Rice resistant starch compound as well as preparation method and application thereof
A rice resistant starch complex was constructed by antisolvent precipitation, octenyl succinic anhydride modification, and enzyme-catalyzed cross-linking reaction. This solved the problems of low resistant starch content and complex process in existing technologies, and achieved efficient and green preparation and multiple mechanisms to delay starch degradation. It is suitable for low-GI staple foods and functional foods.
Patent Information
- Application Number
- CN202511470515.2
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-10-15
- Publication Date
- 2025-12-05
AI Technical Summary
Existing technologies cannot increase the resistant starch content in rice through green processing modification techniques, and physical encapsulation technology has complex processes and risks of biotoxicity, and cannot effectively regulate gut health.
Starch nanoparticles were prepared by antisolvent precipitation. Quercetin was then modified with octenyl succinic anhydride and covalently grafted onto laccase, followed by a cross-linking reaction catalyzed by transglutaminase, to construct a dense protein network encapsulation layer, forming a rice resistant starch complex.
It significantly increases the content of resistant starch in rice, delays starch degradation through multiple mechanisms, and is suitable for low-GI staple foods, functional baked goods, and slow-release nutrient carriers. It is also green and environmentally friendly and suitable for industrial production.
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Figure CN121058905A_ABST
Abstract
Description
Technical Field
[0001] This invention belongs to the field of food starch modification technology, specifically relating to a rice resistant starch complex, its preparation method, and its application. Background Technology
[0002] Starch is a major energy source in the human diet, easily absorbed and rapidly broken down into glucose, leading to a sharp rise in postprandial blood glucose. Rice, a common staple food, is primarily composed of starch (approximately 85%). Therefore, regulating the digestibility of rice starch is beneficial for controlling postprandial blood glucose levels, thereby preventing obesity-related metabolic diseases. Studies have shown that foods rich in resistant starch (RS) can prolong satiety while reducing glucose absorption and insulin requirements, demonstrating dual benefits in blood glucose management and gut microbiota regulation. Therefore, developing starch-based foods with high RS content has become a major approach to lowering the glycemic index.
[0003] In recent years, the preparation of resistant starch (RS4 type) through chemical modification has become a research hotspot, as the introduction of hydrophobic groups can enhance the starch's resistance to enzymatic hydrolysis. For the functional design of starch groups, physical encapsulation technology delays starch digestion by hindering amylase contact. However, existing technologies mostly employ liposomes or synthetic polymers for encapsulation, which are complex processes and pose risks of biotoxicity. Enzyme-mediated biocatalytic synthesis has advantages such as high synthesis efficiency, good reaction condition specificity, and environmental friendliness, but its application in nanoparticle interface modification has not yet been achieved. Therefore, how to increase the resistant starch content in rice through green processing modification technology, thereby regulating gut health, has become a key technological breakthrough. Summary of the Invention
[0004] The main objective of this invention is to provide a rice resistant starch complex, its preparation method, and its application, in order to overcome the shortcomings of the prior art.
[0005] To achieve the aforementioned objectives, the technical solution adopted by this invention includes:
[0006] The first aspect of the present invention provides a method for preparing a rice resistant starch complex, comprising:
[0007] Starch nanoparticles were prepared by antisolvent precipitation of rice starch, and then esterified with octenyl succinic anhydride (OSA) to obtain octenyl succinic anhydride starch nanoparticles.
[0008] Quercetin (QC) was covalently grafted onto the surface of the octenyl succinic anhydride starch nanoparticles using laccase to obtain octenyl succinic anhydride starch-quercetin composite nanoparticles.
[0009] A rice resistant starch complex was prepared by using transglutaminase to catalyze a cross-linking reaction between soybean protein isolate (SPI) and octenyl succinic anhydride starch-quercetin composite nanoparticles, thereby constructing a dense protein network encapsulation layer on the surface of the octenyl succinic anhydride starch-quercetin composite nanoparticles.
[0010] A second aspect of the present invention provides a rice resistant starch complex prepared by the above preparation method, the rice resistant starch complex comprising starch nanoparticles, an interface layer coated on the starch nanoparticles, and a protein coating layer coated on the interface layer.
[0011] In some embodiments, the resistant starch content in the rice resistant starch is 40.75% to 50.87%.
[0012] A third aspect of the present invention provides the application of the above-mentioned rice resistant starch complex in the preparation of low glycemic index foods, functional baked goods, or slow-release nutrient carriers.
[0013] Compared with the prior art, the present invention has at least the following beneficial effects:
[0014] (1) In the preparation method of rice resistant starch complex provided by the present invention, OSA is modified to introduce hydrophobic groups and negatively charged surfaces, which inhibits digestive enzyme adsorption and hinders glycosidic bond hydrolysis; laccase catalyzes quercetin covalently grafted to the OSA nanoparticle interface, which delays starch degradation through a dual mechanism of polyphenol enzyme inhibition activity and steric hindrance; the protein layer cross-linked with glutamine transaminase constructs a dense physical barrier, which blocks enzyme penetration and enhances structural stability in the gastrointestinal environment. The resistant starch content of rice starch is increased through the synergistic effect of chemical modification, enzyme catalytic cross-linking and physical barrier; moreover, the entire preparation process uses all food-grade raw materials and green enzymatic methods, which is green and environmentally friendly and suitable for large-scale industrial production.
[0015] (2) The rice resistant starch complex prepared by the present invention has a triple synergistic structure of "hydrophobic core - covalent cross-linked intermediate layer - protein physical outer layer". Through multiple mechanisms such as enzyme inhibition, spatial barrier and dense barrier, it significantly enhances the anti-digestion stability, which is of great significance for improving intestinal flora and human health. It can be widely used in low-GI staple foods, functional baked foods and slow-release nutrient carriers, and has significant health and industrialization value. Attached Figure Description
[0016] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments recorded in the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.
[0017] Figure 1 This is a schematic diagram of a glucose standard curve in a typical embodiment of the present invention. Detailed Implementation
[0018] In view of the problems existing in the prior art, the inventors of this invention, through extensive and in-depth research, provide a rice resistant starch complex, its preparation method, and its application. The main method involves using a double-layer encapsulation technique to encapsulate nanoparticles, thereby slowing down the digestion rate after consumption. OSA starch nanoparticles are prepared using an antisolvent method and OSA modification. Then, quercetin is covalently grafted onto the surface of the OSA starch nanoparticles using laccase. Finally, soy protein isolate is added, and a cross-linking reaction catalyzed by transglutaminase is formed, creating a dense protein network encapsulation layer on the surface of the composite particles, thus forming the rice resistant starch complex.
[0019] The following will provide a further explanation of the technical solution, its implementation process, and its principles.
[0020] The first aspect of the present invention provides a method for preparing a rice resistant starch complex comprising:
[0021] Starch nanoparticles were prepared by antisolvent precipitation of rice starch, and then esterified with octenyl succinic anhydride to obtain octenyl succinic anhydride starch nanoparticles.
