Equol measurement kit and equol measurement method

Abstract

This invention provides an equol measurement kit that selectively and sensitively measures equol using surface-enhanced Raman scattering, and a method for selectively and sensitively measuring equol using surface-enhanced Raman scattering. [Solution] The equol measurement kit of this disclosure measures equol by surface-enhanced Raman scattering spectroscopy. The method for measuring equol according to this disclosure is a method for measuring equol by surface-enhanced Raman scattering spectroscopy. In this method, 1530-1630 cm⁻¹ -1 , 1230~1330cm -1 , 1140~1240cm -1 , and 535~635cm -1 It is preferable to measure equol by analyzing the peaks that appear in at least one wavenumber region selected from the group.

Description

[Technical Field]

[0001] This disclosure relates to a kit for measuring equol by surface-enhanced Raman scattering spectroscopy and a method for measuring equol. This disclosure claims priority to Japanese Patent Application No. 2020-153720, filed in Japan on September 14, 2020, and the contents of that application are incorporated herein by reference. [Background technology]

[0002] Equol is a compound produced when daidzein, a type of isoflavone, is metabolized by equol-producing bacteria. Equol is a compound with physiological activity similar to female hormones, and its production in the body compensates for the decrease in female hormone levels caused by menopausal symptoms, etc.

[0003] However, some individuals may not possess equol-producing bacteria, and in such cases, it may be necessary to take equol to compensate for the decrease in female hormone levels. Therefore, it is important to check whether or not equol is being produced in the body.

[0004] Whether or not equol is being produced in the body can be determined by measuring the amount of equol contained in urine or blood. However, since urine and blood contain many isoflavones such as daidzein, which have a similar structure to equol (hereinafter sometimes referred to as "equol analogs"), it is difficult to selectively detect and quantify equol.

[0005] Methods for selectively detecting and quantifying equol include separation and analysis methods using large-scale instruments such as HPLC and MS, as well as methods that utilize specific interactions such as antigen-antibody reactions.

[0006] Patent Document 1 describes a method for quantifying equol by contacting a sample with a labeling substance that immunospecifically binds to equol, i.e., a labeled conjugate, and measuring the amount of labeled conjugate bound to equol or the amount of labeled conjugate remaining unbound to equol using immunochromatography. However, the method in Patent Document 1 has problems with its complicated procedure and concerns about accuracy due to the cross-reactivity of the labeled conjugates. [Prior art documents] [Patent Documents]

[0007] [Patent Document 1] Japanese Patent Publication No. 2011-106886 [Overview of the Initiative] [Problems that the invention aims to solve]

[0008] Therefore, the object of this disclosure is to provide an equol measurement kit that selectively and sensitively measures equol by surface-enhanced Raman scattering. Another object of this disclosure is to provide a method for selectively and sensitively measuring equol by surface-enhanced Raman scattering. Another object of this disclosure is to provide a method for testing whether a subject has the ability to produce equol using surface-enhanced Raman scattering. Another object of this disclosure is to provide a surface-enhancing Raman scattering agent for equol measurement. [Means for solving the problem]

[0009] As a result of intensive studies to solve the above problems, the present inventors have found that in surface-enhanced Raman scattering spectroscopy for amplifying Raman scattered light and identifying or quantifying a substance from the spectrum of the amplified Raman scattered light, Raman peaks characteristic of equol can be identified, and it is possible to selectively and highly sensitively identify and quantify equol from a sample in which equol and equol analogs are mixed. The present disclosure has been completed based on these findings.

[0010] That is, the present disclosure provides an equol measurement kit for measuring equol by surface-enhanced Raman scattering spectroscopy.

[0011] The present disclosure also provides a method for measuring equol by surface-enhanced Raman scattering spectroscopy.

[0012] The present disclosure also provides the above method for measuring equol by analyzing peaks that appear in at least one wavenumber region selected from the group of 1530 to 1630 cm -1 、1230 to 1330 cm -1 、1140 to 1240 cm -1 、and 535 to 635 cm -1

[0013] The present disclosure also provides a method for examining whether a subject has the ability to produce equol, the method including measuring equol in a sample collected from the subject by the above method for measuring equol.

[0014] The present disclosure also provides a surface-enhanced Raman scattering agent for equol measurement containing a noble metal component.

