Weissella mesenteroides and application of Weissella mesenteroides in brewing of light soy sauce

Abstract

The invention discloses Weissella mesenteroides and application of the Weissella mesenteroides in brewing of light soy sauce, and belongs to the technical field of microorganisms and the field of food brewing. Due to the existence of a carbon decomposition metabolite repression effect, the invention provides the strain W.parameteroides WP-C8 which still has the capability of efficiently utilizing the xylose under the condition that the glucose concentration is far higher than that of the xylose. The xylose content of the strain can be respectively reduced by 29.5% and 52.4% in a glucose-xylose culture medium system and a koji-forming enzymolysis culture medium system. The W.paramesteroides WP-C8 is used for synergic soy sauce fermentation, and the content of amino acid nitrogen in soy sauce mash can be increased by 14.7%-24.9%.

Description

Technical Field

[0001] The invention relates to a strain of Weissella mesenteroides and application thereof in light soy sauce brewing, belonging to the technical field of microorganisms and the field of food brewing. Background Art

[0002] Soy sauces on the market in my country can be simply divided into two categories based on color: light soy sauce and dark soy sauce. Light soy sauce is lighter in color and is often used for seasoning, while dark soy sauce is darker in color and is mostly used for making marinated food and coloring. In Japan, soy sauce is more finely classified based on color and function, including Koikuchi Shoyu, Usukuchi Shoyu, Tamari Shoyu, Sai-Shikomi Shoyu and Shiro Shoyu. Among them, Koikuchi Shoyu and Usukuchi Shoyu are lighter in color, while Sai-Shikomi Shoyu, Usukuchi Shoyu and Shiro Shoyu are darker in color. To obtain these soy sauces of different shades, it is necessary to regulate and control the soy sauce during the koji fermentation, finished oil extraction and bottling and storage stages.

[0003] The koji making and fermentation stages of the soy sauce production process are the main stages for the formation of the soy sauce color. The regulation of the soy sauce color during the production stage can be achieved by controlling the browning reaction. For example, since low-salt solid soy sauce is fermented at a higher temperature, the Maillard reaction is stronger than at room temperature, and its color is darker than that of high-salt dilute soy sauce. In addition, the regulation of browning during the soy sauce fermentation process can be achieved by adjusting raw materials or intervening with microorganisms. For example, adding lactic acid football bacteria to the soy sauce fermentation process can increase the color rate of the soy sauce by 21.36%, increase the red index by 0.36, and make the color tend to be bright red. Using wheat bran hydrolyzate hydrolyzed by protease and saccharifying enzyme to prepare the brine for soy sauce fermentation can increase the color rate of the soy sauce by 47.6% compared with the control group.

[0004] At the stage of finished oil production, the color of the soy sauce can be directly controlled by post-processing the brewed soy sauce. For example, in the production process of dark soy sauce in my country, caramel color or red yeast rice pigment is added to obtain dark soy sauce with better sensory quality and coloring effect. In addition, there are also methods of using physical means to control the color, such as using adsorption or membrane filtration to decolorize the soy sauce after obtaining the finished soy sauce, so as to obtain light-colored soy sauce.

[0005] During the bottling and storage stage of soy sauce, storage conditions will further affect the color of the soy sauce. The higher the temperature and oxygen content during storage, and the stronger the light, the faster the browning of the soy sauce will deepen. The materials used for soy sauce packaging also affect the browning of soy sauce. Among the packaging materials such as glass bottles, PET bottles, PE bottles, PE films and nylon films, glass bottles have the best color protection effect. The color rate only increased by 0.83% after being stored at room temperature and natural light for 40 days. Adding browning inhibitors such as L-ascorbic acid, calcium chloride and citric acid can also help maintain the color of soy sauce.

[0006] The light-colored soy sauces on the market now mainly include Japanese light-taste soy sauce, white soy sauce, and white soup soy sauce, a traditional specialty soy sauce in northern Jiangsu, my country. Among the three types of soy sauces, light-taste soy sauce is relatively dark in color, while white soy sauce and white soup soy sauce are light yellow. In the process of brewing light-colored soy sauce, raw materials with lower bran content and brewing water with low iron content are usually used, and spores in the koji are removed to reduce the color of the soy sauce. Affected by the brewing process, the total nitrogen and amino acid nitrogen content of light-colored soy sauce are relatively lower than those of ordinary soy sauce. For example, the total nitrogen requirements for white soy sauce and light-taste soy sauce in the Japanese soy sauce standard are lower than those for thick soy sauce. At present, there are two main methods to obtain light-colored soy sauce: decolorization after brewing and adjusting the brewing process to directly brew light-colored soy sauce.

[0007] The main methods of decolorization after brewing are adsorption or membrane filtration. Among the adsorption materials such as activated carbon, activated clay, diatomaceous earth, magnesium oxide and silica gel, activated carbon has the strongest adsorption and decolorization ability for soy sauce, and activated carbon can also better maintain the amino acid and organic acid content of soy sauce. Some studies have used activated carbon adsorption to decolorize soy sauce, with a decolorization rate of 79.68%. Using macroporous adsorption resin instead of activated carbon to decolorize soy sauce can avoid the activated carbon method.

[0008] The environmental pollution and carbon residue caused by the membrane filtration method have been further improved to 86.39%. Compared with the adsorption method, the membrane filtration method is simpler and cleaner, and can sterilize while decolorizing, avoiding the problem of darkening color caused by heating and sterilization after the soy sauce is extracted. The effect of the membrane filtration method is related to the pore size of the membrane. For example, the decolorization rate of nanofiltration is between 86-96% and the decolorization rate of ultrafiltration is only 20%. The corresponding nanofiltration membrane also causes the total nitrogen, free amino acids, salt-free soluble solids and reducing sugars of the soy sauce to lose 5-43%, while these indicators of the soy sauce decolorized by ultrafiltration membrane have no obvious changes.

