or cell from which the host cell is derived is not particularly limited.

[0031] Olfactory receptor genes can be obtained by cloning from organisms that have the olfactory receptor genes. For cloning, nucleic acids such as genomic DNA and cDNA that contain the genes can be used. Olfactory receptor genes can also be obtained by chemical synthesis (Gene, 60(1), 115-127(1987)).

[0032] The obtained olfactory receptor gene can be used as is or after appropriate modification. That is, a variant of the olfactory receptor gene can be obtained by modifying the olfactory receptor gene. Gene modification can be performed by known techniques. For example, a target mutation can be introduced into a target site of DNA by site-directed mutagenesis. That is, for example, a coding region of a gene can be modified by site-directed mutagenesis so that the encoded protein contains substitution, deletion, insertion, and / or addition of amino acid residues at a specific site. A variant of the olfactory receptor gene can also be obtained directly by chemical synthesis.

[0033] The form in which the olfactory receptor polypeptide gene is introduced into the host cell is not particularly limited. The olfactory receptor polypeptide gene may be expressibly retained in the host cell. Specifically, for example, when the olfactory receptor polypeptide gene is introduced in a form that requires transcription of DNA or the like, the olfactory receptor polypeptide gene may be expressibly retained in the host cell under the control of a promoter that functions in the host cell. The promoter for expressing the olfactory receptor polypeptide gene is not particularly limited as long as it functions in the host cell. "Promoter that functions in the host cell" refers to a promoter that has promoter activity in the host cell. The promoter may be a promoter derived from the host cell or a promoter derived from a heterologous species.

[0034] The olfactory receptor polypeptide gene can be introduced into a host cell, for example, using a vector containing the gene. The above vector can be appropriately selected depending on various conditions such as the type of host cell, the mode of introduction of the olfactory receptor gene, etc. The method for introducing the vector into the host cell can be appropriately selected depending on various conditions such as the type of host cell. The olfactory receptor polypeptide may or may not include the N-terminal 20 amino acid residues of mammalian (e.g., human, bovine) rhodopsin (Rho-tag). By incorporating the N-terminal 20 amino acid residues of mammalian rhodopsin, the cell membrane expression of the olfactory receptor polypeptide can be promoted. As an expression vector encoding the N-terminal 20 amino acid residues of human rhodopsin (Rho-tag), the pCI expression vector (Promega) is commercially available.

[0035] In addition, the cell having the olfactory receptor polypeptide may have, for example, a protein involved in signal transduction. In other words, the cell may have a gene encoding a protein involved in signal transduction. Examples of proteins involved in signal transduction include G proteins (e.g., Golf), G protein activators (e.g., Ric8B), adenylate cyclase, and calcium channels. Examples of Golf include animal Golf, such as human Golf (GenBank accession No. NP_892023). Examples of Ric8B include animal Ric8B, such as rat Ric8B (GenBank accession No. NP_783188). Furthermore, the cell having the olfactory receptor polypeptide may have, for example, a component corresponding to a parameter to be measured. Examples of such a component include a probe such as a calcium indicator and a reporter gene such as a luciferase gene. When a probe such as a calcium indicator is expressed from a gene, the cell may have a gene encoding the probe.

[0036] The cell having the olfactory receptor polypeptide may have, for example, a protein that promotes membrane expression of the olfactory receptor polypeptide. In other words, the cell may have a gene that encodes such a protein. Such a protein may include, for example, RTP1s (Zhuang H and Matsunami H, J Biol Chem 282, 15284-15293 (2007)). Examples of RTP1s include animal RTP1s such as human RTP1s (GenBank accession No. AAT70680), mouse RTP1s (GenBank accession No. ABU23737), and bat RTP1s (amino acid sequence from the methionine residue at position 37 to the C-terminus of GenBank accession No. XP_006765914). The host cell is preferably a Hana3A cell.

[0037] (Step (1) of contacting the test substance and the lipid deterioration odor causative substance with the olfactory receptor polypeptide) The above-mentioned olfactory receptor polypeptide can be contacted with the test substance and the lipid-degraded odor-causing substance in any liquid phase atmosphere (e.g., water, aqueous buffer solution, culture medium atmosphere, etc.) or in any gas phase atmosphere (e.g., air atmosphere), and the contact can be carried out via any liquid phase or gas phase. Methods for contacting in any liquid phase atmosphere include causing the olfactory receptor polypeptide to coexist with the gaseous or liquid test substance and the lipid deterioration odor causative substance in the same liquid phase. A method for contacting the olfactory receptor polypeptide in any gaseous atmosphere (for example, in an air atmosphere) includes placing the above-mentioned olfactory receptor polypeptide together with a gaseous or liquid (preferably volatile) test substance and a lipid deterioration odor causative substance in the same container (preferably an airtight container).

[0038] The contact conditions can be appropriately set according to various conditions such as the form of use of the olfactory receptor polypeptide, the type of test substance, and the method of measuring the response of the olfactory receptor polypeptide. For example, known reaction conditions for measuring interactions between substances such as interactions between proteins and ligands may be used as they are or may be appropriately modified. The concentration of the lipid deterioration odor causative substance may be, for example, 0.01 μM to 1 mM, 0.1 μM to 500 μM, or 1 μM to 400 μM. The concentration of the test substance may be, for example, 0.01 μM to 1 mM, 0.1 μM to 500 μM, or 1 μM to 400 μM, or 0.001 μg / mL to 70 μg / mL, 0.01 μg / mL to 50 μg / mL, or 0.1 μg / mL to 40 μg / mL. The concentration of the olfactory receptor polypeptide may be, for example, 1 pg / mL to 10 mg / mL. In addition, when using cells having the olfactory receptor polypeptide, the concentration of the cells having the olfactory receptor protein may be, for example, 10 cells / mL to 10,000,000 cells / mL. The contact of the olfactory receptor polypeptide with the test substance and the lipid deterioration odor causative substance may or may not be terminated at an appropriate time point.

