Lactobacillus plantarum CMRC 1L and application thereof

Abstract

The invention belongs to the technical field of food fermentation, and provides a lactobacillus plantarum CMRC 1L and application thereof. The lactobacillus plantarum CMRC 1L disclosed by the invention is separated from a sour meat sample in Maojia village region in Lijiang City, Yunnan Province, grows well on an MRS culture medium, is high in acid production speed, has the biological characteristics of salt resistance, frostbite resistance, inhibition of growth and reproduction of spoilage microorganisms and the like, has a good fermentation effect on various plant proteins, and is an excellent strain for preparing a leavening agent for production. The invention also provides a leavening agent containing the strain and application of the leavening agent in vegetable protein meat products. Compared with the vegetable protein jerky without the leavening agent, the vegetable protein jerky product prepared by the preparation method provided by the invention has the advantages of stable quality, fresh and sweet fermentation fragrance, no beany flavor and good flavor and taste. The strain CMRC 1L has great potential when being applied to production of vegetable protein meat products.

Description

Technical Field

[0001] The invention belongs to the technical field of food fermentation, and in particular relates to a strain of Lactobacillus plantarum CMRC 1L and an application thereof. Background Art

[0002] Plant protein is a protein with a wider source and more environmentally friendly than animal protein, and is an indispensable part of people's daily diet. With the rapid development of human society, it is estimated that in 2050, the global population will reach 9.8 billion, and the demand for food will continue to grow. In recent years, due to the increasing awareness of low-carbon environmental protection and animal welfare, people's demand for animal protein substitutes has increased, so the market position of plant protein has continued to improve. Plant protein meat is mainly made of plant proteins such as soy protein, corn protein, wheat protein, etc., and supplemented with auxiliary materials. It is a food with nutritional value, taste and flavor similar to meat products through synthesis and processing. Its raw material sources are rich, and a variety of plant proteins provide more possibilities for the production of plant protein meat, such as soy protein, chickpea protein, pea protein in bean protein; corn protein in cereal protein, rapeseed protein in oil protein, etc. can all be used to make plant protein meat. Among them, soy protein is low in price, easy to obtain, and has high nutritional value. It shows excellent food processing properties such as gelling and water retention, and is recognized as a good choice for making plant protein meat. However, soy protein-related plant protein meat products often have a distinct "beany smell" that is difficult to be accepted by consumers. The product texture and flavor quality are poor, and the product form is single. In addition, due to the compact structure of soy protein and the anti-nutritional factors present in beans, the digestibility of soy protein is relatively weak, which does not meet consumers' pursuit of nutritious and healthy food. In recent years, how to improve the flavor quality of plant protein meat products and enhance their nutritional value has received widespread attention.

[0003] As a traditional food processing method, microbial fermentation is a mild and energy-saving processing technology. It does not require complex equipment and processes, and is simple, economical and convenient. Fermented foods have certain advantages in flavor and nutritional value. my country has a long history of fermenting soy protein, such as fermented black beans and fermented bean curd. Bacteria, molds, and yeasts can all be used as fermentation strains for soy protein. Among them, bacteria and molds are the main microorganisms in fermented black beans obtained by natural fermentation. Rhizopus and Mucor are used to make fermented bean curd, while salt-tolerant lactic acid bacteria and yeasts are in a dominant position in the production process of soy sauce. At present, there are relatively few studies on fermentation strains suitable for plant protein meat products with flavor improvement effects, especially the application of plant lactobacillus in the production of plant protein meat-related products to improve product flavor, texture and other characteristics. Summary of the invention

[0004] The invention aims to provide a strain of Lactobacillus plantarum CMRC 1L and application thereof.

[0005] In order to achieve the purpose of the present invention, in the first aspect, the present invention provides a strain of Lactobacillus plantarum CMRC 1L isolated and purified from a farmer's homemade sour meat sample, which is classified and named Lactobacillus plantarum Lactobacillus plantarum , which has been deposited in the General Microbiology Center of China Microorganism Culture Collection Committee, Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Postal Code 100101, with the deposit number CGMCC No. 32440 and the deposit date October 31, 2024.

[0006] In a second aspect, the present invention provides a microbial preparation containing the Lactobacillus plantarum.

[0007] Furthermore, the content of Lactobacillus plantarum in the microbial preparation is 10 9 ~10 11 CFU / g.

[0008] The preparation method of the microbial preparation comprises: fermenting and culturing the plant lactobacillus, centrifuging the obtained fermentation liquid, collecting bacterial precipitates, resuspending the bacterial bodies with sterile physiological saline, centrifuging and discarding the supernatant, adding a small amount of physiological saline again to mix well to obtain concentrated bacterial liquid, adding an appropriate amount of freeze-drying protective agent and freeze-drying to obtain the concentrated bacterial liquid.

