A method to improve gut microbial utilization of high amylose content

Abstract

This invention belongs to the field of starch processing, specifically relating to a method for improving the intestinal microbial utilization rate of high amylose. The method is achieved through the following steps: high amylose pretreatment; low-temperature plasma treatment; spraying with a hydrophobicating agent; washing and centrifuging drying. This invention, by immersing high amylose in the plasma, allows divalent metal ions to be retained on the surface and inside the high amylose granules, enhancing the etching effect of the low-temperature plasma on the high amylose granules and significantly improving the surface roughness of the high amylose granules, thereby promoting the residence time of intestinal microorganisms on the high amylose granules. The low-temperature plasma treatment, using a mixed gas, introduces fluorinated ethane groups onto the starch surface, providing initial hydrophobicity. It also provides reaction sites for subsequent hydrophobication. Shorter-range hydrophobic groups can improve the adhesion and fixation ability of proteins on the surface of intestinal microbial cells, thus improving the utilization rate of intestinal microorganisms.

Description

Technical Field

[0001] This invention belongs to the field of starch processing, and specifically relates to a method for improving the utilization rate of intestinal microorganisms of high amylose. Background Technology

[0002] High-amylose starch is a special type of starch, with a much higher proportion of amylose than ordinary starch, resulting in significant differences in its properties compared to natural starch. It is a novel ingredient used in pharmaceuticals and food processing. Due to its highly crystalline structure and tight interchain interactions, high-amylose starch is not digested in the small intestine, thus it can reach the large intestine as a prebiotic. However, its dense structure also results in low utilization by intestinal microorganisms in the large intestine, leading to a relatively weaker prebiotic effect compared to ordinary resistant starch.

[0003] Domestic and international research indicates that the fermentation of high-amylose in the large intestine primarily occurs under the influence of the gut microbiota. Some microorganisms, such as lactic acid bacteria and bifidobacteria, can utilize high-amylose as a carbon source for fermentation. However, this fermentation process is typically slow and produces relatively low amounts of short-chain fatty acids (such as butyric acid and propionic acid). These short-chain fatty acids are crucial for maintaining gut health, but the low fermentation rate of high-amylose limits their production in the gut.

[0004] The crystalline structure of high-amylose is one of the main reasons why it is difficult for gut microbiota to utilize. The enzyme system of gut microbiota has limited ability to degrade high-amylose. Microorganisms in the large intestine mainly rely on enzymes such as amylase and glucosidase to degrade starch. However, these enzymes are less efficient at breaking down high-amylose because its crystalline structure makes it difficult for amylase and glucosidase to effectively contact and cleave starch molecules, leading to incomplete digestion. Furthermore, the poor swelling and water solubility of high-amylose also limit its contact with gut microbial enzymes. In summary, high-amylose has relatively low utilization by gut microbiota due to its unique structural characteristics. This is mainly due to its crystalline structure and the limited degradation ability of microbial enzymes. Improving the fermentation characteristics of high-amylose in the large intestine has health benefits for human health.

[0005] Improving the utilization efficiency of high-amylose by gut microbiota is a research hotspot in food science and biotechnology. Existing domestic and international literature has proposed various solutions to this problem. Some studies have suggested using physical methods such as grinding and extrusion to disrupt the crystal structure of high-amylose, thereby improving its utilization efficiency by gut microbiota. However, physical methods typically completely alter the crystalline properties of starch, reducing its thermal stability and leading to easy gelatinization during processing, resulting in direct absorption in the small intestine. Chemical modifications such as esterification, etherification, or cross-linking can increase the hydrophobicity of high-amylose, thereby improving its fermentability in the gut. However, common chemical modifications often fail to achieve effective results. Therefore, while ensuring the smooth passage of high-amylose through the small intestine, exploring ways to improve the gut microbiota utilization rate of high-amylose to fully realize its potential for gut health has become an urgent problem to be solved. Summary of the Invention

[0006] To address the problems existing in the prior art, this invention provides a method for improving the intestinal microbial utilization rate of high amylose through plasma-assisted hydrophobic modification. This method can effectively alter the surface structure of high amylose without destroying its internal crystalline structure, and can also increase its fermentability in the large intestine, thereby promoting the proliferation of beneficial bacteria and the secretion of beneficial metabolites in the gut. The technical solution adopted by this invention to achieve the above objectives is as follows:

[0007] This invention provides a method for improving the intestinal microbial utilization of high-amylose content, comprising the following steps:

[0008] (1) Pretreatment of high amylose: High amylose potato starch is soaked in salt solution and then dried.

