Medicine material identification system and method

Abstract

The application belongs to the technical field of molecular detection, and discloses a medicinal material identification system and method; the method comprises the following steps: emitting a topological protection acoustic wave to concentrate DNA of medicinal materials, marking the DNA with a CdTe quantum dot-nucleic acid probe, scraping the sample to extract DNA and performing PCR amplification, designing a logic gate staple chain to prepare a light-controlled origami probe, and injecting a PZT chip nano-pore array to be light-activated, and then, through AFM analysis of free energy, the medicinal materials are identified; the application constructs a complete technical chain from molecular recognition to reliable storage, and comprehensively solves multiple technical bottlenecks of traditional methods in medicinal material identification.

Description

Technical Field

[0001] This invention relates to the field of molecular detection technology, and more specifically, to a system and method for identifying medicinal materials. Background Technology

[0002] Chinese patent application CN106770037A discloses a method for identifying Datura stramonium medicinal material: a two-phase phenotypic pharmacological identification method based on a resonant waveguide grating sensor. (1) Extraction of medicinal material: Accurately weigh 1g of medicinal material powder, place it in an Erlenmeyer flask, add 10mL of 2mol / L hydrochloric acid solution, sonicate at 250W power and 40kHz frequency for 30 minutes, cool, filter, wash the residue and filter with 10mL of the above hydrochloric acid solution several times, combine the filtrate and washings, adjust the pH value to 9 with concentrated ammonia solution, extract with chloroform 4 times by shaking, 10mL each time, combine the chloroform, recover the solvent to dryness, dissolve the residue in buffer solution, transfer it to a 5mL volumetric flask, add buffer solution to the mark. (1) Shake well, filter, and take the filtrate for testing; (2) Use a resonant waveguide grating sensor to detect the pharmacological phenotype of CHO-M2 cells stimulated by the herbal extract. Take 20 μL of CHO-M2 cell suspension and seed it in a cell test plate. Place it in an incubator containing 5% carbon dioxide and a temperature of 37°C and incubate for 14 hours. Take out the cell test plate, remove the culture medium, and add 20 μL of Hank's balanced salt solution to each well with a pipette. Place the test plate in the CorningEpicBT system for signal equilibration. After 1 hour, set the baseline to zero and start a new signal recording program. Record for 2 minutes, pause the program, add 10 μL of extract to each well with a pipette, and continue recording for 1 hour. If the signal detected after adding the herbal extract exceeds 100 pm, the phenotype is recorded as 1; if the signal is between -100 and 100, the phenotype is recorded as 0; if the signal is less than -100, the phenotype is recorded as -1. The dynamic mass redistribution signal generated by the herbal extract at this stage is the first phase phenotype; (3) The integrated pharmacological phenotype generated by the combined stimulation of CHO-M2 cells by acetylcholine and herbal extract is detected by a resonant waveguide grating sensor. After the above recording program is completed, a new program is started. The program is paused after 2 minutes of signal recording. 10 μL of 16 μM acetylcholine Hank's balanced salt solution is added to each well using a pipette. Then the recording program is continued for 1 hour. If the signal exceeds 100 pm after adding the acetylcholine solution, the phenotype is recorded as 1; if the signal is between -100 and 100, the phenotype is recorded as 0; if the signal is less than -100, the phenotype is recorded as -1. The dynamic mass redistribution signal generated in this stage is the second phase phenotype; (4) Integrate the phenotypes of steps 2 and 3 and compare them with the phenotype of Datura stramonium. If (1,0) is satisfied, it is Datura stramonium, where "1" is the phenotype of step 2 and "0" is the phenotype of step 3. Otherwise, it is a counterfeit medicinal material. (5) The combination of the two-phase phenotype pharmacological identification method based on the resonant waveguide grating sensor and HPLC is used to identify the quality of Datura stramonium. Datura stramonium is analyzed by HPLC under the following conditions, and the peak area or content of the components is correlated with the activity value of the medicinal material.The invention ultimately produces phenotypes that are simply similar to (1,0) numbers, which are less dependent on experience, have higher discernibility, and can avoid misjudgment.

[0003] While the above methods can meet the needs of most scenarios, research and practical application of these methods and existing technologies have revealed at least the following shortcomings:

[0004] The above methods can only detect components in medicinal extracts that can act on cell receptors, and cannot distinguish counterfeit products or cases of tissue mismatch.

