MOF (Metal Organic Framework) enzyme, preparation method thereof and method for catalyzing and clarifying fruit juice by using MOF enzyme
By embedding enzymes in Cu-BTC complex and combining vitamin C protective enzymes, the problem of poor activity and stability of biological enzymes in juice is solved, the light transmittance and quality of juice is improved, and the effect of low sugar and low browning is achieved. Meets the need for technological upgrades in the juice process.
Patent Information
- Application Number
- CN202411588403.2
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-11-08
- Publication Date
- 2025-01-14
AI Technical Summary
The existing juice clarification process has problems such as long processing time, high energy consumption, incomplete clarification, and affecting the taste and flavor of juice. Moreover, the activity and stability of biological enzymes in juice are poor, resulting in low enzyme catalytic efficiency and increased production costs.
The Cu-BTC complex embed enzyme consisting of copper sulfate pentahydrate, complex biological enzyme and BTC is used to form MOF enzymes, and the enzyme is immobilized through hydrogen bonds and coordination bonds, reducing the leaching of the enzyme and enhancing its stability. Combining vitamin C protects the enzyme from oxidative damage, provides appropriate catalytic conditions, and improves the activity and efficiency of the enzyme.
Improve the catalytic activity and stability of enzymes, reduce the amount of enzymes, improve the light transmittance and quality of juice, reduce the sugar content and browning degree, extend the shelf life of juice, and meet the consumers' high-quality low-sugar diet need.
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Abstract
Description
Technical Field
[0001] The invention relates to the technical field of agricultural product processing, and in particular to a MOF enzyme and a preparation method thereof, and a method for clarifying fruit juice catalyzed by the MOF enzyme. Background Art
[0002] Clarification is an important process in the processing of fruit and vegetable juice. Generally, fruit juice contains rich nutrients such as polysaccharides, polyphenols, pectin, vitamins and proteins, which cause the juice to be turbid and precipitated. Therefore, juice clarification is crucial to improving the appearance and nutritional value, ensuring consumer perception and market value. At present, there are juice clarification processes such as adsorption clarification, cold and hot treatment clarification and ultrafiltration clarification on the market, but the existing clarification processes have the disadvantages of long processing time, high energy consumption, incomplete clarification, and affecting the taste and flavor of the juice. Enzymatic clarification of fruit and vegetable juice not only overcomes these shortcomings, but also retains the nutrients in the juice to the maximum extent, improves the juice yield and light transmittance, and greatly improves the quality of the juice.
[0003] However, enzyme catalysis is a chemical reaction process, and its efficiency is affected by the amount, activity and catalytic conditions of the biological enzyme. When the conditions cannot meet the catalytic reaction, it will lead to reduced production efficiency and increased production costs. Therefore, in the juice clarification process, efficient use of enzyme activity and improved enzyme reaction efficiency have become a breakthrough point for technological upgrading in the juice process.
[0004] Natural ingredients in juice, such as organic acids and sugars, will have a certain effect on the activity of enzymes. For example, too high or too low pH in juice will affect the expression of enzyme activity, thereby affecting the activity of the enzyme.
[0005] Therefore, how to improve the activity and stability of enzymes in juice, efficiently utilize enzyme activity, and improve enzyme reaction efficiency has become a breakthrough point in the technological upgrade of juice processing. Summary of the invention
[0006] In order to solve the problem that the existing biological enzymes are prone to poor stability and / or activity in the juice clarification process, thereby making the effect of using biological enzymes to clarify juice unsatisfactory, the present invention provides a MOF enzyme and a preparation method thereof and a method for MOF enzyme-catalyzed juice clarification.
[0007] In a first aspect, the present application provides a MOF enzyme:
[0008] A MOF enzyme, whose raw materials include 0.4-2 parts of copper sulfate pentahydrate, 0.3-2 parts of complex biological enzyme and 1-2 parts of BTC ethanol solution with a mass fraction of 50% (v / v).
[0009] The MOF enzyme includes a Cu-BTC complex composed of a metal node copper ion and an organic ligand 1,3,5-benzenetricarboxylic acid. The Cu-BTC complex provides interactions such as hydrogen bonds and coordination bonds to support enzyme immobilization. The Cu-BTC complex with mesopores can reduce enzyme leaching and enhance its stability.
