Device for automatically controlling enzyme reaction and in-situ cleaning

Abstract

The utility model discloses a device for automatically controlling enzyme reaction and in-situ cleaning, which relates to the technical field of biochemical engineering and comprises a central controller, a proportional mixing valve, a temperature control system, a product separation device, an in-situ cleaning system and a sample storage container. And a full-automatic process of enzyme reaction and in-situ cleaning is realized. And after the reaction is completed, the central controller triggers the heating assembly to heat the reaction liquid to 95 DEG C within 5 seconds, so that the enzyme is irreversibly inactivated. After a product is transferred into a separation device, centrifuging for 5 minutes by a miniature centrifuge, carrying out pressure filtration on supernate through a detachable filter element, collecting filtrate into a product bottle, circularly washing an in-situ cleaning system for 10 minutes by using an acidic cleaning solution, carrying out pulse washing for 5 minutes by using an alkaline cleaning solution in sequence, finally rinsing by using deionized water until the pH is neutral, and monitoring the residual quantity to be less than or equal to 0.1 mL in real time by using a pressure sensor. And no cross contamination is ensured. The whole process does not need manual intervention, the enzyme reaction success rate is improved, and the single experiment time is shortened.

Description

Technical Field

[0001] This utility model relates to the field of biochemical engineering technology, and in particular to a device for automatically controlling enzyme reactions and in-situ cleaning. Background Technology

[0002] In the field of biocatalytic processing, traditional enzyme analyzers suffer from problems such as complex operation, large mixing errors, low product separation efficiency, limited reaction termination methods, and difficulty in equipment cleaning. For example, manual mixing can easily lead to deviations in the enzyme-substrate mixing ratio, affecting experimental results; manual centrifugation and filtration are required after the reaction, which is cumbersome and prone to contamination; and residual waste liquid inside the equipment is difficult to completely remove, posing a high risk of cross-contamination.

[0003] This invention addresses these pain points by proposing an integrated device that, through a central controller coordinating a proportional mixing valve, a temperature control system, a centrifugal filtration module, and an in-situ cleaning system, achieves precise control of enzyme reaction parameters, efficient product separation, and full-process automation, thus solving the technical problems of low efficiency and high pollution risk of traditional equipment. Utility Model Content

[0004] To address the shortcomings of existing technologies, this invention provides an automatic control device for enzyme reactions and in-situ cleaning, solving the problems of low efficiency and high risk of contamination associated with traditional equipment.

[0005] To achieve the above objectives, this utility model provides the following technical solution:

[0006] An apparatus for automatically controlling enzyme reactions and in-situ washing, comprising:

[0007] Central controller: Used to coordinate the work of each module, preset enzyme base ratio, reaction time, centrifugation parameters and cleaning procedures;

[0008] Proportional mixing valve: By adjusting the motor speed and opening and closing time, the flow ratio of enzyme solution to substrate can be precisely controlled (accuracy ±1%).

[0009] Temperature control system: Includes reaction tank heating components (heating rate ≥10℃ / s), which can quickly heat to the set temperature (50-100℃) to terminate the reaction;

[0010] Product separation device: integrates a micro high-speed centrifuge (5000-20000rpm) and a pressure-driven filter module (pore size 0.2-5μm);

[0011] In-situ cleaning system: Equipped with multiple cleaning solution storage tanks (acidic / alkaline / enzymatic cleaning solutions), and uses a circulating pump (flow rate 0.5-2 mL / min) to flush pipelines and reaction tanks;

[0012] Sample storage container: directly connected to the proportioning mixing valve.

[0013] Preferably, the proportional mixing valve adopts a dual-path independent control structure, which supports dynamic adjustment of the mixing ratio of enzyme solution and substrate (range 1:100 to 100:1).

[0014] Preferably, the heating component of the temperature control system is a semiconductor thermoelectric module with a response time of ≤5 seconds and a temperature control accuracy of ±0.5℃.

[0015] Preferably, the centrifugal filtration module of the product separation device adopts a detachable filter element design, which supports quick replacement of filter membranes with different pore sizes.

[0016] Preferred: The in-situ cleaning system includes a waste liquid discharge pipeline and is equipped with a pressure sensor to monitor the cleaning fluid flow rate in real time to ensure no residue.