[0009] (2) Low-temperature plasma treatment: The dried high amylose potato starch was subjected to low-temperature plasma treatment. After the treatment was completed, it was taken out and cooled to room temperature.

[0010] (3) Spraying hydrophobic reagent: Spread the cooled high amylose potato starch evenly, spray octadecyl dimethyl chlorosilane solution on the surface, and keep the sprayed high amylose potato starch warm; place the hydrophobically treated high amylose potato starch in ethanol and stir magnetically;

[0011] (4) Collection: The washed high amylose potato starch is collected by centrifugation and then dried naturally.

[0012] Furthermore, in step (1), the concentration of the salt solution is 1-3%; the salt solution is a divalent metal ion salt; most preferably, the salt solution is one or more of ZnCl2, MgCl2, and CuCl2.

[0013] Furthermore, in step (1), the temperature of the salt solution is 60°C; the soaking time is 2-3 hours; and the drying is carried out until the moisture content is 20%-25%.

[0014] Furthermore, in step (2), the parameters of the low-temperature plasma treatment are: time 10-15 min, power 80-100 W, and argon-hexafluoroethane mixed gas as working gas, with hexafluoroethane accounting for 6%-10% in the mixed gas.

[0015] Furthermore, in step (3), the amount of octadecyl dimethyl chlorosilane solution sprayed is 3%-5% of the mass of high amylose potato starch.

[0016] Furthermore, in step (3), the concentration of the octadecyl dimethylchlorosilane solution is 5%-10%.

[0017] Furthermore, in step (3), the heat preservation treatment is performed under dry conditions at 60°C for 4-6 hours; the mass ratio of the hydrophobic high amylose potato starch to ethanol is 1:10-15; and the magnetic stirring time is 10-15 minutes.

[0018] This invention addresses the problem of low gut microbial utilization efficiency of high-amylose starch by providing a method to improve its utilization rate. The specific beneficial effects are as follows:

[0019] (1) By soaking high amylose, divalent metal ions can be preserved on the surface and inside of high amylose particles, which enhances the etching effect of low temperature plasma on high amylose particles and significantly improves the surface roughness of high amylose particles, thereby promoting the residence time of intestinal microorganisms in high amylose particles.

[0020] (2) By treating with a mixed gas using low-temperature plasma, fluorinated ethane groups can be introduced onto the starch surface, providing initial hydrophobicity. This also provides reaction sites for subsequent hydrophobicization. Shorter-range hydrophobic groups can enhance the colonization ability of proteins on the surface of intestinal microbial cells.

[0021] (3) Treatment with octadecyl dimethyl chlorosilane can form certain long-range hydrophobic groups on high amylose particles, which helps to change the environment of the hydration layer on the surface of high amylose particles, which is conducive to the stability of intestinal microbial cells on the surface of high amylose particles and improves the utilization rate of intestinal microorganisms. Attached Figure Description

[0022] Figure 1 A comparison chart of absorbance measurements for different groups; Detailed Implementation

[0023] The technical solution of the present invention will be further explained and described below through specific embodiments.

[0024] Example 1

[0025] (1) Preparation of high amylose: Select pure high amylose potato starch as the treatment object;

[0026] (2) Pretreatment of high amylose: High amylose potato starch was soaked in 2% ZnCl2 solution at 60℃ for 3h;

[0027] (3) Drying of high amylose: Soaked high amylose potato starch was dried to a moisture content of 25%;

[0028] (4) Low-temperature plasma treatment: Dry high-amylose potato starch was placed in a low-temperature plasma reactor and the plasma treatment parameters were set as follows: time 15 min, power 100 W, and argon-hexafluoroethane mixture was used as the working gas with a hexafluoroethane ratio of 8%.

[0029] (5) Cooling: Take out the high amylose potato starch treated with low-temperature plasma and cool it to room temperature;

[0030] (6) Spraying hydrophobic reagent: Spread the cooled high amylose potato starch evenly, and spray octadecyl dimethyl chlorosilane reagent (5% of the mass of high amylose potato starch) on the surface, with the concentration controlled at 10%;

[0031] (7) Hydrophobic treatment: The sprayed high amylose potato starch was kept in a drying oven at 60°C for 6 hours;

[0032] (8) Washing: The hydrophobically treated high amylose potato starch was placed in an ethanol solution at a mass ratio of 1:15 and washed by magnetic stirring for 15 minutes. The washing was repeated twice.

