A pretreatment device for recycling enzymes in oat enzymatic hydrolysate

Abstract

This invention discloses a pretreatment device for regenerating and recovering enzymes from oat hydrolysate, aiming to solve the problems of low enzyme recovery efficiency, easy inactivation, discontinuous operation, and adverse effects on product quality caused by traditional high-temperature enzyme inactivation methods in existing oat hydrolysate solutions. The device mainly includes a pre-cooling circulation loop, a membrane separation unit, an enzyme enrichment unit, and an enzyme reconstitution component. The pre-cooling circulation loop cools the oat hydrolysate to 4°C to 15°C. Utilizing the enzyme's aggregation characteristic at low temperatures, membrane separation is performed using an ultrafiltration membrane component with a molecular weight cutoff of 30kDa to 100kDa, efficiently retaining the enzyme while simultaneously discharging the product. The retained enzyme can be refluxed back to the reaction system for enrichment or added to the enzyme reconstitution component for activity recovery. This invention achieves efficient enzyme recovery and regeneration, significantly reducing the risk of enzyme inactivation and production costs, while avoiding adverse effects on product flavor and quality. It also enables continuous and automated operation, possessing significant industrial application value.

Description

Technical Field

[0001] This utility model relates to the field of enzyme engineering equipment technology, specifically a pretreatment device that can regenerate and recover enzymes from oat hydrolysate. Background Technology

[0002] In the enzymatic hydrolysis of oats, enzymes act as key catalysts in the reaction. However, if the enzymes in the hydrolysate cannot be effectively recovered and reused after the reaction, it will not only waste resources but also increase production costs. Traditionally, after oat enzymatic hydrolysis, the enzymes in the hydrolysate are inactivated by high-temperature plate heat exchange at 95°C. However, this method of enzyme inactivation not only leads to enzyme loss and increases production costs, but the high temperature also affects the flavor of the oat hydrolysate.

[0003] Furthermore, the enzymatic hydrolysate contains a certain amount of starch or macromolecules, which can easily gelatinize during high-temperature plate inactivation of the enzyme. This increases the overall viscosity of the hydrolysate, slowing its flow rate in the pipeline and prolonging the heat exchange time. It may also cause pipe clogging, affecting the final quality of the product. Some enzymatic hydrolysates are inherently sensitive to high temperatures, which can easily destroy some proteins and mucopolysaccharide components, further impacting the quality of the final product.

[0004] To achieve high-quality product production and avoid the negative impact of high temperatures, the enzymes in oat hydrolysate can be recycled. This not only allows for enzyme reuse and reduces costs, but more importantly, it improves the final product quality. Existing enzyme recovery methods generally suffer from low efficiency, easy enzyme inactivation, and co-precipitation of products and enzymes, making it difficult to meet the demands for efficient enzyme recovery and regeneration in industrial production.

[0005] In view of the problems of low enzyme recovery efficiency, easy inactivation, high recovery cost and the significant impact of traditional high-temperature enzyme inactivation on product quality in the existing technology, this utility model provides a pretreatment device that can regenerate and recover enzymes from oat hydrolysate, aiming to achieve efficient and gentle enzyme recovery, reduce production costs and improve product quality. Utility Model Content

[0006] Purpose of this utility model: To achieve efficient recycling of enzymes, significantly reducing production costs; to reduce the risk of enzyme inactivation during the recycling process and maximize enzyme activity; to achieve continuous mechanized operation, reduce labor costs, and improve production efficiency; and to avoid or reduce the impact of high-temperature enzyme inactivation or other recycled solvents on the flavor of enzyme hydrolysate, thereby improving product quality.

[0007] To achieve the above objectives, this utility model provides the following technical solution:

[0008] This pretreatment device mainly consists of a precooling circulation loop, a membrane separation unit, an enzyme enrichment unit, and an enzyme reconstitution component.

[0009] The precooling circulation loop includes a cooling system and a delivery pump, which is used to first enter the oat enzymatic hydrolysate into the cooling system for cooling. Under the action of the cooling system, the enzymatic hydrolysate is cooled to a suitable temperature (preferably 4°C to 15°C), and then pumped into the ultrafiltration membrane module by the delivery pump to prepare for subsequent membrane separation.