Advantages of the Invention

[0015] The equol measurement kit disclosed herein allows for highly sensitive detection of equol. Furthermore, even in samples containing both equol and equol-like substances, the kit can distinguish between equol and equol-like substances, enabling sensitive measurement of the equol concentration in the sample.

[0016] Furthermore, the equol measurement method described herein allows for rapid and highly sensitive measurement of equol without the need for large-scale separation analysis such as HPLC, making it suitable for simple equol testing using gel-type sensors or test strips. [Brief explanation of the drawing]

[0017] [Figure 1] This figure shows the Raman shifts of a 100 μM equol solution and a 100 μM daidzein solution. [Figure 2] This figure shows the Raman shifts of equol solutions with concentrations of 1 μM and 100 μM, and the Raman shift of the solvent used in the solutions (as background). [Figure 3] This is a surface-enhanced Raman scattering spectrum measured by dropping 1 μL of a 100 μM equol solution onto a surface-enhanced Raman scattering substrate. [Modes for carrying out the invention]

[0018] [Equol Measurement Kit] The equol measurement kit described herein is a kit for measuring equol by surface-enhanced Raman scattering spectroscopy. More specifically, the equol measurement kit is a kit for quantifying and / or identifying equol by focusing on the characteristic Raman peak of equol.

[0019] The kit includes at least one device capable of identifying at least one of the characteristic Raman peaks of equol. The device is preferably a Raman spectrometer (for example, including a light source, a filter to remove Rayleigh scattered light, a spectrometer to decompose the Raman scattered light into a spectrum, a detector, etc.).

[0020] As the light source, for example, a light source that emits laser light (preferably near-infrared laser light) is preferred.

[0021] The aforementioned kit preferably further includes a surface-enhancing Raman scattering agent. The surface-enhancing Raman scattering agent will be described in detail later.

[0022] Using the aforementioned kit, selective trace quantification of equol is possible using a method similar to the equol measurement method described later.

[0023] Generally, the limit of equol concentration that can be measured by Raman spectroscopy is at most about 10 mmol / L. However, using the equol measurement kit of this disclosure, equol can be accurately quantified even if the equol content in the sample is very small. The limit of quantification concentration when measuring equol using the equol measurement kit of this disclosure is, for example, 1 μmol / L.

[0024] Furthermore, by using the kit, it is possible to determine whether or not a subject has the ability to produce equol by examining the subject's sample. Therefore, the kit may be an equol production ability test kit.

[0025] [Method for measuring equol] The method for measuring equol according to this disclosure is a method for measuring equol by surface-enhanced Raman scattering spectroscopy.

[0026] In the aforementioned method for measuring equol, by analyzing the peaks that are characteristic of equol, that is, peaks that appear in equol but not in equol-like substances, it is possible to selectively measure (quantify and / or identify) equol in a sample containing both equol and equol-like substances.

[0027] Peaks characteristically appearing in equol may vary slightly depending on experimental conditions such as the measuring instrument used. For example, 1530 - 1630 cm -1 , 1230 - 1330 cm -1 , 1140 - 1240 cm -1 , and at least one wavenumber region selected from the group of 535 - 635 cm -1 , preferably at least one peak appearing in a wavenumber region selected from the group of 1570 - 1590 cm -1 , 1270 - 1290 cm -1 , 1180 - 1200 cm -1 , and 570 - 590 cm -1 (i.e., a peak having a peak position in the said wavenumber region) can be mentioned.

[0028] In the said measuring method, it is preferable to use a surface - enhanced Raman scattering agent as a means to enhance the Raman scattered light of equol. The surface - enhanced Raman scattering agent will be described in detail later.

[0029] The measuring method of equol of the present disclosure includes, for example, the following steps. 1. A test specimen is formed by bringing the said surface - enhanced Raman scattering agent into contact with the specimen. 2. The test specimen is irradiated with irradiation light from a light source to generate scattered light (including Raman scattered light and Rayleigh scattered light). 3. The said scattered light is passed through a filter to remove Rayleigh scattered light. 4. The scattered light after removal of Rayleigh scattered light is introduced into a spectroscope, and the introduced scattered light is decomposed into a spectrum in the spectroscope. 5. The spectrum is detected by a detector.

