A method for improving the aroma of industrial kimchi by multi-strain synergistic fermentation

Abstract

The present invention belongs to the technical field of kimchi preparation, and specifically is a method for enhancing the aroma of industrial kimchi through multi-strain cooperative fermentation. The method comprises the following steps: (1) kimchi preparation; (2) lactic acid bacteria preparation, yeast preparation; (3) low-salt early fermentation; (4) high-salt late fermentation; (5) kimchi packaging. The present application adopts the cooperative regulation and control fermentation process technology of "inoculating lactic acid bacteria + yeast + environmental factors". The process adopts low salt and medium-low temperature in the early stage of fermentation, and the cooperative fermentation of enteric mesenteroides and Hansde Pasteurella candidiasis can quickly produce acid and enhance fragrance; in the middle and late stages of fermentation, high salt and medium temperature are adopted, and the cooperative fermentation of plant lactobacillus and oligosporic Kazakhstan yeast can quickly produce acid and enhance fragrance and simplify the flora, thereby stabilizing the quality. In the late stage of fermentation, the oligosporic Kazakhstan yeast consumes lactic acid to achieve acid control under high acidity, thereby achieving the purpose of preventing post-acidification. After the fermentation is completed, a rapid cooling process is adopted to inhibit microbial and enzyme activity and extend the shelf life.

Description

Technical Field

[0001] The invention belongs to the technical field of pickle preparation, and specifically relates to a method for improving the aroma of industrial pickles through multi-strain cooperative fermentation. Background Art

[0002] Kimchi is a traditional plant-based fermented product in my country. It is produced by microbial fermentation on the surface of vegetables or in the environment. It has a unique flavor and is popular with most consumers. During the natural fermentation of kimchi, the flora is complex, forming a dominant flora dominated by lactic acid bacteria and non-lactic acid bacteria. The dominant lactic acid bacteria include Lactobacillus pentosus, Lactobacillus plantarum, Lactococcus lactis, Lactobacillus brevis, Leuconostoc mesenteroides, Weissella, etc. The non-lactic acid bacteria include Enterobacter cloacae, Acinetobacter calcoaceticus, Raoultia ornithine lysate, Candida tropicalis, Kazakhstan yeast and Debaryomyces hansen. Among them, Enterobacter cloacae, Acinetobacter calcoaceticus and Raoultia ornithine lysate are unfriendly microorganisms in the early stage of kimchi fermentation. They gradually disappear with the increase of acidity in the middle and late stages of fermentation. The above yeasts often run through the entire fermentation process. Due to the use of natural fermentation, the flavor quality of each fermentation container varies greatly. When the temperature is too high and the salinity is too low, kimchi is prone to post-acidification, odor and other problems if it is not handled properly, affecting product quality.

[0003] In recent years, inoculation fermentation has been valued as a method to solve the unstable flavor quality of kimchi. The lactic acid bacteria involved in the research include Leuconostoc mesenteroides, Leuconostoc lactis, Lactococcus lactis, Weissella sinus, Pediococcus pentosaceus and Lactobacillus plantarum, which are mainly used to accelerate the maturation of kimchi, reduce nitrite, extend shelf life, inhibit pathogenic bacteria, improve functional properties and enhance sensory properties. However, current research focuses on the lactic acid bacteria level. The acid production metabolism of lactic acid bacteria accelerates the maturation of kimchi. However, the flavor of a single lactic acid bacteria is insufficient and the metabolites are few. Among them, there are fewer reports on the development of fermented aroma. The metabolic contribution of yeast to aroma is often reported in soy sauce and liquor, but there is less research and development in kimchi. At present, the development of kimchi yeast is still at the level of single strain pure strain fermentation, and there is less technical development on the synergistic fermentation of lactic acid bacteria and yeast.

[0004] Based on the above problems, it is urgent to develop technologies to enhance the aroma of kimchi and prevent the unstable quality of kimchi to solve the bottleneck problems of the kimchi industry. Summary of the invention

[0005] The purpose of the present invention is to provide a method for improving the aroma of industrial kimchi by using multi-strain cooperative fermentation to solve the problems existing in the prior art. In the present invention, based on the excellent strains screened and preserved in the early stage, the cooperative fermentation technology is used to improve the flavor quality of industrial kimchi and solve the problems of insufficient kimchi fermentation aroma, uneven quality and severe post-acidification.