[0010] The invention embeds the biological enzyme in the Cu-BTC complex, provides suitable conditions for enzyme catalysis, thereby enhancing the catalytic activity of the enzyme, improving the efficiency of clarifying the roxburghii juice by the exogenous enzyme activity, reducing the enzyme dosage and improving the catalytic efficiency, thereby achieving the improvement of light transmittance, reducing the sugar content and the browning degree, and reducing the loss of nutrients.
[0011] Furthermore, the method comprises 0.8-1.5 parts of copper sulfate pentahydrate, 0.5-1.5 parts of complexing enzyme and 1-2 parts of BTC in a mass fraction of 50% (v / v) ethanol solution.
[0012] Furthermore, the biological enzyme includes one or more of cellulase, pectinase and glucose oxidase.
[0013] Furthermore, the biological enzyme is a combination of cellulase, pectinase and glucose oxidase.
[0014] Furthermore, the raw material of the MOF enzyme also includes 0.05-0.1 parts by mass of vitamin C.
[0015] Vitamin C is a strong antioxidant. Vitamin C and biological enzymes are embedded in the Cu-BTC complex, which can better protect the enzymes from oxidative damage, thereby maintaining the activity of the enzymes, promoting the expression of enzyme activity, and thus improving the effect of catalytic clarification of juice.
[0016] In a second aspect, the present application provides a method for preparing a MOF enzyme;
[0017] A method for preparing a MOF enzyme comprises the following preparation steps:
[0018] S1. A copper sulfate pentahydrate solution with a mass fraction of 0.3-0.5 wt%, a biological enzyme solution with a mass fraction of 70-90 wt%, and a 50% (v / v) ethanol solution of 1-3 g of BTC are stirred and mixed for 60-90 minutes to obtain an enzyme mixture;
[0019] S2. Centrifuge the enzyme mixture to separate the MOF enzyme solid and liquid, and dry the MOF enzyme solid at 100-105° C. for 12-15 hours to obtain the MOF enzyme.
[0020] The three-dimensional active structure of enzymes is mainly maintained by weak non-covalent interactions, so heat, pH and chemical stress factors can alter or eliminate activity. Metal-organic frameworks (MOFs) are extended porous network materials assembled from metal-based nodes and organic connectors through a bottom-up building block approach and can be used to provide protection for enzymes. Cu-BTC morphology adopts a rod-like structure at moderate temperatures (100-110°C) and transforms into a round structure at higher temperatures.
[0021] Furthermore, 0.05-0.1 parts by mass of vitamin C is added in step S1.
[0022] In a third aspect, the present application provides a method for clarifying juice using MOF enzyme catalysis:
[0023] A method for clarifying juice using MOF enzyme catalysis, comprising the following preparation steps:
[0024] D1. Squeeze the fresh fruit juice, filter out the pomace, and set aside the juice;
[0025] D2, adding MOF enzyme to the juice obtained in D1 and performing enzymolysis, wherein the enzymolysis temperature is 40-60°C and the enzymolysis time is 20-30 min to obtain an enzyme juice mixture; the concentration of MOF enzyme in the juice is 1-10 mg / mL;
[0026] D3, performing enzyme passivation treatment on the MOF enzyme in the enzyme juice mixture after enzymatic hydrolysis;
[0027] D4, centrifugally separating the mixture obtained in step D3, and filtering to obtain clarified juice;
[0028] The MOF enzyme, in parts by mass, comprises raw materials of 1-2 parts of copper sulfate pentahydrate, 0.3-2 parts of complex biological enzyme and 1-2 parts of BTC in 50% (v / v) ethanol solution.
[0029] Furthermore, the concentration of the MOF enzyme in the juice is 2-5 mg / mL; and the fresh fruit is fresh Rosa roxburghii fruit.
[0030] Furthermore, the temperature of the enzymatic hydrolysis is 50°C.