[0022] The octenyl succinic anhydride starch-quercetin composite nanoparticles were prepared by covalently grafting quercetin onto the surface of the octenyl succinic anhydride starch nanoparticles using laccase.
[0023] A rice resistant starch complex was prepared by using transglutaminase to catalyze a cross-linking reaction between soy protein isolate and octenyl succinic anhydride starch-quercetin composite nanoparticles, thereby constructing a dense protein network encapsulation layer on the surface of the octenyl succinic anhydride starch-quercetin composite nanoparticles.
[0024] In some embodiments, the preparation method of the rice resistant starch complex specifically includes: ultrasonically treating a rice starch solution and then adding it to an antisolvent, centrifuging and precipitating to obtain the starch nanoparticles, then preparing a starch nanoparticle solution and adjusting it to a first pH value, adding octenyl succinic anhydride to undergo an esterification reaction, and then adjusting it to a second pH value to obtain the octenyl succinic anhydride starch nanoparticles.
[0025] Furthermore, the concentration of the rice starch solution is 2-15% (w / v).
[0026] Furthermore, the ultrasonic treatment power is 200-400W, and the time is 15-60min.
[0027] Furthermore, the solvent in the antisolvent precipitation method includes water, and the antisolvent includes ethanol.
[0028] Furthermore, the concentration of the starch nanoparticle solution is 1–10% (w / v).
[0029] Furthermore, the first pH value is 8.4 to 8.6, and the second pH value is 6.4 to 6.6.
[0030] Furthermore, the amount of octenyl succinic anhydride added is 1-3% of the dry starch mass.
[0031] Furthermore, the amount of octenyl succinic anhydride added is 3% of the dry starch mass.
[0032] Furthermore, the esterification reaction is carried out at a temperature of 34–36°C for 4–6 hours.
[0033] In the above process, by gelatinizing and cooling rice starch in a water bath, followed by ultrasonic treatment and precipitation with an antisolvent (ethanol), nanoparticles are formed, significantly increasing the structural density of the starch and effectively reducing the accessibility of digestive enzymes. Subsequently, through esterification with OSA, hydrophobic octene and carboxylic acid groups are introduced into the starch molecular chain. The hydrophobic interaction and electrostatic repulsion synergistically prevent enzyme molecules from approaching the glycosidic bond. At the same time, the stable network formed by chemical modification improves the starch's resistance to digestion, thus transforming it into heat-resistant RS4-type resistant starch.
[0034] In some embodiments, the preparation method of the rice resistant starch complex specifically includes: adding the quercetin to an octenyl succinic anhydride starch nanoparticle suspension for a first incubation, then adding the laccase for a second incubation, followed by a first enzyme inactivation, centrifugation precipitation, and dialysis to obtain the octenyl succinic anhydride starch-quercetin composite nanoparticles.
[0035] Furthermore, the mass ratio of quercetin to octenyl succinic anhydride starch nanoparticles is 1 to 3:1.
[0036] Furthermore, the mass ratio of quercetin to octenyl succinic anhydride starch nanoparticles is 1:1.
[0037] Furthermore, each gram of the octenyl succinic anhydride starch nanoparticles uses 50-150 U of laccase.
[0038] Furthermore, the octenyl succinic anhydride starch nanoparticle suspension is weakly acidic, with a pH value of 4.4–4.6.
[0039] Furthermore, the temperature for the first incubation is 28–32°C, and the time is 23–25 hours.
[0040] Furthermore, the second incubation temperature is 43–47°C, and the time is 14–16 hours.
[0041] Furthermore, the temperature for the first enzyme inactivation is 80–100°C, and the time is 10–30 min.
[0042] Furthermore, the dialysis time is 24–72 hours.
[0043] In the above process, under weakly acidic conditions, laccase oxidizes the catechol groups of quercetin to generate active quinone radicals, which covalently cross-link with the carboxyl or hydroxyl groups on the starch surface, forming a dense and stable interfacial layer. This layer not only hinders α-amylase from approaching the starch core through steric hindrance and hydrophobic interactions, but quercetin's own enzyme-inhibiting activity can also directly inhibit digestive enzyme activity, significantly reducing the starch hydrolysis rate through this dual mechanism. Simultaneously, the covalently bonded antioxidant interface protects the starch structure from oxidative degradation, synergistically enhancing the physical stability and digestibility of the particles, ultimately increasing the resistant starch content.
[0044] In some embodiments, the preparation method of the rice resistant starch complex specifically includes: dissolving the soy protein isolate in a buffer solution to obtain a protein solution, adding the octenyl succinic anhydride starch-quercetin composite nanoparticles to the protein solution, stirring and sonicating, and then adding the transglutaminase for a water bath and a second enzyme inactivation to obtain the rice resistant starch complex.
[0045] Furthermore, the concentration of the protein solution is 3-7% (w / v).
[0046] Furthermore, the mass ratio of the octenyl succinic anhydride starch-quercetin composite nanoparticles to soybean protein isolate is 1:0.5-3.
[0047] Furthermore, 5–30 U of transglutaminase is used per gram of soy protein isolate.
[0048] Furthermore, 20U of transglutaminase is used per gram of soy protein isolate.
[0049] Further, the buffer solution comprises a phosphate buffer. In a typical embodiment, the buffer solution is a 0.1M pH 7.0 phosphate buffer.
[0050] Furthermore, the stirring time is 20–120 min, the ultrasonic treatment power is 100–300 W, and the time is 5–40 min.
[0051] Furthermore, the water bath lasts for 3 to 6 hours and the temperature is 48 to 52°C.
[0052] Furthermore, the second enzyme inactivation temperature is 80–100°C, and the time is 10–40 min.
[0053] In the above process, a dense and stable protein coating layer is constructed on the surface of OSA starch-quercetin nanoparticles by the formation of an isopeptide cross-linking network of glutamine transaminase-catalyzed SPI between glutamine and lysine residues. This protein layer restricts the penetration contact of α-amylase to the starch core through a physical barrier effect, while the cross-linking network enhances the mechanical strength and shear resistance of the particles, reducing the risk of structural disintegration during digestion. In addition, the hydrophobic microdomains of SPI and quercetin synergistically inhibit enzyme activity, further delaying starch hydrolysis. This multi-layered protective mechanism enables the rice resistant starch composite particles to maintain structural integrity in a simulated gastrointestinal environment, resulting in a significant increase in resistant starch content.
[0054] In some more specific embodiments, the preparation method of the rice resistant starch complex specifically includes the following steps:
[0055] S1. Preparation of OSA starch nanoparticles: Rice starch solution was magnetically stirred in a water bath at 80°C. After cooling, the solution was ultrasonically treated in an ice bath at 25°C for 15–60 min at a power of 200–400 W. Then, anhydrous ethanol was added dropwise under continuous stirring. The precipitate was collected by centrifugation and dried to obtain starch nanoparticles. A starch nanoparticle solution was prepared and the first pH value of the solution was adjusted to 8.4–8.6. Then, OSA solution was added and esterification reaction was carried out at 34–36°C for 4–6 h. The second pH value of the esterified solution was adjusted to 6.4–6.6 to terminate the reaction and centrifuged and dried to obtain OSA starch nanoparticles. The concentration of the rice starch solution was 2–15% (w / v), the concentration of the starch nanoparticle solution was 1–10% (w / v), and the amount of octenyl succinic anhydride added was 1–3% of the dry weight of starch.
