A method for preparing a banana resistant starch formulation that improves intestinal mucosal damage

Abstract

This invention discloses a method for preparing a banana-resistant starch preparation that improves intestinal mucosal damage, belonging to the field of resistant starch preparation technology. The preparation method includes: (1) peeling and dicing green bananas, adding sterile water containing ascorbic acid, inoculating with *Aspergillus cristatus* fermentation broth, mixing thoroughly, and fermenting to obtain a banana fermentation product; (2) centrifuging the banana fermentation product to remove the supernatant and collecting the precipitate; (3) resuspending the precipitate in sterile water, adding whey protein isolate and mixing thoroughly, adding sodium alginate solution dropwise, centrifuging, and collecting the precipitate; (4) heating the precipitate for crosslinking, and then drying at low temperature to obtain banana-resistant starch. The banana-resistant starch prepared by the method of this invention can significantly improve intestinal mucosal damage and protect the intestinal mucosa.

Description

Technical Field

[0001] This invention relates to the field of resistant starch preparation technology, and in particular to a method for preparing a banana resistant starch preparation that improves intestinal mucosal damage. Background Technology

[0002] Bananas (Musa acuminata) are one of the world's major fruit crops and important food crops, boasting high yields, wide distribution, rich nutritional value, and a sweet, soft texture, making them highly valuable for consumption. Bananas also possess medicinal properties, including clearing heat, moistening the intestines, detoxifying, nourishing yin and moistening dryness, and quenching thirst. They are used to treat symptoms such as feverish thirst, constipation, and hemorrhoidal bleeding. However, bananas are difficult to store, especially ripe bananas which are prone to spoilage and transportation, requiring significant costs for post-harvest preservation and transportation. my country's annual banana production is around 24 billion jin (approximately 12 billion kg), primarily sold fresh, with limited processing.

[0003] With the development of the resistant starch industry, there is undoubtedly a huge business opportunity for natural resistant starch sources such as green bananas, raw potatoes, and raw peas. Currently, the main methods for preparing resistant starch include hydrothermal treatment, debranching degradation, ultrasonic methods, microwave radiation methods, and steam heating methods. However, these methods all have certain drawbacks. For example, hydrothermal treatment, although simple, requires high temperatures and can easily damage the structure of natural green banana resistant starch. Debranching degradation mainly involves pullulanase to enzymatically decompose the branched chains of starch to obtain high-amylose resistant starch. Ultrasonic methods cause cavitation in starch granules during vibration, while microwave radiation can rearrange the starch structure, which is beneficial for the efficient binding of enzymes and substrates in enzymatic reactions. Therefore, microwave radiation treatment and ultrasonic treatment are usually used as auxiliary means to pre-treat starch before preparing resistant starch through enzymatic or pressure heating methods. However, this is generally used in scientific research and is rarely used in production. Therefore, there is an urgent need for a new method for preparing green banana resistant starch suitable for both scientific research and production, which is significant for the development of the banana industry.

[0004] The gut, a vital component of the human digestive system, plays a crucial role in maintaining internal homeostasis, resisting pathogen invasion, and promoting nutrient absorption. In modern life, intestinal mucosal damage is becoming increasingly frequent, closely related to various factors. From a dietary perspective, long-term high-fat, high-sugar diets and excessive intake of irritants such as alcohol alter the gut microecological environment, leading to an increase in harmful bacteria and a decrease in beneficial bacteria, thereby triggering intestinal inflammation and damaging the integrity of the intestinal mucosa. Furthermore, medications are also a significant cause of intestinal mucosal damage. Nonsteroidal anti-inflammatory drugs (NSAIDs) are widely used clinically for anti-inflammatory, analgesic, and antipyretic purposes; however, these drugs often cause gastrointestinal adverse reactions, with intestinal mucosal damage being a common one. In disease states, such as ulcerative colitis and Crohn's disease, a large number of inflammatory cells infiltrate the intestinal mucosa, releasing various inflammatory mediators, such as tumor necrosis factor-α and interleukin-6. These inflammatory mediators not only directly damage intestinal mucosal epithelial cells and vascular endothelial cells, but also interfere with the intestinal mucosal repair process, leaving the intestinal mucosa in a state of continuous damage and repair imbalance. Currently, intestinal mucosal damage is mostly repaired through drugs, and there are no reports of using banana resistant starch to improve intestinal mucosal damage. Summary of the Invention