[0031] Beneficial effects:
[0032] The MOF enzyme in the present invention comprises a Cu-BTC complex composed of a metal node copper ion and an organic ligand 1,3,5-benzenetricarboxylic acid. The biological enzyme is compositely embedded in the Cu-BTC complex. The Cu-BTC complex provides interactions such as hydrogen bonds and coordination bonds, supports enzyme immobilization, and provides suitable conditions for enzyme catalysis. The Cu-BTC complex with mesopores can reduce enzyme leaching and enhance its stability, thereby enhancing the catalytic activity of the enzyme, improving the efficiency of clarifying fruit juice with exogenous enzyme activity, reducing enzyme dosage and improving catalytic efficiency, thereby achieving the purposes of improving light transmittance, reducing sugar content and browning degree, and reducing nutrient loss. DETAILED DESCRIPTION
[0033] In order to make the technical solution of the present invention clearer, the present invention is further described in detail with reference to specific embodiments.
[0034] Example 1-1
[0035] A MOF enzyme comprises 0.8 g of copper sulfate pentahydrate, 0.5 g of complex biological enzyme (a composition of cellulase, pectinase and glucose oxidase in a mass ratio of 1:1:1) and 1 g of BTC in an ethanol solution with a mass fraction of 50% (v / v), and a preparation method thereof comprises the following preparation steps:
[0036] S1. Copper sulfate pentahydrate solution (copper ion mass fraction of 0.3 wt%), biological enzyme solution with a mass fraction of 70 wt% and 1 g of BTC 50% (v / v) ethanol solution (ethanol to water volume ratio of 1:1) were stirred and mixed for 60-90 minutes to obtain an enzyme mixture.
[0037] S2. Centrifuge the enzyme mixture to obtain a MOF enzyme solid and separate the liquid. Dry the MOF enzyme solid at 100-105° C. for 12-15 hours to obtain the MOF enzyme.
[0038] Example 1-2
[0039] A MOF enzyme, which is different from the MOF enzyme in Example 1-1 in that the raw materials of the MOF enzyme also include 0.08 g of vitamin C.
[0040] Examples 1-3
[0041] A MOF enzyme, which is different from the MOF enzyme in Example 1-1 in that pectinase is used in an equal amount to replace glucose oxidase in the biological enzyme.
[0042] Example 2-1
[0043] A method for clarifying juice by MOF enzyme catalysis, comprising the following preparation steps:
[0044] D1. Use a washing device to roll the ripe fresh sea buckthorn fruit to clean the dirt and residual pesticides on the surface; after cleaning, use a belt filter press to squeeze and filter the fruit residue, and the juice is ready for use.
[0045] D2. Add MOF enzyme (using the MOF enzyme of Example 1) to the juice obtained in S1 for enzymolysis, and perform enzymolysis treatment at a constant temperature of 50° C. for 25 min to obtain an enzyme juice mixture; the concentration of MOF enzyme in the juice is 5 mg / mL.
[0046] D3. The MOF enzyme in the enzyme juice mixture obtained in step D2 is deactivated. The enzyme deactivation conditions are 90° C. for 10 min.
[0047] D4. After treatment with MOF enzyme, the clarified roxburghii juice is obtained. The MOF enzyme catalyst is recovered by centrifugation, washed, vacuum dried at 50°C for 2 hours, and recycled;
[0048] Example 2-2
[0049] A method for clarifying fruit juice using MOF enzyme catalysis is different from the method for clarifying fruit juice using MOF enzyme catalysis in Example 2-1 in that the MOF enzyme is the MOF enzyme of Example 1-2.
[0050] Example 2-3
[0051] A method for clarifying fruit juice using MOF enzyme catalysis is different from the method for clarifying fruit juice using MOF enzyme catalysis in Example 2-1 in that the MOF enzyme is the MOF enzyme of Example 1-3.
[0052] Comparative Example 1
[0053] A method for clarifying fruit juice by MOF enzyme catalysis is different from the method for clarifying fruit juice by MOF enzyme catalysis in Example 2-1 in that MOF enzyme is not used in step D2, and other conditions are the same.
[0054] Comparative Example 2
[0055] A method for clarifying fruit juice using MOF enzyme catalysis is different from the method for clarifying fruit juice using MOF enzyme catalysis in Example 2-1 in that in step D2, a composition of cellulase, pectinase and glucose oxidase in a mass ratio of 1:1:1 is used to replace the MOF enzyme in equal amounts.