[0009] Adjusting the brewing process is mainly to adjust the ratio of soybeans and wheat in the soy sauce brewing process, and finally obtain light-colored soy sauce. Compared with decolorization after brewing, adjusting the brewing process avoids the extra costs brought by adsorbents and filter membranes, and also avoids the problem of loss of nutrients and flavor of soy sauce during the decolorization process. Light-taste soy sauce, white soy sauce and white soup soy sauce are all light-colored soy sauces directly brewed. The brewing of light-taste soy sauce is similar to that of ordinary soy sauce, except that the concentration of salt water used is higher, the seed koji used is light-taste seed koji, and fermented glutinous rice is added before pressing out the oil. White soy sauce and white soup soy sauce are different from ordinary soy sauce. In terms of raw material selection, they are not mainly soybeans and supplemented by wheat, but mainly wheat and supplemented by soybeans. Wheat can even account for more than 90% of the raw materials of white soy sauce. In addition to the different proportions of raw materials, the fermentation temperature of white soy sauce is lower and the fermentation time is shorter (60–90d). At the same time, in order to avoid color enhancement, white soy sauce is sterilized by low-temperature heating or no heating after oil is produced. However, due to the low protein content of the raw materials, the flavor of white soy sauce is lacking, and the umami taste is only half of that of light soy sauce.

[0010] Soy sauce is an important condiment originated from China and is well-known around the world. It can be used to flavor and color food. The coloring ability of soy sauce is closely related to the color of the soy sauce itself. Its color is mainly formed by the browning reaction during the fermentation process of soy sauce, including Maillard reaction, caramelization reaction and enzymatic browning reaction. Light soy sauce is a light-colored, shiny soy sauce that can maintain the original color of the ingredients while flavoring. At present, the production of light soy sauce still has the problems of high cost or poor flavor.

[0011] In the actual soy sauce fermentation system, the content of glucose is much higher than that of xylose. Due to the carbon catabolite repression (CCR) effect, when the glucose concentration is much higher than that of xylose, the strain that utilizes xylose is very likely to be unable to consume xylose smoothly. Therefore, it is an urgent problem to find a strain that can still efficiently utilize xylose in the presence of glucose. Summary of the invention

[0012] This study will analyze the browning type of high-salt dilute soy sauce fermentation process. On the basis of preliminarily determining the main browning reaction type of soy sauce fermentation process, the related substances that affect the browning reaction of soy sauce will be studied to provide a basis for the subsequent regulation of soy sauce browning and further reveal the formation mechanism of soy sauce color. At the same time, the related substances that affect the browning reaction of soy sauce will be regulated by screening bacteria and co-fermentation, and the brewing technology of light-colored soy sauce will be established.

[0013] When isolating strains that can utilize xylose, the culture medium used in this study uses xylose as the main carbon source. However, in the actual soy sauce fermentation system, the content of glucose is much higher than xylose, and there is a great difference between the screening culture medium and the actual soy sauce fermentation system. Due to the existence of the carbon catabolite repression (CCR) effect, the present invention provides a strain that still has the ability to efficiently utilize xylose when the glucose concentration is much higher than xylose.

[0014] The main research contents of this paper are as follows:

[0015] (1) To study the browning rules of high-salt dilute soy sauce fermentation process and determine the main types of browning that affect the browning process of soy sauce fermentation.

[0016] (2) Based on the determination of the main browning types in the soy sauce fermentation process, the key substances that affect the main browning types in the soy sauce fermentation process are studied.

[0017] (3) Screening bacteria from high-salt dilute soy mash that can utilize key substances that affect the main browning types in the soy sauce fermentation process, and examining the bacteria's ability to utilize the substances and their ability to use them in soy sauce fermentation, ultimately obtaining strains that can be used in soy sauce fermentation and efficiently utilize key substances that affect the main browning types in the soy sauce fermentation process.

[0018] (4) Using the screened strains to ferment soy sauce, and combining methods such as replacing raw materials to further control the key substances that affect the main browning types in the soy sauce fermentation process, ultimately obtaining light-colored soy sauce.

[0019] The present invention provides a strain of Weissella paramesenteroides WP-C8, which was deposited in the China Center for Type Culture Collection on December 24, 2024, with a deposit number of CCTCC NO: M 20242895.

[0020] The present invention provides a microbial agent, wherein the microbial agent contains the above-mentioned Weissella mesenteroides WP-C8 or its fermentation liquid, or contains the above-mentioned Weissella mesenteroides WP-C8 lysate, or contains living cells of the bacteria of Weissella mesenteroides WP-C8, dry bacteria of Weissella mesenteroides WP-C8 obtained by freeze-drying, immobilized cells of Weissella mesenteroides WP-C8, liquid agent of Weissella mesenteroides WP-C8, solid agent of Weissella mesenteroides WP-C8, or Weissella mesenteroides WP-C8 strain in any other form;

[0021] In one embodiment of the present invention, the amount of Weissella mesenteroides WP-C8 added to the microbial agent is at least: 1×10 8CFU / g.

[0022] The present invention also provides a starter, wheat koji, bran koji or other types of fermentation koji containing the above-mentioned Weissella mesenteroides WP-C8 and / or the above-mentioned microbial preparation.

[0023] The present invention also provides the use of the Weissella mesenteroides WP-C8 and / or the microbial preparation in the preparation of brewing condiments.

[0024] In one embodiment of the present invention, the condiment is soy sauce.

[0025] In one embodiment of the present invention, the application includes but is not limited to at least one of (a) to (e):

[0026] (a) Strengthening the number of lactic acid bacteria in the soy sauce fermentation process system;

[0027] (b) increasing the amino acid nitrogen content in soy sauce;

[0028] (c) increasing the content of organic acid and total acid in the soy sauce system;

[0029] (d) improving the color stability of soy sauce during shelf life;

[0030] (e) promoting the formation of flavor substances in the soy sauce system;

[0031] (f) Reduce the browning index of soy sauce during the fermentation process.

[0032] The present invention also provides a method for reducing the browning index of soy sauce during the fermentation process. The method comprises adding the Weissella mesenteroides WP-C8 or the microbial preparation during the fermentation process of the soy sauce.

[0033] The present invention also provides a method for preparing light-colored soy sauce, which comprises the following steps:

[0034] (1) Koji making process:

[0035] The raw materials are mixed evenly with water, and mixed evenly with wheat flour in a mass ratio of (6-7): (4-3), and after adding 1.5-2‰ of soy sauce koji essence accounting for the total mass of the mixed raw materials and mixing evenly, the mixture is cultured at 28°C-30°C, the koji is turned over in due time, and the koji is made for 40h-48h;

[0036] (2) Fermentation process:

[0037] The koji prepared in step (1) is mixed with saline containing NaCl (22-23%) in a volume ratio of 1:(2-3), so that the final salt concentration of the system is 14-18% (preferably 16%), and the above-mentioned Weissella mesenteroides WP-C8 or the above-mentioned microbial preparation, or the above-mentioned starter, malt koji, bran koji or other types of fermentation koji are added, and fermented at 25°C-30°C for 30-60 days; and, on the 10th day of fermentation, the above-mentioned Weissella mesenteroides WP-C8 or the above-mentioned microbial preparation, or the above-mentioned starter, malt koji, bran koji or other types of fermentation koji are added, and stirring is carried out in time during the fermentation.