[0006] In order to achieve the above invention object, the specific technical solution of the present invention is:

[0007] A method for improving the aroma of industrial kimchi by multi-strain cooperative fermentation, comprising the following steps:

[0008] (1) Preparation of kimchi: Put the vegetables into boiling water for a short time, then dry the surface water and add cold boiled water, and then add appropriate amounts of salt, old ginger, garlic and red pepper;

[0009] (2) Preparation of lactic acid bacteria: activating and culturing Leuconostoc mesenteroides PCY-3R18 and Lactobacillus plantarum PCY-5R4 respectively to obtain activated bacterial suspensions, and then centrifuging the bacterial suspensions to obtain bacterial sludges of Leuconostoc mesenteroides and Lactobacillus plantarum respectively;

[0010] Yeast preparation: activating and culturing oligosporic Kazakhstan yeast PCYWEL-1 and Hansdepasterioyces CICC 33343 respectively to obtain activated bacterial suspensions, and then centrifuging the bacterial suspensions to obtain oligosporic Kazakhstan yeast PCYWEL-1 and Hansdepasterioyces CICC 33343 bacterial muds respectively;

[0011] (3) Low-salt pre-fermentation: add Leuconostoc mesenteroides mud to the kimchi of step (1), shake well, and then ferment for 1-2 days; then add Pasteurella hansdei mud to the kimchi jar, shake well, and ferment;

[0012] (4) High-salt late fermentation: After step (3) is completed, add salt to the kimchi and mix well; add Lactobacillus plantarum PCY-5R4 bacterial slurry and ferment for 2-5 days, then add oligosporic Kazakhstan yeast PCYWEL-1 bacterial slurry and ferment for 2-5 days until fermentation is mature;

[0013] (5) Kimchi packaging: The fermented kimchi is quickly packaged, then quickly cooled at -18°C for 1-3 hours, and then refrigerated at 4°C.

[0014] Furthermore, the detailed steps of step (1) are as follows: removing yellow leaves and rotten leaves from vegetables, selecting fresh, moderately mature vegetables without impurities and debris, washing them clean, draining the water, boiling them in boiling water, taking them out and drying the surface water, and determining the boiling time according to the type of vegetables, and the boiling time is 0.5-2 minutes; mixing the processed vegetables and cold boiled water in a mass ratio of 1:1, and then adding appropriate amounts of old ginger, garlic and red pepper, the amount of salt added is 2-3% of the mass of the vegetables, and the amount of old ginger, garlic and red pepper is appropriate, and the old ginger, garlic and red pepper are washed 3 times with sterilized cold boiled water.

[0015] Furthermore, the detailed steps of preparing lactic acid bacteria in step (2) are as follows: taking Enterobacterium mesenteroides PCY-3R18 and Lactobacillus plantarum PCY-5R4, adding them into MRS medium at a ratio of 1-2% of the volume of the medium for activation, culturing them at 35-38°C for 24h, then taking out 1-2% of the volume of the medium and adding the bacterial solution into MRS medium for reactivation under the same conditions as above, finally obtaining a bacterial suspension after two activations, centrifuging the bacterial suspension at 4000-8000r / min for 10-20min, and obtaining bacterial mud of Enterobacterium mesenteroides PCY-3R18 and Lactobacillus plantarum PCY-5R4.

[0016] Furthermore, the detailed steps of preparing the yeast in step (2) are as follows: taking the oligosporic Kazakhstan yeast PCYWEL-1 and the Hansde Pasteurella CICC 33343, adding them to the PDA medium at a ratio of 1-2% of the volume of the medium for activation, culturing them at 25-28° C. for 24 hours, then taking out the bacterial solution of 1-2% of the volume of the medium and adding it to the PDA medium for reactivation under the same conditions as above, to obtain a bacterial suspension after two activations, and centrifuging the bacterial suspension at 4000-8000 r / min for 10-20 minutes to obtain the oligosporic Kazakhstan yeast PCYWEL-1 and the Hansde Pasteurella CICC 33343 bacterial sludge.

[0017] Furthermore, the specific steps of the low-salt preliminary fermentation in step (3) are as follows: adding the bacterial slurry of Enterococcus mesenteroides to the kimchi, shaking it well, and fermenting it at 10-15°C for 1-2 days; then adding the bacterial slurry of Pasteurella hansdei to the kimchi jar, shaking it well, and fermenting it at 15-20°C for 1-2 days.

[0018] Furthermore, the inoculum of Leuconostoc mesenteroides was shaken and inoculated with 10 6 CFU / g; the inoculation amount of Hansde Pasteurella was 10 5 CFU / g.

[0019] Furthermore, the detailed steps of the high-salt late fermentation in step (4) are as follows: after step (3) is completed, 3-5% of salt is added to the kimchi and mixed; Lactobacillus plantarum is added to the kimchi and fermented at 15-20° C. for 2-5 days; and then oligosporic Kazakhstan yeast is added to the kimchi and fermented at 15-20° C. for 2-5 days.

