A strain of Lactobacillus plantarum and its application

Abstract

The invention discloses a plant lactobacillus and its application, belonging to the field of microorganisms and food. The strain is deposited in China Center for Type Culture Collection, the deposit date is April 28, 2023, the deposit number is CCTCC M 2023667, the bacteria is screened from naturally fermented sourdough, and the strain is classified and named Lactiplantibacillus plantarum G8; The plant lactobacillus of the present invention is applied to rye bran-wheat composite bread, which can significantly improve the hardness, adhesiveness, chewiness and elasticity of the rye bran-wheat composite bread, can improve the baking loss rate, and can also significantly extend the anti-mildew effect period of the bread to 12 days.

Description

Technical Field

[0001] The invention relates to the fields of microorganisms and food, and in particular to a plant lactobacillus strain and application thereof. Background Art

[0002] Rye bran is rich in ferulic acid, dietary fiber and other nutrients. As consumers pay more attention to health, whole grain bread rich in dietary fiber is sought after by consumers. Although whole grain bread is nutritious and healthy, it has a short shelf life and poor taste.

[0003] Specifically, the shelf life of naturally fermented wheat bread containing rye bran is generally no more than 9 days, which brings inconvenience to manufacturers and sellers and consumers. In addition, the texture of this type of bread containing rye bran is poor, including hardness, stickiness, chewiness and elasticity.

[0004] Therefore, adopting certain technical means to extend the shelf life of rye bran bread and improve its quality is of great significance to the entire food industry. Summary of the invention

[0005] One of the objectives of the present invention is to provide a strain of Lactobacillus plantarum to solve the above problems.

[0006] In order to achieve the above object, the technical solution adopted by the present invention is as follows: a strain of Lactobacillus plantarum, the classification of which is named Lactiplantibacillus plantarum G8, the strain was deposited in the China Center for Type Culture Collection on April 28, 2023, with the deposit number CCTCC M 2023667.

[0007] The second object of the present invention is to provide the use of the above-mentioned Lactobacillus plantarum in making bread.

[0008] As a preferred technical solution: the bread is rye bran-wheat composite bread.

[0009] As a further preferred technical solution: the method for making the rye bran-wheat composite bread is:

[0010] (1) Preparation of rye bran sourdough:

[0011] Weigh pure water and Lactiplantibacillus plantarum The G8 bacteria were mixed and then added to rye bran and placed in a constant temperature and humidity incubator at 25-35°C for fermentation for 12-36 h. The initial inoculum amount of the sourdough was 10 8 CFU / g;

[0012] (2) Preparation of rye bran sour dough bread:

[0013] Put wheat flour, water, sugar, salt, yeast and the rye bran sour dough obtained in step (1), excluding butter, into a food processor, mix well, then add butter and continue stirring until the gluten is formed and then take out; then place the bread dough at 25-35°C for primary fermentation; after the primary fermentation is completed, roll the dough into a round shape, ferment it for a secondary time at 25-35°C, and bake it in an oven with an upper / lower heat temperature of 160-180°C to obtain the bread dough.

[0014] As a further preferred technical solution: in step (2), the mass ratio of the wheat flour to the rye bran sour dough is 4:1.

[0015] Compared with the prior art, the advantages of the present invention are: the inventors of the present application screened and obtained a strain of Lactobacillus plantarum with anti-mold activity, and proved that using it as a leavening agent for bread, especially bread containing rye bran, can not only prolong the shelf life of the bread, but also improve the processing and nutritional properties of the bread; using the strain of the present invention, the anti-mold effect period of the bread containing rye bran can be extended to 12 days, which greatly improves the sales convenience of manufacturers and sellers, improves economic benefits, and is also easy for consumers to preserve and eat. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 The obtained by screening in Example 1 Lactiplantibacillus plantarum G8 Gram staining image. DETAILED DESCRIPTION

[0017] The present invention will be further described below in conjunction with the accompanying drawings.

[0018] Example 1

[0019] Strain screening:

[0020] 10 g of naturally fermented sourdough (stored at room temperature for about 5 days) from Sanya Road Vegetable Market in Ya'an City, Sichuan Province was taken and added to 90 mL of physiological saline, mixed and diluted 10 times in a gradient manner. The appropriate dilution was selected to be spread on MRS solid medium containing 0.5% calcium carbonate and cultured in a 37°C incubator for 48 h. The suspected lactic acid bacteria colonies with round, smooth, milky white, raised surfaces and calcium-dissolving rings were selected. After 5 plate streakings, purified single colonies were obtained, which were subjected to Gram staining and catalase tests, and Gram-positive and catalase-negative strains were selected and stored in 20 vol% glycerol tubes.

