Isomaltooligosaccharide, preparation method therefor, and use thereof

Abstract

The present invention relates to isomaltooligosaccharide, a preparation method therefor, and use thereof. The preparation method comprises the following steps: adding water into starch to prepare starch milk, adding α-amylase, liquefying the mixture, and then adding β-amylase and pullulanase for primary saccharification to obtain a crude maltose liquid; adding a transglucosidase for secondary saccharification to obtain a crude isomaltooligosaccharide liquid; and filtering the crude isomaltooligosaccharide liquid, and carrying out ion exchange, decoloration, and evaporation concentration to obtain the isomaltooligosacharide. According to the preparation method, the saccharide proportion of relatively high polymerization degree (DP4, DP5, DP6-9) in isomaltooligosacharide is significantly improved, so that the activity of the isomaltooligosaccharide serving as prebiotics is improved, the transparency of the isomaltooligosaccharide is effectively improved, and the color value of the isomaltooligosaccharide is reduced, thereby reducing the application limitation of the isomaltooligosacharide in product formulas of personal care products (skin, hair, scalp, oral cavity), food, pet care products, and the like.

Description

Isomaltooligosaccharide, preparation method and application thereof TECHNICAL FIELD

[0001] The present application belongs to the field of biotechnology, and particularly relates to isomaltooligosaccharide, a preparation method and application thereof. BACKGROUND

[0002] Isomaltooligosaccharide (IMO) is a mixture of 2 to 9 glucose units, with a degree of polymerization (DP) usually between 2 and 6. Common components of isomaltooligosaccharide include: isomaltose (DP2), isomaltotriose (DP3), panose (DP3), isomaltotetraose (DP4), isomaltopentaose (DP5), isomaltohexaose (DP6). Among these isomaltose, the alpha-1, 6 glycosidic bond is the main connection mode, but it may also contain alpha-1, 4 and a small amount of alpha-1, 3 glycosidic bond.

[0003] When the degree of polymerization of oligomeric isomaltose is relatively high, it generally means that its molecular chain is longer, composed of more monosaccharides. Oligomeric isomaltose with such a structure has the following advantages: (1) better prebiotic properties: oligomeric isomaltose with a high degree of polymerization is more difficult to be absorbed by the small intestine, so a larger proportion reaches the large intestine, where they can be fermented by intestinal microorganisms as prebiotics, promoting the growth of probiotics; (2) lower glycemic index (GI): due to the difficulty of rapid digestion in the small intestine, oligomeric isomaltose with a high degree of polymerization is less likely to cause rapid blood glucose rise, which is particularly beneficial for people with diabetes or those who need to control blood glucose levels; (3) enhanced dietary fiber effect: oligomeric isomaltose with a high degree of polymerization has better dietary fiber properties, which helps to improve intestinal health, improve intestinal motility, and prevent constipation; (4) improved intestinal health: as IMO with a high degree of polymerization is more fermented in the large intestine, they help to produce short-chain fatty acids (SCFAs), which help to maintain intestinal health, reduce the risk of colon cancer, regulate the immune system, and have anti-inflammatory effects; (5) reduced gastrointestinal discomfort: IMO with a high degree of polymerization can reduce some gastrointestinal discomfort that some people may experience when consuming oligosaccharides, such as bloating and diarrhea, as they are less likely to be rapidly fermented in the small intestine. In addition, the application of oligomeric isomaltose in the cosmetics industry is mainly due to its moisturizing, anti-aging, and prebiotic properties; as a prebiotic, oligomeric isomaltose can promote the growth of probiotics on the surface of the skin or in the oral cavity, inhibit harmful bacteria, and thus help to maintain the balance of the skin or oral microecology; for people with sensitive skin, oligomeric isomaltose can also be used as a soothing ingredient due to its mildness, which helps to reduce skin redness and irritation.

[0004] Although the existing preparation technology can produce products with a relatively high content of oligomeric isomaltose, such as oligomeric isomaltose content (mass fraction based on dry matter) can reach more than 90%, but the proportion of oligosaccharides with a relatively low degree of polymerization (DP1, DP2, DP3) in these oligomeric isomaltoses is generally high. Oligosaccharides with a lower degree of polymerization are more likely to be degraded or digested, thereby reducing their activity as prebiotics, and the degradation products of oligomeric isomaltose can also have adverse effects on the human body. Secondly, the oligomeric isomaltose prepared by the existing technology has problems such as deep color, poor transparency, and poor thermal stability, which poses a great challenge to the application of oligomeric isomaltose in cosmetics, oral care products, pet care products, and food formulations.

[0005] Therefore, it is urgent to explore a more scientific preparation process to more effectively expand the application of oligomeric isomaltose with a higher degree of polymerization in the cosmetics, oral care products, pet care products, and food industries.