[0039] The contact of the olfactory receptor polypeptide with the test substance and the lipid deterioration odor causative substance is preferably continued during the measurement of the response of the olfactory receptor polypeptide to the lipid deterioration odor causative substance (and the test substance). The duration of the contact of the olfactory receptor polypeptide with the test substance and the lipid deterioration odor causative substance may be, for example, 0.1 seconds or more, 0.5 seconds or more, 1 second or more, 5 seconds or more, 10 seconds or more, 30 seconds or more, 1 minute or more, 5 minutes or more, 10 minutes or more, 30 minutes or more, 1 hour or more, or 2 hours or more, or 24 hours or less, 12 hours or less, 6 hours or less, 2 hours or less, or 1 hour or less, or a compatible combination thereof. The duration of the contact of the olfactory receptor polypeptide with the test substance and the lipid deterioration odor causative substance may be, for example, 15 minutes to 3 hours. The contact system may contain other components in addition to the olfactory receptor polypeptide (for example, in the form of cells having the olfactory receptor polypeptide as exemplified above), the test substance, and the lipid deterioration odor causative substance, as long as it is possible to screen lipid deterioration odor suppressing materials. The other components can be appropriately set depending on various conditions such as the form of use of the olfactory receptor polypeptide, the type of test substance, and the method of measuring the response of the olfactory receptor polypeptide.

[0040] (Step (2) of measuring the response of the olfactory receptor polypeptide to substances causing lipid deterioration odor) In the step (2) of measuring the response of the olfactory receptor polypeptide to a substance causing an odor of lipid deterioration, the response of the olfactory receptor polypeptide to the substance causing an odor of lipid deterioration is preferably measured in the coexistence of a test substance. The method for measuring the response of the olfactory receptor polypeptide to the lipid deterioration odor causative substance is not particularly limited. The method for measuring the response of the olfactory receptor protein to the lipid deterioration odor causative substance can be appropriately selected according to various conditions such as the form of use of the olfactory receptor polypeptide and the type of response to be measured. The above-mentioned response includes the binding of the above-mentioned olfactory receptor polypeptide to a substance causing a lipid deterioration odor, or the activation of the above-mentioned olfactory receptor polypeptide by a substance causing a lipid deterioration odor.

[0041] The method for measuring the binding between the olfactory receptor polypeptide and the causative substance of the odor of deteriorated lipids (and the test substance) is not particularly limited. The binding between the olfactory receptor polypeptide and the causative substance of the odor of deteriorated lipids (and the test substance) can be measured by a known method for measuring the binding between substances, such as the binding between a protein and a ligand, or the inhibition of the binding by a competitive inhibitor. Examples of such methods include isothermal titration calorimetry (ITC), surface plasmon resonance (SPR), and nuclear magnetic resonance (NMR) fluorescence correlation spectroscopy (FCS).

[0042] The method for measuring the activation of the olfactory receptor polypeptide by the lipid degradation odor causative substance is not particularly limited. The activation of the olfactory receptor polypeptide by the lipid degradation odor causative substance can be measured by a known method for measuring the activity of a receptor such as an olfactory receptor. Examples of such methods include an intracellular calcium amount measurement method and an intracellular cAMP amount measurement method. That is, the activation of the olfactory receptor polypeptide by the lipid degradation odor causative substance can be measured, for example, using the intracellular calcium amount or the intracellular cAMP amount as an indicator. For example, in Hana3A cells or HEK293T cells, it is known that when the olfactory receptor polypeptide is activated by the lipid degradation odor causative substance, it couples with intracellular G protein (Golf, etc.) to activate adenylate cyclase, thereby increasing the intracellular cAMP amount (Kajiya K. et al., Molecular bases of odor discrimination: Reconstitution of olfactory receptors that recognize overlapping sets of odorants. Journal of Neuroscience, 2001, 21:6018-6025). Examples of the method for measuring the amount of intracellular cAMP include ELISA and reporter assay. Examples of the reporter assay include luciferase assay. According to the reporter assay, the amount of intracellular cAMP can be measured by using a reporter gene (luciferase gene, etc.) that is configured to be expressed depending on the amount of cAMP. Examples of the method for measuring the amount of intracellular calcium include calcium imaging. According to the calcium imaging, the amount of intracellular calcium can be measured by using a calcium indicator. Examples of the calcium indicator include calcium-sensitive fluorescent dyes and calcium-sensitive fluorescent proteins.In addition, even when the receptor protein is used in a form having an internal space such as an artificial lipid bilayer vesicle, and the artificial lipid bilayer vesicle is configured so that the calcium amount or cAMP amount in the internal space varies like a cell, the activation of the receptor protein by the test substance can be measured by the same method as above. In that case, the "intracellular calcium amount" and the "intracellular cAMP amount" can be read as the calcium amount and the cAMP amount in the internal space, respectively.

[0043] In addition, by measuring any parameter, it can be used as an index for measuring the response of the olfactory receptor polypeptide to the lipid deterioration odor causative substance (and test substance). In other words, as long as the response can be measured (i.e., it can be determined whether the response is observed), it is sufficient to obtain and use data reflecting the parameter, and it is not necessary to obtain the value of the parameter itself. In other words, when data reflecting the parameter is obtained, it is not necessary to calculate the value of the parameter itself from the data. Specifically, for example, when the amount of intracellular cAMP is measured by luciferase assay and used as an index for measuring the activation of the olfactory receptor polypeptide by the lipid deterioration odor causative substance (and test substance), it is sufficient to obtain and use data reflecting the amount of intracellular cAMP (e.g., fluorescence intensity) as long as the activation can be measured (i.e., it can be determined whether the activation is observed), and it is not necessary to calculate the amount of intracellular cAMP itself from the data.

