Phytobacterium plantarum and application thereof, and preparation method of sauced radish

Abstract

The invention provides plant lactobacillus, application thereof and a preparation method of sauced radishes, and relates to the technical field of biology. The lactobacillus plantarum LH01 provided by the invention is preserved in the China General Microbiological Culture Collection Center (CGMCC), and the preservation number of the lactobacillus plantarum LH01 is CGMCC No.36033. The lactobacillus plantarum LH01 provided by the invention has the advantages that the lactobacillus plantarum LH01 is preserved in the China General Microbiological Culture Collection Center (CGMCC); the strain is derived from a pickled vegetable pickling solution, and the research of the inventor finds that the strain has good acid resistance, cholate resistance and antibacterial performance, can effectively improve the crispness, chewiness and elasticity of the sauced radishes when being used for preparing the sauced radishes, and is outstanding in sensory quality.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, and in particular to a plant lactobacillus and its application, and a method for preparing pickled radishes. Background Technology

[0002] Pickled vegetables refer to various vegetable products made primarily from fresh vegetables through pickling or marinating. This process involves varying degrees of microbial fermentation and boasts a long history and rich cultural significance. Currently, the main production process for vegetable fermentation is traditional natural fermentation. Natural fermentation involves multiple microorganisms, including lactic acid bacteria naturally present in the vegetables, which are crucial for the sensory quality, safety, and nutritional value of fermented vegetables. The presence of yeast and fungi on the surface of fresh vegetables makes the natural fermentation process complex and variable, resulting in unstable quality. Furthermore, traditional fermentation methods are heavily influenced by the natural environment, leading to excessively long fermentation cycles, low efficiency, high levels of unwanted microorganisms, and susceptibility to quality and safety issues. Inoculation fermentation, on the other hand, can significantly shorten fermentation time. It allows the fermentation environment to quickly become low-acid, inhibiting the growth of unwanted microorganisms, reducing the risk of fermentation failure, and decreasing the formation of harmful substances (nitrites, bacterial toxins, and fungal toxins, etc.), thus improving product quality and achieving products with consistent sensory characteristics and nutritional value.

[0003] With the development of molecular biology techniques, researchers both domestically and internationally have conducted in-depth studies on the microbial communities and flavor components in pickled vegetables. These studies have revealed that the dominant microorganisms in pickled vegetables have various beneficial effects on the human body and are significantly correlated with the formation of flavor compounds. On one hand, the microorganisms in pickled vegetables utilize nutrients in vegetable juices to metabolize and produce organic acids, free amino acids, glycols, esters, alcohols, and sulfur-containing compounds, giving pickled vegetable products their unique aroma and flavor. On the other hand, the microorganisms in pickled vegetables, especially lactic acid bacteria, have been identified as a rich source of probiotics, which can promote human health. Furthermore, lactic acid bacteria play a positive role in the fermentation process of pickled vegetables, improving their flavor, enhancing their nutritional value, degrading nitrites, and extending their shelf life.

[0004] Inoculation fermentation is of great significance for the safety and standardization of fermented vegetable processes. However, current research on inoculation fermentation mainly focuses on pickled vegetables, while there is little or no research on wet (such as pickled mustard greens), semi-dry (such as pickled cabbage), dry salted vegetables, as well as pickled vegetables in soy sauce.

[0005] In view of this, the present invention is hereby proposed. Summary of the Invention

[0006] The primary objective of this invention is to provide a plant lactobacillus.Lactiplantibacillus plantarum LH01.

[0007] A second objective of this invention is to provide a microbial agent.

[0008] A third objective of this invention is to provide the aforementioned *Lactobacillus plantarum*. Lactiplantibacillus plantarum Application of LH01 or microbial agents in food preparation.

[0009] The fourth objective of this invention is to provide a method for preparing pickled radish in soy sauce, so as to solve the above-mentioned technical problems.

[0010] To achieve the above objectives, the following technical solution is adopted: In a first aspect, the present invention provides a plant lactobacillus. Lactiplantibacillus plantarum LH01, the taxonomic name of this strain is: *Lactobacillus plantarum*, and its Latin scientific name is: Lactiplantibacillus plantarum It is deposited at the China General Microbiological Culture Collection Center (CGMCC), at No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, on September 23, 2025, with accession number CGMCC No. 36033.

[0011] Secondly, the present invention provides a microbial agent, the microbial agent comprising the aforementioned *Lactobacillus plantarum*. Lactiplantibacillus plantarum LH01.

