Method for producing compounds related to 3-phenylpropionic acid

Abstract

Provided is a method for producing 3-phenylpropionic acid analogue compounds given by formula (I), the production method characteristically comprising a step for culturing a lactic acid bacterium from genus Weissella using a culture medium that contains at least 3 mM of a cinnamic acid analogue compound given by formula (II), wherein the lactic acid bacterium exhibits the following properties (1a) to (3a) when the lactic acid bacterium is inoculated to MRS liquid culture medium containing 3 mM 4-hydroxy-3-methoxycinnamic acid and is cultured for 24 hours at 37°C under anaerobic conditions: (1a) a turbidity of at least 1.0; (2a) a ratio for reduction to 3-(4-hydroxy-3-methoxyphenyl)propionic acid of at least 80%; and (3a) a ratio for decarboxylation to 4-vinylguaiacol of less than 5%. Also provided is a lactic acid bacterium that exhibits these properties. 3-Phenylpropionic acid analogue compounds can be efficiently produced using this method and this lactic acid bacterium.

Description

[Technical Field]

[0001] The present invention relates to a method for producing 3-phenylpropionic acid analogs, and more specifically, to a method for efficiently producing 3-phenylpropionic acid analogs using lactic acid bacteria with cinnamic acid analogs as a raw material. The present invention also relates to lactic acid bacteria capable of carrying out the above production method. [Background technology]

[0002] In recent years, lifestyle-related diseases such as obesity, type 2 diabetes, hypertension, and insulin resistance have become a major problem due to unhealthy lifestyles such as overeating and lack of exercise. In addressing these lifestyle-related diseases, not only are pharmaceutical treatments gaining attention, but preventative and therapeutic approaches using naturally derived or food-derived components are also attracting interest.

[0003] Compounds related to 3-phenylpropionic acid are recognized to have effects that are involved in lifestyle-related diseases, and their functionality is attracting attention. For example, 3-(4-hydroxy-3-methoxyphenyl)propionic acid has been reported to have cAMP phosphodiesterase activity inhibitory effects and dipeptidyl peptidase IV activity inhibitory effects (see Patent Document 1).

[0004] Lactobacillus plantarum, a lactic acid bacterium isolated from pickles and other foods, has been shown to have two pathways for degrading cinnamic acid analogs contained in plant cell walls, such as 4-hydroxy-3-methoxycinnamic acid. For example, when 4-hydroxy-3-methoxycinnamic acid, a type of cinnamic acid analog, is added to a culture medium and cultured, 3-(4-hydroxy-3-methoxyphenyl)propionic acid, which is the reduced propenoic acid portion of 4-hydroxy-3-methoxycinnamic acid, and 4-vinylguaiacol, which is the decarboxylated 4-hydroxy-3-methoxycinnamic acid, are detected.

[0005] When the present inventors added 4-hydroxy-3-methoxycinnamic acid to a medium from which dissolved oxygen had been removed and cultured it, they clarified that the reduction reaction took precedence over the decarboxylation reaction and 3-(4-hydroxy-3-methoxyphenyl)propionic acid was preferentially produced (see Patent Document 2), and they also identified 4-hydroxy-3-methoxycinnamic acid reductase (see Patent Document 3).

Prior Art Documents

Patent Documents

[0006]

Patent Document 1

Patent Document 2

Patent Document 3

Summary of the Invention

Problems to be Solved by the Invention

[0007] An object of the present invention is to provide a production method capable of efficiently producing a 3-phenylpropionic acid analog compound, and to provide a lactic acid bacterium useful for the production method.

Means for Solving the Problems

[0008] As a result of conducting research to solve the above problems, the present inventors found that by using a lactic acid bacterium belonging to the genus Weissella having predetermined properties, a 3-phenylpropionic acid analog compound can be efficiently produced as compared with conventional methods, and thus completed the present invention. Specifically, the present invention is as follows.