[0044] The timing for measuring the response of the olfactory receptor polypeptide to the lipid deterioration odor causative substance (and test substance) is not particularly limited as long as the response of the olfactory receptor polypeptide to the lipid deterioration odor causative substance (and test substance) occurs to a measurable extent. Specifically, the timing for measuring the response of the olfactory receptor polypeptide to the lipid deterioration odor causative substance may be any appropriate time from the start of contact between the olfactory receptor polypeptide and the lipid deterioration odor causative substance to the disappearance of the response of the olfactory receptor polypeptide to the lipid deterioration odor causative substance (and test substance). The timing for measuring the response of the olfactory receptor polypeptide to the lipid deterioration odor causative substance (and test substance) may be, for example, immediately after the start of contact between the olfactory receptor polypeptide and the lipid deterioration odor causative substance, 1 second after the start of contact, 10 seconds after, 30 seconds after, 1 minute after, 5 minutes after, 10 minutes after, 30 minutes after, 1 hour after, or 2 hours after, or up to 24 hours, up to 12 hours, up to 6 hours, up to 2 hours, or up to 1 hour, or any combination thereof that is not contradictory. The timing for measuring the response of the olfactory receptor polypeptide to the lipid deterioration odor causative substance (and test substance) may be, for example, immediately after the start of contact between the olfactory receptor polypeptide and the lipid deterioration odor causative substance (and test substance) to 3 hours.

[0045] (Step (3) of identifying a test substance that inhibits the response of the olfactory receptor polypeptide based on the response) Next, based on the response of the olfactory receptor polypeptide to the lipid-deterioration odor causative substance (and the test substance), it is possible to identify whether the test substance is a substance that reduces the response of the olfactory receptor polypeptide, and when the suppressive response is observed, it is preferable to identify the test substance as a candidate lipid-deterioration odor suppressing material. It is preferable to identify the olfactory receptor polypeptide using as an index the response intensity S1 (preferably the activation intensity S1) of the olfactory receptor polypeptide when the olfactory receptor polypeptide is contacted with a test substance (and a substance causing lipid-degraded odor), and it is more preferable to identify the olfactory receptor polypeptide by comparing the response intensity S1 with the response intensity S2 under control conditions. Here, the response intensities S1, S2, etc. may or may not be standardized values ​​obtained by dividing the response intensity by the response intensity measured in the absence of the olfactory receptor polypeptide.

[0046] Here, the "control condition" refers to a condition in which the olfactory receptor polypeptide is not contacted with the test substance, and more specifically, it is preferable to contact the olfactory receptor polypeptide with the lipid-degraded odor causative substance, but not with the test substance (condition in the absence of the test substance). In other words, it means that the difference in the intensity of the response of the olfactory receptor polypeptide depending on the presence or absence of the test substance is used as an index. The intensity S2 of the response under the control conditions may be a measured value obtained by contacting the olfactory receptor polypeptide with a substance causing a lipid deterioration odor before the addition of the test substance, or it may be a statistical value or range obtained in advance by collecting data from contacting the olfactory receptor polypeptide with a substance causing a lipid deterioration odor.

[0047] More specifically, the above identification is based on the difference between the intensity S1 of the response and the intensity S2 of the response under control conditions, and if intensity S1 is smaller than intensity S2, the test substance can be identified as a substance that reduces the response of the olfactory receptor polypeptide by a masking effect or the like (preferably, a candidate lipid-degradation odor suppressing material). The degree of reduction in the response intensity is preferably a statistically significant reduction, more preferably a reduction of 3 / 4 or less compared to the control condition, even more preferably a reduction of 3 / 5 or less, particularly preferably a reduction of 1 / 2 or less, particularly preferably a reduction of 1 / 3 or less, and most preferably a reduction of 1 / 4 or more. The lower limit of the degree of reduction is not particularly limited, but examples include 1 / 30 or more, 1 / 20 or more, and 1 / 10 or more.

[0048] The first embodiment may or may not include a step of measuring the intensity S2. The intensities S1 and S2 may be measured in a single reaction system with a time lag, or may be measured simultaneously or with a time lag in separate reaction systems. An example of the ratio of intensity S1 to intensity S2 is the S1 value standardized by intensity S2.

[0049] The first aspect may further include a step of evaluating the substance (preferably, the candidate lipid-degraded odor suppressing material) that reduces the response of the olfactory receptor polypeptide. That is, by evaluating the identified substance (preferably, the candidate lipid-degraded odor suppressing material) that reduces the response of the olfactory receptor polypeptide, it is possible to confirm whether the substance (preferably, the candidate lipid-degraded odor suppressing material) actually suppresses lipid-degraded odor by a masking effect or the like. There are no particular limitations on the method of evaluating the identified substance (preferably, the candidate lipid-degraded odor suppressing material) that reduces the response of the olfactory receptor polypeptide, and it can be evaluated by a known method such as evaluation by a sensory test, for example.

[0050] <Odor suppressant for lipid deterioration> A second aspect of the present invention is a lipid-deterioration odor inhibitor, comprising a lipid-deterioration odor inhibitor material obtained by the screening method according to the first aspect. The lipid deterioration odor inhibitor according to the second aspect preferably contains at least one lipid deterioration odor inhibitor material selected from the group consisting of allyl isothiocyanate, dimethylbenzylcarbonyl butyrate, sec-butyl isothiocyanate, indole, and 3,4-dimethyl-1,2-cyclopentanedione.

