Lotus seed juice rich in prebiotics as well as preparation method and application of lotus seed juice

Abstract

The invention belongs to the technical field of food, and particularly relates to lotus seed juice rich in prebiotics as well as a preparation method and application of the lotus seed juice. The preparation method comprises the following steps: softening, soaking and pulping lotus seeds to obtain pulp liquid; the pH value of the slurry liquid is adjusted to 4.5-5, and the slurry liquid is placed for 3-5 min; performing ultrasonic pretreatment on the slurry liquid; compound enzyme enzymolysis treatment of the pulp liquid: adding compound enzyme into the pulp liquid, heating to 35-45 DEG C, treating for 90-120 minutes to obtain enzymatic hydrolysate, and performing enzyme deactivation treatment to obtain the lotus seed juice, the compound enzyme is a mixed enzyme of cellulase, beta-glucanase, mannase, medium temperature alpha-amylase and xylanase. The oligosaccharide can be released to the maximum extent, and the composition of the oligosaccharide and the macromolecular substance structure in the lotus seed juice are optimized, so that the lotus seed juice has good storage stability and antibacterial property.

Description

Technical Field

[0001] The invention belongs to the technical field of food, and particularly relates to a method for preparing lotus seed juice rich in prebiotics. Background Art

[0002] With the growth of global economic strength, the continuous improvement of people's quality of life and dietary requirements, the demand for health foods has also increased. Researching the functional components of food and developing health foods have become hot topics in the field of international and domestic food research. Among many health foods, oligosaccharides have attracted the attention of researchers at home and abroad due to their clear chemical structure and significant physiological effects. Oligosaccharides are low-polymer sugars formed by 3 to 9 monosaccharides connected by glycosidic bonds. They can promote the proliferation of beneficial bacteria in the human intestine-bifidobacteria, and also have excellent physiological properties such as strong anti-caries and lipid-lowering. The production and sales volume ranks among the top among various functional foods.

[0003] The various physiological activities of oligosaccharides have been widely recognized, but because the raw materials of many oligosaccharides are relatively rare and the purification process of oligosaccharides is difficult and complicated, their preparation methods are mainly based on synthesis or extraction. One of the main methods for preparing oligosaccharides is the hydrolysis of polysaccharides, which mainly adopts acid degradation, microbial degradation and enzyme method. When using the acid method, the macromolecular polysaccharides must first be hydrolyzed by acid, and sometimes the corresponding oligosaccharide products are formed through corresponding transformations. The acid hydrolysis process is relatively simple and has been used in production for a long time, but it also has many disadvantages, such as the need for equipment to be resistant to strong acid and high pressure, large waste of water resources, neutralization after hydrolysis, high refining cost, low conversion rate, etc.; microbial degradation is often accompanied by the production of a large amount of organic acids, which affects the taste of the product. The enzymatic degradation technology can overcome the shortcomings of the above two methods. The enzymatic method has the following unique advantages: it can quickly and effectively carry out catalytic reactions at room temperature, pressure and mild acidity, greatly simplifying the equipment; the activation energy required for the enzyme-catalyzed reaction is extremely low, and the catalytic efficiency is much higher than that of inorganic acids; the enzyme hydrolysis is specific and the product purity is high.

[0004] Lotus seeds are the mature fruits or seeds of the plant Nelumbo, which belongs to the Nymphaeaceae family. They have been included in the list of both food and medicine published by the National Health Commission. Modern medical research shows that lotus seeds have antiviral, free radical scavenging, blood-tonifying and anti-inflammatory effects, and also have many health benefits such as lowering blood pressure, anti-oxidation, and protecting the myocardium. Lotus seeds contain a high content of carbohydrates, accounting for more than 60% of the dry weight of lotus seeds, including starch, oligosaccharides and polysaccharides.

[0005] The traditional lotus seed beverage product processing method is simple, the added value is not high, and the starch is easily regenerated after processing, causing the beverage to precipitate and stratify, and the shelf life is short. At present, some documents record the method of using enzymatic hydrolysis of lotus seeds, but the oligosaccharide yield obtained is low and the concentration of oligosaccharides in lotus seed juice is also low. These factors greatly limit the functionality and storage of lotus seed juice. Summary of the invention

[0006] 1. Technical issues to be resolved

[0007] In view of the above-mentioned shortcomings of the prior art, the present invention provides a method for preparing lotus seed juice rich in prebiotics, which can increase the oligosaccharide content in the lotus seed juice through ultrasonic combined with complex enzyme (cellulase, β-glucanase, mannanase, medium-temperature α-amylase, xylanase) degradation, and at the same time solve the problem of regeneration in the development of lotus seed beverages.

[0008] Correspondingly, the present invention also provides a lotus seed juice which is rich in prebiotics and has antibacterial effect.