[0009] 〔1〕 A method for producing a 3-phenylpropionic acid analog compound represented by the following formula (I),

Chemical formula

[0010] According to the method of the present invention, 3-phenylpropionic acid analog compounds can be produced efficiently. Furthermore, the lactic acid bacteria according to the present invention are useful in the above production method. [Modes for carrying out the invention]

[0011] Embodiments of the present invention will be described below. [Lactic acid bacteria] The lactic acid bacteria according to one embodiment of the present invention is a lactic acid bacterium of the genus Weissella, which, when inoculated into a liquid medium containing a high concentration of 4-hydroxy-3-methoxycinnamic acid (hereinafter sometimes abbreviated as HMCA) and cultured under anaerobic conditions, grows sufficiently, has a sufficiently high reduction rate to 3-(4-hydroxy-3-methoxyphenyl)propionic acid (hereinafter sometimes abbreviated as HMPA), and has a sufficiently low decarboxylation rate to 4-vinylguaiacol (hereinafter sometimes abbreviated as 4VG).

[0012] In the process of screening lactic acid bacteria useful for HMPA production, the inventors discovered that, depending on the strain, some lactic acid bacteria of the genus Weissella tend to have their growth inhibited or, even if they do grow, their HMPA reduction rate decreases when the HMPA concentration increases. In contrast, among bacterial strains capable of reducing HMCA to HMPA, strains that can grow sufficiently even at high concentrations of HMCA (e.g., 3 mM or higher) can produce HMPA more efficiently because the reaction of reducing HMCA to HMPA is accelerated as the strain grows. Furthermore, if the decarboxylation rate to 4VG is sufficiently low, HMCA is not consumed unnecessarily, and HMPA can be produced even more efficiently.

[0013] Furthermore, the present inventors have confirmed that 4-hydroxy-3-methoxycinnamate reductase (see Patent Document 3), which was identified in Lactobacillus plantarum, is not found in lactic acid bacteria of the genus Weissella. Therefore, it is believed that the reduction reaction of HMCA by lactic acid bacteria of the genus Weissella is carried out by a different enzyme than that in Lactobacillus plantarum.

[0014] (Culture conditions) The lactic acid bacteria according to this embodiment are identified by their properties when cultured under predetermined culture conditions. The predetermined culture conditions refer to inoculating the bacteria into a liquid culture medium containing a high concentration of HMCA and culturing them under anaerobic conditions.

[0015] The liquid culture medium used under the above culture conditions can be any medium that can be used for culturing lactic acid bacteria, such as MRS medium and GYP medium. Among these, MRS medium is preferred. Specifically, MRS medium has the following composition.

[0016] [Table 1]

[0017] The HMCA concentration under the above culture conditions should be high, specifically 3 mM or higher, with examples including 3 mM, 5 mM, 10 mM, and 50 mM. Furthermore, the above-mentioned predetermined culture conditions are based on the initial turbidity (OD) at the time of inoculation. 660This refers to a case where the ratio is 0.01, under anaerobic conditions, and cultured at 37°C for 24 hours.

[0018] In this embodiment, "anaerobic conditions" means, for example, a state in which the dissolved oxygen concentration of the liquid culture medium is lower than the saturation state (for example, about 6.7 mg / L at 37°C). For example, the dissolved oxygen concentration in the liquid culture medium may be 6 mg / L or less, 5 mg / L or less, 3 mg / L or less, 1 mg / L or less, 0.5 mg / L or less, or 0.2 mg / L or less.

[0019] The method for achieving such anaerobic conditions is not particularly limited. For example, a culture medium with a low dissolved oxygen concentration can be prepared by (pressurized) heating and sterilizing the liquid culture medium to remove dissolved oxygen without contact with oxygen such as air, and then immediately sealing it tightly to maintain a state of not being in contact with oxygen. In addition, known methods can be employed, such as the anaerobic jar method, anaerobic bag method, nitrogen displacement method, steel wool method, etc., which reduce the dissolved oxygen concentration in the culture medium by reducing the oxygen concentration in the gas phase; or the method of adding additives such as deoxidizers and antioxidants to the liquid culture medium to reduce the dissolved oxygen concentration. Furthermore, in order to maintain the above anaerobic conditions during cultivation, it is preferable to either keep the culture medium away from the gas phase or to bring it into contact with a gas phase in which the oxygen concentration is kept lower than that of air.

