Preparation method of OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid

Abstract

The invention relates to a preparation method of OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid, which comprises the following steps: S1, carrying out esterification reaction on porous starch and octenyl succinic anhydride to obtain OSA starch; s2, dissolving sodium butyrate and zein in an alcoholic solution, and stirring for embedding to obtain zein-sodium butyrate; s3, OSA starch and zein-sodium butyrate are subjected to a reaction, and a compound is obtained; s4, dispersing the compound in a sodium alginate solution, dropwise adding into a calcium chloride solution for crosslinking to obtain OSA starch-polysaccharide gel microspheres, soaking the OSA starch-polysaccharide gel microspheres in a chitosan solution with a first concentration for primary coating, and soaking the OSA starch-polysaccharide gel microspheres in a chitosan solution with a second concentration for secondary coating; the first concentration is lower than the second concentration, and the difference between the first concentration and the second concentration is not lower than 1wt%. By constructing a core-carrier-interpenetrating network shell structure, the problems of low encapsulation efficiency, poor targeting property and weak stability in butyric acid delivery are solved.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of biomedical materials and drug delivery technology, and particularly relates to a preparation method of OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid. BACKGROUND

[0002] Butyric acid, as an important short-chain fatty acid, plays a key role in maintaining intestinal barrier function, inhibiting inflammation and preventing colon cancer. However, butyric acid has a small molecular weight and is volatile, and is easily absorbed in the upper digestive tract, making it difficult to effectively reach and act on the colon site, and its bioavailability is extremely low.

[0003] At present, the common carrier system for butyric acid delivery mainly has the following defects: first, microencapsulation technology, such as liposomes or polymer microspheres, is difficult to form efficient and stable interaction with hydrophobic butyric acid, resulting in low encapsulation efficiency of butyric acid; second, lipid-based carriers (such as emulsions, solid lipid nanoparticles) are prone to phase separation during long-term storage, and have insufficient stability; third, inorganic porous materials (such as mesoporous silica) have poor biocompatibility and degradability, and have potential safety risks; fourth, natural polysaccharide gels (such as sodium alginate alone) have low encapsulation efficiency due to the difficulty of hydrophilic carriers to load hydrophobic butyric acid, and their structure is relatively loose, relying only on a single ionic cross-linking response mechanism, which is easily dissolved in the complex gastrointestinal environment (such as gastric acid pH, digestive enzyme action), resulting in premature release of butyric acid and failure to achieve colon targeting; fifth, physical mixing adsorption (such as direct adsorption of butyric acid on starch) has low butyric acid loading due to the limited specific surface area of starch and the lack of effective binding sites, and has almost no protection for butyric acid, with serious burst release.

[0004] In summary, the existing butyric acid delivery system generally has low encapsulation efficiency, insufficient targeting, and poor stability. Therefore, there is an urgent need in the art for a new delivery system that can simultaneously achieve high encapsulation efficiency, good stability, and precise control of butyric acid release in the colon. SUMMARY

[0005] To solve the above technical problems, the present application provides a preparation method of OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid.

[0006] The technical scheme adopted by the present application is as follows:

[0007] The present application provides a preparation method of OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid, characterized by comprising the following steps:

[0008] S1, esterification of porous starch with octenyl succinic anhydride, and then neutralization, washing and freeze-drying of the reaction product to obtain OSA starch;

[0009] S2, sodium butyrate and zein are dissolved in an alcohol solution, stirring to embed, to obtain zein-sodium butyrate;

[0010] S3, the OSA starch and the zein-sodium butyrate are reacted in a buffer solution to obtain a complex;

[0011] S4, the complex is dispersed in a sodium alginate solution, then added dropwise into a calcium chloride solution for crosslinking, solidified, washed to obtain OSA starch-polysaccharide gel microspheres, then immersed in a first concentration of chitosan solution for primary coating, and immersed in a second concentration of chitosan solution for secondary coating to obtain OSA starch-polysaccharide interpenetrating network microspheres;

[0012] The first concentration is lower than the second concentration, and the first concentration and the second concentration differ by no less than 1 wt%.

