Preparation method of RS3 type resistant starch

Abstract

The invention provides a preparation method of RS3 type resistant starch, which comprises the following steps: (1) mixing starch, compound enzyme and deionized water to prepare a starch aqueous solution; sequentially carrying out autoclaving treatment and cooling to obtain starch paste; the compound enzyme comprises a combination of at least two of alpha-amylase, beta-amylase or glucamylase; (2) mixing the starch paste, deionized water, pullulanase and isoamylase, and debranching under the condition that the pH value is 4-6; and (3) after crystallization, carrying out solid-liquid separation, and collecting solids to obtain the RS3 type resistant starch. The RS3 type resistant starch is prepared through a hot-pressing method and an enzymolysis method, and gelatinization and liquefaction reactions are carried out at the same time, so that the reaction efficiency is remarkably improved, and the reaction steps are simplified; the pullulanase and the isoamylase have a synergistic effect, and the alpha-amylase, the beta-amylase and the glucamylase have a synergistic effect, so that the formation of the resistant starch is synergistically promoted.

Description

Technical Field

[0001] The present invention belongs to the technical field of starch processing, and particularly relates to a preparation method of RS3 type resistant starch. Background Art

[0002] RS3 type resistant starch, also known as retrograded starch, refers to starch with a new crystal form generated by the orderly opening of starch molecular chains under certain specific conditions and the retrogradation at a specific temperature, where the starch molecular chains aggregate with each other. Currently, the preparation processes of RS3 type resistant starch mainly include heat-press method, microwave method, ultrasonic method, enzymatic hydrolysis method, etc.

[0003] The autoclave method is to subject the starch milk to high temperature and high pressure treatment, break the hydrogen bonds between starch molecules, and destroy the crystalline structure, thereby forming RS3 type resistant starch; the microwave method utilizes the thermal effect and non-thermal effect of microwaves to enable starch molecules to rapidly absorb energy, change their structure and properties, and form RS3 type resistant starch; the ultrasonic method, through the cavitation effect, mechanical effect, etc. of ultrasonic waves, destroys the crystalline structure of starch granules, breaks the hydrogen bonds between starch molecules, and thus increases the anti-digestibility of starch; the enzymatic hydrolysis method uses specific enzymes to act on starch, change the structure of starch, and make it have anti-digestibility.

[0004] However, the existing technologies have certain limitations, such as uneven reactions, slow reaction rates, the need for special catalysts or complex equipment, large energy consumption, etc., and there is still a need to develop a processing technology for RS3 type resistant starch suitable for actual factory production. Summary of the Invention

[0005] Aiming at the deficiencies of the existing technologies, the purpose of the present invention is to provide a preparation method of RS3 type resistant starch. The present invention prepares RS3 type resistant starch by combining the autoclave method with the enzymatic hydrolysis method, which can significantly increase the content of resistant starch.

[0006] To achieve the purpose of this invention, the following technical solutions are adopted:

[0007] In the first aspect, the present invention provides a preparation method of RS3 type resistant starch, and the preparation method includes the following steps:

[0008] (1) Mix starch, complex enzyme, and deionized water to prepare an aqueous starch solution; successively perform autoclave treatment and cooling to obtain a starch paste; the complex enzyme includes a combination of at least two of α-amylase, β-amylase, or glucoamylase;

[0009] (2) Mix the starch paste, deionized water, pullulanase, and isoamylase, and perform debranching under the condition of a pH value of 4 - 6 (such as 4, 4.2, 4.4, 4.6, 4.8, 5, 5.2, 5.4, 5.6, 5.8, or 6, etc.).

[0010] (3) After crystallization, solid-liquid separation is carried out, and the solid is collected to obtain the RS3 type resistant starch.

[0011] In the present invention, α-amylase can randomly hydrolyze the α-1,4-glycosidic bond inside starch, but cannot hydrolyze the α-1,6-glycosidic bond; if the substrate is amylose, glucose, maltose, and maltotriose are produced by hydrolysis; if the substrate is amylopectin, glucose, maltose, maltotriose, and α-dextrin containing more than 3 glucose residues with α-1,6-glycosidic bond are produced by hydrolysis. β-amylase is a hydrolase containing sulfhydryl groups. Starting from the non-reducing end of starch, it hydrolyzes the α-1,4-glycosidic bond in units of two glucose residues in sequence to produce maltose, but cannot hydrolyze the α-1,6-glycosidic bond. Glucoamylase can not only cleave the α-1,4-glycosidic bond from the non-reducing end of the starch molecule, but also cleave the α-1,6-glycosidic bond, and has a faster hydrolysis rate for the α-1,4-glycosidic bond. Pullulanase and isoamylase can both act on the α-1,6-glycosidic bond of starch to carry out debranching reaction.