[0012] Thirdly, the present invention provides the above-mentioned *Lactobacillus plantarum*. Lactiplantibacillus plantarum Application of LH01 or microbial agents in food preparation.

[0013] As a further technical solution, the food includes pickled radish.

[0014] Fourthly, the present invention provides a method for preparing pickled radish, comprising the following steps: Mix the radish with the soy sauce, then inoculate with the aforementioned Lactobacillus plantarum. Lactiplantibacillus plantarum LH01 was fermented to prepare pickled radish.

[0015] As a further technical solution, the radish is a radish that has undergone salt dehydration treatment.

[0016] As a further technical solution, the sauce is a mixture of sweet bean sauce and water; The ratio of sweet bean sauce to water is 1 g: 9 mL.

[0017] As a further technical solution, the inoculum size for fermentation is 1×10⁻⁶.4 -1×10 7 CFU / mL.

[0018] As a further technical solution, the fermentation material-to-liquid ratio is 1g:(1-2.5 mL).

[0019] As a further technical solution, the fermentation time is 3-6 days.

[0020] Compared with the prior art, the present invention has the following beneficial effects: The LH01 strain of *Lactobacillus plantarum* provided by this invention is derived from the pickling liquid of pickled vegetables. The inventors have found that this strain has good acid resistance, bile salt resistance and antibacterial properties. When used to prepare pickled radishes, it can effectively improve the crispness, chewiness and elasticity of pickled radishes, resulting in outstanding sensory quality. Attached Figure Description

[0021] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0022] Figure 1 : Horizontal community bar graph of cucumber strips and kohlrabi bacteria; Figure 2 : Horizontal community bar graph of bacterial species in cucumber strips and kohlrabi; Figure 3 : Agarose gel electrophoresis image of PCR amplification products of lactic acid bacteria 16S rDNA (1 is LH01, 2 is LH05, 3 is LH09, 4 is LH11, 5 is LS02, 6 is LS06). Figure 4 Results of acid resistance test of lactic acid bacteria; Figure 5 Results of bile salt tolerance test of lactic acid bacteria; Figure 6 pH changes in fermented radish sauce made with lactic acid bacteria; Figure 7 Changes in acidity of pickled radishes fermented with lactic acid bacteria; Figure 8 Results of electronic tongue analysis of fermented pickled radish with lactic acid bacteria (A3-001 is Lactobacillus plantarum Zhang-LL; B3-001 is Lactobacillus plantarum LH01; C3-001 is Lactobacillus paracasei KL1; D3-001 is CK). Figure 9 Sensory evaluation results of lactic acid bacteria fermented pickled radish; Figure 10 Results of single-factor optimization of inoculum quantity for fermented lactic acid bacteria pickled radish; Figure 11 Results of single-factor optimization of the ratio of fermented sauce to pickled radish in lactic acid bacteria fermentation sauce; Figure 12 Results of single-factor optimization of fermentation time for pickled radish in lactic acid bacteria fermented sauce; Figure 13 A graph showing the interaction between inoculum size and liquid-to-material ratio on sensory scores; Figure 14 A graph showing the interaction between the liquid-to-material ratio and fermentation time in affecting sensory scores; Figure 15 A graph showing the interaction between inoculum size and fermentation time on sensory scores. Detailed Implementation

[0023] The embodiments and examples of the present invention will be described in detail below. However, those skilled in the art will understand that the following embodiments and examples are for illustrative purposes only and should not be considered as limiting the scope of the present invention. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the scope of protection of the present invention. Unless otherwise specified, conventional conditions or conditions recommended by the manufacturer shall apply. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products.

[0024] In a first aspect, the present invention provides a plant lactobacillus. Lactiplantibacillus plantarum LH01, the taxonomic name of this strain is: *Lactobacillus plantarum*, and its Latin scientific name is: Lactiplantibacillus plantarum It is deposited at the China General Microbiological Culture Collection Center (CGMCC), at No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, on September 23, 2025, with accession number CGMCC No. 36033.

[0025] The LH01 strain of *Lactobacillus plantarum* provided by this invention is derived from the pickling liquid of pickled vegetables. The inventors have found that this strain has good acid resistance, bile salt resistance and antibacterial properties. When used to prepare pickled radishes, it can effectively improve the crispness, chewiness and elasticity of pickled radishes, resulting in outstanding sensory quality.

