Lactobacillus brevis strain D15 for salt-reduced Pixian thick broad-bean sauce and application of lactobacillus brevis strain D15

Abstract

The invention discloses a lactobacillus brevis strain D15 for salt-reduced Pixian thick broad-bean sauce and application of the lactobacillus brevis strain D15. The preservation registration number of the strain D15 is CGMCC (China General Microbiological Culture Collection Center) No: 27698. The botanical classification of the strain D15 is short-growth-promoting lactobacillus in the bacterial field, the phylum of Thyrium, the Bacillus, the Lactobacillus, the Lactobacillus family and the Lactobacillus genus, and the Latin literature name of the strain D15 is Levibacillus brevis D15. The application of the strain D15 is as follows: firstly, the strain D15 is applied to preparation of salt-reduced Pixian thick broad-bean sauce; 2, application in preparation of salt-reduced chilli sauce; and 3, application in preparation of the salt-reduced broad bean sauce. After inoculation optimization is carried out on chili embryos or broad bean embryos by using the strain D15 disclosed by the invention, a salt-reduced fermentation system which takes chili and broad beans as raw materials and is under low salinity can be kept stable, the pH value is 4.2-4.5, and rancidity of the prepared fermented food is avoided; the enzyme activity can be obviously promoted, the fermentation time is shortened, the preparation cost is reduced, the flavor quality of the Pixian thick broad-bean sauce, the chili sauce and the broad bean sauce is obviously improved, and the industrial application value of the Pixian thick broad-bean sauce is obviously improved.

Description

TECHNICAL FIELD

[0001] The present application relates to a strain of Levilactobacillus brevis and its use. BACKGROUND

[0002] Levilactobacillus brevis is a strict heterofermentative lactic acid bacteria. Previous studies have shown that some Levilactobacillus brevis have the characteristics of antibacterial, acid tolerance, degradation of nitrite, and production of functional ingredients such as mannitol, exopolysaccharide (EPS) and gamma-aminobutyric acid, which help to enrich the sensory quality of fermented food, improve the safety of food, stimulate the function of probiotics, and endow the food with biological activity to promote health.

[0003] Sichuan bean paste (PiXian bean paste) is mainly made of chili, broad bean paste and salt as raw materials, and is a fermented food widely used by people. People like to eat bean paste mainly because of its unique flavor, especially the sauce aroma. Good bean paste has the characteristics of rich sauce aroma. The traditional Sichuan bean paste has a high salt content. The GB / T 20560-2006 Geographical Indication Product PiXian Bean Paste stipulates that the salt content is 15-22%. If 20g of bean paste is consumed per meal, the salt intake caused by bean paste alone is more than 9g / day, which will far exceed the "safety line" of 5g / day of salt intake. Therefore, with the enhancement of people's health consciousness and the need for healthy development of the industry, salt reduction of Sichuan bean paste has become one of the future development trends. However, salt reduction can easily cause deterioration of the flavor quality of bean paste. Screening of microorganisms with probiotic function to improve the flavor of salt-reduced bean paste is the key and effective way to solve this bottleneck problem. SUMMARY

[0004] The first application purpose of the present application is to provide a Levilactobacillus brevis strain D15 for salt-reduced PiXian bean paste, which can maintain and improve the flavor quality of PiXian bean paste under salt-reduced conditions, avoid quality deterioration such as rancidity, and significantly improve the industrial application value of PiXian bean paste.

[0005] The technical solution adopted by the present application to achieve its first application purpose is a Levilactobacillus brevis strain D15 for salt-reduced PiXian bean paste, which has a preservation registration number of CGMCC No: 27698.

[0006] The short-acting lactobacillus (Levilactobacillus brevis) strain D15 of the present application is isolated and screened from the fermented chili sauce prepared by the applicant in June 2018 in Chengdu, Sichuan Province, China, from the Institute of Agricultural Products Processing of Sichuan Academy of Agricultural Sciences (applicant). The applicant's name at that time was the Institute of Agricultural Products Processing of Sichuan Academy of Agricultural Sciences; now it has been renamed as the Institute of Agricultural Products Processing of Sichuan Academy of Agricultural Sciences (the former Institute of Food and Nutrition Health of Sichuan Academy of Agricultural Sciences).

[0007] The short-acting lactobacillus strain D15 of the present application has been preserved in the General Microbiological Center of China Microbial Culture Collection Management Committee on June 25, 2023, the preservation address is No. 1, Beichen West Road, Yard 3, Beijing, the preservation registration number is CGMCC No: 27698, and the classification and naming is Levilactobacillus brevis D15.