[0020] (Properties of lactic acid bacteria) The lactic acid bacteria according to this embodiment, when cultured under the above culture conditions, possess the following properties (1) to (3): (1) To grow sufficiently; (2) The conversion rate from HMCA to HMPA is sufficiently high; and (3) The decarboxylation rate from HMCA to 4VG is sufficiently low.

[0021] Here, (1) "sufficient growth" specifically refers to the turbidity (OD) after culturing for 24 hours under the above culture conditions. 660This means that the turbidity is above a predetermined value. The turbidity after 24 hours of incubation can be set to, for example, 1.0 or higher, 1.2 or higher, or 1.5 or higher.

[0022] Furthermore, the statement above (2) that the reduction rate from HMCA to HMPA is sufficiently high specifically means that the reduction rate to HMPA after 24 hours of incubation under the above culture conditions is above a predetermined value. The HMPA reduction rate after 24 hours of incubation can be set to, for example, 80% or higher, 85% or higher, 90% or higher, or 95% or higher.

[0023] Furthermore, the statement that (3) the decarboxylation rate from HMCA to 4VG is sufficiently low specifically means that the decarboxylation rate to 4VG after 24 hours of incubation under the above culture conditions is below a predetermined value. The 4VG decarboxylation rate after 24 hours of incubation can be set to, for example, less than 5%, less than 4%, or less than 3%.

[0024] The above-mentioned predetermined culture conditions and the properties of (1) to (3) above can be set, for example, as follows. When HMCA is inoculated into a liquid medium containing 3 mM and incubated under anaerobic conditions at 37°C for 24 hours, it possesses the following properties (1a) to (3a): (1a) Turbidity is 1.0 or higher; (2a) Reduction rate to HMPA is 80% or higher; and (3a) Decarboxylation rate to 4VG is less than 5%.

[0025] Furthermore, the above-mentioned predetermined culture conditions and the properties of (1) to (3) above may be set as follows, for example. When HMCA is inoculated into a liquid medium containing 5 mM and incubated under anaerobic conditions at 37°C for 24 hours, it possesses the following properties (1b) to (3b): (1b) Turbidity is 1.0 or higher; (2b) Reduction rate to HMPA is 80% or higher; and (3b) Decarboxylation rate to 4VG is less than 5%.

[0026] Furthermore, lactic acid bacteria of the genus Weissella tend to have their growth inhibited or their HMCA reduction rate decrease as the HMCA concentration increases. In other words, lactic acid bacteria that possess the above properties (1b) to (3b) when the initial HMCA concentration is 5 mM can be said to possess the above properties (1a) to (3a) when the initial HMCA concentration is 3 mM.

[0027] (Types of lactic acid bacteria and how to obtain them) The lactic acid bacteria used in this embodiment are not particularly limited as long as they are lactic acid bacteria of the genus Weissella possessing the above-mentioned properties.

[0028] Examples of lactic acid bacteria belonging to the genus Weissella include Weissella cibaria, Weissella confusa, Weissella hellenica, and Weissella oryzae. Any of these may be used, but Weissella cibaria or Weissella confusa are preferred.

[0029] When using Weissella sibaria, strains Weissella sibaria 011YN2 (accession number: NITE BP-03579) or Weissella sibaria 054YN2 (accession number: NITE BP-03578), used in the examples described later, can be exemplified as particularly suitable strains. These strains were isolated from pickles by the inventors and are deposited at the Patent Organism Depositary Center of the National Institute of Advanced Industrial Science and Technology (AIST).

[0030] Furthermore, when using Weissera Confuser, Weissera Confuser NBRC 3957 strain can be cited as a suitable example. Strain NBRC 3957 is stored at the National Institute of Technology and Evaluation (NITE) Biotechnology Center and is listed in the NBRC catalog as a commercially available strain.