[0009] (II) Technical solution

[0010] In order to achieve the above object, the main technical solutions adopted by the present invention include:

[0011] The present invention provides a method for preparing lotus seed juice rich in prebiotics, which comprises the following steps performed in sequence:

[0012] S1. Softening, soaking and pulping lotus seeds to obtain pulp liquid;

[0013] S2 adjusts the pH of the slurry to 4.5-5 and leaves it for 3-5 minutes;

[0014] S3 slurry ultrasonic pretreatment;

[0015] S4 Compound enzyme enzymatic hydrolysis treatment of slurry: adding compound enzyme to the slurry and heating it to 35-45°C for 90-120 minutes to obtain enzymatic hydrolysis solution, thereby preparing the lotus seed juice; the compound enzyme is a mixed enzyme of cellulase, β-glucanase, mannanase, medium-temperature α-amylase and xylanase.

[0016] The present invention firstly strengthens the chemical reaction between the organic acid system and the organic macromolecular substances of the plant through the strong vibration and cavitation effect of ultrasonic waves, thus creating favorable conditions for the enzymatic preparation of the composite enzyme; the enzymatic treatment of the composite enzyme can eliminate the anti-nutritional components (such as phytic acid) of lotus seeds, improve the absorption rate, provide energy for the body, change the composition of intestinal flora, have a biphasic regulating effect on gastrointestinal motility, relieve constipation symptoms, and promote the absorption function of the small intestine. Among them, the function of the composite enzyme is also to inhibit the growth of harmful bacteria in the storage period of lotus seed juice and promote the growth of beneficial bacteria. It is not only to increase the content of oligosaccharides in lotus seed juice, but also to cooperate with various components in oligosaccharides to promote the prebiotic effect.

[0017] The main component of plant cell walls is cellulose, and it also includes hemicellulose, pectin, lignin and other substances. Cellulose has a stable structure and is difficult to hydrolyze. Traditional extraction methods have high energy and material consumption. The strong vibration and cavitation effect of ultrasound can strengthen the chemical reaction between the organic acid system and the organic macromolecules of plants, creating favorable conditions for the preparation of oligosaccharides.

[0018] Optionally, citric acid is used to adjust the pH in step S2.

[0019] In the traditional acid degradation process, the dosage and reaction time of the inorganic strong acid used are difficult to accurately control, which can easily destroy the activity of the substance and even cause the generation of pigment molecules, thereby increasing the difficulty of the subsequent decolorization process. After the reaction is completed, the acid and alkali solution must be quickly neutralized or dialyzed, otherwise product pollution will be caused. In addition, the use of inorganic acids and alkalis that are not easy to degrade is likely to cause serious environmental pollution during large-scale production. The present invention uses a low-acid environment created by citric acid to assist ultrasonic pretreatment to degrade lotus seed starch and lotus seed polysaccharides at low temperature, while preventing lotus seed juice from browning. Through the effective ratio of the complex enzyme, the production cycle is greatly shortened compared with the microbial fermentation method.

[0020] Optionally, the ultrasonic treatment temperature is 40-50° C., the ultrasonic output power is 800-1200 W, and the ultrasonic treatment time is 20-30 min.

[0021] Optionally, the enzyme addition amounts are 120U / g~140U / g of cellulase, 50U / g~60U / g of β-glucanase, 10U / g~15U / g of mannanase, 180U / g~220U / g of medium-temperature α-amylase, and 25U~30U / g of xylanase based on the weight of fresh lotus seeds.

[0022] Optionally, it also includes the following steps:

[0023] S5: secondary heating enzymolysis: heating the obtained enzymolysis solution to 60-65°C, and continuing the enzymolysis for 20-30 minutes to obtain the lotus seed juice.

[0024] The present invention further adopts a temperature-raising enzymatic hydrolysis method to make the enzymatic hydrolysis process more inclined to the enzymatic hydrolysis of oligosaccharides with a higher content.

[0025] In a second aspect, the present invention further provides a lotus seed juice rich in prebiotics prepared by the preparation method described in any of the above schemes.

[0026] The weight ratio of monosaccharide, disaccharide, trisaccharide and tetrasaccharide in the lotus seed juice is 0.018-0.020: 0.060-0.070: 0.055-0.065: 0.015-0.023.

[0027] Optionally, the weight ratio of monosaccharide, disaccharide, trisaccharide and tetrasaccharide in the lotus seed juice is 0.019:0.068:0.061:0.023.

[0028] In one embodiment, the mass concentrations of the oligosaccharides in the lotus seed juice are:

[0029] Monosaccharide 0.019g / mL, disaccharide 0.068g / mL, trisaccharide 0.061g / mL, tetrasaccharide 0.023g / mL.

[0030] The above ratio of polysaccharides in lotus seed juice can make the lotus seed juice have the function of inhibiting the growth of harmful bacteria in the body and promoting the growth of beneficial bacteria. Among them, only by using the above preparation method can the ratio of oligosaccharides in lotus seed juice reach the above range.