[0005] The purpose of this invention is to provide a method for preparing a banana resistant starch preparation that improves intestinal mucosal damage, in order to solve the problems existing in the prior art. By optimizing the preparation process of banana resistant starch, the obtained banana resistant starch can significantly improve intestinal mucosal damage.

[0006] To achieve the above objectives, the present invention provides the following solution:

[0007] This invention provides a method for preparing banana resistant starch, comprising the following steps:

[0008] (1) Peel the green bananas, dice them, add sterile water containing ascorbic acid, inoculate with the fermentation broth of *Aspergillus cristatus*, mix well, and ferment to obtain banana fermentation product;

[0009] (2) Centrifuge the banana fermentation product to remove the supernatant and collect the precipitate;

[0010] (3) Resuspend the precipitate in sterile water, add whey protein isolate and mix well, add sodium alginate solution dropwise, centrifuge and collect the precipitate;

[0011] (4) The precipitate is heated and crosslinked, and then dried at low temperature to obtain banana resistant starch.

[0012] In the above scheme, adding sterile water containing ascorbic acid after peeling and dicing the green bananas aims to prevent oxidation of the banana surface. Inoculating with *Aspergillus cristatus* utilizes pectin and starch as carbon sources for fermentation, reducing pectin's interference in the extraction of resistant starch from bananas and increasing the extraction rate. Furthermore, *Aspergillus cristatus* fermentation mitigates the damage to the resistant starch structure caused by traditional direct high-speed shearing of bananas into a pulp. Through *Aspergillus cristatus* fermentation, while reducing pectin and starch interference, it also converts them into beneficial soluble monosaccharides, rather than destroying the physical structure of the resistant starch. The precipitate is obtained by centrifugation, resuspended, and then whey protein isolate is added. Immediately afterwards, sodium alginate solution is added dropwise. This process avoids whey protein isolate precipitation and allows it to cross-link with sodium alginate to form a fibrous network structure. The resulting resistant starch is then encapsulated. Brief heating at 80°C denatures the outer whey protein isolate and increases the density of the fibrous network structure without destroying the encapsulated resistant starch. This process essentially yields a novel, modified resistant starch. During digestion, the outer fibrous network structure is degraded in the stomach and small intestine. Upon reaching the colon, the resistant starch is released, achieving effective controlled release of banana resistant starch. Therefore, the banana modified starch prepared under specific conditions according to this invention can fully exert its function in the intestine while avoiding the problem of poor efficacy caused by dissolution in other tissues.

[0013] Optionally, the sterile water containing ascorbic acid is an aqueous solution of 2.0‰-3.0‰ ascorbic acid, and the sterile water containing ascorbic acid is added to the green bananas at a volume of 3-4 times the mass of the green bananas.

[0014] Optionally, the inoculum size of the *Aurogonium cristatum* fermentation broth is 2%-4%, the fermentation temperature is 28-30℃, and the fermentation time is 20-40 hours. Under these conditions, large-scale culture can rapidly obtain *Aurogonium cristatum* fermentation broth, and the cell quality and concentration both meet the requirements of this invention.

[0015] Optionally, the *Aurotriton malignancies* fermentation broth is obtained by inoculating *Aurotriton malignancies* into malt extract medium for large-scale culture, and the concentration of the *Aurotriton malignancies* fermentation broth is 3.5 × 10⁻⁶. 8 CFU / mL.

[0016] Optionally, the amount of whey protein isolate added is 3.0%-4.0% of the total weight of the precipitate.

