Preparation method of Chinese yam powder through synergistic fermentation of lactic acid bacteria and compound enzyme

Abstract

The invention discloses a preparation method of Chinese yam powder cooperatively fermented by lactic acid bacteria and compound enzyme. The preparation method comprises the following steps: weighing Chinese yam powder and germs, and mixing the Chinese yam powder and germs according to a mass ratio of 2: 1; preparing a 2% sugar aqueous solution, and respectively carrying out high-pressure steam sterilization at 115 DEG C for 30 minutes on the sugar aqueous solution and the mixture; after cooling, adding the sweet water according to the mass ratio of the Chinese yam powder to the water of 3: 7, and uniformly mixing; a compound enzyme is added into the mixed solution, the compound enzyme is composed of amylase, glycoprotease and neutral protease, the preferable mass ratio of the amylase to the glycoprotease to the neutral protease is 1: 1: 1, and enzymolysis is conducted for 3-5 h at the temperature of 50 DEG C; inoculating a mixed bacteria solution consisting of lactobacillus bulgaricus and lactobacillus pentosus into the enzymatic hydrolysate, with the inoculation amount being 2-8% of the mass of the material, and fermenting at 30 DEG C for 4 days; and drying, crushing and optional flavor blending are performed to obtain the fermented Chinese yam powder. According to the method, the solubility and water binding capacity of the Chinese yam powder can be remarkably improved, the activity of total flavonoids and SOD (superoxide dismutase) is increased, the nutritional value and flavor are improved, and the method is suitable for development of instant drinks and functional foods.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of food processing and fermentation preparation, and particularly relates to a preparation method of yam powder fermented by lactic acid bacteria and complex enzymes. BACKGROUND

[0002] Yam (Dioscorea opposita) is a traditional food and medicine homologous plant, rich in polysaccharides, proteins, saponins, flavonoids and various vitamins, and has the effects of tonifying the spleen, benefiting qi and nourishing yin, and is widely used in the field of health food and functional food.

[0003] However, the traditional yam powder has problems of poor solubility, rough taste and difficulty in fully releasing nutritional components during processing and brewing, which limits its application in instant drinks and functional foods.

[0004] Previous studies have shown that microbial fermentation can improve the solubility, flavor and functional activity of plant raw materials, but single lactic acid bacteria fermentation or single enzyme treatment has limited effect on the improvement of yam powder. How to further improve the solubility, functional activity and taste of yam powder through the synergistic effect of lactic acid bacteria and complex enzymes has become a technical problem to be solved in the field. SUMMARY

[0005] The application aims to provide a preparation method of yam powder fermented by lactic acid bacteria and complex enzymes, which solves the technical problems of poor solubility, poor taste and low utilization rate of active ingredients of existing yam powder.

[0006] To solve the above technical problems, the application adopts the following technical scheme:

[0007] A preparation method of yam powder fermented by lactic acid bacteria and complex enzymes, comprising the following steps:

[0008] (1) Raw material preparation and sugar water preparation: weigh yam powder and germ, mix them uniformly at a mass ratio of 2:1 to obtain dry materials; additionally, prepare a 2% sugar water solution for standby;

[0009] (2) Sterilization treatment: sterilize the dry materials and the sugar water solution by high-pressure steam sterilization, and the sterilization conditions are 115℃ and 30min;

[0010] (3) Sterilization mixing: after sterilization and cooling, add the sugar water solution to the dry materials and mix them uniformly to form a mixed solution;

[0011] (4) Enzymolysis: add complex enzyme preparation to the mixed solution for enzymolysis, and the complex enzyme is composed of amylase, glycoproteinase and neutral protease;

[0012] (5) Inoculation and fermentation: A mixed lactic acid bacteria solution is inoculated into the enzymatically hydrolyzed material for fermentation. The lactic acid bacteria are composed of Lactobacillus jaguariensis and Lactobacillus pentosus, and the inoculation amount is 2%-8% of the total mass of the material. The fermentation is carried out at a constant temperature of 30°C for 4 days;

[0013] (6) Drying and crushing the prepared materials to obtain the yam powder product.

[0014] Furthermore, after the fermentation in step (5) is completed, the material is flavored by adding sodium citrate and white sugar to the fermented and enzymatically hydrolyzed material and stirring evenly.