[0008] The short-acting lactobacillus strain D15 of the present application is identified as a short-acting lactobacillus of the bacterial kingdom, the Firmicutes phylum, the Bacilli class, the Lactobacillales order, the Lactobacillaceae family, and the Lactobacillus genus through 16S rDNA and whole genome sequence analysis. Its Latin name is Levilactobacillus brevis D15.

[0009] The morphological characteristics of the strain D15 of the present application are as follows:

[0010] After being cultured in MRS (De Man Rogosa Sharpe) liquid medium for 3 days, the morphological characteristics of the strain D15 of the present application are observed under a microscope as follows: the strain is short rod-shaped, single or linearly arranged, non-flagellated, and the size is 0.5-0.8x1-2uM.

[0011] After being cultured in MRS solid medium for 2 days, the morphological characteristics of the strain D15 of the present application are observed under a microscope as follows: the colony is protruding, the surface is smooth, the edge is regular, the color is milky white, and the size is 0.5-2mm.

[0012] After being cultured in MRS solid medium for 6 days, the morphological characteristics of the strain D15 of the present application are observed under a microscope as follows: the colony is protruding, the surface is smooth, the edge is regular, the color is milky white, and the size is 0.5-2mm.

[0013] The specific formula of the MRS (De Man Rogosa Sharpe) liquid medium is as follows: 10g of proteose peptone, 5g of beef powder, 4g of yeast powder, 20g of glucose, 5g of sodium acetate, 2g of potassium phosphate dibasic, 2g of triammonium citrate, 0.2g of magnesium sulfate, 0.05g of manganese sulfate, 1mL of Tween 80, and 1000mL of distilled water.

[0014] The specific formula of the MRS solid culture medium is: 10 g of proteose peptone, 5 g of beef powder, 4 g of yeast powder, 20 g of glucose, 5 g of sodium acetate, 2 g of potassium phosphate dibasic, 2 g of triammonium citrate, 0.2 g of magnesium sulfate, 0.05 g of manganese sulfate, 1 mL of Tween 80, and 20 mL of ethanol.

[0015] Identification of the strain D15 of the present application:

[0016] (1) Genomic DNA extraction. The isolated pure strain is inoculated into 2%vol ethanol MRS liquid medium, and cultured anaerobically at 30 DEG C for 3 days; 50 mL of bacterial liquid is centrifuged at 6000 rpm for 10 minutes to collect the bacterial bodies, which are washed with an equal volume of 0.01 Mol / L phosphate buffer solution, and then centrifuged at 6000 rpm for 10 minutes to collect the bacterial bodies. A bacterial genomic DNA extraction kit is used for DNA extraction.

[0017] (2) 16S rDNA sequence analysis

[0018] The 16S rDNA sequence of the strain D15 of the present application is obtained by using bacterial universal primers 27F (5'-AGAGTTTGATCMTGGCTCAG-3') and 1492R (5'-GGYTACCTTGTTACGACTT-3') to amplify the 16S rDNA sequence of the strain, and sending to Shengong Bioengineering (Shanghai) Co., Ltd. for cloning and sequencing. The 16S rDNA sequence is as follows:

[0019]

[0020] The above sequences were compared in EzBioCloud (https: / / www.ezbiocloud.net / ). The results showed that strain D15 had the highest similarity with the following standard strains of Levilactobacillus: Levilactobacillus brevis ATCC 14869 T , Levilactobacillus angrenensis M1530-1 T , and Levilactobacillus suantsaiihabitans R19 T , with similarities of 99.80%, 98.82%, and 98.79%, respectively. This result supports that strain D15 is Levilactobacillus brevis.

[0021] (3) Whole genome alignment analysis

[0022] The whole genome of strain D15 is 2543880 bp long, with a GC content of 45.71%, containing one genome and ten plasmids. The whole genome is 2364122 bp long, and the genome and six plasmids have formed a ring. The whole genome sequence was uploaded to Calculate ANI (https: / / www.ezbiocloud.net / tools / ani) for average nucleotide identity (ANI) analysis. The ANI value of D15 and the standard strain Levilactobacillus brevis ATCC 367 TT is 99.31%. Using GGDC v2.1 software (https: / / ggdc.dsmz.de / ggdc.php#) for in silico DNA-DNA hybridization analysis (DDH), it was calculated that the DDH value of D15 and the standard strain Levilactobacillus brevis ATCC 367 T is 90.50%. The recommended threshold for ANI species is 95%, and the threshold for DDH is 70%. This shows that it is Levilactobacillus brevis

[0023] Through the above 16S rDNA sequence analysis and whole genome alignment analysis, the strain D15 of the present application is identified as Levilactobacillus brevis.