[0017] Optionally, the amount of sodium alginate solution added is 1.0%-1.5% of the total weight of the precipitate, and the concentration of the sodium alginate solution is 2.0 g / L.

[0018] Cross-linking whey protein isolate and sodium alginate solution within the above range can form a fibrous structure with the best encapsulation effect, which is beneficial for the efficient encapsulation of resistant starch.

[0019] Optionally, the conditions for heating and cross-linking the precipitate are: treatment at 80℃ for 1-5 minutes; and the temperature for low-temperature drying is: 30-45℃. Temperature has a certain impact on the denaturation of whey protein isolate and its cross-linking effect with sodium alginate. If the temperature is too low, it may lead to insufficient cross-linking and an insufficiently compact structure, which is not enough to ensure efficient encapsulation of resistant starch, thus leading to partial degradation of resistant starch. If the temperature is too high, it will affect the denaturation of the outer whey protein isolate and also damage the structure of resistant starch, affecting its performance after use.

[0020] The present invention also provides banana resistant starch prepared by the aforementioned preparation method.

[0021] The present invention also provides a composition comprising the banana resistant starch described above.

[0022] The present invention also provides the use of the banana resistant starch or the composition thereof in the preparation of a medicament for improving the intestinal mucosa.

[0023] The present invention discloses the following technical effects:

[0024] This invention utilizes *Aspergillus cristatus* to ferment green bananas to extract resistant starch, which is then encapsulated with whey protein isolate and sodium alginate cross-linking structures. Further optimization of the cross-linking structure yields a modified banana resistant starch. Animal experiments showed that the banana resistant starch prepared by this invention significantly reduced inflammatory factors in blood and intestinal tissue, increased the content of tight junction proteins in intestinal tissue, and significantly improved weight gain and stool formation. This indicates that the banana resistant starch prepared by this invention can significantly improve intestinal mucosal damage and plays an important role in protecting the intestinal mucosa. Attached Figure Description

[0025] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0026] Figure 1 The relative mRNA expression level of the tight junction protein ZO-1;

[0027] Figure 2 The relative mRNA expression level of the tight junction protein occludin;

[0028] In the figure above, CK: blank control group; Sham: intestinal mucosal injury model group; S1: banana resistant starch experimental group (Example 1); S2: banana resistant starch experimental group (Example 2); S3: banana resistant starch experimental group (Example 3); D1: banana resistant starch experimental group (Comparative Example 1); D2: banana resistant starch experimental group (Comparative Example 2); D3: banana resistant starch experimental group (Comparative Example 3); D4: banana resistant starch experimental group (Comparative Example 4); * indicates significant difference compared with CK, P < 0.05; # indicates significant difference compared with Sham, P < 0.05. Detailed Implementation

[0029] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0030] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Any stated value or intermediate value within a stated range, as well as each smaller range between any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0031] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0032] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be readily apparent to those skilled in the art. This specification and embodiments are merely exemplary.

[0033] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0034] Resistant starch, also known as enzyme-resistant starch or indigestible starch, cannot be broken down by enzymes in the small intestine, but it can undergo fermentation with volatile fatty acids in the human gastrointestinal tract, specifically the colon.

[0035] Resistant starch is found in some natural foods, such as potatoes, bananas, and rice. Corn starch, in particular, is high in amylose and contains up to 60% resistant starch. This type of starch is more difficult to break down than other starches, digests slowly in the body, and is absorbed and enters the bloodstream more gradually. Its properties are similar to soluble fiber, offering some slimming effects, and it has become increasingly popular among those concerned with weight loss in recent years.