[0030] There are no particular limitations on the method for bringing the surface-enhancing Raman scattering agent into contact with the sample, as long as the surface-enhancing Raman scattering agent (preferably the surface of the noble metal component in the surface-enhancing Raman scattering agent) can be brought into contact with the sample. Examples include fixing the surface-enhancing Raman scattering agent to the surface of a substrate and dropping a liquid sample (or a liquid preparation if the sample is not liquid) onto it, or mixing the surface-enhancing Raman scattering agent and the liquid sample on a sample stage or the like.

[0031] According to the above-described method for measuring equol, the surface-enhancing Raman scattering agent is used to amplify the Raman scattering of equol contained in the sample, thereby enabling selective and highly sensitive quantification of equol.

[0032] [Surface-enhancing Raman scattering agent] The surface-enhanced Raman scattering agent is a surface-enhanced Raman scattering agent used for measuring equol by surface-enhanced Raman scattering spectroscopy. Therefore, the surface-enhanced Raman scattering agent may also be a surface-enhanced Raman scattering agent for equol measurement.

[0033] The surface-enhancing Raman scattering agent preferably contains a noble metal component. In addition to the noble metal component, the surface-enhancing Raman scattering agent may also contain one or more selected from, for example, solvents, thickeners, surfactants, and dispersion stabilizers (e.g., citric acid).

[0034] The surface-enhancing Raman scattering agent may have the noble metal component fixed to the substrate or dispersed in a dispersion medium. However, it is preferable that the noble metal component be dispersed in a dispersion medium, in particular in terms of enhancing the Raman scattering light enhancement effect and improving identification accuracy, and more preferably in a viscous dispersion medium, and especially preferably in a gel-like dispersion medium. Therefore, the surface-enhancing Raman scattering agent is preferably a noble metal dispersion, and more preferably a gel-like noble metal dispersion.

[0035] The viscosity of the dispersion medium (preferably a viscous dispersion medium, and particularly preferably a gel-like dispersion medium) at 25°C and a shear rate of 10¹ / s is preferably, for example, 50 mPa·s or higher, in that it can enhance the Raman scattering light enhancement effect.

[0036] Furthermore, the precious metal components may be dispersed in a colloidal state within the precious metal dispersion. That is, the precious metal dispersion may be a precious metal colloidal solution.

[0037] The viscosity of the aforementioned noble metal dispersion (especially the gel-like noble metal dispersion) at 25°C and a shear rate of 10¹ / s is, for example, 50 to 100,000 mPa·s, preferably 100 to 50,000 mPa·s, and particularly preferably 200 to 30,000 mPa·s.

[0038] Furthermore, the viscosity of the aforementioned dispersion or dispersion medium can be measured using a viscosity-viscoelasticity measuring device (rheometer) (product name "RheoStress600", manufactured by HAAKE).

[0039] The surface-enhanced Raman scattering agent allows for accurate measurement of equol content even when the amount of equol in a sample (e.g., a subject's urine, blood, or intestinal contents, or the supernatant thereof) is very small.

[0040] When adult subjects have the ability to produce equol, the equol concentration in their urine is usually between 1 and 75 μmol / L. Therefore, by measuring the amount of equol in a subject's urine using the surface-enhanced Raman scattering agent, it is easy to determine whether or not the subject has the ability to produce equol (or whether or not the subject has equol-producing bacteria).

[0041] (Precious metal component) The aforementioned precious metal component is preferably at least one precious metal component selected from gold, silver, and copper.

[0042] (solvent) The aforementioned solvent is a component of the dispersion medium. It is preferable to use a solvent that can highly disperse the noble metal component and has low Raman activity, as this improves the accuracy of equol quantification.

[0043] The aforementioned solvent includes water-soluble solvents and oil-based solvents. Specific examples of the aforementioned solvents include water, alcohol, and oils. These can be used individually or in combination of two or more.

[0044] Examples of the aforementioned alcohols include methanol, ethanol, propanol, butanol, and other alcohols; and polyhydric alcohols such as ethylene glycol, propylene glycol, butylene glycol, and glycerin.