[0012] The present invention prepares RS3 type resistant starch by the autoclaving method combined with the enzymatic method, which can fully rearrange and crystallize the starch molecular chain, significantly increase the content of resistant starch; at the same time, gelatinization and liquefaction reactions of starch are carried out, without the need for additional microwave or ultrasonic assistance, significantly improving the reaction efficiency and simplifying the reaction steps; pullulanase and isoamylase act synergistically on the starch molecule, having a better debranching effect and increasing the content of resistant starch.

[0013] Preferably, the ratio of the complex enzyme to starch is (0.01-4) U:1 g.

[0014] The specific point values in (0.01-4) can be 0.01, 0.05, 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, or 0.4, etc.

[0015] Other specific point values within the above numerical ranges can be selected and will not be elaborated one by one here.

[0016] Preferably, the complex enzyme includes α-amylase, β-amylase, and glucoamylase.

[0017] In the present invention, α-amylase, β-amylase, and glucoamylase are all high-temperature resistant enzymes, which can simultaneously carry out gelatinization and liquefaction reactions of starch; α-amylase, β-amylase, and glucoamylase act synergistically on the starch molecule, having a better starch molecular chain separation effect and promoting the formation of resistant starch.

[0018] Preferably, the enzyme activity ratio of α-amylase, β-amylase, and glucoamylase is (0.1-2):(0.1-2):(0.1-1.5).

[0019] Specific point values within the first range (0.1 - 2) can be 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.2, 1.4, 1.6, 1.8, 2, etc.; specific point values within the second range (0.1 - 2) can be 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.2, 1.4, 1.6, 1.8, 2, etc.; specific point values within the range (0.1 - 1.5) can be 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.1, 1.2, 1.3, 1.4, 1.5, etc.

[0020] Other specific point values within the above numerical ranges can be selected, and will not be elaborated one by one here.

[0021] Preferably, the mass percentage content of starch in the aqueous starch solution is 20 - 80%, for example, it can be 20%, 30%, 40%, 50%, 60%, 70%, 80%, etc.

[0022] Other specific point values within the above numerical ranges can be selected, and will not be elaborated one by one here.

[0023] Preferably, the starch includes any one or a combination of at least two of rice starch, wheat starch, corn starch, sweet potato starch, cassava starch, mung bean starch, pea starch, or potato starch.

[0024] The starch described in the present invention is a conventional starch raw material in the art, not limited to the starch raw materials listed above, and is also applicable to other unlisted starch raw materials.

[0025] Preferably, the process of pressure heat treatment includes first maintaining at 72 - 80 °C (for example, it can be 72 °C, 73 °C, 74 °C, 75 °C, 76 °C, 77 °C, 78 °C, 79 °C, or 80 °C, etc.) for 30 - 60 min (for example, it can be 30 min, 35 min, 40 min, 45 min, 50 min, 55 min, or 60 min, etc.), and then maintaining at 90 - 120 °C (for example, it can be 90 °C, 95 °C, 100 °C, 105 °C, 110 °C, 115 °C, or 120 °C, etc.) for 30 - 60 min (for example, it can be 30 min, 35 min, 40 min, 45 min, 50 min, 55 min, or 60 min, etc.).

[0026] In the present invention, the gelatinization reaction and the liquefaction reaction are carried out simultaneously. α - amylase, β - amylase, and glucoamylase are all high - temperature - resistant enzymes. β - amylase and glucoamylase carry out the liquefaction reaction at 60 - 80 °C, and α - amylase carries out the liquefaction reaction at 90 - 120 °C.

[0027] Preferably, the cooling temperature is 50 - 60°C, for example, it can be 50°C, 51°C, 52°C, 53°C, 54°C, 55°C, 56°C, 57°C, 58°C, 59°C or 60°C, etc.; the time is 30 - 90 min, for example, it can be 30 min, 40 min, 50 min, 60 min, 70 min, 80 min or 90 min, etc.