[0038] In one embodiment of the present invention, in step (1), the raw materials include but are not limited to soybean meal and gluten powder.

[0039] In one embodiment of the present invention, in step (1), the raw material is soybean meal;

[0040] In one embodiment of the present invention, the raw materials are gluten powder and soybean meal.

[0041] In one embodiment of the present invention, the gluten powder and soybean meal are mixed in a mass ratio of 1:1.

[0042] In one embodiment of the present invention, the amount of Weissella mesenteroides WP-C8 added is at least: 1×10 8 CFU / g.

[0043] The present invention also provides the use of the Weissella mesenteroides WP-C8 and / or the microbial preparation in improving the flavor, color and storage stability of soy sauce.

[0044] The invention also provides light-colored soy sauce obtained by applying the preparation method.

[0045] Beneficial Effects

[0046] (1) 17 strains belonging to 5 genera that can utilize xylose were isolated from the dilute soy sauce mash of the present invention. A strain Weissella paramesenteroides WP-C8 that can efficiently utilize xylose was obtained by screening. The strain can reduce the xylose content by 29.5% and 52.4% in the glucose-sugar culture medium system and the koji enzymatic hydrolysis culture medium system, respectively. When W. paramesenteroides WP-C8 is used in the coordinated soy sauce fermentation, the amino acid nitrogen content in the mash can be increased by 14.7-24.9%.

[0047] (2) The present invention uses low pentosan raw material gluten to replace 50% soybean meal or strengthens the efficient xylose utilization strain to reduce the browning reaction in the high-salt dilute soy sauce fermentation process. Among them, replacing the raw materials and strengthening the fermentation of W. paramesenteroides WP-C8 at the same time has the best effect on reducing the browning of soy sauce, which can reduce the browning index and color rate of soy sauce by 86.7% and 94.8% respectively compared with commercially available light soy sauce. The amino acid nitrogen content of light soy sauce brewed by this technology is 0.96g / 100g -1 , the content of alcohol and ester in its volatile flavor substances increased significantly. Under simulated storage conditions, the increase in browning index of the light-colored soy sauce brewed in this study was 33.7% and 30.8% lower than that of commercially available light soy sauce and white soup soy sauce, respectively.

[0048] Biomaterial Deposit

[0049] A strain of Weissella paramesenteroides, classified and named Weissella paramesenteroides WP-C8, was deposited in the China Center for Type Culture Collection on December 24, 2024, with the deposit number CCTCC NO:M 20242895, and the deposit address is Wuhan University, Wuhan, China. BRIEF DESCRIPTION OF THE DRAWINGS

[0050] Figure 1 : Phylogenetic tree of strain W. paramesenteroides WP-C8.

[0051] Figure 2 : Changes in viable bacterial counts in co-fermented soy sauce.

[0052] Figure 3 : Changes of amino acid nitrogen and total acid in co-fermented soy sauce.

[0053] Figure 4 : Changes in viable bacterial counts in soy sauce after co-fermentation with raw materials replacement; LC1: only W. paramesenteroides WP-C8 was enhanced; LC2: only raw materials were replaced; LC3: raw materials were replaced and W. paramesenteroides WP-C8 was enhanced.

[0054] Figure 5 : Changes in amino acid nitrogen and total acid in soy sauce by replacing the co-fermented raw materials; among them, LC1 is: only strengthening W. paramesenteroides WP-C8; LC2 is: only replacing the raw materials; LC3 is: replacing the raw materials and strengthening W. paramesenteroides WP-C8.

[0055] Figure 6: Changes in browning index in soy sauce after co-fermentation with raw material replacement. LC1: only fortification of W. paramesenteroides WP-C8; LC2: only replacement of raw materials; LC3: replacement of raw materials and fortification of W. paramesenteroides WP-C8.

[0056] Figure 7 : Changes in the content of two sugars in soy sauce by replacing the co-fermented raw materials; among them, LC1 is: only strengthening W. paramesenteroides WP-C8; LC2 is: only replacing the raw materials; LC3 is: replacing the raw materials and strengthening W. paramesenteroides WP-C8.

[0057] Figure 8 : Comparison of the content of volatile flavor substances in soy sauce replaced by co-fermented raw materials; among them, LC1 is: only W. paramesenteroides WP-C8 is enhanced; LC2 is: only raw materials are replaced; LC3 is: raw materials are replaced and W. paramesenteroides WP-C8 is enhanced.

[0058] Fig. 9 : Comparison of the composition of volatile flavor substances in soy sauce replaced by synergistic fermentation raw materials; among them, LC1 is: only W. paramesenteroides WP-C8 is enhanced; LC2 is: only raw materials are replaced; LC3 is: raw materials are replaced and W. paramesenteroides WP-C8 is enhanced.

[0059] Fig.10 : Comparison of light soy sauce color.

[0060] Fig.11 : Changes of browning index of different soy sauces under simulated storage conditions. DETAILED DESCRIPTION

[0061] The culture medium involved in the following examples is as follows:

[0062] Xylose-enriched medium (g·L -1 ): 40 xylose, 1 magnesium sulfate monohydrate, 2 dipotassium hydrogen phosphate, 2 ammonium sulfate, 2 sodium chloride, 0.04 manganese sulfate.

[0063] Glucose-xylose medium (g·L -1 ): xylose 10, glucose 10, magnesium sulfate monohydrate 1, dipotassium hydrogen phosphate 2, ammonium sulfate 2, sodium chloride 2, manganese sulfate 0.04, yeast powder 3, solid culture medium supplemented with 1.5% agar powder.

[0064] Semisynthetic xylose medium (g·L -1): 20% xylose, 1% magnesium sulfate monohydrate, 2% dipotassium hydrogen phosphate, 2% ammonium sulfate, 2% sodium chloride, 0.04% manganese sulfate, 3% yeast powder, and 1.5% agar powder added to the solid culture medium.