[0020] Furthermore, the detailed steps of kimchi packaging in step (5) are: quickly packaging the fermented and mature kimchi, and then quickly cooling it at -18°C for 1-3h, and then refrigerating it at 4°C.

[0021] Furthermore, the inoculum amount of Lactobacillus plantarum was 10 5CFU / g; the inoculum amount of oligosporic Kazakhstan yeast was 10 5 -10 6 CFU / g.

[0022] The present application also protects the pickles obtained by any of the above methods.

[0023] Compared with the prior art, the positive effects of the present invention are embodied in:

[0024] Traditional kimchi is mainly fermented naturally, and its fermentation is affected by many factors of the natural environment. Its flora is unstable and complex, the quality of the fermentation process is unstable and it is easy to spoil and "flower". In the late fermentation period, kimchi will have problems such as post-acidification and softening, unstable quality, and insufficient fermentation aroma. At present, some patents use inoculation of lactic acid bacteria to increase the acidity and fermentation cycle of kimchi, reduce nitrite, etc., but there is less development of aroma. This application uses "inoculation of lactic acid bacteria + yeast + environmental factors" to coordinately regulate the fermentation process technology. In the early stage of fermentation, the process uses low salt and medium and low temperature, and intestinal mesenteroides and Hansde Pasteurella co-ferment to quickly produce acid and enhance fragrance; in the middle and late stages of fermentation, high salt and medium temperature are used, and plant lactobacillus and oligosporic Kazakhstan yeast co-ferment to quickly produce acid and enhance fragrance and simplify the flora, thereby stabilizing the quality. In the late fermentation period, oligosporic Kazakhstan yeast consumes lactic acid to achieve acid control under high acidity, thereby preventing post-acidification. After the fermentation is completed, a rapid cooling process is used to inhibit microbial and enzyme activity and extend the shelf life. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 This is a curve chart of the analysis of fermentation characteristics of four lactic acid bacteria single bacteria and mixed bacteria in the examples of this application;

[0026] Figure 2 This is a curve chart of the analysis of fermentation characteristics of six non-lactic acid bacteria single bacteria and mixed bacteria in the Examples section of this application;

[0027] Figure 3 This is a curve diagram for analyzing the acid production capacity of the synergistic fermentation of lactic acid bacteria and yeast in some examples of this application;

[0028] Figure 4 This is a sensory analysis curve diagram of kimchi group and control group prepared by the method described in Example 1 in the examples section of this application. DETAILED DESCRIPTION

[0029] In order to make the purpose, technical scheme and advantages of the present invention clearer, the present invention is further described in detail below in conjunction with specific embodiments, but this should not be understood as the scope of the above-mentioned subject matter of the present invention being limited to the following embodiments. Without departing from the above-mentioned technical ideas of the present invention, various substitutions and changes are made according to common technical knowledge and customary means in the art, which should all be included in the scope of the present invention.

[0030] In this application document, % means wt% unless otherwise specified; the raw materials used are commercially available products unless otherwise specified.

[0031] The amount of each substance added in the following examples is based on the weight of the vegetable raw material after being cleaned and drained in step (1). The Leuconostoc mesenteroides PCY-3R18, Lactobacillus plantarum PCY-5R4, and oligosporous Kazakhstan yeast PCYWEL-1 in the present application are all prior art; among them, Leuconostoc mesenteroides PCY-3R18 is classified and named Leuconostoc mesenteroides, with a preservation number of GDMCC No.60197 and a preservation date of June 8, 2017; Lactobacillus plantarum PCY-5R4 is classified and named Lactobacillus plantarum, with a preservation number of GDMCC No.60196 and a preservation date of June 5, 2017; oligosporous Kazakhstan yeast PCYWEL-1 is classified and named Kazachstania exigua, with a preservation number of CGMCC No.12791 and a preservation date of July 15, 2016; Hansdepasteurella CICC 33343, a commercial product, was purchased from China Industrial Microbiology Culture Collection Center (CICC), website: http: / / m.china-cicc.org / .

[0032] Embodiment 1:

[0033] (1) Preparation of kimchi: Select fresh, moderately mature, and free of impurities and debris, clean and drain, cut into 2-5 cm pieces, and then boil in boiling water for 0.5 minutes, remove the surface moisture, add pre-treated vegetables and cold boiled water in a mass ratio of 1:1, and then add appropriate amounts of salt, old ginger, garlic and red pepper, the salt addition amount is 3%, and the old ginger, garlic and red pepper are washed 3 times with sterilized cold boiled water to reduce the microorganisms carried on the surface. The amount of old ginger, garlic and red pepper added is a conventional choice, and can be appropriately adjusted according to personal taste.

