A sustained-release synergistic carboxymethyl active pachyman polysaccharide composition and a preparation method thereof

Abstract

The application discloses a slow-release synergistic carboxymethyl active Pachyman polysaccharide composition and a preparation method thereof. The method comprises the following steps: dissolving and pre-activating carboxymethyl active Pachyman polysaccharide, pre-complexing chitosan and tea polyphenol under weak acid conditions; adding the pre-activated polysaccharide liquid into chitooligosaccharide-tea polyphenol pre-complexing liquid to perform electrostatic association, then adding a phytate component, and then adding a calcium salt component dropwise to perform ion bridge connection aging, so as to obtain a composite microgel dispersion liquid; adding a drying protection component and drying, so as to obtain a composition which can form a composite microgel particle after rehydration. The application improves the embedding rate and rehydration stability of carboxymethyl active Pachyman polysaccharide through step-by-step bridge connection of electrostatic association and calcium phytate, reduces the burst release in a gastric juice environment, realizes sustained release in an intestinal juice environment, and improves the antioxidant synergistic performance.

Description

Technical Field

[0001] This invention relates to the field of Poria cocos polysaccharide composition technology, and in particular to a sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition and its preparation method. Background Technology

[0002] Poria cocos polysaccharides are one of the main active components of Poria cocos, possessing potential applications in immunomodulation, antioxidation, anti-inflammation, and intestinal regulation, and have attracted attention in the fields of functional foods, nutritional preparations, and natural active ingredient delivery. Due to the poor water solubility, easy aggregation of molecular chains, and insufficient system stability of natural Poria cocos polysaccharides, carboxymethylation modification is often used in practical applications to improve their hydrophilicity and dispersibility. Carboxymethylated active Poria cocos polysaccharides, by introducing carboxymethyl groups, can improve solubility while retaining certain polysaccharide activity, and have become an important direction for the deep processing of Poria cocos polysaccharides. Existing technologies also include research on improving the stability and release behavior of active polysaccharides using emulsification, encapsulation, nanocarriers, or polysaccharide complexation.

[0003] However, existing technologies for carboxymethyl Poria cocos polysaccharides mainly focus on degree of substitution, molecular weight, extraction and purification, or single-activity evaluation, with insufficient attention paid to their burst release control in the gastrointestinal environment, compound stability, and sustained-release effect. Directly mixing carboxymethyl active Poria cocos polysaccharides with conventional excipients can easily lead to problems such as excessively rapid release, decreased activity retention, unstable rehydration and dispersion, and batch-to-batch particle size fluctuations. If ordinary encapsulation or gelation systems are used, there may be defects such as insufficient encapsulation rate, premature release under acidic conditions, and insufficient release in the intestinal environment. Therefore, there is an urgent need to develop a method for preparing carboxymethyl active Poria cocos polysaccharide compositions that combine rehydration stability, low burst release in gastric juice, sustained release in intestinal juice, and antioxidant synergistic effects. Summary of the Invention

[0004] To address the aforementioned issues, this invention proposes a sustained-release synergistic carboxymethyl active poria cocos polysaccharide composition and its preparation method. The method involves electrostatic association of carboxymethyl active poria cocos polysaccharide and chitosan oligosaccharide-tea polyphenol pre-composite solution at a specific pH, followed by adsorption by phytate and ion bridging by the addition of calcium salt to form a composite microgel dispersion. After protective drying, a composition that can form microgel particles after rehydration is obtained.

[0005] This invention can be achieved through the following technical solutions: A method for preparing a sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition includes the following steps: S1. Dissolve carboxymethyl active Poria cocos polysaccharide in water and adjust the pH to 6.0-7.2 to obtain a pre-activated polysaccharide solution; combine chitosan oligosaccharide and tea polyphenols under pH 4.0-5.5 conditions to obtain a chitosan oligosaccharide-polyphenol pre-complex solution; S2. The pre-activated polysaccharide solution is added to the chitosan oligosaccharide-tea polyphenol pre-composite solution, and electrostatic association is carried out under the condition of pH 5.0-6.2 to obtain a composite microparticle dispersion. Then, phytate component is added to the composite microparticle dispersion first, and then calcium salt component is added dropwise. Ion bridging aging is carried out under the condition of pH 5.8-6.8 to obtain a composite microgel dispersion. The mass ratio of carboxymethyl active poria cocos polysaccharide to chitosan oligosaccharide is 100:(15-35), and the mass ratio of phytate component to calcium salt component is 1:(0.5-2.0). S3. Add a drying protection component to the composite microgel dispersion, mix and dry to obtain a sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition that can form composite microgel particles after rehydration.