[0065] The detection methods involved in the following embodiments are as follows:

[0066] Maillard reaction:

[0067] 1 mL of the solution containing sugar and amino acid was placed in a 90°C metal bath for 4 h, then quickly cooled to room temperature and its color index was measured. When studying the effect of sugar and amino acid types on the browning index, 10 g·L of sugar and amino acid were used. -1 The amino acid concentration used to compare the browning ability of glucose and xylose was 50 g·L -1 Glycine.

[0068] Determination of color index

[0069] Take 2g of mash in a 5mL centrifuge tube and centrifuge at 12000r / min. -1 Centrifuge for 5 minutes. Take the supernatant and filter it through a 0.45 μm filter membrane. Dilute it 10 times and measure its absorbance at 420, 460, 510, and 610 nm, and calculate it according to formula (1)-formula (4).

[0070] Browning index = OD 420 (1)

[0071]

[0072]

[0073] Determination of total bacterial count and lactic acid bacteria count in mash

[0074] The total bacterial count and lactic acid bacteria count were determined by the live bacteria counting method. Accurately weigh 1g of mash and add it to 9mL of physiological saline, and mix thoroughly on a vortex oscillator. Then dilute and spread on MRS and LB culture media, and count after culturing at 30℃ for 1-2 days.

[0075] Determination of physical and chemical indexes and flavor of soy sauce

[0076] (1) Determination of total acid and amino acid nitrogen in soy sauce

[0077] The determination method of total acid and amino acid nitrogen in soy sauce shall refer to the national standard GB5009.235-2016.

[0078] (2) Liquid chromatography method used for free amino acid detection:

[0079] The content of free amino acids in soy sauce was determined by o-phthalaldehyde (OPA) pre-column derivatization method. Liquid phase system: Agilent 1260; chromatographic column: Hypersil ODS-2 (250 mm × 4.6 mm, 2.5 μm); injection volume: 10 μL; mobile phase A: 5 g·L -1 Sodium acetate solution (containing 0.2mL·L -1 Triethylamine, 5 mL·L -1 Tetrahydrofuran); mobile phase B: 5 g·L -1 Sodium acetate-methanol-acetonitrile (volume ratio 1:2:2) solution; elution rate: 1 mL min -1 ; Elution time: 40min; Column temperature: 40℃; Detector: UV 338nm.

[0080] (3) Determination of volatile flavor substances in soy sauce

[0081] The volatile flavor compounds in soy sauce were determined by headspace solid phase microextraction (HS-SPME) and gas chromatography-mass spectrometry (GC-MS). Sample pretreatment: 5 mL of soy sauce sample was added to a 20 mL headspace bottle, and 2-octanol was added as an internal standard (final concentration was 100 μg / L -1 ). Extraction conditions: After the soy sauce sample was kept at 50℃ for 30min, the extraction head was placed 1cm below the liquid surface for 30min of adsorption, followed by thermal desorption at 250℃ for 15min at the gas chromatography injection port; GC-MS conditions for volatile flavor substance detection: GC-MS system: Pegasus BT; chromatographic column: HP-5ms (30m×250μm×0.25μm); heating program: initial 32℃, 5℃·min -1 Raise to 180℃, maintain for 2min, then 12℃·min -1 Increase to 240℃; carrier gas flow rate: 0.8mL·min -1 ; Ion source energy: 70eV.

[0082] Determination of sugar content

[0083] (1) Determination of pentosan content

[0084] Using xylose as the standard, the orcinol-hydrochloric acid method was used to determine the pentosan content. The specific operation is as follows:

[0085] Sample preparation: Accurately weigh 0.1 g of sample and add 20 mL of 2 mol·L -1 Boil HCl in water for 2 h, filter and dilute 10 times for later use.

[0086] Pentosan content detection: Take 3 mL of the treated sample and add 3 mL of 0.1% FeCl 3Mix with 0.3 mL of 1% orcinol, boil in a boiling water bath for 30 min, cool and add 3.7 mL of deionized water. After mixing, measure the absorbance at 580 nm and 670 nm, and calculate the pentosan content based on the difference between the two values ​​in the standard curve formula.

[0087] (2) Determination of glucose and xylose content

[0088] The contents of glucose and xylose were determined by high performance liquid chromatography. Liquid chromatography method used for glucose and xylose detection: Liquid phase system: LC-20A; Chromatographic column: ThermoFisher 69008-307780 (300mm×7.7mm, 8μm); Injection volume: 10μL; Mobile phase: 5mmol·L -1 H 2 SO 4 ; Elution rate: 0.6mL·min -1 ; Elution time: 15min; Column temperature: 40℃; Detector: differential detector.

[0089] Polyphenol oxidase activity assay

[0090] Extraction of polyphenol oxidase: Accurately weigh 30g of soy sauce koji into a mortar, add 0.6g of cross-linked polyvinyl pyrrolidone. Grind evenly in an ice bath, transfer to a beaker, add 60mL of PBS buffer (pH 6.8) pre-cooled at 4℃, and extract in a 4℃ refrigerator for 12h. Then, heat at 4℃ and 9000r·min. -1 Centrifuge for 30 minutes and take the supernatant to obtain the crude polyphenol oxidase enzyme solution.

[0091] Enzyme activity determination: Add 3.8 mL of catechol solution (0.3 mol·L -1 ) and 0.2 mL crude enzyme solution, and continuously monitor its absorbance at 420 nm at 40°C for 20 min. The blank control is 3.8 mL catechol solution with 0.2 mL crude enzyme solution treated with 10% TCA. The enzyme activity calculation formula is shown in formula (5).

[0092]

[0093] Wherein: V: volume of crude enzyme solution (mL); t: reaction time (min); OD420 value increase of 0.001 per milliliter of sample per minute is one unit of enzyme activity.

[0094] Investigation on the Storage Stability of Soy Sauce Color

[0095] Soy sauce was diluted 10 times and then placed in a 90°C metal bath for 4 hours to simulate the color formed by the Maillard reaction after long-term storage of soy sauce. After cooling to room temperature, its browning index was measured.

[0096] Soy sauce simulated fermentation system (enzymatic hydrolysis culture medium system):

[0097] (1) Koji making:

[0098] Add 1.4 times the volume of water (w / v) to the soybean meal to moisten the material for 4 hours, then sterilize at 121℃ for 20 minutes. After cooling to room temperature, mix the soybean meal and flour in a ratio of 6:4 (w / w), then add 0.15% soy sauce koji essence, mix well, and culture in a constant temperature and humidity incubator (temperature is 30℃, humidity is 95%) for 48 hours, turn the koji intermittently (about once every 16 hours), and the koji surface is covered with yellow-green hyphae.