[0034] (2) Preparation of lactic acid bacteria: Take Enterobacter mesenteroides PCY-3R18 and Lactobacillus plantarum PCY-5R4, add them to MRS medium at a ratio of 1% of the volume of the culture medium for activation, and culture them at 37°C for 24 hours. Then, take out 1% of the volume of the culture medium from the bacterial solution and add it to MRS medium for reactivation under the same conditions as before. Finally, a bacterial suspension after two activations is obtained. The bacterial suspension is centrifuged at 6000 r / min for 10 minutes to obtain Leuconostoc mesenteroides and Lactobacillus plantarum mud.

[0035] Yeast preparation: Take oligosporic Kazakhstan yeast PCYWEL-1 and Hansde Pasteurella CICC 33343, add them to PDA medium at a ratio of 1% of the volume of the culture medium for activation, and culture them at 28°C for 24 hours. Then, take out 1% of the volume of the culture medium and add it to PDA medium for re-activation under the same conditions as before to obtain a bacterial suspension after two activations. The bacterial suspension is centrifuged at 6000r / min for 10min to obtain oligosporic Kazakhstan yeast and Hansde Pasteurella slurry.

[0036] (3) Low-salt pre-fermentation: Add Leuconostoc mesenteroides slurry to kimchi and shake well. The inoculation amount is 10 6 CFU / g, fermented at 15℃ for 2 days, using high inoculation amount and low temperature fermentation, low temperature can prevent the rapid growth of bacteria, and can make the inoculated bacteria ferment quickly, increase the acidity in the environment, mainly produce lactic acid, acetic acid and carbon dioxide, reduce the oxygen concentration in the environment, inhibit the growth of bacteria, and provide conditions for the growth of Lactobacillus plantarum; then add the sludge of Hansde Pasteurella to inoculate in the kimchi jar, shake well, and the inoculation amount is 10 5 CFU / g, fermented at 20℃ for 2d, the acid resistance of Hansde Pasteurella is weak, it exists in the early and middle stages of naturally fermented kimchi, and plays a synergistic fermentation role with Enterococcus mesenteroides, which can prolong the stable period of Enterococcus mesenteroides, produce isoamyl acetate and isoamyl alcohol, and provide vanilla and fusel flavors.

[0037] (4) High-salt late fermentation: After step (3), add 4% salt to the kimchi and mix well. Add Lactobacillus plantarum slurry with an inoculation amount of 10 5 CFU / g, fermented at 20℃ for 2 days, using low inoculation and medium temperature growth conditions, mainly to prolong the growth time of Leuconostoc mesenteroides and Pasteurella hansdei, slow down the growth rate of Lactobacillus plantarum, and prevent acidification of kimchi. This bacterium mainly produces lactic acid, which can increase the acidity of kimchi, gradually simplify the microbial flora of the kimchi system, and stabilize the quality of kimchi. Then add oligosporic Kazakhstan yeast, inoculated at 10 6 CFU / g, fermented at 20℃ for 3 days. Lactobacillus plantarum and oligosporic Kazakhstan yeast can synergistically ferment. The two strains have strong salt resistance, acid resistance and ethanol resistance, which can stabilize the quality of kimchi and increase the aroma of kimchi in the later stage of kimchi fermentation. Oligosporic Kazakhstan yeast can metabolize residual sugar and consume lactic acid as a substrate in the later stage to produce ester aroma substances such as ethyl acetate and ethyl butyrate, which can prevent the acidification of kimchi and increase the aroma of kimchi.

[0038] (5) Kimchi packaging: The fermented kimchi is quickly packaged and then rapidly cooled at -18°C for 2 hours to quickly reduce the enzyme activity and growth and metabolic activity of the microorganisms to achieve precise quality control. The kimchi product is then refrigerated at 4°C to extend the shelf life of the kimchi.

[0039] Embodiment 2:

[0040] (1) Preparation of kimchi: Remove yellow and rotten leaves from vegetables, select fresh, moderately mature, and free of impurities and debris, wash and drain, and boil in boiling water for 1 minute. Add the pretreated vegetables and cold boiled water in a mass ratio of 1:1, and then add appropriate amounts of salt, old ginger, garlic and red pepper, with the mass ratio of salt added being 3%. The old ginger, garlic and red pepper are washed three times with sterilized cold boiled water to reduce the microorganisms carried on the surface.