[0013] The preparation method of the OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid proposed in the application constructs a three-layer structure of a hydrophobic inner core, an amphiphilic carrier and a pH-responsive interpenetrating network shell from inside to outside, and solves the technical problems of low encapsulation efficiency, poor targeting and weak stability in butyric acid delivery. Specifically, first, the hydrophobicity of zein is used to form an inner core by emulsifying and crosslinking sodium butyrate, to construct a chemical barrier and effectively prevent butyric acid (the form of sodium butyrate acting in the body is butyric acid) from being released and degraded in advance in the stomach acid environment. Second, the porous starch is modified by grafting octenyl succinic anhydride (OSA) to introduce hydrophilic carboxyl groups and hydrophobic long chains on the molecular chain of the porous starch at the same time, to form OSA starch with an amphiphilic structure; the carrier is tightly combined with the inner core through hydrophobic interaction (the hydrophobic octenyl chain of OSA can be combined with the hydrophobic alkyl chain of butyric acid), to encapsulate butyric acid inside the OSA starch, and the hydrophilic carboxyl groups on the outside of the carrier build a hydrophilic shell, to isolate adverse environmental factors such as light and oxygen and significantly enhance the stability. Third, a gradient concentration of chitosan and sodium alginate are used to construct an interpenetrating network through electrostatic interaction, and the structure has pH responsiveness and a structure gradient: in the stomach juice (pH 1-3), the sodium alginate gel network shrinks, and the dense inner layer formed by the low-concentration chitosan cooperatively realizes strong protection and blocks the invasion of gastric acid; after entering the intestinal juice (pH 7-8), the sodium alginate gel network swells, and the loose outer layer formed by the high-concentration chitosan gradually disintegrates under the regulation of the colon environment, to trigger the precise controlled release of butyric acid and realize colon targeting. In addition, the interpenetrating network structure as a whole enhances the mechanical strength of the microspheres, and can effectively resist the physical shear force caused by gastrointestinal peristalsis.

[0014] Preferably, in step S1, the preparation method of the porous starch specifically comprises: preparing starch milk with a concentration of 18-25wt%, adding liquid enzyme with a concentration of 80-200U / mL, and enzymatically hydrolyzing at 37°C for 6-24h to obtain the porous starch.

[0015] The liquid enzyme is composed of α-amylase and α-amylase glucosidase.

[0016] Preferably, in step S1, the mass ratio of the porous starch to the octenyl succinic anhydride is 10-20:1.

[0017] Preferably, in step S1, the esterification conditions are as follows: the esterification temperature is 20-40°C, the esterification time is 2-6h, and the system pH is 8-10.

[0018] Preferably, in step S2, the mass ratio of the sodium butyrate to the zein is 1:3-5.

[0019] Preferably, in step S2, the stirring temperature is 30-40°C, the stirring time is 1-2h, and the system pH is 3.

[0020] Preferably, in step S3, the reaction temperature is 45-55°C, the reaction time is 3-5h, and the pH of the buffer solution is 7-8.

[0021] Preferably, in step S4, the concentration of the sodium alginate solution is 1.5-2.5wt%, the concentration of the calcium chloride solution is 1.5-2wt%, and the mass ratio of the complex to the sodium alginate solution is 1:1-3.

[0022] The dropping speed is 1 drop / s, and the solidification time is 25-35min.

[0023] Preferably, in step S4, the first concentration is 0.5-0.8wt%, and the first coating time is 15-25min.