[0005] In view of this, the present invention proposes a medicinal material identification system and method to solve the above problems. Summary of the Invention

[0006] To overcome the aforementioned deficiencies of the prior art and to achieve the above objectives, the present invention provides the following technical solution: a method for identifying medicinal materials, comprising the following steps:

[0007] Topologically protected acoustic waves are emitted into the medicinal material, and the DNA of the medicinal material is labeled by CdTe quantum dots-nucleic acid probes in the microcavity of the medicinal material to obtain the target medicinal material;

[0008] A sample of the target medicinal material was scraped from the microcavity, and DNA was extracted from the sample using the CTAB method. The DNA target gene was amplified by PCR, and a logic gate staple chain was designed based on the DNA target gene sequence to prepare a light-controlled origami probe solution.

[0009] The optically controlled origami probe solution is injected into the nanopore array of the PZT chip and photoactivated to obtain an activated origami structure. An axial force is applied to the activated origami structure using an AFM probe, and the free energy of the activated origami probe is analyzed. Based on the free energy and a preset database, medicinal materials are identified to obtain identification labels.

[0010] Furthermore, methods for obtaining the target medicinal material include:

[0011] Emit 8-12MHz topological protection acoustic waves for 80-120ms to the medicinal materials, then place the medicinal materials in PBS buffer and shake to clean them. Repeat 2-3 times.

[0012] Add 1%-5% BSA solution to the cleaned medicinal materials and incubate at room temperature for 30-60 minutes;

[0013] Wash the medicinal materials again with PBS buffer 3-5 times to obtain the medicinal materials to be processed;

[0014] In a centrifuge tube, add CdTe quantum dot solution and antibody solution in a molar ratio of 10:1-50:1, mix well, and shake on a shaker at room temperature for 1-2 hours.

[0015] After shaking, place the centrifuge tubes in a centrifuge and centrifuge at 10,000-15,000 rpm for 10-15 minutes.

[0016] The precipitate was resuspended in PBS buffer to obtain the CdTe quantum dot-antibody probe solution;

[0017] Place the medicinal material to be treated into a container containing CdTe quantum dot-antibody probe solution; incubate at room temperature for 1-3 hours with shaking at 100-200 rpm on a shaker. After incubation, wash the medicinal material 3-5 times with PBS buffer to obtain the target medicinal material.

[0018] Furthermore, methods for obtaining the light-controlled origami probe solution include:

[0019] Origami structures were prepared based on medicinal material samples;

[0020] 3-8 nm gold nanoparticles were prepared, Ni-NTA was modified on the surface of the gold nanoparticles, and His-tag was mixed with Ni-NTA in a molar ratio of 3:1-5:1. The mixture was reacted in the dark at 20-30℃ for 10-15 h to form directional coupling.

[0021] The photolytic linker was dissolved in DMSO and mixed with the origami structure at a molar ratio of 1:1. The mixture was reacted at room temperature in the dark for 1-3 hours, and then purified by magnetic beads to obtain the purified structure.

[0022] The biotinylated DNA short strand and the vertex sequence of the purified structure are incubated at 35-38℃ for 20-40 min in a molar ratio of 4:1-6:1 to form the biotinylated exposed end and obtain the labeled structure.

[0023] Streptavidin-modified CdSe@ZnS quantum dots were added to the labeled structure at a molar ratio of 1:1. The reaction was carried out at 4-6℃ in the dark for 1-2 hours. The mixture was then filtered using a 90-120kDa membrane at a flow rate of 1-3mL / min to obtain the light-controlled origami structure.

[0024] Furthermore, methods for obtaining origami structures include:

[0025] Design a 110-130nt circular DNA template containing a latch strand sequence, a logic gate targeting sequence, and a complementary sequence to the M13mp18 phage backbone strand;

[0026] Add 80-150 μL of amplification reaction system and react at 25-35℃ for 15-18 hours to obtain long-chain backbone DNA containing the latching strand;

[0027] Add 80-120 nM Cas12a and 180-220 nM gRNA to the amplification product, incubate at 35-38℃ for 1-2 h, cleave the long backbone containing the locked strand, and then purify it with streptavidin magnetic beads to obtain the backbone strand.