[0099] (2) Mix the koji with distilled water at a ratio of 1:4 (w / w -1 ) in a certain ratio, and then heated at 40°C, 220 r·min -1 Oscillating enzyme hydrolysis for 5 h, 8000 r·min -1 Centrifuge for 20 minutes and take the supernatant. Then add 6% (w·v -1 ) of NaCl and sterilized at 115℃ for 20min.

[0100] Example 1: Isolation and identification of bacteria in mash

[0101] The specific steps are as follows:

[0102] (1) Screening of strains:

[0103] Koji making: Add 1.4 times the volume of water (w·v -1 ) for 4 hours, and then sterilized at 121℃ for 20 minutes. After cooling to room temperature, soybean meal and flour were mixed at a ratio of 6:4 (w·w -1 ) in a ratio of 0.15% soy sauce koji essence (purchased from Shanghai Difa Brewing Biological Products Co., Ltd.) based on the total mass of the mixed raw materials, mix thoroughly and culture in a constant temperature and humidity incubator (temperature: 30°C, humidity: 95%) for 48h, turning the koji intermittently (about once every 16h), and the koji is ready when its surface is covered with yellow-green mycelium.

[0104] Fermentation: Mix the koji with salt concentration of 19% (w·v -1 ) of saline at a ratio of 1:2.5 (w·w -1 ) in a ratio of 1:1 and 2:1, and ferment at a constant temperature of 30°C for 60 days. Stir every other day for the first 5 days of fermentation, and stir every 5 days thereafter to obtain the sauce mash.

[0105] (2) Bacterial isolation and screening:

[0106] 1) Take 1g of mash, add it to 10mL of xylose enriched medium, culture it at 30℃ for 24h, then transfer it to 5mL of xylose enriched medium with 4% (v / v) inoculation, and culture it at 30℃ for 24h. Then spread it on a semi-synthetic xylose medium plate, culture it at 30℃, and the larger colonies grown on the plate are the strains with potential strong xylose utilization ability.

[0107] 2) Screening of strains that efficiently utilize xylose:

[0108] The strain obtained in step 1) is placed in a glucose-xylose fermentation medium, cultured for 48 hours, and then centrifuged to obtain the supernatant, and the sugar content thereof is determined and a Maillard reaction is performed.

[0109] 3) Rescreening of simulated soy sauce fermentation system:

[0110] The strains screened in step 2) were cultured with lactic acid bacteria in MRS medium and Bacillus in LB medium until the absorbance of the bacterial solution at 600 nm was 1.0, and then inoculated into the soy sauce simulated fermentation system at an inoculum amount of 10% (v / v), and cultured at 30° C. for 14 days, and shaken once a day. After 14 days, the residual xylose content and browning index in the system were measured.

[0111] (3) Identification of bacterial species:

[0112] The genomic DNA of the strain was extracted using the bacterial genome extraction kit of Shanghai Shenggong Biological Company, and PCR amplification was performed using the 16SrRNA universal primers 27F (5'-AGAGTTTGATCCTGGCTCAG-3') and 1492R (5'-GGTTACCTTGTTACGACTT-3'). The PCR product was sent to Tianlin Biotechnology Wuxi Co., Ltd. for sequencing, and the sequencing results were compared by BLAST in NCBI. The phylogenetic tree of the strain was constructed using MEGA-X software ( Figure 1 ).

[0113] The results show that:

[0114] Two strains of Weissella paramesenteroides (W. paramesenteroides) were screened from the mash and named Weissella paramesenteroides WP-F8 and Weissella paramesenteroides WP-C8 respectively.

[0115] The Weissella paramesenteroides WP-C8 has been deposited in the General Microbiological Center of China National Committee for the Preservation of Microbiological Cultures, with the deposit number being CCTCC NO.M 20242895.

[0116] Example 2: Screening of strains that efficiently utilize xylose

[0117] When isolating strains that can utilize xylose, the culture medium used in this study uses xylose as the main carbon source. However, in the actual soy sauce fermentation system, the content of glucose is much higher than xylose, and there is a huge difference between the screening culture medium and the actual soy sauce fermentation system. Due to the existence of the carbon catabolite repression (CCR) effect, when the glucose concentration is much higher than xylose, the strain that utilizes xylose is very likely to be unable to consume xylose smoothly. Therefore, it is necessary to examine the ability of the strain to utilize xylose in the presence of glucose.

[0118] 1. Xylose utilization capacity and browning index

[0119] (1) The Weissella paramesenteroides WP-F8 and Weissella paramesenteroides WP-C8 strains obtained in Example 1 were respectively placed in a glucose-xylose fermentation medium, cultured at 30° C. for 48 h, and then centrifuged to determine the sugar content of the supernatant.

[0120] (2) the supernatant is subjected to Maillard reaction;

[0121] The specific steps are:

[0122] 1 mL of the supernatant obtained in step (1) and 50 g·L -1 The glycine solution was placed in a 90°C metal bath for 4 h, then rapidly cooled to room temperature, and its color index was measured.

[0123] Blank group: 1 mL glucose-xylose fermentation medium and 50 g·L -1 The glycine solution was placed in a 90°C metal bath for 4 h, then rapidly cooled to room temperature, and its color index was measured.

[0124] The results show:

[0125] (1) After 2 days of fermentation, the xylose content in the fermentation broth of all strains decreased, indicating that these strains have a certain ability to utilize xylose.

[0126] W. paramesenteroides WP-C8 can utilize xylose most efficiently, and the xylose content in its fermentation broth is reduced by 29.5%; the glucose content is reduced by 20.0%;

[0127] The xylose content in the fermentation broth of W. paramesenteroides WP-F8 decreased by 15.6%, and the glucose content decreased by 20.8%;

[0128] (2) The fermentation broth of strain W. paramesenteroides WP-C8 showed the highest reduction in browning index after Maillard reaction, which was 40.9% lower than that of blank culture medium.

[0129] The reduction in the browning index of the fermentation broth of strain W. paramesenteroides WP-F8 after Maillard reaction was 38.2% lower than that of the blank culture medium.

[0130] 2. Effect of strains on browning of enzymatic hydrolysis medium system

[0131] Lactic acid bacteria (Weissella paramesenteroides WP-F8, Weissella paramesenteroides WP-C8) were cultured using MRS medium under the following conditions: the bacterial solution was cultured in a test tube at 37°C until the absorbance of the bacterial solution at 600nm was 1.0, and then the prepared bacterial solution was inoculated into the soy sauce simulated fermentation system at an inoculum amount of 10% (v / v), and cultured at 30°C for 14 days, with shaking once a day. After 14 days, the residual xylose content and browning index in the system were measured.