[0009] Furthermore, the mass ratio of the concentrated bacterial liquid to the lyophilization protective agent is (0.1-10):1.

[0010] The lyoprotectant may be selected from at least one of skim milk, glycerol, sucrose and the like.

[0011] In a third aspect, the present invention provides the use of the Lactobacillus plantarum or the microbial preparation in food fermentation.

[0012] In a fourth aspect, the present invention provides a method for preparing a plant protein meat product, comprising the following steps: 1) Soaking and untying plant texture protein: Soak the plant texture protein in water for 1-2 hours to fully rehydrate it and then drain the water; 2) Raw material mixing and flavoring: Mix the vegetable protein powder, vegetable textured protein, seasoning and water evenly according to the proportion; 3) Inoculating the starter: adding the microbial preparation according to the ratio; 4) Fermentation: Place in a constant temperature and humidity chamber for fermentation; 5) Molding and drying: according to production needs, the fermented raw materials are squeezed into corresponding shapes; 6) Baking: put it in the oven for baking; 7) Cooling and packaging: Cut according to production needs and package after cooling to room temperature.

[0013] Furthermore, the plant textured protein can be selected from soybean textured protein and / or pea textured protein, etc.

[0014] Furthermore, the plant protein powder can be selected from at least one of soy protein isolate powder, soy protein powder, wheat protein powder, rice protein powder, pea protein powder, etc.; Furthermore, the seasoning can be selected from at least one of plant-derived seasonings such as salt, sugar, brewed soy sauce, five-spice powder, yeast extract, etc.

[0015] Preferably, the ingredients in step 2) are as follows, by weight: 50-80 parts of soy textured protein, 20-50 parts of soy protein isolate powder, 20-50 parts of gluten powder, 2-20 parts of white sugar, 2-6 parts of salt, 2-10 parts of brewed soy sauce, 1-3 parts of spice powder or five-spice powder, 1-5 parts of yeast extract and 100-150 parts of water.

[0016] Preferably, the inoculation amount of Lactobacillus plantarum in step 3) is 10 4 CFU / g raw material, the microbial preparation is dissolved in advance with 2% glucose solution, wherein the amount of glucose solution is 5-10% of the total weight of the raw material.

[0017] Preferably, the fermentation conditions in step 4) are: temperature 20-25° C., 12-36 hours; Preferably, in step 5), the fermented raw material blanks are squeezed into a tray in the shape of meat strips, and placed in a smoking oven to dry the moisture, and the drying conditions are: temperature 50-55° C., 1-2 hours; Preferably, the baking conditions in step 6) are: temperature 120-150° C., 10-20 min.

[0018] In a fifth aspect, the present invention provides a plant protein meat product prepared according to the method.

[0019] By means of the above technical solution, the present invention has at least the following advantages and beneficial effects: (I) Lactobacillus plantarum provided by the present invention ( Lactobacillus plantarum )CMRC 1L, this strain was isolated from the sour meat sample in Maojia Village, Lijiang City, Yunnan Province. It grows well on MRS medium, produces acid quickly, has biological characteristics such as salt resistance, frostbite resistance, and inhibition of the growth and reproduction of corruptive microorganisms. It is an excellent strain for making starter for production.

[0020] (ii) The plant lactobacillus CMRC 1L provided by the present invention has a good fermentation effect on various plant proteins such as soy protein, peanut protein, pea protein, etc., can grow and reproduce with plant protein as a substrate, and produce a specific fermentation flavor during the fermentation process. It can effectively improve the unpleasant odors such as "beany smell" in plant proteins such as soy protein isolate and wheat protein, improve the quality of plant protein meat products, promote protein degradation and flavor formation, and improve the nutritional value of the product.

[0021] The product produced by the method for preparing artificially fermented plant protein jerky provided by the present invention has good elasticity and chewiness, significantly improved flavor, improved overall acceptance, and good market prospects. The application of Lactobacillus plantarum CMRC 1L in the production of plant protein meat products has great potential. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 It is the colony morphology of Lactobacillus plantarum CMRC 1L in a specific embodiment of the present invention.

[0023] Figure 2 It is the strain morphology of Lactobacillus plantarum CMRC 1L in a specific embodiment of the present invention.

[0024] Figure 3 It is the growth curve of Lactobacillus plantarum CMRC 1L strain in a specific embodiment of the present invention.

[0025] Figure 4 It is the acid production curve of Lactobacillus plantarum CMRC 1L strain in a specific embodiment of the present invention.

[0026] Figure 5 The figure shows the effect of different pH values ​​on the growth of Lactobacillus plantarum CMRC 1L strain in a specific embodiment of the present invention.

[0027] Figure 6 The figure shows the effect of different ethanol concentrations on the growth of CMRC 1L strain and CICC 21792 strain in a specific embodiment of the present invention.