[0021] The preserved strain was inoculated into fresh MRS broth at a ratio of 2 vol%, and the pH of the MRS broth was measured after culturing at 37°C for 24 h, and the strain with the largest acid production was selected. The genome of the isolated strain was extracted using a DNA extraction kit, and its 16S rDNA was amplified using universal primers 27F (5'-AGAGTTTGATCMTGGCTCAG-3') and 1492R (5'-GGTTACCTTGTTACGACTT-3'). The PCR amplification product was sent to Sangon Biotech (Shanghai) Co., Ltd. for sequencing. The sequencing results were compared using the NCBI BLAST analysis tool (http: / / www.ncbi.nlm.nih.gov / blast), and the plant lactobacillus ( Lactiplantibacillus plantarum G8),

[0022] The 16S rDNA sequence is shown in SEQ ID NO: 1:

[0023]

[0024] Its Gram staining is shown in Figure 1 and deposited it in the China Center for Type Culture Collection on April 28, 2023, with the deposit number CCTCC M 2023667.

[0025] Example 2

[0026] The preparation of rye bran-wheat composite bread comprises the following steps:

[0027] (1) Preparation of rye bran sourdough:

[0028] The screened samples obtained in Example 1 Lactiplantibacillus plantarum G8 was cultured in MRS broth until the logarithmic growth phase, and the cells were collected by centrifugation at 5000 g for 10 min. The cells were washed twice with sterile saline and then collected. An appropriate amount of purified water was weighed and mixed with the cells, and then added to rye bran (the initial inoculum of sourdough was 10 8 CFU / g), DY=250, the DY value was calculated according to formula (1), and the mixture was placed in a constant temperature and humidity incubator at 30°C for 24 h (hereinafter referred to as "Example");

[0029] DY = (1)

[0030] Where: m 粉 represents the mass of flour (g), m 水 represents the mass of water (g);

[0031] Another strain of the same lactic acid bacteria from fresh sourdough at Sanya Road Vegetable Market was screened by the same screening method as in Example 1. Lactiplantibacillus plantarum G12 (medium acid-producing ability) was inoculated into sourdough made from rye bran (hereafter referred to as “control A”), and naturally fermented rye bran sourdough without inoculation with any strain served as the control group (hereafter referred to as “control B”);

[0032] (2) Preparation of rye bran sour dough bread:

[0033] Put 80 g wheat flour, 40 g water, 6 g sugar, 1 g salt, 1.5 g yeast and 20 g rye bran sourdough (except butter) into a chef machine, mix well, add 4 g butter and continue stirring until the gluten is formed and then take it out. First, let the bread dough ferment at 30℃ for 1 hour; after the first fermentation, roll the dough into a round shape and ferment at 30℃ for 1.5 hours; bake in an oven with a top / bottom temperature of 170℃ for 30 minutes.

[0034] The preparation method of rye bran sourdough bread of control A and control B is exactly the same as step (2) above, except that the sourdough added is different;

[0035] Three kinds of rye bran-wheat composite breads were obtained, which were respectively named "Example", "Control A" and "Control B" according to the corresponding sourdough used.

[0036] Example 3

[0037] Texture testing of rye bran sourdough bread

[0038] For test methods, refer to Characteristics of sourdough bread fermented with Pediococcus pentosaceus and Saccharomyces cerevisiae and its bio-preservative effect against Aspergillus flavus.

[0039] The test results are shown in Table 1.

[0040] Table 1 Hardness, stickiness, chewiness and elasticity of different breads

[0041]

[0042] As can be seen from Table 1, the Lactiplantibacillus plantarum The hardness, stickiness and chewiness of G8 rye bran sour dough bread (Example) were the smallest, and there were significant differences compared with the two control groups. Lactiplantibacillus plantarum G8 rye bran sourdough bread had the highest elasticity value. Lactiplantibacillus plantarum G8 improves the textural properties of bread, resulting in a softer crumb structure.

[0043] Example 4

[0044] Baking loss test of rye bran sourdough bread

[0045] Test method: Weigh the bread dough before baking and the bread after baking, and calculate the baking loss rate of each group of bread according to the following formula.

[0046] Baking loss rate (%) =;

[0047] The results are shown in Table 2 below:

[0048] Table 2 Baking loss rate of different breads

[0049]

[0050] As can be seen from Table 2, the Lactiplantibacillus plantarum The baking loss rate of G8 rye bran sourdough bread (Example) was the smallest, which was significantly different from the two control groups.

[0051] Example 5

[0052] Test on the mold-proof period of rye bran sourdough bread

[0053] Test method: Fresh rye bran sourdough bread samples cooled to room temperature were cut into slices with length, width and height of 100 mm, 100 mm and 15 mm respectively, placed in sterile sampling bags and stored at room temperature.

[0054] The mold colonies on the surface of each group of bread slices were monitored every day to evaluate the anti-mold effectiveness of the bread. The results are shown in Table 3.

[0055] Table 3 Anti-mold effect period of different breads

[0056]

[0057] As can be seen from Table 3, the Lactiplantibacillus plantarum The anti-mildew effect period of G8 rye bran sourdough bread (Example) was the longest, and there were significant differences compared with the two control groups. The anti-mildew effect period of the Example was extended to 12 days, which was extended by 33.3% compared with the naturally fermented bread.