[0051] The lipid deterioration odor inhibitor may contain any component as long as it does not impair the lipid deterioration odor inhibitory effect. As the optional component, any component that is acceptable depending on the application of the lipid deterioration odor inhibitor, such as an orally ingestible component, may be used. As the optional component, for example, a component that is blended with seasonings, foods and beverages, or medicines may be used. Specific examples of the optional component include sugars such as sugar, honey, maple syrup, sucrose, glucose, fructose, isomerized sugar, and oligosaccharides; sugar alcohols such as xylitol and erythritol; high-intensity sweeteners; inorganic salts such as salt, sodium chloride, and potassium chloride; organic acids such as acetic acid and citric acid and their salts; amino acids such as glutamic acid and glycine and their salts; nucleic acids such as inosinic acid, guanylic acid, and xanthylic acid and their salts; dietary fiber, pH buffers, excipients, bulking agents, flavorings, edible oils, ethanol, and water. As the optional component, one component may be used, or two or more components may be used in combination.

[0052] <<Method of manufacturing food and beverage products, and food and beverage products>> A third aspect of the present invention is a method for producing a food or drink containing a lipid deterioration odor inhibitor, including a lipid deterioration odor inhibitor material obtained by the screening method according to the first aspect, which preferably includes adding the lipid deterioration odor inhibitor containing the lipid deterioration odor inhibitor material obtained by the screening method according to the first aspect to any food or drink, It is more preferable that the method includes adding a lipid deterioration odor inhibitor containing at least one lipid deterioration odor inhibitor selected from the group consisting of allyl isothiocyanate, dimethylbenzylcarbonyl butyrate, sec-butyl isothiocyanate, indole, and 3,4-dimethyl-1,2-cyclopentanedione to any food or beverage. By adding the lipid deterioration odor inhibitor to any food or drink, the lipid deterioration odor can be suppressed.

[0053] Furthermore, a fourth aspect of the present invention is a food or beverage containing a lipid deterioration odor inhibitor, including a lipid deterioration odor inhibitor material obtained by the screening method according to the first aspect, and any food or beverage containing a lipid deterioration odor inhibitor containing at least one lipid deterioration odor inhibitor material selected from the group consisting of allyl isothiocyanate, dimethylbenzylcarbonyl butyrate, sec-butyl isothiocyanate, indole, and 3,4-dimethyl-1,2-cyclopentanedione is preferred. Specific examples of food and beverages include beverages such as carbonated drinks, soft drinks, fruit juice drinks, dairy drinks, lactic acid bacteria drinks, energy drinks, soy milk, and tea drinks; alcoholic beverages such as chuhai, cocktail drinks, sparkling wine, fruit liquor, and condiment liquor; desserts such as ice cream, ice milk, lacto ice cream, frozen desserts, yogurt, pudding, jelly, and daily desserts; sweets such as caramel, candy, tablet candy, crackers, biscuits, cookies, pies, chocolate, snacks, and chewing gum; soups such as Japanese-style soup and Western-style soup; jams; flavor seasonings; various instant beverages; and various instant foods. In foods and beverages that contain oil (particularly foods and beverages that contain a lot of oil), the odor of deteriorated lipids acts as an off-flavor and is one factor that impairs the palatability of the foods and beverages, so the foods and beverages according to the fourth aspect are preferable. For example, conventional lemon water, lemon carbonated water, etc., sometimes contain lemon peel oil, which can cause a lipid deterioration odor. In response to this, the lipid deterioration odor can be suppressed by adding the lipid deterioration odor inhibitor.

[0054] The amount of the lipid deterioration odor inhibitor added to a food or beverage varies depending on the type and form of the product, but can be, for example, within a concentration range of 0.05 to 500 ppb, preferably 0.1 to 300 ppb, more preferably 0.5 to 100 ppb, and even more preferably 1 to 50 ppb, based on the mass of the food or beverage before the lipid deterioration odor inhibitor is added. EXAMPLES

[0055] The present invention will be described in more detail below with reference to examples thereof. However, the present invention is not limited to these examples, and various applications are possible within the scope of the technical concept of the present invention.

[0056] <Method for measuring olfactory receptor response> (1) Cloning of olfactory receptor genes Each of the 388 types of human olfactory receptors was obtained by cloning from Human Genomic DNA:Female (Promega) by PCR based on the sequence information registered in GenBank. Each gene amplified by PCR was inserted into a pCI expression vector (Promega) encoding the N-terminal 20 amino acid residues of human rhodopsin (Rho-tag), to obtain a human olfactory receptor gene expression vector.

[0057] (2) Luciferase assay (Day 1: Cell seeding) Hana3A cells were seeded in each well at 50 μL at a cell number that would result in confluence after 48 hours. (Day 2: Introduction of olfactory receptor gene) Human olfactory receptor gene expression vector 80ng, RTP1s vector 5ng, pGloSensor TM 10 ng of -22F plasmid (Promega) and 0.2 μL of ViaFect (registered trademark) Transfection Reagent (Promega) were dissolved in 10 μL of D-MEM (Fujifilm Wako Pure Chemical Industries, Ltd.) to prepare a gene solution (for one well), which was then left to stand in a clean bench for 15 minutes. The gene solution after standing was replaced with the culture medium of Hana3A cells 24 hours after cell seeding, and the gene was introduced into the cells. After gene transfer, the cells were cultured at 37° C. under 5% CO 2 for 24 hours.

[0058] (Day 3: Luciferase assay) Using the GloSensor cAMP assay (Promega), changes in cAMP concentration caused by receptors activated by binding to ligands were measured in real time according to the product manual. After removing the culture medium, 25 μL of HBSS(+) (Fujifilm Wako Pure Chemical Industries, Ltd.) buffer solution (containing 10 mM HEPES, 1 mM glucose, 0.75 mM CuCl2, and GloSensor cAMP Reagent) was added and equilibrated at room temperature for 2 hours in the dark.