[0028] Other specific point values within the above numerical ranges can be selected and will not be elaborated one by one here.

[0029] Preferably, the ratio of the total dosage of pullulanase and isoamylase to starch is (20 - 50) U:1 g.

[0030] The specific point values in (20 - 50) can be 20, 25, 30, 35, 40, 45, 50, 55 or 60, etc.

[0031] Other specific point values within the above numerical ranges can be selected and will not be elaborated one by one here.

[0032] Preferably, the enzyme activity ratio of pullulanase to isoamylase is (1 - 3):(1 - 3).

[0033] The specific point values in the first (1 - 3) can be 1, 1.2, 1.5, 1.8, 2, 2.2, 2.5, 2.8 or 3, etc.; the specific point values in the second (1 - 3) can be 1, 1.2, 1.5, 1.8, 2, 2.2, 2.5, 2.8 or 3, etc.

[0034] Other specific point values within the above numerical ranges can be selected and will not be elaborated one by one here.

[0035] Preferably, the pH-adjusting solution includes any one or a combination of at least two of acetic acid, lactic acid, citric acid or hydrochloric acid.

[0036] Preferably, the temperature for debranching is 50 - 70°C, for example, it can be 50°C, 52°C, 54°C, 56°C, 58°C, 60°C, 62°C, 64°C, 66°C, 68°C or 70°C, etc.; the time is 0.5 - 10 h, for example, it can be 0.5 h, 1 h, 2 h, 3 h, 4 h, 5 h, 6 h, 7 h, 8 h, 9 h or 10 h, etc.

[0037] Preferably, the crystallization temperature is 0 - 50°C, for example, it can be 0°C, 5°C, 10°C, 15°C, 20°C, 25°C, 30°C, 35°C, 40°C, 45°C or 50°C, etc.; the time is 24 - 72 h, for example, it can be 24 h, 30 h, 36 h, 42 h, 48 h, 54 h, 60 h, 66 h or 72 h, etc.

[0038] Other specific point values within the above numerical ranges can be selected and will not be elaborated one by one here.

[0039] Preferably, the solid-liquid separation method includes centrifugation or filtration.

[0040] The method of first crystallizing and then performing solid-liquid separation in the present invention can improve the crystallization efficiency and increase the content of resistant starch.

[0041] Preferably, the rotation speed of the centrifugation is 4000 - 8000 rpm, for example, it can be 4000 rpm, 5000 rpm, 6000 rpm, 7000 rpm or 8000 rpm, etc.; the time is 3 - 10 min, for example, it can be 3 min, 4 min, 5 min, 6 min, 7 min, 8 min, 9 min or 10 min, etc.

[0042] Other specific point values within the above numerical ranges can be selected and will not be elaborated one by one here.

[0043] Preferably, after collecting the solid, it further includes the steps of washing with water, drying and pulverizing.

[0044] Preferably, the drying temperature is 100 - 200 °C, for example, it can be 100 °C, 110 °C, 120 °C, 130 °C, 140 °C, 150 °C, 160 °C, 170 °C, 180 °C, 190 °C or 200 °C, etc.; the time is 2 - 24 h, for example, it can be 2 h, 4 h, 6 h, 8 h, 10 h, 12 h, 14 h, 16 h, 18 h, 20 h, 22 h or 24 h, etc.

[0045] Other specific point values within the above numerical ranges can be selected and will not be elaborated one by one here.

[0046] Preferably, the preparation method specifically includes the following steps:

[0047] (1) Mix starch, complex enzyme and deionized water to prepare a starch aqueous solution with a starch mass percentage of 20 - 80%; the ratio of the complex enzyme to starch is (0.01 - 4) U:1 g, the complex enzyme includes α-amylase, β-amylase and glucoamylase, and the enzyme activity ratio of α-amylase, β-amylase and glucoamylase is (0.1 - 2):(0.1 - 2):(0.1 - 1.5); perform autoclaving treatment, first maintain at 72 - 80 °C for 30 - 60 min, then maintain at 90 - 120 °C for 30 - 60 min, and cool to 50 - 60 °C after 30 - 90 min to obtain a starch paste;

[0048] (2) Mix the starch paste, deionized water, pullulanase and isoamylase, adjust the pH value to 4 - 6 with any one or a combination of at least two of acetic acid, lactic acid, citric acid or hydrochloric acid, and carry out debranching at 50 - 70 °C for 0.5 - 10 h; the total dosage ratio of pullulanase and isoamylase to starch is (20 - 50) U:1 g, and the enzyme activity ratio of pullulanase and isoamylase is (1 - 3):(1 - 3);

[0049] (3) Crystallize at 0 - 50 °C for 24 - 72 h, centrifuge at 4000 - 8000 rpm for 3 - 10 min, collect the solid, wash with water, dry at 100 - 200 °C for 2 - 24 h, and pulverize to obtain the RS3 type resistant starch.