[0031] The lactic acid bacteria used in this embodiment can be obtained by known screening methods. For example, the lactic acid bacteria of this embodiment can be obtained by culturing Lactobacillus weissella as a sample under the culture conditions described above and determining whether or not it satisfies the above properties (1) to (3). In such screening, it is also possible to pre-screen whether the plants grow sufficiently (corresponding to property (1) above) by culturing them in a solid medium containing a high concentration of HMCA, and then determine again whether they satisfy properties (1) to (3) above using a liquid medium. The term "high concentration" here is the same as the definition used for the liquid medium used when determining properties (1) to (3) above.

[0032] Furthermore, by repeatedly culturing in a medium containing a high concentration of HMCA, it is possible to accumulate Weissella lactic acid bacteria with improved HMCA production efficiency. For example, if there are Weissella lactic acid bacteria that possess the above properties (1a) to (3a) when the initial HMCA concentration is 3 mM, but do not possess the above properties (1b) to (3b) when the initial HMCA concentration is 5 mM, then by repeatedly culturing these lactic acid bacteria in a medium containing 5 mM or more of HMCA, strains that can efficiently utilize HMCA can be selected from among these lactic acid bacteria. Among the strains selected in this way, there are strains that have come to satisfy the above properties (1b) to (3b). Therefore, by determining whether or not these selected strains satisfy the above properties (1b) to (3b), Weissella lactic acid bacteria with improved HMCA production efficiency can be obtained.

[0033] As described above, the Lactobacillus weissella species can efficiently reduce HMCA to HMPA, making it particularly useful for HMPA production and, consequently, for the production of 3-phenylpropionic acid-related compounds.

[0034] [Method for producing compounds related to 3-phenylpropionic acid] The method for producing a 3-phenylpropionic acid analog compound according to an embodiment of the present invention includes a step of culturing lactic acid bacteria of the genus Weissella according to the above embodiment using a medium containing a cinnamic acid analog compound at a high concentration.

[0035] The 3-phenylpropionic acid analog compound produced in this embodiment is a compound represented by the following formula (I) (hereinafter, may be simply abbreviated as "3-phenylpropionic acids").

[0036]

Chemical formula

[0037] In the above formula (I), R 11 ~R 15 are each independently a hydrogen atom, a hydroxy group, an alkoxy group having 1 to 3 carbon atoms, or an alkyl group having 1 to 3 carbon atoms.

[0038] Among these, a hydrogen atom, a hydroxy group, and a methoxy group are preferable. In particular, it is preferable that R 11 、R 14 and R 15 are hydrogen atoms, and it is preferable that R 12 and R 13 are each independently a hydrogen atom, a hydroxy group, and a methoxy group.

[0039] Exemplary compounds that are particularly preferable as the 3-phenylpropionic acids produced by the method of this embodiment are as follows. HMPA: R 11 =H, R 12 =OCH3, R 13 =OH, R 14 =H, R 15 =H 3-Phenylpropionic acid: R 11 =H, R 12 =H, R 13 =H, R 14 =H, R 15 =H 3-(3,4-Dihydroxyphenyl)propionic acid: R 11=H,R 12 =OH,R 13 =OH,R 14 =H,R 15 =H 3-(4-hydroxyphenyl)propionic acid:R 11 =H,R 12 =H,R 13 =OH,R 14 =H,R 15 =H

[0040] Furthermore, in the production method of this embodiment, the cinnamic acid analog compound used as a raw material is the compound represented by the following formula (II) (hereinafter sometimes simply referred to as "cinnamic acid compounds").

[0041] [ka]

[0042] In the above equation (II), R 21 ~R 25 This corresponds to R in equation (I) above. 11 ~R 15 These are identical to each other. Here, R in equation (I) 11 ~R 15 and R in equation (II) 21 ~R 25 Terms that have the same substitution position are called "corresponding," for example, R in equation (I) 11 The corresponding part is R in equation (II). 21 That is the case.