[0006] SUMMARY

[0007] The present application aims to overcome the shortcomings and deficiencies of the prior art and provide an oligoisomaltose and a preparation method thereof, so as to increase the proportion of sugar with relatively high degree of polymerization (DP4, DP5, DP6-9) in oligoisomaltose, thereby improving the activity of oligoisomaltose as a prebiotic, and effectively improving the transparency, water solubility, and thermal stability of oligoisomaltose, and reducing its color, thereby reducing the application limitations of oligoisomaltose in cosmetic, oral care products, pet care products, and food formulations.

[0008] To achieve the above object, the technical scheme adopted by the present application comprises:

[0009] In a first aspect, the present application provides a preparation method of oligoisomaltose, comprising the following steps:

[0010] (1) starch is added with water to prepare a starch milk, and alpha-amylase is added for liquefaction to obtain a starch liquefaction solution;

[0011] (2) beta-amylase and pullulanase are added to the liquefaction solution for primary saccharification to obtain a crude maltose solution;

[0012] (3) transglucosidase is added to the crude maltose solution for secondary saccharification to obtain a crude oligoisomaltose solution;

[0013] (4) the crude oligoisomaltose solution is filtered and ion exchanged to obtain a refined oligoisomaltose solution;

[0014] (5) the refined oligoisomaltose solution is decolorized and evaporated and concentrated to obtain the oligoisomaltose.

[0015] The present application significantly increases the content of oligoisomaltose with DP4-9 by the above preparation method, solves the problem that the proportion of sugar with relatively low degree of polymerization (DP1, DP2, DP3) is generally high in oligoisomaltose prepared by the existing process, thereby reducing the problem that the short molecular chain and simple structure of oligoisomaltose affect its own prebiotic activity due to its degradation during application; and the present application optimizes the decolorization and purification process, so that the prepared oligoisomaltose has good transparency, water solubility, and thermal stability, and is lighter in color, so that it is better applied in cosmetic, oral care products, pet care products, and food formulations.

[0016] Preferably, in the step (1), the addition amount of alpha-amylase is 0.1-0.5% of the weight of starch.

[0017] Alpha-amylase is one of the key enzymes for catalyzing starch hydrolysis, which can hydrolyze the alpha-1,4-glucosidic bonds in starch molecules. The addition amount of alpha-amylase directly affects the degree of starch hydrolysis and thus the polymerization degree of the product. By controlling the addition amount of alpha-amylase, the polymerization degree of oligoisomaltose can be adjusted, and the production cycle can be reduced while the yield is improved. The inventors have found through experiments that when the addition amount of alpha-amylase is 0.1-0.5% of the weight of starch, it is more conducive to the generation of oligoisomaltose with a higher polymerization degree.

[0018] More preferably, in step (1), the addition amount of alpha-amylase is 0.3% of the weight of starch.

[0019] Preferably, in step (2), the addition amount of beta-amylase is 0.1-0.3% of the weight of starch, and the addition amount of pullulanase is 0.01-0.1% of the weight of starch. The DE value of the crude maltose solution obtained after the first saccharification is 40-50%. In step (3), the addition amount of transglucosidase is 0.1-0.3% of the weight of starch.

[0020] The DE value (mashing degree) of the crude maltose solution significantly affects the polymerization degree of oligoisomaltose. When the DE value is too low, the degree of hydrolysis of the crude maltose solution is low, and the hydrolysis reaction rate is slow, which reduces the yield, increases the production cycle and raw material cost, and the sugar chains in the crude maltose solution with a too low DE value are relatively long, which may result in a higher average polymerization degree of oligoisomaltose, making it difficult to control the polymerization degree and affecting the quality and performance of the product. When the DE value is too high, the degree of hydrolysis of the crude maltose solution is relatively high, which is not conducive to the formation of oligoisomaltose with a high polymerization degree.

[0021] Beta-amylase can selectively hydrolyze the alpha-1,4-glucosidic bonds within starch molecules, breaking down starch into shorter oligosaccharide monomers. When the addition amount of beta-amylase is too high, the starch molecules may be hydrolyzed too quickly, producing a large amount of oligosaccharide monomers, which reduces the average polymerization degree of oligoisomaltose and makes it difficult to obtain a product with a high polymerization degree. When the addition amount is too low, the starch may not be completely hydrolyzed, affecting the completeness of the reaction and the yield.

[0022] Pullulanase is an enzyme that can hydrolyze the alpha-1,6-glucosidic bonds in starch molecules. Its role is to hydrolyze the branches in starch molecules into shorter oligosaccharides, further increasing the diversity and solubility of the hydrolysis products. Too high an addition amount of pullulanase may lead to non-specific hydrolysis, not only hydrolyzing the target alpha-1,6-glucosidic bonds but also possibly hydrolyzing alpha-1,4-glucosidic bonds, which reduces the selectivity and polymerization degree of the product. Too low an addition amount may result in incomplete hydrolysis of the branches in starch, affecting the diversity and solubility of the product.