[0026] Secondly, the present invention provides a microbial agent, the microbial agent comprising the aforementioned *Lactobacillus plantarum*. Lactiplantibacillus plantarum LH01 may also include other strains or excipients, and this bacterial agent has all the beneficial effects of the *Lactobacillus plantarum* of the present invention.

[0027] Thirdly, the present invention provides the above-mentioned *Lactobacillus plantarum*. Lactiplantibacillus plantarum Application of LH01 or microbial agents in food preparation.

[0028] In some alternative embodiments, the food includes, but is not limited to, pickled radishes, or other Lactobacillus plantarum fermented foods known to those skilled in the art.

[0029] Fourthly, the present invention provides a method for preparing pickled radish, comprising the following steps: Mix the radish with the soy sauce, then inoculate with the aforementioned Lactobacillus plantarum. Lactiplantibacillus plantarum LH01 was fermented to prepare pickled radish.

[0030] This fermentation method is simple and efficient, and the fermented pickled radishes have good crispness, chewiness, and elasticity, with outstanding sensory quality.

[0031] In some alternative embodiments, the radish is a radish that has undergone salt dehydration treatment.

[0032] In some alternative embodiments, the salt dehydration treatment is 6% salt dehydration for 4 hours.

[0033] In some alternative embodiments, the sauce is a mixture of sweet bean sauce and water; The ratio of sweet bean sauce to water is 1 g: 9 mL.

[0034] In some alternative embodiments, the radish and soy sauce mixture also includes the addition of 3% NaCl.

[0035] In some optional embodiments, the inoculum size for fermentation is 1 × 10⁻⁶. 4 -1×10 7 CFU / mL, preferably 1×10⁻⁶ 6 CFU / mL.

[0036] In some optional embodiments, the fermentation liquid-to-material ratio is 1 g:(1-2.5 mL), preferably 1 g:2 mL.

[0037] In some alternative implementations, the fermentation time is 3-6 days, preferably 4 days.

[0038] The present invention will be further illustrated below with specific embodiments. However, it should be understood that these embodiments are merely for the purpose of more detailed illustration and should not be construed as limiting the present invention in any way.

[0039] Example 1 1. Experimental reagents, bacterial strains and instruments 1.1 Experimental strains Table 1. Main strains and culture conditions

[0040] 1.2 Experimental Reagents Table 2. Main experimental reagents and their manufacturers

[0041] 1.3 Experimental Instruments and Equipment Table 3. Main Experimental Instruments and Manufacturers

[0042] 2 Experimental Methods 2.1 Analysis of Microbial Diversity in Pickled Vegetables Pickling liquid from Liubiju pickled vegetables (cucumber strips and kohlrabi) was collected separately, and microbial genomic DNA was extracted using a column extraction microbial genome extraction kit (EE401-01, Beijing TransGen Biotech Co., Ltd.). The DNA samples were then amplified by PCR and sequenced using the Illumina platform. Bacterial DNA was compared using the Silva 123 / 16S rRNA database.

[0043] 2.2 Isolation and culture of lactic acid bacteria strains Samples of the pickling liquid from pickled vegetables were collected, and lactic acid bacteria were isolated by streak plating on MRS solid medium using a serial dilution method and incubated at 37°C for 72 h. Single colonies were picked and inoculated into MRS liquid medium and cultured with shaking at 37°C for 24 h for subsequent genomic DNA extraction.

[0044] 2.3 Extraction of genomic DNA from lactic acid bacteria Take 1 mL of lactic acid bacteria culture into a sterile centrifuge tube, centrifuge at 5000 rpm for 5 minutes, discard the supernatant, and collect the bacterial pellet. Extract genomic DNA from the lactic acid bacteria using a column extraction microbial genome extraction kit (EE401-01, Beijing TransGen Biotech Co., Ltd.).

[0045] 2.4 Identification of Lactic Acid Bacteria 2.4.1 PCR amplification of 16S rDNA from lactic acid bacteria Using extracted lactic acid bacteria genomic DNA as a template, PCR amplification was performed using universal primers 27F and 1492R for bacterial 16S rDNA sequences. A 25 μL PCR amplification system was prepared, with the following added sequentially: 12.5 μL of 2×taq DNA polymerase buffer, 1 μL of primer 27F, 1 μL of primer 1492R, 1 μL of lactic acid bacteria genomic DNA, and finally 9.5 μL of ddH2O. PCR amplification conditions were: 95°C pre-denaturation for 5 min; cycling parameters: 95°C denaturation for 30 s, 55°C annealing for 30 s, 72°C extension for 1 min, for a total of 35 cycles; after the 72°C extension for 5 min, the product was stored at 4°C. After PCR, the PCR products were sent to Beijing Qingke Biotechnology Co., Ltd. for DNA sequencing, and BLAST analysis was performed based on the sequencing results.