[0017] Preferred configuration: The central controller integrates a human-machine interface, supporting real-time parameter display, historical data query, and fault alarm functions.

[0018] Compared with the prior art, the present invention has the following beneficial effects:

[0019] This invention achieves a fully automated process for enzyme reaction and in-situ cleaning through a central controller that coordinates the control of various modules. First, the user inputs the enzyme-to-substrate ratio (e.g., 1:5), reaction temperature (50℃), and time (30 minutes) via a human-machine interface. The central controller drives the proportional mixing valve to dynamically adjust the motor speed (1000-5000 rpm), precisely controlling the flow ratio of enzyme solution (0.1-10 mL / min) to substrate (0.1-10 mL / min) (accuracy ±1%). After the mixture enters the reaction tank, a semiconductor thermoelectric module (TEC1-12706) maintains a constant temperature (temperature control accuracy ±0.3℃). After the reaction is complete, the central controller triggers the heating component (heating rate 15℃ / s) to heat the reaction solution to 95℃ within 5 seconds, causing irreversible enzyme inactivation (inactivation rate 99.75%). After the product is transferred to the separation device, it is centrifuged for 5 minutes using a microcentrifuge (5000-20000 rpm). The supernatant is then pressure filtered (0.3 MPa) through a removable filter cartridge (0.22 μm polyethersulfone), and the filtrate is collected in the product bottle. Subsequently, the in-situ cleaning system is sequentially rinsed with acidic cleaning solution (0.1 M HCl) circulating (1 mL / min) for 10 minutes, followed by alkaline cleaning solution (0.1 M NaOH + 0.1% SDS) pulsed rinsing (1.5 mL / min) for 5 minutes. Finally, it is rinsed with deionized water until the pH is neutral. The pressure sensor monitors the residual amount in real time to ensure it is ≤0.1 mL, preventing cross-contamination. The entire process requires no manual intervention, improving the success rate of enzyme reactions and shortening the time required for a single experiment. Attached Figure Description

[0020] The above description is only an overview of the technical solution of this utility model. In order to better understand the technical means of this utility model and to implement it in accordance with the contents of the specification, the preferred embodiments of this utility model are described in detail below with reference to the accompanying drawings.

[0021] Figure 1 This is a structural diagram of the present invention. Detailed Implementation

[0022] This application provides an automatic control device for enzyme reactions and in-situ cleaning, which effectively solves the problems of low efficiency and high risk of contamination of traditional equipment.

[0023] Example

[0024] like Figure 1 As shown, the overall technical solution in this application embodiment is as follows:

[0025] To address the problems existing in the prior art, this utility model provides an apparatus for automatically controlling enzyme reactions and in-situ washing, comprising:

[0026] Central controller: presets enzyme base ratio, reaction time, centrifugation parameters, and cleaning procedures;

[0027] Proportional mixing valve: controls the flow ratio of enzyme solution to substrate;

[0028] Temperature control system: includes the heating components for the reaction tank;

[0029] Product separation device: integrates a miniature high-speed centrifuge and a pressure-driven filtration module;

[0030] In-situ cleaning system: Equipped with multiple cleaning fluid storage tanks, and uses a circulating pump to flush pipelines and reaction tanks;

[0031] Sample storage container: directly connected to the proportioning mixing valve.

[0032] The core functions of the central controller include:

[0033] Receive sensor data (temperature, pressure, flow rate);

[0034] Control the motor speed of the proportional mixing valve (1000-5000 rpm);

[0035] Adjust the heating power of the temperature control system (0-200 W);

[0036] Trigger centrifuge start / stop (5000-20000 rpm).

[0037] Human-computer interaction:

[0038] A 7-inch touchscreen displays real-time parameters (enzyme base ratio, temperature profile, centrifugation time).

[0039] Historical data storage (supports exporting to Excel);

[0040] Fault alarm (abnormal pressure, excessive temperature).

[0041] Structural design of proportional mixing valve:

[0042] Dual-channel independent solenoid control valve (material: 316L stainless steel);

[0043] Flow rate range: 0.1-10 mL / min (single stream);

[0044] Dynamic adjustment of mixing ratio: 1:100 to 100:1 (accuracy ±1%).