[0031] In a third aspect, the present invention further provides a use of the above-mentioned lotus seed juice rich in prebiotics in preparing a lotus seed beverage, comprising adding auxiliary materials to the obtained lotus seed juice for preparation, and subjecting the mixture to high pressure homogenization treatment at 35-45 MPa for refinement and uniform mixing; and filling the mixed juice and sterilizing it at 90° C.-100° C. for 10-15 minutes to obtain a lotus seed beverage;

[0032] The auxiliary materials are a combination of various components, and the added amounts of the various components are 0.3%-0.5% CMC-Na, 1%-2% honey, 0.03%-0.05% xanthan gum, and 3%-5% skim milk powder, respectively, based on the weight of fresh lotus seeds.

[0033] (III) Beneficial effects

[0034] The present invention can release and prepare oligosaccharides with prebiotic activity (selectively absorbed by probiotics) to the greatest extent, and optimize the composition structure of oligosaccharides to make them have good antibacterial properties. The complex enzyme is composed of cellulase, β-glucanase, mannanase, medium-temperature α-amylase, and xylanase, which can comprehensively degrade different polysaccharide components in lotus seed juice. Cellulase mainly acts on cellulose, xylanase mainly acts on xylan, β-glucanase mainly acts on β-glucan, etc., so that oligosaccharides can be released to the greatest extent;

[0035] The present invention uses ultrasonic treatment to break the cell barrier and release the small and macromolecular contents in the lotus seeds. Then, the lotus seed starch is acted on by ultrasonic method to open the agglomerated structure of the branched starch in the lotus seed starch and transform the branched starch into a spherical shape, while avoiding the mutual entanglement of the straight chain starch. The surface area and effective action sites of starch and various types of enzymes are increased, thereby providing a good basis for the preparation of oligosaccharides with specific structures. Ultrasonic treatment can also denature the endogenous enzymes in the lotus seed juice and reduce the progress of the enzymatic browning reaction;

[0036] Optimize oligosaccharide composition: Due to the diversity of complex enzyme combinations, different enzymes have different characteristics of action on different polysaccharides. Therefore, by adjusting the enzymatic hydrolysis conditions, such as temperature and time, the production amount and proportion of different types of oligosaccharides during the enzymatic hydrolysis process can be changed, thereby optimizing the composition of oligosaccharides and making it more reasonable;

[0037] Improve enzymatic hydrolysis efficiency: The operation of secondary heating enzymatic hydrolysis can further improve the enzymatic hydrolysis efficiency and further optimize the composition of oligosaccharides. By heating the enzymatic hydrolysis solution to 60-65°C and continuing the enzymatic hydrolysis for 20-30 minutes, the higher temperature can be used to accelerate the activity of the enzyme and promote the degradation of the substrate, thereby further increasing the yield of oligosaccharides.

[0038] Increase the functionality and nutritional value of the product: Due to the modification of the complex enzyme, the carbohydrates with α-1, 4 glycosidic bonds in the substrate that are easily used by harmful bacteria are consumed, which easily increases the proportion of a large number of oligosaccharides with prebiotic activity containing α-1, 3 glycosidic bonds or α-1, 6, and components with molecular weights below 10, thereby retaining oligosaccharides that have a proliferation effect on probiotics in the human body. By fully degrading multiple polysaccharides and optimizing the composition of oligosaccharides, the content and types of oligosaccharides in lotus seed juice are enriched, with more prebiotics and functional ingredients, and the nutritional value and health functions of the product are improved;

[0039] Improved storage stability of the product: The high osmotic pressure of prebiotic functional oligosaccharides is used to effectively inhibit the growth of bacteria, yeast and mold in the lotus seed juice, which can lead to deterioration and stratification. In addition, the linear macromolecules formed by the transformation in the juice effectively protect the antioxidant active ingredients dissolved from the lotus seed cells, forming a rod-shaped inclusion compound protective layer to inhibit these ingredients from being oxidized and destroyed during storage. For example, the enzymatic browning of polyphenols oxidized to quinones is inhibited to avoid discoloration of lotus seed juice during storage; the light transmittance of lotus seed juice is reduced to avoid light accelerating the oxidation reaction of lotus seed juice; this macromolecular structure can also avoid the problem of excessive viscosity caused by excessive increase in viscosity. Finally, the network and particle interface layer structure formed by the substances in the juice can prevent the aggregation and sedimentation of suspended particles, effectively increasing the storage stability of the product.

