Method for extracting insoluble dietary fibers from Dangshan pear pomace based on synergistic effect of double enzymes

Abstract

The invention discloses a method for extracting insoluble dietary fibers from Dangshan pear pomace based on a synergistic effect of double enzymes, and relates to the technical field of food processing and comprehensive utilization of resources. The method adopts double enzymes (alpha-amylase and protease) for preparation, and comprises the following steps: firstly, mixing 40-mesh Dangshan pear pomace with ultrapure water according to the ratio of 1: 50g / mL, adjusting the pH value of the solution to 6.0, and adding the alpha-amylase; then putting into a shaking table at 60 DEG C to react for 50 minutes, setting the rotating speed to be 200r / min, and taking out after the reaction is finished; when the temperature is reduced to room temperature, adjusting the pH value of the solution to 7.5, adding protease, putting the solution into a shaking table at 45 DEG C to react for 50 minutes, setting the rotating speed to be 200r / min, and taking out the solution at 100 DEG C after the reaction is finished; the method comprises the following steps: extracting fruit residues of Dangshan pears, then deactivating enzymes, filtering, collecting precipitates, and drying in a drying oven at 75 DEG C to obtain dried precipitates, namely the Dangshan pear fruit residues IDF, and by adopting a synergistic effect of double enzymes, starch and protein in the fruit residues are effectively degraded, so that the extraction rate of the IDF is improved and reaches 89.44%, and the method is superior to most single-enzyme extraction methods in the market.

Description

Technical Field

[0001] The invention relates to the technical field of food processing and comprehensive resource utilization, and in particular to a method for extracting insoluble dietary fiber from Dangshan pear pomace based on the synergistic action of two enzymes. Background Art

[0002] Dangshan pear pomace is rich in dietary fiber, minerals and bioactive ingredients, but its traditional processing methods are mainly feed or direct disposal, resulting in resource waste and environmental pollution. Therefore, how to efficiently extract insoluble dietary fiber from pomace and improve its utilization value is a research hotspot in the current food processing industry. The extraction of dietary fiber mainly relies on mechanical crushing, alkaline extraction or single enzymatic hydrolysis. The mechanical method has the problems of serious fiber structure destruction and low yield; although the alkaline extraction method can increase the yield of dietary fiber, it is easy to cause the loss of some nutrients and increase environmental pollution; the single enzymatic hydrolysis method is limited by the selectivity of the enzyme, resulting in low extraction efficiency. Since the pomace contains more starch and protein, the single enzyme method is often difficult to effectively degrade these components, affecting the extraction rate of IDF. In addition, the traditional extraction process does not fully consider the optimization of process parameters, resulting in high energy consumption and long time in the extraction process, which affects industrial application. Therefore, an efficient and environmentally friendly IDF extraction method is urgently needed to improve the yield and optimize the process conditions.

[0003] However, the existing technologies mainly rely on mechanical methods, alkaline extraction or single enzyme methods, but these methods either destroy the fiber structure and have low yields (mechanical methods), or affect product quality and cause environmental pollution (alkaline extraction methods), or have low extraction efficiency due to enzyme selectivity limitations (single enzyme methods); when extracting IDF, there is often a lack of optimization of process parameters, resulting in high energy consumption and long time, which makes it difficult to promote on a large scale; traditional alkaline extraction methods will leave residual chemical substances, affecting the safety and taste of dietary fiber. In response to the above problems, the inventors proposed a method for extracting insoluble dietary fiber from Dangshan pear pomace based on the synergistic action of two enzymes to solve the above problems. Summary of the invention

[0004] In order to solve the problems of easy destruction of fiber structure and low yield during IDF extraction, affecting product quality and causing environmental pollution, the present invention aims to provide a method for extracting insoluble dietary fiber from Dangshan pear pomace based on the synergistic action of two enzymes.