[0023] Transglucosidase can carry out transglycosylation reaction to transfer the glucosyl of substrate molecules (such as maltose) to starch molecules or other oligosaccharide molecules to form longer chains to generate high polymeric isomaltulose, which helps to achieve the desired high degree of polymerization; but too high transglucosidase addition amount can cause non-specific transglycosylation reaction, resulting in too many isomers in the product and increasing the impurities of the product, while insufficient addition amount can limit the transglycosylation reaction, thereby reducing the formation of high polymeric isomaltulose and affecting the quality and yield of the product.

[0024] It is found through experiments that when the addition amount of β-amylase is 0.1-0.3% of the weight of starch, the addition amount of pullulanase is 0.01-0.1% of the weight of starch, the DE value of the crude maltose solution obtained after the initial saccharification is 40-50%, and the addition amount of transglucosidase is 0.1-0.3% of the weight of starch, the formation of high polymeric oligomeric isomaltulose can be significantly increased, and the quality and yield of oligomeric isomaltulose can be effectively improved.

[0025] More preferably, in the step (2), the addition amount of β-amylase is 0.2% of the weight of starch, the addition amount of pullulanase is 0.05% of the weight of starch, and the DE value of the crude maltose solution obtained after the initial saccharification is 45%; in the step (3), the addition amount of transglucosidase is 0.2% of the weight of starch.

[0026] Preferably, the weight ratio of the β-amylase and the pullulanase is 4:1.

[0027] It is found through experiments that the dosage ratio of β-amylase and pullulanase has an important influence on promoting the production of high polymeric oligomeric isomaltulose; when the weight ratio of the two is too high, it will cause excessive hydrolysis, reduce the degree of polymerization of oligomeric isomaltulose, and possibly cause excessive reaction, affecting the total yield of oligomeric isomaltulose; when the weight ratio of the two is too low, it can cause the reaction to be too strong, resulting in too high degree of polymerization of oligomeric isomaltulose, which can increase the viscosity of the product and is not conducive to subsequent processing and application; when the weight ratio of β-amylase and pullulanase is 4:1, the sugar solution can be stably reacted to effectively control the degree of polymerization of oligomeric isomaltulose, so as to ensure the effective preparation of high polymeric oligomeric isomaltulose and obtain stable and high-quality oligomeric isomaltulose product.

[0028] Preferably, in the step (1), the specific process of liquefaction is to adjust the pH value to 6.5-7.5, and then spray liquefy at 100-110°C for 2.5-3.5h.

[0029] Preferably, in the step (2), the specific process of the primary saccharification is adjusting the pH value to 7.5-8.5, and then incubating at 45-55℃ for 25-35h; in the step (3), the specific process of the secondary saccharification is adjusting the pH value to 7.5-8.5, and then incubating at 45-55℃ for 85-95h.

[0030] The liquefied liquid is hydrolyzed by primary saccharification to generate shorter oligosaccharide monomers, and then the oligosaccharide molecules are subjected to transglycosylation reaction under the action of transglucosidase to extend the polymer chain, so that the preparation of oligoisomaltose with higher polymerization degree is realized. The inventors have found through experiments that, when the above-mentioned preferred saccharification conditions are within the range, better enzymatic efficiency and good transglycosylation reaction can be achieved, thereby effectively improving the polymerization degree of oligoisomaltose and reducing the problem that the oligoisomaltose is more easily degraded due to its short molecular chain and simple structure, thereby affecting its own prebiotic activity in the application process.

[0031] Preferably, in the step (4), the filtration is performed by using a drum filter, the filtration pressure is 1.5-4.5PSI, and the filtration accuracy of the filter cloth in the drum filter is 20-30μm; in the step (4), the ion exchange is performed by using a strong acid cation exchange resin to treat the sugar liquid, and the flow rate of the sugar liquid is 3-5 bed volumes / hour; in the step (5), the decolorization is performed by using 1-5%(w / v) granular activated carbon for 3-5h; and in the step (5), the evaporation compression is performed by using a mechanical steam compressor, specifically, the feeding temperature of the sugar liquid is kept below 50℃, the compression pressure is 0.5-1.5PSI, and the exhaust temperature is 55-65℃.

[0032] The present application filters the crude oligoisomaltose liquid by using a drum filter to remove larger particulate impurities mixed in the sugar liquid, further removes impurity ions in the sugar liquid by using a cation exchange resin, and finally removes pigments in the sugar liquid by adsorption or oxidation-reduction mechanism of granular activated carbon, and the inventors optimize the parameters and conditions of the decolorization and purification process, so that the prepared oligoisomaltose has high transparency and light color.