[0041] (2) Preparation of lactic acid bacteria: Take Enterobacter mesenteroides PCY-3R18 and Lactobacillus plantarum PCY-5R4, add them to MRS medium at a ratio of 1-2% of the culture medium volume for activation, and culture them at 37°C for 24 hours. Then, take out 1% of the culture medium volume of the bacterial solution and add it to MRS medium for reactivation under the same conditions as before. Finally, a bacterial suspension after two activations is obtained. The bacterial suspension is centrifuged at 6000 r / min for 15 minutes to obtain Leuconostoc mesenteroides and Lactobacillus plantarum mud.

[0042] (3) Yeast preparation: Kazakhstan oligosporic yeast PCYWEL-1 and Hansde Pasteurella CICC 33343 were added to PDA medium at a ratio of 1% of the volume of the medium for activation, and cultured at 28°C for 24 h. Then, 1% of the volume of the culture medium was taken out and added to PDA medium for reactivation under the same conditions as before to obtain a bacterial suspension after two activations. The bacterial suspension was centrifuged at 4000 r / min for 10 min to obtain Kazakhstan oligosporic yeast and Hansde Pasteurella slurry.

[0043] (3) Low-salt pre-fermentation: Add Leuconostoc mesenteroides slurry to kimchi and shake well. The inoculation amount is 10 6 CFU / g, fermented at 12℃ for 2 days; then added Hansde Pasteurella slurry to the kimchi jar, shaken well, and the inoculation amount was 10 5 CFU / g, fermented at 20℃ for 1 day.

[0044] (4) High-salt late fermentation: After step 3, add 3% salt to the kimchi and mix well. Add Lactobacillus plantarum slurry with an inoculation amount of 10 5 CFU / g, and fermented at 15°C for 4 days. Then, Kazakhstan oligosporic yeast was added, with an inoculation amount of 105 CFU / g, fermented at 15℃ for 3 days.

[0045] (5) Kimchi packaging: The fermented kimchi is quickly packaged and then rapidly cooled at -18°C for 1.5 hours to quickly reduce the enzyme activity and growth and metabolic activity of the microorganisms to achieve precise quality control. It is then refrigerated at 4°C to extend the shelf life of the kimchi.

[0046] Embodiment 3:

[0047] (1) Preparation of kimchi: Select fresh, non-corrupted radishes, clean them, drain them, and boil them in boiling water for 0.5 minutes. Add the pre-treated vegetables and cold boiled water in a mass ratio of 1:1, and then add appropriate amounts of old ginger, garlic and red pepper. The mass ratio of salt is 4%. The old ginger, garlic and red pepper are washed three times with sterilized cold boiled water to reduce the microorganisms carried on the surface.

[0048] (2) Preparation of lactic acid bacteria: Take Enterobacter mesenteroides PCY-3R18 and Lactobacillus plantarum PCY-5R4, add them to MRS medium at a ratio of 2% of the volume of the culture medium for activation, and culture them at 37°C for 24 hours. Then, take out 1% of the volume of the culture medium and add it to MRS medium for reactivation under the same conditions as before. Finally, a bacterial suspension after two activations is obtained. The bacterial suspension is centrifuged at 6000 r / min for 15 minutes to obtain Leuconostoc mesenteroides and Lactobacillus plantarum mud.

[0049] (3) Yeast preparation: Kazakhstan oligosporic yeast PCYWEL-1 and Hansde Pasteurella CICC 33343 were added to PDA medium at a ratio of 1% of the volume of the medium for activation, and cultured at 28°C for 24 h. Then, 2% of the volume of the culture medium was taken out and added to PDA medium for reactivation under the same conditions as before to obtain a bacterial suspension after two activations. The bacterial suspension was centrifuged at 5000 r / min for 10 min to obtain Kazakhstan oligosporic yeast and Hansde Pasteurella slurry.

[0050] (3) Low-salt pre-fermentation: Add Leuconostoc mesenteroides slurry to kimchi and shake well. The inoculation amount is 10 6 CFU / g, fermented at 15℃ for 1 day; then added Hansde Pasteurella slurry to the kimchi jar, shook well, and the inoculation amount was 10 5 CFU / g, fermented at 15℃ for 2 days.

[0051] (4) High-salt late fermentation: After step 3, add 3% salt to the kimchi and mix well. Add Lactobacillus plantarum slurry with an inoculation amount of 10 5CFU / g, and fermented at 20℃ for 1 day. Then, Kazakhstan oligosporic yeast was added, with an inoculation amount of 10 5 CFU / g, fermented at 20℃ for 2 days.

