Degreasing technology for tooth and bone crusher

Abstract

The invention relates to the technical field of front-end treatment detection of bone DNA (deoxyribonucleic acid) detection, in particular to a degreasing technology for a tooth and bone crusher, which comprises the following steps of: a) placing teeth or bones to be treated in a microwave, ultrasonic and lipase synergistic device; b) after setting and starting the number of circular treatment times through a control circuit, executing the following treatment units according to a preset cycle: adding lipase into the to-be-treated object or enabling the to-be-treated object to be in contact with a lipase solution; c) simultaneously or successively applying microwave irradiation in each treatment unit to generate local heating and non-thermal effect and applying ultrasound to generate cavitation effect for enhancing fat release and improving the contact efficiency of lipase and a substrate; according to the method, the degreasing rate and efficiency are remarkably improved under the mild catalysis condition, release of fat from a bone / tooth matrix can be accelerated, the contact probability of enzyme and a substrate can be improved, and therefore the pretreatment time is shortened, and the chemical damage risk is reduced.

Description

Technical Field

[0001] This invention relates to the field of front-end processing and detection technology for bone DNA detection, specifically a degreasing technology for a tooth and bone crusher. Background Technology

[0002] In the existing front-end processing flow for DNA testing of bones / teeth, the sample needs to be crushed to a certain particle size before subsequent extraction and testing can be carried out. However, bone tissue and teeth often contain a certain amount of fat. Fat not only affects the grinding efficiency during the grinding process, but may also interfere with the subsequent DNA extraction and detection results. Therefore, bone and tooth samples need to be degreased before grinding. Traditional degreasing methods (such as strong acid / alkali treatment or high-temperature overall heating) can easily damage proteins and nucleic acids in samples, or require a long time and complex equipment, making it difficult to meet the requirements for DNA integrity and detection sensitivity. Therefore, a degreasing technology for tooth and bone crushers is proposed to address the above problems. Summary of the Invention

[0003] The purpose of this invention is to provide a degreasing technology for tooth and bone crushers, in order to solve the problems that traditional degreasing methods (such as strong acid / strong alkali treatment or high-temperature overall heating) easily damage the proteins and nucleic acids in the sample, or require a long time and complex equipment, making it difficult to meet the requirements for DNA integrity and detection sensitivity.

[0004] To achieve the above objectives, the present invention provides the following technical solution: A degreasing technique for a tooth and bone shredder includes the following steps: a) Place the teeth or bones to be treated in a microwave + ultrasound + lipase synergistic device; b) After setting and starting the number of cyclic processing times through the control circuit, the following processing unit is executed according to the preset cycle: adding lipase to the substance to be processed or bringing it into contact with lipase solution; c) Simultaneous or sequential application of microwave irradiation to produce local heating and non-thermal effects and application of ultrasound to produce cavitation effects in each processing unit, to enhance fat release and improve the contact efficiency between lipase and substrate. d) After a predetermined number of cycles, the processed material is removed and further pulverized for DNA testing.

[0005] As a further optimization of the present invention, the lipase is pancreatic lipase or a lipase of microbial origin.

[0006] As a further optimization of the present invention, the frequency of the ultrasound is greater than 20 kHz.

[0007] As a further optimization of the present invention, the frequency range of the microwave is 300 MHz to 300 GHz.

[0008] As a further optimization of the present invention, the control circuit includes a microcontroller (MCU), a loop processing count display, and touch buttons, used to set the loop count and start / stop the synergistic effect of microwave and ultrasound.

[0009] As a further optimization of the present invention, the number of cyclic processing cycles is 7 to 9.

[0010] As a further optimization of the present invention, the treatment system is controlled within the appropriate catalytic conditions for lipase under the action of microwave and lipase, with a temperature of 30-50°C and a pH of 6-8, in order to enhance enzyme catalytic activity and avoid damage to proteins and nucleic acids in bone tissue.

[0011] As a further optimization of the present invention, the method is carried out at room temperature and does not require a traditional overall high-temperature heating device. The microwave is only used for local and short-term mild heating and non-thermal electromagnetic field effects.

[0012] As a further optimization of the present invention, the bone powder / tooth powder processed by microwave + ultrasound + lipase does not affect the integrity of DNA and the detection results during subsequent DNA extraction and detection, and is suitable for the preprocessing of bone DNA detection.