[0045] Examples of the aforementioned oils include hydrocarbon oils such as mineral oil and liquid paraffin; sunflower oil, macadamia nut oil, avocado oil, almond oil, wheat germ oil, rice germ oil, olive oil, soybean oil, corn oil, castor oil, beef tallow, jojoba oil, evening primrose oil, coconut oil, camellia oil, rosehip oil, squalane, turtle oil, mink oil, egg yolk oil, lanolin, whale wax, candelilla wax, montan wax, rice wax, lanolin wax, shellac, and other animal and vegetable oils; and carbonated water such as hexane, cyclohexane, isododecane, benzene, toluene, poly-alpha-olefin, and liquid paraffin. Examples include raw oils; ethers such as tetrahydrofuran; halogenated hydrocarbons such as carbon tetrachloride and chlorobenzene; petroleum components such as kerosene, gasoline, diesel fuel, and heavy oil; silicone oils such as dimethylpolysiloxane and methylphenylpolysiloxane; ester oils such as octyldodecyl oleate, cetyl ethylhexanoate, glyceryl triisooctanate, and neopentyl glycol diisooctanate; higher alcohols such as hexadecyl alcohol and oleyl alcohol; higher fatty acids such as lauric acid, isostearic acid, and oleic acid; aromatic carboxylic acids, pyridine, etc.

[0046] Examples of solvents with high Raman activity include N,N-dimethylformamide (DMF), dimethyl sulfoxide (DMSO), dimethylacetamide (DMAA), N-methylpyrrolidone (NMP), tetrahydrofuran (THF), cyclohexanone, lactam, lactone, and N,N,N,N-tetramethylurea. When using these solvents, the amount used (or the total amount if two or more are used in combination) is preferably 30% by weight or less, more preferably 20% by weight or less, particularly preferably 10% by weight or less, most preferably 5% by weight or less, and especially preferably 1% by weight or less, of the total amount of solvent contained in the surface-enhancing Raman scattering agent.

[0047] (Thickening agent) If the viscosity of the solvent is low, a thickening agent may be added to increase its viscosity. The thickening agent can be appropriately selected depending on the type of solvent.

[0048] When a water-soluble solvent is used as the solvent, examples of thickeners include cellulosic polymer compounds such as hydroxyethylcellulose, methylcellulose, ethylcellulose, and carboxymethylcellulose; plant-derived natural polymer compounds such as carrageenan and guar gum; microbial-derived natural polymer compounds such as xanthan gum; animal-derived natural polymer compounds such as casein and gelatin; starch-based polymer compounds such as carboxymethyl starch; vinyl-based polymer compounds such as polyvinyl alcohol, polyvinylpyrrolidone, sodium polyacrylate, and carboxyvinyl polymer; polyether-based polymer compounds such as polyethylene glycol, polypropylene glycol, and polyglycerin; and polyoxyalkylene-modified sugars such as polyoxyethylene methyl glucoside and polyoxypropylene methyl glucoside.

[0049] When an oily solvent is used as the solvent, examples of thickeners include sugar fatty acid esters, 12-hydroxystearic acid, 1,3,2,4-dibenzylidene-D-sorbitol, N-lauroyl-L-glutamic acid-α,γ-bis-n-butylamide, benzene fatty acid amide, and cyclohexane fatty acid amide. The sugar fatty acid esters include, for example, dextrin fatty acid esters such as dextrin palmitate and dextrin myristate; and inulin fatty acid esters such as inulin stearate.

[0050] [Method for testing equol production ability] The method for testing equol production ability is a method for testing whether or not a subject has the ability to produce equol. The method for testing equol production ability also includes measuring equol in a sample taken from the subject using the method for measuring equol described above.

[0051] If a subject has the ability to produce equol, they will have equol-producing bacteria in their intestines. When a subject ingests daidzein, a type of isoflavone, these bacteria metabolize daidzein to produce equol. The produced equol is then absorbed from the intestinal tract into the bloodstream, and some of it is excreted in the urine. Therefore, by using urine, blood, or intestinal contents, or the supernatant obtained by centrifugation of these, as a sample and measuring the amount of equol contained in the sample, it is easy to determine whether a subject has the ability to produce equol or whether they have equol-producing bacteria.

[0052] Examples of equol-producing bacteria include microorganisms belonging to the genera Lactococcus, Slackia, Adlercreutzia, Asaccharobacter, and Eggerthella. More specifically, examples include Lactococcus garvieae, Adlercreutzia equolifaciens, Asaccharobacter celatus, and Eggerthella sp.YY7918.

[0053] For example, if an adult subject has the ability to produce equol, the equol concentration in their urine is usually between 1 and 75 μmol / L. Therefore, by measuring the equol contained in the subject's urine, if the equol concentration is above this range, it can be determined that the subject has the ability to produce equol, and if the equol concentration is below this range, it can be determined that the subject does not have the ability to produce equol.