[0024] Experiments prove that the Levilactobacillus brevis strain D15 of the present application can keep the salt-reduced fermentation system (such as Pixian bean sauce) with chili pepper and / or broad bean as raw materials stable, with pH value between 4.2-4.5, avoiding rancidity; in the salt-reduced fermentation system with chili pepper or broad bean as raw materials, inoculation of the strain can significantly promote enzyme activity, shorten fermentation time, and improve the flavor quality of chili (bean) sauce, which is an excellent functional strain for preparing bean sauce and chili sauce, and has obvious industrial application value.

[0025] The second object of the present application is to provide the use of the above-mentioned Levilactobacillus brevis strain D15.

[0026] The first technical solution adopted by the present application to achieve the second object of the present application is the use of the above-mentioned Levilactobacillus brevis strain D15, which is applied in the preparation of salt-reduced Pixian bean sauce.

[0027] Further, the above-mentioned application of the Levilactobacillus brevis strain D15 in the preparation of salt-reduced Pixian bean sauce has the following specific methods:

[0028] (1) Strain activation: inoculate the Levilactobacillus brevis strain D15 into MRS liquid medium, and culture at 28-32℃ for 3-4 days to obtain D15 activated seed culture.

[0029] (2) Inoculation and fermentation: add the D15 activated seed culture to the intermediate raw material, i.e., zero-salt fermented chili embryo, in the preparation of salt-reduced Pixian bean sauce at a mass ratio of 1-2%, and then ferment at 30-37℃ for 48-72 hours to obtain optimized zero-salt fermented chili embryo.

[0030] (3) Optimization of salt-reduced Pixian bean sauce: in the preparation of salt-reduced Pixian bean sauce, use the optimized zero-salt fermented chili embryo to replace the zero-salt fermented chili embryo in the preparation step of the zero-salt fermented chili embryo, to obtain the optimized salt-reduced Pixian bean sauce.

[0031] The second technical solution adopted by the present application to achieve the second object of the present application is the use of the above-mentioned Levilactobacillus brevis strain D15, which is applied in the preparation of salt-reduced chili sauce.

[0032] The use of the above-mentioned Levilactobacillus brevis strain D15 is applied in the preparation of salt-reduced chili sauce.

[0033] Furthermore, the application of the above-mentioned Lactobacillus breve strain D15 in the preparation of reduced-salt chili sauce is specifically carried out as follows:

[0034] (1) Strain activation: Inoculate the Lactobacillus breve strain D15 into MRS liquid culture medium and culture at 28-32°C for 3-4 days to obtain D15 activated strain culture medium.

[0035] (2) Preparation of reduced-salt pepper embryos: fresh peppers are washed, drained, and the stems and peduncles are removed, and then crushed with a crusher to obtain pepper material; salt is added to the pepper material to make the salinity reach 8-10%, thereby obtaining reduced-salt pepper embryos;

[0036] (3) The D15 activated bacterial culture solution is inoculated onto the salt-reduced pepper embryos at a mass ratio of 1-2%, and natural fermentation is carried out for 25-30 days to obtain the improved salt-reduced pepper sauce.

[0037] The third technical solution adopted by the present invention to achieve the second invention object is to use the above-mentioned Lactobacillus breve growth-promoting strain D15 in the preparation of reduced-salt broad bean paste.

[0038] Furthermore, the application of the above-mentioned Lactobacillus breve strain D15 in the preparation of salt-reduced broad bean paste is specifically carried out as follows:

[0039] (1) Strain activation: Inoculate the Lactobacillus breve strain D15 into MRS liquid culture medium and culture at 28°C-32°C for 3-4 days to obtain D15 activated strain culture medium.

[0040] (2) Preparation of reduced-salt broad bean embryos: 10 parts by weight of shelled and soaked broad bean petals are mixed with 2.8-3.2 parts by weight of wheat flour and 0.1-0.12 parts by weight of Shanghai-brewed 3.042% Aspergillus oryzae, and koji is made at 34-36°C for 45-50 hours to obtain broad bean embryos; salt and water are then added to adjust the salinity of the broad bean embryos to 10-12% and the moisture to 60-65% to obtain reduced-salt broad bean embryos;

[0041] (3) Inoculate the D15 activated bacterial culture solution into the salt-reduced broad bean embryos at a mass ratio of 1.0-1.5%, and allow to ferment naturally for 18-25 days to obtain the salt-reduced broad bean paste.

[0042] Compared with the prior art, the present invention has the following beneficial effects:

[0043] I. Experiments have proved that inoculating the strain D15 of the application on optimized pepper embryos or chickpea embryos can keep the low-salinity salt-reducing fermentation system stable, with a pH value of 4.2-4.5, avoiding rancidity, and can significantly promote enzyme activity, shorten fermentation time, and significantly improve the flavor quality of fermented foods such as Pixian bean sauce using fermented pepper sauce and / or fermented chickpea sauce as raw materials. Strain D15 is an extremely rare beneficial microorganism that can significantly improve the application value of Pixian bean sauce industry.