[0056] Experimental Results
[0057] The roxburghii juice obtained by the method of MOF enzyme catalytic juice clarification of Example 2-1, Example 2-2, Example 2-3, Comparative Example 1 and Comparative Example 2 was used as the experimental object, and the transmittance, sugar content and browning degree were tested (n=3 for each experiment, and the average value was taken).
[0058] 1. Transmittance value
[0059] The absorbance was measured at 660 nm using a UV spectrophotometer using the Folin-Ciocalteu method to evaluate the transmittance percentage of the experimental samples. The experimental results are shown in Table 1.
[0060] Table 1. Transmittance values of roxburghii juice obtained by the method of MOF enzyme catalysis to clarify juice according to Example 2-1, Example 2-2, Example 2-3, Comparative Example 1 and Comparative Example 2
[0061]
[0062] The light transmittance of the sea buckthorn juice treated with MOF enzyme was significantly improved compared with that of Comparative Examples 1 and 2, indicating that the catalytic activity of the enzyme was enhanced after the enzyme was immobilized by MOF, which not only enhanced the visual appearance of the juice, but also improved the quality and shelf life of the juice.
[0063] 2. Sugar content
[0064] A handheld refractometer (Erma, Japan) was used to measure TSS at 25±1°C and expressed as Brix%. The results are shown in Table 2.
[0065] Table 2. Sugar content of roxburghii juice obtained by the method of MOF enzyme catalysis clarification of juice in Example 2-1, Example 2-2, Example 2-3, Comparative Example 1 and Comparative Example 2
[0066]
[0067] After treatment with the combined enzyme alone, the sugar content increased significantly (Comparative Example 2), while the sugar content increased slightly after treatment with the MOF enzyme, indicating that the MOF enzyme had less effect on the sugar content compared with no treatment, indicating the selectivity or potential action preference of specific components in the fruit substrate, emphasizing the specificity driven by the MOF enzyme. This result also meets the current consumers' demand for healthy and low sugar.
[0068] Cellulose-pectin complex enzyme acts on fruit substrates. Cellulase can decompose cellulose in plant cell walls and hydrolyze it into simple sugars such as glucose. Cellulose is the main component of plant cell walls. Through the action of cellulase, sugars wrapped in cell walls can be released. Pectinase is responsible for decomposing pectin in the intercellular matrix, destroying the connection between cells, reducing the viscosity of the substrate, and increasing the clarity and yield of juice.
[0069] Through the action of these enzymes, plant cell walls and intercellular matrix are broken down, and sugars originally bound in cells are released, thereby increasing the sugar content of the food. MOF enzyme treatment showed a slight increase in sugar content, and compared with direct treatment with enzyme composition, the effect on sugar content was limited, indicating its selective effect or potential preference for specific components in the sea buckthorn juice substrate. These results emphasize the specificity of MOF enzyme drive. This is also conducive to the healthy low-sugar diet habits of current consumers, which not only improves the taste of juice but also can accurately control sugar.
[0070] 3. Browning degree
[0071] Spectrophotometer method: The degree of browning is evaluated by measuring the absorbance of the sample at a specific wavelength. Take the test sample and add 50% (v / v) ethanol. After centrifugation, use a spectrophotometer to measure the absorbance at 420nm. The absorbance directly indicates the degree of browning. The experimental results are shown in Table 3.
[0072] Table 3. Browning degree of roxburghii juice obtained by the method of MOF enzyme catalyzed juice clarification in Example 2-1, Example 2-2, Example 2-3, Comparative Example 1 and Comparative Example 2
[0073] distinguish Example 2-1 Example 2-2 Example 2-3 Comparative Example 1 Comparative Example 2 Rosa roxburghii juice 0.067 0.065 0.070 0.085 0.078
[0074] During the juice processing, non-enzymatic browning is very easy to occur, and the dark substances produced have a great impact on the appearance, color, flavor and nutritional value of the juice. Non-enzymatic browning mainly includes four types of reactions: Maillard reaction, polyphenol oxidation condensation reaction, vitamin C oxidation decomposition reaction and caramelization reaction.