[0010] The membrane separation unit uses an ultrafiltration membrane module with a molecular weight cutoff of 30kDa to 100kDa. Under low temperature conditions, enzymes will aggregate to form larger particles. Utilizing this characteristic, the ultrafiltration membrane can effectively retain enzymes, while the products are discharged as permeate, thereby achieving the initial separation of enzymes and products.

[0011] The enzyme enrichment unit returns the enzymes retained by the membrane separation unit to the reaction system through a specific pipeline, thereby enriching the enzymes and enabling them to continue to function in the reaction system, thus improving enzyme utilization. At the same time, the permeate (product) is discharged from the device.

[0012] The enzyme reconstitution assembly includes a container for holding a suitable buffer solution (such as phosphate buffer) and a mixing device. The enzyme precipitated at low temperature is transferred to the container, and the mixing device is used to fully mix it with the buffer solution to achieve enzyme reconstitution. It can also be equipped with a detection device to detect the enzyme activity recovery rate.

[0013] In addition, the device may also include:

[0014] Activity protection component: After the low-temperature treatment process, a stabilizer (such as 5% - 10% glycerol, 1 - 2mM EDTA) is added to the separated enzyme solution through an addition device to reduce enzyme conformational damage and protect enzyme activity.

[0015] Recycling monitoring component: Monitors the activity of enzymes that have been reconstituted and reused in the enzymatic hydrolysis reaction, and records the activity decay after repeated use, so as to adjust process parameters or replace enzymes in a timely manner.

[0016] Temperature control components: Employing a precisely temperature-controlled refrigeration system, the optimal precipitation temperature for the target enzyme (typically 4℃ - 15℃) can be experimentally determined, maintaining a stable low-temperature environment to prevent irreversible enzyme denaturation due to excessively low temperatures. Simultaneously, a gradient cooling design can be implemented, allowing for staged cooling (e.g., from 25℃ → 15℃ → 5℃) to reduce the impact of sudden temperature changes on the enzyme.

[0017] pH adjustment component: Before low-temperature treatment, the pH of the enzyme hydrolysate is adjusted to near the isoelectric point (pI) of the enzyme by adding acid and base reagents, thereby enhancing the precipitation effect and improving the enzyme separation efficiency.

[0018] To further improve the recovery rate, the device also has reserved interfaces to combine with other technologies, such as combining ultrafiltration or magnetic separation components after low-temperature pretreatment, or adding specific adsorption separation components related to affinity tags (such as His-Tag).

[0019] Compared with the prior art, the beneficial effects of this utility model are:

[0020] 1. High-efficiency recovery: By utilizing the low-temperature membrane filtration system and the aggregation characteristics of enzymes at low temperatures, combined with an ultrafiltration membrane with a suitable molecular weight cutoff, the enzymes in oat hydrolysate can be efficiently retained and recovered, significantly improving the enzyme recovery rate.

[0021] 2. Reduce the risk of enzyme inactivation: Low-temperature treatment effectively reduces thermal denaturation, oxidative degradation and autohydrolysis of enzymes during the recycling process, ensuring enzyme activity, improving enzyme regeneration and utilization rate, and avoiding the damage to enzyme activity caused by traditional high-temperature enzyme inactivation.

[0022] 3. Continuous operation: The coordinated operation of the precooling circulation loop, membrane separation unit and enzyme enrichment unit enables continuous operation of enzyme recovery from oat hydrolysate, which significantly improves production efficiency and reduces labor costs.

[0023] 4. Flexible condition optimization: Specialized adjustment components are set up for key conditions of low-temperature separation, such as temperature, pH and additives, which can be flexibly optimized according to different enzymes and reaction conditions, thereby improving the applicability of the device.

[0024] 5. Protects product flavor: It avoids the impact of traditional high-temperature enzyme inactivation methods on the flavor of oat hydrolysate, while reducing problems such as high-temperature sticking, which helps improve the quality and stability of the final product. Attached Figure Description

[0025] Figure 1 This is a schematic diagram of the overall structure of this utility model.