[0044] II. The strain D15 of the application is screened and isolated from salt-reducing fermented pepper sauce and belongs to Levilactobacillus brevis without genetic modification. It is extremely easy to popularize and implement in fermented foods (pepper sauce, bean sauce) with low implementation cost.

[0045] The application will be further described in detail below in combination with the accompanying drawings and specific embodiments. BRIEF DESCRIPTION OF DRAWINGS

[0046] Figure 1 is the flagella staining morphological diagram of D15 strain. DETAILED DESCRIPTION

[0047] Example 1

[0048] One specific way of the application is a Levilactobacillus brevis strain D15 for salt-reducing Pixian bean sauce, which has a preservation registration number of CGMCC No: 27698.

[0049] The strain D15 is isolated and screened from the fermented pepper sauce prepared by the applicant, Sichuan Provincial Academy of Agricultural Sciences (the applicant), in Chengdu, Sichuan Province, China in June 2018.

[0050] The Levilactobacillus brevis strain D15 has been preserved in the China General Microbiological Culture Collection Center on June 25, 2023, with a preservation address of No. 1, Beichen West Road, Yard 3, Beijing, Chaoyang District, and a preservation registration number of CGMCC No: 27698.

[0051] The strain D15 is identified as Levilactobacillus brevis of the bacterial kingdom, the Firmicutes phylum, the Bacilli class, the Lactobacillales order, the Lactobacillaceae family, and the Lactobacillus genus through 16S rDNA and whole genome sequence analysis. Its Latin name is Levilactobacillus brevis D15.

[0052] The use of the Levilactobacillus brevis strain D15 is in the preparation of salt-reducing Pixian bean sauce, and the specific method is as follows:

[0053] (1) Strain activation: Lactobacillus brevis strain D15 was inoculated into MRS liquid medium and cultured at 28°C for 3 days to obtain D15 activated seed culture;

[0054] (2) Inoculation fermentation: The D15 activated seed culture was added to the intermediate raw material, zero-salt fermented chili embryo, for preparing reduced-salt Pi Xian bean paste at a mass ratio of 1%, and then fermented at 30°C for 72 hours to obtain optimized zero-salt fermented chili embryo;

[0055] (3) Optimization of reduced-salt Pi Xian bean paste: In the preparation step of reduced-salt Pi Xian bean paste, the optimized zero-salt fermented chili embryo was used to replace the zero-salt fermented chili embryo, thereby obtaining the optimized reduced-salt Pi Xian bean paste.

[0056] Tests showed that in the entire preparation and fermentation period of the reduced-salt Pi Xian bean paste inoculated with strain D15, the pH value of the fermentation system was maintained between 4.2 and 4.5, which could effectively stabilize the microbial community in the product, promote enzyme activity, and avoid rancidity; and could significantly promote enzyme activity and significantly improve the flavor quality of the reduced-salt Pi Xian bean paste product.

[0057] Example 2

[0058] The Lactobacillus brevis strain D15 used in the reduced-salt Pi Xian bean paste of this example is the same as the strain D15 of Example 1.

[0059] The use of the Lactobacillus brevis strain D15 of this example is in the preparation of reduced-salt Pi Xian bean paste, and the specific method is as follows:

[0060] (1) Strain activation: Lactobacillus brevis strain D15 was inoculated into MRS liquid medium and cultured at 28°C for 3 days to obtain D15 activated seed culture;

[0061] (2) Inoculation fermentation: The D15 activated seed culture was added to the intermediate raw material, zero-salt fermented chili embryo, for preparing reduced-salt Pi Xian bean paste at a mass ratio of 1%, and then fermented at 30°C for 72 hours to obtain optimized zero-salt fermented chili embryo;

[0062] (3) Optimization of reduced-salt Pi Xian bean paste: In the preparation step of reduced-salt Pi Xian bean paste, the optimized zero-salt fermented chili embryo was used to replace the zero-salt fermented chili embryo, thereby obtaining the optimized reduced-salt Pi Xian bean paste.

[0063] The fermentation period of the reduced-salt Pi Xian bean sauce prepared after inoculating the strain D15 in the example is tested, the pH value of the fermentation system is maintained between 4.2-4.5, the acidity condition can effectively stabilize the microbial community in the product, promote enzyme activity, and avoid rancidity; and can significantly promote enzyme activity, and significantly improve the flavor quality of the reduced-salt Pi Xian bean sauce product.