[0033] In the second aspect, the present application provides an oligoisomaltose prepared by the above-mentioned preparation method.

[0034] In the third aspect, the present application further provides the use of the above-mentioned oligoisomaltose in personal care products, food or pet care products, wherein the personal care products include skin care products, hair care products, scalp care products and oral care products.

[0035] In the fourth aspect, the present application further provides a personal care product, food or pet care product containing oligoisomaltose, which comprises the above-mentioned oligoisomaltose.

[0036] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0037] (1) By optimizing the range and ratio of enzymes used in the enzymatic hydrolysis process, this invention significantly increases the content of isomaltooligosaccharides with DP≥4, which solves the problem that the proportion of sugars with relatively low degree of polymerization (DP1, DP2, DP3) in isomaltooligosaccharides prepared by existing processes is generally high, thereby reducing the problem that isomaltooligosaccharides are more easily degraded during application due to their short molecular chains and simple structure, thus affecting their prebiotic activity.

[0038] (2) By optimizing the decolorization and purification process, the present invention makes the obtained isomaltooligosaccharide more transparent, water-soluble, and thermally stable, and lighter in color, making it easier to apply in the formulations of cosmetics, oral care products, pet care products and food. Attached Figure Description

[0039] Figure 1 shows the condition of different parts of the scalp of a subject in Effect Example 6 after 7 and 14 days of using the sample. Detailed Implementation

[0040] To better illustrate the content of this invention, the invention will be further described below in conjunction with the accompanying drawings and specific embodiments.

[0041] The α-amylase described in this invention (brand name: 2X L), β-amylase (brand name: pullulanase (brand name: Dextrozyme) All of the above were purchased from Novozymes; the transglucosidase (brand name: Transglucosidase L "Amano") was purchased from Amano Enzyme Co., Ltd.

[0042] Example 1

[0043] This embodiment provides a method for preparing isomaltooligosaccharide, including the following steps:

[0044] (1) Add water to corn starch to make starch milk, add α-amylase (0.3% of the weight of corn starch), adjust the pH value to 7, and then spray liquefy at 100℃ for 3 hours to obtain starch liquefaction liquid;

[0045] (2) Add β-amylase (0.2% of the weight of corn starch) and pullulanase (0.05% of the weight of corn starch) to the liquefied liquid, adjust the pH value to 8, and then keep it at 50℃ for 30h for primary saccharification to obtain crude maltose solution. The DE value of the crude maltose solution obtained is 45%.

[0046] (3) adding transglucosidase (0.2% of the weight of corn starch) to the crude maltose solution, adjusting the pH to 8, and then performing secondary saccharification at 50°C for 90h to obtain a crude oligoisomalto-oligosaccharide solution;

[0047] (4) filtering the crude oligoisomalto-oligosaccharide solution through a drum filter at a filtration pressure of 3PSI and a filter cloth having a filtration accuracy of 25μm, and then treating the sugar solution with a strongly acidic cation exchange resin at a flow rate of 4 bed volumes / hour to obtain a refined oligoisomalto-oligosaccharide solution;

[0048] (5) decolorizing the refined oligoisomalto-oligosaccharide solution with 3% (w / v) granular activated carbon for 4h, and then performing evaporation compression with a mechanical vapor compressor, the feed temperature of the sugar solution being maintained below 50°C, the compression pressure being 1PSI, and the exhaust temperature being 60°C to obtain the oligoisomalto-oligosaccharide.

[0049] Example 2

[0050] The present embodiment provides a method for preparing oligoisomalto-oligosaccharide, comprising the following steps:

[0051] (1) preparing a starch slurry by adding water to corn starch, adding α-amylase (0.1% of the weight of corn starch), adjusting the pH to 6.5, and then jet-liquefying at 100°C for 2.5h to obtain a starch liquefied solution;

[0052] (2) adding β-amylase (0.1% of the weight of corn starch) and pullulanase (0.025% of the weight of corn starch) to the liquefied solution, adjusting the pH to 7.5, and then performing primary saccharification at 45°C for 25h to obtain a crude maltose solution, the DE value of the obtained crude maltose solution being 40%;

[0053] (3) adding transglucosidase (0.1% of the weight of corn starch) to the crude maltose solution, adjusting the pH to 8, and then performing secondary saccharification at 50°C for 85h to obtain a crude oligoisomalto-oligosaccharide solution;

[0054] (4) filtering the crude oligoisomalto-oligosaccharide solution through a drum filter at a filtration pressure of 1.5PSI and a filter cloth having a filtration accuracy of 20μm, and then treating the sugar solution with a strongly acidic cation exchange resin at a flow rate of 3 bed volumes / hour to obtain a refined oligoisomalto-oligosaccharide solution;

[0055] (5) decolorizing the refined oligoisomalto-oligosaccharide solution with 1% (w / v) granular activated carbon for 3h, and then performing evaporation compression with a mechanical vapor compressor, the feed temperature of the sugar solution being maintained below 50°C, the compression pressure being 1PSI, and the exhaust temperature being 60°C to obtain the oligoisomalto-oligosaccharide.