[0059] After equilibration, the sample, a substance causing the odor of deteriorated lipids ((E,E)-2,4-decadienal), was diluted with HBSS(+) buffer to a test concentration, and 25 μL was added to each well. The plate was quickly set in a luminometer, and the luminescence intensity derived from firefly luciferase immediately after stimulation was measured for 15 cycles at 120 s intervals. All luminescence values ​​were divided by the luminescence value at the same cycle number of cells transfected with the Rho-pCI vector (vector control: empty vector), and ROC analysis was performed to calculate the response intensity as AUC (area under the curve) value.

[0060] <Screening and identification of olfactory receptors that respond to (E,E)-2,4-decadienal> Using the above cells expressing 388 types of human olfactory receptors, the receptor response to 10 μM of (E,E)-2,4-decadienal was measured by the above luciferase assay method (n=2). We again prepared cells expressing the 31 types of human olfactory receptors that showed an AUC value of 1 or more, and then measured the receptor response strength to various concentrations (0 / 1 / 10 / 100 μM) of (E,E)-2,4-decadienal using a luciferase assay (n=3). The measurement results showing the above responses are shown in Figure 1(a) and (b). The values ​​in the figures are the average values ​​of n=3. As shown in Figure 1(a) and (b), OR2J3 and OR51E1 respond to (E,E)-2,4-decadienal in a concentration-dependent manner. OR2J3 was more responsive to (E,E)-2,4-decadienal than OR51E1, and OR2J3 was finally identified as the human olfactory receptor that responds to (E,E)-2,4-decadienal.

[0061] <Measurement of the response of the olfactory receptor OR2J3 to various substances causing the odor of deteriorated lipids> Human olfactory receptor OR2J3 expressing cells were prepared, and the response strength of the receptor OR2J3 to various concentrations of heptanal (0 / 0.1 / 1 / 3 / 10 / 30 / 100 / 300 μM) was measured as an AUC value by luciferase assay (n=3). The measurement results showing the above responses are shown in Figure 2(a). The values ​​in the figure are the average values ​​of n=3, and Rho-pCI is the measurement result of the vector control (empty vector). The response intensity of the receptor OR2J3 was measured as an AUC value by luciferase assay in the same manner except that the lipid deterioration odor causative substance was changed from heptanal to octanal, nonanal, decanal, undecanal, dodecanal, (E,E)-2,4-hexadienal, (E,E)-2,4-heptadienal, (E,E)-2,4-octadienal, (E,E)-2,4-nonadienal, (E,Z)-2,6-nonadienal, (E,E)-2,4-undecadienal, (E)-2-nonenal, (E)-2-decenal, or (E,E)-2,4-decadienol (n=3). The measurement results showing the above responses are shown in Figures 2(b)-(f) and 3(a)-(i). The values ​​in the figure are the average values ​​of n=3.

[0062] As is clear from the results shown in Figures 2(a) to (f) and Figures 3(a) to (i), OR2J3 responds to heptanal, octanal, nonanal, decanal, undecanal, dodecanal, (E,E)-2,4-hexadienal, (E,E)-2,4-heptadienal, (E,E)-2,4-octadienal, (E,E)-2,4-nonadienal, (E,Z)-2,6-nonadienal, (E,E)-2,4-undecadienal, (E)-2-nonenal, (E)-2-decenal, or (E,E)-2,4-decadienol in a concentration-dependent manner. In other words, OR2J3 was identified as a human olfactory receptor that responds to lipid degradation odors such as not only (E,E)-2,4-decadienal, but also heptanal, octanal, nonanal, decanal, undecanal, dodecanal, (E,E)-2,4-hexadienal, (E,E)-2,4-heptadienal, (E,E)-2,4-octadienal, (E,E)-2,4-nonadienal, (E,Z)-2,6-nonadienal, (E,E)-2,4-undecadienal, (E)-2-nonenal, (E)-2-decenal, and (E,E)-2,4-decadienol.

[0063] <Confirmation of the in vitro lipid deterioration odor masking effect of conventional masking agents on (E,E)-2,4-decadienal> (1) Using a conventional masking agent whose masking effect against (E,E)-2,4-decadienal has been confirmed by sensory evaluation, receptor response intensities against various concentrations (0 / 0.1 / 1 / 10 / 30 μM) of the conventional masking agent were measured by the luciferase assay method described above in cells expressing OR2J3 (specifically, Hap4 OR2J3 represented by SEQ ID NO: 4) (n=3). The results are shown in Figure 4. The values ​​in the figure are the average values ​​of n=3. For comparison, FIG. 4 also shows the receptor response to various concentrations of (E,E)-2,4-decadienal. As is clear from the results shown in FIG. 4, it was confirmed that OR2J3 showed a high response only to (E,E)-2,4-decadienal, and showed a weak response or no response to the above-mentioned conventional masking agents.

[0064] (2) Next, the attenuation effect (masking effect) of the above-mentioned conventional masking agents on the receptor response to (E,E)-2,4-decadienal was evaluated. The concentration of (E,E)-2,4-decadienal at this time was determined by the EC 50 Concentrations close to the concentration of (E,E)-2,4-decadienal were used, i.e., the response of OR2J3 to 3 μM (E,E)-2,4-decadienal was used as the blank. The above conventional masking agents were co-administered with (E,E)-2,4-decadienal at molar concentration ratios of 1 / 10 (0.3 μM), 1x (3 μM), 3x (9 μM), 10x (30 μM), and 20x (60 μM), and the cell response was measured by the above luciferase assay method (n=3). The results are shown in Figure 5. The values ​​in the figure are the average values ​​of n=3. As is clear from the results shown in FIG. 5, the intensity of the OR2J3 response was attenuated to 51% by co-addition of the conventional masking agent at a molar concentration 20-fold (60 μM) higher than that of the OR2J3-containing agent. As is clear from the above results, the response of OR2J3 is elicited by stimulation with (E,E)-2,4-decadienal, the intensity of the response is reduced in a concentration-dependent manner by co-addition of the above-mentioned conventional masking agents, and the above-mentioned conventional masking agents have been confirmed to have a masking effect on (E,E)-2,4-decadienal by sensory evaluation, suggesting that OR2J3 is a receptor involved in the recognition of the lipid deterioration odor caused by (E,E)-2,4-decadienal.