[0024] Preferably, in step S4, the second concentration is 2-2.5wt%, and the second coating time is 5-15min. BRIEF DESCRIPTION OF DRAWINGS

[0025] Figure 1 Figure 1 is a schematic diagram of the preparation process of OSA starch-polymer interpenetrating network microspheres of Example 1, wherein NPS is raw potato starch, PRS is porous potato starch, OSAS is OSA starch (i.e. porous potato starch grafted with OSA), NaB is sodium butyrate, and OSAS-NaB is OSA starch-polymer interpenetrating network microspheres (OSAS loaded with zein-sodium butyrate, which is finally obtained by cross-linking the sodium alginate and chitosan);

[0026] Figure 2 Performance test chart of Example 1, A is FT-IR chart, B is butyric acid encapsulation rate, C is SEM chart of OSAS-NaB, D is gastrointestinal release rate of butyric acid, wherein NPS-NaB is original potato starch loaded sodium butyrate, PRS-NaB is porous potato starch loaded sodium butyrate, and the rest of NPS, PRS, OSAS, OSAS-NaB have the same meaning as Figure 1 . DETAILED DESCRIPTION

[0027] In order to better understand the above technical solutions, the exemplary embodiments of the present application will be described in more detail below with reference to the accompanying drawings. Although the exemplary embodiments of the present application are shown in the drawings, it should be understood that the present application can be implemented in various forms and should not be limited by the embodiments described herein. On the contrary, these embodiments are provided so that the present application can be more clearly, thoroughly understood, and the scope of the present application can be completely conveyed to those skilled in the art.

[0028] The present application provides a preparation method of OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid, characterized in that it comprises the following steps:

[0029] S1, esterification reaction of porous starch and octenyl succinic anhydride, and then neutralization, washing and freeze-drying of the reaction product to obtain OSA starch;

[0030] S2, dissolving sodium butyrate and zein in an alcohol solution, stirring to embed, to obtain zein-sodium butyrate;

[0031] S3, reacting OSA starch and zein-sodium butyrate in a buffer solution to obtain a complex;

[0032] S4, dispersing the complex in a sodium alginate solution, then adding dropwise into a calcium chloride solution for crosslinking, solidifying, washing to obtain OSA starch-polysaccharide gel microspheres, then soaking in a first concentration of chitosan solution for primary coating, and then soaking in a second concentration of chitosan solution for secondary coating to obtain OSA starch-polysaccharide interpenetrating network microspheres;

[0033] The first concentration is lower than the second concentration, and the difference between the first concentration and the second concentration is not less than 1wt%.

[0034] In the present application, based on its unique "core-carrier-shell" three-layer structure, through the synergistic effect of pH response and enzyme triggering in different segments of the digestive tract environment, the multi-level release of "stomach protection, small intestine transition, colon release" is realized, and the efficient targeted delivery of butyric acid is realized. Specifically: (1) In the stomach environment, the carboxyl group of sodium alginate in the shell is protonated in a strong acid environment, causing the entire gel network to shrink dramatically, and the pores close. At the same time, the dense inner layer formed by low-concentration chitosan is also in a protonated state and is tightly combined with the shrinking sodium alginate network through electrostatic interaction to form a dense barrier that effectively blocks gastric acid invasion. (2) In the small intestine environment, as the environmental pH rises to neutral, the carboxyl group of sodium alginate ionizes, and the originally contracted gel network begins to swell fully, opening the internal pores and forming a preliminary release channel. However, the overall structure of the shell is relatively complete at this time, thereby avoiding burst release. (3) In the colon environment, the interpenetrating network further disintegrates, and amylase begins to attack and degrade OSA starch, triggering the release of the core, which is further degraded by proteases in the colon, thereby releasing the sodium butyrate encapsulated therein. Sodium butyrate dissociates in the colon microenvironment to form biologically active butyric acid molecules, which directly act on colon mucosal cells to exert their physiological functions. The OSA starch-polymer interpenetrating network microspheres prepared by the present application can be applied in the fields of food additives, feed, medicine, etc., and are particularly useful for the treatment or prevention of intestinal diseases such as colitis.

[0035] In a preferred embodiment of the present application, in step S1, the preparation method of the porous starch specifically comprises: preparing starch milk with a concentration of 18-25wt%, and adding liquid enzyme with a concentration of 80-200U / mL for enzymatic hydrolysis at 37℃ for 6-24h.

[0036] The liquid enzyme is composed of alpha-amylase and alpha-amylase glucosidase.