[0036] Several existing technologies have disclosed methods for extracting resistant starch from green bananas. For example, CN105852138B describes a method for extracting natural resistant starch from bananas. This patent uses a combination of freezing and blanching for pretreatment to facilitate separation of the peel and pulp. Then, it utilizes microbial fermentation combined with enzymatic treatment to remove other components and extract high-purity natural resistant starch from bananas. This method combines cold and heat treatment, microbial fermentation, enzymatic hydrolysis, and ultrasound. While it can obtain natural resistant starch from bananas, the process is complex, and the control of multiple steps, such as ultrasound conditions, is very strict. Another example is CN101427803B, which describes pectin-modified resistant starch, its composition, and a method for preparing resistant starch. This method involves cross-linking starch with pectin via a pectin esterase reaction. The resistant starch has low amylase digestibility and can therefore be used in foods, including nutritional supplements, to reduce calories and increase fiber content. This method mainly utilizes pectin to modify resistant starch. However, this resistant starch introduces another type of dietary fiber, and it remains unknown whether the modified resistant starch will perform its original dietary fiber function in the intestines. Therefore, given the various existing methods for extracting resistant starch, but the different levels of dietary fiber function they achieve, exploring new methods for extracting resistant starch is still very necessary.

[0037] The *Eurotium cristatum* used in this embodiment of the invention has the accession number CICC 2099 and can be routinely purchased from the China Industrial Microbial Culture Collection Center. This invention is not limited to this strain, and the public can purchase *Eurotium cristatum* with the same function through the above platform or other channels.

[0038] Example 1: A method for preparing banana resistant starch

[0039] (1) Select bananas with green peels, peel them, chop them into small cubes (e.g., 1×1×cm cubes), add 3 times the volume of sterile water containing ascorbic acid (2.5‰ ascorbic acid was added to prevent the bananas from discoloring), inoculate with 2% Aspergillus cristatus fermentation broth, mix well, ferment at 28℃ for 20h to obtain banana fermentation products.

[0040] Among them, *Eurotium cristatum* was activated with malt extract agar medium, then inoculated into malt extract culture medium and cultured at 30℃ for 3 days to obtain a bacterial count ≥10. 8 Fermentation broth with CFU / mL; the bacterial concentration was adjusted to 3.5 × 10⁻⁶ CFU / mL in this experiment. 8 Inoculate with fermentation broth containing CFU / mL;

[0041] (2) Centrifuge the banana fermentation product to remove the supernatant and collect the precipitate;

[0042] (3) Resuspend the precipitate with sterile water, add 3.5% whey protein isolate by weight of the precipitate and mix well, then add sodium alginate solution (the amount of sodium alginate is 1.0% of the total weight of the precipitate and the concentration is 2.0 g / L), centrifuge and collect the precipitate.

[0043] (4) The precipitate was treated at 80°C for 1 min, and then dried at 30°C to form granules, which yielded banana resistant starch.

[0044] The natural resistant starch of bananas prepared in this embodiment can be observed under a polarizing microscope and a scanning electron microscope. The polarized cross of the natural resistant starch of bananas is obvious, mostly X-shaped, and the starch hilum is located in the center of the smaller end of the starch granule. It has an oval structure, with one end of the starch granule being larger and the other end being smaller.

[0045] Example 2: A method for preparing banana resistant starch

[0046] (1) Select bananas with green peels, peel them, chop them into small cubes (e.g., 1×1×cm cubes), add 4 times the volume of sterile water containing ascorbic acid (2.0‰ ascorbic acid was added to prevent the bananas from discoloring), inoculate with 3% Aspergillus cristatus fermentation broth, mix well, ferment at 29℃ for 30h to obtain banana fermentation products.

[0047] Among them, *Eurotium cristatum* was activated with malt extract agar medium, then inoculated into malt extract culture medium and cultured at 30℃ for 3 days to obtain a bacterial count ≥10. 8 Fermentation broth with CFU / mL; the bacterial concentration was adjusted to 3.5 × 10⁻⁶ CFU / mL in this experiment. 8 Inoculate with fermentation broth containing CFU / mL;

[0048] (2) Centrifuge the banana fermentation product to remove the supernatant and collect the precipitate.