[0056] Example 3

[0057] The present example provides a method for preparing oligoisomaltose, comprising the following steps:

[0058] (1) corn starch is added with water to prepare starch milk, and then α-amylase (0.5% of the weight of corn starch) is added, and the pH value is adjusted to 7, and then the starch is liquefied at 100°C for 3.5h to obtain a starch liquefied solution;

[0059] (2) β-amylase (0.3% of the weight of corn starch) and pullulanase (0.075% of the weight of corn starch) are added to the liquefied solution, and then the pH value is adjusted to 8.5, and then the solution is incubated at 55°C for 35h for primary saccharification to obtain a crude maltose solution, and the DE value of the obtained crude maltose solution is 50%;

[0060] (3) transglucosidase (0.3% of the weight of corn starch) is added to the crude maltose solution, and then the pH value is adjusted to 8, and then the solution is incubated at 50°C for 95h for secondary saccharification to obtain a crude oligoisomaltose solution;

[0061] (4) the crude oligoisomaltose solution is filtered through a drum filter, the filtration pressure is 4.5PSI, the filter cloth in the drum filter has a filtration accuracy of 30μm, and then the sugar solution is treated with strong acid cation exchange resin, the flow rate of the sugar solution is 5 bed volumes / hour, and a refined oligoisomaltose solution is obtained;

[0062] (5) the refined oligoisomaltose solution is decolorized with 5% (w / v) granular activated carbon for 5h, and then evaporated and compressed by a mechanical vapor compressor, the feeding temperature of the sugar solution is kept below 50°C, the compression pressure is 1PSI, and the exhaust temperature is 60°C, and then the solution is evaporated and concentrated by a mechanical vapor recompression machine to obtain the oligoisomaltose.

[0063] Comparative Example 1

[0064] The difference between the present comparative example and Example 1 is that in step (1), the amount of α-amylase added is 0.05% of the weight of corn starch.

[0065] Comparative Example 2

[0066] The difference between the present comparative example and Example 1 is that in step (1), the amount of α-amylase added is 0.6% of the weight of corn starch.

[0067] Comparative Example 3

[0068] The difference between the present comparative example and Example 1 is that in step (2), the amount of β-amylase added is 0.5% of the weight of starch, and the volume ratio of the added β-amylase to pullulanase is 10:1.

[0069] Comparative Example 4

[0070] The difference between this comparative example and Example 1 is that in step (3), the amount of transglucosidase added is 0.05% by weight of the starch.

[0071] Comparative Example 5

[0072] The difference between this comparative example and Example 1 is that in step (3), the amount of transglucosidase added is 0.5% by weight of the starch.

[0073] Comparative Example 6

[0074] The difference between this comparative example and Example 1 is that in step (2), no pullulanase is added.

[0075] Comparative Example 7

[0076] The difference between this comparative example and Example 1 is that in step (4), the filter pressure used is 1 PSI.

[0077] Comparative Example 8

[0078] The difference between this comparative example and Example 1 is that in step (4), the sugar solution is treated with strongly acidic cation exchange resin, and the flow rate of the sugar solution is 2 bed volumes per hour; in step (5), the decolorization time with activated carbon is 2 h.

[0079] Comparative Example 9

[0080] The difference between this comparative example and Example 1 is that in step (4), the filter pressure used is 6 PSI.

[0081] Comparative Example 10

[0082] The difference between this comparative example and Example 1 is that in step (4), the sugar solution is treated with strongly acidic cation exchange resin, and the flow rate of the sugar solution is 6 bed volumes per hour; in step (5), the amount of activated carbon used for decolorization is 6% (w / v).

[0083] Comparative Example 11

[0084] The difference between this comparative example and Example 1 is that in step (4), the filter precision of the filter cloth in the drum filter is 15 μm; in step (5), the amount of activated carbon used for decolorization is 0.5% (w / v).

[0085] Comparative Example 12

[0086] The difference between this comparative example and Example 1 is that in step (4), the filter precision of the filter cloth in the drum filter is 35 μm; in step (5), the decolorization time with activated carbon is 6 h.