[0040] Optimize production process: The use of compound enzymes and secondary temperature enzymatic hydrolysis methods can obtain high-quality products in a shorter time, optimize the production process, and improve production efficiency and economic benefits. BRIEF DESCRIPTION OF THE DRAWINGS

[0041] Figure 1 This is a graph showing the experimental results of intestinal flora in the lotus seed beverage of Example 2 in Experiment 1;

[0042] Figure 2 This is a graph showing the experimental results of intestinal flora in the lotus seed beverage (blank group) of Example 7 in Experiment 1;

[0043] Figure 3 This is the oligosaccharide composition diagram of the lotus seed beverage obtained in Example 7;

[0044] Figure 4 This is the oligosaccharide composition diagram of the lotus seed beverage obtained in Example 2;

[0045] Figure 5 This is the molecular conformation diagram of the solute in lotus seed juice;

[0046] Figure 6 This is the stability analysis diagram of lotus seed beverage A;

[0047] Figure 7 B is the stability analysis diagram of lotus seed beverage;

[0048] Figure 8 This is the stability analysis diagram of lotus seed beverage D. DETAILED DESCRIPTION

[0049] In order to better explain the present invention and facilitate understanding, the present invention is described in detail below through specific implementation modes, but is not limited thereto.

[0050] Example 1

[0051] This embodiment provides a method for preparing a lotus seed beverage rich in prebiotics, the steps of which are:

[0052] S1. Lotus seed screening and pulping: Select fresh or quick-frozen lotus seeds with full particles, no damage and complete core, screen and core them, add 3 times the volume of deionized water and soak them for 20 minutes; put the soaked and softened lotus seeds and the soaking liquid into a high-speed tissue pounder and pulp them for 2 minutes to obtain a slurry liquid;

[0053] S2 Acid adjustment: Add citric acid solution to the slurry to adjust the pH to 4.5 and leave it for 3 minutes;

[0054] S3 ultrasonic pretreatment: ultrasonically treat the slurry liquid, the ultrasonic temperature is 50°C, the ultrasonic output power is 800W, and the treatment time is 20min;

[0055] S4 compound enzyme enzymatic treatment: add compound enzyme to the slurry and heat to 35°C for 90 minutes; the compound enzyme is a mixed enzyme of multiple enzymes, and the added enzyme amounts are 120U / g cellulase, 50U / g β-glucanase, 10U / g mannanase, 180U / g medium-temperature α-amylase and 25U / g xylanase;

[0056] S5 secondary heating enzymolysis: the slurry liquid is heated to 60°C and the enzymolysis is continued for 20 minutes to obtain the enzymolysis liquid;

[0057] S6 Preparation: Add auxiliary materials to the enzymatic hydrolysate and mix them by high pressure homogenization at 35MPa to refine and evenly mix them; after filling the mixed juice, sterilize it at 90℃ and keep it for 15 minutes to obtain the finished lotus seed beverage;

[0058] The auxiliary material comprises the following components in percentage: 0.3% CMC-Na, 1% honey, 0.03% xanthan gum and 3% skim milk powder.

[0059] Example 2

[0060] This embodiment provides a method for preparing a lotus seed beverage rich in prebiotics, the steps of which are:

[0061] S1. Lotus seed screening and pulping: Select fresh or quick-frozen lotus seeds with full particles, no damage and complete core, screen and core them, add 4 times the volume of deionized water and soak them for 25 minutes; put the soaked and softened lotus seeds and the soaking liquid into a high-speed tissue pounder and pulp them for 1 minute to obtain a slurry liquid;

[0062] S2 Acid adjustment: Add citric acid solution to the slurry to adjust the pH to 5 and leave it for 5 minutes;

[0063] S3 ultrasonic pretreatment: ultrasonically treat the slurry liquid, the ultrasonic temperature is 40°C, the ultrasonic output power is 1200W, and the treatment time is 30min;

[0064] S4 compound enzyme enzymatic treatment: add compound enzyme to the slurry and heat to 45°C for 120 minutes; the compound enzyme is a mixed enzyme of multiple enzymes, and the added enzyme amounts are 140U / g cellulase, 60U / g β-glucanase, 15U / g mannanase, 220U / g medium-temperature α-amylase and 30U / g xylanase;

[0065] S5 secondary heating enzymolysis: the slurry liquid is heated to 65°C and the enzymolysis is continued for 30 minutes to obtain the enzymolysis liquid;

[0066] S6 Preparation: Add auxiliary materials to the enzymatic hydrolysate for preparation, and perform high pressure homogenization at 45MPa to refine and evenly mix; after filling the mixed juice, sterilize at 100℃ and keep it for 10 minutes to obtain the finished lotus seed beverage;

[0067] The auxiliary material comprises the following components in percentage: 0.5% CMC-Na, 2% honey, 0.05% xanthan gum and 5% skim milk powder.