[0015] Furthermore, the amount of sodium citrate added is 0.6% of the total mass of the material, and the amount of white sugar added is 10% of the total mass of the material.

[0016] Furthermore, the amount of the sugar aqueous solution added in step (3) is such that the mass ratio of the total mass of water in the system to the mass of the yam powder is 7:3.

[0017] Furthermore, the inoculation amount of the mixed lactic acid bacteria solution in step (5) is 4% of the total mass of the material.

[0018] Furthermore, in step (5), the lactic acid bacteria are prepared by mixing Lactobacillus jaguar and Lactobacillus pentosus in a ratio of 1:1.

[0019] Furthermore, the amount of compound enzyme added in step (4) is 0.1 g of compound enzyme per 10 g of yam powder.

[0020] Furthermore, in step (4), the complex enzyme consists of amylase, glycoproteinase and neutral proteinase in a mass ratio of 1:1:1.

[0021] Furthermore, in step (4), the complex enzyme and the mixed solution are enzymatically hydrolyzed at 50° C. for 3 to 5 hours.

[0022] Furthermore, the enzymatic hydrolysis time in step (4) is 4 hours.

[0023] Compared with the prior art, the beneficial effects of the present invention are as follows: the present invention utilizes a complex enzyme (amylase, glycoproteinase, neutral protease) to pre-enzymatically hydrolyze the yam matrix, effectively breaking down the cell wall structure and releasing small molecule sugars and amino acids, providing an efficient substrate for subsequent lactic acid bacteria (Lactobacillus jaguar and Lactobacillus pentosus) fermentation, greatly promoting bacterial growth and metabolic activity, and shortening the fermentation cycle; the organic acid produced during the fermentation process cooperates with the enzymatic hydrolysis to further improve the solubility and dissolution properties of the product, avoids agglomeration, and effectively converts the functional components in the raw materials, so that the total flavonoid content, SOD activity and amino acid composition of the final product are significantly improved; in addition, the soft sour aroma produced by fermentation and the sweetness scientifically formulated in the later stage are coordinated with each other, completely covering up the inherent earthy smell and astringency of the yam, and finally obtaining a high-quality yam powder product that is rich in nutrition, mellow in flavor, has good solubility and is easier to be absorbed by the human body. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] The accompanying drawings are used to provide a further understanding of the present invention and constitute a part of the specification. Together with the embodiments of the present invention, they are used to explain the present invention and do not constitute a limitation of the present invention. In the accompanying drawings:

[0025] Figure 1 is the standard curve of rutin;

[0026] Figure 2 is the standard curve of Coomassie brilliant blue protein;

[0027] Figure 3 The following is a table showing the amino acid content of unfermented yam flour;

[0028] Figure 4 The following is a table showing the amino acid content of fermented yam flour;

[0029] Figure 5 This is the thermal spectrum of unfermented yam flour;

[0030] Figure 6 This is the thermal spectrum of fermented yam flour. DETAILED DESCRIPTION

[0031] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of the present invention.

[0032] The present invention is described in further detail below with reference to the examples.

[0033] Specific Example 1 of the method for preparing yam powder by cooperative fermentation of lactic acid bacteria and complex enzymes provided by the present invention:

[0034] A method for preparing yam powder by cooperative fermentation of lactic acid bacteria and complex enzymes comprises the following steps:

[0035] (1) Raw material preparation and sugar solution preparation: Weigh yam powder and germ, mix them evenly in a mass ratio of 2:1 to obtain dry material; prepare a 2% sugar solution for later use;

[0036] (2) Sterilization: The dry material and the sugar solution are sterilized by high-pressure steam, respectively, at 115° C. for 30 min;

[0037] (3) Sterilization and mixing: After sterilization and cooling, add the sugar solution to the dry material in an amount such that the total mass of water in the system is 7:3 to the mass ratio of yam powder, and mix well to form a mixed solution;

[0038] (4) Enzymatic hydrolysis: A composite enzyme preparation was added to the mixed solution for enzymatic hydrolysis. The composite enzyme was composed of amylase, glycoproteinase, and neutral proteinase in a mass ratio of 1:1:1. The amount of composite enzyme added was 0.1 g for every 10 g of yam powder. The composite enzyme and the mixed solution were enzymatically hydrolyzed at 50°C for 4 h.