[0005] In order to solve the above technical problems, the present invention adopts the following technical solution: a method for extracting insoluble dietary fiber from Dangshan pear pomace based on the synergistic action of two enzymes, characterized in that it comprises the following steps:

[0006] S1. Experimental materials: Take the pomace of Dangshan Pear (the peel and the flesh near the core), spread the peel directly into a dried fruit electric oven at 65℃ for drying, and grind it into powder with a pulverizer;

[0007] S2, the pulp near the core is mixed with pure water, broken up using a high-speed emulsifying homogenizer, filtered and precipitated, the residue is spread flat in a 65°C dried fruit electric oven for drying, and ground into powder using a pulverizer;

[0008] S3, after mixing the two powders, sieve them according to the mesh, put the obtained pomace powder into a sealed can, and store it in a refrigerator at 4°C;

[0009] S4. Experimental process: Double enzymes (α-amylase and protease) were used for preparation. First, 40-mesh 'Dangshan Pear' pomace was mixed with ultrapure water at a ratio of 1:50 g / mL, the pH of the solution was adjusted to 6.0, and α-amylase was added;

[0010] S5, then put it into a shaking table at 60°C for 50 minutes, set the speed to 200r / min, and take it out after the reaction is completed;

[0011] S6, when the temperature drops to room temperature, adjust the solution pH to 7.5, add protease, put it in a 45℃ shaker for reaction for 50min, set the speed to 200r / min, and take it out at 100℃ after the reaction is completed;

[0012] S7, then inactivate the enzyme, filter, collect the precipitate and put it into a 75°C oven for drying, the dried precipitate is the 'Dangshan Pear' pomace IDF;

[0013] S8. Single factor experiment: With particle size, liquid-to-liquid ratio, enzyme addition and enzymatic hydrolysis time as single factors, single factor experiments were designed for particle size (40, 60, 80, 100, 120 mesh), liquid-to-liquid ratio (30:1, 40:1, 50:1, 60:1, 70:1 g / mL), enzyme addition (0.5%, 1.0%, 1.5%, 2.0%, 2.5%) and enzymatic hydrolysis time (10, 30, 50, 70, 90 min) to explore the effects of the four factors on DIDF yield;

[0014] S9. Response surface optimization experiment: Based on the results of single-factor experiments, four factors, including particle size, liquid-to-solid ratio, enzyme addition amount and enzymatic hydrolysis time, were designed as independent variables, and IDF yield was taken as the dependent variable. Data analysis was used to design a four-factor three-level response surface experiment, and then the optimized extraction process of D IDF yield was obtained.

[0015] Compared with the prior art, the present invention has the following beneficial effects:

[0016] 1. The synergistic effect of double enzymes is adopted to effectively degrade starch and protein in pomace, and the extraction rate of IDF is increased to 89.44%, which is better than most single enzyme extraction methods on the market;

[0017] 2. Through single-factor experiments and response surface optimization, the optimal process conditions (particle size, liquid-to-solid ratio, enzyme addition amount and enzymatic hydrolysis time) were clarified to ensure that high-purity IDF can be obtained under the premise of low cost and low energy consumption, thereby improving the feasibility of industrial production.

[0018] 3. It uses pure biological enzymatic hydrolysis, without the need to add additional chemical reagents, which reduces environmental pollution and improves the safety and natural properties of dietary fiber, which is in line with the development trend of the food health industry;

[0019] 4. The introduction of high-speed emulsification homogenization and screening technology optimizes the enzymatic hydrolysis process by graded treatment of the peel and the flesh near the core. The final IDF product has better uniformity and functional properties, which is more in line with market demand. It provides an efficient and high value-added utilization method, allowing the pomace to be converted into high-quality dietary fiber, broadening the application areas of the pomace, such as its wide application in functional foods, health foods, food ingredients and other fields. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0021] Figure 1 This is a schematic diagram showing the effect of particle size on DIDF yield in the present invention;

[0022] Figure 2 Schematic diagram showing the effect of the liquid-to-solid ratio on the DIDF yield of the present invention;

[0023] Figure 3 Schematic diagram showing the effect of the enzyme addition amount on the DIDF yield of the present invention;

[0024] Figure 4 This is a schematic diagram showing the effect of the enzymatic hydrolysis time on the DIDF yield of the present invention;

[0025] Figure 5 It is a schematic diagram of the response surface AB of the present invention;

[0026] Figure 6 It is a schematic diagram of the contour map AB of the present invention;

[0027] Figure 7 It is a schematic diagram of the response surface AD ​​of the present invention;

[0028] Figure 8 It is a schematic diagram of the contour map AD of the present invention;

[0029] Fig. 9It is a schematic diagram of the response surface BD of the present invention;

[0030] Fig.10 It is a schematic diagram of the contour map BD of the present invention;

[0031] Fig.11 It is a CD schematic diagram of the response surface of the present invention;

[0032] Fig.12 It is a schematic diagram of the contour map CD of the present invention.