[0068] Example 3

[0069] This embodiment provides a method for preparing a lotus seed beverage rich in prebiotics, the steps of which are:

[0070] S1. Lotus seed screening and pulping: Select fresh or quick-frozen lotus seeds with full particles, no damage and complete core, screen and core them, add 3.5 times the volume of deionized water and soak them for 23 minutes; put the soaked and softened lotus seeds and the soaking liquid into a high-speed tissue pounder and pulp them for 1.5 minutes to obtain a slurry liquid;

[0071] S2 Acid adjustment: Add citric acid solution to the slurry to adjust the pH to 5 and leave it for 4 minutes;

[0072] S3 ultrasonic pretreatment: ultrasonically treat the slurry liquid, the ultrasonic temperature is 45°C, the ultrasonic output power is 1000W, and the treatment time is 25min;

[0073] S4 compound enzyme enzymatic treatment: add compound enzyme to the slurry and heat to 40°C for 100 minutes; the compound enzyme is a mixed enzyme of multiple enzymes, and the enzyme addition amounts are 130U / g cellulase, 55U / g β-glucanase, 13U / g mannanase, 200U / g medium-temperature α-amylase and 28U / g xylanase;

[0074] S5 secondary heating enzymolysis: the slurry liquid is heated to 65°C and the enzymolysis is continued for 30 minutes to obtain the enzymolysis liquid;

[0075] S6 Preparation: Add auxiliary materials to the enzymatic hydrolysate for preparation, and perform high pressure homogenization at 45MPa to refine and evenly mix; after filling the mixed juice, sterilize at 100℃ and keep it for 10 minutes to obtain the finished lotus seed beverage;

[0076] The auxiliary material comprises the following components in percentage: 0.5% CMC-Na, 2% honey, 0.05% xanthan gum and 5% skim milk powder.

[0077] Example 4

[0078] This embodiment provides a method for preparing a lotus seed beverage rich in prebiotics, wherein the steps are the same as S1-S5 of Embodiment 2: the enzymatic hydrolyzate obtained in step S5 is subjected to enzyme inactivation treatment to obtain lotus seed juice (without S6 treatment).

[0079] Example 5

[0080] This embodiment provides a method for preparing a lotus seed beverage rich in prebiotics, and the steps are the same as those of Embodiment 2, except that the secondary heating enzymatic hydrolysis step of step S5 is not performed, and the enzymatic hydrolyzate obtained in step S4 is inactivated before step S6 (without step S5).

[0081] Example 6

[0082] This embodiment provides a method for preparing a lotus seed beverage rich in prebiotics, and the steps are the same as those in Embodiment 2, except that the temperature of the secondary heating enzymatic hydrolysis in step S5 is still 35°C.

[0083] Example 7

[0084] This embodiment provides a method for preparing a lotus seed beverage rich in prebiotics, and its steps are the same as those of Example 2, except that: without the enzymatic hydrolysis treatment of steps S4 and S5, the slurry liquid after ultrasonic pretreatment in step S3 is directly used to replace the enzymatic hydrolysis liquid in step S6, and auxiliary materials are added for preparation.

[0085] Example 8

[0086] This embodiment provides a method for preparing a lotus seed beverage rich in prebiotics, and the steps are the same as those of Example 2, except that the β-glucanase is limited to endo-β-1,4-glucanase.

[0087] In order to verify that the lotus seed beverage obtained in Examples 1-7 of the present invention has the property of prebiotic activity, the following experiments were conducted:

[0088] The following is an experimental verification that the lotus seed juice obtained by the invention has prebiotic activity.

[0089] Experiment 1

[0090] Experimental animals: Mice were about 8 weeks old and weighed 20±2g each. The room temperature was maintained at 18℃, relative humidity was 50%, artificial light time was 12 hours / day, automatic exhaust was used, food, water and disinfected bedding were changed once every morning, and mice were kept in separate cages and observed for 7 days. Those that could eat and drink normally were included in the experiment.

[0091] Example 1-Example 7 Lotus seed beverage solution preparation, so that each mouse gavage volume is: Lotus seed beverage 1000mL kg -1 BW, ensure that each mouse was gavaged with 0.4 mL (calculated based on 20 g body weight of each mouse); the blank group was gavaged with 0.4 mL of water, and the lotus seed beverage solution used for gavage was refrigerated at 4°C.

[0092] Feeding experiment: The mice were divided into 3 groups, each with 10 mice, one of which was a blank control group (without gavage), and the other two groups were lotus seed beverage groups of Examples 1 to 7. All mice were fed with a basic diet ad libitum and were raised in separate cages. The feces of each mouse were collected daily and frozen for later use. The mice were raised for a total of 14 days.