[0028] Design a logic gate staple chain, introduce an azobenzene photoresponsive group at the 5' end of the logic gate staple chain, and modify the 3' end with a His-tag;

[0029] Add a self-assembly system, denature at 90-100℃ for 3-8 minutes, anneal at 55-65℃ for 1-2 hours; slowly cool to 25-30℃ for 48-50 hours; then irradiate with 365nm ultraviolet light for 8-12 seconds to obtain the origami structure.

[0030] Furthermore, the logic gate targeting sequence is complementary to the DNA of the medicinal sample;

[0031] The amplification reaction system consists of 8-12 U Phi29 DNA polymerase, 1-3 mM dNTPs containing 10% dUTP, 0.08-0.12 nM circular template, and 0.08-0.12 nM 5'-phosphorylated primers, in a solution containing 8-12 mM Mg 2 It was prepared in 18-22 mM Tris-HCl buffer at pH 7.8-8.2 to activate Cas12a;

[0032] The logic gate staple chain includes an 18-22nt target segment complementary to the DNA of the medicinal sample, a 13-18nt CRISPR response segment containing a PAM sequence recognized by Cas12a, and a preset anchor point.

[0033] The self-assembled system includes an 8-12 nM backbone chain, an 8-12 μM logic gate stapled chain pool, 18-22 mM Tris-HCl with a pH of 7.8-8.2, 10 mM MgCl2, and 50 mM NaCl.

[0034] Furthermore, methods for obtaining identification tags include:

[0035] An axial force is applied to activate the origami structure using an AFM probe, and the AFM force curve is recorded. The stretching curve corresponds to the molecular unfolding process, and the retraction curve corresponds to the molecular folding process.

[0036] When the molecules in the folded region are not stretched, the conformational length corresponding to the displacement of the molecule is taken as the folding displacement; in the completely unfolded region, the conformational length when the force change is lower than a preset threshold, corresponding to the length of the stretched molecule, is taken as the completely unfolding displacement.

[0037] The free energy of the medicinal material is obtained by integrating the AFM force curve by combining the folded displacement and the completely defolded displacement.

[0038] The system detects whether the free energy of medicinal materials is within the range of the free energy of genuine medicinal materials in the preset database. If it is, the material is identified as genuine and marked with an identification label of 1; otherwise, it is identified as counterfeit and marked with an identification label of 0.

[0039] Furthermore, it also includes:

[0040] A topology protection acoustic wave is emitted to the PZT chip, the acoustic energy spectrum is collected, features are extracted, and an acoustic hash is obtained.

[0041] CdTe quantum dots were excited, fluorescence decay curves were collected, and optical hashes were obtained by feature extraction.

[0042] After applying chaotic encryption to acoustic and optical hashes, the evidence is stored on the blockchain in conjunction with identification tags.

[0043] Methods for obtaining acoustic hashes include:

[0044] The time-domain pressure signal is acquired, discretized, and a discrete signal s[n] is obtained.

[0045] Perform a discrete Fourier transform on s[n] to convert it into a frequency domain amplitude spectrum S[k];

[0046] From S[k], obtain the K frequency points with the largest amplitude, corresponding to the spectral line indices, and calculate the K frequency values; obtain the K frequency points with the second largest amplitude, corresponding to the spectral line indices, and calculate the K frequency values.

[0047] Normalize the 2K frequency values ​​to [0, 2]. K / 2 The range is -1], and 2K K / 2-bit integers are obtained;

[0048] 2K K / 2-bit integers are concatenated into a binary string, and then compressed into a 256-bit hash value using the SHA-256 hash function; this yields the acoustic hash.

[0049] Furthermore, methods for obtaining the optical hash Hl include:

[0050] The time-domain decay signal is acquired and discretized to obtain the discrete signal l[n].

[0051] DWT is performed on l[n] using the Morlet wavelet basis. At F preset scales, the E wavelet coefficients W with the largest amplitudes are selected at each scale, for a total of 2K W. For each selected W, the phase is calculated, and the 2K phases are normalized to [0, 2]. K / 2 The range is -1], and 2K K / 2-bit integers are obtained;

[0052] 2K K / 2-bit integers are concatenated into a binary string, and then compressed into a 256-bit hash value using the SHA-256 hash function; this yields the optical hash.