[0006] Preferably, by weight, the carboxymethyl active Poria cocos polysaccharide is 40-70 parts, the chitosan oligosaccharide is 8-20 parts, the tea polyphenols are 0.5-5 parts, the phytate component is 1-6 parts, the calcium salt component is 1-5 parts, and the drying protection component is 10-30 parts.

[0007] Preferably, the carboxymethyl active Poria cocos polysaccharide has a carboxymethyl substitution degree of 0.25 to 0.58, a weight-average molecular weight of 50 to 220 kDa, and a total sugar content of not less than 70%.

[0008] Preferably, in step S1, the mass concentration of carboxymethyl active Poria cocos polysaccharide in the pre-activated polysaccharide solution is 0.5% to 3.0%, the pre-activation temperature is 35 to 55°C, and the pre-activation time is 20 to 60 min.

[0009] Preferably, the degree of deacetylation of the chitosan oligosaccharide is not less than 85%, and the molecular weight is 1-10 kDa; the compounding time of the chitosan oligosaccharide and tea polyphenols is 15-45 min.

[0010] Preferably, in step S2, the pre-activated polysaccharide solution is added to the chitosan oligosaccharide-tea polyphenol pre-complex solution by dropwise addition, with a dropwise addition time of 10–40 min, an electrostatic association stirring speed of 300–1000 rpm, and an electrostatic association time of 20–60 min.

[0011] Preferably, the phytate component is one or two of sodium phytate and potassium phytate; the calcium salt component is one or more of calcium lactate, calcium gluconate, and calcium chloride; and the ion bridging aging temperature is 20–40°C, and the time is 30–120 min.

[0012] Preferably, the drying and protective component is one or more of trehalose, mannitol, maltodextrin, and fructooligosaccharides; the drying is freeze-drying or spray drying.

[0013] Preferably, when freeze-drying is used, the pre-freezing temperature is -40 to -20°C, the pre-freezing time is 4 to 12 hours, and the vacuum drying time is 18 to 36 hours; when spray drying is used, the inlet air temperature is 120 to 160°C, and the outlet air temperature is 60 to 85°C.

[0014] Preferably, the composition, after rehydration, forms composite microgel particles with an average particle size of 200–900 nm, a PDI of no more than 0.35, and an encapsulation rate of carboxymethyl active Poria cocos polysaccharide of no less than 80%; the cumulative release rate in simulated gastric juice is no more than 30% in 2 hours and in simulated intestinal juice is 65%–90% in 8 hours.

[0015] The beneficial effects of this invention are: This invention constructs a sustained-release synergistic composition that forms stable composite microgel particles after rehydration by electrostatic association of carboxymethyl active Poria cocos polysaccharide and chitosan oligosaccharide-tea polyphenol pre-complex solution under specific pH conditions, and further employs a stepwise ion bridging method of phytate adsorption followed by calcium salt addition. Compared with direct mixing or ordinary bridging methods, this invention can improve the encapsulation rate of carboxymethyl active Poria cocos polysaccharide, improve the particle size uniformity and dispersion stability after rehydration, reduce the initial burst release in an acidic simulated gastric juice environment, and achieve continuous release in a simulated intestinal juice environment. Tea polyphenols, after pre-complexation with chitosan oligosaccharide, can be well immobilized in the microgel structure, synergistically exerting antioxidant and synergistic effects with carboxymethyl active Poria cocos polysaccharide. Performance tests show that the sample in the example has a high encapsulation rate, low gastric juice release rate after 2 hours, sufficient release in intestinal juice after 8 hours, and no significant sedimentation after 48 hours of rehydration, indicating that this invention effectively solves the problems of excessively rapid release, poor rehydration stability, and insufficient synergistic effect in existing carboxymethyl Poria cocos polysaccharide compositions. Attached Figure Description

[0016] The accompanying drawings are provided to further illustrate the invention and form part of the specification. They are used in conjunction with embodiments of the invention to explain the invention and do not constitute a limitation thereof. In the drawings: Figure 1 The results are from tests simulating the cumulative release rate of gastric and intestinal fluids. Detailed Implementation

[0017] The following provides a detailed description of the embodiments of the present invention: These embodiments are implemented based on the technical solution of the present invention, and provide detailed implementation methods and processes. However, the scope of protection of the present invention is not limited to the following embodiments. Experimental methods in the following embodiments that do not specify specific conditions are generally performed under conventional conditions.