[0052] (5) Kimchi packaging: The fermented kimchi is quickly packaged and then rapidly cooled at -18°C for 1.5 hours to quickly reduce the enzyme activity and growth and metabolic activity of the microorganisms to achieve precise quality control. It is then refrigerated at 4°C to extend the shelf life of the kimchi.

[0053] Experiment 1 Analysis of fermentation characteristics of single bacteria inoculation

[0054] (1) Lactic acid bacteria inoculation and fermentation

[0055] Kimchi production: Cabbage purchased from the local farmers' market in Meishan was cleaned, dried, cut into 2-3 cm blocks, blanched in boiling water for 30 seconds, and then put into a 2L glass jar. Then hot salt water was added to the jar, and the edge of the jar was sealed with water. The mass ratio of the vegetable to the salt water was 1:2, and the salinity was 3% of the total mass. Four lactic acid bacteria, Leuconostoc mesenteroides PCY-3R18 (L.eu), Lactococcus lactis PCYTY-13 (Ll), Lactobacillus brevis PCYTY-7 (Lb) and Lactobacillus plantarum PCY-5R4 (Lp), were activated twice in MRS medium and then subjected to refrigerated centrifugation (4°C, 6000r / min, 10min). The centrifuged bacterial sludge was added to kimchi for constant temperature fermentation at 20°C. The four strains were added in the same proportion as mixed bacterial fermentation kimchi, and the total initial bacteria in each jar of kimchi was 10 6 CFU / g, mixed bacteria were matched 1:1 for each strain, 3 parallel samples. According to the fermentation rules, 0, 1, 2, 3, 4, 5 days of kimchi liquid were analyzed, and pH, total acid, colony count, organic acid and flavor components were determined immediately after sampling, and the remaining samples were stored at -20℃ for subsequent analysis.

[0056] Results: At the beginning of fermentation, L.eu, Ll and Lb started fermentation quickly, and the number of lactic acid bacteria reached the maximum value after 2 days of fermentation and then decreased rapidly, indicating that its tolerance was poor. In contrast, Lp started slowly and reached the peak value after 3 days of fermentation. The number was the largest in the late fermentation and decreased slowly, indicating that it had strong growth ability and good tolerance. The number of lactic acid bacteria in mixed fermented kimchi increased rapidly in the early stage and remained stable in the late fermentation. As the fermentation progressed, the acidity gradually increased, among which the acid production of L.eu, Ll and Lb increased rapidly, and the acid production rate slowed down after 2 days of fermentation. Lp produced acid slowly in the early stage of fermentation, and the acidity increased continuously after 2 days. The acid production of the four lactic acid bacteria in the late fermentation showed Lp>Lb>L.eu>Ll. Mixed fermented kimchi produced acid quickly in the early stage, and the acid production slowed down in the later stage. It shows that strain fermentation is conducive to the rapid acid production of kimchi and accelerates the maturation of kimchi, but the acid production capacity of each lactic acid bacteria is significantly different.

[0057] Lactic acid is the main organic acid in kimchi, and the lactic acid content directly affects the stability of microorganisms in kimchi. Lp produces a large amount of lactic acid in the middle and late stages of fermentation, while the other three lactic acid bacteria produce lactic acid rapidly in the early stages of fermentation, and the content is stable in the later stages. The lactic acid content in the late stages of kimchi fermentation showed Lp>mixed bacteria>Lb>L.eu>Ll. Acetic acid is one of the metabolites of heterofermentative lactic acid bacteria, and an appropriate amount of acetic acid has a promoting effect on the flavor of kimchi. L.eu and Lb have strong acetic acid production capabilities, while Ll has a relatively weak acetic acid production capability. Lp is a facultative heterofermentative lactic acid bacterium, and gradually produces a small amount of acetic acid after 2 days of fermentation. L.eu and Lb produce a large amount of mannitol, a flavor substance, after 1 day of fermentation, and Ll produces a small amount of mannitol after 2 days of fermentation. They all become stable in the later stages, showing L.eu>Lb>Ll.

[0058] The above results indicate that Leuconostoc mesenteroides and Lactobacillus plantarum are the dominant microorganisms in the early and late stages of kimchi fermentation. Leuconostoc mesenteroides ferments quickly, has weak post-acidification, and can produce flavor substances such as mannitol; while Lactobacillus plantarum starts slowly, but produces high lactic acid and has strong acid resistance. It is the dominant bacteria in the late stage of fermentation and is prone to post-acidification.