[0056] S2. Preparation of OSA starch-quercetin nanoparticles under a weakly acidic environment (pH = 4.4–4.6): Quercetin (QC) was added to the OSA starch nanoparticle suspension, and incubated at 28–32°C for 23–25 h. Then, laccase was added, and the mixture was incubated at 43–47°C for 14–16 h. After inactivating the enzyme at 80–100°C for 10–30 min, the precipitate was centrifuged and resuspended in deionized water. The mixture was dialyzed for 24–72 h and then freeze-dried to obtain OSA starch-quercetin nanoparticle powder. The mass ratio of quercetin to octenyl succinic anhydride starch nanoparticles was 1–3:1, and 50–150 U of laccase was used per gram of octenyl succinic anhydride starch nanoparticles.
[0057] S3. Preparation of low-digestibility rice resistant starch complex: Soy protein isolate (SPI) was dissolved in phosphate buffer to prepare a protein solution, which was stirred in a 50°C water bath for 1 hour. OSA starch-quercetin nanoparticles were added to the SPI solution and magnetically stirred for 20–120 min. Then, the mixture was sonicated at 100–300 W (ice bath) for 5–40 min. Glutamine transaminase was then added and the mixture was incubated in a 48–52°C water bath for 3–6 h. After inactivation, the enzyme was inactivated at 80–100°C for 10–40 min. The mixture was then centrifuged and freeze-dried to obtain the composite particles. The concentration of the protein solution was 3–7% (w / v). The mass ratio of the octenyl succinic anhydride starch-quercetin composite nanoparticles to the soy protein isolate was 1:0.5–3. 5–30 U of glutamine transaminase was used per gram of soy protein isolate.
[0058] In step S1, the first pH value is preferably 8.5, and the second pH value is preferably 6.5.
[0059] In step S2, the first incubation temperature is preferably 30°C for 24 hours, and the second incubation temperature is preferably 45°C for 15 hours.
[0060] The second aspect of the present invention provides a rice resistant starch complex prepared by the above preparation method, wherein the rice resistant starch complex comprises starch nanoparticles, an interface layer coated on the starch nanoparticles, and a protein coating layer coated on the interface layer.
[0061] In some embodiments, the resistant starch content in the rice resistant starch complex is 40.75% to 50.87%.
[0062] In some embodiments, the interface layer is a covalently cross-linked structure, and the protein coating layer is an isopeptide cross-linked network structure.
[0063] The third aspect of the present invention provides the application of the above-mentioned rice resistant starch complex in the preparation of low glycemic index foods, functional baked foods, or slow-release nutrient carriers.
[0064] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the invention. Modifications or equivalent substitutions made by those skilled in the art based on their understanding of the technical solutions of this invention, without departing from the spirit and scope of the invention, should be covered within the protection scope of this invention.
[0065] For experiments not specifically described in the examples, the procedures or conditions can be performed according to the conventional experimental procedures described in the literature in this field. Reagents or instruments whose manufacturers are not specified are all commercially available. Other unmentioned raw materials and instruments are all conventionally chosen and do not involve the core technical means of this invention.
[0066] For example, the rice starch used in the embodiments and comparative examples of the present invention was purchased from Shanghai Yuanye Biotechnology Co., Ltd.
[0067] Octenyl succinic anhydride, model number O160036, is sourced from Shanghai Aladdin Biochemical Technology Co., Ltd.
[0068] The quercetin, model number S25567, is sourced from Shanghai Yuanye Biotechnology Co., Ltd.
[0069] Laccase is sourced from Shanghai Yuanye Biotechnology Co., Ltd.
[0070] The transglutaminase was obtained from Shanghai Yuanye Biotechnology Co., Ltd.
[0071] Soy protein isolate is sourced from Shanghai Yuanye Biotechnology Co., Ltd.
[0072] Example 1
[0073] (1) Add 3g of rice starch to 100ml of 0.1M sodium hydroxide solution, preheat (80℃), and magnetically stir for 30min. After cooling, place the starch suspension in an ultrasonic cell disruptor and ultrasonically treat the solution (300W) at 25℃ using an ice bath for 30min. Then, add anhydrous ethanol dropwise at a ratio of 1:2 with continuous stirring. Subsequently, centrifuge at 3000g for 15min, collect the precipitate, and dry to obtain starch nanoparticles. Prepare a 3% (w / v) starch nanosolution and magnetically stir at 35℃ for 30min. Adjust the pH of the solution (2M NaOH) to 8.5. Add 3% OSA (based on the mass of starch nanoparticles) diluted 5 times with anhydrous ethanol to the starch nanoparticle suspension. After esterification reaction for 4h, adjust the pH of the esterified solution to 6.5 with 1M hydrochloric acid solution to terminate the reaction, centrifuge, and dry to obtain OSA starch nanoparticles (OS).
[0074] (2) Quercetin (QC) (quercetin to OSA starch nanoparticles mass ratio 1:1) was added to a 3% (w / v) OSA starch nanoparticle suspension (pH = 4.5). After shaking at 30°C for 24 hours, laccase (50 U / g based on dry starch) was added to initiate the enzyme reaction. The mixture was incubated at 45°C at 120 rpm for 15 hours. The enzyme was inactivated by water bath at 80°C for 10 min. After cooling to room temperature, the mixture was centrifuged, and the precipitate was resuspended in deionized water. The precipitate was then dialyzed for 48 hours (water changed every 6 hours) and then freeze-dried to obtain OSA starch-quercetin nanoparticle powder (OS-QC).
[0075] (3) Soy protein isolate powder (SPI) was dissolved in 0.1M phosphate buffer at pH 7.0 to prepare a 5% (w / v) protein solution, and stirred in a 50°C water bath for 1 hour. OSA starch-quercetin nanoparticles were added to the SPI solution at a 1:1 mass ratio, magnetically stirred for 30 minutes, and then sonicated (300W power, 20kHz frequency, ice bath) for 5 minutes. Then, transglutaminase (20U / g soy protein isolate) was added and stirred in a 50°C water bath for 4 hours. The enzyme was inactivated by heating at 85°C for 15 minutes, cooled to room temperature, centrifuged, and freeze-dried to obtain composite particles (OS-QC-SPI).