[0049] (3) Resuspend the precipitate with sterile water, add 4.0% whey protein isolate by weight of the precipitate and mix well, then add sodium alginate solution (the amount of sodium alginate is 1.25% of the total weight of the precipitate and the concentration is 2.0 g / L), centrifuge and collect the precipitate;

[0050] (4) The precipitate was treated at 80°C for 3 minutes, and then dried at 40°C to form granules, thus obtaining banana resistant starch.

[0051] The banana resistant starch prepared in this embodiment exhibits the same starch granule structure as in Example 1.

[0052] Example 3: A method for preparing banana resistant starch

[0053] (1) Select bananas with green peels, peel them, chop them into small cubes (e.g., 1×1×cm cubes), add 5 times the volume of sterile water containing ascorbic acid (2.5‰ ascorbic acid was added to prevent the bananas from discoloring), inoculate with 4% Aspergillus cristatus fermentation broth, mix well, ferment at 30℃ for 40h to obtain banana fermentation products.

[0054] Among them, *Eurotium cristatum* was activated with malt extract agar medium, then inoculated into malt extract culture medium and cultured at 30℃ for 3 days to obtain a bacterial count ≥10. 8 Fermentation broth with CFU / mL; the bacterial concentration was adjusted to 3.5 × 10⁻⁶ CFU / mL in this experiment. 8 Inoculate with fermentation broth containing CFU / mL;

[0055] (2) Centrifuge the banana fermentation product to remove the supernatant and collect the precipitate;

[0056] (3) Resuspend the precipitate with sterile water, add 3.0% whey protein isolate by weight of the precipitate and mix well, then add sodium alginate solution (the amount of sodium alginate is 1.5% of the total weight of the precipitate and the concentration is 2.0 g / L), centrifuge and collect the precipitate;

[0057] (4) The precipitate was treated at 80°C for 5 minutes, and then dried at 45°C to form granules, thus obtaining banana resistant starch.

[0058] The banana resistant starch prepared in this embodiment exhibits the same starch granule structure as in Example 1.

[0059] Comparative Example 1

[0060] The difference between this comparative example and Example 1 is that *Aspergillus cristatus* was not inoculated, and pectinase was used instead for the enzymatic hydrolysis reaction. Details are as follows:

[0061] (1) Select bananas with green peels, peel them, chop them into small cubes (e.g., 1×1×cm cubes), add 3 times the volume of sterile water containing ascorbic acid (3.0‰ ascorbic acid was added to prevent the bananas from discoloring), add pectinase for enzymatic hydrolysis; the amount of pectinase added is 30U / mL, the reaction conditions are: pH=4.0, 55℃, and the hydrolysis time is 1h to obtain the banana hydrolysis product;

[0062] (2) Centrifuge the banana enzymatic hydrolysis product to remove the supernatant, collect the precipitate, and wash it 3 times;

[0063] (3) After washing, the precipitate was resuspended in sterile water, and 3.5% whey protein isolate was added to the total weight of the precipitate and mixed evenly. Then, sodium alginate solution was added dropwise (the amount of sodium alginate was 1.0% of the total weight of the precipitate, and the concentration was 2.0 g / L). The precipitate was centrifuged and collected.

[0064] (4) The precipitate was treated at 80℃ for 5 minutes, and then dried at 45℃ to form granules, thus obtaining banana resistant starch.

[0065] The banana resistant starch prepared in this comparative example exhibits the same typical starch granule structure as in Example 1.

[0066] Comparative Example 2

[0067] The difference between this comparative example and Example 1 is that the method of adding whey protein isolate and sodium alginate is changed, as shown below:

[0068] (1) Same as (1) in Example 1;

[0069] (2) Same as (2) in Example 1;

[0070] (3) Resuspend the precipitate in sterile water, add 3.5% whey protein isolate and 1.0% sodium alginate by weight of the precipitate, mix well, centrifuge, and collect the precipitate;

[0071] (4) Same as (4) in Example 1.