[0043] The lactic acid bacteria of the genus Weissella used in this embodiment can efficiently reduce HMCA to HMPA, but as shown in the examples described later, these lactic acid bacteria can also reduce cinnamic acids other than HMCA. This is thought to be because the HMCA reductase of Weissella lactic acid bacteria is relatively tolerant of the substituents on the benzene ring of cinnamic acids. Therefore, the above-mentioned Weissella lactic acid bacteria can also be used in reactions to reduce cinnamic acids other than HMCA and produce 3-phenylpropionic acids other than HMPA. The raw material, cinnamic acid, is, in principle, R in formula (II). 21 ~R 25 However, in the compound of formula (I) (3-phenylpropionic acid) that we intend to produce, the corresponding R 11 ~R 15 We use compounds that are identical to each of the others.

[0044] Here, the culture medium used in the process of culturing lactic acid bacteria of the genus Weissella is not particularly limited as long as it is a culture medium that can be used for culturing lactic acid bacteria. For example, it may be the same as the liquid medium used to identify the properties of the lactic acid bacteria in the above embodiment, or it may be a different medium. Examples of such media include MRS medium and GYP medium. Furthermore, a culture medium containing plant materials that can be used as food ingredients can also be suitably used, and when such a medium is used, the resulting culture solution can be used directly as a food ingredient.

[0045] The pH of the culture medium is preferably 2 to 9, and more preferably 3 to 8. The culture temperature is preferably 20 to 45°C, more preferably 25 to 40°C, and particularly preferably 30 to 38°C. The pH and culture temperature of the culture medium being within this range is suitable for the growth of Lactobacillus weissella and results in a desirable reduction rate from cinnamic acids to 3-phenylpropionic acids.

[0046] The concentration of cinnamic acid compounds in the culture medium used in the above process can preferably be 3 mM or higher, 5 mM or higher, 10 mM or higher, 20 mM or higher, 50 mM or higher, or 100 mM or higher, and can be appropriately set considering the above properties (1) to (3) of the Weissella lactic acid bacteria used. By increasing the concentration of cinnamic acid compounds, a culture with a high concentration of 3-phenylpropionic acid compounds can be obtained in this process.

[0047] Here, the cinnamic acids used in this process may be purified products or compositions containing cinnamic acids. Examples of compositions containing cinnamic acids include extracts of plants containing cinnamic acids. Examples of such plants include crushed materials and extracts of plants such as rice, wheat, barley, corn, bamboo, coffee, tomato, mate, mugwort, and burdock. Furthermore, since cinnamic acids are components of lignin in woody and herbaceous plants, lignin or compositions containing it may also be used. Among these, rice, barley, and bamboo (bamboo shoots) are preferred from the viewpoint of ease of availability and cinnamic acid content, and among rice, rice bran is particularly preferred.

[0048] In this process, cultivation is preferably carried out under anaerobic conditions. Since Lactobacillus weissella is a facultative anaerobic bacterium, culturing under anaerobic conditions is suitable for the growth of Lactobacillus weissella. Furthermore, culturing under anaerobic conditions results in a desirable reduction rate from cinnamic acids to 3-phenylpropionic acids. The meaning of anaerobic conditions and the method for creating anaerobic conditions are as described in the embodiments above. However, unlike the embodiments above, it is not necessary to strictly control the anaerobic conditions in this process. For example, even if the conditions are not met in the initial stages of culture, the culture conditions may become anaerobic as dissolved oxygen is consumed as the Weissella lactic acid bacteria grow.

[0049] The culture obtained in this cultivation process contains a high concentration of 3-phenylpropionic acids. The obtained culture may be used as is as a food ingredient, etc., depending on the subsequent purpose, or it may be subjected to appropriate treatments such as decolorization and clarification. Examples of such treatments include activated carbon treatment and resin treatment. Furthermore, the above culture may be further fractionated and purified to obtain a composition with an increased concentration of 3-phenylpropionic acids. The method of fractionation and purification is not particularly limited, and known methods such as column chromatography, HPLC, and recrystallization can be used as appropriate.