[0132] According to the test, the xylose content in the soy sauce simulated fermentation system before inoculation was 10 g / L, and the glucose content was 10 g / L.

[0133] The results show:

[0134] The strain W. paramesenteroides WP-C8 showed a strong ability to reduce browning in the enzymatic hydrolysis medium system. The browning index of the system was reduced by 9.4% and the xylose content was reduced by 52.4%.

[0135] The browning index of strain W. paramesenteroides WP-F8 in the enzymatic hydrolysis medium system was reduced by 7.8%, and the xylose content was reduced by 47.5%.

[0136] Based on the performance of the strain in the glucose-xylose medium system and the koji enzymatic hydrolysis medium system, this study selected W. paramesenteroides WP-C8 as the strain used for the co-fermentation of light soy sauce for subsequent experiments.

[0137] Example 3: Effect of adding W. paramesenteroides WP-C8 to fermentation on soy sauce fermentation

[0138] Strengthening W. paramesenteroides WP-C8 in soy sauce system is the prerequisite for successful synergistic fermentation. In order to confirm whether the purpose of strengthening this strain can be achieved by adding W. paramesenteroides WP-C8, the total bacterial count, lactic acid bacteria count, total acid and amino acid nitrogen in soy sauce mash were investigated.

[0139] The specific steps are as follows:

[0140] Experimental Group:

[0141] Koji making: add 1.4 times volume of water (w·v -1 ) for 4 hours, and then sterilized at 121℃ for 20 minutes. After cooling to room temperature, soybean meal and flour were mixed at a ratio of 6:4 (w·w -1 ) in a ratio of 1:1 to 2:1, then add 0.15% soy sauce koji essence based on the total mass of the mixed raw materials, mix thoroughly and culture in a constant temperature and humidity incubator (temperature is 30°C, humidity is 95%) for 48h, turning the koji intermittently (about once every 16h), and the koji is ready when the surface is covered with yellow-green mycelium.

[0142] Fermentation: Mix the koji with salt concentration of 19% (w·v -1 ) of saline at a ratio of 1:2.5 (w·w -1 ) and fermented at a constant temperature of 30°C for 60 days. Stir every other day for the first 5 days of fermentation and every 5 days thereafter; at the same time, add W. paramesenteroides WP-C8 on the 0th and 10th days of fermentation, respectively, at an addition rate of 10 8 CFU·g -1 Sauce and mash.

[0143] The culture method of W.paramesenteroides WP-C8 bacterial agent is as follows: using MRS medium to culture lactic acid bacteria Weissella paramesenteroides WP-C8, and the culture conditions are: placing it in a test tube and culturing it at 37° C. until the absorbance value of the bacterial solution at 600 nm is 1.0.

[0144] Control group:

[0145] The specific implementation method is the same as the experimental group, except that no W. paramesenteroides WP-C8 bacterial agent is added during the fermentation process.

[0146] The results show:

[0147] (1) Figure 2As shown, the number of lactic acid bacteria in the experimental group with the addition of W. paramesenteroides WP-C8 was 1-2 orders of magnitude higher than that in the control group during the entire fermentation process (0-60d), and the total number of bacteria was 0.5-1 orders of magnitude higher than that in the control group, indicating that the addition of W. paramesenteroides WP-C8 during the soy sauce fermentation process can successfully strengthen the strain.

[0148] (2) Figure 3 As shown in the figure, the amino acid nitrogen content in the mash of the experimental group fortified with W. paramesenteroides WP-C8 was 14.7-24.9% higher than that of the control group. And the amino acid nitrogen increased faster: the amino acid nitrogen content of the fortified W. paramesenteroides WP-C8 rose rapidly to 0.8 g / 100 g in 0-20 days. -1 Then increase slowly.

[0149] The amino acid nitrogen content in the control group began to slowly increase at 30 days.

[0150] (3) Since W. paramesenteroides WP-C8 produces organic acids during the fermentation process, the total acid content of the experimental group fortified with W. paramesenteroides WP-C8 increased by 6.7-18.0% compared with the control.

[0151] The above results showed that W. paramesenteroides WP-C8 can be used for soy sauce fermentation and can improve the quality of soy sauce to a certain extent.

[0152] Example 4: Raw material adjustment and co-fermentation of light-colored soy sauce with W. paramesenteroides WP-C8

[0153] The specific steps are as follows:

[0154] 1. Preparation of light soy sauce

[0155] (1) Koji making:

[0156] Add 1.4 times the volume of water (w·v -1 ) for 4 hours, then sterilize at 121℃ for 20 minutes. After cooling to room temperature, mix the raw materials with flour at a ratio of 6:4 (w·w -1 ) in a ratio of 0.15% soy sauce koji essence (purchased from Shanghai Difa Brewing Biological Products Co., Ltd.) based on the total mass of the mixed raw materials, mix thoroughly and culture in a constant temperature and humidity incubator (temperature: 30°C, humidity: 95%) for 48h, turning the koji intermittently (about once every 16h), and the koji is ready when its surface is covered with yellow-green mycelium.

[0157] (2) Fermentation:

[0158] The koji was mixed with salt at a concentration of 19% (w·v -1 ) of saline at a ratio of 1:2.5 (w·w -1 ) in a ratio of 1:1 and 2:1, and ferment at a constant temperature of 30°C for 60 days. Stir every other day for the first 5 days of fermentation, and stir every 5 days thereafter to obtain the sauce mash.

[0159] 2. Experimental group settings

[0160] LC1: W. paramesenteroides WP-C8 (W. paramesenteroides WP-C8 + 100% soybean meal group) was strengthened to explore whether the enhanced xylose-utilizing strain can reduce the browning of soy sauce; the specific preparation method is the same as step 1, except that the raw material is adjusted to soybean meal; at the same time, in step (2), W. paramesenteroides WP-C8 was added on the 0th and 10th days of fermentation, respectively, with an addition amount of 10 8 CFU·g -1 The preparation method of W. paramesenteroides WP-C8 bacterial agent is the same as that in Example 3.