[0028] Figure 7 The figure shows the effect of different sodium chloride concentrations on the growth of CMRC 1L strain and CICC 21792 strain in a specific embodiment of the present invention.

[0029] Figure 8 The figure shows the effect of different glucose concentrations on the growth of CMRC 1L strain and CICC 21792 strain in the specific examples of the present invention.

[0030] Figure 9-13 The following are radar charts for sensory evaluation of fermented products of soybean protein, rice protein, peanut protein, wheat protein and pea protein in specific embodiments of the present invention. DETAILED DESCRIPTION

[0031] The present invention isolates and selects a plant lactobacillus strain suitable for production operation and capable of effectively improving the flavor and other qualities of plant proteins such as soy protein, and based on this, establishes a processing method for plant protein jerky, which is of great significance for developing nutritious and delicious plant protein meat products.

[0032] The present invention adopts the following technical solution: The inventors collected a variety of traditional homemade sour meat samples from Maojia Village, Lijiang City, Yunnan Province, Jialiang Town, Libo County, Guizhou Province and other areas. Through separation, screening, purification and further biological and fermentation characteristics tests, a plant lactobacillus suitable for use as a starter for plant protein meat products was obtained and sent to the General Microbiology Center of the China Microbiological Culture Collection Committee. The strain collection number is CGMCC No. 32440. The strain was isolated from the Maojia sour meat sample.

[0033] The invention also provides a method for separating, screening and identifying the plant lactobacillus strain.

[0034] The collected samples were enriched and cultured. After 10-fold series gradient dilution, the sample solution of appropriate concentration was aspirated and inoculated into MRS medium (containing 6% NaCl by mass) by pouring plate method. The samples were mixed and cultured at 37°C for 48 h after solidification. Single colonies were picked according to colony morphology, size, transparency, etc. and repeatedly streaked and purified until a single strain was obtained. Single colonies in the plate were picked and inoculated into MRS broth and cultured at 37°C and 200 r / min for 24 h. The culture solution was dripped onto a precision test paper of pH 5.4-7.0, and the strains corresponding to the yellow color of the test paper (pH < 5.4) were selected for Gram staining to observe the bacterial morphology, and the strains were preserved.

[0035] The fermentation performance of the plant lactobacillus obtained above was initially screened, including acid production characteristics experiment, glucose fermentation gas production experiment, viscosity production experiment, H production experiment, 2 S test, Oxford cup method antibacterial test (Staphylococcus aureus and Escherichia coli). Select bacteria with strong acid production ability, no stickiness, no gas production during glucose fermentation, and no H production. 2 S, and the strains with obvious inhibitory effect on spoilage bacteria were rescreened. The strains obtained from the initial screening were inoculated into a liquid culture medium with soy protein isolate powder and glucose, and cultured at 37°C for 24 hours, and then the strains were screened by olfactory method. Based on the results of the initial screening and rescreening, plant lactobacillus CMRC 1L with good growth, strong fermentation ability, and the ability to ferment with soy protein isolate as a substrate to produce characteristic flavors was comprehensively selected for subsequent identification and application research.

[0036] The colony morphology of the plant lactobacillus provided by the present invention is: it grows well on solid MRS medium, the colonies are white, round or oval, convex, with smooth edges and surfaces, and the colony diameter is about 1 to 2 mm. Bacterial morphology: Gram staining positive, short rod-shaped, without spores and flagella. The colony morphology of plant lactobacillus CMRC 1L is shown in Figure 1 , strain morphology see Figure 2 .

[0037] Molecular biological identification of strain CMRC 1L: Bacterial genomic DNA was extracted using a bacterial genome extraction kit, and the quality of DNA extraction and fragment size were detected by 0.8% agarose gel electrophoresis. The universal primers 27F and 1492R were used to amplify the 16S rRNA sequence using genomic DNA as a template. The PCR product was analyzed by gel electrophoresis and sent to Yingwei Jieji Trading Co., Ltd. (Beijing) for sequencing. The sequencing results were compared with the sequences in the GenBank database by Blast homology. The results showed that the strain sequence was consistent with the strain Lactobacillus plantarum The MLG-17 gene sequence was highly homologous and was identified as Lactobacillus plantarum ( Lactobacillus plantarum ), the 16S rRNA gene sequence of the strain is shown in SEQ ID NO:1.

[0038] The invention also provides a plant protein meat product starter containing the Lactobacillus plantarum CMRC 1L.

[0039] After the bacteria preserved in the glycerol tube are transferred and activated twice, they are inoculated into MRS broth culture medium at an inoculation ratio of 2-4%, and cultured at 37°C with shaking at 100-150 r / min for 18-24 hours to obtain a bacterial suspension. The upper culture liquid is discarded after centrifugation at 3000-5000 rpm for 10-15 min, and the bacteria are resuspended in sterile saline and centrifuged to discard the supernatant. A small amount of saline is added again to mix well to obtain a concentrated bacterial solution, and a certain volume of lyophilization protective agent is added and freeze-dried to obtain a starter.