[0033] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. 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 creative work are within the scope of protection of the present invention.

[0034] Example: Figure 1-12 As shown, the present invention provides a method for extracting insoluble dietary fiber from Dangshan pear pomace based on the synergistic action of two enzymes, characterized in that it comprises the following steps:

[0035] S1. Experimental materials: Take the pomace of Dangshan Pear (the peel and the flesh near the core), spread the peel directly into a dried fruit electric oven at 65℃ for drying, and grind it into powder with a pulverizer;

[0036] S2, the pulp near the core is mixed with pure water, broken up using a high-speed emulsifying homogenizer, filtered and precipitated, the residue is spread flat in a 65°C dried fruit electric oven for drying, and ground into powder using a pulverizer;

[0037] S3, after mixing the two powders, sieve them according to the mesh size, put the obtained pomace powder into a sealed can, and store it in a refrigerator at 4°C;

[0038] S4. Experimental process: Double enzymes (α-amylase and protease) were used for preparation. First, 40-mesh 'Dangshan Pear' pomace was mixed with ultrapure water at a ratio of 1:50 g / mL, the pH of the solution was adjusted to 6.0, and α-amylase was added;

[0039] S5, then put it into a shaking table at 60°C for 50 minutes, set the speed to 200r / min, and take it out after the reaction is completed;

[0040] S6, when the temperature drops to room temperature, adjust the solution pH to 7.5, add protease, put it in a 45℃ shaker for reaction for 50min, set the speed to 200r / min, and take it out at 100℃ after the reaction is completed;

[0041] S7, then inactivate the enzyme, filter, collect the precipitate and put it into a 75°C oven for drying, the dried precipitate is the 'Dangshan Pear' pomace IDF;

[0042] S8. Single factor experiment: With particle size, liquid-to-liquid ratio, enzyme addition and enzymatic hydrolysis time as single factors, single factor experiments were designed for particle size (40, 60, 80, 100, 120 mesh), liquid-to-liquid ratio (30:1, 40:1, 50:1, 60:1, 70:1 g / mL), enzyme addition (0.5%, 1.0%, 1.5%, 2.0%, 2.5%) and enzymatic hydrolysis time (10, 30, 50, 70, 90 min) to explore the effects of the four factors on DIDF yield;

[0043] See attached Figure 1 Effect of particle size on DIDF yield:

[0044] Under the conditions of liquid-to-solid ratio of 30:1 g / mL, enzyme addition of 0.5%, and enzymatic hydrolysis time of 50 min, the particle size was set to 40, 60, 80, 100, and 120 meshes, and the IDF yield was determined. This experiment was divided into five groups, and three parallel samples were designed for each group of experiments. The mean of the three samples was then taken to explore the effect of different mesh sizes on the D IDF extraction effect. As the particle size increased, the IDF extraction rate tended to decrease. Since the minimum particle size of the 'Dangshan Pear' pomace was 40 meshes when sieving, the minimum particle size of the particle size single factor experiment was set to 40 meshes. When the particle size was 40 meshes, the IDF extraction rate was the highest, at 75.12%; when the particle size exceeded 60 meshes, the IDF extraction rate showed a significant decrease. The reason may be that as the particle size increased, part of the connection structure of the IDF in the 'Dangshan Pear' pomace was destroyed during the crushing process, resulting in a significant decrease in the IDF extraction rate. In summary, the particle size selection is more appropriate as 40 mesh;

[0045] See attached Figure 2 Effect of liquid-to-solid ratio on DIDF yield:

[0046] Under the conditions of a particle size of 40 mesh, an enzyme addition amount of 0.5%, and an enzymatic hydrolysis time of 50 min, the liquid-to-solid ratio was set to 30:1, 40:1, 50:1, 60:1, and 70:1 g / mL, respectively, to determine the yield of IDF. This experiment was divided into five groups, and three parallel samples were designed for each group of experiments. The mean of the three samples was then taken to explore the effect of different liquid-to-solid ratios on the extraction effect of D IDF. As can be seen from the figure: the IDF extraction rate first increased and then decreased with the increase of the liquid-to-solid ratio, reaching a peak of 82.22% at a liquid-to-liquid ratio of 50 g / mL. When the liquid-to-solid ratio is in the range of 30-50 g / mL, the IDF extraction rate tends to increase. The reason may be that when the liquid-to-solid ratio is too low, as the ratio continues to increase, the enzymatic hydrolysis reaction between the enzyme and the substrate becomes intense; in the range of 50-70 g / mL, the IDF extraction rate tends to decrease, which may be due to the fact that as the ratio is too high, the substrate concentration is diluted, resulting in insufficient contact between the enzyme and the substrate. In summary, it is more appropriate to select a solid-liquid ratio of 50 g / mL;

[0047] See attached Figure 3 Effect of enzyme addition on DIDF yield:

[0048] Under the conditions of particle size of 40 mesh, liquid-to-solid ratio of 50 g / mL, and enzymatic hydrolysis time of 50 min, the enzyme addition amount was set to 0.5, 1.0, 1.5, 2.0, and 2.5%, respectively, and the IDF yield was determined. The experiment was divided into five groups, and three parallel samples were designed for each group of experiments. The mean of the three samples was then taken to explore the effect of different enzyme addition amounts on the D IDF extraction efficiency. Figure 3 It can be seen that with the increase of enzyme addition, the IDF extraction rate shows a trend of first increasing and then decreasing. When the enzyme addition amount is between 0.5 and 1.5%, the IDF extraction rate shows a trend of continuous increase, reaching a peak of 88.34% when the enzyme addition amount is 1.5%. The reason may be that α-amylase will enzymatically break part of the structure of IDF in the 'Dangshan Pear' pomace into SDF components with a smaller degree of polymerization. With the continuous increase of enzyme addition, the influence of α-amylase on IDF gradually decreases, and the IDF extraction rate continues to increase. When the enzyme addition amount is between 1.5 and 2.5%, the extraction rate slowly decreases. This is because the enzyme concentration and substrate concentration reach saturation. As the enzyme amount increases, the extraction efficiency of IDF is inhibited. In summary, it is more appropriate to choose an enzyme addition amount of 1.5%;

[0049] See attached Figure 4 Effect of enzymatic hydrolysis time on DIDF yield:

[0050] Under the conditions of particle size of 40 mesh, liquid-to-solid ratio of 50g / mL, and enzyme addition of 1.5%, the enzymatic hydrolysis time was set to 10, 30, 50, 70, and 90min, respectively, and the yield of IDF was determined. The experiment was divided into five groups, and three parallel samples were designed for each group of experiments. Then the mean of the three samples was taken to explore the effect of different enzymatic hydrolysis times on the extraction efficiency of D IDF. The results are shown in the figure below. The IDF extraction rate first increased and then decreased with the increase of enzymatic hydrolysis time. When the enzymatic hydrolysis time was in the range of 10 to 90min, the IDF extraction rate tended to increase, and the extraction rate reached a peak of 89.24% at 70min. The reason for this may be that as the reaction time increased, the starch and protein in the 'Dangshan Pear' pomace were fully enzymatically hydrolyzed, which was conducive to the analysis of IDF components. When the enzymatic hydrolysis time is in the range of 70 to 90 minutes, the extraction rate tends to decrease. The reason may be that as the reaction time increases, the probability of the hydroxyl groups in the solution and the hydrogen atoms of IDF combine increases, resulting in partial hydrolysis of IDF, which leads to a decrease in the extraction rate. In summary, the enzymatic hydrolysis time of 70 minutes is more appropriate.