[0037] In the present application, the starch is pretreated by complex enzymatic hydrolysis to form starch with high specific surface area and porous structure. On the one hand, the significantly increased specific surface area increases the physical adsorption capacity of sodium butyrate, laying a foundation for achieving high drug loading. On the other hand, the porous structure exposes more hydroxyl groups in the reaction environment, thereby significantly improving the grafting efficiency of the subsequent esterification reaction with OSA. In addition, the present application does not limit the ratio between alpha-amylase and alpha-amylase glucosidase in the liquid enzyme.

[0038] In one preferred embodiment of the present application, in step S1, the mass ratio of porous starch to octenyl succinic anhydride is 10-20:1. In the present application, the mass ratio of porous starch to OSA is defined to regulate the grafting degree of OSA, to ensure the balance of the amphiphilicity of OSA starch, and to further ensure that the subsequent stable and uniform compound of OSA starch and the inner core of zein-sodium butyrate is formed through hydrophobic interaction. For example, the mass ratio of porous starch to OSA can be 10:1, 12:1, 14:1, 16:1, 18:1, 20:1, etc., without particular limitation.

[0039] In one preferred embodiment of the present application, in step S1, the esterification conditions are as follows: the esterification temperature is 20-40℃, the esterification time is 2-6h, and the pH of the system is 8-10. In the present application, controlling the esterification conditions is conducive to ensuring the grafting efficiency of OSA starch. For example, the esterification conditions include, but are not limited to, 25℃×2h, pH=9, 30℃×4h, pH=10, 35℃×6h, pH=8, etc., wherein NaOH solution can be used to adjust the pH of the system.

[0040] In one preferred embodiment of the present application, in step S2, the mass ratio of sodium butyrate to zein is 1:3-5. In the present application, under this ratio, zein can provide an appropriate amount of hydrophobic wrapping sites, so as to coat sodium butyrate molecules and form a compact and uniformly stable inner core, which not only avoids incomplete embedding due to insufficient zein, but also prevents excessive compact structure due to excessive zein, thereby laying a foundation for the subsequent efficient and stable combination with OSA starch and even the construction of a complete interpenetrating network system. For example, the mass ratio of sodium butyrate to zein can be any value in 1:3, 1:3.5, 1:4, 1:4.5, 1:5, etc.

[0041] In one preferred embodiment of the present application, in step S2, the stirring temperature is 30-40℃, the stirring time is 1-2h, and the pH of the system is 3. In the present application, the embedding reaction is carried out under specific stirring conditions to ensure that zein fully and uniformly embeds sodium butyrate. For example, the stirring conditions can be 30℃×2h, 32℃×1.8h, 35℃×1.5h, 40℃×1h, etc.

[0042] In one preferred embodiment of the present application, in step S3, the reaction temperature is 45-55℃, the reaction time is 3-5h, and the pH of the buffer solution is 7-8. In the present application, the reaction conditions of OSA starch and zein-sodium butyrate are controlled to promote the hydrophobic interaction between them (the hydrophobic octenyl chain in OSA starch binds with the hydrophobic interface of the core of zein-sodium butyrate), so as to anchor the core in the porous pores of OSA starch and form a stable complex. For example, the reaction conditions include, but are not limited to, 45℃×3h, pH=7, 50℃×3.5h, pH=8, 55℃×5h, pH=7.5, etc.

[0043] In one preferred embodiment of the present application, in step S4, the concentration of the sodium alginate solution is 1.5-2.5wt%, the concentration of the calcium chloride solution is 1.5-2wt%, and the mass ratio of the complex to the sodium alginate solution is 1:1-3.

[0044] The dropping speed is 1 drop / s, and the solidification time is 25-35min.

[0045] In the present application, the concentrations of the sodium alginate solution and the calcium chloride solution are limited to ensure sufficient ionic crosslinking; the mass ratio of the complex to the sodium alginate solution is limited to ensure uniform dispersion of the complex and improve the uniformity of subsequent release; and the dropping speed and the solidification time are limited to precisely control the process of forming the microspheres and obtain microspheres with uniform particle size and complete crosslinking and solidification.