[0054] In the aforementioned method for testing equol production ability, equol is measured selectively and with high sensitivity using the equol measurement method described above, making it possible to accurately determine whether or not equol production ability is present, or whether or not equol-producing bacteria are present.

[0055] The configurations and combinations thereof described herein are examples only, and additions, omissions, substitutions, and modifications are permitted as appropriate, without departing from the spirit of this disclosure. Furthermore, this disclosure is not limited by the embodiments, but is limited solely by the claims. [Examples]

[0056] The present disclosure will be described in more detail below with reference to examples, but the present disclosure is not limited to these examples.

[0057] Preparation Example 1 (Sample Preparation) A 10 mM ethanol solution of equol was diluted with water and ethanol to prepare a 1 μM equol solution and a 100 μM equol solution.

[0058] Preparation Example 2 (Sample Preparation) A 100 μM daidzein solution was prepared by diluting a 420 μM daidzein solution with water and ethanol.

[0059] Example 1 Equol was measured according to the method described in H. Segawa, T. Fukuoka, T. Itoh, Y. Imai, YT Iwata, T. Yamamuro, K. Kuwayama, K. Tsujikawa, T. Kanamori and H. Inoue, Analyst, 144, 6928-6935 (2019). Specifically, 3 μL of the 100 μM equol solution obtained in Preparation Example 1, 3 μL of the 1 M sodium chloride solution, and 54 μL of the gold colloid solution were added to a container and mixed. The container was placed in the measurement chamber of Raman spectrometer 1 (manufactured by LambdaVision, Inc., product name "RAM mini"), and Raman scattered light was generated by repeatedly irradiating it with a 1-second laser beam (laser wavelength: 785 nm) four times, and the average was recorded. The time required for mixing and measurement was less than 30 seconds. The same procedure was also performed on the solvent used for sample preparation to create the background.

[0060] Furthermore, for surface-enhanced Raman scattering measurements, the co-aggregation method, a wet analytical method suitable for initial screening, was used.

[0061] Comparative Example 1 The procedure was the same as in Example 1, except that the 100 μM daidzein solution obtained in Preparation Example 2 was used instead of the 100 μM equol solution.

[0062] The results for Example 1 and Comparative Example 1 are shown in Figure 1. As can be seen from Figure 1, in the 100 μM equol solution, 1580 cm³ -1 , 1280cm-1 , 1190cm -1 , and 590cm -1 A characteristic peak appeared in the vicinity. On the other hand, no peak appeared in the vicinity of the aforementioned wavenumber region in the 100 μM daidzein solution.

[0063] Example 2 The procedure was carried out in the same manner as in Example 1, except that a 1 μM equol solution was used instead of a 100 μM equol solution.

[0064] The results of Example 1 and Example 2 are shown in Figure 2. As can be seen from Figure 2, even at a concentration of 1 μM, equol produced 1580 cm³, similar to the case of 100 μM. -1 , 1280cm -1 , 1190cm -1 , and 590cm -1 A peak characteristic of equol was observed in the vicinity. This confirmed that the method disclosed herein allows for the selective quantitative determination of trace amounts of equol.

[0065] Example 3 A substrate containing gold nanorods as a precious metal component (NIDEK Corporation, Wavelet substrate) was used as a surface-enhanced Raman scattering substrate. One μL of the 100 μM equol solution obtained in Preparation Example 1 was dropped onto the surface-enhanced Raman scattering substrate and allowed to air dry. The substrate was placed in the measurement section of Raman spectrometer 2 (LambdaVision, Inc., product name "RAM200") equipped with a 785 nm laser, and the average of four 1-second laser irradiations was recorded. 1570 cm² -1 , 1270cm -1 1174cm -1 , 570cm -1 A distinctive peak was observed in the vicinity (Figure 3).

[0066] Here, the Raman spectrum of diamond was measured using the Raman spectrometer 2 as a standard material. The result was 1322 cm⁻¹. -1 A sharp peak was observed. The single-crystal diamond was 1332 cm³. -1Since a Raman peak is known to appear in the vicinity, the peak position observed by the Raman spectrometer 2 is approximately 10 cm. -1 It was found that the frequency shifted to a lower wavenumber.

[0067] Considering the above, the peak position is 1580cm. -1 , 1280cm -1 , 1184cm -1 , 580cm -1 It can be corrected to be in the vicinity.