[0161] LC2: The raw material was set to 50% gluten + 50% soybean meal group to determine the effect of adjusting the raw material alone on reducing the browning of soy sauce. The specific preparation method is the same as step 1, except that the raw material was adjusted to 50% gluten + 50% soybean meal group;

[0162] LC3: The strain was strengthened and the raw material was replaced (W. paramesenteroides WP-C8 + 50% gluten + 50% soybean meal group) to explore the effect of strengthening the xylose strain in the fermentation with raw material replacement. The specific preparation method is the same as step 1, except that the raw material is adjusted to 50% gluten + 50% soybean meal; at the same time, in step (2), W. paramesenteroides WP-C8 was added on the 0th and 10th days of fermentation, respectively, and the addition amount was 10 8 CFU·g -1 The preparation method of W. paramesenteroides WP-C8 bacterial agent is the same as that in Example 3.

[0163] Control group: the raw material was set to 100% soybean meal; the specific preparation method was the same as step 1, except that the raw material was adjusted to 100% soybean meal;

[0164] According to the method of step 1, light-colored soy sauce is prepared.

[0165] 3. Experimental results

[0166] (1) Effect of adding W. paramesenteroides WP-C8 on microorganisms

[0167] like Figure 4 As shown, when W. paramesenteroides WP-C8 was not fortified, the number of lactic acid bacteria and the total number of bacteria in the gluten replacement group (LC2 group) were only 18.6-52.3% and 12.5-64.1% of the control group, respectively.

[0168] After fortification with W. paramesenteroides WP-C8, the lactic acid bacteria count and total bacterial count of the gluten replacement group (LC3 group) were 0.5-1 orders of magnitude higher than those of the control, and there was no significant difference in the lactic acid bacteria count and total bacterial count compared with the moromi group (LC1 group) only fortified with W. paramesenteroides WP-C8.

[0169] This indicates that strengthening W. paramesenteroides WP-C8 can restore the decrease in viable bacteria count caused by raw material replacement.

[0170] (2) Dynamic changes of total acid and amino acid nitrogen

[0171] like Figure 5 As shown, the contents of amino acid nitrogen in the four groups of sauce mash were LC3>LC2>LC1>control group from high to low.

[0172] Among them, the amino acid nitrogen content of LC3 (0.96g·100g -1 ) were 3.4-9.5% higher than those of LC2 and 10.1-28.0% higher than those of the control group.

[0173] In terms of total acid, the total acid content of LC3 was 5.1-21.1% higher than that of LC2 and 11.4-20.7% higher than that of the control group.

[0174] This shows that W. paramesenteroides WP-C8 can also play a role in the soy sauce fermentation process when gluten replaces soybean meal, and increase the amino acid nitrogen and total acid of soy sauce.

[0175] (3) Dynamic changes of browning index during soy sauce fermentation

[0176] like Figure 6 As shown, the browning index of LC3 was the lowest among the four groups. The browning index of LC3 was 9.4%-15.7% lower than that of the control group, and the browning index of LC3 was 5.2%-12.2% lower than that of LC1. The browning index of LC3 was 1.2%-4.3% lower than that of LC2. Figure 6It can be seen that this is because the xylose content in their (LC3, LC1, LC2) systems decreased by 24.2%-54.3%, 24.6%-46.3% and 9.3%-16.9%, respectively. The effect of strengthening W. paramesenteroides WP-C8 in reducing browning in the raw material replacement group is not as good as that in the control group. This may be because the xylose content in soy sauce in the raw material replacement group is lower, and W. paramesenteroides WP-C8 uses xylose relatively slowly, so the effect of reducing browning is relatively weak.

[0177] (4) Color index of soy sauce

[0178] The color indexes of the final soy sauce are shown in Table 1.

[0179] Table 1: Comparison of soy sauce color indicators

[0180]

[0181] Among the four groups, LC3 had the lowest browning index and color rate, which were 13.0% and 9.2% lower than those of the control. At the same time, LC3 also had the highest red index, which was 8.3% higher than that of the control.

[0182] The above experimental results show that using low pentosan raw materials for soy sauce fermentation and strengthening the strains that can utilize xylose can reduce the concentration of xylose in the soy sauce fermentation system, inhibit the Maillard reaction by reducing the substrate, and finally successfully obtain light soy sauce LC3 ( Figure 7 ).

[0183] (5) Effects of raw material replacement and strain synergy on volatile flavor compounds in soy sauce

[0184] In order to investigate the effects of raw material replacement and fortification with W. paramesenteroides WP-C8 on the flavor of soy sauce, the composition and content of volatile flavor compounds in four groups of soy sauce were investigated.

[0185] A total of 44 volatile flavor compounds were detected in all soy sauce samples, including 11 alcohols, 7 aldehydes, 8 ketones, 7 esters, 6 phenols, and 5 others such as furans and pyrazines.

[0186] Among them, the total amount of volatile flavor substances in soy sauce LC1 fortified with W. paramesenteroides WP-C8 was the highest, which was 266.88 μg·L -1 ( Figure 8). The total amount of volatile flavor substances in soy sauce increased after the fortification of W. paramesenteroides WP-C8. The total amount of volatile flavor substances in LC1 increased by 4.3% compared with the control. The main categories of volatile flavor substances that increased were alcohols and esters.

[0187] The replacement of raw materials reduced the total amount of volatile flavor substances in soy sauce, and the total amount of volatile flavor substances in LC3 decreased by 11.3% compared with the control. The strengthening of W. paramesenteroides WP-C8 could reduce the decrease in the content of volatile flavor substances caused by the replacement of raw materials to a certain extent, and the total amount of volatile flavor substances in LC3 increased by 6.3% compared with LC2.

[0188] Depend on Fig. 9 It can be seen that there are certain differences in the composition of volatile flavor substances among the four groups. LC3 has relatively high contents of flavor substances such as ethanol, 2-furan methanol, and ethyl acetate; LC2 has relatively high contents of flavor substances such as 3-acetylpyrrole and 2,4-dimethyl-3-carbonyl-pentanoic acid methyl ester; LC1 has relatively high contents of flavor substances such as 2,6-dimethylpyrazine and 3-octanone, while the control group has relatively high contents of flavor substances such as 4-ethylguaiacol and 4-ethylphenol.

[0189] (6) Quality and color analysis of light soy sauce

[0190] In order to investigate the difference between the light soy sauce LC3 obtained in this study and other soy sauces, the light soy sauce LC3 was compared with commercially available Haitian soy sauce and commercially available white soup soy sauce brewed with wheat as the main raw material. The results are shown in Table 2.