[0040] Optionally, the content of Lactobacillus plantarum in the starter is 10 9 ~10 11 CFU / g.

[0041] Optionally, the mass ratio of the concentrated bacterial solution to the lyophilization protective agent is (0.1-10):1.

[0042] Optionally, the volume of physiological saline added for the second time is 1 / 5 to 1 / 10 of the volume of the initial bacterial suspension, so that the bacterial concentration in the final concentrated bacterial solution reaches 10 10 CFU / mL.

[0043] Optionally, the lyoprotectant composition may include skim milk, glycerol and sucrose.

[0044] Optionally, the culture conditions for inoculation in MRS broth medium are pH 6.0-6.4 and culture temperature 30-37°C.

[0045] The present invention also provides application of the Lactobacillus plantarum CMRC 1L or a microbial agent containing the Lactobacillus plantarum in producing plant protein meat products.

[0046] The present invention further provides a method for preparing artificially fermented plant protein jerky containing the plant lactobacillus strain, comprising the following steps: 1) Soaking and stripping plant tissue protein: Soak the plant tissue protein in water for 1 to 2 hours in advance, fully rehydrate it, and then drain the water.

[0047] 2) Raw material mixing and flavoring: Mix the plant protein powder, plant tissue protein, seasoning and water evenly according to the ratio.

[0048] 3) Inoculate the starter: Add the starter bacteria according to the ratio.

[0049] 4) Fermentation: Place in a constant temperature and humidity chamber for fermentation.

[0050] 5) Molding and drying: According to production needs, the fermented raw materials are squeezed into corresponding shapes.

[0051] 6) Bake: Place in oven for baking.

[0052] 7) Cooling and packaging: It can be cut and cooled to room temperature before packaging according to needs.

[0053] Optionally, in step 1), the plant textured protein can be selected from one or more of soybean textured protein, pea textured protein, etc., and can be subjected to a detangling process according to the shape of the textured protein.

[0054] Optionally, in step 2), the plant protein powder can be selected from one or more of soy protein isolate powder, soy protein powder, wheat protein powder, rice protein powder, pea protein powder, etc. The seasoning can be selected from at least one of plant-derived seasonings such as salt, sugar, brewed soy sauce, five-spice powder, yeast extract, etc.

[0055] Preferably, the ingredients in step 2) are as follows: 50-80 parts of soy textured protein, 20-50 parts of soy protein isolate powder, 20-50 parts of gluten powder, 2-20 parts of white sugar, 2-6 parts of salt, 2-10 parts of brewed soy sauce, 1-3 parts of spice powder (or five-spice powder), 1-5 parts of yeast extract and 100-150 parts of water. The parts are by weight.

[0056] Preferably, in step 3), the inoculation amount of Lactobacillus plantarum is 10 4CFU / g raw material, the starter is dissolved in advance with 2% glucose solution, wherein the amount of glucose solution is 5-10% of the total weight of the raw material.

[0057] Preferably, in step 4), the fermentation conditions are 20-25° C. for 12-36 hours. To ensure uniform distribution of the starter in the raw materials, intermittent stirring may be performed during the fermentation.

[0058] Optionally, in step 5), the fermented raw material blanks are squeezed into a tray in the shape of meat strips, and placed in a smoking oven to dry out moisture. The drying stage conditions are a temperature of 50-55° C. for 1-2 hours.

[0059] Preferably, in step 6), the baking conditions are a temperature of 120-150° C. for 10-20 min.

[0060] The following examples are used to illustrate the present invention, but are not intended to limit the scope of the present invention. Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art, and the raw materials used are all commercially available products.

[0061] The unit "portion" expressing weight in the present invention can be gram, kilogram, kilo, jin, liang, ton etc., depending on the production scale.

[0062] Example 1 Screening by olfactory method The fermentation performance of the plant lactobacillus obtained above was initially screened, including acid production characteristics experiment, glucose fermentation gas production experiment, viscosity production experiment, H production experiment, 2 S test, Oxford cup method antibacterial test (Staphylococcus aureus and Escherichia coli). Select bacteria with strong acid production ability, no stickiness, no gas production during glucose fermentation, and no H production. 2 S, and the strains with obvious inhibitory effect on spoilage bacteria were rescreened. The strains obtained in the initial screening were inoculated into liquid culture medium supplemented with soy protein isolate powder and glucose. After culturing at 37℃ for 24h, the fermentation effect of lactic acid bacteria on soy protein was analyzed by olfactory aroma. The results were expressed in terms of the intensity of fermentation aroma and beany flavor. The fermentation aroma flavor characteristics included sweet aroma, fruity aroma, fat aroma and cream aroma. The intensity was expressed by "-" (no aroma), "+" (slight aroma), "++" (relatively fragrant) and "+++" (strong aroma); the intensity of beany flavor was expressed by "-" (none), "+" (light beany flavor) and "++" (strong beany flavor). The blank liquid culture medium without any starter inoculated was used as the negative control, and the fermentation liquid culture medium inoculated with Lactobacillus plantarum CICC 21792 was used as the positive control.