[0059] The volatile flavor substances in the fermentation process of kimchi fermented by single bacteria and mixed bacteria were analyzed by HS-SPME-GC-MS. Among them, L.eu has a strong ability to degrade the raw material flavor, but cannot degrade allyl nitrile and dimethyl trisulfide. The degradation ability shows L.eu>Lp>Lb>Ll. L.eu can prolong the stability period of Leuconostoc mesenteroides, produce isoamyl acetate and isoamyl alcohol, and provide vanilla and fusel flavors.

[0060] (2) Non-lactic acid bacteria inoculation and fermentation

[0061] The kimchi preparation process is the same as the above-mentioned single-bacteria inoculation lactic acid bacteria experiment, wherein the added bacteria are Enterobacter cloacae1B12 (EC), Acinetobacter calcoaceticus 0B7 (AC), Raoultella ornithinolytica 1B4 (RO), Candida tropicalis 5F2 (CT), Kazachstania exigua PCYWEL-1 (KE) and Debaryomyces hansenii CICC 33343 (DH). Studies have found that non-lactic acid bacteria microorganisms produce almost no organic acid in kimchi, pH>4, and cannot make kimchi mature, but studies have found that Kazachstania exigua PCYWEL-1 and Debaryomyces hansenii CICC 33343 can make kimchi produce ethanol, and Kazachstania exigua can produce ester aroma substances such as ethyl acetate and ethyl butyrate, which increase the aroma of kimchi.

[0062] Table 1 Differences in aroma components OAV of 6 non-lactic acid bacteria fermentations

[0063]

[0064]

[0065] (3) Multi-bacteria synergistic fermentation

[0066] The kimchi preparation process is similar to the above-mentioned single-bacteria inoculation lactic acid bacteria experiment. Leuconostoc mesenteroides and Lactobacillus plantarum are selected for synergistic fermentation with Pasteurella hansde and Kazakhstan oligosporus respectively. The study found that Leuconostoc mesenteroides and Pasteurella hansde have synergistic ability, which may be that Pasteurella hansde provides free amino acids as a growth nutrient factor for Leuconostoc mesenteroides, and there is a competitive relationship with Kazakhstan oligosporus, but this relationship is weak; Lactobacillus plantarum and Pasteurella hansde can not inhibit the growth of Lactobacillus plantarum, which may cause the acidification of kimchi, while Lactobacillus plantarum and Kazakhstan oligosporus have a competitive relationship in the synergistic fermentation, and the metabolism of Lactobacillus plantarum is inhibited, which may be related to factors such as the consumption of lactic acid by Kazakhstan oligosporus, reducing the total acid content.

[0067] (4) Sensory evaluation and shelf life of kimchi

[0068] Six trained panelists who were familiar with the properties of kimchi conducted sensory analysis on the fermented kimchi samples. Four sensory parameters were evaluated, namely crispness intensity, sourness intensity, aroma intensity, and comprehensive score, ranging from 1 to 5 points, equivalent to negligible, small, medium, strong, and very strong. The mature kimchi samples were packaged and stored at 4°C, and the time for the samples to change color, soften, and swell was recorded every one week.

[0069] The results show that yeast can increase the aroma of kimchi, and KE can reduce the acidity of kimchi. Lactobacillus plantarum can soften kimchi and reduce its crispness. The use of this patented synergistic fermentation can better improve the sensory quality of kimchi.

[0070] Table 2 Quality changes of kimchi samples during storage

[0071]

[0072] The above examples are only preferred implementations of this patent, but the protection scope of this patent is not limited thereto. It should be pointed out that, for ordinary technicians in this technical field, without departing from the principle of this patent, according to the technical solution and patent concept of this patent, several improvements and modifications can be made, and these improvements and modifications should also be considered within the protection scope of this patent.