[0053] Furthermore, methods for chaotic encryption of acoustic hashes and optical hashes include:

[0054] Select the Lorentz parameter, iteratively calculate the Lorentz parameter in M ​​steps, extract the x value of the chaotic trajectory, map it to the [0,1] interval through the sigmoid function, and generate an M-bit chaotic key stream;

[0055] The acoustic hash and optical hash are converted into 256-bit hexadecimal strings to obtain preprocessed data. The identification tag and preprocessed data are XORed byte by byte with the chaotic key stream to obtain encrypted data.

[0056] For encrypted data, the SHA-256 algorithm is used to calculate the acoustically enhanced hash; the SHA-3 algorithm is used to calculate the optically enhanced hash.

[0057] The SHA-256 algorithm is then used to compute and encrypt the acoustic enhanced hash, optical enhanced hash, and chaotic key stream.

[0058] A medicinal herb identification system, implementing the aforementioned medicinal herb identification method, including:

[0059] Enrichment sampling module: Emits topologically protected acoustic waves to the medicinal material, and modifies the medicinal material DNA with CdTe quantum dots-nucleic acid probes in the microcavity of the medicinal material to obtain a medicinal material sample;

[0060] Sample processing module: scrape the medicinal sample from the microcavity of the medicinal material, extract DNA from the medicinal sample by CTAB method, amplify the DNA target gene by PCR, design the logic gate staple chain based on the DNA target gene sequence, and prepare the light-controlled origami probe solution;

[0061] Authenticity identification module: The optically controlled origami probe solution is injected into the nanopore array of the PZT chip and photoactivated to obtain an activated origami structure. An axial force is applied to the activated origami structure through an AFM probe, and the free energy of the activated origami probe is analyzed. Based on the free energy and a preset database, the medicinal materials are identified to obtain an identification label.

[0062] First extraction module: Emits topology protection acoustic waves to the PZT chip, collects the acoustic energy spectrum, performs feature extraction, and obtains acoustic hash;

[0063] The second extraction module: excites CdTe quantum dots, collects fluorescence decay curves, and performs feature extraction to obtain optical hashes;

[0064] Encrypted Evidence Preservation Module: After performing chaotic encryption on acoustic hash and optical hash, it combines the identification tag to perform on-chain evidence preservation.

[0065] The technical effects and advantages of the medicinal herb identification system and method of this invention are as follows:

[0066] This invention utilizes topologically protected acoustic waves to penetrate the epidermis of medicinal materials, forming an acoustic vortex field that directionally enriches deep DNA into surface microcavities. This solves the problems of traditional methods being unable to reach deep genetic material, distinguishing counterfeit products, and confusing tissue locations. By employing CdTe quantum dot-nucleic acid probes to specifically label DNA and combining this with optically controlled origami probes for single-molecule recognition, it overcomes the challenges of insufficient sensitivity in detecting trace samples and interference from impurities in the extract. Furthermore, by analyzing free energy using AFM force spectroscopy and combining it with multi-dimensional signal verification using acoustic-optical dual-modal hashing, it replaces single refractive index detection, avoiding interference from cell state and environmental factors, and improving detection stability and accuracy. Finally, by employing chaotic encryption and blockchain notarization to ensure data immutability, it constructs a complete technology chain from molecular recognition to trusted notarization, comprehensively solving multiple technical bottlenecks in traditional methods for medicinal material identification. Attached Figure Description

[0067] Figure 1 This is a schematic diagram of the medicinal material identification method of the present invention;

[0068] Figure 2 This is a schematic diagram of the method for obtaining the light-controlled origami probe solution according to the present invention;

[0069] Figure 3 This is a schematic diagram of the encrypted on-chain process of the present invention;

[0070] Figure 4 This is a schematic diagram of the medicinal material identification system of the present invention. Detailed Implementation

[0071] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0072] Example 1

[0073] Please see Figure 1 As shown, this embodiment provides a method for identifying medicinal materials, including the following steps:

[0074] Topologically protected acoustic waves are emitted into the medicinal material, and the DNA of the medicinal material is labeled by CdTe quantum dots-nucleic acid probes in the microcavity of the medicinal material to obtain the target medicinal material;

[0075] Methods for obtaining the target medicinal material include:

[0076] Emit 8-12MHz topological protection acoustic waves at 80-120ms to the medicinal materials, which penetrate the epidermis and form an acoustic vortex field inside the medicinal materials, thereby directionally enriching deep DNA into the surface microcavities. Then, place the medicinal materials in PBS buffer and shake to wash them. Repeat 2-3 times.