[0018] Example 1: A method for preparing a sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition, comprising the following steps: S1. Weigh 40 g of carboxymethyl active Poria cocos polysaccharide. The degree of carboxymethyl substitution of the carboxymethyl active Poria cocos polysaccharide is 0.42, the weight-average molecular weight is 120 kDa, and the total sugar content is 82%. Add it to purified water to prepare a 2.0% carboxymethyl active Poria cocos polysaccharide solution. Adjust the pH to 6.8 and stir at 45℃ for 40 min to pre-activate the polysaccharide solution. Separately weigh 8 g of chitosan oligosaccharide and 0.5 g of tea polyphenols. The degree of deacetylation of the chitosan oligosaccharide is 90% and the molecular weight is 5 kDa. Add the chitosan oligosaccharide to purified water to prepare a 1.0% chitosan oligosaccharide solution. Add the tea polyphenols and adjust the pH to 4.8. Stir and compound in the dark for 30 min to obtain a chitosan oligosaccharide-tea polyphenol pre-composite solution. S2. Under the condition of stirring speed of 600 rpm, the pre-activated polysaccharide solution obtained in step S1 is added dropwise to the chitosan oligosaccharide-tea polyphenol pre-composite solution for 25 min. The pH of the system is adjusted to 5.6, and electrostatic association is continued for 40 min to obtain a composite microparticle dispersion. Then, 1 g of sodium phytate is added to the composite microparticle dispersion, and after stirring for 10 min, an aqueous solution containing 1 g of calcium lactate is added dropwise to adjust the pH of the system to 6.3. The system is then aged at 30℃ for 60 min to obtain a composite microgel dispersion. The mass ratio of carboxymethyl active poria cocos polysaccharide to chitosan oligosaccharide is 100:20, and the mass ratio of sodium phytate to calcium lactate is 1:1. S3. Add 10 g of trehalose to the composite microgel dispersion obtained in step S2, stir and mix evenly, and then freeze-dry. The freeze-drying conditions are: pre-freezing at -30℃ for 8 h, followed by vacuum drying for 24 h, to obtain a sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition that can form composite microgel particles after rehydration.

[0019] Example 2: A method for preparing a sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition, comprising the following steps: S1. Weigh 55 g of carboxymethyl active Poria cocos polysaccharide. The degree of carboxymethyl substitution of the carboxymethyl active Poria cocos polysaccharide is 0.42, the weight-average molecular weight is 120 kDa, and the total sugar content is 82%. Add it to purified water to prepare a 2.0% carboxymethyl active Poria cocos polysaccharide solution. Adjust the pH to 6.8 and stir at 45℃ for 40 min to pre-activate the polysaccharide solution. Separately weigh 14 g of chitosan oligosaccharide and 2.5 g of tea polyphenols. The degree of deacetylation of the chitosan oligosaccharide is 90% and the molecular weight is 5 kDa. Add the chitosan oligosaccharide to purified water to prepare a 1.0% chitosan oligosaccharide solution. Add the tea polyphenols and adjust the pH to 4.8. Stir and compound in the dark for 30 min to obtain a chitosan oligosaccharide-tea polyphenol pre-composite solution. S2. Under the condition of stirring speed of 600 rpm, the pre-activated polysaccharide solution obtained in step S1 is added dropwise to the chitosan oligosaccharide-tea polyphenol pre-composite solution for 25 min. The pH of the system is adjusted to 5.6, and electrostatic association is continued for 40 min to obtain a composite microparticle dispersion. Then, 3 g of sodium phytate is added to the composite microparticle dispersion, and after stirring for 10 min, an aqueous solution containing 4 g of calcium lactate is added dropwise to adjust the pH of the system to 6.3. The system is then aged at 30℃ for 60 min by ion bridging to obtain a composite microgel dispersion. The mass ratio of carboxymethyl active poria cocos polysaccharide to chitosan oligosaccharide is 100:25.5, and the mass ratio of sodium phytate to calcium lactate is 1:1.33. S3. Add 20 g of trehalose to the composite microgel dispersion obtained in step S2, stir and mix evenly, and then freeze-dry. The freeze-drying conditions are: pre-freezing at -30℃ for 8 h, followed by vacuum drying for 24 h, to obtain a sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition that can form composite microgel particles after rehydration.

