A salt-containing probiotic composition and a method for preparing the same

Abstract

This invention relates to the field of microbial preparation technology, specifically to a salt-containing probiotic composition and its preparation method, comprising the following components: refined coarse salt, modified carboxymethyl chitosan, lactic acid-gluconolactone composite acidifier, sodium alginate, calcium chloride, freeze-dried Lactobacillus plantarum powder, and trehalose; the coarse salt is treated with hydrogen peroxide to remove Fe... 2+ Oxidized to Fe 3+ Citric acid chelates Fe 3+ and Cu 2+ Tannic acid displaces Fe 3+ A coordination reaction occurs, forming a metal-polyphenol network on the surface of probiotics, providing a physical barrier protection. The lactic acid-gluconolactone complex acidifier gradually decreases the pH, allowing tannic acid to systematically capture Fe. 3+ The MPNs network grows uniformly. Sodium alginate-Ca 2+ The gel network forms a composite gel network with modified carboxymethyl chitosan, and the double protection of the MPNs network significantly improves the survival rate and functional activity of probiotics.

Description

Technical Field

[0001] This invention belongs to the field of microbial preparation technology, specifically a salt-containing probiotic composition and its preparation method. Background Technology

[0002] Probiotics are live microorganisms that are beneficial to the host and have been widely used in the food, health product, and pharmaceutical fields. Copper salt, as a natural mineral salt, is rich in various trace elements and has unique medicinal and health-promoting functions. It can be used as a probiotic enhancer; the mineral elements in copper salt can promote microbial growth and enhance cell membrane stability.

[0003] However, traditional coarse salt particles are relatively large, making it difficult to form an effective synergistic effect with probiotics at the microscopic level. Furthermore, probiotic metabolism may produce hydrogen peroxide, and coarse salt contains transition metal ions (such as Fe). 2+ Cu 2+ When probiotics (such as bacteria) are in a free state, they undergo the Fenton reaction to produce reactive oxygen species, which can cause oxidative damage to the cell membrane of probiotics, leading to a sharp decline in the survival rate of probiotics. Summary of the Invention

[0004] (1) Technical problems to be solved

[0005] The purpose of this invention is to provide a salt-containing probiotic composition and its preparation method, so as to solve the problem of Fenton reaction damage to probiotic cell membranes caused by transition metal ions in cobalt salt.

[0006] (2) Technical solution

[0007] To achieve the above objectives, on the one hand, the present invention provides a salt-containing probiotic composition comprising the following components in parts by weight: 2-5 parts refined coarse salt, 8-15 parts modified carboxymethyl chitosan, 1.5-3 parts lactic acid-gluconolactone complex acidifier, 1.5-3 parts sodium alginate, 0.5-1.5 parts calcium chloride, 5-15 parts freeze-dried Lactobacillus plantarum powder, and 2-5 parts trehalose;

[0008] The refined coarse salt is treated with hydrogen peroxide to remove Fe. 2+ Oxidized to Fe 3+ Citric acid is used to bind free Fe. 3+ and Cu 2+ Finally, it is passivated with stearic acid for hydrophobic treatment;

[0009] The modified carboxymethyl chitosan was modified with tannic acid and gallic acid.

[0010] Furthermore, the preparation method of the refined coarse salt includes the following steps:

[0011] S11. Load coarse salt powder into a ball mill jar, add anhydrous ethanol and grinding balls, grind and crush to obtain coarse salt slurry;

[0012] S12. Hydrogen peroxide was added to anhydrous ethanol, and the resulting solution was slowly added dropwise to the slurry of cobalt salt. The mixture was then magnetically stirred to obtain the first compound.

[0013] S13. Dissolve citric acid in anhydrous ethanol, and slowly add the resulting citric acid solution dropwise to the first compound while stirring continuously to obtain the second compound;

[0014] S14. Stearic acid is dissolved in anhydrous ethanol, and the resulting stearic acid solution is added to the second compound. The mixture is stirred continuously, and the resulting slurry is concentrated by vacuum rotary evaporation, vacuum dried, ground and dispersed to obtain refined coarse salt.

[0015] Furthermore, the mass ratio of the coarse salt powder to citric acid is 100:5~8.