Claims

1. A method for improving the aroma of industrial kimchi by multi-strain cooperative fermentation, characterized in that The following steps are involved: (1) Preparation of kimchi: Boil the vegetables in boiling water for a short time, then dry the surface water and add cold boiled water. Then add appropriate amounts of salt, ginger, garlic and red pepper. The amount of salt added is 2-3% of the weight of the vegetables. (2) Preparation of lactic acid bacteria: activating and culturing Leuconostoc mesenteroides PCY-3R18 and Lactobacillus plantarum PCY-5R4, respectively, to obtain activated bacterial suspensions, and then centrifuging the bacterial suspensions to obtain bacterial sludges of Leuconostoc mesenteroides PCY-3R18 and Lactobacillus plantarum PCY-5R4, respectively; Yeast preparation: activating and culturing the oligosporic Kazakhstan yeast PCYWYL-1 and the pastoria hansdei yeast CICC 33343 with the deposit number of CGMCC No. 12791 to obtain activated bacterial suspensions, and then centrifuging the bacterial suspensions to obtain the oligosporic Kazakhstan yeast PCYWYL-1 and the pastoria hansdei yeast CICC 33343 bacterial muds; (3) Low-salt pre-fermentation: add the bacterial slurry of Leuconostoc mesenteroides PCY-3R18 to the kimchi prepared in step (1), shake well, and then ferment for 1-2 days; then inoculate the bacterial slurry of Pasteurella hansde CICC 33343 into the kimchi jar, shake well, and ferment; (4) High-salt late fermentation: After step (3) is completed, add salt to the kimchi and mix well; add Lactobacillus plantarum PCY-5R4 bacterial slurry and ferment for 2-5 days, then add oligosporic Kazakhstan yeast PCYWYL-1 and ferment for 2-5 days until the fermentation is mature; (5) Kimchi packaging: The fermented kimchi is quickly packaged, then quickly cooled at -18°C for 1-3 h, and then refrigerated at 4°C; The specific steps of the low-salt preliminary fermentation in step (3) are as follows: adding the bacterial slurry of Leuconostoc mesenteroides PCY-3R18 to the kimchi, shaking it well, and fermenting it at 10-15° C. for 1-2 days; then adding the bacterial slurry of Pasteurella hansde CICC 33343 to the kimchi jar, shaking it well, and fermenting it at 15-20° C. for 1-2 days; The detailed steps of the high-salt late fermentation in step (4) are as follows: after step (3) is completed, 3-5% of table salt is added to the kimchi and mixed; Lactobacillus plantarum PCY-5R4 bacterial slurry is added and fermented at 15-20° C. for 2-5 days; then, oligosporic Kazakhstan yeast PCYWYL-1 bacterial slurry is added and fermented at 15-20° C. for 2-5 days.

2. The method for improving the aroma of industrial kimchi by multi-strain cooperative fermentation as claimed in claim 1, characterized in that: The detailed steps of step (1) are as follows: removing yellow leaves and rotten leaves from vegetables, selecting fresh, moderately mature vegetables without impurities and debris, washing them clean, draining the water, boiling them in boiling water, taking them out and drying the surface water, and determining the boiling time according to the type of vegetables, the boiling time is 0.5-2 minutes; mixing the processed vegetables and cold boiled water in a mass ratio of 1:

1.

3. The method for improving the aroma of industrial kimchi by multi-strain cooperative fermentation as claimed in claim 1, characterized in that: The detailed steps of preparing lactic acid bacteria in step (2) are as follows: taking Enterobacter mesenteroides PCY-3R18 and Lactobacillus plantarum PCY-5R4, adding them to MRS medium at a ratio of 1-2% of the volume of the culture medium for activation, culturing them at 35-38° C. for 24 hours, then taking out 1-2% of the volume of the culture medium and adding the bacterial solution to MRS medium for reactivation under the same conditions as above, finally obtaining a bacterial suspension after two activations, centrifuging the bacterial suspension at 4000-8000 r / min for 10-20 minutes, and obtaining bacterial slurry of Enterobacter mesenteroides PCY-3R18 and Lactobacillus plantarum PCY-5R4.

4. The method for improving the aroma of industrial kimchi by multi-strain cooperative fermentation as claimed in claim 1, characterized in that: The detailed steps of preparing yeast in step (2) are as follows: taking oligosporic Kazakhstan yeast PCYWYL-1 and Hansdepasterioyces CICC33343, adding them to PDA medium at a ratio of 1-2% of the volume of the medium for activation, culturing them at 25-28° C. for 24 h, then taking out 1-2% of the volume of the culture medium from the culture solution and adding it to PDA medium for reactivation under the same conditions as above, obtaining a bacterial suspension after two activations, centrifuging the bacterial suspension at 4000-8000 r / min for 10-20 min, and obtaining oligosporic Kazakhstan yeast PCYWYL-1 and Hansdepasterioyces CICC 33343 bacterial sludge respectively.

5. The method for improving the aroma of industrial kimchi by multi-strain cooperative fermentation as claimed in claim 1, characterized in that: The inoculum size of Leuconostoc mesenteroides PCY-3R18 was 10 6 CFU / g; the inoculum size of Hansde Pasteurella CICC 33343 was 10 5 CFU / g.

6. The method for improving the aroma of industrial kimchi by multi-strain cooperative fermentation as claimed in claim 1, characterized in that: The inoculum size of Lactobacillus plantarum PCY-5R4 was 10 5 CFU / g; the inoculum size of the oligosporic Kazakh yeast PCYWYL-1 was 10 5 -10 6 CFU / g.

7. Pickles obtained by the method according to any one of claims 1 to 6.

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