[0046] 2.42 Physiological and Biochemical Identification Single colonies from MRS plates were streaked onto BUA+B plates (containing 5% sterile defibrinated sheep blood) and incubated at 37°C. After obtaining single colonies, the turbidity of the bacterial suspension was adjusted to 65±2% T using Biolog's dedicated inoculation solution. The suspension was then inoculated into AN micro-identification plates and anaerobically incubated at 35–37°C for 24–48 hours. The identification results were then read on a Biolog fully automated microbial identification instrument.

[0047] 2.5 Evaluation of lactic acid bacteria tolerance 2.5.1 Acid Resistance Evaluation Six selected lactic acid bacteria strains were cultured in MRS liquid medium at 37°C for 16 h. The cells were collected by centrifugation, washed with sterile physiological saline, and resuspended in simulated gastric fluid (pH 3.0) for 2 h at 37°C. After incubation, the bacterial suspension was appropriately diluted, and the survival rate was calculated before and after treatment using the plate count method (CFU / mL) (survival rate = (number of viable bacteria after incubation / initial number of viable bacteria) × 100%).

[0048] 2.5.2 Evaluation of bile salt tolerance MRS media containing 0.3% and 0.5% bile salts were prepared, with a bile-free MRS medium used as a control. Lactic acid bacteria were inoculated into the media at a concentration of 5% (v / v) and incubated at 37°C for 48 hours. After incubation, a suitable amount of the bacterial suspension was serially diluted. Different dilutions were plated onto bile-free MRS plates, with 3-5 parallel plates for each dilution. Results were expressed as viable bacteria count (CFU / mL) using the plate count method.

[0049] 2.6 Analysis of the antibacterial properties of lactic acid bacteria The Oxford cup method was used to analyze the antibacterial properties of the fermentation broth against some foodborne pathogens and some acid bacteria for the six selected lactic acid bacteria and the laboratory-preserved Lactobacillus plantarum Zhang-LL, Lactobacillus plantarum Lp2, and Lactobacillus paracasei KL1.

[0050] Using sterile forceps, place the Oxford cup vertically on the culture medium surface. Mix a 1% indicator bacterial suspension with beef extract peptone agar medium and pour the mixture into the plate. After the medium has completely solidified, remove the Oxford cup. Under aseptic conditions, pipette 100 μL of the bacterial supernatant into each well and incubate at 37°C for 24–48 h. Repeat each group three times. Measure the diameter (d) of the inhibition zone using the cross-hatching method. The antibacterial performance of the isolated strain is determined by comparing the diameters of the inhibition zones.

[0051] 2.7 Process flow and key points of lactic acid bacteria fermented pickled radish sauce Cut fresh radishes into chunks → dehydrate with 6% salt for 4 hours → wash → air dry → pack into jars → add soy sauce → inoculate with bacteria → seal and ferment at 25℃.

[0052] Take Liubiju sweet bean sauce, add sterile water at a ratio of 1:9 (g / mL) and mix well. Add 3% NaCl and ferment fresh radish with Lactobacillus plantarum Zhang-LL, Lactobacillus plantarum LH01 and Lactobacillus paracasei KL1 respectively. Use the sample without added bacteria as the control group. Ferment at 25℃ for 3 days. Detect the pH, acidity, texture, taste and sensory quality of the pickled radish. Compare and analyze the effects of different lactic acid bacteria fermentation on the quality of pickled radish.

[0053] 2.8 Evaluation Methods for the Physicochemical Properties of Pickled Vegetables 2.8.1 pH Measurement The pH value of the sample brine was measured using a pH meter.

[0054] 2.8.2 Determination of total acidity Total acidity was determined according to GB 12456-2021, "National Food Safety Standard - Determination of Total Acidity in Food".

[0055] 2.8.3 Determination of texture Cut the fermented pickled radishes into 4 cm lengths, keeping the shape and size as uniform as possible. The crispness, elasticity, and chewiness of the pickled vegetables were measured using an LFRA-100 texture analyzer (Brookfield Laboratories, USA).