[0016] Furthermore, the method for preparing the modified carboxymethyl chitosan includes the following steps:

[0017] S21. Dissolve carboxymethyl chitosan in deionized water, adjust the pH with HCl to obtain a carboxymethyl chitosan solution;

[0018] S22. Dissolve gallic acid in deionized water, add EDC·HCl and NHS, stir to activate, and slowly add the resulting gallic acid solution to the carboxymethyl chitosan solution. Adjust the pH with NaOH, stir the reaction, and dialysis the resulting reaction solution to obtain gallic acid-carboxymethyl chitosan.

[0019] S23. Dissolve tannic acid in deionized water, adjust the pH with NaOH, add the resulting tannic acid solution to gallic acid-carboxymethyl chitosan, stir to react, add the resulting reaction solution to anhydrous ethanol to precipitate, collect the precipitate by centrifugation, wash with anhydrous ethanol, redissolve in deionized water, freeze dry to obtain modified carboxymethyl chitosan.

[0020] Furthermore, the mass ratio of carboxymethyl chitosan, gallic acid, and tannic acid is 1:0.3~0.6:0.5~1.5.

[0021] Furthermore, the preparation method of the lactic acid-gluconolactone composite acidifier includes the following steps:

[0022] S31. Dissolve sodium lactate in deionized water, add lactic acid, stir well, and adjust the pH with NaOH to obtain a lactate-sodium lactate buffer solution;

[0023] S32. Dissolve gluconolactone in deionized water, add it to lactate-sodium lactate buffer solution, and stir until homogeneous to obtain lactate-gluconolactone composite acidifier.

[0024] On the other hand, based on the same inventive concept, the present invention also provides a method for preparing a salt-containing probiotic composition, applied to the aforementioned salt-containing probiotic composition, comprising the following steps:

[0025] S1. Disperse the freeze-dried Lactobacillus plantarum powder in PBS buffer to obtain a probiotic suspension;

[0026] S2. Disperse refined coarse salt and modified carboxymethyl chitosan separately in deionized water, add them sequentially to the probiotic suspension, mix evenly, and obtain the first mixture;

[0027] S3. Slowly add the lactic acid-gluconolactone complex acidifier to the first mixture, stir evenly, and let it stand to react to obtain the second mixture;

[0028] S4. Add sodium alginate solution to the second mixture and stir until homogeneous. Add the resulting mixture dropwise to CaCl2 solution to solidify the reaction. Wash the product with deionized water, add trehalose, and freeze-dry to obtain a salt-containing probiotic composition.

[0029] In summary, due to the adoption of the above technical solution, the beneficial effects of the present invention are:

[0030] 1. In the process of preparing coarse salt, hydrogen peroxide is first used to react the highly catalytically active Fe... 2+ Oxidation to Fe with lower catalytic activity 3+ Then chelate Fe with citric acid 3+ and Cu 2+ This process fixes the free transition metal ions in the cobalt salt and passesivates it with stearic acid to prevent moisture absorption.

[0031] 2. Modified carboxymethyl chitosan grafted with tannic acid and gallic acid, the tannic acid will... 3+ Fe was displaced from citric acid. 3+ It rapidly undergoes a high-intensity coordination reaction with polyphenols (tannic acid and gallic acid), forming a dense, uniform, and stable metal-polyphenol network (MPNs) on the surface of probiotics, providing physical barrier protection.

[0032] 3. Using lactic acid-gluconolactone composite acidifiers as a slow-release acidification system, gluconolactone undergoes slow hydrolysis, gradually decreasing the pH, while citric acid-Fe... 3+ The coordination gradually dissociates, and tannic acid orderly captures Fe. 3+ MPNs networks grow uniformly.

[0033] 4. Sodium alginate-Ca 2+The gel network forms a composite gel network with modified carboxymethyl chitosan, and the double protection of the MPNs network significantly improves the survival rate of probiotics. Attached Figure Description

[0034] Figure 1 The pH change curves of the reaction systems in Example 1 and Comparative Example 5 of this invention are shown over time. Detailed Implementation

[0035] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0036] Example 1: This example discloses a salt-containing probiotic composition, comprising the following components in parts by weight: 3.5 parts refined coarse salt, 11 parts modified carboxymethyl chitosan, 2 parts lactic acid-gluconolactone composite acidifier, 2 parts sodium alginate, 1 part calcium chloride, 10 parts freeze-dried Lactobacillus plantarum powder, and 3.5 parts trehalose.