[0050] Compared with the prior art, the present invention has the following beneficial effects:

[0051] (1) The present invention prepares RS3 type resistant starch by the autoclave method combined with the enzymatic method, which can fully rearrange and crystallize the starch molecular chains, significantly increase the content of resistant starch; at the same time, the gelatinization and liquefaction reactions of starch are carried out, without the need for additional microwave or ultrasonic assistance, significantly improving the reaction efficiency and simplifying the reaction steps; pullulanase and isoamylase act synergistically on the starch molecules, having a better debranching effect and increasing the content of resistant starch.

[0052] (2) In the present invention, α - amylase, β - amylase and glucoamylase are all high - temperature - resistant enzymes, which can carry out the gelatinization and liquefaction reactions of starch simultaneously; α - amylase, β - amylase and glucoamylase act synergistically on the starch molecules, having a better separation effect of starch molecular chains and promoting the formation of resistant starch. Specific embodiments

[0053] To further elaborate on the technical means and effects adopted by the present invention, the following preferred embodiments of the present invention are used to further illustrate the technical solutions of the present invention, but the present invention is not limited to the scope of the embodiments.

[0054] For those not specifying specific techniques or conditions in the embodiments, they shall be carried out according to the techniques or conditions described in the literature in the field or according to the product specifications. For the reagents or instruments not indicating the manufacturer, they are all conventional products that can be obtained through regular channels.

[0055] The sources of the materials used in the following specific embodiments are as follows:

[0056] Raw material name Purchasing manufacturer α-Amylase Ningxia Xiasheng Biotechnology Co., Ltd. β-Amylase Hunan Hongyingxiang Biotechnology Co., Ltd. Glucoamylase Hunan Hongyingxiang Biotechnology Co., Ltd. Pullulanase Shandong Longkete Enzyme Preparation Co., Ltd. Isoamylase Hunan Hongyingxiang Biotechnology Co., Ltd.

[0057] Example 1

[0058] This example provides a method for preparing RS3 type resistant starch, and the preparation method includes the following steps:

[0059] (1) Mix 200 g of corn starch, 1 U / g of α-amylase, 1 U / g of β-amylase, 1 U / g of glucoamylase and deionized water (the ratio of the complex enzyme to starch is 3 U:1 g) to prepare a starch aqueous solution with a starch mass percentage of 20%; conduct pressure heat treatment, first hold at 75 °C for 45 min, then hold at 105 °C for 45 min, and cool to 55 °C after 60 min to obtain a starch paste;

[0060] (2) Mix the starch paste, 22 U / g of pullulanase, 18 U / g of isoamylase and deionized water (the total dosage of pullulanase and isoamylase and starch ratio is 40 U:1 g), adjust the pH value to 5 with 1 M acetic acid, and carry out debranching at 60 °C for 3 h;

[0061] (3) Crystallize at 25 °C for 48 h, centrifuge at 6000 rpm for 5 min, collect the solid, wash with water, dry at 150 °C for 12 h, and pulverize to obtain the RS3 type resistant starch.

[0062] Example 2

[0063] This example provides a method for preparing RS3 type resistant starch, and the preparation method includes the following steps:

[0064] (1) Mix 200 g of corn starch, 1.5 U / g of α-amylase, 0.3 U / g of β-amylase, 0.7 U / g of glucoamylase and deionized water (the ratio of the complex enzyme to starch is 2.5 U:1 g) to prepare a starch aqueous solution with a starch mass percentage of 20%; conduct pressure heat treatment, first hold at 72 °C for 60 min, then hold at 120 °C for 30 min, and cool to 60 °C after 30 min to obtain a starch paste;

[0065] (2) Mix the starch paste, 18 U / g of pullulanase, 12 U / g of isoamylase and deionized water (the total dosage of pullulanase and isoamylase and starch ratio is 30 U:1 g), adjust the pH value to 6 with 1 M lactic acid, and carry out debranching at 50 °C for 6 h;

[0066] (3) Crystallize at 50 °C for 24 h, centrifuge at 4000 rpm for 10 min, collect the solid, wash with water, dry at 100 °C for 24 h, and pulverize to obtain the RS3 type resistant starch.