[0056] 2.8.4 Electronic tongue Take 40 g of fermented pickled radish paste made from different strains, mash it into a 150 mL beaker, and add 80 mL of deionized water. After mixing, pour 60 mL of the sample solution into a special measuring cup for the electronic tongue to be tested, and use taste analysis to convert the data into taste values ​​for analysis.

[0057] 2.9 Sensory Quality Evaluation Methods After the radishes fermented and matured, 10 food professionals were invited to conduct a sensory evaluation of the samples. A weighted scoring method was used to rigorously evaluate the color, aroma, texture, and taste of the samples, and the results were recorded according to the scoring criteria.

[0058] Table 4 Sensory Evaluation Table for Pickled Radishes

[0059] 2.10 Single-factor experiment on fermentation of pickled radish with lactic acid bacteria LH01 Take Liubiju sweet bean sauce, add sterile water at a ratio of 1:9 (g / mL) and mix well. Add 3% NaCl and ferment fresh radish with Lactobacillus plantarum Zhang-LL, Lactobacillus plantarum LH01 and Lactobacillus paracasei KL1 respectively. Use the sample without added bacteria as the control group. Ferment at 25℃.

[0060] 2.10.1 Single-factor optimization of inoculum quantity The inoculation amount was set to 1×10 4 CFU / mL, 1×10 5 CFU / mL, 1×10 6 CFU / mL, 1×10 7 The inoculum concentration was CFU / mL, the material-to-liquid ratio was 1:2, the fermentation temperature was 25℃, and the fermentation time was 3 days. After pickling and fermentation, the samples were taken out and subjected to sensory evaluation. Based on the sensory evaluation results, the optimal level of inoculum concentration was determined.

[0061] 2.10.2 Single-factor optimization of feed-liquid ratio The material-to-liquid ratios were set to 1:1, 1:1.5, 1:2, and 1:2.5, respectively, with an inoculum quantity of 1×10⁻⁶. 6 The optimal range of material-liquid ratio was determined by comparing the results of sensory evaluation after fermentation, using CFU / mL, fermentation temperature of 25℃, fermentation time of 3 days, and the results of sensory evaluation.

[0062] 2.10.3 Single-factor optimization of fermentation time Fermentation times were set to 3, 4, 5, and 6 days, with an inoculum size of 1×10⁻⁶. 6 The optimal fermentation time was determined by comparing the results of sensory evaluation after fermentation, using CFU / mL, a material-to-liquid ratio of 1:2, a fermentation temperature of 25℃, and the results of the sensory evaluation.

[0063] 2.11 Response Surface Experiment Based on the influence of single factors on the sensory evaluation of the fermentation process of pickled radish, three single factors—inoculum quantity, liquid-to-material ratio, and fermentation time—were selected to further optimize the conditions using response surface methodology. The design factors and levels of the response surface experiment are shown in the table below.

[0064] Table 5 Box-Behnken Design Factors and Levels

[0065] 3 Results and Analysis 3.1 Results of bacterial diversity analysis in pickled vegetables Samples of pickled vegetables (cucumber strips and kohlrabi) from Liubiju pickled vegetables were sent to Shanghai Meiji Company for microbial diversity sequencing. Results showed that bacterial diversity was at the genus level ( Figure 1 Bacterial diversity was significantly higher in cucumber strips (HGT) than in kohlrabi (SK). The dominant genera in cucumber strips included *Lactobacillus*, *Pseudomonas*, *Halomonas*, and *Chromohalobacter*, while endemic genera included *Cobelia* and *Pediococcus*. In kohlrabi, the main dominant genera included *Sphingomonas*, *Enterobacter*, and *Staphylococcus*, while endemic genera included *Aureimonas*, *Alkali bacillus*, and *Methylobacterium-Methylo*.

[0066] At the seed level ( Figure 2 The dominant bacterial species in cucumber strips (HGT) include Lactobacillus plantarum, Weissella dibaria, and Chromohalobacter canadensis; the main dominant bacteria in kohlrabi include Halomonas jeotgali, Staphylococcus sciuri, and Alkalibacillus salliacus.

[0067] 3.2 Isolation and culture of microbial strains in naturally pickled vegetables 3.2.1 PCR identification of strain-type lactic acid bacteria Lactic acid bacteria were isolated from Liubiju pickled cucumber strips using MRS medium. Six strains of lactic acid bacteria were preliminarily identified through morphological identification. Genomic DNA was extracted from these six strains, and 16S rDNA PCR amplification was performed using the genomic DNA as a template. PCR products of approximately 1500 bp in length were successfully obtained, and electrophoresis showed the products to be a single bright band (see...). Figure 3 ).