[0037] The refined coarse salt is treated with hydrogen peroxide to remove Fe. 2+ Oxidized to Fe 3+ Citric acid is used to bind free Fe. 3+ and Cu 2+ Finally, it is passivated with stearic acid for hydrophobic treatment;

[0038] The modified carboxymethyl chitosan was modified with tannic acid and gallic acid.

[0039] The method for preparing the refined coarse salt includes the following steps:

[0040] S11. Put 100g of coarse salt powder into a ball mill jar, add 200mL of anhydrous ethanol and grinding balls, grind and crush to obtain a coarse salt slurry with D50 of 200~500nm.

[0041] S12. Add 1 mL of 30% hydrogen peroxide to 50 mL of anhydrous ethanol, and slowly add the resulting solution dropwise to the slurry of cochinchinensis. Stir magnetically at room temperature for 1 h to obtain the first compound.

[0042] S13. Dissolve 5g of citric acid in 50mL of anhydrous ethanol, and slowly add the resulting citric acid solution to the first compound. Stir continuously for 2h to obtain the second compound.

[0043] S14. Dissolve 1.5g of stearic acid in 50mL of hot anhydrous ethanol, add the resulting stearic acid solution to the second compound, stir continuously for 1h, concentrate the resulting slurry under vacuum at 50℃, dry under vacuum at 50~60℃ for 12~24h, grind and disperse, and pass through a 100-mesh sieve to obtain refined coarse salt.

[0044] It should be noted that in the preparation of refined cobalt salt, the solvent for treating the cobalt salt slurry with hydrogen peroxide must be anhydrous ethanol. A low concentration of hydrogen peroxide should be added slowly dropwise while magnetically stirred at room temperature. This process is more conducive to the production of Fe. 2+ Hydroxygenated by peroxide to Fe 3+ .

[0045] The mass ratio of coarse salt powder to citric acid is 100:5.

[0046] The method for preparing the modified carboxymethyl chitosan includes the following steps:

[0047] S21. Dissolve 10g of carboxymethyl chitosan in 400mL of deionized water, and adjust the pH to 5-5.5 with HCl to obtain a carboxymethyl chitosan solution.

[0048] S22. Dissolve 3g gallic acid in 80mL deionized water, add 2.5g EDC·HCl and 1.5g NHS, stir and activate at room temperature for 30min, slowly add the resulting gallic acid solution to carboxymethyl chitosan solution, adjust the pH to 6~6.5 with NaOH, stir and react at room temperature for 12~24h, and dialysis the resulting reaction solution to obtain gallic acid-carboxymethyl chitosan;

[0049] S23. Dissolve 5g of tannic acid in deionized water, adjust the pH to 8-8.5 with NaOH, add the resulting tannic acid solution to gallic acid-carboxymethyl chitosan, stir the reaction at room temperature for 6-12 hours, adjust the pH of the resulting reaction solution to 6-6.5, add anhydrous ethanol to precipitate, collect the precipitate by centrifugation, wash with anhydrous ethanol, redissolve in deionized water, freeze dry to obtain modified carboxymethyl chitosan.

[0050] The mass ratio of carboxymethyl chitosan, gallic acid, and tannic acid is 1:0.3:0.5.

[0051] The preparation method of the lactic acid-gluconolactone composite acidifier includes the following steps:

[0052] S31. Dissolve 0.4g of sodium lactate in 20mL of deionized water, add 0.4mL of 85% lactic acid, stir well, and adjust the pH to 6~6.5 with NaOH to obtain lactate-sodium lactate buffer solution;

[0053] S32. Dissolve 1.8g of gluconolactone in 10mL of deionized water, add it to lactate-sodium lactate buffer solution, and stir until homogeneous to obtain lactate-gluconolactone composite acidifier.