[0063] The results showed that the flavors of fermented products of different lactic acid bacteria strains were significantly different. Some strains could not adapt well to the fermentation of soy protein, and may not convert the beany compounds in soy protein into other flavor components, resulting in only a weak fermentation aroma or no fermentation aroma. The fermentation broth of Lactobacillus plantarum CMRC 1L had a sweet, fruity and creamy aroma, a strong fermentation aroma, and a significantly weakened beany smell. As shown in Table 1, compared with the control without inoculation of the fermentation agent, Lactobacillus plantarum CMRC 1L effectively improved the overall flavor characteristics of the sample, indicating that this strain plays an important role in improving the sensory quality of plant protein meat products with soy protein as the main raw material. Lactobacillus plantarum CICC 21792 has a normal sweet fermentation aroma, but also has a stimulating and sour smell, a weaker sweet aroma, and a more prominent fragrance.

[0064] Table 1

[0065] Example 2 Lactobacillus plantarum ( Lactobacillus plantarum )Biological characteristics of CGMCC No. 32440 1. Growth curve determination The Lactobacillus plantarum ( Lactobacillus plantarum ) CGMCC No. 32440 was inoculated into MRS nutrient broth for activation twice, and then transferred to fresh MRS broth medium at a 2% inoculum. The culture was incubated at 37°C and 200 r / min for 48 h. The turbidity (OD600) of the bacterial suspension was measured by sampling every 2 h, and 3 replicates were performed at each time point. Figure 3 It is the growth curve of the Lactobacillus plantarum CMRC 1L strain of the present invention.

[0066] 2. Determination of acid production curve The strain culture and regular sampling were the same as above. The pH value of the bacterial suspension was measured with a pH meter, and 3 replicates were performed at each time point. Figure 4 It is the acid production curve of the Lactobacillus plantarum CMRC 1L strain of the present invention.

[0067] Example 3 A fermented vegetable protein jerky product and its processing method 1. Soak and shred soy protein: Soak 500g of soy protein in water for 1 hour, rehydrate it and drain it.

[0068] 2. Mix and flavor the raw materials: Mix 600g of soy textured protein, 400g of soy protein isolate powder, 400g of gluten powder, 100g of white sugar, 40g of salt, 40g of brewed soy sauce, 20g of five-spice powder, 20g of yeast extract and 1200g of water according to the ratio.

[0069] 3. Inoculation of starter: Accurately weigh 0.005g of starter powder, dissolve it in 200g of 2% glucose solution, and add it to the raw materials and stir evenly to make the plant lactobacillus ( Lactobacillus plantarum )The addition amount of CGMCC No. 32440 is 10 4 CFU / g raw material.

[0070] 4. Fermentation: Place in a constant temperature and humidity chamber for fermentation at 20°C for 16 hours. To ensure uniform distribution of the starter in the raw materials, perform intermittent stirring during fermentation.

[0071] 5. Forming and drying: squeeze the fermented raw materials into meat strips and put them into a smoke oven to dry out the moisture. The drying stage conditions are 55°C for 1 hour.

[0072] 6. Baking: Put it in the oven for baking at a temperature of 130°C for 10 minutes.

[0073] 7. Cooling packaging: Pack after cooling to room temperature.

[0074] Comparative Example 1 A fermented vegetable protein jerky product and its processing method Same as Example 3, except that: 3. Inoculation of starter: Accurately weigh 0.005g of starter powder, dissolve it in 200g of 2% glucose solution, and add it to the raw material and stir evenly, so that the amount of Lactobacillus plantarum CICC 21792 added to the raw material is 10 4 CFU / g raw material.

[0075] Comparative Example 2 A plant protein jerky product and its processing method Same as Example 3, except that: No starter culture was inoculated.

[0076] Experimental Example 1 Sensory Evaluation A sensory evaluation panel consisting of 15 experienced professionals aged between 20 and 40 was selected to evaluate the color, smell, taste and mouthfeel of the plant protein jerky products through blind scoring. The total score for each item was 25 points. The acceptance was the sum of the scores of the above four items. The mouth was rinsed with water between the evaluations of each sample. The sensory evaluation standards are shown in Table 2, and the sensory evaluation results are shown in Table 3.