[0026] Figure 2 This is a cross-sectional view of the present invention.

[0027] In the diagram: 1. Cooling system; 2. Transfer pump; 3. Ultrafiltration membrane module; 4. Enzyme enrichment unit; 5. Enzyme reconstitution module; 6. Activity protection module; 7. Mixing device; 10. pH adjustment module; 11. Recycling monitoring module. Detailed Implementation

[0028] In the following description, numerous specific details are set forth in order to provide a more thorough understanding of the present invention. However, it will be apparent to those skilled in the art that the present invention can be practiced without one or more of these details. In other instances, certain technical features well-known in the art have not been described in order to avoid confusion with the present invention.

[0029] like Figure 1-2 As shown, the pretreatment device for enzymes in oat hydrolysate of this invention mainly includes a precooling circulation loop, a membrane separation unit, an enzyme enrichment unit 4, and an enzyme reconstitution component 5.

[0030] Precooling loop:

[0031] The enzymatically hydrolyzed oat hydrolysate first enters cooling system 1. This cooling system 1 employs a precisely temperature-controlled refrigeration device, capable of gradually or directly cooling the hydrolysate from its initial hydrolysis temperature (e.g., 25℃-30℃) to 4℃-15℃, preferably 8℃. Low temperature effectively inhibits enzyme thermal denaturation, oxidative degradation, and autohydrolysis, and promotes appropriate aggregation of enzyme molecules, creating favorable conditions for subsequent membrane separation. The cooled hydrolysate is then stably pumped into the membrane separation unit via transfer pump 2. This transfer pump 2 can be a peristaltic pump or a centrifugal pump, ensuring a stable flow rate and minimizing shear force on the enzyme.

[0032] Membrane separation unit:

[0033] The membrane separation unit is the core component of this device, containing at least one ultrafiltration membrane module 3. The selected ultrafiltration membrane has a molecular weight cutoff (MWCO) of 30 kDa to 100 kDa, specifically optimized based on the molecular weight and aggregation characteristics of the target enzyme. At a low temperature of 4℃-15℃, enzyme molecules in the oat hydrolysate will moderately aggregate due to weak interactions such as hydrogen bonding and hydrophobic interactions, forming particles with larger effective molecular weights, thus making them easier to retain by the ultrafiltration membrane. Meanwhile, the enzymatic hydrolysis products (such as small peptides and oligosaccharides), due to their small molecular weight, can freely permeate the membrane and are discharged as permeate. This physical filtration method is gentle and avoids the damage to the enzyme structure caused by traditional chemical precipitation or high-speed centrifugation.

[0034] Enzyme enrichment unit 4:

[0035] The concentrated enzyme solution retained by the membrane separation unit enters the enzyme enrichment unit 4. This unit can directly reflux the concentrated enzyme solution back into the oat enzymatic hydrolysis reaction system through a specific circulation pipeline, realizing real-time enzyme enrichment and reuse, significantly improving enzyme utilization and reducing enzyme consumption costs. If further enzyme processing or storage is required, the retained enzyme solution can also be guided to the enzyme reconstitution component 5.

[0036] Enzyme reconstitution component 5:

[0037] The enzyme reconstitution assembly 5 includes a container holding a suitable buffer solution (e.g., phosphate buffer with a pH of 6.0-7.0) and a mixing device 7 (e.g., a magnetic stirrer or a gentle mechanical stirrer). Enzymes obtained from the membrane separation unit or enzyme enrichment unit 4, which have been concentrated or precipitated at low temperatures, are transferred to this container. Through thorough mixing with the buffer solution, the enzymes are reconstituted from their aggregated or precipitated state, restoring their activity. This assembly can be equipped with online or offline detection devices for real-time monitoring of the enzyme activity recovery rate, ensuring the quality of the recovered enzymes.