[0093] Intestinal flora analysis: 0.2 g of mouse cecal contents were aseptically taken and placed in a sterile vial with glass beads. An appropriate amount of diluent was added at a ratio of 1:10. The vial was then placed on an oscillator and fully oscillated for 0.5 min to homogenize it. Then, 10-fold serial dilutions were performed in sequence to 10 -7 . Select the appropriate dilution according to the selected culture medium and take 1mL for inoculation, and then perform aerobic and anaerobic culture respectively. Bifidobacteria and lactobacilli use MRS medium, 37℃, 48h anaerobic culture; Enterobacteriaceae use EMB medium, 37℃, 24h; Enterococci use EC medium, 37℃, 24h culture. Finally, count the number of colonies of each bacteria. Number of bacteria per 1g of feces CFU g -1 = colony count × dilution factor, logarithmic value of bacterial colony count per gram of feces Log CFU g -1 express.

[0094] Table 1 shows the number of intestinal flora (log CFU / g wet stool) after intragastric administration of the lotus seed beverage of Examples 1-4 for 14 days

[0095] Bacteria Blank group Lotus seed beverage of embodiment 1 Lotus seed beverage of embodiment 2 Lotus seed beverage of embodiment 3 Lotus seed beverage of embodiment 4 Enterobacteriaceae 6.78±0.34 5.78±0.18 5.59±0.27 5.71±0.02 5.67±0.12 Enterococci 5.61±0.38 5.38±0.06 5.26±0.21 5.34±0.07 5.28±0.18 Bifidobacterium 6.88±0.19 7.3±0.07 7.50±0.11 7.38±0.03 7.68±0.08 Lactobacilli 7.41±0.42 10.71±0.2 10.95±0.35 10.81±0.13 10.29±0.06

[0096] Table 2 shows the number of intestinal flora (log CFU / g wet stool) after intragastric administration of lotus seed beverage in Example 5-7 for 14 days

[0097] Bacteria Lotus seed beverage of embodiment 5 Lotus seed beverage of embodiment 6 Lotus seed beverage of embodiment 7 Enterobacteriaceae 5.87±0.12 5.82±0.02 5.93±0.17 Enterococci 5.56±0.2 5.43±0.12 5.52±0.13 Bifidobacterium 7.2±0.05 7.27±0.06 7.23±0.23 Lactobacilli 10.33±0.17 10.55±0.06 10.30±0.47

[0098] After 14 days of intragastric administration of the lotus seed beverage solution, the number of intestinal flora in mice changed significantly. The intestinal flora of the lotus seed beverage group (lotus seed beverage of Example 2) was as follows: Figure 2 As shown in the blank group, the lotus seed beverage of Example 7 (such as Figure 1As shown in Table 2), the number of bifidobacteria and lactobacilli increased significantly (p<0.05), while the number of enterobacteria and enterococci decreased significantly (Table 2) (p<0.05). This shows that lotus seed beverage can promote the growth of beneficial bacteria in the intestines of mice, and has an inhibitory effect on the growth of other bacteria and even harmful bacteria. The functional oligosaccharide content of the lotus seed beverage in Example 2 is relatively high. The number of live bacteria in the lotus seed juice prepared in Example 4 after oral administration to mice is close to that of the four live bacteria in Example 2. It can be seen that the functional characteristics of the lotus seed beverage are mainly contributed by the lotus seed juice, and the effect of changing the process conditions (Examples 1 and 3) is second; the above effect of lotus seed juice without secondary temperature treatment is relatively poor (Example 5), indicating that secondary enzymatic hydrolysis can further improve the prebiotic effect of lotus seed juice, which may be related to the secondary enzymatic hydrolysis to increase the yield of oligosaccharides with specific digestion-resistant glycosidic bond structure and degree of polymerization. In summary, the lotus seed juice treated with ultrasound and enzymatic hydrolysis (lotus seed juice prepared under optimal process conditions, such as Examples 2 and 4) has a better effect on the proliferation of beneficial bacteria and the inhibition of harmful bacteria. Ultrasound and appropriate enzymatic hydrolysis conditions and enzymatic hydrolysis times have an important influence on the probiotic effect of the final lotus seed juice.

[0099] Experiment 2

[0100] The oligosaccharide composition in lotus seed juice was determined by HPLC.

[0101] Test method: Take the same volume of lotus seed beverage samples obtained in Example 2 and Example 7, and add 3 times the volume of ethanol. After centrifugation, take the supernatant and concentrate it by rotary evaporation to remove the ethanol, add deionized water to make the volume 100mL, and take a small amount to inject into high performance liquid chromatography analysis. The column model is Agilent Hi-plex Na (Octo) column; the mobile phase is pure water; the detector is a differential detector; the flow rate is 0.6mL / min; the column temperature is 85℃.