[0065] Furthermore, as is clear from the results shown in Figure 5, OR2J3 is highly responsive to (E,E)-2,4-decadienal, and is suitable for screening new lipid deterioration odor suppressing materials (new masking materials). The following screening was carried out using the response intensity of OR2J3 as an indicator of the receptor involved in the recognition of the lipid deterioration odor caused by (E,E)-2,4-decadienal.

[0066] <Screening of new lipid deterioration odor suppressing materials against lipid deterioration odor causative substance ((E,E)-2,4-decadienal)> As candidates for new lipid degradation odor suppression materials (new masking materials), 200 compounds used as fragrance ingredients and 20 natural essential oils were listed and used as test substances in the following screening. Hana3A cells expressing OR2J3 were equilibrated with 25 μL of HBSS(+) buffer for 2 hours. After equilibration, 12.5 μL of HBSS(+) buffer solution was added to each well, either as a blank (containing neither (E,E)-2,4-decadienal nor the above test substances) or to each well containing 400 μM of the 200 compounds or 40 μg / mL of the 20 natural essential oils as test substances.

[0067] In addition, 12.5 μL of HBSS(+) buffer containing 40 μM (E,E)-2,4-decadienal was added to each well. A blank and a positive control (also simply called "Posicon") were also provided. The plate was quickly placed in the luminometer and measured again for 10 cycles at 120 s intervals. The results are shown in Figure 6. Figure 6 shows the OR2J3 response (AUC value) when (E,E)-2,4-decadienal was further added to the above test substances (220 flavoring raw materials that can be used in food), and the relative AUC value % is shown when the positive control below (containing only (E,E)-2,4-decadienal) is represented as 100%. In FIG. 6, the blank does not contain (E,E)-2,4-decadienal or the above test substances (the 220 flavoring raw materials). The positive control did not contain the test substances (the 220 fragrance raw materials) and contained only (E,E)-2,4-decadienal.

[0068] As is clear from the results shown in FIG. 6, allyl isothiocyanate had a relative AUC value of 27%, Dimethyl benzyl carbinyl butyrate (2-methyl-1-phenyl-2-propyl butyrate) had a relative AUC value of 30%. Sec-butyl isothiocyanate has a relative AUC value of 30%. Indole has a relative AUC value of 23%. 3,4-dimethyl-1,2-cyclopentanedione had a relative AUC value of 34%, significantly reducing the OR2J3 response, and these five compounds were selected as new lipid degradation odor suppression materials (new masking materials).

[0069] <Sensory evaluation test of the lipid deterioration odor suppression effect of a new lipid deterioration odor suppression material against the lipid deterioration odor causative substance ((E,E)-2,4-decadienal)> (1) Sensory evaluation test (cotton balls) A cotton ball with a diameter of 1 cm was placed in a 20 cc glass vial, and 10 μL of 0.1% by mass (E,E)-2,4-decadienal in ethanol and 10 μL of 0.01 to 0.1% by mass of the new lipid deterioration odor suppression material (new masking material) in ethanol were impregnated into the vial, and the vial was then capped and allowed to equilibrate for 1 hour. After equilibration, the lid was opened and the nose was brought close to the container, and the odor intensity of (E,E)-2,4-decadienal was evaluated on a scale of 0 to 10 for each item shown in Table 2 below. The odor intensity of (E,E)-2,4-decadienal was defined as follows, and the panel was informed of this in advance before participating in the evaluation. The odor intensity of (E,E)-2,4-decadienal alone was rated as 8, and the odor intensity of (E,E)-2,4-decadienal when mixed with the new masking material was rated from 0 (not noticeable) to 10 (very strong). The samples were presented in the form of three-digit random numbers, and the order of presentation was random. The results are shown in Figure 7 as the average of n=25.

[0070] [Table 2]

[0071] As is clear from the cotton ball test results shown in Figure 7, allyl isothiocyanate, dimethylbenzylcarbinylbutyrate, sec-butyl isothiocyanate, indole, and 3,4-dimethyl-1,2-cyclopentanedione all have the effect of suppressing (masking effect) the odor of lipid deterioration ((E,E)-2,4-decadienal odor). In addition, allyl isothiocyanate, dimethylbenzylcarbinylbutyrate, sec-butyl isothiocyanate, indole, and 3,4-dimethyl-1,2-cyclopentanedione are all similar in that they respond not only to (E,E)-2,4-decadienal but also to the human olfactory receptor OR2J3 and its variants, i.e., to heptanal, octanal, nonanal, decanal, undecanal, dodecanal, and (E It is presumed that this has an effect of suppressing (masking effect) the odor of lipid deterioration produced by compounds such as (E,E)-2,4-hexadienal, (E,E)-2,4-heptadienal, (E,E)-2,4-octadienal, (E,E)-2,4-nonadienal, (E,Z)-2,6-nonadienal, (E,E)-2,4-undecadienal, (E)-2-nonenal, (E)-2-decenal, and (E,E)-2,4-decadienol.