[0058] Example 6

[0059] Sensory evaluation of rye bran sourdough bread

[0060] Evaluation method: The sensory evaluation panel consisted of 20 trained members. Different bread samples were placed in the same type of plastic boxes and randomly presented to the sensory evaluation panel members. Pure water was provided for mouthwash during the process. A 1-9 point hedonic scale (9: very like, 1: very dislike) was used to evaluate the color, taste, smell, texture and overall acceptability of the bread; the results are shown in Table 4;

[0061] Table 4 Sensory evaluation of different breads

[0062]

[0063] As can be seen from Table 4, the Lactiplantibacillus plantarum The sensory scores of G8 rye bran sourdough bread (Example) in terms of smell and texture were significantly different from those of the control group, with the highest scores. Lactiplantibacillus plantarum The overall acceptability of G8 rye bran sourdough bread was higher than that of control B, but slightly lower than that of control A.

[0064] Example 7

[0065] In vitro protein digestibility testing of rye bran sourdough bread

[0066] Test method:

[0067] (1) In vitro digestion

[0068] The method of Minekus et al. (A standardised static in vitro digestion methodsuitable for food-an international consensus) was used with slight modifications. The specific operation was as follows: first, 2 g of bread was thoroughly mixed with 10 mL of SSF, 0.5 mL of SSF containing 1500 U / mL α-amylase, 25 μL of 0.3 mol / L CaCl2, and 975 μL of ultrapure water, and reacted at 37°C for 2 min. 10 mL of the mixture was thoroughly mixed with 7.5 mL of SGF and 5 μL of 0.3 M CaCl2, the pH was adjusted to 3 with 1 mol / L HCl solution, and 1.6 mL of SGF containing 25000 U / mL porcine pepsin was added, and the mixture was reacted at 37°C for 2 h. After the reaction, 20 mL of gastric chyme, 11 mL of SIF and 40 μL of 0.3 mol / L CaCl2 were mixed thoroughly, the pH of the solution was adjusted to 7 with 1 mol / L NaOH solution, 5 mL of SIF containing 800 U / mL porcine trypsin and 0.4 g of bile salt (about 10 mmol / L in chyme) were added, and the reaction was carried out at 37°C for 2 h. This was repeated in triplicate. After the simulated digestion was completed, the reaction was terminated by heating at 95°C for 5 min.

[0069] (2) Protein digestibility

[0070] The gastric and intestinal phase reaction solutions were mixed with three volumes of ethanol respectively and placed at 4°C for 12 h to precipitate protein (reference reference Real meat and plant-based meat analogues have different in vitro protein digestibility properties). The samples were centrifuged at 10,000 g at 4°C for 20 min, the separated precipitate was suspended in 5 mL of protein extraction buffer, and the mixture was centrifuged at 4,000 g at 4°C for 5 min. The protein content in the supernatant was determined by the BCA protein assay kit. The protein digestibility of each group of bread was calculated according to formula (8).

[0071] Protein digestibility (%) = (8)

[0072] Where W1 is the undigested protein content (g); W0 is the protein content of the sample before digestion (g);

[0073] The results are shown in Table 5:

[0074] Table 5 In vitro protein digestibility of different breads (%)

[0075]

[0076] As can be seen from Table 5, the Lactiplantibacillus plantarum The in vitro protein digestibility of G8 rye bran sourdough bread (Example) was the highest and significantly different from both control groups.

[0077] The above is only a preferred embodiment of the present invention. It should be understood that the present invention is not limited to the form disclosed herein, and should not be regarded as excluding other embodiments, but can be used in various other combinations, modifications and environments, and can be modified within the scope of the concept described herein through the above teachings or the technology or knowledge of the relevant field. The changes and modifications made by those skilled in the art shall not deviate from the spirit and scope of the present invention, and shall be within the scope of protection of the claims attached to the present invention.

Claims

1. A strain of Lactobacillus plantarum G8, characterized in that: The bacterium was deposited in the China Center for Type Culture Collection on April 28, 2023, with the deposit number CCTCC M 2023667.

2. Application of the plant lactobacillus according to claim 1 in making bread.

3. The use according to claim 2, characterized in that: The bread is rye bran-wheat composite bread.

4. The use according to claim 3, characterized in that: The preparation method of the rye bran-wheat composite bread is as follows: (1) Preparation of rye bran sourdough: Weigh purified water and mix it with the Lactiplantibacillus plantarum G8 bacteria, then add it into rye bran and place it in a constant temperature and humidity incubator at 25-35°C for fermentation for 12-36 hours. The initial inoculum amount of the sourdough is 10 8 CFU / g; (2) Preparation of rye bran sour dough bread: Put wheat flour, water, sugar, salt, yeast and the rye bran sour dough obtained in step (1), excluding butter, into a food processor, mix them evenly, then add butter and continue stirring until the gluten is formed and then take them out; then place the bread dough at 25-35°C for primary fermentation; after the primary fermentation is completed, roll the dough into a round shape, ferment it for a secondary time at 25-35°C, and bake it in an oven with an upper / lower heat temperature of 160-180°C to obtain the bread dough.

5. The use according to claim 4, characterized in that: In step (2), the mass ratio of the wheat flour to the rye bran sour dough is 4:1.

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