[0102] Example 7 Oligosaccharide composition in lotus seed beverage Figure 3 , and the oligosaccharide composition in the lotus seed beverage in Example 2 is shown in Figure 4 According to the analysis of high performance liquid chromatography, the oligosaccharides in the lotus seed juice obtained by the present invention are mainly composed of a series of oligosaccharides with different polymerization degrees. Figure 3 Peak 1 is a tetrasaccharide, peak 2 is a trisaccharide, and peak 3 is a diosaccharide; Figure 2 Peak 1 is hepta-saccharide, peaks 2 and 3 are pentasaccharide and tetrasaccharide, peaks 4 and 5 are trisaccharide and disaccharide, and peak 6 is monosaccharide. The lotus seed beverage obtained by ultrasonic combined with complex enzyme treatment has a large amount of oligosaccharide produced, and its content is higher than that of the lotus seed beverage without enzymatic hydrolysis treatment, that is, the lotus seed beverage in Example 7.

[0103] Experiment 3

[0104] Four kinds of lotus seed beverages were prepared. Lotus seed beverage A was not treated with ultrasound and composite enzyme method. The other steps were the same as those in Example 2, except that the slurry after acidification was directly prepared. Lotus seed beverage B was prepared by the method of Example 7. Lotus seed beverage C was prepared by the method of Example 2. Lotus seed beverage D was prepared by the method of Example 8. The other treatment processes were the same.

[0105] After the four beverages were sterilized, they were placed in an incubator at 37°C for 15 days and then taken out. The number of bacteria and molds in the lotus seed juice was counted using colony counting medium (plate count agar medium), Escherichia coli counting medium (VRB-MUG agar medium) and mold counting medium (Bengal red medium). At the same time, the quality changes of each lotus seed juice were observed.

[0106] Table 2 Comparison of preservation effects of three lotus seed beverages

[0107] Lotus seed juice type Bacterial count logCFU / mL Fungal count logCFU / mL Escherichia coli count logCFU / mL Appearance, smell and taste A 8.79±0.27 3.72±0.17 6.23±0.261 The juice has a putrid smell, the juice is sour and smelly, the juice color becomes lighter, bacteria and mold colonies float on the surface, and a large amount of sediment appears at the bottom B 2.33±0.06 0.64±0.05 0.85±0.09 No significant change in appearance and taste C 2.10±0.14 0.57±0.03 0.74±0.16 No significant change in appearance and taste D 0.82±0.33 0.28±0.18 0.34±0.12 No significant change in appearance and taste

[0108] As can be seen from the above table, the lotus seed juice prepared by the above steps has a good preservation effect, which may be related to the high concentration of oligosaccharides (0.196g / mL) and the proportion of various polysaccharides contained in this lotus seed beverage, which inhibits the growth of harmful microorganisms. More specifically, the mass concentration of each oligosaccharide in the lotus seed beverage in Example 2 is: monosaccharide 0.019g / mL, disaccharide 0.068g / mL, trisaccharide 0.061g / mL, and tetrasaccharide 0.023g / mL. In Example 8, the weight ratio of monosaccharide, disaccharide, trisaccharide, and tetrasaccharide in the lotus seed beverage is 0.018: 0.065: 0.064: 0.018 (corresponding mass concentrations are: monosaccharide 0.0576g / mL, disaccharide 0.208g / mL, trisaccharide 0.2048g / mL, and tetrasaccharide 0.0576g / mL). Compared with Beverage B, Beverage C increases the effect of compound enzyme enzymolysis, but the antibacterial effect does not change significantly. Compared with beverage C, after beverage D further defines β-glucanase as endo-β-1,4-glucanase, the proportion of polysaccharides changes, and its antibacterial effect is significantly improved compared with beverages B and C. It has undergone a qualitative change and has unexpected technical effects.

[0109] In addition, the present application further confirmed through experiments that the various enzymes in the D beverage have a synergistic effect on the antibacterial effect of the beverage. This is not only because the different cleavage sites between the various enzymes result in a higher content of oligosaccharides, but also because of the types of sugars in the obtained oligosaccharides and their ratios to each other.

[0110] Experiment 4

[0111] The two samples of lotus seed juice obtained in Experimental Example 7 and Example 8 and the untreated lotus seed juice were diluted 100 times with ionized water and fixed to volume, and then injected into a high-performance size exclusion chromatography and multi-angle laser scattering (with a differential refractive index detector) system for analysis. The mobile phase of the system was 0.1 mol NaNO 3 Aqueous solution was used to test the weight-average molecular weight and gyration radius of macromolecules in lotus seed juice, with an injection volume of 100 μL.

[0112] Figure 5 It shows that the macromolecules in the lotus seed juice (red blank group) are partially U-shaped and have a partially branched structure, indicating that the initial macromolecules in the lotus seed juice are mainly branched starch. The slope of Example 7 (green) is 0.29, which is a solid sphere conformation, indicating that the macromolecules in the lotus seed juice are transformed into spheres after ultrasonic treatment. Ultrasonic treatment breaks the branching of the macromolecules and promotes the transformation of the macromolecules into spheres, increasing the surface area, thereby providing more directional binding sites for the enzyme molecules. The slope of Example 8 (blue) is 0.50, which is a rod-shaped conformation. It shows that the treatment of the mixed enzyme promotes the extension of the molecular chain and exposes free hydroxyl groups, forming a macromolecular conformation that is conducive to directional degradation into a mixture of oligosaccharides with anti-digestion prebiotics.