[0068] Sequence alignment (Table 6) revealed that four strains of *Lactobacillus plantarum* constituted the largest proportion among the six strains and can be considered the dominant species. Additionally, two strains of *Lactobacillus fermentum* were identified through screening.

[0069] Table 6 shows the sequence alignment and identification results of some strains.

[0070] 3.2.2 Physiological and biochemical identification The physiological and biochemical characteristics of the selected lactic acid bacteria LH01 were analyzed using the Biolog fully automated microbial identification system. The results are shown in Table 7. Lactic acid bacteria LH01 can utilize various monosaccharides, oligosaccharides and other carbon sources, and the identification result is Lactobacillus plantarum.

[0071] Table 7. Physiological and biochemical identification results of Lactobacillus plantarum LH01

[0072] Note: "+" indicates that more than 95% of the strains are positive; "-" indicates that more than 95% of the strains are negative.

[0073] 3.3 Acid Resistance Evaluation The acid resistance results of the six strains are as follows: Figure 4 As shown, Lactobacillus plantarum LH01 and Lactobacillus plantarum LS02 maintained a high proportion of viable bacteria, with a survival rate of over 95%, which was significantly higher than (p<0.05) the other four lactic acid bacteria strains, indicating that these two strains have good acid resistance (pH 3.0).

[0074] 3.4 Evaluation of bile salt tolerance In the determination of bile salt tolerance of lactic acid bacteria ( Figure 5 All six strains showed certain bile salt tolerance characteristics. Among them, Lactobacillus plantarum LH01 had a survival rate of 79% in a medium containing 0.5% ox bile salt, which was significantly higher than (p<0.05) the other five lactic acid bacteria strains.

[0075] 3.5 Analysis of the antibacterial properties of lactic acid bacteria Six lactic acid bacteria strains screened and identified in this study, and three lactic acid bacteria strains (Lactobacillus plantarum Zhang-LL, Lactobacillus plantarum Lp2, and Lactobacillus paracasei KL1) preserved in the laboratory, were used to detect the antibacterial properties of the lactic acid bacteria fermentation broth against some foodborne pathogens and some acid bacteria using the Oxford cup method. The results are shown in Table 7. Among them, Lactobacillus plantarum Zhang-LL, Lactobacillus plantarum Lp2, Lactobacillus plantarum LH01, and Lactobacillus paracasei KL1 showed certain antibacterial effects against Escherichia coli, Enterococcus faecalis, Listeria monocytogenes, and Salmonella. Meanwhile, Lactobacillus plantarum LH01 also showed antibacterial effects against some Lactobacillus plantarum, Lactobacillus paracasei, and other lactic acid bacteria. Considering the results of tolerance evaluation and antibacterial characteristic analysis, Lactobacillus plantarum LH01 will be used as the fermentation strain for subsequent research on the fermentation and quality analysis of pickled radishes.

[0076] Table 7 Antibacterial spectrum of lactic acid bacteria

[0077] Inhibition zone diameter (mm): +++, 15–21 mm; ++, 9–14 mm; +, 1–8 mm; –, noinhibition.

[0078] 3.6 Quality Analysis of Lactic Acid Bacteria Fermented Pickled Radish 3.6.1 Comparative Analysis of pH Changes like Figure 6 During the 3-day fermentation process of the three lactic acid bacteria-fermented pickled radish and naturally fermented pickled radish, the overall pH trend was first decreasing and then increasing, reaching the lowest pH point on the second day of fermentation. Among them, Lactobacillus plantarum LH01 reached the lowest pH value of 3.18 on the second day of fermentation, which was significantly lower than the other two lactic acid bacteria and the CK group, indicating that Lactobacillus plantarum LH01 had the strongest acid-producing ability.

[0079] 3.6.2 Comparative Analysis of Acidity Changes like Figure 7 During the 3-day fermentation process, the overall acidity of the three lactic acid bacteria-fermented pickled radish strains and the naturally fermented pickled radish showed a trend of first increasing and then decreasing. The acidity of Lactobacillus plantarum LH01 reached its highest value (close to 0.6g / 100g) on ​​the second day of fermentation, which was significantly higher than that of the other two lactic acid bacteria strains and the CK group, indicating that Lactobacillus plantarum LH01 had the strongest acid-producing capacity.