[0020] Example 3: A method for preparing a sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition, comprising the following steps: S1. Weigh 70 g of carboxymethyl active Poria cocos polysaccharide. The degree of carboxymethyl substitution of the carboxymethyl active Poria cocos polysaccharide is 0.42, the weight-average molecular weight is 120 kDa, and the total sugar content is 82%. Add it to purified water to prepare a 2.0% carboxymethyl active Poria cocos polysaccharide solution. Adjust the pH to 6.8 and stir at 45℃ for 40 min to pre-activate the polysaccharide solution. Separately weigh 20 g of chitosan oligosaccharide and 5 g of tea polyphenols. The degree of deacetylation of the chitosan oligosaccharide is 90% and the molecular weight is 5 kDa. Add the chitosan oligosaccharide to purified water to prepare a 1.0% chitosan oligosaccharide solution. Add the tea polyphenols and adjust the pH to 4.8. Stir and compound in the dark for 30 min to obtain a chitosan oligosaccharide-tea polyphenol pre-composite solution. S2. Under the condition of stirring speed of 600 rpm, the pre-activated polysaccharide solution obtained in step S1 is added dropwise to the chitosan oligosaccharide-tea polyphenol pre-composite solution for 25 min. The pH of the system is adjusted to 5.6, and electrostatic association is continued for 40 min to obtain a composite microparticle dispersion. Then, 5 g of sodium phytate is added to the composite microparticle dispersion, and after stirring for 10 min, an aqueous solution containing 5 g of calcium lactate is added dropwise to adjust the pH of the system to 6.3. The system is then aged at 30℃ for 60 min to obtain a composite microgel dispersion. The mass ratio of carboxymethyl active poria cocos polysaccharide to chitosan oligosaccharide is 100:28.6, and the mass ratio of sodium phytate to calcium lactate is 1:1. S3. Add 30 g of trehalose to the composite microgel dispersion obtained in step S2, stir and mix evenly, and then freeze-dry. The freeze-drying conditions are: pre-freezing at -30℃ for 8 h, followed by vacuum drying for 24 h, to obtain a sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition that can form composite microgel particles after rehydration.

[0021] Comparative Example 1: The difference between this comparative example and Example 1 is that chitosan oligosaccharide is not added, and the electrostatic association structure between carboxymethyl active poria cocos polysaccharide and chitosan oligosaccharide is not formed.

[0022] The preparation method of the Poria cocos polysaccharide composition in this comparative example includes the following steps: S1. Weigh 40 g of carboxymethyl active Poria cocos polysaccharide. The degree of carboxymethyl substitution of carboxymethyl active Poria cocos polysaccharide is 0.42, the weight-average molecular weight is 120 kDa, and the total sugar content is 82%. Add it to purified water to prepare a 2.0% (w / w) carboxymethyl active Poria cocos polysaccharide solution. Adjust the pH to 6.8 and stir at 45℃ for 40 min to pre-activate the polysaccharide solution. Separately, weigh 0.5 g of tea polyphenols, add the tea polyphenols to purified water to dissolve them, adjust the pH to 4.8, and stir in the dark for 30 min to obtain a tea polyphenol solution. S2. Under the condition of stirring speed of 600 rpm, the pre-activated polysaccharide solution obtained in step S1 is added dropwise to the tea polyphenol solution for 25 min. The pH of the system is adjusted to 5.6, and stirring is continued for 40 min to obtain a mixed dispersion. Then, 1 g of sodium phytate is added to the mixed dispersion, and after stirring for 10 min, an aqueous solution containing 1 g of calcium lactate is added dropwise to adjust the pH of the system to 6.3. The mixture is then treated at 30℃ for 60 min to obtain the Poria cocos polysaccharide dispersion. S3. Add 10 g of trehalose to the poria polysaccharide dispersion obtained in step S2, stir and mix evenly, and then freeze-dry. The freeze-drying conditions are: pre-freezing at -30℃ for 8 h, followed by vacuum drying for 24 h, to obtain the poria polysaccharide composition of Comparative Example 1.

[0023] Comparative Example 2: The difference between this comparative example and Example 1 is that sodium phytate and calcium lactate are not added, and calcium phytate ion bridging aging is not performed.