[0046] In one preferred embodiment of the present application, in step S4, the first concentration is 0.5-0.8wt%, and the time for one-time coating is 15-25min. In the present application, the OSA starch-polymer gel microspheres are first coated with a low-concentration chitosan solution of the first concentration, and the purpose is to form a dense inner layer that is tightly combined with sodium alginate through electrostatic interaction. When the microspheres enter the strong acidic environment of the stomach, the inner layer structure can cooperate with the contracted sodium alginate gel network to build a strong physical and chemical barrier, thereby achieving efficient protection of the core. For example, the first concentration can be 0.5wt%, 0.6wt%, 0.7wt%, 0.8wt%, etc., and the time for one-time coating can be 15min, 18min, 20min, 22min, 25min, etc., without particular limitation.

[0047] In one preferred embodiment of the present application, in step S4, the second concentration is 2-2.5 wt%, and the time for the secondary coating is 5-15 min. In the present application, the secondary coating is performed using a high-concentration chitosan solution of the second concentration, and the purpose is to construct a loose and porous outer layer structure. In the colon environment, the outer layer can be efficiently degraded, cooperating with the internal swollen sodium alginate gel, to accurately trigger the rapid and complete release of sodium butyrate, and finally achieve efficient colon-targeted delivery. For example, the second concentration can be 2.1 wt%, 2.2 wt%, 2.3 wt%, 2.4 wt%, 2.5 wt%, etc., and the time for the secondary coating can be 5 min, 8 min, 10 min, 12 min, 15 min, etc., all without particular limitation.

[0048] Example 1

[0049] Reference Figure 1 A preparation method of OSA starch-polymer interpenetrating network microspheres for targeted delivery of butyric acid, characterized in that it comprises the following steps:

[0050] S1, mixing porous starch and octenyl succinic anhydride at a mass ratio of 10:1, and performing esterification reaction at a temperature of 35℃ and a system pH of 8.5 for 2h, continuously stirring during the reaction, and then sequentially neutralizing, washing, and freeze-drying the reaction product to obtain OSA starch, which is denoted as OSAS.

[0051] The preparation method of the porous starch specifically comprises: preparing potato starch into a starch milk with a concentration of 25 wt%, adding liquid enzyme (composed of α-amylase and α-amylase glucosidase) with a concentration of 200 U / mL, and performing enzymatic hydrolysis at 37℃ for 8h, and then washing and drying.

[0052] S2, dissolving sodium butyrate and zein in an ethanol solution with a volume fraction of 75% at a mass ratio of 1:3, embedding at a temperature of 35℃ and a system pH of 3 for 2h, removing ethanol by rotary evaporation, and then dispersing in water with a pH of 3 to obtain zein-sodium butyrate.

[0053] S3, reacting OSAS and zein-sodium butyrate in a potassium dihydrogen phosphate buffer solution with a pH of 7.2 at 45℃ for 3h to obtain a complex.

[0054] S4, the complex was dispersed in a 1.5wt% sodium alginate solution (the mass ratio of the complex to the sodium alginate solution was 1:2), and then was added dropwise to a 1.5wt% calcium chloride solution at a rate of 1 drop per second for cross-linking, and was solidified for 25 min, washed, to obtain OSA starch-polymer gel microspheres, and then was immersed in a 0.5wt% chitosan solution for primary coating for 20 min, and was immersed in a 2wt% chitosan solution for secondary coating for 10 min, to obtain OSA starch-polymer interpenetrating network microspheres, which were denoted as OSAS-NaB.

[0055] Reference should be made to Figure 2 A, the OSA starch obtained in step S1 was subjected to FT-IR detection, which proved that the OSA groups were successfully grafted onto the starch molecules, and the degree of substitution was 12.0±0.1×10 -3 ; it can be seen from Figure 2 C that the OSAS-NaB obtained in step S4 was observed by SEM, and it was found that a relatively uniform product structure was formed.