[0051] S9, response surface optimization experiment: Based on the results of single factor test, four factors, including particle size, liquid-to-solid ratio, enzyme addition amount and enzymatic hydrolysis time, were designed as independent variables, and IDF yield was used as the dependent variable. A four-factor three-level response surface experiment was designed using data analysis to obtain the optimized extraction process of D IDF yield.

[0052]

[0053]

[0054]

[0055]

[0056] By fitting the data with the statistical response surface, the four-variable quadratic regression equation for the IDF extraction rate (Y) is obtained as follows:

[0057] Y=82.19-10.84A+0.83B-0.80C-2.24D-1.37AB-0.36AC-1.50AD-0.29BC+1.12BD-2.68CD-3.85A2-5.61B2-3.97C2-1.73D2.

[0058] The F test was used to determine whether the factors in the regression model were significant for the optimized response value. P value <0.05 indicated that the factor had a significant difference, and P value <0.001 indicated that the factor had an extremely significant difference. According to the figure above, the regression model had an extremely significant difference (P <0.0001), and the lack of fit term was not significant (P = 0.3965 > 0.05), indicating that the model was less contingent. The coefficient of determination R2 = 0.9924, and the adjusted coefficient of determination R2adj = 0.9849, indicating that the regression model had a high degree of fit, good feasibility, and the model was established. The variance analysis data showed that the linear terms A and D, the quadratic terms A2, B2, C2, D2, and the interaction term CD had an extremely significant effect on the IDF extraction rate; the linear terms B and C, and the interaction terms AB, AD, and BD had a significant effect on the IDF extraction rate; the interaction terms AC and BC had no significant effect on the IDF extraction rate. The factors affecting IDF extraction rate can be ranked as: A>D>B>C, i.e., particle size>enzymatic hydrolysis time>liquid-to-solid ratio>enzyme addition amount;

[0059] Statistical analysis was performed based on the quadratic regression equation. Figure 5-12 The three-dimensional response surface and contour map shown in the figure reflect the central value of any two of the four fixed factors and the influence of the interaction between the remaining two factors on the IDF extraction process. The response surface and contour map can intuitively reflect the degree of influence of the interaction between particle size, liquid-to-solid ratio, enzyme addition amount, and enzymatic hydrolysis time on the response value. The steeper the surface and the denser the contour lines, the more significant the influence. The closer the contour lines are to the ellipse, the stronger the interaction between the two factors. By comparing the two-factor response surface maps in the above figure, it can be seen that the response surface AD ​​is steeper than AB, indicating that the interaction AD has a greater impact on the IDF extraction rate. Analysis of each contour map shows that the CD contour map is the densest and the contour lines are closest to the ellipse, indicating that the interaction between CD has the most significant effect on the response value among the interaction items. The order of influence of each interaction on IDF is: CD>AD>AB>BD, which is consistent with the results of variance analysis table 3. After the response surface optimization test, the optimal extraction conditions for the dual enzyme method of D IDF extraction were obtained by regression model point prediction: particle size 40 mesh, liquid-to-solid ratio 54:1 g / mL, enzyme addition amount 1.46%, and enzymatic hydrolysis time 67.60 min. Under this condition, the DIDF yield was 89.44%. In order to detect the reliability of the regression model point prediction and verify the convenience of the experimental operation, the optimized extraction conditions were adjusted to: particle size 40 mesh, liquid-to-solid ratio 54:1 g / mL, enzyme addition amount 1.50%, and enzymatic hydrolysis time 68 min. The experiment was repeated 3 times and the average value was taken. The final D IDF extraction rate was 89.26%, which was only 0.18% different from the theoretical prediction value, indicating that the model fits the response surface optimization experiment well and the optimized extraction process is very reliable. Effects and advantages of the present invention: 1. The extraction rate is significantly improved: the IDF yield of the traditional mechanical method is generally between 50% and 70%, and the single enzyme method usually does not exceed 80%. The dual enzyme synergistic process of the present invention increases the IDF extraction rate to 89.44%, far exceeding the existing technology on the market. 2. Better product quality: The synergistic effect of dual enzymes reduces the residual non-fiber impurities, making the final product higher in purity, better in color, and more complete in structure, making it more suitable for application in the fields of food and health products. 3. Lower energy consumption and higher production efficiency: By optimizing the enzymatic hydrolysis conditions (particle size 40 mesh, liquid-to-solid ratio 54:1 g / mL, enzyme addition 1.46%, enzymatic hydrolysis time 67.60 min), the enzyme dosage and enzymatic hydrolysis time are reduced, saving about 20%-30% of energy consumption compared to traditional processes and shortening the production cycle. 4. Process optimization and easy operation: The traditional alkaline extraction method requires high temperature and high alkalinity treatment, complicated steps, and neutralization treatment, which affects the convenience of operation. This method adopts mild pH regulation and temperature control conditions (pH 6.0-7.5, temperature 45℃-60℃), avoiding complex chemical treatment, making the operation easier and the production more stable. 5. Green and environmentally friendly, reducing pollution: The traditional alkaline extraction method uses strong alkali, which will produce a large amount of waste liquid that pollutes the environment. This method does not use any chemical reagents throughout the process, but only uses biological enzyme catalysis, which not only reduces the use of chemical reagents, but also reduces the cost of wastewater treatment, meeting the requirements of green production.