[0024] The preparation method of the Poria cocos polysaccharide composition in this comparative example includes the following steps: S1. Weigh 40 g of carboxymethyl active Poria cocos polysaccharide. The degree of carboxymethyl substitution of the carboxymethyl active Poria cocos polysaccharide is 0.42, the weight-average molecular weight is 120 kDa, and the total sugar content is 82%. Add it to purified water to prepare a 2.0% carboxymethyl active Poria cocos polysaccharide solution. Adjust the pH to 6.8 and stir at 45℃ for 40 min to pre-activate the polysaccharide solution. Separately weigh 8 g of chitosan oligosaccharide and 0.5 g of tea polyphenols. The degree of deacetylation of the chitosan oligosaccharide is 90% and the molecular weight is 5 kDa. Add the chitosan oligosaccharide to purified water to prepare a 1.0% chitosan oligosaccharide solution. Add the tea polyphenols and adjust the pH to 4.8. Stir and compound in the dark for 30 min to obtain a chitosan oligosaccharide-tea polyphenol pre-composite solution. S2. Under the condition of stirring speed of 600 rpm, the pre-activated polysaccharide solution obtained in step S1 is added dropwise to the chitosan oligosaccharide-tea polyphenol pre-composite solution for 25 min. The pH of the system is adjusted to 5.6, and electrostatic association is continued for 40 min to obtain a composite microparticle dispersion. The pH of the composite microparticle dispersion is adjusted to 6.3, and the mixture is stirred at 30℃ for 60 min to obtain a composite dispersion without calcium phytate bridging. The mass ratio of carboxymethyl active poria cocos polysaccharide to chitosan oligosaccharide is 100:20. S3. Add 10g of trehalose to the composite dispersion obtained in step S2, stir and mix evenly, and then freeze-dry. The freeze-drying conditions are: pre-freeze at -30℃ for 8 h, followed by vacuum drying for 24 h, to obtain the Poria cocos polysaccharide composition of Comparative Example 2.

[0025] Comparative Example 3: The difference between this comparative example and Example 1 is that sodium phytate and calcium lactate were premixed and then added to the composite microparticle dispersion in one go.

[0026] The preparation method of the Poria cocos polysaccharide composition in this comparative example includes the following steps: S1. Weigh 40 g of carboxymethyl active Poria cocos polysaccharide. The degree of carboxymethyl substitution of the carboxymethyl active Poria cocos polysaccharide is 0.42, the weight-average molecular weight is 120 kDa, and the total sugar content is 82%. Add it to purified water to prepare a 2.0% carboxymethyl active Poria cocos polysaccharide solution. Adjust the pH to 6.8 and stir at 45℃ for 40 min to pre-activate the polysaccharide solution. Separately weigh 8 g of chitosan oligosaccharide and 0.5 g of tea polyphenols. The degree of deacetylation of the chitosan oligosaccharide is 90% and the molecular weight is 5 kDa. Add the chitosan oligosaccharide to purified water to prepare a 1.0% chitosan oligosaccharide solution. Add the tea polyphenols and adjust the pH to 4.8. Stir and compound in the dark for 30 min to obtain a chitosan oligosaccharide-tea polyphenol pre-composite solution. S2. Under the condition of stirring speed of 600 rpm, the pre-activated polysaccharide solution obtained in step S1 was added dropwise to the chitosan oligosaccharide-tea polyphenol pre-composite solution for 25 min. The pH of the system was adjusted to 5.6, and electrostatic association was continued for 40 min to obtain a composite microparticle dispersion. Separately, 1 g of sodium phytate and 1 g of calcium lactate were dissolved in purified water and mixed to obtain a sodium phytate-calcium lactate mixture. The sodium phytate-calcium lactate mixture was added to the composite microparticle dispersion at one time, the pH of the system was adjusted to 6.3, and the mixture was stirred at 30℃ for 60 min to obtain a control composite dispersion. The mass ratio of carboxymethyl active poria cocos polysaccharide to chitosan oligosaccharide was 100:20, and the mass ratio of sodium phytate to calcium lactate was 1:1. S3. Add 10 g of trehalose to the control composite dispersion obtained in step S2, stir and mix evenly, and then freeze-dry. The freeze-drying conditions are: pre-freeze at -30℃ for 8 h, followed by vacuum drying for 24 h to obtain the Poria cocos polysaccharide composition of Comparative Example 3.