[0077] Add 1%-5% BSA solution to the cleaned medicinal materials and incubate at room temperature for 30-60 minutes to block non-specific binding sites on the surface of the medicinal materials.

[0078] Wash the herbs again with PBS buffer 3-5 times to remove unbound blocking agent.

[0079] In a centrifuge tube, add CdTe quantum dot solution and antibody solution in a molar ratio of 10:1-50:1, mix well, and shake on a shaker at room temperature for 1-2 hours to allow the quantum dots and antibody to be fully coupled.

[0080] After the reaction is complete, place the centrifuge tubes in a centrifuge and centrifuge at 10,000-15,000 rpm for 10-15 minutes to remove unbound quantum dots and antibodies.

[0081] The precipitate was resuspended in an appropriate amount of PBS buffer to obtain the CdTe quantum dot-antibody probe solution.

[0082] Place the pretreated medicinal materials into a container containing CdTe quantum dot-antibody probe solution, ensuring that the medicinal materials are completely submerged.

[0083] At room temperature, place the container on a shaker and incubate at 100-200 rpm for 1-3 hours to allow the CdTe quantum dot-antibody probe to specifically bind to the target DNA location in the microcavities on the surface of the medicinal material.

[0084] After incubation, the medicinal material was washed 3-5 times with PBS buffer to obtain the medicinal material bound to the CdTe quantum dot-antibody probe.

[0085] By emitting a 10MHz topologically protected acoustic wave at 100ms onto the medicinal material, utilizing its ability to penetrate the epidermis and form an acoustic vortex field within, deep DNA is directionally enriched into surface microcavities. This solves the technical challenge of efficient extraction of deep DNA in traditional methods. Surface impurities are then removed by oscillation washing with PBS buffer. 1%-5% BSA solution is added to block non-specific binding sites on the medicinal material surface, followed by another wash to reduce background interference during subsequent labeling. Finally, CdTe quantum dots are coupled to antibodies at a molar ratio of 10:1-50:1 and purified by centrifugation to obtain the required amount of CdTe. The quantum dot-antibody probe solution is incubated with pretreated medicinal materials at room temperature on an oscillator for 1-3 hours, allowing the probe to specifically bind to the target DNA sites in the microcavities on the surface of the medicinal materials. Finally, the medicinal materials bound to the probe are obtained by washing. The above method overcomes the spatial limitations of sample processing through the directional enrichment of topological acoustic waves. Combined with the specific recognition of quantum dot-antibody probes, it achieves efficient capture and accurate labeling of deep DNA in medicinal materials. It effectively solves the technical bottlenecks of low extraction efficiency and insufficient labeling specificity of trace samples in medicinal material identification, and provides a reliable sample basis for subsequent multimodal signal detection and anti-counterfeiting evidence preservation.

[0086] A sample of the target medicinal material was scraped from the microcavity, and DNA was extracted from the sample using the CTAB method. The DNA target gene was amplified by PCR, and a logic gate staple chain was designed based on the DNA target gene sequence to prepare a light-controlled origami probe.

[0087] Reference Figure 2 Methods for obtaining optically controlled origami probe solutions include:

[0088] Design a 110-130nt circular DNA template containing a latching strand sequence, a logic gate targeting sequence (complementary to the DNA of the medicinal sample), and a complementary sequence to the M13mp18 phage backbone strand;

[0089] Add 80-150 μL of amplification reaction mixture and react at 25-35 °C for 15-18 hours to obtain long-chain backbone DNA containing the latching strand. The amplification reaction mixture consists of 8-12 U Phi29 DNA polymerase, 1-3 mM dNTPs containing 10% dUTP, 0.08-0.12 nM circular template, and 0.08-0.12 nM 5'-phosphorylated primers, in a solution containing 8-12 mM Mg... 2 It was prepared by activating Cas12a in 18-22 mM Tris-HCl buffer at pH 7.8-8.2.