[0054] The method for preparing the salt-containing probiotic composition includes the following steps:

[0055] S1. Disperse the freeze-dried Lactobacillus plantarum powder in pre-cooled 0.1M PBS buffer to obtain a probiotic suspension;

[0056] S2. Disperse refined coarse salt and modified carboxymethyl chitosan separately in deionized water, and add them sequentially to the probiotic suspension. Mix them evenly at 25~30℃ to obtain the first mixture;

[0057] S3. Slowly add the lactic acid-gluconolactone complex acidifier to the first mixture, stir evenly, and let it stand at 37°C to react, thus obtaining the second mixture;

[0058] S4. Add sodium alginate solution to the second mixture and stir until homogeneous. Add the resulting mixture dropwise to CaCl2 solution and allow it to solidify for 20-30 minutes. Wash the product with deionized water, add trehalose, and freeze-dry to obtain a salt-containing probiotic composition.

[0059] It should be noted that the lactic acid-gluconolactone complex acidifier must be prepared and used immediately, and its weight fraction is based on solid content.

[0060] Example 2: This example is based on Example 1, but differs from Example 1 in that it discloses a salt-containing probiotic composition, comprising the following components in parts by weight: 2 parts refined coarse salt, 8 parts modified carboxymethyl chitosan, 1.5 parts lactic acid-gluconolactone composite acidifier, 1.5 parts sodium alginate, 0.5 parts calcium chloride, 5 parts freeze-dried Lactobacillus plantarum powder, and 2 parts trehalose.

[0061] The other components and preparation methods are the same as in Example 1.

[0062] Example 3: This example is based on Example 1, but differs from Example 1 in that it discloses a salt-containing probiotic composition, comprising the following components in parts by weight: 5 parts refined coarse salt, 15 parts modified carboxymethyl chitosan, 3 parts lactic acid-gluconolactone composite acidifier, 3 parts sodium alginate, 1.5 parts calcium chloride, 15 parts freeze-dried Lactobacillus plantarum powder, and 5 parts trehalose.

[0063] The other components and preparation methods are the same as in Example 1.

[0064] Example 4: This example is based on Example 1, but differs from Example 1 in that the mass ratio of coarse salt powder to citric acid is 100:8.

[0065] The other components and preparation methods are the same as in Example 1.

[0066] Example 5: This example is based on Example 1, but differs from Example 1 in that the mass ratio of carboxymethyl chitosan, gallic acid and tannic acid in this example is 1:0.6:1.5.

[0067] The other components and preparation methods are the same as in Example 1.

[0068] Comparative Example 1: This comparative example is based on Example 1, but differs from Example 1 in that the refined coarse salt described in this comparative example is not treated with hydrogen peroxide.

[0069] The method for preparing the refined coarse salt includes the following steps:

[0070] S11. Put 100g of coarse salt powder into a ball mill jar, add 200mL of anhydrous ethanol and grinding balls, grind and crush to obtain a coarse salt slurry with D50 of 200~500nm.

[0071] S12. Dissolve 5g of citric acid in 50mL of anhydrous ethanol, and slowly add the resulting citric acid solution to the slurry of codonopsis pilosula while stirring continuously for 2h to obtain the third compound.

[0072] S13. Dissolve 1.5g of stearic acid in 50mL of hot anhydrous ethanol, add the resulting stearic acid solution to the third compound, stir continuously for 1h, concentrate the resulting slurry under vacuum at 50℃, dry under vacuum at 50~60℃ for 12~24h, grind and disperse, and pass through a 100-mesh sieve to obtain refined coarse salt.

[0073] The other components and preparation methods are the same as in Example 1.

[0074] Comparative Example 2: This comparative example is based on Example 1, but differs from Example 1 in that the refined coarse salt described in this comparative example is not treated with citric acid.

[0075] The method for preparing the refined coarse salt includes the following steps:

[0076] S11. Put 100g of coarse salt powder into a ball mill jar, add 200mL of anhydrous ethanol and grinding balls, grind and crush to obtain a coarse salt slurry with D50 of 200~500nm.