[0039] (5) Inoculation and fermentation: A mixed lactic acid bacteria solution was inoculated into the enzymatically hydrolyzed material for fermentation. The lactic acid bacteria were prepared by mixing Lactobacillus jaguar and Lactobacillus pentosus in a ratio of 1:1. The inoculation amount was 4% of the total mass of the material, and the fermentation was carried out at a constant temperature of 30°C for 4 days.

[0040] (6) Drying and crushing the prepared materials to obtain the yam powder product.

[0041] After fermentation in step (5) is complete, the material can be flavored: 4-day fermented yam powder was removed from the constant temperature incubator, and different ammonium citrate addition amounts (0.2%, 0.4%, 0.6%, and 0.8%) and different white sugar addition amounts (6%, 8%, 10%, and 12%) were designed. The different amounts of ammonium citrate and white sugar were added to the fermented yam powder. Five people were randomly invited to taste the flavors. Ultimately, the addition of 0.6% ammonium citrate and 10% white sugar was found to be the most suitable for human taste.

[0042] Specific Example 2 of the method for preparing yam powder by cooperative fermentation of lactic acid bacteria and complex enzymes provided by the present invention:

[0043] This example measures the physical and chemical properties of yam flour before and after fermentation based on Example 1;

[0044] 1. Determination of water binding capacity

[0045] Weigh 2.5 g (dry weight) of each unfermented and fermented yam flour, add 37.5 mL of distilled water, stir the suspension in a constant-temperature, constant-speed magnetic stirrer (20°C, 860 rpm) for 1 hour, centrifuge at 3000 rpm for 10 minutes, discard the supernatant, and weigh the suspension (wet weight). The water binding capacity (WBC) is calculated as follows:

[0046] WBC(%)=((Mw-Md) / Md)×100

[0047] Where: Mw is the wet powder mass, unit is g; Md is the dry powder mass, unit is g; the water binding capacity of fermented and unfermented yam powder is shown in Table 1:

[0048] Table 1 Water binding capacity of fermented and unfermented yam flour

[0049]

[0050] 2. Solubility determination

[0051] Aqueous suspensions of fermented and unfermented yam flour were heated in a shaking water bath at 60°C, 70°C, and 80°C for 1 hour. The suspensions were then centrifuged at 3800 rpm for 15 minutes. The samples were then cooled to ambient temperature, and the supernatant was poured into an evaporating dish and dried in an oven at 110°C to constant weight. The solubility calculation formula is:

[0052] SOL (%) = (mass of dried supernatant / dry weight of yam powder) × 100

[0053] The effect of lactic acid bacteria fermentation on the solubility of yam powder is shown in Table 2:

[0054] Table 2 Solubility of fermented and unfermented yam flour

[0055]

[0056] 3. Determination of total flavonoid content

[0057] 2.0 mL of unfermented yam powder and fermented yam powder extract were added to 10 mL stoppered test tubes, and then 0.4 mL of 5% sodium nitrite solution was added. The mixture was allowed to stand for 5 minutes, and 0.4 mL of 5% aluminum nitrate solution was added; it was allowed to stand for another 5 minutes, and 4 mL of 4% NaOH solution and 3.2 mL of ethanol were added. The mixture was shaken vigorously, left for 15 minutes, and the absorbance was measured at 510 nm. Rutin was used as the standard for the calibration curve, and a linear equation was derived from it to determine the total flavonoid content of the sample. The total flavonoid content was expressed in mg rutin equivalent / g dry basis, as follows: Figure 1 Shown is the standard curve of rutin.

[0058] The effect of lactic acid bacteria fermentation on the total flavonoids content of yam flour is shown in Table 3:

[0059] Table 3 Total flavonoids content of fermented and unfermented yam flour

[0060]

[0061] 4. Protein content determination

[0062] Weigh 0.05 g of fermented yam powder and unfermented yam powder in three groups respectively, add distilled water to make up to 10 mL, water bath at 35 ° C for 10 min, shake well and use a pipette to draw 1 mL into a test tube, add 5 mL of Coomassie Brilliant Blue G-250 solution, shake well, let it stand for 5 min, and then measure its absorbance at 595 nm with a UV-visible spectrophotometer. Repeat five times for each group.