[0033] (9) Collection: After washing, high amylose potato starch is collected by centrifugation and then dried naturally to obtain the sample.

[0034] Example 2

[0035] (1) Preparation of high amylose: Select pure high amylose potato starch as the treatment object;

[0036] (2) Pretreatment of high amylose: High amylose potato starch was soaked in 3% MgCl2 solution at 60℃ for 2h;

[0037] (3) Drying of high amylose: Soaked high amylose potato starch was dried to a moisture content of 25%;

[0038] (4) Low-temperature plasma treatment: Dry high-amylose potato starch was placed in a low-temperature plasma reactor and the plasma treatment parameters were set as follows: time 12 min, power 100 W, argon-hexafluoroethane mixture was used as working gas, and the proportion of hexafluoroethane was 10%.

[0039] (5) Cooling: Take out the high amylose potato starch treated with low-temperature plasma and cool it to room temperature;

[0040] (6) Spraying hydrophobic reagent: Spread the cooled high amylose potato starch evenly, and spray octadecyl dimethyl chlorosilane reagent (3% of the mass of high amylose potato starch) on the surface, with the concentration controlled at 8%;

[0041] (7) Hydrophobic treatment: The sprayed high amylose potato starch was kept in a drying oven at 60°C for 5 hours;

[0042] (8) Washing: Place the hydrophobically treated high amylose potato starch in an ethanol solution and stir magnetically for 15 minutes. Repeat the above steps twice.

[0043] (9) Collection: After washing, high amylose potato starch is collected by centrifugation and then dried naturally to obtain the sample.

[0044] Comparative Example 1

[0045] (1) Preparation of high amylose: Select pure high amylose potato starch as the treatment object;

[0046] (2) Drying of high amylose: Adjust the moisture content of high amylose potato starch to 25%;

[0047] The other steps are the same as in Example 1.

[0048] Comparative Example 2

[0049] (1) Preparation of high amylose: Select pure high amylose potato starch as the treatment object;

[0050] (2) Pretreatment of high amylose: High amylose potato starch was soaked in 2% ZnCl2 solution at 60℃ for 3h;

[0051] (3) Drying of high amylose: Soaked high amylose potato starch was dried to a moisture content of 25%;

[0052] (4) Spraying hydrophobic reagent: Spread the dried high amylose potato starch evenly, and spray octadecyl dimethyl chlorosilane reagent on the surface, with the concentration controlled at 10%;

[0053] The other steps are the same as in Example 1.

[0054] Comparative Example 3

[0055] (1) Preparation of high amylose: Select pure high amylose potato starch as the treatment object;

[0056] (2) Pretreatment of high amylose: High amylose potato starch was soaked in 2% ZnCl2 solution at 60℃ for 3h;

[0057] (3) Drying of high amylose: Soaked high amylose potato starch is dried to a moisture content of 20%-25%;

[0058] (4) Low-temperature plasma treatment: Dry high amylose potato starch was placed in a low-temperature plasma reactor and the plasma treatment parameters were set as follows: time 15 min, power 100 W, and argon gas was used as the working gas.

[0059] The other steps are the same as in Example 1.

[0060] Comparative Example 4

[0061] (1) Preparation of high amylose: Select pure high amylose potato starch as the treatment object;

[0062] (2) Pretreatment of high amylose: High amylose potato starch was soaked in 2% ZnCl2 solution at 60℃ for 3h;

[0063] (3) Drying of high amylose: Soaked high amylose potato starch was dried to a moisture content of 25%;

[0064] (4) Low-temperature plasma treatment: Dry high-amylose potato starch was placed in a low-temperature plasma reactor and the plasma treatment parameters were set as follows: time 15 min, power 100 W, and argon-hexafluoroethane mixture was used as the working gas with a hexafluoroethane ratio of 8%.

[0065] (5) Cooling: Take out the high amylose potato starch treated with low-temperature plasma and cool it to room temperature;

[0066] (8) Washing: Place the cooled high amylose potato starch in an ethanol solution and stir magnetically for 15 minutes. Repeat the above steps twice.

[0067] (9) Collection: After washing, high amylose potato starch is collected by centrifugation and then dried naturally to obtain the sample.