[0075] Rosa roxburghii is rich in phenolic substances and vitamin C. Studies have shown that phenolic substances and vitamin C are the only factors that affect the browning of juice. The content of phenolic substances increases during the juice squeezing and sterilization process, which enhances the self-polymerization and mutual oxidation reaction of phenolic substances and leads to browning. Studies have shown that glucose oxidase can reduce the content of dissolved oxygen and reducing sugar in juice, while slowing down the Maillard reaction and the oxidation reaction of vitamin C, to achieve the effect of inhibiting browning.
[0076] The browning degree of Example 2-2 is better than that of Example 2-1, indicating that the use of a small amount of vitamin C in the MOF enzyme reduces the browning degree. This may be because the vitamin C in the MOF enzyme enhances the peroxidase-like catalytic performance and enhances the specificity of the formation of the metal organic framework intermediate, thereby achieving the goal of reducing the dissolved oxygen and reducing sugar content in the juice and the browning reaction.
[0077] The above-mentioned embodiments only express several implementation methods of the present invention, and the description thereof is relatively specific and detailed, but it cannot be understood as limiting the scope of the patent of the present invention. It should be pointed out that, for ordinary technicians in this field, several variations and improvements can be made without departing from the concept of the present invention, which all belong to the protection scope of the present invention. Therefore, the protection scope of the patent of the present invention shall be subject to the attached claims.
Claims
1. A MOF enzyme, characterized in that Calculated by weight, the raw materials include 0.4-2 parts of copper sulfate pentahydrate, 0.3-2 parts of complex biological enzyme and 1-2 parts of BTC ethanol solution with a mass fraction of 50% (v / v).
2. A MOF enzyme according to claim 1, characterized in that: The invention comprises, by weight, 0.8-1.5 parts of copper sulfate pentahydrate, 0.5-1.5 parts of complex biological enzyme and 1-2 parts of BTC in a 50% (v / v) ethanol solution.
3. A MOF enzyme according to claim 1, characterized in that: The biological enzymes include one or more of cellulase, pectinase and glucose oxidase.
4. A MOF enzyme according to claim 2, characterized in that: The biological enzyme is a combination of cellulase, pectinase and glucose oxidase.
5. The MOF enzyme according to claim 1, characterized in that: The raw materials of the MOF enzyme also include 0.05-0.1 parts by mass of vitamin C.
6. A method for preparing the MOF enzyme according to any one of claims 1 to 5, characterized in that: The method comprises the following preparation steps: S1. A copper sulfate pentahydrate solution with a mass fraction of 0.3-0.5 wt%, a biological enzyme solution with a mass fraction of 70-90 wt%, and a 50% (v / v) ethanol solution of 1-3 g of BTC are stirred and mixed for 60-90 min to obtain an enzyme mixture; S2. Centrifuge the enzyme mixture to separate the MOF enzyme solid and liquid, and dry the MOF solid at 100-105° C. for 12-15 hours to obtain the MOF enzyme.
7. The method for preparing the MOF enzyme according to claim 6, characterized in that: In the step S1, 0.05-0.1 parts by mass of vitamin C is added.
8. A method for clarifying juice using MOF enzyme catalysis, characterized in that: The method comprises the following preparation steps: D1. Squeeze the fresh fruit juice, filter out the pomace, and set aside the juice; D2, adding MOF enzyme to the juice obtained in D1 and performing enzymolysis, wherein the enzymolysis temperature is 40-60°C and the enzymolysis time is 20-30 min to obtain an enzyme juice mixture; the concentration of MOF enzyme in the juice is 1-10 mg / mL; D3, performing enzyme passivation treatment on the MOF enzyme in the enzyme juice mixture after enzymatic hydrolysis; D4, centrifugally separating the mixture obtained in step D3, and filtering to obtain clarified juice; The MOF enzyme, in parts by mass, comprises raw materials of 1-2 parts of copper sulfate pentahydrate, 0.3-2 parts of complex biological enzyme and 1-2 parts of BTC in 50% (v / v) ethanol solution.
9. The method for clarifying juice using MOF enzyme catalysis according to claim 8, characterized in that: The concentration of the MOF enzyme in the fruit juice is 2-5 mg / mL; and the fresh fruit is fresh roxburghii fruit.
10. The method for clarifying juice using MOF enzyme catalysis according to claim 9, characterized in that: The temperature of the enzymatic hydrolysis is 50°C.