[0064] Example 3

[0065] The short Lactobacillus brevis strain D15 for preparing reduced-salt Pi Xian bean sauce in the example is the same as the strain D15 in Example 1.

[0066] The use of the short Lactobacillus brevis strain D15 in the example is the application in the preparation of reduced-salt Pi Xian bean sauce, and the specific method is as follows:

[0067] (1) Strain activation: inoculate the short Lactobacillus brevis strain D15 into MRS liquid medium, and culture at 30℃ for 3.5 days to obtain D15 activated seed culture liquid;

[0068] (2) Inoculation and fermentation: add the D15 activated seed culture liquid to the intermediate raw material, i.e., zero-salt fermented chili embryo, in the preparation of reduced-salt Pi Xian bean sauce at a mass ratio of 1.5%, and then ferment at 34℃ for 60 hours to obtain optimized zero-salt fermented chili embryo;

[0069] (3) Optimization of reduced-salt Pi Xian bean sauce: in the preparation of reduced-salt Pi Xian bean sauce, the optimized zero-salt fermented chili embryo is used to replace the zero-salt fermented chili embryo in the preparation step of the zero-salt fermented chili embryo, and the optimized reduced-salt Pi Xian bean sauce is obtained.

[0070] The fermentation period of the reduced-salt Pi Xian bean sauce prepared after inoculating the strain D15 in the example is tested, the pH value of the fermentation system is maintained between 4.2-4.5, the acidity condition can effectively stabilize the microbial community in the product, promote enzyme activity, and avoid rancidity; and can significantly promote enzyme activity, and significantly improve the flavor quality of the reduced-salt Pi Xian bean sauce product.

[0071] The preparation process of reduced-salt Pi Xian bean sauce described in the application is a disclosed prior art, such as the Chinese patent with the name of "a preparation method of reduced-salt Pi Xian bean sauce" and the patent number of 202111359175.8.

[0072] Obviously, the intermediate raw material, i.e., zero-salt fermented chili embryo, in the preparation process of reduced-salt Pi Xian bean sauce is the "zero-salt fermented chili embryo" obtained in step B in the Chinese patent with the number of 202111359175.8.

[0073] The preparation steps of the zero-salt fermented pepper embryo required in the preparation of reduced-salt Pixian bean paste are C, D, and E steps of Chinese Patent No. 202111359175.8; the optimized zero-salt fermented pepper embryo is used to replace the zero-salt fermented pepper embryo, that is, the "zero-salt fermented pepper embryo" in the C, D, and E steps of Chinese Patent No. 202111359175.8 is replaced by "optimized zero-salt fermented pepper embryo".

[0074] Example 4

[0075] The short Lactobacillus brevis strain D15 for reduced-salt Pixian bean paste in this example is the same as the strain D15 in Example 1.

[0076] The use of the short Lactobacillus brevis strain D15 in this example is in the preparation of reduced-salt pepper sauce, and the specific method is as follows:

[0077] (1) Strain activation: inoculate the short Lactobacillus brevis strain D15 into MRS liquid medium and cultivate at 32°C for 4 days to obtain D15 activated seed culture.

[0078] (2) Preparation of reduced-salt pepper embryo: wash, drain, remove the stem and peduncle, and then crush the fresh peppers with a crusher to obtain pepper material; add salt to the pepper material to make the salinity reach 10% to obtain a reduced-salt pepper embryo.

[0079] (3) Inoculate the D15 activated seed culture into the reduced-salt pepper embryo at a mass ratio of 1% and conduct natural fermentation for 30 days to obtain improved reduced-salt pepper sauce.

[0080] After testing, the reduced-salt pepper sauce in this example can maintain the stability of the reduced-salt fermentation system, and the pH value during the entire fermentation period is between 4.2 and 4.5. This acidity condition can effectively stabilize the microbial community in the product, promote enzyme activity, and prevent the rancidity of the pepper sauce. At the same time, the amino acid nitrogen concentration reaches 0.3 g / 100 g, which can produce unique flavor substances, including isopropyl acetic acid, ethyl acetate, valeric acid with fruity aroma, sabinene with aromatic and woody odor, β-myrcene with sweet orange flavor and balsamic aroma, and p-ethylphenol with strong guaiacol odor, etc. The product has moderate acidity, balanced flavor, and uniform taste.

[0081] Example 5

[0082] The short Lactobacillus brevis strain D15 for reduced-salt Pixian bean paste in this example is the same as the strain D15 in Example 1.