[0077] S12. Add 2 mL of 30% hydrogen peroxide to 50 mL of anhydrous ethanol, and slowly add the resulting solution dropwise to the slurry of cobalt salt. Stir magnetically at room temperature for 1 h to obtain the first compound.

[0078] S13. Dissolve 1.5g of stearic acid in 50mL of hot anhydrous ethanol, add the resulting stearic acid solution to the first compound, stir continuously for 1h, concentrate the resulting slurry under vacuum at 50℃, dry under vacuum at 50~60℃ for 12~24h, grind and disperse, and pass through a 100-mesh sieve to obtain refined coarse salt.

[0079] The other components and preparation methods are the same as in Example 1.

[0080] Comparative Example 3: This comparative example is based on Example 1, but differs from Example 1 in that the modified carboxymethyl chitosan in this comparative example is not modified with gallic acid.

[0081] The method for preparing the modified carboxymethyl chitosan includes the following steps:

[0082] S21. Dissolve 10g of carboxymethyl chitosan in 400mL of deionized water, and adjust the pH to 5-5.5 with HCl to obtain a carboxymethyl chitosan solution.

[0083] S22. Dissolve 5g of tannic acid in deionized water, adjust the pH to 8-8.5 with NaOH, add the resulting tannic acid solution to the carboxymethyl chitosan solution, stir and react at room temperature for 6-12 hours, adjust the pH of the resulting reaction solution to 6-6.5, add anhydrous ethanol to precipitate, collect the precipitate by centrifugation, wash with anhydrous ethanol, redissolve in deionized water, freeze dry to obtain modified carboxymethyl chitosan.

[0084] The other components and preparation methods are the same as in Example 1.

[0085] Comparative Example 4: This comparative example is based on Example 1, but differs from Example 1 in that the modified carboxymethyl chitosan in this comparative example is not modified with tannic acid.

[0086] The method for preparing the modified carboxymethyl chitosan includes the following steps:

[0087] S21. Dissolve 10g of carboxymethyl chitosan in 400mL of deionized water, and adjust the pH to 5-5.5 with HCl to obtain a carboxymethyl chitosan solution.

[0088] S22. Dissolve 3g gallic acid in 80mL deionized water, add 2.5g EDC·HCl and 1.5g NHS, stir and activate at room temperature for 30min, slowly add the resulting gallic acid solution dropwise to the carboxymethyl chitosan solution, adjust the pH to 6~6.5 with NaOH, stir and react at room temperature for 12~24h, dialysis and purify the resulting reaction solution, freeze dry to obtain modified carboxymethyl chitosan.

[0089] The other components and preparation methods are the same as in Example 1.

[0090] Comparative Example 5: This comparative example is based on Example 1, but differs from Example 1 in that the lactic acid-gluconolactone complex acidifier is replaced with an equal amount of dilute HCl.

[0091] The other components and preparation methods are the same as in Example 1.

[0092] Comparative Example 6: This comparative example is based on Example 1, but unlike Example 1, no modified carboxymethyl chitosan is added.

[0093] The other components and preparation methods are the same as in Example 1.

[0094] Comparative Example 7: This comparative example is based on Example 1, but unlike Example 1, this comparative example does not add lactic acid-gluconolactone complex acidifier.

[0095] The other components and preparation methods are the same as in Example 1.

[0096] Comparative Example 8: This comparative example is based on Example 1, but unlike Example 1, this comparative example does not add sodium alginate and calcium chloride.

[0097] The other components and preparation methods are the same as in Example 1.

[0098] Comparative Example 9: This comparative example is based on Example 1, but differs from Example 1 in that it includes the following components in parts by weight: 7 parts refined coarse salt, 10 parts freeze-dried Lactobacillus plantarum powder, and 3 parts trehalose.

[0099] The refined coarse salt is commercially available refined coarse salt powder that has not been treated with hydrogen peroxide and citric acid.

[0100] The method for preparing the salt-containing probiotic composition includes the following steps:

[0101] S1. Disperse the freeze-dried Lactobacillus plantarum powder in pre-cooled 0.1M PBS buffer to obtain a probiotic suspension;

[0102] S2. Disperse refined coarse salt in deionized water, add it to the probiotic suspension, mix evenly at 25~30℃, add trehalose, freeze dry, and obtain a salt-containing probiotic composition.