[0072] The banana resistant starch prepared in this comparative example exhibits the same typical starch granule structure as in Example 1.

[0073] Comparative Example 3

[0074] The difference between this comparative example and Example 1 is that the treatment temperature of the collected precipitate is changed, as shown below:

[0075] (1) Same as (1) in Example 1;

[0076] (2) Same as (2) in Example 1;

[0077] (3) Same as (3) in Example 1;

[0078] (4) The precipitate was treated at 90°C for 1 min, and then dried at 30°C to form granules, thus obtaining banana resistant starch.

[0079] The banana resistant starch prepared in this comparative example exhibits the same typical starch granule structure as in Example 1.

[0080] Comparative Example 4

[0081] The difference between this comparative example and Example 1 is that the treatment temperature of the collected precipitate is changed, as shown below:

[0082] (1) Same as (1) in Example 1;

[0083] (2) Same as (2) in Example 1;

[0084] (3) Same as (3) in Example 1;

[0085] (4) The precipitate was treated at 50°C for 1 min, and then dried at 30°C to form granules, thus obtaining banana resistant starch.

[0086] The banana resistant starch prepared in this comparative example exhibits the same typical starch granule structure as in Example 1.

[0087] Animal experiments

[0088] The banana resistant starch prepared in Examples 1-3 and Comparative Examples 1-4 was used to conduct mouse model experiments of intestinal mucosal injury.

[0089] Male Balb / c mice of the same age, weighing 20±1g, were selected and randomly divided into 9 groups after one week of acclimatization:

[0090] (1) Blank control group;

[0091] (2) Banana resistant starch experimental group (Examples 1-3);

[0092] (3) Banana resistant starch control group (comparative examples 1-4);

[0093] (4) Intestinal mucosal injury model group.

[0094] Eight mice were used in each group. The blank control group was administered sterile distilled water by gavage, while the other groups were administered sterile distilled water containing 8% DSS by gavage. The model was established in 8 days. The banana resistant starch experimental group and the banana resistant starch control group were administered the corresponding resistant starch solution by gavage once a day at a dose of 100 μg / kg / day. The intestinal mucosal injury model group and the blank control group were administered the same volume of physiological saline by gavage daily. The administration was continued for one week. After the last administration, the mice were fasted for 24 hours, weighed, and their fecal characteristics were observed. After the experiment, blood was collected from the inner canthal vein of the eye, and serum was collected to detect the levels of pro-inflammatory factors IL-1β and TNF-α in the blood. Subsequently, the mice were anesthetized and sacrificed, and small intestinal tissue was collected to detect the levels of inflammatory factors IL-1β and TNF-α in the small intestinal tissue, as well as the relative mRNA expression levels of tight junction proteins ZO-1 and occludin.

[0095] Table 1. Mouse body weight and fecal characteristics

[0096] Blank control group 115.0 normal Intestinal mucosal injury model group 21.9 Diarrhea, bloody stool Banana resistant starch experimental group (Example 1) 97.0 normal Banana resistant starch experimental group (Example 2) 93.0 normal Banana resistant starch experimental group (Example 3) 93.5 normal Banana resistant starch control group (Comparative Example 1) 52.5 normal Banana resistant starch control group (Comparative Example 2) 59.7 normal Banana resistant starch control group (Comparative Example 3) 66.5 normal Banana resistant starch control group (Comparative Example 4) 64.3 normal

[0097] Note: Weight gain rate (%) = (Final weight - Initial weight) / Initial weight × 100

[0098] Table 1 shows that, compared with the intestinal mucosal injury model group, the banana resistant starch administered by gavage in Examples 1-3 and Comparative Examples 1-4 significantly improved diarrhea and bloody stools in mice, and also significantly improved weight gain. Furthermore, the banana resistant starch in Examples 1-4 showed a more significant improvement effect than that in Comparative Examples 1-4.