[0045] The Python working principle of a proportional mixing valve is as follows:

[0046] defcalculate_flow_rate(ratio,total_flow):

[0047] enzyme_flow=total_flow*ratio / (ratio+1)

[0048] substrate_flow=total_flow-enzyme_flow

[0049] returnenzyme_flow,substrate_flow

[0050] In the heating components of the temperature control system:

[0051] Semiconductor thermoelectric module (TEC1-12706);

[0052] Reaction tank volume: 50 mL (material: polytetrafluoroethylene);

[0053] Temperature control range: 25-100℃ (accuracy ±0.3℃);

[0054] Heating rate: 15℃ / s (25℃→95℃ takes 4.7 seconds).

[0055] The temperature control logic code is as follows:

[0056] graph TD

[0057] A [Preset Temperature] --> B {PID Algorithm Adjustment}

[0058] B --> C [Heating / Cooling]

[0059] C --> D [Temperature sensor feedback]

[0060] D --> B

[0061] In the centrifuge module of the product separation device:

[0062] Mini centrifuge (Model: MiniSpinPlus);

[0063] Maximum speed: 20,000 rpm (centrifugal force 21,130 × g);

[0064] Centrifuge chamber capacity: 15 mL.

[0065] Filtering module:

[0066] Removable filter element (material: polyethersulfone);

[0067] Pore ​​size options: 0.22μm (protein separation), 5μm (cell debris filtration);

[0068] Pressure-driven system: air pressure 0.1-0.5MPa.

[0069] In-situ cleaning system cleaning fluid preparation:

[0070] Acidic cleaning solution (0.1M HCl);

[0071] Alkaline cleaning solution (0.1M NaOH + 0.1% SDS);

[0072] Enzymatic hydrolysis washing solution (1 mg / mL proteinase K).

[0073] Cleaning process:

[0074] Central controller ->> Cleaning pump: Start acidic cleaning solution;

[0075] Cleaning pump -> reaction tank: Circulate and rinse for 10 minutes;

[0076] Central controller -> Waste liquid valve: Discharges waste liquid;

[0077] Central controller -> Cleaning pump: Switch to alkaline cleaning solution;

[0078] Cleaning pump -> pipeline: pulse flush for 5 minutes.

[0079] Workflow and control logic:

[0080] Enzyme reaction stage:

[0081] Parameter input:

[0082] The central controller is set to input the enzyme-base ratio (e.g., 1:5), reaction temperature (50℃), and time (30 minutes).

[0083] Mixing process:

[0084] The proportioning valve delivers enzyme solution (1 mL / min) and substrate (5 mL / min) at a total flow rate of 6 mL / min.

[0085] The mixture enters the reaction tank, and the stirring motor maintains uniformity at 200 rpm.

[0086] Reaction terminated:

[0087] Once the time is up, the temperature control system starts heating to 95°C and maintains it for 2 minutes to irreversibly inactivate the enzyme.

[0088] Product separation stage:

[0089] Centrifugation clarification:

[0090] The reaction solution was transferred to a centrifuge and centrifuged at 10,000 rpm for 5 minutes.

[0091] Pressure filtration:

[0092] The supernatant was passed through a 0.22 μm filter membrane at a pressure of 0.3 MPa, and the filtrate was collected into a product bottle.

[0093] In-situ cleaning stage:

[0094] Acidic cleaning:

[0095] The reaction tank was flushed with 0.1M HCl at a rate of 1 mL / min for 10 minutes.

[0096] Enzymatic washing:

[0097] Rinse with proteinase K solution using a pulsed flow rate (1.5 mL / min, 30-second intervals) for 5 minutes.

[0098] Rinse with pure water:

[0099] Rinse with deionized water until pH neutral, and monitor the residual amount with a pressure sensor to ≤0.1mL.

[0100] Proportional mixing accuracy test:

[0101]

[0102] Reaction termination efficiency test:

[0103]

[0104] Cleaning residue test:

[0105]

[0106] Comparative Example 1: Traditional manual operation:

[0107] Mixing error: ±8-15% (causing product yield fluctuation of 20%);

[0108] Reaction termination time: 3-5 minutes (residual enzyme activity 5-8 U / mL);

[0109] Cleaning time: Manual disassembly and cleaning takes 20 minutes per session.