[0063] Proteins form complexes with the hydrophobic bonds of Coomassie Brilliant Blue G-250, resulting in a dye-protein complex with a maximum absorbance at 595 nm. Within this range, the absorbance is linearly related to protein concentration. A concentration gradient standard curve is prepared using bovine serum albumin (BSA) as a standard sample, and the protein content in the extract is calculated using a regression equation. This method is commonly used for protein determination and offers high sensitivity.

[0064] The method for preparing the bovine serum albumin (BSA) standard curve is as follows:

[0065] (a) Prepare a bovine serum albumin (BSA) standard solution by weighing 100 mg of BSA into a 100 mL volumetric flask and adding distilled water to the volume to obtain the stock solution.

[0066] (b) Take an appropriate amount of BSA stock solution and dilute it to 0.1 mg / mL.

[0067] (c) Prepare Coomassie Brilliant Blue solution by dissolving 100 mg of the dye in 50 mL of 95% ethanol, adding 100 mL of 85% phosphoric acid, and finally making up to 1000 mL with distilled water. Protect from light.

[0068] (d) Take 7 clean test tubes and add the corresponding reagents according to the requirements of Table 3.

[0069] (e) Mix the reagents in each tube evenly on an oscillator. After standing for 5 minutes, measure the absorbance of each tube at 595 nm using a spectrophotometer and record the values. Draw a standard curve with the absorbance as the ordinate and the BSA concentration as the abscissa, as shown in the following example: Figure 2 Shown is the Coomassie brilliant blue protein standard curve.

[0070] The effect of lactic acid bacteria fermentation on the protein content of yam flour is shown in Table 4:

[0071] Table 4 Protein content of fermented and unfermented yam powder

[0072]

[0073] 5. SOD determination

[0074] The effect of lactic acid bacteria fermentation on SOD of yam powder was determined by the method provided in the SOD kit instruction manual of Nanjing Jiancheng Bioengineering Institute, as shown in Table 5:

[0075] Table 5 SOD activity of fermented and unfermented yam powder

[0076]

[0077] 6. Determination of microbial indicators

[0078] The total number of colonies of fermented yam powder was determined according to the national standards GB4789.2-2016, GB4789.3-2016 and GB4789.4-2016. 1 g of fermented yam powder was placed in a sterile bottle with 10 mL of physiological saline to prepare a 1:10 sample solution; 1 mL of the 1:10 sample solution was slowly injected along the wall of a sterile test tube containing 9 mL of physiological saline, taking care not to touch the dilution liquid surface with the pipette, and the test tube was shaken to mix evenly to prepare a 1:100 sample solution; 1 mL of the 1:100 sample solution was slowly injected along the wall of a sterile test tube containing 9 mL of physiological saline, and the test tube was shaken to mix evenly to prepare a 1:1000 sample solution. According to the principle that each living bacterium can grow a colony, 1 mL of the 1:10, 1:100 and 1:1000 sample solutions were respectively taken and placed in sterile plates, with 3 plates in parallel for each concentration. At the same time, 1 mL of blank diluent was taken and added to 3 sterile plates as a blank control, and 1 mL of standard bacterial solution was taken and added to 3 sterile plates as a positive control. Agar medium was added and mixed evenly, and incubated at 37°C for 48 h. According to the number of colonies cultured, the number of viable bacteria was calculated. According to the number of colonies cultured, the number of viable lactic acid bacteria in the fermented yam powder was 146000 CFU / g.

[0079] 7. Determination of amino acid content

[0080] Accurately weigh 20-200mg of solid sample (adjust according to protein content) and add it to a hydrolysis tube. Add 10ml of analytical grade hydrochloric acid (approximately 6M) in a 1:1 ratio, blow nitrogen into the tube for 30 seconds, seal it, and place it in an oil bath at 110°C for hydrolysis for 22-24 hours. After the hydrolysis is completed, cool it to room temperature, filter it through a 0.45μm filter into a 50ml volumetric flask, and adjust the volume. Pipette 2ml of the adjusted sample and deacidify it on a rotary evaporator at 45°C. Dry it until a small amount of solid or trace remains at the bottom of the flask. Add 2ml of sodium citrate buffer to fully dissolve it. Filter it through a 0.45μm filter, and analyze it on a liquid analyzer. The amino acid content of unfermented yam powder is shown in Table 3, and the amino acid content of fermented yam powder is shown in Table 4.