[0050] According to the above-described method for producing 3-phenylpropionic acids, 3-phenylpropionic acids can be obtained in high yield by culturing Lactobacillus weissella, which possesses the above-mentioned properties, in a culture medium containing a high concentration of cinnamic acids.

[0051] The embodiments described above are provided to facilitate understanding of the present invention and are not intended to limit it. Accordingly, each element disclosed in the above embodiments is intended to include all design modifications and equivalents that fall within the technical scope of the present invention. [Examples]

[0052] The present invention will be described in more detail below with reference to test examples, etc., but the present invention is not limited in any way to the test examples, etc. described below.

[0053] <Test Example 1> Culture Test-1 Using the lactic acid bacteria shown in Table 2, cultures were performed in MRS medium containing HMCA, and turbidity, HMCA retention rate, HMPA production rate, and 4VG were measured as follows.

[0054] The lactic acid bacteria used in this study are as follows. Strains 011YN2 and 054YN2 were isolated from pickles by the inventors and were identified as belonging to the lactic acid bacteria Weissella cibaria by determining their 16S rDNA base sequences according to conventional methods.

[0055] [Table 2]

[0056] In this culture test, MRS medium (Merck) was used to culture lactic acid bacteria, and the pH was adjusted to 7.5. HMCA was manufactured by Tokyo Chemical Industry Co., Ltd. Anaerobic conditions were maintained using Aneropack anaerobic culture medium (Mitsubishi Gas Chemical Company).

[0057] The bacteria grown on MRS agar were inoculated into MRS liquid medium and pre-cultured under anaerobic conditions at 37°C for 24 hours. The obtained pre-culture medium was then mixed with MRS liquid medium to measure turbidity (OD). 660 The turbidity (OD) was adjusted to 1.0, and 1 mL of the adjusted pre-culture medium was inoculated into 99 mL of MRS liquid medium containing HMCA at each concentration (initial turbidity (OD)). 660 The culture was performed under anaerobic conditions at pH 7.5, 37°C for 24 hours, with a final HMCA concentration of 0.01 (see Tables 3-5).

[0058] After 24 hours, the turbidity (OD) of the culture medium was measured. 660 The ) was measured, and the concentrations of HMCA, HMPA, and 4-vinylguaiacol (4VG) in the culture medium were analyzed by HPLC. The standards used were HMCA (manufactured by Tokyo Chemical Industry Co., Ltd.), HMPA (manufactured by Tokyo Chemical Industry Co., Ltd.), and 4VG (manufactured by Alfa Aesar). The obtained HMCA concentration, HMPA concentration, and 4VG concentration (all in mM) were divided by the initial HMCA concentration to calculate the HMCA retention rate, HMPA reduction rate, and 4VG generation rate, respectively. The results are shown in Tables 3-5.

[0059] =HPLC conditions= Column: Wakosil II 5C18HG φ4.6mm × 250mm Column temperature: 40℃ Injection volume: 10μL Mobile phase: Solution A: 0.1% TFA aqueous solution, Solution B: Acetonitrile 0 min → 12 min: Mixture of solution A and solution B (82:18) 12 min → 20 min: Mixture of solution A and solution B (82:18) → (75:25) 20 min → 35 min: Mixture of solution A and solution B (75:25) → (70:30) 35 min → 40 min: Mixture of solution A and solution B (70:30) 40 min → 50 min: Mixture of solution A and solution B (10:90) 50 min → 65 min: Mixture of solution A and solution B (82:18) Flow rate: 0min→40min:1.0mL / min 40 min → 55 min: 1.4 mL / min 55 min → 65 min: 1.0 mL / min Detector: UV detector Detection wavelength: HMPA, 4VG: 280nm HMCA: 320nm

[0060] [Table 3]

[0061] [Table 4]

[0062] [Table 5]

[0063] Our inventors have found that, depending on the bacterial strain, growth is inhibited or, even if growth occurs, the HMPA reduction rate tends to decrease when the HMCA concentration is high. By using lactic acid bacteria of the genus Weissella that meet the requirements of the present invention, it has become clear that HMCA can be efficiently reduced to HMPA even at high concentrations (3 mM or higher). On the other hand, strains that could not reduce HMCA to HMPA even at an initial HMCA concentration of 1 mM (such as strain JCM7777) could not reduce it to HMPA even at 3 mM or 5 mM.