[0080] 3.6.3 Comparative Analysis of Texture Changes As shown in Table 8, measurements using a texture analyzer revealed that the crispness of *Lactobacillus plantarum* LH01 was 45.33 ± 3.16 g, significantly higher than that of *Lactobacillus plantarum* Zhang-LL (37.91 ± 3.16 g), *Lactobacillus paracasei* KL1 (37.76 ± 3.16 g), and CK (25.93 ± 3.16 g), indicating that *Lactobacillus plantarum* LH01 was the most effective in maintaining crispness. The chewiness of LH01 was 2.12 ± 0.16 mj, significantly higher than that of KL1 (1.75 ± 0.12 mj), Zhang-LL (0.68 ± 0.06 mj), and CK (0.81 ± 0.07 mj), indicating that the fermented product of *Lactobacillus plantarum* LH01 had the strongest chewiness. The elasticity of LH01 was 3.96±0.29 mm, significantly higher than that of Zhang-LL (2.71±0.22 mm), but not significantly different from that of KL1 (3.25±0.24 mm) and CK (3.24±0.31 mm). This indicates that the elasticity of LH01 is superior to that of Zhang-LL and comparable to that of KL1 and CK. In summary, LH01 of Lactobacillus plantarum showed outstanding performance in improving the crispness, chewiness, and elasticity of pickled radish, especially in crispness and chewiness, which were significantly better than other samples. This suggests that LH01 of Lactobacillus plantarum is the most effective in improving the texture characteristics of pickled radish.

[0081] Table 8. Texture changes of pickled radishes fermented with lactic acid bacteria

[0082] Note: In the same column, a, b, and c represent significant differences (p<0.05). 3.6.4 Comparative Analysis of Electronic Tongues like Figure 8 The four groups of experimental samples shown exhibit a certain distribution across various taste dimensions. Compared to other groups, Lactobacillus plantarum LH01 has a higher sourness intensity. Due to its low pH value and strong acid production capacity, it indicates that the pickled radish fermented by this strain has a more prominent sour taste.

[0083] 3.7 Comparative Analysis of Sensory Evaluation After sensory evaluation of the samples by 10 food professionals, the results were as follows: Figure 9 As shown in the figure, *Lactobacillus plantarum* LH01 scored 94.85 in sensory evaluation, which was not significantly different from *Lactobacillus plantarum* Zhang-LL (91.92), but slightly higher; it was significantly higher than *Lactobacillus paracasei* KL1 (84.13) and the CK group (71.38). In conclusion, *Lactobacillus plantarum* LH01 is the most recognized and effective in improving the sensory quality of pickled radishes.

[0084] 3.8 Single-factor optimization experiment of fermented sauce and pickled radish made with lactic acid bacteria LH01 3.8.1 Single-factor optimization of inoculum quantity The inoculation amount was set to 1×10 4 CFU / mL, 1×10 5 CFU / mL, 1×10 6 CFU / mL, 1×10 7 The inoculum concentration was CFU / mL, the material-to-liquid ratio was 1:2, the fermentation temperature was 25℃, and the fermentation time was 3 days. After pickling and fermentation, the samples were removed and subjected to sensory evaluation. Ten food professionals conducted the sensory evaluation, and the optimal inoculum level was determined based on the sensory evaluation results. The results are as follows: Figure 10 As shown, when the inoculation dose is 1×10 6 At CFU / mL, the sensory evaluation score reached the highest at 96.4, significantly higher than other inoculum amounts (p<0.05). Therefore, the optimal substrate-to-liquid ratio for single-factor optimization of inoculum amount for pickled radishes is 1×10⁻⁶. 6 CFU / mL.

[0085] 3.8.2 Single-factor optimization of feed-liquid ratio The material-to-liquid ratios were set to 1:1, 1:1.5, 1:2, and 1:2.5, respectively, with an inoculum quantity of 1×10⁻⁶. 6 The fermentation concentration was CFU / mL, the fermentation temperature was 25℃, and the fermentation time was 3 days. Sensory evaluation results after fermentation are as follows: Figure 11 When the material-to-liquid ratio is 1:2, the sensory evaluation score reaches the highest of 96.7, which is significantly higher (p<0.05) than the sensory evaluation scores of other inoculation amounts. Therefore, the optimal material-to-liquid ratio for pickled radish is 1:2.