[0077] Table 2 Sensory evaluation standards

[0078] Table 3 Sensory evaluation results

[0079] As can be seen from Table 3, the products made in Example 3 and Comparative Example 1 are significantly higher than those in Comparative Example 2 in terms of smell, flavor, mouthfeel and overall acceptance, which are specifically manifested as having a sweet fermented aroma without a beany smell; the taste is palatable and the aftertaste is mellow. The flavor of Comparative Example 2 is relatively simple, the taste is average, and there is a significant beany smell. The plant protein jerky product made in Example 3 is superior to that in Comparative Example 1 in terms of smell and taste, and is manifested as a richer aroma without unpleasant odors such as irritation and acridity. There is no significant difference in color between Example 3 and Comparative Examples 1-2. It can be seen that the present invention uses Lactobacillus plantarum CMRC 1L to prepare fermented plant protein jerky, which can significantly improve the flavor characteristics of the product, enhance the taste and quality of plant protein meat products, and make it easier for consumers to accept.

[0080] Experimental Example 2: Environmental tolerance test of Lactobacillus plantarum The pH tolerance, ethanol tolerance, glucose tolerance and sodium chloride tolerance of the strains were determined respectively: the strains were transferred and activated twice and then inoculated with 2% inoculum in MRS liquid medium containing different ethanol volume fractions (0%, 3%, 6%, 9%, 12%, 15%), pH values ​​(2, 3, 4, 5, 6, 7, 8, 9, 10), glucose mass concentrations (100, 200, 300, 400, 500, 600, 700 g / L) and sodium chloride mass concentrations (0, 20, 40, 60, 80, 100 g / L). The culture was carried out at 36°C, 120 r / min for 1 day, and the OD value was measured at 600 nm using an enzyme marker ( Figure 5 , Figure 6 Figure 7 and Figure 8 ).

[0081] like Figure 5 As shown in the figure, the OD values ​​at different pH values ​​increased first and then decreased. The growth of the strain was inhibited by an overly acidic or alkaline environment. The OD value of Lactobacillus plantarum CMRC 1L was close to 0 at pH 3 and pH 9, and the highest OD value was measured at pH 5. The OD value of CICC 21792 strain was close to 0 at pH 4 and pH 9, and the highest OD value was measured at pH 6.

[0082] like Figure 6 As shown in the figure, as the amount of ethanol added increases, the OD value gradually decreases, indicating that the growth and reproduction ability of the strain gradually decreases. The higher the ethanol concentration, the more obvious the inhibitory effect on its growth, and it may even stop growing. The maximum ethanol tolerance concentration of the CMRC 1L strain is 12%, and the maximum ethanol tolerance concentration of the CICC 21792 strain is 9%.

[0083] like Figure 7As shown, when the mass concentration of sodium chloride is less than 40g / L, it has basically no effect on the growth and reproduction of the two strains of bacteria. When the mass concentration of sodium chloride is greater than 40g / L, the growth and reproduction ability of the strains gradually decreases. When the mass concentration of sodium chloride increases to 80g / L, both strains of bacteria basically stop growing.

[0084] like Figure 8 As shown in the figure, with the increase of initial glucose mass concentration, the growth ability of the strains gradually decreased; in 600 g / L glucose solution, the two strains were basically unable to grow; in 500 g / L glucose solution, the CMRC 1L strain was able to grow slowly, with an OD600nm of 0.994, while the OD600nm of the CICC 21792 strain was 0.688 under this condition, indicating that the CMRC 1L strain had good tolerance to glucose.

[0085] Experimental Example 3 Growth and reproduction of Lactobacillus plantarum using different plant proteins (1) Preparation of plant protein culture medium: Add 15% soy protein powder and 2% glucose to 100 mL of water, shake and mix, sterilize at 105°C for 10 min, and use it as soy protein liquid culture medium. Similarly, prepare rice protein, wheat protein, pea protein, and peanut protein liquid culture medium.

[0086] (2) Plant protein liquid fermentation method: After activation, the lactic acid bacteria are inoculated into the above culture medium at a 2% inoculation rate. At the same time, the uninoculated plant protein liquid culture medium is used as a control. The culture is carried out at 36°C, 120 r / min, and the culture period is 24-48 h.

[0087] (3) Live lactic acid bacteria counts of different plant protein fermentations The plate counting method was used to count lactic acid bacteria. The plant protein fermentation products with different dilution gradients were spread on MRS agar medium and counted after being cultured at 36°C for 48 hours.

[0088] According to the plate count method, the fermentation products of the two lactic acid bacteria cultured in different plant protein media for 24 h were gradiently diluted and coated with mRS medium. The results are shown in Table 4. The two lactic acid bacteria grew poorly in rice protein and had good utilization of other plant proteins. By comparing the number of live bacteria in the fermentation products, it was found that the fermentation effect of CMRC 1L strain on different plant proteins was better than that of CICC 21792 strain, that is, the number of live bacteria was higher.