[0113] Experiment 5

[0114] A lotus seed beverage is not treated with ultrasound and complex enzyme method, and the other steps are the same as Example 2, and the slurry liquid after acid adjustment is directly prepared. B is a lotus seed beverage made by the method of Example 7, and D is a lotus seed beverage made by the method of Example 8. The rest of the processing technology is the same (this experiment is the sample of Experimental Example 3, one to one correspondence). The three lotus seed juices were tested after being stored in an incubator at 37°C for 15 days. The sample was analyzed using a stability analyzer. 400 μL of lotus seeds prepared by the above method was transferred to a rectangular sample tube container, and the sample was centrifuged at a speed of 4000 rpm at 25°C using a near-infrared wavelength (880 nm) beam sample, and the sample was scanned at a scanning rate of 60 s / time, and the position change curve of the integrated transmitted light over time was plotted.

[0115] The red to green curves represent the transmittance distribution at different positions in the lotus seed juice with gradually increasing time. Figure 6 (A sample), Figure 7 (Sample B), Figure 8The clarification indexes of lotus seed juice (sample D) were 0.688, 0.638 and 0.452, respectively, and the clarification index was inversely proportional to the stability of the sample. This indicates that the system of lotus seed juice of sample D is still relatively stable after being stored at 37°C for 15 days; sample A showed water separation and stratification without ultrasonic and enzymatic treatment, and the suspended matter in the lotus seed juice decreased significantly after storage due to the decomposition of microorganisms and the accelerated molecular movement at high temperature; the stability of sample B lotus seed juice was partially improved after ultrasonic treatment, but the effect was still somewhat different from that of sample D. This indicates that the oligosaccharides formed by the lotus seed juice prepared in the steps of Example 8 of the present invention can effectively inhibit the reproduction of microorganisms stored at high temperatures. In addition, the network structure formed by the macromolecular straight chains formed in the juice also contributes to the stability of lotus seed juice and the improvement of sensory indicators.

[0116] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for preparing lotus seed juice rich in prebiotics, characterized in that: It includes the following steps performed in sequence: S1. Softening, soaking and pulping lotus seeds to obtain pulp liquid; S2 adjusts the pH of the slurry to 4.5-5 and leaves it for 3-5 minutes; S3 slurry ultrasonic pretreatment; S4 Compound enzyme enzymatic hydrolysis treatment of slurry: adding compound enzyme to the slurry and heating it to 35-45°C for 90-120 minutes to obtain enzymatic hydrolysis solution, thereby preparing the lotus seed juice; the compound enzyme is a mixed enzyme of cellulase, β-glucanase, mannanase, medium-temperature α-amylase and xylanase.

2. The method for preparing the lotus seed juice rich in prebiotics as claimed in claim 1, characterized in that: The temperature of ultrasonic treatment is 40~50℃, the ultrasonic output power is 800~1200W, and the ultrasonic treatment time is 20~30min.

3. The method for preparing the lotus seed juice rich in prebiotics as claimed in claim 1, characterized in that: The added amounts of various enzymes are 120U / g~140U / g of cellulase, 50U / g~60U / g of β-glucanase, 10U / g~15U / g of mannanase, 180U / g~220U / g of medium-temperature α-amylase, and 25U~30U / g of xylanase based on the weight of fresh lotus seeds.

4. The method for preparing the lotus seed juice rich in prebiotics as claimed in claim 1, characterized in that: It also includes the following steps: S5: secondary heating enzymolysis: heating the obtained enzymolysis solution to 60-65°C, and continuing the enzymolysis for 20-30 minutes to obtain the lotus seed juice.

5. A lotus seed juice rich in prebiotics prepared by the preparation method according to any one of claims 1 to 4, characterized in that: The weight ratio of monosaccharide, disaccharide, trisaccharide and tetrasaccharide in the lotus seed juice is 0.018-0.020: 0.060-0.070: 0.055-0.065: 0.015-0.

023.

6. Use of the lotus seed juice rich in prebiotics as claimed in claim 5 in preparing a lotus seed beverage, characterized in that: Add auxiliary materials to the obtained lotus seed juice for preparation, and perform high pressure homogenization at 35-45 MPa to refine and evenly mix the juice; after filling the mixed juice, sterilize it at 90-100°C and keep it for 10-15 minutes to obtain the lotus seed beverage; The auxiliary materials are a combination of various components, and the added amounts of the various components are 0.3%-0.5% CMC-Na, 1%-2% honey, 0.03%-0.05% xanthan gum, and 3%-5% skim milk powder, respectively, based on the weight of fresh lotus seeds.

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