[0090] Add 80-120 nM Cas12a and 180-220 nM gRNA to the amplification product, incubate at 35-38℃ for 1-2 h, cleave the long backbone containing the locked strand, and then purify it with streptavidin magnetic beads to obtain the backbone strand.

[0091] Design a logic gate staple chain, introduce an azobenzene photoresponsive group at the 5' end of the logic gate staple chain, and modify the 3' end with a His-tag;

[0092] The logic gate staple chain contains an 18-22nt target segment complementary to the DNA of the medicinal sample, a 13-18nt CRISPR response segment containing a PAM sequence recognized by Cas12a, and a pre-defined anchor point.

[0093] Add a self-assembly system, denature at 90-100℃ for 3-8 minutes, anneal at 55-65℃ for 1-2 hours; slowly cool to 25-30℃ for 48-50 hours; then irradiate with 365nm ultraviolet light for 8-12 seconds to cut the latch chain, trigger structural transformation, and obtain the origami structure.

[0094] The self-assembled system includes an 8-12 nM backbone chain, an 8-12 μM logic gate stapled chain pool, 18-22 mM Tris-HCl with a pH of 7.8-8.2, 10 mM MgCl2, and 50 mM NaCl;

[0095] 3-8 nm gold nanoparticles were prepared, Ni-NTA was modified on the surface of the gold nanoparticles, and His-tag was mixed with Ni-NTA in a molar ratio of 3:1-5:1. The mixture was reacted in the dark at 20-30℃ for 10-15 h to form directional coupling.

[0096] The photolytic linker (o-nitrobenzyl ester) was dissolved in DMSO (azobenzene photoresponsive group), mixed with the origami structure at a molar ratio of 1:1, reacted at room temperature in the dark for 1-3 hours, and then purified by magnetic beads to obtain the purified structure.

[0097] The biotinylated DNA short strand and the vertex sequence of the purified structure are incubated at 35-38℃ for 20-40 min in a molar ratio of 4:1-6:1 to form the biotinylated exposed end and obtain the labeled structure.

[0098] Streptavidin-modified CdSe@ZnS quantum dots were added to the labeled structure at a molar ratio of 1:1. The reaction was carried out at 4-6℃ in the dark for 1-2 hours. The mixture was then filtered using a 90-120kDa membrane at a flow rate of 1-3mL / min to obtain the light-controlled origami structure.

[0099] By scraping microcavity samples from medicinal materials and extracting DNA using the CTAB method, combined with PCR amplification of target genes, specific DNA sequences were efficiently obtained from trace samples. This solved the problems of low sample DNA extraction efficiency and insufficient specificity in traditional methods, providing precise templates for subsequent molecular recognition. A 120nt circular DNA template containing a latching strand sequence, a logic gate targeting sequence, and a complementary backbone strand sequence was designed. After Phi29 polymerase rolling circle amplification and Cas12a enzyme digestion and purification, a long-chain backbone DNA that can be photo-controlled was prepared. dUTP was used to enhance the enzyme digestion efficiency and ensure precise processing of the backbone strand, solving the problem of preparation accuracy of complex nanostructure assembly precursors. Azobenzene photoresponsive groups and His-tags were introduced into the logic gate stapled strand, combined with the design of a CRISPR response segment containing a PAM sequence, enabling the stapled strand to both respond to UV-triggered structure transitions and be photosensitive. By coupling gold nanoparticles to a self-assembled system, thermal annealing, and photoactivation, a light-controlled origami structure with enhanced mechanical signal is formed, overcoming the bottlenecks of weak signals and poor controllability of traditional probes. Ni-NTA surface modification of the gold nanoparticles and directional coupling with His-tag enhance the stability of the mechanical signal in AFM force spectroscopy detection. Photolytic linkers coupled with the origami structure enable light-controlled release. The specific binding of biotinylated DNA short chains with quantum dots provides optical signal output. Membrane filtration purification ensures probe homogeneity. This method constructs a force-optical dual-modal signal detection system, solving the problem of insufficient reliability in single-signal detection. Ultimately, through multi-step synergy, a complete detection chain is achieved, from specific identification of medicinal material DNA and light-controlled structure conversion to multimodal signal output, providing an innovative solution for identifying genuine and counterfeit medicinal materials that combines high efficiency, specificity, and traceability.