[0089] Table 4 Viable bacteria counts of different plant proteins

[0090] Experimental Example 4 Sensory evaluation of the fermentation effect of Lactobacillus plantarum on different plant proteins Sensory evaluation by olfactory method: The sensory evaluation panel consisted of 20 experienced tasters aged 24 to 40 years old, and the fermented samples were randomly presented in the sensory test room. Determine the aroma characteristics of plant protein: The sensory evaluation panel developed and determined the aroma characteristics descriptors by directly smelling and comparing with plant protein fermentation products. The final descriptors were: nutty, green, greasy, sour, sweet, creamy, winey and overall acceptance. The flavors were scored and divided into ten grades from weak to strong (Table 5).

[0091] Table 5 Sensory intensity scoring criteria for fermented products

[0092] The sensory evaluation of different protein fermentations and controls was carried out by olfactory method. Figure 9~Figure 13 As shown (soy protein fermentation corresponds to Fig. 9 , rice protein fermentation corresponds to Fig.10 , Peanut protein fermentation corresponds to Fig.11 , wheat protein fermentation corresponds to Fig.12 , pea protein fermentation corresponds to Fig.13 ). In the rice protein fermentation experiment, the flavor profiles of the lactic acid bacteria fermentation group and the control group were similar, with no significant difference. In other plant protein fermentation experiments, the green and astringent taste of the beans in the unfermented control group was more obvious, while the nutty and oily tastes were more prominent, and the overall flavor was weaker. After lactic acid bacteria fermentation, the green and astringent taste, nutty taste, and oily taste were all weakened to varying degrees, or even disappeared. Compared with the control, the sour taste characteristics in the fermentation group were more easily detected, and the sweet and creamy aromas of the fermentation increased, and some fermented products had a little wine aroma. The acceptance of different plant proteins has increased. Among them, peanut protein has the highest acceptance, followed by soy protein, and the improvement effect of soy protein is the best.

[0093] In summary, lactic acid bacteria fermentation can greatly improve the flavor of soy protein, wheat protein, pea protein and peanut protein, and the pleasant flavor characteristics such as sweet and creamy aroma produced by CMRC 1L strain fermentation are more obvious, and the overall acceptance is better than that of CICC 21792 strain fermentation group.

[0094] Experimental Example 5 Comparison of volatile flavor substances in soybean protein fermented with Lactobacillus plantarum The olfactory method preliminarily judged that lactic acid bacteria fermentation had the most significant effect on improving the flavor quality of soy protein, and further analyzed the volatile flavor substances of soy protein fermentation. Accurately weigh 5mL of the sample to be tested and place it in a headspace sample bottle, add 5µL of 2-methyl-3-heptanone with a concentration of 0.0816μg / µL, tighten the bottle cap, insert a 50 / 30µm DVB / CAR / PDMS extraction head, place it in a 50℃ water bath for 10min, then push out the fiber head and start extraction for 30min. Then insert the extraction head into the GC-MS injection port for desorption for 10min, and start the instrument to collect data at the same time. Repeat each sample 3 times and take the average value. The culture medium without lactic acid bacteria inoculation is used as the control.

[0095] GC conditions: TG-WaxmS polar column (30m×0.25mm, 0.25µm) in splitless mode; carrier gas was high-purity helium; flow rate was maintained at 1.2mL / min; injection port temperature was 230℃. Heating program: column temperature was maintained at 40℃ for 3min; increased to 200℃ at a rate of 5℃ / min; then increased to 230℃ at a rate of 10℃ / min and maintained for 5min.

[0096] MS conditions: Electron ionization source (EI): ionization energy 70 eV, ion source temperature 300 °C; transfer line temperature 250 °C; full scan mode, mass scan range 40-400 m / z, start scan time 3 min, scan time 48 min.

[0097] Qualitative analysis: Compare the compounds in the mass spectrum with databases such as NIST and Wiley, and delete high-boiling point compounds with retention times greater than 45 minutes and no odor. The determination of the compounds is based on the positive and negative similarity index greater than 800.

[0098] Quantitative analysis: The content of each flavor substance relative to the internal standard 2-methyl-3-heptanone is calculated by the peak area and content of the internal standard. The calculation formula is as follows:

[0099] Where: C is the content of the volatile substance to be measured / (μg / kg); Ax is the peak area of ​​the volatile substance to be measured / (AU·min); C 0 is the internal standard concentration (0.0816 μg / μL); A 0 is the peak area of ​​the internal standard / (AU·min); V is the injection volume of the internal standard / μL; m is the mass of the sample to be measured / g.

[0100] The results in Table 6 show that the types and total amount of flavor substances in the fermented soybean protein inoculated with lactic acid bacteria increased significantly, specifically in the significantly increased content of acids, alcohols, aldehydes, and esters. In the fermentation groups of CMRC 1L strain and CICC 21792 strain, the acid substances were 711.40 and 652.90 μg / kg, respectively, which were significantly higher than 86.14 μg / kg in the control group. At the same time, small molecular aldehydes and esters often have low thresholds and play an important role in the formation of the overall flavor of the sample.