[0076] Example 2
[0077] (1) Add 3g of rice starch to 100ml of 0.1M sodium hydroxide solution, preheat (80℃), and magnetically stir for 30min. After cooling, place the starch suspension in an ultrasonic cell disruptor and ultrasonically treat the solution (300W) at 25℃ using an ice bath for 30min. Then, add anhydrous ethanol dropwise at a ratio of 1:2 with continuous stirring. Subsequently, centrifuge at 3000g for 15min, collect the precipitate, and dry to obtain starch nanoparticles. Prepare a 3% (w / v) starch nanosolution and magnetically stir at 35℃ for 30min. Adjust the pH of the solution (2M NaOH) to 8.5. Add 3% OSA (based on the mass of starch nanoparticles) diluted 5 times with anhydrous ethanol to the starch nanoparticle suspension. After esterification reaction for 4h, adjust the pH of the esterified solution to 6.5 with 1M hydrochloric acid solution to terminate the reaction, centrifuge, and dry to obtain OSA starch nanoparticles (OS).
[0078] (2) Quercetin (QC) (quercetin to OSA starch nanoparticles mass ratio 1:1) was added to a 3% (w / v) OSA starch nanoparticle suspension (pH = 4.5). After shaking at 30°C for 24 hours, laccase (100 U / g based on dry starch) was added to initiate the enzyme reaction. The mixture was incubated at 45°C at 120 rpm for 15 hours. The enzyme was then inactivated by water bath at 80°C for 10 minutes. After cooling to room temperature, the mixture was centrifuged, and the precipitate was resuspended in deionized water. The precipitate was then dialyzed for 48 hours (water changed every 6 hours) and then freeze-dried to obtain OSA starch-quercetin nanoparticle powder (OS-QC).
[0079] (3) Soy protein isolate powder (SPI) was dissolved in 0.1M phosphate buffer at pH 7.0 to prepare a 5% (w / v) protein solution, and stirred in a 50°C water bath for 1 hour. OSA starch-quercetin nanoparticles were added to the SPI solution at a 1:1 mass ratio, magnetically stirred for 30 minutes, and then sonicated (300W power, 20kHz frequency, ice bath) for 5 minutes. Then, transglutaminase (20U / g soy protein isolate) was added and stirred in a 50°C water bath for 4 hours. The enzyme was inactivated by heating at 85°C for 15 minutes, cooled to room temperature, centrifuged, and freeze-dried to obtain composite particles (OS-QC-SPI).
[0080] Example 3
[0081] (1) Add 3g of rice starch to 100ml of 0.1M sodium hydroxide solution, preheat (80℃), and magnetically stir for 30min. After cooling, place the starch suspension in an ultrasonic cell disruptor and ultrasonically treat the solution (300W) at 25℃ using an ice bath for 30min. Then, add anhydrous ethanol dropwise at a ratio of 1:2 with continuous stirring. Subsequently, centrifuge at 3000g for 15min, collect the precipitate, and dry to obtain starch nanoparticles. Prepare a 3% (w / v) starch nanosolution and magnetically stir at 35℃ for 30min. Adjust the pH of the solution (2M NaOH) to 8.5. Add 3% OSA (based on the mass of starch nanoparticles) diluted 5 times with anhydrous ethanol to the starch nanoparticle suspension. After esterification reaction for 4h, adjust the pH of the esterified solution to 6.5 with 1M hydrochloric acid solution to terminate the reaction, centrifuge, and dry to obtain OSA starch nanoparticles (OS).
[0082] (2) Quercetin (QC) (quercetin to OSA starch nanoparticles mass ratio 1:1) was added to a 3% (w / v) OSA starch nanoparticle suspension (pH = 4.5). After shaking at 30°C for 24 hours, laccase (150 U / g based on dry starch) was added to initiate the enzyme reaction. The mixture was incubated at 45°C at 120 rpm for 15 hours. The enzyme was inactivated by water bath at 80°C for 10 min. After cooling to room temperature, the mixture was centrifuged, and the precipitate was resuspended in deionized water, placed in a dialysis bag, and dialyzed for 48 hours (changing the water every 6 hours). Then, the mixture was freeze-dried to obtain OSA starch-quercetin nanoparticle powder (OS-QC).
[0083] (3) Soy protein isolate powder (SPI) was dissolved in 0.1M phosphate buffer at pH 7.0 to prepare a 5% (w / v) protein solution, and stirred in a 50°C water bath for 1 hour. OSA starch-quercetin nanoparticles were added to the SPI solution at a 1:1 mass ratio, magnetically stirred for 30 minutes, and then sonicated (300W power, 20kHz frequency, ice bath) for 5 minutes. Then, transglutaminase (20U / g soy protein isolate) was added and stirred in a 50°C water bath for 4 hours. The enzyme was inactivated by heating at 85°C for 15 minutes, cooled to room temperature, centrifuged, and freeze-dried to obtain composite particles (OS-QC-SPI).
[0084] Example 4
[0085] (1) Add 3g of rice starch to 100ml of 0.1M sodium hydroxide solution, preheat (80℃), and magnetically stir for 30min. After cooling, place the starch suspension in an ultrasonic cell disruptor and ultrasonically treat the solution (300W) at 25℃ using an ice bath for 30min. Then, add anhydrous ethanol dropwise at a ratio of 1:2 with continuous stirring. Subsequently, centrifuge at 3000g for 15min, collect the precipitate, and dry to obtain starch nanoparticles. Prepare a 3% (w / v) starch nanosolution and magnetically stir at 35℃ for 30min. Adjust the pH of the solution (2M NaOH) to 8.5. Add 3% OSA (based on the mass of starch nanoparticles) diluted 5 times with anhydrous ethanol to the starch nanoparticle suspension. After esterification reaction for 4h, adjust the pH of the esterified solution to 6.5 with 1M hydrochloric acid solution to terminate the reaction, centrifuge, and dry to obtain OSA starch nanoparticles (OS).
[0086] (2) Quercetin (QC) (quercetin to OSA starch nanoparticles mass ratio 1:1) was added to a 3% (w / v) OSA starch nanoparticle suspension (pH = 4.5). After shaking at 30°C for 24 hours, laccase (100 U / g based on dry starch) was added to initiate the enzyme reaction. The mixture was incubated at 45°C at 120 rpm for 15 hours. The enzyme was then inactivated by water bath at 80°C for 10 minutes. After cooling to room temperature, the mixture was centrifuged, and the precipitate was resuspended in deionized water. The precipitate was then dialyzed for 48 hours (water changed every 6 hours) and then freeze-dried to obtain OSA starch-quercetin nanoparticle powder (OS-QC).
[0087] (3) Soy protein isolate powder (SPI) was dissolved in 0.1M phosphate buffer at pH 7.0 to prepare a 5% (w / v) protein solution, and stirred in a 50°C water bath for 1 hour. OSA starch-quercetin nanoparticles were added to the SPI solution at a mass ratio of 1:0.5, and the mixture was magnetically stirred for 30 minutes, followed by sonication (300W power, 20kHz frequency, ice bath) for 5 minutes. Then, transglutaminase (20U / g soy protein isolate) was added and stirred in a 50°C water bath for 4 hours. The enzyme was inactivated by heating at 85°C for 15 minutes, cooled to room temperature, centrifuged, and freeze-dried to obtain composite particles (OS-QC-SPI).