[0067] Example 3

[0068] This example provides a method for preparing RS3 type resistant starch, and the preparation method includes the following steps:

[0069] (1) Mix 200 g of corn starch, 0.2 U / g of α - amylase, 1.5 U / g of β - amylase, 0.3 U / g of glucoamylase and deionized water (the ratio of the complex enzyme to starch is 2 U:1 g) to prepare a starch aqueous solution with a starch mass percentage of 20%. Conduct pressure - heat treatment, first hold at 80 °C for 30 min, then hold at 90 °C for 60 min, and cool to 50 °C after 90 min to obtain a starch paste;

[0070] (2) Mix the starch paste, 10 U / g of pullulanase, 10 U / g of isoamylase and deionized water (the total dosage of pullulanase and isoamylase and starch ratio is 20 U:1 g), adjust the pH value to 4 with 1 M citric acid, and carry out de - branching at 70 °C for 2 h;

[0071] (3) Crystallize at 4 °C for 72 h, centrifuge at 8000 rpm for 3 min, collect the solid, wash with water, dry at 200 °C for 8 h, and pulverize to obtain the RS3 - type resistant starch.

[0072] Example 4

[0073] This example provides a method for preparing RS3 - type resistant starch, which is only different from Example 1 in that: in step (1), α - amylase is not added, and its reduced amount is proportionally distributed to β - amylase and glucoamylase, and the ratio of the total enzyme activity of the complex enzyme to starch is 3 U:1 g, and other steps remain unchanged.

[0074] Example 5

[0075] This example provides a method for preparing RS3 - type resistant starch, which is only different from Example 1 in that: in step (1), β - amylase is not added, and its reduced amount is proportionally distributed to α - amylase and glucoamylase, and the ratio of the total enzyme activity of the complex enzyme to starch is 3 U:1 g, and other steps remain unchanged.

[0076] Example 6

[0077] This example provides a method for preparing RS3 - type resistant starch, which is only different from Example 1 in that: in step (1), glucoamylase is not added, and its reduced amount is proportionally distributed to α - amylase and β - amylase, and the ratio of the total enzyme activity of the complex enzyme to starch is 3 U:1 g, and other steps remain unchanged.

[0078] Comparative Example 1

[0079] This comparative example provides a method for preparing RS3 - type resistant starch, which is only different from Example 1 in that: in step (1), α - amylase, β - amylase and glucoamylase are not added, and other steps remain unchanged.

[0080] Comparative Example 2

[0081] This comparative example provides a method for preparing RS3 type resistant starch, and the difference from Example 1 is only that: in step (2), pullulanase is not added, and its reduced amount is supplemented by isoamylase, and the ratio of isoamylase to starch is 40 U:1 g, and other steps remain unchanged.

[0082] Comparative Example 3

[0083] This comparative example provides a method for preparing RS3 type resistant starch, and the difference from Example 1 is only that: in step (2), isoamylase is not added, and its reduced amount is supplemented by pullulanase, and the ratio of pullulanase to starch is 40 U:1 g, and other steps remain unchanged.

[0084] Comparative Example 4

[0085] This comparative example provides a method for preparing RS3 type resistant starch, and the difference from Example 1 is only that: in step (1), 200 g of corn starch and deionized water are mixed to prepare a starch aqueous solution with a starch mass percentage of 20%; pressure heat treatment is carried out for gelatinization reaction, first kept at 75 °C for 45 min, and then kept at 105 °C for 45 min; 1 U / g of α-amylase, 1 U / g of β-amylase, and 1 U / g of glucoamylase (the ratio of the complex enzyme to starch is 3 U:1 g) are added for liquefaction reaction, first kept at 70 °C for 45 min, and then kept at 105 °C for 45 min, and other steps remain unchanged.

[0086] Comparative Example 5

[0087] This comparative example provides a method for preparing RS3 type resistant starch, and the difference from Example 1 is only that: in step (3), first centrifuge at 6000 rpm for 5 min, and then crystallize at 25 °C for 48 h, and other steps remain unchanged.