[0081] It should be noted that, in this document, terms such as "comprises", "includes" or any other variations thereof are intended to cover non-exclusive inclusion, so that a process, method, article or apparatus that includes a series of elements includes not only those elements, but also other elements not explicitly listed, or also includes elements that are inherent to such process, method, article or apparatus.

[0082] While embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions and variations may be made to these embodiments without departing from the principles and spirit of the invention, and that the scope of the invention is defined by the appended claims and their equivalents.

Claims

1. A method for preparing yam powder by synergistic fermentation of lactic acid bacteria and complex enzymes, characterized in that: The following steps are involved: (1) Raw material preparation and sugar solution preparation: Weigh yam powder and germ, mix them evenly in a mass ratio of 2:1 to obtain dry material; prepare a 2% sugar solution for later use; (2) Sterilization: The dry material and the sugar aqueous solution are sterilized by high-pressure steam, respectively, at 115°C for 30 min; (3) Sterilization and mixing: After sterilization and cooling, add the sugar solution to the dry material and mix well to form a mixed solution; (4) Enzymatic hydrolysis: adding a composite enzyme preparation to the mixed solution for enzymatic hydrolysis, wherein the composite enzyme comprises amylase, glycoproteinase and neutral protease; (5) Inoculation and fermentation: A mixed lactic acid bacteria solution is inoculated into the enzymatically hydrolyzed material for fermentation. The lactic acid bacteria consist of Lactobacillus jaguariensis and Lactobacillus pentosus, and the inoculation amount is 2%-8% of the total mass of the material. The fermentation is carried out at a constant temperature of 30°C for 4 days. (6) Drying and crushing the prepared materials to obtain the yam powder product.

2. The method for preparing yam powder by cooperative fermentation of lactic acid bacteria and complex enzyme according to claim 1, characterized in that: After the fermentation is completed in step (5), the material is flavored: sodium citrate and white sugar are added to the fermented and enzymatically hydrolyzed material and stirred evenly.

3. The method for preparing yam powder by cooperative fermentation of lactic acid bacteria and complex enzyme according to claim 2, characterized in that: The amount of sodium citrate added is 0.6% of the total mass of the material, and the amount of white sugar added is 10% of the total mass of the material.

4. The method for preparing yam powder by cooperative fermentation of lactic acid bacteria and complex enzyme according to claim 1, characterized in that: The amount of the sugar aqueous solution added in step (3) is such that the mass ratio of the total mass of water in the system to the mass of the yam powder is 7:

3.

5. The method for preparing yam powder by cooperative fermentation of lactic acid bacteria and complex enzyme according to claim 1, characterized in that: The inoculation amount of the lactic acid bacteria mixed liquid in step (5) is 4% of the total mass of the material.

6. The method for preparing yam powder by cooperative fermentation of lactic acid bacteria and complex enzyme according to claim 5, characterized in that: In the step (5), the lactic acid bacteria are prepared by mixing Lactobacillus jaguar and Lactobacillus pentosus in a ratio of 1:

1.

7. The method for preparing yam powder by cooperative fermentation of lactic acid bacteria and complex enzyme according to claim 1, characterized in that: The amount of compound enzyme added in step (4) is 0.1 g of compound enzyme per 10 g of yam powder.

8. The method for preparing yam powder by cooperative fermentation of lactic acid bacteria and complex enzyme according to claim 7, characterized in that: In step (4), the complex enzyme is composed of amylase, glycoproteinase and neutral proteinase in a mass ratio of 1:1:

1.

9. The method for preparing yam powder by cooperative fermentation of lactic acid bacteria and complex enzyme according to claim 8, characterized in that: In the step (4), the complex enzyme and the mixed solution are enzymatically hydrolyzed at 50° C. for 3 to 5 hours.

10. The method for preparing yam powder by cooperative fermentation of lactic acid bacteria and complex enzymes according to claim 9, characterized in that: The enzymatic hydrolysis time in step (4) is 4 hours.

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