[0072] (2) Sensory evaluation test (ion-exchanged water) 10 μL of 0.01% by mass (E,E)-2,4-decadienal in ethanol and 10 μL of 0.001 to 0.1% by mass new lipid deterioration odor suppressing material (new masking material) in ethanol were added to 200 mL of ion-exchanged water and stirred thoroughly. The sample was taken in a mouthful and then immediately swallowed, and the odor intensity of (E,E)-2,4-decadienal as it escaped through the nose was evaluated on a scale of 0 to 10 for each item shown in Table 3 below. The odor intensity of (E,E)-2,4-decadienal was defined as follows, and the panel was informed of this in advance before participating in the evaluation. The odor intensity of (E,E)-2,4-decadienal alone was rated as 8, and the odor intensity of (E,E)-2,4-decadienal when mixed with the new masking material was rated from 0 (not noticeable) to 10 (very strong). The samples were presented in three-digit random numbers, and the order of presentation was random. The results are shown in Figure 8 as the average value of n = 24-26.

[0073] [Table 3]

[0074] As is clear from the test results for ion-exchanged water shown in Figure 8, allyl isothiocyanate, dimethylbenzylcarbinylbutyrate, sec-butyl isothiocyanate, indole, and 3,4-dimethyl-1,2-cyclopentanedione all have the effect of suppressing (masking effect) the odor of lipid deterioration ((E,E)-2,4-decadienal odor). In addition, allyl isothiocyanate, dimethylbenzylcarbinylbutyrate, sec-butyl isothiocyanate, indole, and 3,4-dimethyl-1,2-cyclopentanedione are all similar in that they respond not only to (E,E)-2,4-decadienal but also to the human olfactory receptor OR2J3 and its variants, i.e., to heptanal, octanal, nonanal, decanal, undecanal, dodecanal, and (E It is presumed that this has an effect of suppressing (masking effect) the odor of lipid deterioration produced by compounds such as (E,E)-2,4-hexadienal, (E,E)-2,4-heptadienal, (E,E)-2,4-octadienal, (E,E)-2,4-nonadienal, (E,Z)-2,6-nonadienal, (E,E)-2,4-undecadienal, (E)-2-nonenal, (E)-2-decenal, and (E,E)-2,4-decadienol.

[0075] (3) Sensory evaluation test (lemon carbonated water) To 200 mL of commercially available lemon carbonated water (sparkling lemon natural water), 10 μL of 0.01 mass% (E,E)-2,4-decadienal in ethanol and 10 μL of 0.01 to 0.1 mass% new lipid deterioration odor suppression material (new masking material) in ethanol were added and stirred well. The concentration of the new masking material was set to an optimal concentration based on the results of the sensory evaluation in the above (2) ion-exchanged water test. The sample was taken in a mouthful and then immediately swallowed, and the odor intensity of (E,E)-2,4-decadienal as it escaped through the nose was evaluated on a scale of 0 to 10 for each item shown in Table 4 below. The odor intensity of (E,E)-2,4-decadienal was defined as follows, and the panel was informed of this in advance before participating in the evaluation. The odor intensity of (E,E)-2,4-decadienal alone was rated as 8, and the odor intensity of (E,E)-2,4-decadienal when mixed with the new masking material was rated from 0 (not noticeable) to 10 (very strong). The samples were presented in the form of three-digit random numbers, and the order of presentation was random. The results are shown in Figure 9 as the average value of n=23.

[0076] [Table 4]

[0077] As is clear from the test results of ion-exchanged water shown in Figure 9, allyl isothiocyanate, dimethylbenzylcarbinylbutyrate, sec-butyl isothiocyanate, indole, and 3,4-dimethyl-1,2-cyclopentanedione all have the effect of suppressing (masking effect) the odor of lipid deterioration ((E,E)-2,4-decadienal odor). In addition, allyl isothiocyanate, dimethylbenzylcarbinylbutyrate, sec-butyl isothiocyanate, indole, and 3,4-dimethyl-1,2-cyclopentanedione are all similar in that they respond not only to (E,E)-2,4-decadienal but also to the human olfactory receptor OR2J3 and its variants, i.e., to heptanal, octanal, nonanal, decanal, undecanal, dodecanal, and (E It is presumed that this has an effect of suppressing (masking effect) the odor of lipid deterioration produced by compounds such as (E,E)-2,4-hexadienal, (E,E)-2,4-heptadienal, (E,E)-2,4-octadienal, (E,E)-2,4-nonadienal, (E,Z)-2,6-nonadienal, (E,E)-2,4-undecadienal, (E)-2-nonenal, (E)-2-decenal, and (E,E)-2,4-decadienol.

[0078] From the above results, it can be said that the screening method of the present invention, which measures the polypeptide response of the olfactory receptor OR2J3, is capable of screening new lipid deterioration odor suppression materials that suppress lipid deterioration odor ((E,E)-2,4-decadienal odor). Furthermore, the screening method of the present invention, which measures the response of the polypeptide of the olfactory receptor OR2J3, can be said to be capable of screening for novel lipid deterioration odor suppression materials that suppress lipid deterioration odors, such as not only (E,E)-2,4-decadienal, but also heptanal, octanal, nonanal, decanal, undecanal, dodecanal, (E,E)-2,4-hexadienal, (E,E)-2,4-heptadienal, (E,E)-2,4-octadienal, (E,E)-2,4-nonadienal, (E,Z)-2,6-nonadienal, (E,E)-2,4-undecadienal, (E)-2-nonenal, (E)-2-decenal, and (E,E)-2,4-decadienol, which have in common that they respond to the human olfactory receptor OR2J3 and its variants.