[0056] Example 2

[0057] A preparation method of OSA starch-polymer interpenetrating network microspheres for targeted delivery of butyric acid, characterized in that it comprises the following steps:

[0058] S1, porous starch was mixed with octenyl succinic anhydride at a mass ratio of 18:1, and esterification reaction was carried out at a temperature of 25℃ and a pH of 9.5 for 3h, and the reaction product was continuously stirred during the reaction, and then was sequentially neutralized with dilute hydrochloric acid, washed and freeze-dried, to obtain OSA starch, which was denoted as OSAS.

[0059] The preparation method of the porous starch specifically comprises the following steps: potato starch was prepared into a starch milk with a concentration of 18wt%, and liquid enzyme (consisting of α-amylase and α-amylase glucosidase) with a concentration of 120U / mL was added, and the mixture was enzymatically hydrolyzed at 37℃ for 24h, and then was washed and dried.

[0060] S2, sodium butyrate and zein were dissolved in an ethanol solution with a volume fraction of 75% at a mass ratio of 1:4, and embedding was carried out at a temperature of 38℃ and a pH of 3 for 1.8h, and then ethanol was removed by rotary evaporation, and the mixture was dispersed into water with a pH of 3, to obtain zein-sodium butyrate.

[0061] S3, OSA starch and zein-sodium butyrate were reacted in a potassium dihydrogen phosphate buffer solution with a pH of 7.8 at 52℃ for 4.5h, to obtain a complex.

[0062] S4, the complex was dispersed in a 2.5wt% sodium alginate solution (mass ratio of the complex to the sodium alginate solution was 1:1), and then was added dropwise to a 2wt% calcium chloride solution at a rate of 1 drop per second for crosslinking, and was solidified for 35 min, washed, to obtain OSA starch-polymer gel microspheres, and then was immersed in a 0.8wt% chitosan solution for primary coating for 20 min, and was immersed in a 2.2wt% chitosan solution for secondary coating for 10 min, to obtain OSA starch-polymer interpenetrating network microspheres, which were denoted as OSAS-NaB.

[0063] In this embodiment, the OSA starch obtained in step S1 was subjected to FT-IR detection, which confirmed that the OSA groups were successfully grafted onto the starch molecules, and the degree of substitution was 11.8±0.2×10 -3 The OSAS-NaB obtained in step S4 was subjected to SEM observation, and it was found that the structure thereof was relatively uniform.

[0064] Example 3

[0065] A preparation method of OSA starch-polymer interpenetrating network microspheres for targeted delivery of butyric acid, characterized in that the method comprises the following steps:

[0066] S1, porous starch was mixed with octenyl succinic anhydride at a mass ratio of 14:1, and esterification reaction was carried out at a temperature of 38℃ and a pH of 10 for 2.5h, and stirring was continuously performed during the reaction, and then the reaction product was sequentially subjected to neutralization with dilute hydrochloric acid, washing and freeze-drying, to obtain OSA starch, which was denoted as OSAS.

[0067] The preparation method of the porous starch specifically comprises the following steps: potato starch was prepared into a starch milk with a concentration of 22wt%, and liquid enzyme (consisting of α-amylase and α-amylase glucosidase) with a concentration of 180U / mL was added, and enzyme hydrolysis was carried out at 37℃ for 12h, and then washing and drying were performed.

[0068] S2, sodium butyrate and zein were dissolved in an ethanol solution with a volume fraction of 75% at a mass ratio of 1:5, embedding was carried out at a temperature of 33℃ and a pH of 3 for 2h, ethanol was removed by rotary evaporation, and then the mixture was dispersed in water with a pH of 3, to obtain zein-sodium butyrate.

[0069] S3, OSA starch and zein-sodium butyrate were reacted in a potassium dihydrogen phosphate buffer solution with a pH of 7.6 at 50℃ for 4h, to obtain a complex.