[0064] <Test Example 2> Culture Test-2 Weissella cibaria strains 011YN2 and 054YN2 were used in a jar fermenter for culture testing.

[0065] In this culture test, MRS medium (Merck) was used to culture lactic acid bacteria. Rice bran extract (Tsukuno Foods Industry Co., Ltd.) (HMCA content 80% by mass) was used as the HMCA source. A 2L jar fermenter was used for cultivation. The dissolved oxygen concentration was measured during cultivation and confirmed to be maintained at 0.2 mg / L or less.

[0066] Bacteria grown on MRS agar were inoculated into MRS liquid medium containing 50 mM HMCA, and pre-cultured under anaerobic conditions at 37°C for 24 hours. 20 mL of the resulting pre-culture was inoculated into 2 L of MRS liquid medium (HMCA: 50 mM) packed in the jar fermenter, and cultured at 37°C. HMCA and HMPA concentrations were measured 24 hours after inoculation of jar fermenters, in the same manner as in Test Example 1. The results are shown in Table 6.

[0067] [Table 6]

[0068] <Test Example 3> Culture Test-3 Weissella cibaria strain 054YN2 was used for culture tests in a jar fermenter.

[0069] In this culture test, the procedure was the same as in Test Example 2, except for changing the conditions for adding rice bran extract as the HMCA source. From 0 hours to 12 hours after starting culture in the jar fermenter, a total of 63 g of rice bran extract (HMCA concentration: 130 mM) or a total of 131 g (HMCA concentration: 270 mM) was added. The results are shown in Table 7.

[0070] [Table 7]

[0071] <Test Example 4> Culture Test-4 We tested the production of 3-phenylpropionic acids other than HMPA using the Weissella cibaria 054YN2 strain.

[0072] In this culture experiment, MRS medium (Merck) was used to culture lactic acid bacteria, and cinnamic acid and caffeic acid (both manufactured by Fujifilm Wako Pure Chemical Industries) were used as cinnamic acids. Anaerobic conditions were maintained using Aneropack for anaerobic culture (manufactured by Mitsubishi Gas Chemical Company).

[0073] Bacteria grown on MRS agar were inoculated into MRS liquid medium and pre-cultured at 37°C for 24 hours under anaerobic conditions. 1 mL of the resulting pre-culture was inoculated into 99 mL of MRS liquid medium containing cinnamic acids (see Table 9 for the type and final concentration of cinnamic acids), and cultured at pH 7.5, 37°C for 24 hours under anaerobic conditions.

[0074] After 24 hours, the turbidity (OD) of the culture medium was measured. 660 In addition to measuring the ) values, the concentrations of cinnamic acids and their reduced products, 3-phenylpropionic acid (PPA), in the culture medium were analyzed by HPLC. The HPLC was performed under the same conditions as in Test Example 1, and the standards and detection wavelengths are as shown in Table 8.

[0075] [Table 8]

[0076] The obtained concentrations of cinnamic acids and PPAs (both in mM) were divided by the initial concentration of cinnamic acids to calculate the residual cinnamic acid concentration and PPA reduction rate, respectively. The results are shown in Table 9.

[0077] [Table 9]

[0078] As shown in Table 9, Lactobacillus weissella was able to reduce cinnamic acids other than HMCA. Therefore, it was confirmed that the Lactobacillus weissella of the present invention can also be used for the production of 3-phenylpropionic acids other than HMCA.