[0088] Example 5
[0089] (1) Add 3g of rice starch to 100ml of 0.1M sodium hydroxide solution, preheat (80℃), and magnetically stir for 30min. After cooling, place the starch suspension in an ultrasonic cell disruptor and ultrasonically treat the solution (300W) at 25℃ using an ice bath for 30min. Then, add anhydrous ethanol dropwise at a ratio of 1:2 with continuous stirring. Subsequently, centrifuge at 3000g for 15min, collect the precipitate, and dry to obtain starch nanoparticles. Prepare a 3% (w / v) starch nanosolution and magnetically stir at 35℃ for 30min. Adjust the pH of the solution (2M NaOH) to 8.5. Add 3% OSA (based on the mass of starch nanoparticles) diluted 5 times with anhydrous ethanol to the starch nanoparticle suspension. After esterification reaction for 4h, adjust the pH of the esterified solution to 6.5 with 1M hydrochloric acid solution to terminate the reaction, centrifuge, and dry to obtain OSA starch nanoparticles (OS).
[0090] (2) Quercetin (QC) (quercetin to OSA starch nanoparticles mass ratio 1:1) was added to a 3% (w / v) OSA starch nanoparticle suspension (pH = 4.5). After shaking at 30°C for 24 hours, laccase (100 U / g based on dry starch) was added to initiate the enzyme reaction. The mixture was incubated at 45°C at 120 rpm for 15 hours. The enzyme was then inactivated by water bath at 80°C for 10 minutes. After cooling to room temperature, the mixture was centrifuged, and the precipitate was resuspended in deionized water. The precipitate was then dialyzed for 48 hours (water changed every 6 hours) and then freeze-dried to obtain OSA starch-quercetin nanoparticle powder (OS-QC).
[0091] (3) Soy protein isolate powder (SPI) was dissolved in 0.1M phosphate buffer (pH 7.0) to prepare a 5% (w / v) protein solution, and stirred in a 50°C water bath for 1 hour. OSA starch-quercetin nanoparticles were added to the SPI solution at a mass ratio of 1:2, and the mixture was magnetically stirred for 30 minutes, followed by sonication (300W power, 20kHz frequency, ice bath) for 5 minutes. Then, transglutaminase (20U / g soy protein isolate) was added and stirred in a 50°C water bath for 4 hours. The enzyme was inactivated by heating at 85°C for 15 minutes, cooled to room temperature, centrifuged, and freeze-dried to obtain composite particles (OS-QC-SPI).
[0092] Example 6
[0093] (1) Add 3g of rice starch to 100ml of 0.1M sodium hydroxide solution, preheat (80℃), and magnetically stir for 30min. After cooling, place the starch suspension in an ultrasonic cell disruptor and ultrasonically treat the solution (300W) at 25℃ using an ice bath for 30min. Then, add anhydrous ethanol dropwise at a ratio of 1:2 with continuous stirring. Subsequently, centrifuge at 3000g for 15min, collect the precipitate, and dry to obtain starch nanoparticles. Prepare a 3% (w / v) starch nanosolution and magnetically stir at 35℃ for 30min. Adjust the pH of the solution (2M NaOH) to 8.5. Add 3% OSA (based on the mass of starch nanoparticles) diluted 5 times with anhydrous ethanol to the starch nanoparticle suspension. After esterification reaction for 4h, adjust the pH of the esterified solution to 6.5 with 1M hydrochloric acid solution to terminate the reaction, centrifuge, and dry to obtain OSA starch nanoparticles (OS).
[0094] (2) Quercetin (QC) (quercetin to OSA starch nanoparticles mass ratio 1:1) was added to a 3% (w / v) OSA starch nanoparticle suspension (pH = 4.5). After shaking at 30°C for 24 hours, laccase (100 U / g based on dry starch) was added to initiate the enzyme reaction. The mixture was incubated at 45°C at 120 rpm for 15 hours. The enzyme was then inactivated by water bath at 80°C for 10 minutes. After cooling to room temperature, the mixture was centrifuged, and the precipitate was resuspended in deionized water. The precipitate was then dialyzed for 48 hours (water changed every 6 hours) and then freeze-dried to obtain OSA starch-quercetin nanoparticle powder (OS-QC).
[0095] (3) Soy protein isolate powder (SPI) was dissolved in 0.1M phosphate buffer (pH 7.0) to prepare a 5% (w / v) protein solution, and stirred in a 50°C water bath for 1 hour. OSA starch-quercetin nanoparticles were added to the SPI solution at a mass ratio of 1:3, and the mixture was magnetically stirred for 30 minutes, followed by sonication (300W power, 20kHz frequency, ice bath) for 5 minutes. Then, transglutaminase (20U / g soy protein isolate) was added and stirred in a 50°C water bath for 4 hours. The enzyme was inactivated by heating at 85°C for 15 minutes, cooled to room temperature, centrifuged, and freeze-dried to obtain composite particles (OS-QC-SPI).
[0096] Comparative Example 1
[0097] (1) Add 3g of rice starch to 100ml of 0.1M sodium hydroxide solution, preheat (80℃), and magnetically stir for 30min. After cooling, place the starch suspension in an ultrasonic cell disruptor and ultrasonically treat the solution (300W) at 25℃ using an ice bath for 30min. Then, add anhydrous ethanol dropwise at a ratio of 1:2 with continuous stirring. Subsequently, centrifuge at 3000g for 15min, collect the precipitate, and dry to obtain starch nanoparticles. Prepare a 3% (w / v) starch nanosolution and magnetically stir at 35℃ for 30min. Adjust the pH of the solution (2M NaOH) to 8.5. Add 3% OSA (based on the mass of starch nanoparticles) diluted 5 times with anhydrous ethanol to the starch nanoparticle suspension. After esterification reaction for 4h, adjust the pH of the esterified solution to 6.5 with 1M hydrochloric acid solution to terminate the reaction, centrifuge, and dry to obtain OSA starch nanoparticles.
[0098] (2) Quercetin (QC) (the mass ratio of quercetin to OSA starch nanoparticles was 1:1) was added to a 3% (w / v) OSA starch nanoparticle suspension (pH=4.5). After shaking at 30°C for 24 hours, the mixture was incubated at 45°C at 120 rpm for 15 hours. After cooling to room temperature, the mixture was centrifuged, and the precipitate was resuspended in deionized water. The precipitate was then placed in a dialysis bag and dialyzed for 48 hours (with water changed every 6 hours). Finally, the mixture was freeze-dried to obtain OSA starch-quercetin nanoparticles.
[0099] (3) Soy protein isolate powder (SPI) was dissolved in 0.1M phosphate buffer (pH 7.0) to prepare a 5% (w / v) protein solution, and stirred in a 50°C water bath for 1 hour. OSA starch-quercetin nanoparticles were added to the SPI solution at a mass ratio of 1:2, magnetically stirred for 30 minutes, and then sonicated (300W power, 20kHz frequency, ice bath) for 5 minutes. The mixture was then stirred in a 50°C water bath for 4 hours, cooled to room temperature, centrifuged, and freeze-dried to obtain composite particles.