[0079] In addition, allyl isothiocyanate, dimethylbenzylcarbonyl butyrate, sec-butyl isothiocyanate, indole, and 3,4-dimethyl-1,2-cyclopentanedione are all new lipid deterioration odor suppression materials that can suppress lipid deterioration odor ((E,E)-2,4-decadienal odor) and can be used as lipid deterioration odor suppressants. In addition, allyl isothiocyanate, dimethylbenzylcarbonyl butyrate, sec-butyl isothiocyanate, indole, and 3,4-dimethyl-1,2-cyclopentanedione are all new lipid degradation odor suppressing materials, and are similar to (E,E)-2,4-decadienal, but also to heptanal, octanal, nonanal, decanal, undecanal, dodecanal, and other olfactory odor suppressants that respond to the human olfactory receptor OR2J3 and its variants. It can be seen that the lipid deterioration odor suppressant can be used as a lipid deterioration odor suppressant, for example, (E,E)-2,4-hexadienal, (E,E)-2,4-heptadienal, (E,E)-2,4-octadienal, (E,E)-2,4-nonadienal, (E,Z)-2,6-nonadienal, (E,E)-2,4-undecadienal, (E)-2-nonenal, (E)-2-decenal, and (E,E)-2,4-decadienol. It can also be said that the addition of allyl isothiocyanate, dimethylbenzylcarbonyl butyrate, sec-butyl isothiocyanate, indole, or 3,4-dimethyl-1,2-cyclopentanedione to food or beverages (e.g., water, lemon carbonated water) can suppress the odor of deteriorated lipids in the food or beverage.

[0080] <Evaluation of the response of various variants (e.g., haplotypes) of the human olfactory receptor OR2J3 to the odor of lipid degradation ((E,E)-2,4-decadienal odor)> Cells expressing various haplotypes (Hap1 to Hap5) of OR2J3 were prepared, and then the response strength of the various haplotypes of OR2J3 to (E,E)-2,4-decadienal at a concentration (100 μM) was measured by luciferase assay. The measurement results showing the above response strength are shown in FIG. 10. In FIG. 10, the vertical axis is the relative response strength (%) when the response strength of Hap4 is set to 100%. As is clear from the results shown in FIG. 10, all of the various haplotypes of OR2J3 (Hap1 to Hap5) respond to (E,E)-2,4-decadienal. In addition, the various haplotypes of OR2J3 (Hap1 to Hap5) all share the common feature of responding not only to (E,E)-2,4-decadienal, but also to the human olfactory receptor OR2J3 and its variants, i.e., heptanal, octanal, nonanal, decanal, undecanal, dodecanal, (E,E)-2,4-hexadienal, (E,E)-2,4-heptadienal, and (E,E) It is presumed that the compound responds to linear or branched, saturated or unsaturated aliphatic alcohols or aldehydes having 4 to 20 carbon atoms, such as (E,E)-2,4-octadienal, (E,E)-2,4-nonadienal, (E,Z)-2,6-nonadienal, (E,E)-2,4-undecadienal, (E)-2-nonenal, (E)-2-decenal, and (E,E)-2,4-decadienol. From the above results, it can be said that the screening method of the present invention can screen for novel lipid-degraded odor suppression materials that suppress lipid-degraded odor ((E,E)-2,4-decadienal odor) using polypeptides of various variants (e.g., haplotypes) of the olfactory receptor OR2J3. In addition, the screening method of the present invention uses polypeptides of various variants (e.g., haplotypes) of the olfactory receptor OR2J3 to detect not only (E,E)-2,4-decadienal, but also heptanal, octanal, nonanal, decanal, undecanal, dodecanal, (E,E)-2,4-hexadienal, and the like, which have in common that they respond to the human olfactory receptor OR2J3 and its variants. It is possible to screen for new lipid deterioration odor suppression materials that suppress lipid deterioration odors such as (E,E)-2,4-heptadienal, (E,E)-2,4-octadienal, (E,E)-2,4-nonadienal, (E,Z)-2,6-nonadienal, (E,E)-2,4-undecadienal, (E)-2-nonenal, (E)-2-decenal, and (E,E)-2,4-decadienol.

Claims

1. A method for screening a lipid-deterioration odor suppressing material, comprising the following steps (1) to (3): (1) contacting a test substance and the causative substance of lipid deterioration odor with an olfactory receptor polypeptide that contains an amino acid sequence of OR2J3 and / or an amino acid sequence having at least 90% identity with the amino acid sequence of OR2J3 and is responsive to the causative substance of lipid deterioration odor; (2) measuring the response of the olfactory receptor polypeptide to the lipid deterioration odor causative substance; and (3) identifying a test substance that inhibits the response of the olfactory receptor polypeptide based on the measured response; The method, wherein the substance causing the odor of deteriorated lipids is at least one selected from the group consisting of (E,E)-2,4-decadienal, heptanal, octanal, nonanal, decanal, undecanal, dodecanal, (E,E)-2,4-hexadienal, (E,E)-2,4-heptadienal, (E,E)-2,4-octadienal, (E,E)-2,4-nonadienal, (E,Z)-2,6-nonadienal, (E,E)-2,4-undecadienal, (E)-2-nonenal, (E)-2-decenal, and (E,E)-2,4-decadienol.

2. The method according to claim 1, wherein the lipid deterioration odor causative substance is (E,E)-2,4-decadienal.

3. The method according to claim 1, wherein the amino acid sequence of OR2J3 is an amino acid sequence set forth in any one of SEQ ID NOs: 1 to 5 in the sequence listing.

4. The lipid deterioration odor inhibitor contains at least one lipid deterioration odor inhibitor material selected from the group consisting of allyl isothiocyanate, dimethylbenzylcarbonyl butyrate, sec-butyl isothiocyanate, indole, and 3,4-dimethyl-1,2-cyclopentanedione.

5. A method for producing a lipid deterioration odor inhibitor produced by a method including the method according to claim 1, or a food or drink containing the lipid deterioration odor inhibitor according to claim 4.

6. A food or drink containing the lipid deterioration odor inhibitor according to claim 4.

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