[0110] Comparative Example 2: Commercial ELISA reader:

[0111] Temperature control accuracy: ±2℃ (±0.3℃ for this device);

[0112] Centrifugation function: None (this device integrates centrifugal filtration);

[0113] Cross-contamination rate: 5-8% (<0.1% for this device).

[0114] Working principle:

[0115] This invention achieves a fully automated process for enzyme reaction and in-situ cleaning through a central controller that coordinates the control of various modules. First, the user inputs the enzyme-to-substrate ratio (e.g., 1:5), reaction temperature (50℃), and time (30 minutes) via a human-machine interface. The central controller drives the proportional mixing valve to dynamically adjust the motor speed (1000-5000 rpm), precisely controlling the flow ratio of enzyme solution (0.1-10 mL / min) to substrate (0.1-10 mL / min) (accuracy ±1%). After the mixture enters the reaction tank, a semiconductor thermoelectric module (TEC1-12706) maintains a constant temperature (temperature control accuracy ±0.3℃). After the reaction is complete, the central controller triggers the heating component (heating rate 15℃ / s) to heat the reaction solution to 95℃ within 5 seconds, causing irreversible enzyme inactivation (inactivation rate 99.75%). After the product is transferred to the separation device, it is centrifuged for 5 minutes using a microcentrifuge (5000-20000 rpm). The supernatant is then pressure filtered (0.3 MPa) through a removable filter cartridge (0.22 μm polyethersulfone), and the filtrate is collected in the product bottle. Subsequently, the in-situ cleaning system is sequentially rinsed with acidic cleaning solution (0.1 M HCl) circulating (1 mL / min) for 10 minutes, followed by alkaline cleaning solution (0.1 M NaOH + 0.1% SDS) pulsed rinsing (1.5 mL / min) for 5 minutes. Finally, it is rinsed with deionized water until the pH is neutral. The pressure sensor monitors the residual amount in real time to ensure it is ≤0.1 mL, preventing cross-contamination. The entire process requires no manual intervention, improving the success rate of enzyme reactions and shortening the time required for a single experiment.

[0116] Finally, it should be noted that the above embodiments are merely examples for clearly illustrating the present invention and are not intended to limit the implementation. Those skilled in the art can make other variations or modifications based on the above description. It is neither necessary nor possible to exhaustively list all possible implementations. However, obvious variations or modifications derived therefrom are still within the protection scope of this invention.

Claims

1. An apparatus for automatically controlling an enzyme reaction and in-situ cleaning, characterized by, Comprise: Central controller: preset enzyme substrate ratio, reaction time, centrifugal parameter and cleaning program; Proportioning valve: control the flow ratio of enzyme liquid and substrate; Temperature control system: including reaction tank heating assembly; Product separation device: integrated micro high-speed centrifuge and pressure-driven filtration module; In-situ cleaning system: equipped with multiple cleaning liquid storage tanks, through circulating pump to flush pipeline and reaction tank; Sample storage container: directly connected to the proportioning valve.

2. The apparatus for automatically controlling an enzyme reaction and in-situ cleaning according to claim 1, wherein: The proportioning valve adopts a double-path independent control structure, supporting dynamic adjustment of the mixing ratio of enzyme liquid and substrate.

3. The apparatus for automatically controlling an enzyme reaction and in-situ cleaning according to claim 1, wherein: The heating assembly of the temperature control system is a semiconductor thermoelectric module, with a response time ≤5 seconds and a temperature control accuracy of ±0.5℃.

4. The apparatus for automatically controlling an enzyme reaction and in-situ cleaning according to claim 1, wherein: The centrifugal filtration module of the product separation device adopts a detachable filter core design.

5. The apparatus for automatically controlling an enzyme reaction and in-situ cleaning according to claim 1, wherein: The in-situ cleaning system includes a waste liquid discharge pipeline, equipped with a pressure sensor to monitor the cleaning liquid flow in real time.

6. The apparatus for automatically controlling an enzyme reaction and in-situ cleaning according to claim 1, wherein: The central controller integrates a human-computer interaction interface, supporting real-time parameter display, historical data query and fault alarm functions.

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