[0191] Table 2: Comparison of physical, chemical and color indicators of soy sauce

[0192]

[0193] As can be seen from Table 2, the amino acid nitrogen content of light soy sauce LC3 is roughly the same as that of light soy sauce, and is 1 times higher than that of white soup soy sauce.

[0194] The browning index and color rate of light-colored soy sauce LC3 are 86.7% and 94.8% lower than those of ordinary light soy sauce, which is close to that of white soup soy sauce.

[0195] Compared with white soup soy sauce, the red index of light soy sauce LC3 increased by 4%, and it is brighter and more lustrous overall, while white soup soy sauce is darker and has a relatively poor appearance ( Fig.10 )

[0196] (7) Storage stability of soy sauce color

[0197] Since there are a lot of reducing sugars and amino acids in soy sauce, the Maillard reaction will continue to proceed during the storage of soy sauce. The browning of soy sauce during storage is affected by various factors such as packaging materials, temperature and light. Therefore, the present invention accelerates the Maillard reaction of the three soy sauces by reacting in a metal bath for 4 hours, simulates the final browning index of the three soy sauces after long-term storage, and reduces the impact caused by storage conditions.

[0198] The specific methods to accelerate the Maillard reaction of the three soy sauces are:

[0199] The light-colored soy sauce LC3 obtained by the above method, commercially available light soy sauce and commercially available white soup soy sauce were reacted in a metal bath at 90° C. for 4 h. After the reaction, the browning index of each soy sauce was detected.

[0200] The results are as follows Fig.11 As shown, after the accelerated reaction, the browning index of the three soy sauces increased significantly. The browning index of light soy sauce and white soup soy sauce increased by 4.75 and 0.36, respectively, an increase of 221.1% and 212.0% compared with the original.

[0201] In contrast, the browning index of light-colored soy sauce LC3 only increased by 0.24, an increase of 146.6% compared with the original value, which is 33.7% lower than that of light soy sauce and 30.8% lower than that of white soup soy sauce.

[0202] This shows that compared with the other two soy sauces, LC3 has better color stability and the degree of color darkening after long-term storage is less.

[0203] The present invention combines the two methods of replacing low pentosan raw materials and strengthening the use of xylose strains, while reducing the source of xylose and increasing the consumption of xylose, to obtain light-colored soy sauce LC3. Compared with ordinary light soy sauce on the market, the browning index and color rate of LC3 are 86.7% and 94.8% lower. In addition, the amino acid nitrogen content is twice that of white soup soy sauce. While the color is close to that of white soup soy sauce, it is more shiny and more transparent, and the browning index is lower after simulating long-term storage.

[0204] Although the present invention has been disclosed as above in the form of a preferred embodiment, it is not intended to limit the present invention. Anyone familiar with this technology can make various changes and modifications without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention should be based on the definition of the claims.

Claims

1. A strain of Weissella paramesenteroides WP-C8 was deposited in the China Center for Type Culture Collection on December 24, 2024, with the deposit number CCTCC NO:M 20242895.

2. A microbial agent, characterized in that: The microbial agent contains the Weissella mesenteroides WP-C8 or its fermentation liquid according to claim 1, or contains the lysate of the Weissella mesenteroides WP-C8, or contains living cells of the Weissella mesenteroides WP-C8, dry cells of the Weissella mesenteroides WP-C8 obtained by freeze-drying, immobilized cells of the Weissella mesenteroides WP-C8, liquid agent of the Weissella mesenteroides WP-C8, solid agent of the Weissella mesenteroides WP-C8, or the Weissella mesenteroides WP-C8 strain in any other form; Preferably, in the microbial agent, the amount of Weissella mesenteroides WP-C8 added is at least: 1×10 8 CFU / g.

3. A starter, wheat koji, bran koji or other types of fermentation koji containing the Weissella mesenteroides WP-C8 according to claim 1 and / or the microbial preparation according to claim 2.

4. Use of Weissella mesenteroides WP-C8 according to claim 1 and / or the microbial preparation according to claim 2 in soy sauce production.

5. The use according to claim 4, characterized in that: The application includes but is not limited to at least one of (a) to (e): (a) Strengthening the number of lactic acid bacteria in the soy sauce fermentation process system; (b) increasing the amino acid nitrogen content in soy sauce; (c) increasing the content of organic acid and total acid in the soy sauce system; (d) improving the color stability of soy sauce during shelf life; (e) promoting the formation of flavor substances in the soy sauce system; (f) Reduce the browning index of soy sauce during the fermentation process.

6. A method for reducing the browning index of soy sauce during fermentation, characterized in that: During the soy sauce fermentation process, the Weissella mesenteroides WP-C8 described in claim 1 or the microbial preparation described in claim 2 is added.

7. A method for preparing light soy sauce, characterized in that: The method comprises the following steps: (1) Koji making process: The raw materials are mixed evenly with water, and mixed evenly with wheat flour at a mass ratio of 6-7:4-3, and after adding 1.5-2‰ of soy sauce koji essence accounting for the total mass of the mixed raw materials and mixing evenly, the mixture is cultured at 28℃-30℃, the koji is turned over in due time, and the koji is made for 40h-48h; (2) Fermentation process: The koji prepared in step (1) is mixed with saline containing NaCl in a volume ratio of 1:(2-3), and the Weissella mesenteroides WP-C8 described in claim 1 or the microbial preparation described in claim 2, or the starter, malt koji, bran koji or other types of fermentation koji described in claim 3 are added, and fermented at 25° C.-30° C. for 30-60 days; and, on the 10th day of fermentation, the Weissella mesenteroides WP-C8 described in claim 1 or the microbial preparation described in claim 2, or the starter, malt koji, bran koji or other types of fermentation koji described in claim 3 are added, and stirring is performed in a timely manner during the fermentation; Preferably, in step (1), the raw materials include but are not limited to soybean meal and gluten.

8. The preparation method according to claim 7, characterized in that: In step (1), the raw material is soybean meal; Preferably, the raw materials are gluten powder and soybean meal. More preferably, the gluten powder and soybean meal are mixed in a mass ratio of 1:

1. Preferably, the amount of Weissella mesenteroides WP-C8 added is at least: 1×10 8 CFU / g.

9. Use of Weissella mesenteroides WP-C8 according to claim 1 and / or the microbial preparation according to claim 2 in improving the flavor, color and storage stability of soy sauce.

10. Light-colored soy sauce obtained by the preparation method according to claim 7 or 8.

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