[0088] Test Example 1

[0089] (1) Test samples: RS3 type resistant starch prepared in Examples 1-6 and Comparative Examples 1-5;

[0090] (2) Test method: The Englyst method is used to determine resistant starch (RS), and the determination kit (product number "K-DSTR", manufactured by Megazyme Company);

[0091] 0.5 g of pancreatin was suspended in 50 mL of 50 mM maleic acid buffer (pH 6.0, 2 mM CaCl2) for 5 minutes, and 0.5 mL of amyloglucosidase solution (300 U / mL) was added; 4 mL of the supernatant was centrifuged and added to a capped test tube containing 100 mg (± 5 mg) of the sample (i.e., the starch-containing material) and fully suspended using a vortex mixer;

[0092] The mixture was shaken horizontally at a shaking speed of 200 times / minute and simultaneously subjected to enzymatic digestion at 37°C for 4 hours. After digestion, 4 mL of 99.5% ethanol was added and mixed thoroughly, and the mixture was centrifuged at 3000 rpm for 10 minutes, and the supernatant was removed. 8 mL of 50% ethanol was added to the precipitate in two portions, and the precipitate was resuspended and centrifuged again. The above steps were repeated once more to recover the precipitate, i.e., the resistant starch fraction.

[0093] Immerse the capped test tube containing the precipitate in ice water, add 2 mL of 2 M KOH solution, and stir and mix with a star-shaped stirring bar for 20 minutes to completely dissolve the resistant starch component; add 8 mL of 1.2 M sodium acetate buffer (pH 3.8) for neutralization, and add 0.1 mL of amyloglucosidase (3300 U / mL); incubate in a 50°C water bath for 30 minutes to digest the resistant starch component into glucose;

[0094] The reaction solution after amyloglucosidase digestion was centrifuged at 3000 rpm for 10 minutes, and 0.5 mL of the supernatant was diluted with 4.5 mL of distilled water. 0.1 mL of the diluted reaction solution was mixed with 3 mL of GOPOD reagent and incubated at 50°C for 20 minutes. After cooling to room temperature, the absorbance at 510 nm was measured by spectrophotometry, using the standard contained in the kit as a reference for quantifying glucose. The resistant starch content is the weight of resistant starch per unit dry weight of the sample (w / w), and is determined by converting the amount of glucose derived from the free resistant starch component produced by the final amyloglucosidase digestion into the amount of starch.

[0095] (3) Test results: As shown in Table 1, the present invention prepares RS3 resistant starch by combining the autoclave method with the enzymatic hydrolysis method, which can fully rearrange and crystallize the starch molecular chains and significantly increase the content of resistant starch; α-amylase, β-amylase and glucoamylase have a synergistic effect, and pullulanase and isoamylase have a synergistic effect, which synergistically promote the formation of resistant starch; the gelatinization and liquefaction reactions of starch are carried out simultaneously, without the need for additional microwave or ultrasonic assistance, which significantly improves the reaction efficiency and simplifies the reaction steps; the method of crystallization first and then solid-liquid separation can improve the crystallization efficiency and increase the content of resistant starch.

[0096] Table 1

[0097] Sample RS3 type resistant starch content (%) Yield (%) Example 1 63.34 57.12 Example 2 60.34 56.34 Example 3 58.34 55.42 Example 4 28.46 33.99 Example 5 31.34 36.39 Example 6 33.22 37.90 Comparative Example 1 15.06 16.21 Comparative Example 2 45.35 44.33 Comparative Example 3 40.45 43.21 Comparative Example 4 35.34 34.32 Comparative Example 5 50.34 46.73

[0098] The applicant declares that the present invention illustrates a method for preparing RS3 type resistant starch of the present invention through the above embodiments, but the present invention is not limited to the above embodiments, that is, it does not mean that the present invention must rely on the above embodiments to be implemented. Those skilled in the art should understand that any improvement to the present invention, the equivalent substitution of each raw material of the product of the present invention, the addition of auxiliary components, the selection of specific methods, etc. all fall within the protection scope and the disclosure scope of the present invention.

[0099] The preferred embodiments of the present invention have been described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the technical concept scope of the present invention, various simple modifications can be made to the technical solution of the present invention, and these simple modifications all belong to the protection scope of the present invention.