[0100] Comparative Example 2
[0101] (1) Add 3g of rice starch to 100ml of 0.1M sodium hydroxide solution, preheat (80℃), and magnetically stir for 30min. After cooling, place the starch suspension in an ultrasonic cell disruptor and ultrasonically treat the solution (300W) at 25℃ using an ice bath for 30min. Then, add anhydrous ethanol dropwise at a ratio of 1:2 with continuous stirring. Subsequently, centrifuge at 3000g for 15min, collect the precipitate, and dry to obtain starch nanoparticles. Prepare a 3% (w / v) starch nanosolution and magnetically stir at 35℃ for 30min. Adjust the pH of the solution (2M NaOH) to 8.5. Add 3% OSA (based on the mass of starch nanoparticles) diluted 5 times with anhydrous ethanol to the starch nanoparticle suspension. After esterification reaction for 4h, adjust the pH of the esterified solution to 6.5 with 1M hydrochloric acid solution to terminate the reaction, centrifuge, and dry to obtain OSA starch nanoparticles.
[0102] (2) Quercetin (QC) (the mass ratio of quercetin to OSA starch nanoparticles was 1:1) was added to a 3% (w / v) OSA starch nanoparticle suspension (pH=4.5). After shaking at 30°C for 24 hours, the mixture was incubated at 45°C at 120 rpm for 15 hours. After cooling to room temperature, the mixture was centrifuged, and the precipitate was resuspended in deionized water. The precipitate was then placed in a dialysis bag and dialyzed for 48 hours (with water changed every 6 hours). Finally, the mixture was freeze-dried to obtain OSA starch-quercetin nanoparticles.
[0103] (3) Soy protein isolate powder (SPI) was dissolved in 0.1M phosphate buffer (pH 7.0) to prepare a 5% (w / v) protein solution, and stirred in a 50°C water bath for 1 hour. OSA starch-quercetin nanoparticles were added to the SPI solution at a mass ratio of 1:2, and the mixture was magnetically stirred for 30 minutes, followed by sonication (300W power, 20kHz frequency, ice bath) for 5 minutes. Then, transglutaminase (20U / g soy protein isolate) was added and stirred in a 50°C water bath for 4 hours. The enzyme was inactivated by heating at 85°C for 15 minutes, cooled to room temperature, centrifuged, and freeze-dried to obtain composite particles.
[0104] Comparative Example 3
[0105] (1) Add 3g of rice starch to 100ml of 0.1M sodium hydroxide solution, preheat (80℃), and magnetically stir for 30min. After cooling, place the starch suspension in an ultrasonic cell disruptor and ultrasonically treat the solution (300W) at 25℃ using an ice bath for 30min. Then, add anhydrous ethanol dropwise at a ratio of 1:2 with continuous stirring. Subsequently, centrifuge at 3000g for 15min, collect the precipitate, and dry it to obtain ordinary starch nanoparticles.
[0106] (2) Quercetin (QC) (quercetin to ordinary starch nanoparticles mass ratio 1:1) was added to a 3% (w / v) suspension of ordinary starch nanoparticles (pH=4.5). After shaking at 30℃ for 24 hours, laccase (100 U / g based on dry starch) was added to initiate the enzyme reaction. The mixture was incubated at 45℃ and 120 rpm for 15 hours. The enzyme was inactivated by water bath at 80℃ for 10 min. After cooling to room temperature, the mixture was centrifuged, and the precipitate was resuspended in deionized water, placed in a dialysis bag, and dialyzed for 48 hours (changing the water every 6 hours). Then, the mixture was freeze-dried to obtain ordinary starch-quercetin nanoparticles.
[0107] (3) Soy protein isolate powder (SPI) was dissolved in 0.1M phosphate buffer (pH 7.0) to prepare a 5% (w / v) protein solution, and stirred in a 50°C water bath for 1 hour. Ordinary starch-quercetin nanoparticles were added to the SPI solution at a mass ratio of 1:2, and the mixture was magnetically stirred for 30 minutes, followed by sonication (300W power, 20kHz frequency, ice bath) for 5 minutes. Then, transglutaminase (20U / g soy protein isolate) was added and stirred in a 50°C water bath for 4 hours. The enzyme was inactivated by heating at 85°C for 15 minutes, cooled to room temperature, centrifuged, and freeze-dried to obtain composite particles.
[0108] Comparative Example 4
[0109] (1) Add 3g of rice starch to 100ml of 0.1M sodium hydroxide solution, preheat (80℃), and magnetically stir for 30min. After cooling, place the starch suspension in an ultrasonic cell disruptor and ultrasonically treat the solution (300W) at 25℃ using an ice bath for 30min. Then, add anhydrous ethanol dropwise at a ratio of 1:2 with continuous stirring. Subsequently, centrifuge at 3000g for 15min, collect the precipitate, and dry to obtain starch nanoparticles. Prepare a 3% (w / v) starch nanosolution and magnetically stir at 35℃ for 30min. Adjust the pH of the solution (2M NaOH) to 8.5. Add 3% OSA (based on the mass of starch nanoparticles) diluted 5 times with anhydrous ethanol to the starch nanoparticle suspension. After esterification reaction for 4h, adjust the pH of the esterified solution to 6.5 with 1M hydrochloric acid solution to terminate the reaction, centrifuge, and dry to obtain OSA starch nanoparticles.
[0110] (2) Quercetin (QC) (quercetin to OSA starch nanoparticles mass ratio 1:1) was added to a 3% (w / v) OSA starch nanoparticle suspension (pH = 4.5). After shaking at 30°C for 24 hours, laccase (100 U / g based on dry starch) was added to initiate the enzyme reaction. The mixture was incubated at 45°C at 120 rpm for 15 hours. The enzyme was inactivated by water bath at 80°C for 10 min. After cooling to room temperature, the mixture was centrifuged, and the precipitate was resuspended in deionized water. The precipitate was then dialyzed for 48 hours (water changed every 6 hours) and then freeze-dried to obtain OSA starch-quercetin nanoparticles.
[0111] (3) Soy protein isolate powder (SPI) was dissolved in 0.1M phosphate buffer (pH 7.0) to prepare a 5% (w / v) protein solution, and stirred in a 50°C water bath for 1 hour. OSA starch-quercetin nanoparticles were added to the SPI solution at a mass ratio of 1:2, magnetically stirred for 30 minutes, and then sonicated (300W power, 20kHz frequency, ice bath) for 5 minutes. The mixture was then stirred in a 50°C water bath for 4 hours, cooled to room temperature, centrifuged, and freeze-dried to obtain composite particles.