[0027] Performance testing 1. Measurement of rehydrated microgel particle size, PDI and Zeta potential Appropriate amounts of the dried samples obtained in Examples 1-3 and Comparative Examples 1-3 were taken and dispersed in purified water to control the mass concentration of carboxymethyl active Poria cocos polysaccharide in the samples to 1.0 mg / mL. The samples were magnetically stirred at 25℃ for 10 min, then dispersed using an ultrasonic water bath for 3 min. After standing for 5 min, the upper uniform dispersion was collected for testing. The average particle size and PDI of the samples were determined using a dynamic light scattering particle size analyzer at 25℃ and a scattering angle of 173°. Each sample was tested in triplicate, and the average value was taken. The Zeta potential was determined using electrophoretic light scattering, with each sample tested in triplicate.

[0028] 2. Carboxymethyl active Poria cocos polysaccharide encapsulation rate test Samples of the composite microgel dispersions obtained in Examples 1-3 and Comparative Examples 1-3 were taken before freeze-drying. After sampling, the samples were centrifuged at 10000 r / min for 10 min, and the supernatant was collected. The content of unencapsulated carboxymethyl active Poria cocos polysaccharides in the supernatant was determined by the phenol-sulfuric acid method. Separately, samples from the same batch were subjected to thorough demulsification / dissociation treatment, and the total carboxymethyl active Poria cocos polysaccharide content in the samples was determined. Specifically, the samples were added to pH 7.4 phosphate buffer and sonicated for 10 min to fully dissociate the composite microgel structure, and then the total sugar content was determined by the phenol-sulfuric acid method.

[0029] The embedding rate is calculated using the following formula: Encapsulation rate (%) = (Total carboxymethyl active poria polysaccharide content - Unencapsulated carboxymethyl active poria polysaccharide content in supernatant) / Total carboxymethyl active poria polysaccharide content × 100%.

[0030] Each group of samples was measured in parallel three times, and the average value of the results was taken.

[0031] Table 1. Results of tests on particle size, PDI, Zeta potential, and encapsulation efficiency of the rehydrated microgels.

[0032] 3. Simulated gastric and intestinal fluid release performance test Take appropriate amounts of the dried samples obtained in Examples 1-3 and Comparative Examples 1-3, respectively, and calculate the amount of carboxymethyl active Poria cocos polysaccharide added to each group of samples based on the content of carboxymethyl active Poria cocos polysaccharide in the samples. Add the sample to 50 mL of simulated gastric juice (pH 1.2 hydrochloric acid-sodium chloride solution) and place it in a constant temperature shaker at 37℃ with a shaking speed of 100 r / min. Take 2 mL samples at 0.5 h, 1 h, and 2 h, respectively, and immediately add an equal volume of simulated gastric juice at the same temperature after sampling. Centrifuge the collected samples at 10000 r / min for 10 min, collect the supernatant, and determine the content of carboxymethyl active Poria cocos polysaccharide released into the supernatant using the phenol-sulfuric acid method, and calculate the cumulative release rate in the simulated gastric juice stage.

[0033] After 2 h of treatment with simulated gastric fluid, the remaining sample was centrifuged to collect the precipitate. The precipitate was gently washed once with a small amount of purified water and then transferred to 50 mL of simulated intestinal fluid (pH 6.8 phosphate buffer). The sample was then placed in a 37°C constant-temperature shaker at 100 r / min. Samples of 2 mL were taken at 4 h, 6 h, and 8 h, and an equal volume of simulated intestinal fluid at the same temperature was added after each sample collection. The sample solution was centrifuged at 10000 r / min for 10 min, and the supernatant was used to determine the content of carboxymethyl active Poria cocos polysaccharide. The stage and total cumulative release rate of the simulated intestinal fluid were calculated.

[0034] The cumulative release rate is calculated using the following formula: Cumulative release rate (%) = (cumulative release of carboxymethyl active poria polysaccharide at each time point) / (total release of carboxymethyl active poria polysaccharide in the sample) × 100%.

[0035] Each group of samples was tested in parallel three times, and the average value of the results was taken.

[0036] Table 2. Results of cumulative release rate tests for simulated gastric and intestinal fluids.