[0068] Effect Example

[0069] (a) Growth curve

[0070] Methods: Human fecal extract was inoculated onto culture medium at a 1% inoculum. An equal volume of glucose in the medium was replaced with high-amylose starch under different treatment conditions. The mixture was anaerobically cultured at 37°C, and OD values ​​were measured using a microplate reader. 600 The absorbance was measured every 1 hour.

[0071] Figure 1 In the comparison chart of the treatment method provided in Example 1 of this invention (Example 1), no pretreatment (Comparative Example 1), no plasma treatment (Comparative Example 2), argon plasma treatment during low-temperature plasma treatment (Comparative Example 3), and no hydrophobic modification treatment - without octadecyl dimethylchlorosilane (Comparative Example 4), it can be seen that the untreated high amylose (control) has a certain proliferative effect on intestinal microorganisms, but it is the lowest among all the subjects studied. The treatment method provided by this invention can significantly increase the proliferation rate of intestinal microorganisms and reduce the OD of the final bacterial suspension. 600 It also reached its maximum value (1.74).

[0072] (ii) Surface hydrophobicity

[0073] Methods: The hydrophobicity of different high-amylose starches was characterized using contact angle. Contact angles were measured using a video optical contact angle analyzer, and the water resistance of the samples was determined using the seated drop method at ambient temperature, with water as the test solvent. Starch samples were processed into circular starch flakes approximately 1 cm in diameter under a pressure of 20 MPa. 2 μL of ultrapure water was dropped onto the surface of the flake using a 5 μL microsyringe. Photography was taken after the water droplet had fully contacted the sample, with a delay of 10 s. Each sample was tested three times, and the average value was taken. Specific results are shown in Table 1.

[0074] (iii) Surface roughness

[0075] High-amylose content was dissolved in ultrapure water to prepare a 10 g / L solution. After stirring for 10 min, the solution was uniformly dropped onto a clean mica sheet and dried at room temperature. Observation was performed using an atomic force microscope (AFM). The probe tip curvature was less than 10, the typical force constant was 2 N / m, and the scanning range was 2 μm. The average surface roughness was calculated using NanoScope Analysis. Specific results are shown in Table 1.

[0076] Table 1

[0077]

Claims

1. A method for improving the intestinal microbial utilization rate of high amylose content, characterized in that, Includes the following steps: (1) Pretreatment of high amylose: Soak high amylose potato starch in salt solution and then dry the soaked high amylose potato starch; (2) Low-temperature plasma treatment: The dried high amylose potato starch was subjected to low-temperature plasma treatment. After the treatment was completed, it was taken out and cooled to room temperature. (3) Spraying hydrophobic reagent: Cooled high amylose potato starch is spread out and sprayed with octadecyl dimethyl chlorosilane solution. The sprayed high amylose potato starch is kept warm to obtain hydrophobic high amylose potato starch. The hydrophobic high amylose potato starch is washed in ethanol and magnetically stirred. (4) Collection: The washed high amylose potato starch is collected by centrifugation and then naturally dried to obtain the product; In step (1), the concentration of the salt solution is 1-3%; the salt solution is a divalent metal ion salt; In step (2), the low-temperature plasma treatment uses an argon-hexafluoroethane mixture as the working gas, with hexafluoroethane accounting for 6%-10% of the mixture.

2. The method according to claim 1, characterized in that, In step (1), the salt solution is one or more of ZnCl2, MgCl2, and CuCl2.

3. The method according to claim 1 or 2, characterized in that, In step (1), the temperature of the salt solution is 60°C; the soaking time is 2-3 hours; and the drying is carried out until the moisture content is 20%-25%.

4. The method according to claim 1, characterized in that, In step (2), the parameters for the low-temperature plasma treatment are: time 10-15 min, power 80-100 W.

5. The method according to claim 1, characterized in that, In step (3), the amount of octadecyl dimethyl chlorosilane solution sprayed is 3%-5% of the mass of high amylose potato starch.

6. The method according to claim 5, characterized in that, In step (3), the concentration of the octadecyl dimethylchlorosilane solution is 5%-10%.

7. The method according to claim 6, characterized in that, In step (3), the heat preservation treatment is to keep warm at 60°C for 4-6 hours; the mass ratio of the hydrophobic high amylose potato starch to ethanol is 1:10-15; and the magnetic stirring time is 10-15 minutes.

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