[0086] 3.8.3 Single-factor optimization of fermentation time Fermentation times were set to 3, 4, 5, and 6 days, with an inoculum size of 1×10⁻⁶. 6 The concentration of CFU / mL was 1:2 (solid-to-liquid ratio), and the fermentation temperature was 25℃. Sensory evaluation results after fermentation are as follows: Figure 12 When the fermentation time was 4 days, the sensory evaluation score reached the highest of 96.8, which was significantly higher (p<0.05) than the sensory evaluation scores of other inoculum amounts. Therefore, the optimal material-liquid ratio for the single-factor optimization of fermentation time for pickled radishes was 4 days.

[0087] 3.9 Response Surface Optimization Experiment of Lactic Acid Bacteria LH01 Fermented Pickled Radish Sauce Based on the single-factor model, the inoculation amount was designed to be 1×10⁻⁶ according to the Box-Behnken procedure. 5 CFU / mL, 1×10 6 CFU / mL, 1×10 7 The sensory evaluation results of the three-factor, three-level experiment with CFU / mL, material-to-liquid ratio of 1:1.5, 1:2, 1:2.5, and fermentation time of 3 days, 4 days, and 5 days are shown in Table 9.

[0088] Table 9 Response Surface Experimental Design Scheme and Results

[0089] Further methodological analysis, based on the model analysis and variance analysis results of the fermentation process conditions for pickled radishes (Table 10), showed that the regression model was highly significant (p<0.01), indicating a good fit to the response values. The p-value for the lack-of-fit term was 0.7403, indicating that the lack-of-fit term was not significant. The influence of the three factors on fermented radishes was A > C > B. This demonstrates that the regression equation has small experimental error and the lack-of-fit term is not significant. Therefore, this regression model can be used to accurately analyze and predict the fermentation process parameters of Lactobacillus plantarum LH01 fermented pickled radishes.

[0090] Table 10. Analysis of variance for regression models using sensory scores as response variables.

[0091] Response surface methodology can intuitively reflect the impact of changes in fermentation conditions on the sensory quality of fermented radishes, such as... Figures 13-15 This is a principal response surface plot and contour plot showing the pairwise relationships between inoculum size, liquid-to-material ratio, fermentation time, and sensory score. The 3D response surface plot all point downwards, indicating that the sensory evaluation score has a maximum value. As shown in the figure, AB ( Figure 13 The surface slopes are all convex and the contour lines are elliptical, indicating a strong interaction between factors, BC ( Figure 14 ) and AC ( Figure 15 Although the surface is relatively steep, the contour ellipses are not very obvious, and their interaction is weak. Further analysis using this model yielded the optimal process conditions: inoculum quantity of 1×10⁻⁶. 6.15 With a CFU / mL concentration, a solid-liquid ratio of 1:2.15, and a fermentation time of 3.86 days, the optimal process combination for ease of implementation was determined to be an inoculum quantity of 1×10⁻⁶. 6 With a CFU / mL concentration, a material-to-liquid ratio of 1:2, and a fermentation time of 4 days, the sensory evaluation value of fermented radishes produced using this process can reach 95.2.

[0092] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A type of *Lactobacillus plantarum* Lactiplantibacillus plantarum LH01, characterized in that, The plant lactobacillus Lactiplantibacillus plantarum LH01 was deposited with China General Microbiological Culture Collection Center, and the deposit number is CGMCC No. 36033.

2. A bacterial agent, characterized in that, The bacterial agent comprises the Lactobacillus plantarum of claim 1 Lactiplantibacillus plantarum LH01.

3. The Lactobacillus plantarum of claim 1 Lactiplantibacillus plantarum Use of the LH01 or the bacterial agent of claim 2 in the manufacture of a food product.

4. Use according to claim 3, characterized in that, The food includes pickled radish.

5. A method of preparing chutneyed radish characterized in that, Including the following steps: Mixing radish with brine, and then inoculating the plant lactobacillus of claim 1 Lactiplantibacillus plantarum LH01, after fermentation, to prepare pickled radish.

6. The production method according to claim 5, wherein The radish is a radish treated by salt dehydration.

7. The preparation method according to claim 5, characterized in that, The sauce water is a mixed solution of sweet bean sauce and water. The ratio of sweet bean sauce to water is 1 g:9 mL.

8. The production method according to claim 5, characterized by, The fermentation was carried out at a 1 x 10 4 -1 x 10 7 CFU / mL.

9. The production method according to claim 5, characterized by, The ratio of the fermented material to liquid is 1 g:(1-2.5 mL).

10. The method of claim 5, wherein, The fermentation time is 3-6 days.

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