[0037] 4. Rehydration and Dispersion Stability Test The dried samples obtained in Examples 1-3 and Comparative Examples 1-3 were rehydrated and dispersed in purified water, with the concentration of carboxymethyl active Poria cocos polysaccharide in the samples controlled at 1.0 mg / mL. The rehydrated solution was placed at 25℃ and allowed to stand. The samples were observed at 0 h, 6 h, 24 h, and 48 h to determine whether obvious sedimentation, flocculation, or stratification occurred, and the average particle size and PDI of the samples were measured at each time point. Another sample after 24 h of rehydration was centrifuged at 3000 r / min for 5 min. The samples were observed to determine whether obvious precipitation occurred before and after centrifugation, and the change in transmittance of the supernatant at 600 nm before and after centrifugation was measured.

[0038] Stability retention rate is calculated using the following formula: Stability retention rate (%) = Absorbance of supernatant at 600 nm after centrifugation / Absorbance of dispersion at 600 nm before centrifugation × 100%.

[0039] Each group of samples was tested in parallel three times, and the average value of the results was taken.

[0040] 5. Antioxidant synergistic performance test The antioxidant capacity of the samples obtained in Examples 1-3 and Comparative Examples 1-3 was evaluated using a DPPH free radical scavenging assay. Each group of samples was rehydrated and dispersed in purified water, and then uniformly diluted to 0.5 mg / mL according to the content of carboxymethyl active Poria cocos polysaccharide. Two mL of the sample solution was mixed with two mL of 0.1 mmol / L DPPH ethanol solution, and the mixture was reacted in the dark for 30 min. The absorbance was then measured at 517 nm. Purified water was used as a blank control instead of the sample, and a DPPH-free system was used for background correction. Each group of samples was measured in triplicate.

[0041] DPPH free radical scavenging rate is calculated using the following formula: DPPH free radical scavenging rate (%) = [1 - (A sample - A background) / A blank] × 100%.

[0042] Table 3. Test results of rehydration dispersion stability and antioxidant properties

[0043] As shown in Tables 1 to 3, all three examples formed relatively stable composite microgel particles after rehydration, with an average particle size of 438.7–526.4 nm, a PDI of 0.218–0.255, a narrow particle size distribution, and an encapsulation rate of carboxymethyl active Poria cocos polysaccharide of 84.6%–88.9%, all exceeding 80%. Meanwhile, the cumulative release rate of Examples 1–3 in simulated gastric fluid after 2 hours was only 20.2%–24.7%, indicating that it can effectively reduce burst release under acidic conditions. In simulated intestinal fluid, the cumulative release rate reached 72.4%–82.3% after 8 hours, demonstrating good sustained release performance. Regarding rehydration stability, all three examples showed no significant sedimentation after 48 hours, with a particle size change rate of less than 10% and a stability retention rate of 92.5%–95.1%. In terms of antioxidant performance, the DPPH free radical scavenging rate was 70.8%–80.2%.

[0044] Compared to Example 1, Comparative Example 1, lacking the addition of chitosan oligosaccharide, lacked the electrostatic association structure between carboxymethyl active poria cocos polysaccharide and chitosan oligosaccharide, resulting in an increased average particle size of 1286.5 nm, a PDI increase of 0.612, a decrease in encapsulation efficiency to 54.8%, and a cumulative release rate of 61.8% in simulated gastric juice over 2 hours. Significant flocculation and sedimentation were observed after 48 hours of rehydration, indicating that relying solely on carboxymethyl active poria cocos polysaccharide, tea polyphenols, and calcium phytate is insufficient to form a stable microgel system. Comparative Example 2, lacking the addition of sodium phytate and calcium lactate, lacked the ion bridging enhancement effect of calcium phytate, resulting in an encapsulation efficiency of only 73.5%. The cumulative release rate of simulated gastric juice over 2 hours increased to 47.9%, and the particle size change rate reached 31.8% over 48 hours, indicating that the unbridged composite microparticle structure was relatively loose, with insufficient resistance to acid release and rehydration stability. Comparative Example 3 involved the pre-mixing and simultaneous addition of calcium phytate and calcium lactate, resulting in uneven local bridging. The average particle size was 918.2 nm, the PDI was 0.476, the encapsulation rate was 70.3%, and the cumulative release rate of simulated gastric juice over 2 hours was 42.5%. Significant flocculation was observed after rehydration, indicating that the stepwise bridging sequence of phytate adsorption followed by calcium salt addition plays a crucial role in forming a uniform, stable, and sustained-release composite microgel structure.