[0083] The use of the short Lactobacillus brevis strain D15 in this example is in the preparation of reduced-salt pepper sauce, and the specific method is as follows:

[0084] (1) Strain activation: Lactobacillus brevis strain D15 was inoculated into MRS liquid medium and cultured at 30°C for 3 days to obtain D15 activated seed culture.

[0085] (2) Preparation of reduced-salt pepper embryo: Fresh peppers were washed, drained, de-stemmed, de-stalked, and then crushed with a crusher. Salt was added to the pepper material to achieve a salinity of 9%. The reduced-salt pepper embryo was obtained.

[0086] (3) The D15 activated seed culture was inoculated into the reduced-salt pepper embryo at a mass ratio of 2%, and natural fermentation was carried out for 28 days to obtain reduced-salt pepper paste.

[0087] Tests showed that the reduced-salt pepper paste of this example could maintain the stability of the reduced-salt fermentation system, with a pH value of 4.2-4.5 during the entire fermentation period. This acidity condition could effectively stabilize the microbial community in the product, promote enzyme activity, and prevent the rancidity of the pepper paste. The amino acid nitrogen concentration reached 0.3 g / 100 g, which could produce unique flavor substances, including isopropyl acrylate, ethyl acetate, and valeric acid, which have a fruity aroma; sabinene, which has an aromatic and woody odor; β-myrcene, which has a sweet orange flavor and balsamic aroma; and p-ethylphenol, which has a strong guaiacol odor. This resulted in a moderate sour taste, a harmonious flavor, and a uniform mouthfeel.

[0088] Example 6

[0089] The Lactobacillus brevis strain D15 used in this example for reduced-salt Pixian bean paste is the same as strain D15 in Example 1.

[0090] The use of the Lactobacillus brevis strain D15 in this example is in the preparation of reduced-salt pepper paste, and the specific method is as follows:

[0091] (1) Strain activation: Lactobacillus brevis strain D15 was inoculated into MRS liquid medium and cultured at 28°C for 3.5 days to obtain D15 activated seed culture.

[0092] (2) Preparation of reduced-salt pepper embryo: Fresh peppers were washed, drained, de-stemmed, de-stalked, and then crushed with a crusher. Salt was added to the pepper material to achieve a salinity of 8%. The reduced-salt pepper embryo was obtained.

[0093] (3) The D15 activated seed culture was inoculated into the reduced-salt pepper embryo at a mass ratio of 1.5%, and natural fermentation was carried out for 25 days to obtain reduced-salt pepper paste.

[0094] The test shows that the reduced-salt chili sauce of the example can keep the reduced-salt fermentation system stable, the pH value in the whole fermentation cycle is between 4.2 and 4.5, the acidity condition can effectively stabilize the microbial community in the product, promote enzyme activity, and avoid the rancidity of chili sauce; and the amino acid nitrogen concentration reaches 0.3 g / 100 g under the acidity condition, can produce unique flavor substances, including fruit aroma of isopropylene acetate, ethyl acetate, valeric acid, cymene with aromatic and woody odor, β-lime has sweet orange flavor and balsamic gas, p-ethyl phenol has strong guaiacol odor, etc., so that the product has moderate sour taste, coordinated flavor, and uniform taste. Thus, the flavor and quality of the reduced-salt chili sauce are significantly improved.

[0095] Examples 4-6 prove that after the strain D15 is inoculated into the reduced-salt chili embryo with a salinity as low as 8-10%, the reduced-salt chili sauce prepared has significantly improved flavor and quality:

[0096] The reduced-salt fermentation system can be kept stable, the pH value in the whole fermentation cycle is between 4.2 and 4.5, the acidity condition can effectively stabilize the microbial community in the product, promote enzyme activity, and avoid the rancidity of chili sauce. And the amino acid nitrogen concentration reaches 0.3 g / 100 g under the acidity condition, can produce unique flavor substances, including fruit aroma of isopropylene acetate, ethyl acetate, valeric acid, cymene with aromatic and woody odor, β-lime has sweet orange flavor and balsamic gas, p-ethyl phenol has strong guaiacol odor, etc., so that the product has moderate sour taste, coordinated flavor, and uniform taste. Thus, the flavor and quality of the reduced-salt chili sauce are significantly improved.

[0097] Meanwhile, the reduced-salt chili sauce inoculated with the strain D15 has accelerated acid production speed, the fermentation cycle is shortened from 40 days to 30 days, 30% of the fermentation cycle is shortened, the preparation cost is obviously reduced, and the preparation benefit is improved.

[0098] Example 7

[0099] The short Lactobacillus brevis strain D15 for reduced-salt Pixian chili bean sauce of the example is the same as the strain D15 of Example 1.