[0100] The photo-controlled origami probe solution is injected into the nanopore array of the PZT chip and photoactivated to obtain an activated origami structure. An axial force is applied to the activated origami structure using an AFM probe, and the free energy of the activated origami probe is analyzed. Based on the free energy and a preset database, medicinal materials are identified to obtain identification labels.

[0101] Methods for obtaining activated origami structures include:

[0102] The PZT chip was immersed in anhydrous ethanol, sonicated for 5 minutes, and dried with nitrogen gas; then immersed in 0.1% PLL solution (polylysine solution), incubated at room temperature for 30 minutes, rinsed 3 times with deionized water, and dried with nitrogen gas.

[0103] A 10 nM optically controlled origami probe solution was dropped into the center of a pre-set PZT chip microcavity array and left to stand at 4 °C for 12 h (the origami is bonded to the PLL by electrostatic interaction and fixed to the microcavity).

[0104] Rinse the chip microcavities with PBS buffer containing 0.01% Tween-20 to remove any unfixed origami structures.

[0105] The photosensitive gRNA and Cas12a were mixed at a molar ratio of 2:1, and 1×NEBuffer 3.1 containing 10mM MgCl2 was added. The mixture was reacted at room temperature in the dark for 30 min. The mixture was then purified by a Superdex 200 gel filter column, and the elution peak was collected. The mixture was then concentrated to 100nM by dialyzing with PBS buffer to obtain the complex solution.

[0106] The composite solution was dropped onto the surface of the PZT chip and incubated at 4°C in the dark for 1 hour.

[0107] Irradiate the surface of the PZT chip with 365nm ultraviolet light for 10s, and immediately rinse the chip three times with 100μL of PBS buffer preheated to 37℃ to obtain the activated origami structure.

[0108] The PZT chip was first ultrasonically treated with anhydrous ethanol and dried with nitrogen to clean the surface, and then incubated with 0.1% PLL solution for modification. The positively charged property of PLL enhanced the electrostatic adsorption capacity for negatively charged DNA origami structures, solving the technical problem of weak bonding between the chip surface and the origami structure, ensuring stable fixation of the origami structure in the microcavity. A 10 nM light-controlled origami probe solution was dropped into the center of the chip microcavity array and incubated at 4°C for 12 hours. By optimizing the fixation temperature and time, the origami and PLL interacted fully while avoiding structural denaturation caused by prolonged high temperatures, achieving precise positioning and stable anchoring of the origami structure in the microcavity. The chip microcavity was rinsed with PBS buffer containing 0.01% Tween-20, using surfactants to remove unfixed origami structures and non-specific adsorbed impurities, effectively reducing background signal interference and improving the specificity of subsequent detections. Photosensitive gRNA and Cas12a were mixed at a 2:1 molar ratio and incubated in NE Buffer containing 10 mM MgCl2. In reaction 3.1, the efficiency of complex formation was improved by optimizing the ratio and buffer system. After purification by Superdex 200 gel filtration column and concentration by PBS dialysis, a highly active Cas12a-gRNA complex was obtained, solving the problems of low activity and many impurities in the complex preparation. The complex solution was dropped onto the chip surface and incubated in the dark to ensure its specific binding to the latching strand on the origami structure. Finally, irradiation with 365nm ultraviolet light for 10 seconds triggered the unwinding of the photosensitive gRNA, activating Cas12a to cleave the latching strand. At the same time, the reaction was immediately terminated and the cleaved fragments were removed by rinsing with PBS buffer preheated to 37°C. By precisely controlling the photoactivation time and temperature, efficient and specific conformational conversion of the origami structure was achieved, solving the technical bottleneck of low efficiency and non-specific cleavage in traditional activation methods. This provides a stable and reliable activated origami structure for subsequent AFM force spectroscopy analysis of free energy and multimodal signal detection, ensuring the accuracy of molecular calculations and signal output in medicinal material identification.

[0109] Methods for obtaining identification tags include:

[0110] An axial force is applied to activate the origami structure using an AFM probe, and the AFM force curve is recorded. The stretching curve corresponds to the molecular unfolding process, and the retraction curve corresponds to the molecular folding process.