[0060] The standard parts used in the present invention can all be purchased from the market, and special-shaped parts can be customized according to the instructions and the drawings. The specific connection methods of each part adopt conventional means such as mature bolts, rivets, welding, etc. in the prior art. Machinery, parts and equipment all adopt conventional models in the prior art, and the circuit connection adopts the conventional connection method in the prior art, which will not be described in detail here.

[0061] Obviously, those skilled in the art can make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if these modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include these modifications and variations.

Claims

1. A method for extracting insoluble dietary fiber from Dangshan pear pomace based on the synergistic action of two enzymes, characterized in that: The following steps are involved: S1. Experimental materials: Take the pomace of Dangshan Pear (the peel and the flesh near the core), spread the peel directly into a dried fruit electric oven at 65℃ for drying, and grind it into powder with a pulverizer; S2, the pulp near the core is mixed with pure water, broken up using a high-speed emulsifying homogenizer, filtered and precipitated, the residue is spread flat in a 65°C dried fruit electric oven for drying, and ground into powder using a pulverizer; S3, after mixing the two powders, sieve them according to the mesh, put the obtained pomace powder into a sealed can, and store it in a refrigerator at 4°C; S4. Experimental process: Double enzymes (α-amylase and protease) were used for preparation. First, 40-mesh 'Dangshan Pear' pomace was mixed with ultrapure water at a ratio of 1:50 g / mL, the pH of the solution was adjusted to 6.0, and α-amylase was added; S5, then put it into a shaking table at 60°C for 50 minutes, set the speed to 200r / min, and take it out after the reaction is completed; S6, when the temperature drops to room temperature, adjust the solution pH to 7.5, add protease, put it in a 45℃ shaker for reaction for 50min, set the speed to 200r / min, and take it out at 100℃ after the reaction is completed; S7, then inactivate the enzyme, filter, collect the precipitate and put it into a 75°C oven for drying, the dried precipitate is the 'Dangshan Pear' pomace IDF; S8. Single factor experiment: With particle size, liquid-to-liquid ratio, enzyme addition and enzymatic hydrolysis time as single factors, single factor experiments were designed for particle size (40, 60, 80, 100, 120 mesh), liquid-to-liquid ratio (30:1, 40:1, 50:1, 60:1, 70:1 g / mL), enzyme addition (0.5%, 1.0%, 1.5%, 2.0%, 2.5%) and enzymatic hydrolysis time (10, 30, 50, 70, 90 min) to explore the effects of the four factors on DIDF yield; S9. Response surface optimization experiment: Based on the results of single-factor experiments, four factors, including particle size, liquid-to-solid ratio, enzyme addition amount and enzymatic hydrolysis time, were designed as independent variables, and IDF yield was used as the dependent variable. Data analysis was used to design a four-factor three-level response surface experiment, and then the optimized extraction process of DIDF yield was obtained.

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