[0100] In addition, it should be noted that in the above specific embodiments, the various specific technical features described can be combined in any suitable manner without conflict. In order to avoid unnecessary repetition, the present invention will not separately describe various possible combination methods.

Claims

1. A method for preparing RS3 type resistant starch, characterized in that, The preparation method comprises the following steps: (1) Mix starch, complex enzyme and deionized water to prepare a starch aqueous solution; successively carry out autoclaving treatment and cooling to obtain a starch paste; the complex enzyme comprises a combination of at least two of α-amylase, β-amylase or glucoamylase; (2) Mix the starch paste, deionized water, pullulanase and isoamylase, and carry out debranching under the condition that the pH value is 4-6; (3) After crystallization, carry out solid-liquid separation, collect the solid to obtain the RS3 type resistant starch.

2. The preparation method according to claim 1, characterized in that, The ratio of the complex enzyme to starch is (0.01-4) U: 1 g; Preferably, the complex enzyme comprises α-amylase, β-amylase and glucoamylase; Preferably, the enzyme activity ratio of α-amylase, β-amylase and glucoamylase is (0.1-2): (0.1-2): (0.1-1.5).

3. The preparation method according to claim 1 or 2, characterized in that, The mass percentage content of starch in the starch aqueous solution is 20-80%; Preferably, the starch comprises any one or a combination of at least two of rice starch, wheat starch, corn starch, sweet potato starch, cassava starch, mung bean starch, pea starch or potato starch.

4. The preparation method according to any one of claims 1-3, characterized in that, The autoclaving treatment process comprises maintaining at 72-80 °C for 30-60 min first, and then maintaining at 90-120 °C for 30-60 min; Preferably, the cooling temperature is 50-60 °C and the time is 30-90 min.

5. The preparation method according to any one of claims 1-4, characterized in that, The total dosage of pullulanase and isoamylase and the ratio of starch is (20-50) U: 1 g; Preferably, the enzyme activity ratio of pullulanase and isoamylase is (1-3): (1-3).

6. The preparation method according to any one of claims 1-5, characterized in that, The pH value regulating solution comprises any one or a combination of at least two of acetic acid, lactic acid, citric acid or hydrochloric acid.

7. The preparation method according to any one of claims 1-6, characterized in that, The debranching temperature is 50-70 °C and the time is 0.5-10 h.

8. The preparation method according to any one of claims 1-7, characterized in that, The crystallization temperature is 0-50 °C and the time is 24-72 h.

9. The preparation method according to any one of claims 1-8, characterized in that, The solid-liquid separation method comprises centrifugation or filtration; Preferably, the centrifugation speed is 4000-8000 rpm and the time is 3-10 min; Preferably, after collecting the solid, it further comprises the steps of washing with water, drying and pulverizing; Preferably, the drying temperature is 100-200 °C and the time is 2-24 h.

10. The preparation method according to any one of claims 1-9, characterized in that, The preparation method specifically comprises the following steps: (1) Mix starch, complex enzyme and deionized water to prepare a starch aqueous solution with a starch mass percentage content of 20-80%; the ratio of the complex enzyme to starch is (0.01-4) U: 1 g, the complex enzyme comprises α-amylase, β-amylase and glucoamylase, and the enzyme activity ratio of α-amylase, β-amylase and glucoamylase is (0.1-2): (0.1-2): (0.1-1.5); carry out autoclaving treatment, maintain at 72-80 °C for 30-60 min first, then maintain at 90-120 °C for 30-60 min, and cool to 50-60 °C after 30-90 min to obtain a starch paste; (2) Mix the starch paste, deionized water, pullulanase and isoamylase, and adjust the pH value to 4 - 6 with any one or a combination of at least two of acetic acid, lactic acid, citric acid or hydrochloric acid, and carry out debranching at 50 - 70 °C for 0.5 - 10 h; the total dosage ratio of the pullulanase and isoamylase to the starch is (20 - 50) U:1 g, and the enzyme activity ratio of the pullulanase and isoamylase is (1 - 3):(1 - 3); (3) Crystallize at 0 - 50 °C for 24 - 72 h, centrifuge at 4000 - 8000 rpm for 3 - 10 min, collect the solid, wash with water, dry at 100 - 200 °C for 2 - 24 h, and pulverize to obtain the RS3 type resistant starch.