[0100] The use of the short Lactobacillus brevis strain D15 of the example is the application in the preparation of reduced-salt broad bean sauce, and the specific method is as follows:

[0101] (1) Strain activation: inoculate the short Lactobacillus brevis strain D15 into MRS liquid medium, and culture at 28℃ for 3 days to obtain D15 activated seed culture.

[0102] (2) Preparation of reduced-salt broad bean embryo: mix 10 parts by mass of shelled and soaked broad bean with 3.0 parts by mass of wheat flour and 0.12 parts by mass of Aspergillus oryzae Hujiang 3.042, and make koji at 35℃ for 48 h to obtain broad bean embryo; then add salt and water to make the salinity of the broad bean embryo 11% and the moisture content 65% to obtain reduced-salt broad bean embryo.

[0103] (3) The D15 activated seed culture liquid is inoculated into the salt-reduced broad bean embryo at a mass ratio of 1.0%, and naturally fermented for 20 days. Thus, the salt-reduced broad bean paste is obtained.

[0104] Tests show that the salt-reduced broad bean paste of this example has a pH value of 4.2-4.5 during the whole fermentation period, an amino acid nitrogen concentration of 0.3 g / 100 g, and can produce ester flavor substances such as isopropenyl acetate and ethyl acetate with fruit aroma. These substances are unique after inoculation of D15.

[0105] Example 8

[0106] The short Brevibacterium lacto D15 used in the salt-reduced Pixian broad bean paste of this example is the same as the strain D15 of Example 1.

[0107] The use of the short Brevibacterium lacto D15 of this example is in the preparation of salt-reduced broad bean paste, and the specific method is as follows:

[0108] (1) Strain activation: The short Brevibacterium lacto D15 is inoculated into MRS liquid medium and cultured at 32°C for 4 days to obtain the D15 activated seed culture liquid.

[0109] (2) Preparation of salt-reduced broad bean embryo: 10 parts by mass of shelled and soaked broad bean paste are mixed with 3.2 parts by mass of wheat flour and 0.10 parts by mass of Aspergillus oryzae Hujiang 3.042, and koji is prepared at 34°C for 45 h to obtain broad bean embryo. Salt and water are then added to the broad bean embryo to make the salt content 10% and the water content 63%. Thus, the salt-reduced broad bean embryo is obtained.

[0110] (3) The D15 activated seed culture liquid is inoculated into the salt-reduced broad bean embryo at a mass ratio of 1.5%, and naturally fermented for 18 days. Thus, the salt-reduced broad bean paste is obtained.

[0111] Tests show that the salt-reduced broad bean paste of this example has a pH value of 4.2-4.5 during the whole fermentation period, an amino acid nitrogen concentration of 0.3 g / 100 g, and can produce ester flavor substances such as isopropenyl acetate and ethyl acetate with fruit aroma. These substances are unique after inoculation of D15.

[0112] Example 9

[0113] The short Brevibacterium lacto D15 used in the salt-reduced Pixian broad bean paste of this example is the same as the strain D15 of Example 1.

[0114] The use of the short Brevibacterium lacto D15 of this example is in the preparation of salt-reduced broad bean paste, and the specific method is as follows:

[0115] (1) Strain activation: Lactobacillus brevis strain D15 was inoculated into MRS liquid medium and incubated at 30°C for 3.5 days to obtain D15 activated seed culture.

[0116] (2) Preparation of reduced-salt broad bean embryo: 10 parts by mass of shelled and soaked broad bean halves were mixed with 2.8 parts by mass of wheat flour and 0.11 parts by mass of Aspergillus oryzae HuLiang 3.042, and koji-making was performed at 36°C for 50 h to obtain broad bean embryo. Salt and water were then added to the broad bean embryo to obtain a salt concentration of 12% and a water content of 60%.

[0117] (3) The D15 activated seed culture was inoculated into the reduced-salt broad bean embryo at a mass ratio of 1.2%, and natural fermentation was performed for 25 days to obtain reduced-salt broad bean paste.

[0118] Tests showed that the pH value of the reduced-salt broad bean paste of this example was between 4.2 and 4.5, the concentration of amino acid nitrogen reached 0.3 g / 100 g, and ester flavor substances such as isopropenyl acetate and ethyl acetate with fruit aroma could be produced. These substances were unique after inoculation of D15.

[0119] Examples 7-9 proved that:

[0120] The reduced-salt broad bean paste prepared by inoculating the reduced-salt broad bean embryo with strain D15 had a pH of 4.2-4.5, and this acidity condition could effectively stabilize the microbial community in the product, promote enzyme activity, and prevent spoilage of the broad bean paste.