[0087] Effect Example 1

[0088] In this effect example, the oligomaltose prepared in Examples 1-3 and Comparative Examples 1-6 was used as a test sample, and the content of sugars of different polymerization degrees was determined by high performance liquid chromatography to analyze the yield of each polymerization degree sugar and oligomaltose with DP≥4 in the prepared oligomaltose sample. The specific test method is as follows:

[0089] 1. Draw a standard curve

[0090] After sampling with a series of standard solutions of various sugars, a standard curve was drawn with the standard solution concentration against the peak area. The linear correlation coefficient should be 0.9990 or higher. The test sugars included:

[0091] DP1 Glucose

[0092] DP2 Maltose / Isomaltose / Nigerose / Kojibiose

[0093] DP3 Maltotriose / Panose / Isomaltotriose

[0094] DP4 Isomaltotetraose / Maltotetraose

[0095] DP5 Isomaltopentaose

[0096] DP6 Isomaltohexaose

[0097] DP7 Isomaltoheptaose

[0098] DP8 Isomaltooctaose

[0099] DP9 Nonasaccharide

[0100] 2. Preparation of sample solution

[0101] About 0.5 g of sample (accurate to 0.0001 g) was weighed, dissolved in water, transferred to a 100 ml volumetric flask, and diluted to the mark with water, then filtered with a 0.2 μm or 0.45 μm water phase microporous membrane, and the filtrate was used as needed.

[0102] 3. Determination

[0103] The prepared sample solution was injected, and the chromatographic peaks of various sugars in the sample were qualitatively determined according to the retention time of the standard sample. The mass fraction of each sugar was calculated by the peak area normalization method based on the peak area of the sample.

[0104] The results are shown in Table 1.

[0105] Table 1

[0106] Effect Example 2

[0107] In this effect example, the isomaltooligosaccharides prepared in Examples 1-3 and Comparative Examples 7-12 were used as samples, and their transmittance and color were tested.

[0108] 1. The transmittance was tested using a spectrophotometer, and the specific method was as follows:

[0109] According to the instrument manual, the zero point and transmittance of the instrument were adjusted at a wavelength of 440 nm. An appropriate amount of sample was weighed, and a new boiling cooled distilled water with a pH of 5.0-7.0 was used to prepare an isomaltooligosaccharide sample solution with a dry matter content of 30%. Then, the sample solution was injected into a 1 cm cuvette, and the transmittance of the sample solution was measured using a spectrophotometer at a wavelength of 440 nm with the same batch of water as the reference. The results were retained to one decimal place.

[0110] 2. The color was tested using a Gardner colorimeter, and the specific method was as follows:

[0111] (1) Sample preparation: The sample to be tested was stirred evenly to ensure uniform distribution of color and transparency.

[0112] (2) Calibration of the instrument: According to the instrument manual, the instrument was calibrated to ensure accurate measurement results.

[0113] (3) Measurement: The sample was injected into a transparent glass cuvette using a Gardner colorimeter. The cuvette was placed in the instrument, and the measurement button was pressed to measure.

[0114] The test results of the transmittance and color of the samples are shown in Table 2.

[0115] Table 2

[0116] Effect Example 3

[0117] In this effect example, the isomaltooligosaccharides prepared in Example 1 were used as samples, and their antibacterial test against Malassezia furfur, Propionibacterium acnes, and Staphylococcus aureus was tested.

[0118] (1) Test strains:

[0119] Malassezia furfur (ATCC 44344) was provided by Shanghai Jitai Yikosai Biotechnology Co., Ltd., the 4th generation, and the bacterial liquid concentration was: ① 7.4 x 10 5 , ② 7.2 x 10 5 , ③ 7.5 x 10 5 .

[0120] Propionibacterium acnes (GDMCC 1.243) was provided by Guangdong Microbial Culture Collection Center, the 4th generation, and the bacterial liquid concentration was: ① 1.0 x 10 6 , ② 9.8 x 10 5 , ③ 1.1 x 10 6 .

[0121] Staphylococcus aureus (ATCC 6538) was provided by Beinaelieli Biotechnology Co., Ltd., the 4th generation, and the bacterial liquid concentration was: ① 7.6 x 10 5 , ② 8.1 x 10 5 , ③ 7.5 x 10 5 .

[0122] (2) Test steps: Malassezia furfur, Propionibacterium acnes and Staphylococcus aureus bacterial suspension were diluted with PBS to about 5.0 x 10 5 CFU / mL~4.5 x 10 6 CFU / mL, respectively, 5.0 mL of oligoisomaltose prepared in Example 1 (dilution 5%) was added to a sterile test tube, 20℃±1℃ water bath for 5 min, then 0.1 mL of bacterial suspension was added, mixed quickly and immediately timed, after 0h, 8h, 24h, 0.5mL was taken and added to 4.5mL PBS, mixed thoroughly, diluted appropriately, then 2~3 dilutions were taken, 1.0mL of each was inoculated into 2 plates, and the medium was poured; at the same time, PBS was used instead of the test substance to perform a parallel test as a positive control; the same batch of PBS and medium were used as a negative control group; the test was repeated 3 times, and the culture was incubated at 30℃ for 72h. The results are shown in Tables 3-5.