[0013] As a further optimization of the present invention, it includes: a treatment cavity for placing the tooth or bone to be treated; Microwave generator and coupling unit; Ultrasonic transmitter; Setup for adding or maintaining lipase / lipase solution; The control circuit includes a microcontroller (MCU), a cycle count display, and touch buttons, and is configured to control microwave power, ultrasonic operation, and record / display the cycle count according to the process of the method described in claim 1, for implementing the degreasing technology.

[0014] Compared with the prior art, the beneficial effects of the present invention are: This invention significantly improves the defatting rate and efficiency under mild catalytic conditions, accelerating the release of fat from the bone / dental matrix and increasing the probability of enzyme-substrate contact. This shortens pretreatment time and reduces the risk of chemical damage. Simultaneously, the physical effects of localized short-term microwave heating and ultrasonic cavitation enhance mass transfer and mechanical stripping, reducing reliance on overall high-temperature treatment of the sample. This helps maintain the integrity and detectability of DNA in bone / teeth. Furthermore, it is compatible with the front-end processing flow (batch / cycle processing) of ordinary tooth / bone pulverizers, allowing for seamless integration with subsequent pulverization and extraction steps, thereby improving detection throughput and repeatability. Attached Figure Description

[0015] Figure 1 This is a flowchart illustrating the degreasing technology used in a tooth and bone pulverizer according to the present invention. Figure 2 This is a block diagram of the microwave + ultrasound + lipase synergistic device of the present invention; Figure 3 This is a flowchart of the microwave + ultrasound + lipase synergistic processing of the present invention. Detailed Implementation

[0016] Please see Figures 1-3 The present invention provides a technical solution: A degreasing technique for a tooth and bone shredder includes the following steps: a) Place the teeth or bones to be treated in a microwave + ultrasound + lipase synergistic device; b) After setting and starting the number of cyclic processing times through the control circuit, the following processing unit is executed according to the preset cycle: adding lipase to the substance to be processed or bringing it into contact with lipase solution; c) Simultaneous or sequential application of microwave irradiation to produce local heating and non-thermal effects and application of ultrasound to produce cavitation effects in each processing unit, to enhance fat release and improve the contact efficiency between lipase and substrate. d) After a predetermined number of cycles, the processed material is removed and further pulverized for DNA detection. Through the synergy of physical fields (microwave, ultrasound) and biocatalysis (lipase), the degreasing efficiency can be significantly improved under low-damage conditions, the pretreatment time can be shortened and the integrity of DNA in bone / teeth can be maintained, making it suitable for high-throughput pretreatment before detection.

[0017] As a further technical solution for implementing this scheme, the lipase is pancreatic lipase or microbial lipase. Using pancreatic lipase or industrially produced microbial lipase can ensure catalytic specificity and stability, improve the efficiency of triglyceride hydrolysis, and is easy to obtain and control in terms of quality. As a further implementation of this scheme, the frequency of ultrasound is greater than 20 kHz. High-frequency ultrasound (>20 kHz) produces significant cavitation and microfluidic field effects, which can enhance mechanical stripping and mass transfer rates, thereby improving the efficiency of enzyme-fat contact and accelerating the degreasing process. As a further implementation of this solution, the microwave frequency range is 300 MHz to 300 GHz. Within this frequency range, short-term localized mild heating and non-thermal electromagnetic effects can be achieved, which can enhance enzyme activity while avoiding the damage of nucleic acids and proteins to overall high temperature, thereby improving safety and controllability. As a further implementation of this solution, the control circuit includes a microcontroller (MCU), a cycle processing count display, and touch buttons, which are used to set the cycle count and start / stop the synergistic effect of microwave and ultrasound. The integrated control circuit facilitates accurate and repeatable setting of processing parameters and cycle count, improves process stability, user-friendliness, and traceability, and is beneficial for batch applications. As a further implementation of this solution, the number of cycles is 7 to 9. The preferred range of cycles can serve as a convenient process guide value to achieve a balance between degreasing effect and processing time, while also facilitating operators to quickly set parameters. As a further technical solution for implementing this scheme, under the action of microwave and lipase, the processing system is controlled within the suitable catalytic conditions of lipase, with a temperature of 30-50°C and a pH of 6-8. This is to enhance enzyme catalytic activity and avoid damage to proteins and nucleic acids in bone tissue. The well-defined temperature and pH range can balance enzyme activity and the biomolecular stability of the sample, thereby improving catalytic efficiency, reducing the risk of DNA degradation, and increasing the detection success rate. As a further implementation of this solution, this method is carried out at room temperature and does not require a traditional overall high-temperature heating device. Microwaves are only used for local and short-term mild heating and non-thermal electromagnetic field effects, avoiding overall high-temperature processing, reducing equipment energy consumption and the risk of thermal damage to the sample, while simplifying the equipment structure and improving the adaptability to sensitive samples. As a further implementation of this solution, the bone powder / tooth powder processed by microwave + ultrasound + lipase does not affect the integrity of DNA and the detection results during subsequent DNA extraction and detection. It is suitable for the front-end preprocessing of bone DNA detection, clearly indicating that it has no adverse effect on downstream DNA detection. It provides a reliable preprocessing method for judicial identification, forensic or scientific research applications, reduces the sample redo rate and improves the reliability of detection. As a further implementation of this solution, the technical solution includes: a treatment cavity for placing the tooth or bone to be treated; Microwave generator and coupling unit; Ultrasonic transmitter; Setup for adding or maintaining lipase / lipase solution; The control circuit includes a microcontroller (MCU), a cycle count display, and touch buttons. It is configured to control microwave power and ultrasonic operation according to the process of claim 1, and record / display the cycle count to achieve degreasing technology. The integrated device design realizes an automated and controllable pretreatment process, improves operational safety and repeatability, and facilitates rapid deployment in the laboratory or on-site and linkage with a grinder.