[0121] At the same time, the acidity condition of pH 4.2-4.5 maintained a moderate sour taste, coordinated flavor, and uniform mouthfeel in the product, and the content of organic acids such as citric acid, malic acid, succinic acid, lactic acid, and pyroglutamic acid was effectively increased. Among them, the total amino acid content was significantly higher than that of commonly used high-salt fermented broad bean paste, and the production of terpenes and alcohols (benzene ethanol) was also significantly increased. The production of main flavor free amino acids such as aspartic acid (Asp) and glutamic acid (Glu) was also increased. A large number of taste-producing dipeptide substances (Prolyl-Glutamate, Glycyl-Valine, Gly-Norleucine) were also produced. In particular, unique flavor substances such as esters (e.g., isopropenyl acetate and ethyl acetate) were produced. In the control experiment under the same process conditions except for inoculation of strain D15, no flavor substances such as esters (e.g., isopropenyl acetate and ethyl acetate) were detected. Thus, the flavor and quality of the broad bean paste were significantly improved.

[0122] After inoculation of strain D15, the acid production rate was accelerated, and the fermentation period was shortened to 18-25 days, a 25% reduction in the fermentation period. The production efficiency of reduced-salt broad bean paste was significantly improved, and the economic benefits were improved.

Claims

1. A growth-promoting Lactobacillus breve strain D15 for reducing salt in Pixian fermented bean paste, the registration number of which is CGMCC No: 27698.

2. A use of the Lactobacillus breve growth-promoting strain D15 according to claim 1 in the preparation of salt-reduced Pixian bean paste.

3. The use of the Lactobacillus breve growth-promoting strain D15 according to claim 2, characterized in that The specific method of the application in the preparation of salt-reduced Pixian bean paste is as follows: (1) Strain activation: inoculate the Lactobacillus breve strain D15 into MRS liquid culture medium and culture at 28-32°C for 3-4 days to obtain D15 activated strain culture medium; (2) Inoculation and fermentation: Add the D15 activated bacterial culture solution at a mass ratio of 1-2% to the zero-salt fermented pepper embryos, an intermediate raw material in the preparation of reduced-salt Pixian bean paste, and then ferment at 30-37°C for 48-72 hours to obtain optimized zero-salt fermented pepper embryos; (3) Optimization of reduced-salt Pixian bean paste: In the preparation step of the reduced-salt Pixian bean paste, which requires the use of zero-salt fermented pepper embryos, the optimized zero-salt fermented pepper embryos are used to replace the zero-salt fermented pepper embryos, thereby obtaining the optimized reduced-salt Pixian bean paste.

4. A use of the Lactobacillus breve growth-promoting strain D15 according to claim 1 in the preparation of reduced-salt chili sauce.

5. The use of the Lactobacillus breve growth-promoting strain D15 according to claim 4, characterized in that The application of the reduced-salt chili sauce in the preparation process is as follows: (1) Strain activation: inoculate the Lactobacillus breve strain D15 into MRS liquid culture medium and culture at 28-32°C for 3-4 days to obtain D15 activated strain culture medium; (2) Preparation of reduced-salt pepper embryos: fresh peppers are washed, drained, and the stems and peduncles are removed, and then crushed with a crusher to obtain pepper material; salt is added to the pepper material to make the salinity reach 8-10%, thereby obtaining reduced-salt pepper embryos; (3) The D15 activated bacterial culture solution is inoculated onto the salt-reduced pepper embryos at a mass ratio of 1-2%, and natural fermentation is carried out for 25-30 days to obtain the improved salt-reduced pepper sauce.

6. A use of the Lactobacillus breve growth-promoting strain D15 according to claim 1 in the preparation of reduced-salt broad bean paste.

7. The use of the Lactobacillus breve growth-promoting strain D15 according to claim 6, characterized in that The specific method of applying the method in the preparation of reduced-salt broad bean paste is as follows: (1) Strain activation: inoculate the Lactobacillus breve strain D15 into MRS liquid culture medium and culture at 28°C-32°C for 3-4 days to obtain D15 activated strain culture medium; (2) Preparation of reduced-salt broad bean embryos: 10 parts by weight of shelled and soaked broad bean petals are mixed with 2.8-3.2 parts by weight of wheat flour and 0.1-0.12 parts by weight of Shanghai-brewed 3.042% Aspergillus oryzae, and koji is made at 34-36°C for 45-50 hours to obtain broad bean embryos; salt and water are then added to adjust the salinity of the broad bean embryos to 10-12% and the moisture to 60-65% to obtain reduced-salt broad bean embryos; (3) Inoculate the D15 activated bacterial culture solution into the salt-reduced broad bean embryos at a mass ratio of 1.0-1.5%, and allow to ferment naturally for 18-25 days to obtain the salt-reduced broad bean paste.

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