[0123] Table 3 Growth rate of Malassezia furfur

[0124] Table 4 Growth rate of Propionibacterium acnes

[0125] Table 5 Growth rate of Staphylococcus aureus

[0126] As shown in Tables 3-5, compared with the microbial growth rate of the positive control group, the oligoisomaltose prepared in the present application can effectively inhibit the growth of P. acnes and M. furfur, and promote the growth of S. epidermidis, indicating that the oligoisomaltose prepared in the present application can promote the growth of probiotics on the skin surface or in the oral cavity, inhibit harmful bacteria, thereby helping to maintain the balance of the skin or oral microecology, and expanding its application in the field of oral and skin care products.

[0127] Effect Example 4

[0128] In this effect example, the oligoisomaltose prepared in Example 1 was used as a sample to test its antibacterial effect on two oral bacteria, S. mutans and P. gingivalis, at different action concentrations (0.2%, 0.5%, and 1%).

[0129] In this effect example, the standard method T / COCIA 18-2022 Evaluation Method for Antibacterial Effect of Oral Cleaning and Care Products (7.1 Suspension Quantitative Method) was referred to, and the antibacterial rate was used to judge whether the sample had antibacterial ability. The antibacterial judgment standard is: if the antibacterial rate is ≥50% to 90%, it is judged to have antibacterial effect; if the antibacterial rate is ≥90%, it is judged to have strong antibacterial effect. The results are shown in Table 6.

[0130] Table 6

[0131] As shown in Table 6, the oligoisomaltose prepared in the present application has an antibacterial rate of >99.9% at different action concentrations (0.2%, 0.5%, and 1%), indicating that it has a strong inhibitory effect on S. mutans and P. gingivalis. Therefore, the oligoisomaltose prepared by the technical scheme of the present application can inhibit the growth of harmful bacteria in the oral cavity to a certain extent, thereby inhibiting the generation of halitosis and reducing bad breath. When it is applied to oral care products, it can effectively improve the oral health status and oral odor problem.

[0132] Effect Example 5

[0133] In this effect example, the oligoisomaltose prepared in Example 1 was used as a sample to test its moisturizing effect, and the specific method was as follows.

[0134] Number of test persons: 30, 13 males and 18 females; age between 18 and 65 years old, average age 22 years old, and volunteers meet the selection criteria of human skin patch test in the “Cosmetic Safety Technology Specification” (2015 edition).

[0135] Test method: The test sample was applied to the arm calibration area at a dosage of (2.0 ± 0.1) mg / cm for a single application, and the test sample was evenly applied to the dry test area using a latex glove, and the actual application amount was recorded. After adjusting the instrument according to the instructions for the capacitance method skin moisture meter, the product area and the control area were measured, and each area was measured at least 3 times in parallel. The initial value of each test area (before use of the sample) was measured, and then the skin moisture content of the test area and the control area was measured after a set time, and the data was averaged. The same instrument must be used by the same measurement personnel to test the same subject, and the measurement probe should be cleaned between two measurements. The test sample grouping is shown in Table 7, and the transdermal water loss is shown in Table 8.

[0136] Table 7

[0137] Table 8 Transdermal water loss

[0138] As shown in Table 8, after 6h moisturizing test, compared with the blank control group, the oligoisomaltose aqueous solution prepared by the prepared oligoisomaltose of Example 1 at a concentration of 5% showed a better effect of reducing transdermal water loss, thereby exerting good moisturizing performance, while the moisturizing effect of the oligoisomaltose aqueous solution at a concentration of 1% was relatively not obvious.

[0139] Effect Example 6

[0140] In this effect example, the oligoisomaltose prepared in Example 1 was used as the sample, and the scalp serum containing 2% dilution concentration of oligoisomaltose was tested for its dandruff removal and itching relief effect, and the specific method was as follows:

[0141] (1) The scalp serum includes the following components by mass percentage: 2% oligoisomaltose (Example 1), 0.5% p-hydroxyacetophenone, 0.15% sodium carbomer, 5% butylene glycol, 0.5% 1,2-hexanediol, and the balance: water.

[0142] (2) Test subjects: A total of 13 subjects were selected, and the test population met the following conditions: healthy men or women aged 18-60 years old, at least 2 areas of the scalp with a dandruff density of 2-5 points according to expert assessment, and self-perceived itching problem. The scoring standard is shown in Table 9.