[0112] Comparative Example 5
[0113] (1) Add 3g of rice starch to 100ml of 0.1M sodium hydroxide solution, preheat (80℃), and magnetically stir for 30min. After cooling, place the starch suspension in an ultrasonic cell disruptor and ultrasonically treat the solution (300W) at 25℃ using an ice bath for 30min. Then, add anhydrous ethanol dropwise at a ratio of 1:2 with continuous stirring. Subsequently, centrifuge at 3000g for 15min, collect the precipitate, and dry to obtain starch nanoparticles. Prepare a 3% (w / v) starch nanosolution and magnetically stir at 35℃ for 30min. Adjust the pH of the solution (2M NaOH) to 8.5. Add 3% OSA (based on the mass of starch nanoparticles) diluted 5 times with anhydrous ethanol to the starch nanoparticle suspension. After esterification reaction for 4h, adjust the pH of the esterified solution to 6.5 with 1M hydrochloric acid solution to terminate the reaction, centrifuge, and dry to obtain OSA starch nanoparticles.
[0114] (2) Soy protein isolate powder (SPI) was dissolved in 0.1M phosphate buffer (pH 7.0) to prepare a 5% (w / v) protein solution, and stirred in a 50°C water bath for 1 hour. OSA starch nanoparticles were added to the SPI solution at a mass ratio of 1:2, and the mixture was magnetically stirred for 30 minutes, followed by sonication (300W power, 20kHz frequency, ice bath) for 5 minutes. Then, transglutaminase (20U / g soy protein isolate) was added and stirred in a 50°C water bath for 4 hours. The enzyme was inactivated by heating at 85°C for 15 minutes, cooled to room temperature, centrifuged, and freeze-dried to obtain composite particles.
[0115] Comparative Example 6
[0116] (1) Add 3g of rice starch to 100ml of 0.1M sodium hydroxide solution, preheat (80℃), and magnetically stir for 30min. After cooling, place the starch suspension in an ultrasonic cell disruptor and ultrasonically treat the solution (300W) at 25℃ using an ice bath for 30min. Then, add anhydrous ethanol dropwise at a ratio of 1:2 with continuous stirring. Subsequently, centrifuge at 3000g for 15min, collect the precipitate, and dry to obtain starch nanoparticles. Prepare a 3% (w / v) starch nanosolution and magnetically stir at 35℃ for 30min. Adjust the pH of the solution (2M NaOH) to 8.5. Add 3% OSA (based on the mass of starch nanoparticles) diluted 5 times with anhydrous ethanol to the starch nanoparticle suspension. After esterification reaction for 4h, adjust the pH of the esterified solution to 6.5 with 1M hydrochloric acid solution to terminate the reaction, centrifuge, and dry to obtain OSA starch nanoparticles.
[0117] (2) Quercetin (QC) (quercetin to OSA starch nanoparticles mass ratio 1:1) was added to a 3% (w / v) OSA starch nanoparticle suspension (pH = 4.5). After shaking at 30°C for 24 hours, laccase (100 U / g based on dry starch) was added to initiate the enzyme reaction. The mixture was incubated at 45°C at 120 rpm for 15 hours. The enzyme was then inactivated by water bath at 80°C for 10 minutes. After cooling to room temperature, the mixture was centrifuged, and the precipitate was resuspended in deionized water, placed in a dialysis bag, and dialyzed for 48 hours (changing the water every 6 hours). Finally, the mixture was freeze-dried.
[0118] Comparative Example 7
[0119] It uses natural rice starch that has not undergone any technical or method processing.
[0120] Performance testing
[0121] To further illustrate the technical effects of the present invention, measurements were performed on the related products obtained in Examples 1-6 and Comparative Examples 1-7.
[0122] The steps for plotting the glucose standard curve in the above process are as follows:
[0123] (1) Preparation of glucose standard solution: Weigh 50 mg of D(+)-anhydrous glucose standard, dissolve it in distilled water, transfer it to a volumetric flask and make up to 50 mL (1 mg / mL).
[0124] (2) Take six 50mL stoppered test tubes and accurately pipette 0, 0.2, 0.4, 0.6, 0.8, and 1.0mL of glucose standard solution, respectively. Then add 1.0, 0.8, 0.6, 0.4, 0.2, and 0mL of distilled water to each tube. Next, add 1mL of 3,5-dinitrosalicylic acid (DNS) reagent to each test tube. Incubate in a boiling water bath for 5 minutes, then cool to room temperature. Dilute to 50mL with distilled water and mix thoroughly. Using sample No. 1 as a blank, measure the absorbance of the solution at 540nm using a UV-Vis spectrophotometer. Plot a standard curve with glucose content on the x-axis and the corresponding absorbance value on the y-axis, and fit a linear equation, as shown below. Figure 1 As shown. The fitted equation is y = 0.2041x + 0.0054, and the correlation coefficient R0 is... 2 =0.9974.
[0125] In the above steps, 3,5-dinitrosalicylic acid (DNS) was used to determine the content of rapidly digestible starch (RDS), slowly digestible starch (SDS), and resistant starch (RS) in the sample, as follows:
[0126] (1) Weigh 200mg of starch sample and mix it with 20ml of sodium acetate buffer (0.2mol / L, pH=5.2) for 5min, then heat and stir in a boiling water bath for 20min.
[0127] (2) The gelatinized sample was continuously stirred at 200 rpm in a water bath at 37°C for 10 min to reach equilibrium.
[0128] (3) Take 5 mL of mixed enzyme (α-amylase and amyloglucosidase) solution and add it to the above solution for enzymatic hydrolysis. At 0 min, 20 min and 120 min, take 0.5 mL of hydrolysate and put it into a centrifuge tube containing 2 mL of anhydrous ethanol to terminate the enzyme reaction. After inactivating the enzyme, centrifuge at 4000 rpm for 15 minutes.
[0129] (4) After centrifugation, take 1 mL of supernatant and add 2.5 mL of DNS reagent. After boiling in a water bath for 5 minutes, cool to room temperature and use a UV-Vis spectrophotometer to measure the absorbance at 540 nm.
[0130] Calculate the content of rapidly digestible starch (RDS), slowly digestible starch (SDS), and resistant starch (RS) using the following formula.
[0131]
[0132] RS(%) = 1 - RDS% - SDS%
[0133] Among them, G0, G 20 and G 120The values represent the glucose content (mg) released by the sample during the intestinal digestion phase at 0 min, 20 min, and 120 min, respectively; Ts represents the total starch content (mg) of the sample.
[0134] The determination of total starch content refers to the acid hydrolysis method in GB 5009.9-2016 "Determination of Starch in Food".
[0135] Characterization:
[0136] Table 1 shows the starch composition of the rice starch samples prepared in Examples 1-6 and Comparative Examples 1-7.