[0045] The above description is only a preferred embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any equivalent substitutions or modifications made by those skilled in the art within the scope of the technology disclosed in the present invention, based on the technical solution and inventive concept of the present invention, should be covered within the scope of protection of the present invention.

Claims

1. A method for preparing a sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition, characterized in that, Includes the following steps: S1. Dissolve carboxymethyl active Poria cocos polysaccharide in water and adjust the pH to 6.0-7.2 to obtain a pre-activated polysaccharide solution; combine chitosan oligosaccharide and tea polyphenols under pH 4.0-5.5 conditions to obtain a chitosan oligosaccharide-polyphenol pre-complex solution; S2. The pre-activated polysaccharide solution is added to the chitosan oligosaccharide-tea polyphenol pre-composite solution, and electrostatic association is carried out under the condition of pH 5.0-6.2 to obtain a composite microparticle dispersion. Then, phytate component is added to the composite microparticle dispersion first, and then calcium salt component is added dropwise. Ion bridging aging is carried out under the condition of pH 5.8-6.8 to obtain a composite microgel dispersion. The mass ratio of carboxymethyl active poria cocos polysaccharide to chitosan oligosaccharide is 100:(15-35), and the mass ratio of phytate component to calcium salt component is 1:(0.5-2.0). S3. Add a drying protection component to the composite microgel dispersion, mix and dry to obtain a sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition that can form composite microgel particles after rehydration.

2. The method for preparing the sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition according to claim 1, characterized in that, By weight, the carboxymethyl active Poria cocos polysaccharide is 40-70 parts, the chitosan oligosaccharide is 8-20 parts, the tea polyphenols are 0.5-5 parts, the phytate component is 1-6 parts, the calcium salt component is 1-5 parts, and the drying protection component is 10-30 parts.

3. The method for preparing the sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition according to claim 1, characterized in that, The carboxymethyl active Poria cocos polysaccharide has a carboxymethyl substitution degree of 0.25-0.58, a weight-average molecular weight of 50-220 kDa, and a total sugar content of not less than 70%.

4. The method for preparing the sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition according to claim 1, characterized in that, In step S1, the mass concentration of carboxymethyl active Poria cocos polysaccharide in the pre-activated polysaccharide solution is 0.5%–3.0%, the pre-activation temperature is 35–55℃, and the pre-activation time is 20–60 min.

5. The method for preparing the sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition according to claim 1, characterized in that, The degree of deacetylation of the chitosan oligosaccharide is not less than 85%, and the molecular weight is 1-10 kDa; the compounding time of the chitosan oligosaccharide and tea polyphenols is 15-45 min.

6. The method for preparing the sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition according to claim 1, characterized in that, In step S2, the pre-activated polysaccharide solution is added dropwise to the chitosan oligosaccharide-tea polyphenol pre-complex solution for 10–40 min, the electrostatic association stirring speed is 300–1000 rpm, and the electrostatic association time is 20–60 min.

7. The method for preparing the sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition according to claim 1, characterized in that, The phytate component is one or two of sodium phytate and potassium phytate; the calcium salt component is one or more of calcium lactate, calcium gluconate and calcium chloride; the ion bridging aging temperature is 20-40℃ and the time is 30-120 min.

8. The method for preparing the sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition according to claim 1, characterized in that, The drying and protective components are one or more of trehalose, mannitol, maltodextrin, and fructooligosaccharides; the drying is performed by freeze drying or spray drying.

9. The method for preparing the sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition according to claim 1, characterized in that, When freeze-drying is used, the pre-freezing temperature is -40 to -20°C, the pre-freezing time is 4 to 12 hours, and the vacuum drying time is 18 to 36 hours; when spray drying is used, the inlet air temperature is 120 to 160°C, and the outlet air temperature is 60 to 85°C.

10. The sustained-release synergistic carboxymethyl active Poria cocos polysaccharide composition prepared by the preparation method according to any one of claims 1-9, characterized in that, The composition, after rehydration, forms composite microgel particles with an average particle size of 200–900 nm, a PDI of no more than 0.35, and an encapsulation rate of carboxymethyl active Poria cocos polysaccharide of no less than 80%. The cumulative release rate in simulated gastric juice is no more than 30% in 2 hours and in simulated intestinal juice is 65%–90% in 8 hours.

Images