[0070] S4, the complex was dispersed in a 2.5wt% sodium alginate solution (the mass ratio of the complex to the sodium alginate solution was 1:3), and then was added dropwise to a 1.5wt% calcium chloride solution at a rate of 1 drop per second for cross-linking, and was solidified for 20 min, washed, to obtain OSA starch-polymer gel microspheres, which were then immersed in a 0.5wt% chitosan solution for primary coating for 20 min, and then were immersed in a 2.5wt% chitosan solution for secondary coating for 10 min, to obtain OSA starch-polymer interpenetrating network microspheres, which were denoted as OSAS-NaB.

[0071] In this example, the OSA starch obtained in step S1 was subjected to FT-IR detection, which confirmed that the OSA groups were successfully grafted to the starch molecules, and the degree of substitution was 12.0±0.5×10 -3 The OSAS-NaB obtained in step S4 was subjected to SEM observation, and it was found that the structure thereof was relatively uniform.

[0072] Comparative Example 1

[0073] The difference between this comparative example and Example 1 was that, in step S4, a mixed solution of a sodium alginate solution, a calcium chloride solution and a chitosan solution was directly used to encapsulate the complex, and the specific steps included the following: 1.5wt% of a sodium alginate solution, 1.5wt% of a calcium chloride solution and 2.5wt% of a chitosan solution were mixed uniformly to obtain a mixed solution, and the complex obtained in step S3 was dispersed in the mixed solution and reacted for 1 h. The remaining steps were unchanged.

[0074] Comparative Example 2

[0075] The difference between this comparative example and Example 1 was that, in step S4, a single low-concentration chitosan solution was used, and the “immersion in a 0.5wt% chitosan solution for primary coating for 20 min, and then immersion in a 2wt% chitosan solution for secondary coating for 10 min” was adjusted to “immersion in a 0.5wt% chitosan solution for coating for 30 min”. The remaining steps were unchanged.

[0076] Comparative Example 3

[0077] The difference between this comparative example and Example 1 was that, in step S4, a single high-concentration chitosan solution was used, and the “immersion in a 0.5wt% chitosan solution for primary coating for 20 min, and then immersion in a 2wt% chitosan solution for secondary coating for 10 min” was adjusted to “immersion in a 2wt% chitosan solution for coating for 30 min”. The remaining steps were unchanged.

[0078] Comparative Example 4

[0079] The present comparative example differs from Example 1 in that step S2 is omitted, and no zein is used to embed sodium butyrate (i.e. sodium butyrate is directly reacted with OSA starch to obtain the complex). The remaining steps remain unchanged.

[0080] Comparative Example 5

[0081] The present comparative example differs from Example 1 in that step S2 is omitted, and step S4 uses a single concentration of chitosan solution. Specifically, the following steps are included: OSA starch is prepared according to step S1 of Example 1, sodium butyrate is reacted with OSA starch in a potassium dihydrogen phosphate buffer solution having a pH of 7.2 at 45°C for 3h to obtain a complex; the complex is dispersed in a 1.5wt% sodium alginate solution (the mass ratio of the complex to the sodium alginate solution is 1:2), and is then added dropwise to a 1.5wt% calcium chloride solution at a rate of 1 drop per second for crosslinking, and is solidified for 25min, washed, and finally the obtained gel microspheres are immersed in a 2.5wt% chitosan solution for coating for 30min.

[0082] The microspheres obtained in Examples 1-3 and Comparative Examples 1-5 are subjected to performance testing, and the results are recorded in Table 1 and Figure 2 .

[0083] Table 1

[0084]

[0085]