[0101] Comparative analysis of the two fermentation groups found that the total amount of volatile flavor substances in the CMRC 1L fermentation group was 1864.68 μg / kg, which was significantly higher than the 1593.56 μg / kg in the CICC 21792 fermentation group. The main manifestations were alcohols, aldehydes and other substances. For example, 2-n-pentylfuran with green bean and butter smell and trans-2-octenal with nutty smell were significantly higher in the CMRC 1L group than in the CICC 21792 group; for example, hexanoic acid with a sweet smell and hexanal with a tallow smell were only detected in the CMRC 1L group. Combined with sensory evaluation, it can be seen that the CMRC 1L strain can improve the flavor of plant protein products with soy protein as the main raw material, and it has great potential for application in the production of plant protein meat products.

[0102] Table 6 Comparison of volatile flavor substances in soybean protein fermented by Lactobacillus plantarum

[0103] Although the present invention has been described in detail above with general descriptions and specific embodiments, it is obvious to those skilled in the art that some modifications or improvements can be made based on the present invention. Therefore, these modifications or improvements made without departing from the spirit of the present invention all fall within the scope of protection claimed by the present invention.

Claims

1. Lactobacillus plantarum Lactobacillus plantarum CMRC 1L, the deposit number is CGMCC No. 32440.

2. A microbial preparation containing the plant lactobacillus according to claim 1; The content of plant lactobacillus in the microbial preparation is 10 9 -10 11 CFU / g.

3. The microbial preparation according to claim 2, characterized in that The preparation method of the microbial preparation comprises: fermenting and culturing the plant lactobacillus, centrifuging the obtained fermentation liquid, collecting bacterial precipitates, resuspending the bacterial bodies with sterile physiological saline, centrifuging and discarding the supernatant, adding a small amount of physiological saline again to mix well to obtain concentrated bacterial liquid, adding an appropriate amount of freeze-drying protective agent and freeze-drying to obtain the concentrated bacterial liquid.

4. The microbial preparation according to claim 3, characterized in that The mass ratio of the concentrated bacterial solution to the freeze-drying protective agent is (0.1-10):1; The lyoprotectant is selected from at least one of skim milk, glycerol and sucrose.

5. Use of the plant lactobacillus according to claim 1 or the microbial preparation according to any one of claims 2 to 4 in food fermentation.

6. A method for preparing a vegetable protein meat product, characterized in that: The following steps are involved: 1) Soaking and untying plant texture protein: Soak the plant texture protein in water for 1-2 hours to fully rehydrate it and then drain the water; 2) Raw material mixing and flavoring: Mix the vegetable protein powder, vegetable textured protein, seasoning and water evenly according to the proportion; 3) Inoculating the starter: adding the microbial preparation according to any one of claims 2 to 4 according to the ratio; 4) Fermentation: Place in a constant temperature and humidity chamber for fermentation; 5) Molding and drying: according to production needs, the fermented raw materials are squeezed into corresponding shapes; 6) Baking: put it in the oven for baking; 7) Cooling and packaging: Cut according to production needs and package after cooling to room temperature.

7. The method according to claim 6, characterized in that The plant textured protein is selected from soybean textured protein and / or pea textured protein; The plant protein powder is selected from at least one of soy protein isolate powder, soy protein powder, wheat protein powder, rice protein powder and pea protein powder; The seasoning is selected from at least one of salt, sugar, brewed soy sauce, five-spice powder and yeast extract.

8. The method according to claim 7, characterized in that In terms of weight, the ingredients of step 2) are as follows: 50-80 parts of soy textured protein, 20-50 parts of soy protein isolate powder, 20-50 parts of gluten powder, 2-20 parts of white sugar, 2-6 parts of salt, 2-10 parts of brewed soy sauce, 1-3 parts of spice powder or five-spice powder, 1-5 parts of yeast extract and 100-150 parts of water; In step 3), the inoculum amount of Lactobacillus plantarum is 10 4 CFU / g raw material, the microbial preparation is dissolved in advance with 2% glucose solution, wherein the amount of glucose solution is 5-10% of the total weight of the raw material.

9. The method according to claim 8, characterized in that Step 4) The fermentation conditions are: temperature 20-25°C, 12-36 hours; Step 5) squeeze the fermented raw material into a tray in the shape of meat strips, and put it into a smoker to dry out the moisture. The drying conditions are: temperature 50-55°C, 1-2h; The baking conditions in step 6) are: temperature 120-150° C., 10-20 min.

10. A vegetable protein meat product prepared according to the method according to any one of claims 6 to 9.

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