[0038] Active protection component 6:

[0039] After the enzyme solution is separated from the membrane separation unit, a stabilizer, such as 5% to 10% glycerol or 1 to 2 mM EDTA, can be added to the enzyme solution through the activity protection component 6. Glycerol, as a polyol, can stabilize the three-dimensional structure of the enzyme protein; EDTA, as a chelating agent, can chelate metal ions, reduce metal ion-catalyzed oxidation reactions, further protect the enzyme activity, and reduce its degradation during recovery and storage.

[0040] Recycling monitoring component 11:

[0041] This device is also equipped with a recycling monitoring component 11, which is used to periodically or in real-time monitor the activity of enzymes that have been reconstituted and reused in the enzymatic hydrolysis reaction. By recording the number of times the enzyme is reused and the degree of activity decay, data support can be provided for adjusting process parameters, replenishing or replacing enzymes, and ensuring the stability of enzymatic hydrolysis efficiency.

[0042] pH adjustment component 10:

[0043] The pH adjustment component 10 can adjust the pH value of the oat hydrolysate to near the isoelectric point (pI) of the target enzyme by automatically or manually adding acid or alkali reagents before low-temperature treatment. This helps to enhance the aggregation and precipitation of the enzyme, thereby improving the membrane separation efficiency.

[0044] Reserved interface:

[0045] The device also has reserved interfaces to facilitate subsequent technology upgrades or joint use with other separation technologies (such as higher precision ultrafiltration, magnetic separation components, or affinity tag-related specific adsorption separation components for specific enzymes) to further improve enzyme recovery rate and purity and meet different production needs.

[0046] Through the above structural and functional design, this utility model achieves efficient, mild, and continuous recovery and regeneration of enzymes in oat hydrolysate, greatly reducing production costs and avoiding the negative impact of traditional enzyme inactivation methods on product flavor and quality, thus having significant industrial application value.

[0047] It will be apparent to those skilled in the art that this invention is not limited to the details of the exemplary embodiments described above, and that it can be implemented in other specific forms without departing from the spirit or essential characteristics of this invention. Therefore, the embodiments should be considered illustrative and non-limiting in all respects, and the scope of this invention is defined by the appended claims rather than the foregoing description. Thus, it is intended that all variations falling within the meaning and scope of equivalents of the claims be included within this invention. No reference numerals in the claims should be construed as limiting the scope of the claims.

Claims

1. A pretreatment device for regenerating and recovering enzymes from oat hydrolysate, characterized in that, include: A cooling system is used to cool the oat hydrolysate to a suitable temperature. A delivery pump is used to pump the cooled oat hydrolysate into the membrane separation unit; A membrane separation unit, connected to the delivery pump, includes at least one ultrafiltration membrane module, the ultrafiltration membrane module having a molecular weight cutoff of 30kDa to 100kDa, used to retain enzymes at low temperatures and discharge the permeate; An enzyme enrichment unit, connected to the membrane separation unit, is used to reflux the enzyme retained by the membrane separation unit back to the reaction system or send it to the enzyme reconstitution assembly; Enzyme reconstitution unit, used to reconstitute recovered enzymes.

2. The pretreatment apparatus according to claim 1, characterized in that: The cooling system lowers the temperature of the oat hydrolysate to 4°C to 15°C.

3. The pretreatment apparatus according to claim 1, characterized in that, The enzyme reconstitution assembly includes a container holding a suitable buffer solution and a mixing device.

4. The pretreatment apparatus according to claim 1, characterized in that, The device also includes an activity protection component for adding a stabilizer to the separated enzyme solution after membrane separation.

5. The pretreatment apparatus according to claim 1, characterized in that, The device also includes a recycling monitoring component for monitoring the activity of enzymes that have been reconstituted and re-introduced into the enzymatic hydrolysis reaction.

6. The pretreatment apparatus according to claim 1, characterized in that, The device also includes a pH adjustment component for adjusting the pH of the enzymatic hydrolysate to near the isoelectric point of the enzyme before low-temperature treatment.

7. The pretreatment apparatus according to claim 1, characterized in that, The device has a reserved interface for use in conjunction with ultrafiltration, magnetic separation or affinity adsorption separation components.

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