[0079] TIFF0007905110000014.tif236155

Claims

1. A method for producing a 3-phenylpropionic acid analog compound represented by the following formula (I), 【Chemistry 1】 (In formula (I), R １１ ~R １５ These are, independently, a hydrogen atom, a hydroxyl group, an alkoxy group having 1 to 3 carbon atoms, or an alkyl group having 1 to 3 carbon atoms. The process includes a step of culturing lactic acid bacteria of the genus Weissella using a culture medium containing 3 mM or more of a cinnamic acid analog compound represented by the following formula (II), 【Chemistry 2】 (In formula (II), R ２１ ~R ２５ In the above equation (I), the corresponding R is... １１ ~R １５ These are identical to each other. The lactic acid bacteria were inoculated into MRS liquid medium containing 3 mM 4-hydroxy-3-methoxycinnamic acid, and the initial turbidity (OD) was measured. ６６０ A production method characterized by having the following properties (1a) to (3a) when cultured at 37°C for 24 hours under anaerobic conditions where the dissolved oxygen concentration in the liquid medium is 0.5 mg / L or less, with the value set to 0.01: (1a) Turbidity of 1.0 or higher; (2a) Reduction rate to 3-(4-hydroxy-3-methoxyphenyl)propionic acid is 80% or more; and (3a) Decarboxylation rate to 4-vinylguaiacol is less than 5%.

2. The production method according to claim 1, characterized in that the lactic acid bacteria is Weissella cibaria or Weissella confusa.

3. The production method according to claim 1, characterized in that the lactic acid bacteria is Weissella cibaria strain 011YN2 (accession number: NITE BP-03579), Weissella cibaria strain 054YN2 (accession number: NITE BP-03578), or Weissella confusa strain NRBC3957.

4. The production method according to claim 1, characterized in that the lactic acid bacteria, when inoculated into MRS liquid medium containing 5 mM 4-hydroxy-3-methoxycinnamic acid and cultured under anaerobic conditions at 37°C for 24 hours, possesses the following properties (1b) to (3b): (1b) Turbidity of 1.0 or higher; (2b) Reduction rate to 3-(4-hydroxy-3-methoxyphenyl)propionic acid is 80% or more; and (3b) Decarboxylation rate to 4-vinylguaiacol is less than 5%.

5. Lactic acid bacteria of the genus Weissella (excluding NBRC 3957, which belongs to Weissella confusa), Inoculate into MRS liquid medium containing 3 mM 4-hydroxy-3-methoxycinnamic acid and measure the initial turbidity (OD ６６０ A lactic acid bacterium characterized by having the following properties (1a) to (3a) when cultured at 37°C for 24 hours under anaerobic conditions where the dissolved oxygen concentration in the liquid medium is 0.5 mg / L or less, with the value of ) set to 0.01: (1a) Turbidity of 1.0 or higher; (2a) Reduction rate to 3-(4-hydroxy-3-methoxyphenyl)propionic acid is 80% or more; and (3a) Decarboxylation rate to 4-vinylguaiacol is less than 5%.

6. Lactobacillus of the genus Weissella, Inoculated into MRS liquid medium containing 5 mM of 4-hydroxy-3-methoxycinnamic acid, with an initial turbidity (OD ６６０ ), when cultured at 37 °C for 24 hours under anaerobic conditions where the dissolved oxygen concentration in the liquid medium is 0.5 mg / L or less, a lactic acid bacterium characterized by having the following properties (1b) to (3b): (1b) Turbidity of 1.0 or higher; (2b) Reduction rate to 3-(4-hydroxy-3-methoxyphenyl)propionic acid is 80% or more; and (3b) Decarboxylation rate to 4-vinylguaiacol is less than 5%.

7. The lactic acid bacterium according to claim 5, characterized in that it is Weissella cibaria.

8. The lactic acid bacterium according to claim 5, characterized in that it is Weissella cibaria strain 011YN2 (accession number: NITE BP-03579) or Weissella cibaria strain 054YN2 (accession number: NITE P-03578).

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