[0068] Although different types of spectrometers cause shifts in peak positions, it was found that at least one peak position can be matched by correcting for the shift in the peak position of the standard substance. Furthermore, it was found that equol can be measured from the Raman peak by using the peak with the matched position.

[0069] In summary, the structure of this disclosure and its variations are described below. [1] An equol measurement kit that measures equol by surface-enhanced Raman scattering spectroscopy. [2] The equol measurement kit described in [1], including a Raman spectrometer. [3] An equol measurement kit according to [1] or [2], comprising a surface-enhancing Raman scattering agent. [4] An equol measurement kit according to [1] or [2], comprising a surface-enhancing Raman scattering agent containing a precious metal component. [5] An equol measurement kit according to [1] or [2], comprising a surface-enhancing Raman scattering agent containing at least one precious metal component selected from gold, silver, and copper. [6] An equol measurement kit according to [1] or [2], comprising a surface-enhancing Raman scattering agent in which a noble metal component is dispersed in a viscous dispersion medium. [7] A surface-enhancing Raman scattering agent comprising a noble metal component dispersed in a gel-like dispersion medium. [8] An equol measurement kit according to [1] or [2], comprising a gel-like dispersion of precious metals. [9] An equol measurement kit according to [1] or [2], comprising a gel-like dispersion of noble metals having a viscosity of 50 mPa·s or more at 25℃ and a shear rate of 10(1 / s).

[10] An equol measurement kit according to [1] or [2], comprising a precious metal colloid solution.

[11] The equol measurement kit according to [1] or [2], comprising a substrate on which a precious metal component is immobilized.

[12] A method for measuring equol, which involves measuring equol by surface-enhanced Raman scattering spectroscopy.

[13] 1530~1630cm -1 , 1230~1330cm -1 , 1140~1240cm -1 , and 535~635cm -1 A method for measuring equol as described in

[12] , wherein equol is measured by analyzing a peak appearing in at least one wavenumber region selected from the group.

[14] 1570~1590cm -1 , 1270~1290cm -1 , 1180~1200cm -1 , and 570~590cm -1 A method for measuring equol as described in

[12] , wherein equol is measured by analyzing a peak appearing in at least one wavenumber region selected from the group.

[15] A method for testing whether a subject has the ability to produce equol, comprising measuring equol in a sample taken from the subject by any one of the methods for measuring equol described in

[12] to

[14] .

[16] A surface-enhancing Raman scattering agent containing a precious metal component for equol measurement.

[17] A surface-enhanced Raman scattering agent for equol measurement according to

[16] , comprising a surface-enhanced Raman scattering agent containing at least one precious metal component selected from gold, silver, and copper.

[18] A surface-enhancing Raman scattering agent for equol measurement according to

[16] , comprising a noble metal component and a viscous dispersion medium.

[19] A surface-enhancing Raman scattering agent for equol measurement according to

[16] , comprising a noble metal component and a gel-like dispersion medium.

[20] A surface-enhancing Raman scattering agent for equol measurement according to

[16] , comprising a gel-like dispersion of noble metals.

[21] A surface-enhancing Raman scattering agent for equol measurement according to

[16] , comprising a gel-like noble metal dispersion having a viscosity of 50 mPa·s or more at 25℃ and a shear rate of 10(1 / s).

[22] A surface-enhancing Raman scattering agent for equol measurement according to

[16] , comprising a noble metal colloid solution. [Industrial applicability]

[0070] The equol measurement kit disclosed herein allows for rapid and highly sensitive detection of equol. Furthermore, even in samples containing both equol and equol-like substances, the kit can distinguish between equol and equol-like substances, enabling sensitive measurement of the equol concentration in the sample.

Claims

1. An equol measurement kit that measures equol using surface-enhanced Raman scattering spectroscopy.

2. A method for measuring equol using surface-enhanced Raman scattering spectroscopy.

3. 1530-1630cm －１ , 1230-1330cm －１ , 1140-1240cm －１ , and 535-635 cm －１ A method for measuring equol according to claim 2, comprising measuring equol by analyzing a peak appearing in at least one wavenumber region selected from the group.

4. A method for testing whether a subject has the ability to produce equol, comprising measuring equol in a sample taken from the subject using the method for measuring equol described in claim 2 or 3.

5. A surface-enhancing Raman scattering agent containing precious metal components for equol measurement.

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