[0137] Table 1
[0138]
[0139] As shown in Table 1, the rice starch prepared in Example 5 had the highest content of resistant components, with an RS content of 50.87%. In this study, the resistant starch content of all examples (40.75–50.87%) was significantly higher than that of untreated natural rice starch (Comparative Example 7, 25.43%), indicating that the synergistic strategy of OSA modification-laccase-catalyzed cross-linking-glutamine transaminase construction of a ternary complex can effectively enhance the formation of resistant starch. Among them, Example 5 (OS-QC to SPI mass ratio 1:2, laccase addition 100 U / g) showed the best effect, with a content of 50.87%. The comparison revealed that laccase catalysis is a key pre-step in the formation of a high-resistance complex, and its dosage has an optimal value (100 U / g is better than 50 U / g and 150 U / g); the composite ratio of OS-QC to SPI significantly affects the product structure, with a 1:2 ratio showing the best effect. Comparative analysis further showed that omitting any key process (e.g., Comparative Examples 1 and 2 did not use laccase, Comparative Examples 1 and 4 did not use transglutaminase, Comparative Example 3 did not undergo OSA modification, Comparative Example 5 did not add quercetin, and Comparative Example 6 did not combine with SPI) all resulted in a significant decrease in resistant starch content (27.57%–37.03%), confirming that OSA modification, covalent cross-linking of quercetin with laccase, and the combination of transglutaminase-mediated protein are all indispensable for constructing a stable enzymatically resistant structure and have a significant synergistic effect.
[0140] In summary, this invention improves the resistant starch content of rice starch through the synergistic effect of chemical modification, enzymatic cross-linking, and physical barriers. OSA modification introduces hydrophobic groups and a negatively charged surface, inhibiting digestive enzyme adsorption and hindering glycosidic bond hydrolysis. Laccase catalyzes the covalent grafting of quercetin onto the OSA nanoparticle interface, delaying starch degradation through a dual mechanism of polyphenol enzyme inhibition and steric hindrance. The protein layer cross-linked with glutamine transaminase constructs a dense physical barrier, blocking enzyme penetration and enhancing structural stability in the gastrointestinal environment. Furthermore, the high-resistant rice starch prepared by this invention employs a fully enzymatic process, eliminating chemical cross-linking agents and using food-grade raw materials, providing a safe, stable, and multifunctional solution for developing low-GI foods, applicable to large-scale industrial production.
[0141] In addition, the inventors of this case also conducted experiments with other raw materials, process operations, and process conditions described in this specification, referring to the aforementioned embodiments, and obtained relatively ideal results in all cases.
[0142] Although the invention has been described with reference to illustrative embodiments, those skilled in the art will understand that various other changes, omissions, and / or additions can be made without departing from the spirit and scope of the invention, and that elements of the embodiments can be substituted with substantially equivalents. Furthermore, many modifications can be made without departing from the scope of the invention to adapt particular situations or materials to the teachings of the invention. Therefore, this invention is not intended to be limited to the specific embodiments disclosed for carrying out the invention, but rather is intended to encompass all embodiments falling within the scope of the appended claims. Moreover, unless specifically stated otherwise, any use of the terms first, second, etc., does not indicate any order or importance, but is used to distinguish one element from another.
Claims
1. A method for preparing a rice resistant starch complex, characterized in that, include: Starch nanoparticles were prepared by antisolvent precipitation of rice starch, and then esterified with octenyl succinic anhydride to obtain octenyl succinic anhydride starch nanoparticles. The octenyl succinic anhydride starch-quercetin composite nanoparticles were prepared by covalently grafting quercetin onto the surface of the octenyl succinic anhydride starch nanoparticles using laccase. A rice resistant starch complex was prepared by using transglutaminase to catalyze a cross-linking reaction between soy protein isolate and octenyl succinic anhydride starch-quercetin composite nanoparticles, thereby constructing a dense protein network encapsulation layer on the surface of the octenyl succinic anhydride starch-quercetin composite nanoparticles.
2. The preparation method according to claim 1, characterized in that, Specifically, it includes: The rice starch solution was ultrasonically treated and then added to an antisolvent. The starch nanoparticles were obtained by centrifugation and precipitation. The starch nanoparticle solution was then prepared and adjusted to a first pH value. Octenyl succinic anhydride was added to induce an esterification reaction. The pH value was then adjusted to a second pH value to obtain the octenyl succinic anhydride starch nanoparticles.
3. The preparation method according to claim 2, characterized in that: The concentration of the rice starch solution is 2-15% (w / v); And / or, the power of the ultrasonic treatment is 200~400W, and the time is 15~60min; And / or, the solvent in the antisolvent precipitation method includes water, and the antisolvent includes ethanol; And / or, the concentration of the starch nanoparticle solution is 1~10% (w / v); And / or, the first pH value is 8.4~8.6, and the second pH value is 6.4~6.6; And / or, the amount of octenyl succinic anhydride added is 1-3% of the dry weight of starch; And / or, the esterification reaction is carried out at a temperature of 34~36℃ and for a time of 4~6h.
4. The preparation method according to claim 2, characterized in that, Specifically, it includes: The quercetin was added to an octenyl succinic anhydride starch nanoparticle suspension for a first incubation, followed by the addition of laccase for a second incubation. After enzyme inactivation, centrifugation, precipitation, and dialysis, the octenyl succinic anhydride starch-quercetin composite nanoparticles were obtained.
5. The preparation method according to claim 4, characterized in that: The mass ratio of quercetin to octenyl succinic anhydride starch nanoparticles is 1~3:1; And / or, 50-150 U of laccase is used per gram of the octenyl succinic anhydride starch nanoparticles; And / or, the octenyl succinic anhydride starch nanoparticle suspension is weakly acidic, with a pH value of 4.4~4.6; And / or, the first incubation temperature is 28~32℃, and the time is 23~25h; And / or, the second incubation temperature is 43~47℃, and the time is 14~16h; And / or, the temperature for the first enzyme inactivation is 80~100℃, and the time is 10~30min; And / or, the dialysis time is 24~72h.
6. The preparation method according to claim 4, characterized in that, Specifically, it includes: The soybean protein isolate was dissolved in a buffer solution to obtain a protein solution. The octenyl succinic anhydride starch-quercetin composite nanoparticles were added to the protein solution. After stirring and sonication, the transglutaminase was added and subjected to a water bath for a second enzyme inactivation to obtain the rice resistant starch complex.
7. The preparation method according to claim 6, characterized in that: The concentration of the protein solution is 3-7% (w / v); And / or, the mass ratio of the octenyl succinic anhydride starch-quercetin composite nanoparticles to soy protein isolate is 1:0.5~3; And / or, use 5-30 U of transglutaminase per gram of soy protein isolate; And / or, the buffer solution includes a phosphate buffer solution; And / or, the stirring time is 20~120min, the ultrasonic treatment power is 100~300W, and the time is 5~40min; And / or, the water bath time is 3-6 hours and the temperature is 48-52°C; And / or, the temperature for the second enzyme inactivation is 80~100℃, and the time is 10~40min.
8. The rice resistant starch complex prepared by any one of claims 1-7, characterized in that: The rice resistant starch complex includes starch nanoparticles, an interface layer coated on the starch nanoparticles, and a protein coating layer coated on the interface layer.
9. The rice resistant starch complex according to claim 8, characterized in that: The interface layer is a covalently cross-linked structure, and the protein coating layer is a heteropeptide cross-linked network structure; And / or, the content of resistant starch in the rice resistant starch complex is 40.75%~50.87%.
10. The application of the rice resistant starch complex according to claim 8 or 9 in the preparation of low glycemic index foods, functional baked foods, or slow-release nutrient carriers.