[0086] As can be seen from Table 1 and Figure 2 , the present application can achieve (1) an encapsulation rate of ≥38.6% (traditional starch carrier <10%); (2) a butyric acid release rate of <5% in simulated gastric fluid for 0.5h, and a butyric acid release rate of <30% in simulated intestinal fluid for 3h; (3) a butyric acid retention rate of >90% after storage for 2-3 months, and no obvious deterioration or performance decline. Comparative Example 1 directly uses a mixed solution of sodium alginate solution, calcium chloride solution and chitosan solution for encapsulation, and the gel structure formed is disordered, loose and unstable, and cannot achieve effective encapsulation, nor can it effectively resist gastric acid and intestinal fluid; Comparative Example 2 has a faster release rate in the intestinal tract due to the lack of a loose outer layer formed by high-concentration chitosan; Comparative Example 3 has a faster release rate in the stomach due to the lack of a dense inner layer formed by low-concentration chitosan, and the structure formed by high-concentration chitosan is not dense enough to form a strong barrier with sodium alginate; Comparative Example 4 has a lower encapsulation rate and a faster release of sodium butyrate in the gastrointestinal tract due to the lack of hydrophobic encapsulation by zein, on the one hand, and the lack of a hydrophobic barrier, on the other hand; and Comparative Example 5 has a significantly lower encapsulation rate and a faster burst release of sodium butyrate in the gastrointestinal tract due to the lack of encapsulation of sodium butyrate by zein and the lack of a gradient structure of the chitosan layer.

[0087] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present application, and are not intended to limit the present application; although the present application has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that the technical solutions described in the foregoing embodiments can still be modified, or some or all of the technical features can be replaced by equivalents; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the scope of the technical solutions of the embodiments of the present application.

Claims

1. A method for preparing OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid, characterized in that, Includes the following steps: S1. Porous starch is esterified with octenyl succinic anhydride, and the reaction product is then neutralized, washed and freeze-dried to obtain OSA starch. S2. Sodium butyrate and zein are dissolved in an alcohol solution and stirred to encapsulate the mixture, thus obtaining zein-sodium butyrate. S3. The OSA starch is reacted with the zein-sodium butyrate in a buffer solution to obtain a complex; S4. The complex is dispersed in sodium alginate solution and then added dropwise to calcium chloride solution for cross-linking. After curing and washing, OSA starch-polysaccharide gel microspheres are obtained. Then, they are immersed in a chitosan solution of a first concentration for primary coating and then immersed in a chitosan solution of a second concentration for secondary coating to obtain OSA starch-polysaccharide interpenetrating network microspheres. The first concentration is lower than the second concentration, and the difference between the first concentration and the second concentration is not less than 1 wt%.

2. The method for preparing OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid as described in claim 1, characterized in that, In step S1, the method for preparing the porous starch specifically includes: preparing starch into a starch milk with a concentration of 18-25 wt%, adding liquid enzyme with a concentration of 80-200 U / mL, and enzymatically hydrolyzing at 37°C for 6-24 hours to obtain the product; The liquid enzyme consists of α-amylase and α-amyl glucosidase.

3. The method for preparing OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid as described in claim 1, characterized in that, In step S1, the mass ratio of the porous starch to the octenyl succinic anhydride is 10-20:

1.

4. The method for preparing OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid as described in claim 1, characterized in that, In step S1, the esterification reaction conditions are: esterification temperature of 20℃~40℃, esterification time of 2~6h, and system pH of 8~10.

5. The method for preparing OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid as described in claim 1, characterized in that, In step S2, the mass ratio of sodium butyrate to zein is 1:3 to 5.

6. The method for preparing OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid as described in claim 1, characterized in that, In step S2, the stirring temperature is 30℃~40℃, the stirring time is 1~2h, and the pH of the system is 3.

7. The method for preparing OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid as described in claim 1, characterized in that, In step S3, the reaction temperature is 45℃~55℃, the reaction time is 3~5h, and the pH of the buffer solution is 7~8.

8. The method for preparing OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid as described in claim 1, characterized in that, In step S4, the concentration of the sodium alginate solution is 1.5-2.5 wt%, the concentration of the calcium chloride solution is 1.5-2 wt%, and the mass ratio of the complex to the sodium alginate solution is 1:1-3. The dripping rate is 1 drop / second, and the curing time is 25-35 minutes.

9. The method for preparing OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid as described in claim 1, characterized in that, In step S4, the first concentration is 0.5 to 0.8 wt%, and the coating time is 15 to 25 min.

10. The method for preparing OSA starch-polysaccharide interpenetrating network microspheres for targeted delivery of butyric acid as described in claim 1, characterized in that, In step S4, the second concentration is 2 to 2.5 wt%, and the secondary coating time is 5 to 15 min.

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