[0143] (3) Test method: after cleaning the head, part the hair, and evenly spray the prepared serum product on the scalp (especially the part with severe dandruff), and gently massage with hands or a comb until the product is absorbed, with the standard of slightly wetting the scalp. At home, use twice a day, both on dry and wet hair, once in the morning and once in the evening; after 14 days of continuous use of the test product, compare the relevant scalp indicators before and after use, and determine whether there is statistical significance within a 95% confidence interval. The results are shown in Table 10.

[0144] Table 9

[0145] Table 10

[0146] As shown in Table 10, compared with the base value, the dandruff adhesion density on the scalp of the subjects was significantly reduced (p<0.05) after 7 days and 14 days of continuous use of the oligoisomaltose aqueous solution (2% concentration) prepared in Example 1 of the present application, and the degree of scalp itching was also significantly improved (p<0.05) after 7 days and 14 days of continuous use of the product. Figure 1 shows the use of samples from different parts of the scalp of one of the subjects.

[0147] In summary, the present application optimizes the enzyme dosage range and ratio used in the enzymatic process, and optimizes the decolorization and purification process, to prepare oligoisomaltose with high polymerization degree, which can effectively reduce the problem of affecting its own activity due to its short molecular chain and simple structure which is more easily degraded during application. In addition, the oligoisomaltose prepared by the present application has light color, good transparency, and good moisturizing, dandruff-removing, and soothing itching effects, expanding its application in hair, scalp, oral and skin care products, and other fields.

[0148] Finally, it should be noted that the above examples are only used to illustrate the technical solutions of the present application and not to limit the scope of protection of the present application. Although the present application has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solutions of the present application can be modified or replaced by equivalents without departing from the essence and scope of the technical solutions of the present application.

Claims

1. A method for producing oligoisomalto- sides, characterized by, The method comprises the following steps: (1) preparing starch milk by adding water to starch, adding alpha-amylase, and liquefying to obtain a starch liquefied solution; (2) adding beta-amylase and pullulanase to the liquefied solution to perform primary saccharification, and obtaining a crude maltose solution; (3) adding transglucosidase to the crude maltose solution to perform secondary saccharification, and obtaining a crude oligoisomaltose solution; (4) filtering and ion exchanging the crude oligoisomaltose solution to obtain a refined oligoisomaltose solution; (5) decolorizing and evaporating and concentrating the refined oligoisomaltose solution to obtain the oligoisomaltose.

2. The production method according to claim 1, characterized by, In the step (1), the alpha-amylase is added in an amount of 0.1-0.5% by weight of the starch.

3. The preparation method according to claim 1, characterized in that, In the step (2), the beta-amylase is added in an amount of 0.1-0.3% by weight of the starch, the pullulanase is added in an amount of 0.01-0.1% by weight of the starch, and the obtained crude maltose solution has a DE value of 40-50%; In the step (3), the transglucosidase is added in an amount of 0.1-0.3% by weight of the starch.

4. The production method according to claim 3, characterized by, The weight ratio of the beta-amylase to the pullulanase is 4:

1.

5. The method of claim 1, wherein, In the step (1), the specific process of liquefaction is as follows: adjusting the pH value to 6.5-7.5, and then performing jet liquefaction at 100-110℃ for 2.5-3.5h.

6. The method of claim 1, wherein, In the step (2), the specific process of primary saccharification is as follows: adjusting the pH value to 7.5-8.5, and then incubating at 45-55℃ for 25-35h; In the step (3), the specific process of secondary saccharification is as follows: adjusting the pH value to 7.5-8.5, and then incubating at 45-55℃ for 85-95h.

7. The preparation method according to claim 1, characterized in that, In the step (4), the filtration is performed by using a rotary drum filter, the filtration pressure is 1.5-4.5PSI, and the filtration accuracy of the filter cloth in the rotary drum filter is 20-30μm; In the step (4), the ion exchange is performed by using a strong acid cation exchange resin to treat the sugar solution, and the flow rate of the sugar solution is 3-5 bed volumes / hour; In the step (5), the decolorization is performed by using 1-5% (w / v) granular activated carbon, and the decolorization time is 3-5h； In the step (5), the evaporation compression is performed by using a mechanical vapor compressor, specifically, the feeding temperature of the sugar solution is below 50℃, the compression pressure is 0.5-1.5PSI, and the exhaust temperature is 55-65℃.

8. An oligoisomaltose, characterized in that, The oligoisomaltose is prepared by the method of any one of claims 1-7.

9. Use of oligomeric isomaltulose according to claim 8 in personal care products, food products or pet care products, characterized in that, The personal care product comprises skin care products, hair care products, scalp care products, and oral care products.

10. A personal care product, food or pet care product comprising oligomeric isomaltulose, characterized in that, The personal care product comprises skin care products, hair care products, scalp care products, and oral care products.

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