[0099] Table 2. Levels of pro-inflammatory factors in blood

[0100]

[0101]

[0102] Table 2 shows that, compared with the intestinal mucosal injury model group, the banana resistant starch administered by gavage in Examples 1-3 and Comparative Examples 1-4 significantly reduced the levels of pro-inflammatory factors IL-1β and TNF-α in the blood, and the banana resistant starch in Examples 1-4 had a more significant effect than that in Comparative Examples 1-4.

[0103] Table 3. Expression levels of inflammatory factors in intestinal tissue

[0104]

[0105] Table 3 shows that, compared with the intestinal mucosal injury model group, the banana resistant starch administered by gavage in Examples 1-3 and Comparative Examples 1-4 can significantly reduce the content of pro-inflammatory factors IL-1β and TNF-α in intestinal tissue, and the banana resistant starch in Examples 1-4 has a more obvious effect than the banana resistant starch in Comparative Examples 1-4.

[0106] like Figure 1 and Figure 2 As shown, compared with the intestinal mucosal injury model group, the banana resistant starch administered by gavage in Examples 1-3 and Comparative Examples 1-4 significantly increased the relative expression levels of ZO-1 and Occludin mRNA in intestinal tight junctions (P < 0.05), and the banana resistant starch in Examples 1-4 had a more significant effect than that in Comparative Examples 1-4.

[0107] The above experimental results demonstrate that the banana resistant starch prepared in this invention can significantly promote the decrease of verification factors in the blood and intestines, and significantly improve intestinal inflammation. It can also improve intercellular tight junctions by upregulating the relative expression levels of ZO-1 and Occludin mRNA, thereby facilitating the restoration of the integrity and permeability of the intestinal barrier and preventing intestinal molecules from entering the bloodstream. Through the adjustment of intestinal tissue, significant improvements were observed in appearance, mouse weight, and stool formation. Therefore, this invention demonstrates that the banana resistant starch prepared in this invention can improve intestinal mucosal damage and plays an important role in protecting the intestinal mucosa.

[0108] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. A method for preparing banana resistant starch, characterized in that, Includes the following steps: (1) Peel the green bananas, dice them, add sterile water containing ascorbic acid, inoculate with the fermentation broth of *Aspergillus cristatus*, mix well, and ferment to obtain banana fermentation product; (2) Centrifuge the banana fermentation product to remove the supernatant and collect the precipitate; (3) Resuspend the precipitate in sterile water, add whey protein isolate and mix well, add sodium alginate solution dropwise, centrifuge and collect the precipitate; (4) The precipitate is heated and crosslinked, and then dried at low temperature to obtain banana resistant starch; The sterile water containing ascorbic acid is added to the green bananas at a volume of 3-4 times the weight of the green bananas. The inoculum size of the *Eurotium cristatum* fermentation broth is 2%-4%, the fermentation temperature is 28-30℃, and the fermentation time is 20-40 hours. The amount of whey protein isolate added is 3.0%-4.0% of the total weight of the precipitate; The amount of sodium alginate solution added is 1.0%-1.5% of the total weight of the precipitate; The conditions for cross-linking the precipitate by heating are: treatment at 80℃ for 1-5 minutes; the temperature for low-temperature drying is: 30-45℃.

2. The preparation method according to claim 1, characterized in that, The sterile water containing ascorbic acid is an aqueous solution of ascorbic acid at a concentration of 2.0‰-3.0‰.

3. The preparation method according to claim 1, characterized in that, The *Aureobasidium cristatum* fermentation broth was obtained by inoculating *Aureobasidium cristatum* into malt extract medium and expanding the culture. The bacterial concentration of the *Aureobasidium cristatum* fermentation broth was 3.5 × 10⁻⁶. 8 CFU / mL.

4. The preparation method according to claim 1, characterized in that, The concentration of the sodium alginate solution is 2.0 g / L.

5. The use of banana resistant starch prepared by the method according to any one of claims 1-4 in the preparation of a drug for improving intestinal mucosal damage.

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