[0103] Experimental verification:

[0104] Experiment 1:

[0105] (1) Determination of free metal ion content: The refined coarse salt of Examples 1, 4, Comparative Examples 1-2 and 8 was dissolved in deionized water, filtered, and the Fe content in the filtrate was determined by ICP-OES. 2+ Fe 3+ Cu2+ content.

[0106]

[0107] Table 1 shows the free metal ion content in refined cobalt salt. As can be seen from the table, the free Fe ions in refined cobalt salt treated with hydrogen peroxide and citric acid are... 2+ Fe 3+ Cu 2+ The content decreased significantly.

[0108] (2) In step S3 of the preparation process of the salt-containing probiotic composition, the pH change curve of the reaction system of Example 1 and Comparative Example 5 was monitored in real time after the addition of lactic acid-gluconolactone composite acidifier. Furthermore, at 0, 2, 4, and 6 hours of reaction, equal volumes of reaction solutions from Example 1, Example 5, and Comparative Examples 3-6 were taken, centrifuged at high speed, and the supernatant was collected. Fe was detected using ICP-OES. 3+ concentration.

[0109]

[0110] like Figure 1 The figure shows the pH change curves of the reaction systems of Example 1 and Comparative Example 5 over time. It can be seen from the figure that the pH decrease in the reaction system with the addition of lactic acid-gluconolactone composite acidifier is relatively gradual, which is more conducive to the orderly and uniform formation of MPNs networks. Table 2 shows the Fe in S3 during the preparation process of the salt-containing probiotic composition. 3+ The concentration changes, as shown in the table, indicate that the lactic acid-gluconolactone complex acidifier causes the system pH to decrease slowly, thereby allowing the citrate carboxyl groups to react with Fe. 3+ The coordination gradually dissociates, and tannic acid and gallic acid orderly capture Fe. 3+ and Fe 3+ A coordination reaction occurs, forming a dense, uniform, and stable metal-polyphenol network (MPNs) on the surface of probiotics.

[0111] Experiment 2:

[0112] (1) Storage stability: The salt-containing probiotic compositions prepared in the examples and comparative examples were stored at room temperature of 25°C for 90 days, and the survival rate of the probiotics was tested.

[0113] (2) Thermal stability: The salt-containing probiotic compositions prepared in the examples and comparative examples were placed in a 60°C water bath for 30 min and the survival rate of the probiotics was tested.

[0114] (3) Gastric acid tolerance: The salt-containing probiotic compositions prepared in the examples and comparative examples were placed in simulated gastric juice (containing 3 g / L pepsin) at pH 2.0 and incubated at 37°C for 2 h. The survival rate of the probiotics was then detected.

[0115] (4) Intestinal fluid tolerance: After being treated with simulated gastric fluid (containing 3 g / L pepsin) at pH 2.0 for 2 h, the samples were transferred to simulated intestinal fluid (pH 6.8, containing trypsin) for 4 h, and the survival rate of probiotics was detected.

[0116]

[0117] Table 3 shows the survival rate of probiotics in the salt-containing probiotic composition. As can be seen from the table, Fe... 3+ It undergoes coordination reactions with polyphenols (tannic acid and gallic acid) to form a dense, uniform, and stable MPN network, providing physical barrier protection and enhancing the tolerance of probiotics to stresses such as acid and heat treatment. Furthermore, sodium alginate-Ca... 2+ The gel network and modified carboxymethyl chitosan form a composite gel network. Under the dual protection of the composite gel network and the MPNs network, the survival rate of probiotics can be significantly improved.