[0018] Example 1: Standard Batch Degreasing Process (Preferred Example) Pretreatment: Weigh the tooth or bone samples to be degreased (to remove large soft tissue residues) and place them in a microwave-safe container. Add an appropriate amount of buffer (e.g., phosphate buffer at pH 7.0) to maintain enzyme activity.

[0019] Add lipase: Add lipase solution (preferably pancreatic lipase or microbial lipase) to the system to make the final enzyme concentration a suitable range (in this example, the enzyme activity or manufacturer's recommended amount is used, and the specific value can be determined in subsequent optimization). Lipase enables specific hydrolysis of triglycerides under mild conditions to produce glycerol and fatty acids. Place the container in the processing chamber of the synergistic device (i.e., the microwave chamber with a coupled configuration of an ultrasonic transmitter), close the chamber door, and set the number of cycles to 7 to 9 on the control panel (preferably 7 to 9, but can be adjusted according to the fat content of the sample). Each cycle consists of a combination of microwave and ultrasound treatment and an intermediate settling / mixing phase: Microwave irradiation is initiated to provide a short-term, localized, and gentle temperature increase (the local temperature is controlled within the range of approximately 30–50 °C to enhance enzyme activity while avoiding thermal damage); Simultaneously or synchronously before / after the microwave treatment, ultrasound emission (frequency greater than 20 kHz) is initiated to generate cavitation and a microfluidic field, mechanically peeling away the encapsulated fat and enhancing mass transfer. The duration of the combined action can be set to a short interval (e.g., 30s to 180s for each combined action, depending on the sample volume and equipment power). After that, microwave and ultrasound are stopped and a short period of stilling / light stirring is performed to promote enzyme catalysis and product diffusion (stilling for 30s to 120s). After repeating the above cycle 7 to 9 times, remove the processed material and rinse it with deionized water or buffer to remove soluble hydrolysis products (glycerol, free fatty acids); then proceed with the standard procedure to grind (e.g., grind to the target particle size using a tooth / bone grinder) and proceed to the DNA extraction step.

[0020] After defatting, the powder was subjected to oil content testing and DNA extraction, and the extraction volume / integrity was compared (electrophoresis or qPCR) to confirm that the method did not damage the DNA and achieved the defatting goal.

[0021] Example 2: Continuous small-batch automated processing (equipment-side optimization) Device composition: A movable or multi-point ultrasonic transmitter is arranged in the microwave cavity. The processing cavity is equipped with a micro-dosing system (for automatic addition of enzyme solution), temperature / power sensors and a single-chip microcomputer (MCU) control panel (with cycle count display and touch buttons) to realize automatic control of microwave power, ultrasonic start / stop, cycle count and time window. Operating procedure: Select “Sample type → Sample amount → Degreasing program” through the MCU preset program. The device automatically adds enzyme solution and executes preset microwave / ultrasonic cycles. After the cycle is completed, it automatically rinses and discharges the processing solution or transfers it to the pulverizing chamber. This embodiment is suitable for detection pretreatment scenarios that require batch processing and high repeatability.