[0118] The above description is merely a preferred embodiment of the present invention and is not intended to limit the scope of protection of the present invention. Any modifications, equivalent substitutions, and improvements made by those skilled in the art within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A salt-containing probiotic composition, characterized in that, The product comprises the following components in parts by weight: 2-5 parts refined coarse salt, 8-15 parts modified carboxymethyl chitosan, 1.5-3 parts lactic acid-gluconolactone composite acidifier, 1.5-3 parts sodium alginate, 0.5-1.5 parts calcium chloride, 5-15 parts freeze-dried Lactobacillus plantarum powder, and 2-5 parts trehalose. The refined coarse salt is treated with hydrogen peroxide to remove Fe. 2+ Oxidized to Fe 3+ Citric acid is used to bind free Fe. 3+ and Cu 2+ Finally, it is passivated with stearic acid for hydrophobic treatment; The modified carboxymethyl chitosan was modified with tannic acid and gallic acid. The refined coarse salt and the modified carboxymethyl chitosan are reacted via Fe... 3+ A metal-polyphenol network is formed by bridging, and the metal-polyphenol network together with the sodium alginate-calcium chloride gel network constitutes a double-layer protective structure. The lactic acid-gluconolactone complex acidifier is used to control Fe. 3+ The release rate is adjusted to form a uniform MPNs network; The method for preparing the refined coarse salt includes the following steps: S11. Load coarse salt powder into a ball mill jar, add anhydrous ethanol and grinding balls, grind and crush to obtain coarse salt slurry; S12. Hydrogen peroxide was added to anhydrous ethanol, and the resulting solution was slowly added dropwise to the slurry of cobalt salt. The mixture was then magnetically stirred to obtain the first compound. S13. Dissolve citric acid in anhydrous ethanol, and slowly add the resulting citric acid solution dropwise to the first compound while stirring continuously to obtain the second compound; S14. Stearic acid is dissolved in anhydrous ethanol, and the resulting stearic acid solution is added to the second compound. The mixture is stirred continuously, and the resulting slurry is concentrated by vacuum rotary evaporation, vacuum dried, ground and dispersed to obtain refined coarse salt. The method for preparing the modified carboxymethyl chitosan includes the following steps: S21. Dissolve carboxymethyl chitosan in deionized water, adjust the pH with HCl to obtain a carboxymethyl chitosan solution; S22. Dissolve gallic acid in deionized water, add EDC·HCl and NHS, stir to activate, and slowly add the resulting gallic acid solution to the carboxymethyl chitosan solution. Adjust the pH with NaOH, stir the reaction, and dialysis the resulting reaction solution to obtain gallic acid-carboxymethyl chitosan. S23. Dissolve tannic acid in deionized water, adjust the pH with NaOH, add the resulting tannic acid solution to gallic acid-carboxymethyl chitosan, stir to react, add the resulting reaction solution to anhydrous ethanol to precipitate, collect the precipitate by centrifugation, wash with anhydrous ethanol, redissolve in deionized water, freeze dry to obtain modified carboxymethyl chitosan. The mass ratio of carboxymethyl chitosan, gallic acid, and tannic acid is 1:0.3~0.6:0.5~1.

5.

2. The salt-containing probiotic composition according to claim 1, characterized in that, The mass ratio of coarse salt powder to citric acid is 100:5~8.

3. The salt-containing probiotic composition according to claim 1, characterized in that, The preparation method of the lactic acid-gluconolactone composite acidifier includes the following steps: S31. Dissolve sodium lactate in deionized water, add lactic acid, stir well, and adjust the pH with NaOH to obtain a lactate-sodium lactate buffer solution; S32. Dissolve gluconolactone in deionized water, add it to lactate-sodium lactate buffer solution, and stir until homogeneous to obtain lactate-gluconolactone composite acidifier.

4. A method for preparing a salt-containing probiotic composition, applied to the preparation of a salt-containing probiotic composition as described in any one of claims 1 to 3, characterized in that, The method includes the following steps: S1. Disperse the freeze-dried Lactobacillus plantarum powder in PBS buffer to obtain a probiotic suspension; S2. Disperse refined coarse salt and modified carboxymethyl chitosan separately in deionized water, add them sequentially to the probiotic suspension, mix evenly, and obtain the first mixture; S3. Slowly add the lactic acid-gluconolactone complex acidifier to the first mixture, stir evenly, and let it stand to react to obtain the second mixture; S4. Add sodium alginate solution to the second mixture and stir until homogeneous. Add the resulting mixture dropwise to CaCl2 solution to solidify the reaction. Wash the product with deionized water, add trehalose, and freeze-dry to obtain a salt-containing probiotic composition.

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