[0022] Example 3: Optimization of enzyme species and system pH / temperature (preferred catalytic conditions) Enzyme selection: Pancreatic lipase or industrially produced microbial lipase are preferred as catalysts because they have high specificity for triglycerides and good activity under mild conditions. Catalytic conditions: The pH of the system was controlled at about 6 to 8, and the temperature was controlled at about 30 to 50°C by short-term local heating with microwave. These conditions can take into account both the requirements of enzyme activity and DNA protection. The ultrasonic frequency was better than 20 kHz to ensure that the cavitation effect was significant.

[0023] The proposed microwave + ultrasound + lipase synergistic degreasing technology utilizes physical fields (microwave and ultrasound) to enhance mass transfer and mechanical stripping, while using a biocatalyst (lipase) to specifically hydrolyze fat. It can achieve efficient degreasing of tooth / bone samples under room temperature and gentle conditions without compromising the sensitivity and integrity of subsequent DNA extraction and detection, and is suitable for the front-end pretreatment requirements of tooth and bone pulverizers.

[0024] This article uses specific examples to illustrate the principles and implementation methods of the present invention. The above examples are only for the purpose of helping to understand the method and core ideas of the present invention. The above descriptions are only preferred embodiments of the present invention. It should be noted that due to the limitations of textual expression, while there are objectively infinite specific structures, those skilled in the art can make several improvements, modifications, or changes without departing from the principles of the present invention, and can also combine the above technical features in an appropriate manner. These improvements, modifications, changes, or combinations, or the direct application of the inventive concept and technical solution to other situations without modification, should all be considered within the scope of protection of the present invention.

Claims

1. A degreasing technology for a tooth and bone pulverizer, characterized in that, Includes the following steps: a) Place the teeth or bones to be treated in a microwave + ultrasound + lipase synergistic device; b) After setting and starting the number of cyclic processing times through the control circuit, the following processing unit is executed according to the preset cycle: adding lipase to the substance to be processed or bringing it into contact with lipase solution; c) Simultaneous or sequential application of microwave irradiation to produce local heating and non-thermal effects and application of ultrasound to produce cavitation effects in each processing unit, to enhance fat release and improve the contact efficiency between lipase and substrate. d) After a predetermined number of cycles, the processed material is removed and further pulverized for DNA testing.

2. The degreasing technology for a tooth and bone pulverizer according to claim 1, characterized in that: The lipase is pancreatic lipase or a lipase of microbial origin.

3. The degreasing technology for a tooth and bone pulverizer according to claim 1, characterized in that: The frequency of the ultrasound is greater than 20 kHz.

4. The degreasing technology for a tooth and bone pulverizer according to claim 1, characterized in that: The frequency range of the microwave is 300 MHz to 300 GHz.

5. The degreasing technology for a tooth and bone pulverizer according to claim 1, characterized in that: The control circuit includes a microcontroller (MCU), a cycle count display, and touch buttons, used to set the cycle count and start / stop the combined action of microwave and ultrasound.

6. The degreasing technology for a tooth and bone pulverizer according to claim 1, characterized in that: The number of cycles is 7 to 9.

7. The degreasing technology for a tooth and bone pulverizer according to claim 1, characterized in that: Under the action of microwaves and lipase, the treatment system is controlled within the suitable catalytic conditions for lipase, with a temperature of 30–50°C and a pH of 6–8, to enhance enzyme catalytic activity and avoid damage to proteins and nucleic acids in bone tissue.

8. The degreasing technology for a tooth and bone pulverizer according to claim 1, characterized in that: This method is performed at room temperature and does not require a traditional overall high-temperature heating device. The microwaves are used only for local and short-term mild heating and non-thermal electromagnetic field effects.

9. The degreasing technology for a tooth and bone pulverizer according to claim 1, characterized in that: The bone / tooth powder processed by microwave, ultrasound, and lipase does not affect the integrity of the DNA or the detection results during subsequent DNA extraction and testing, and is suitable for preprocessing in bone DNA detection.

10. A device for synergistic action of microwave, ultrasound, and lipase, characterized in that, include: A treatment cavity used to hold the teeth or bone to be treated; Microwave generator and coupling unit; Ultrasonic transmitter; Setup for adding or maintaining lipase / lipase solution; The control circuit includes a microcontroller (MCU), a cycle count display, and touch buttons, and is configured to control microwave power, ultrasonic operation, and record / display the cycle count according to the process of the method described in claim 1, for implementing the degreasing technology.

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