[0111] When the folded region molecules are not stretched, the conformational length corresponding to the displacement molecules is obtained as the folding displacement; in the fully unfolded region, the conformational length when the force change is lower than a preset threshold, corresponding to the length of the stretched molecules, is taken as the fully unfolding displacement; the preset threshold can be set based on the experimental noise level, force curve characteristics and physical characteristics of molecular unfolding behavior.

[0112] The free energy of the medicinal material is obtained by integrating the AFM force curve by combining the folded displacement and the completely defolded displacement.

[0113] The system detects whether the free energy of medicinal materials is within the range of the free energy of genuine medicinal materials in the preset database. If it is, the material is identified as genuine and marked with an identification label of 1; otherwise, it is identified as counterfeit and marked with an identification label of 0.

[0114] By applying axial force to the activated origami structure using an AFM probe and recording the force curve, the molecular conformational change process is accurately captured by utilizing the properties of stretching corresponding to molecular unfolding and retraction corresponding to folding in the curve. By determining the fold displacement when the folded state is not stretched and the fully unfolded displacement where the force value change in the fully unfolded region is below a threshold, key parameters are provided for subsequent thermodynamic analysis. The free energy of the medicinal material is then calculated by integrating the AFM force curve using these two displacements and comparing it with the free energy range of genuine medicinal materials in a preset database. This achieves a quantitative conversion from molecular mechanical signals to thermodynamic characteristics, solving the problems of low sensitivity and insufficient specificity caused by relying on macroscopic features or single chemical indicators in traditional medicinal material identification. By capturing the subtle differences in free energy when the DNA of genuine and counterfeit medicinal materials interacts with the origami structure using single-molecule force spectroscopy, and replacing subjective experience judgment with thermodynamic quantitative standards, this method overcomes technical bottlenecks such as micro-sample detection and interference from complex matrices, providing a precise and digital solution at the molecular level for the identification of genuine and counterfeit medicinal materials.

[0115] A topology protection acoustic wave is emitted to the PZT chip, the acoustic energy spectrum is collected, features are extracted, and an acoustic hash is obtained.

[0116] Reference Figure 3 Methods for obtaining the acoustic hash Hs include:

[0117] The time-domain pressure signal s(t) is acquired and discretized to obtain the discrete signal s[n], where t represents time and n represents discrete points;

[0118] Perform a discrete Fourier transform on s[n] to convert it into a frequency domain amplitude spectrum S[k], where k represents a frequency domain point;

[0119] From S[k], obtain the K frequency points with the largest amplitude, corresponding to the spectral line indices, and calculate the K frequency values; obtain the K frequency points with the second largest amplitude, corresponding to the spectral line indices, and calculate the K frequency values.

[0120] Normalize the 2K frequency values ​​to [0, 2]. K / 2 The range is -1], and 2K K / 2-bit integers are obtained;

[0121] 2K K / 2-bit integers are concatenated into a binary string, and then compressed into a 256-bit hash value using the SHA-256 hash function; this yields the acoustic hash.

[0122] By acquiring the time-domain pressure signal of the PZT chip under the action of topological protection acoustic waves and discretizing it, the signal is converted into a frequency domain amplitude spectrum through discrete Fourier transform. The K frequency points with the largest and second largest amplitudes are extracted to capture the main characteristic components of the signal and effectively filter noise interference. The frequency values ​​are then normalized and quantized into K / 2-bit integers, concatenated into a binary string, and compressed using the SHA-256 hash function to generate a 256-bit acoustic hash. The above method solves the technical problems of ambiguous signal features and poor noise resistance in traditional acoustic detection through frequency domain analysis and feature optimization. The unidirectionality and collision resistance of the hash algorithm ensure the integrity and immutability of acoustic feature data, providing a physically unique acoustic fingerprint for medicinal material identification. After the hash value is fused with multimodal features such as optical hash, a high-dimensional medicinal material identity identifier can be constructed, effectively solving the industry pain points of insufficient accuracy of single-modal identification and low data storage credibility. This realizes a closed-loop technology from acoustic signal acquisition to blockchain trusted storage.

[0123] CdTe quantum dots were excited, fluorescence decay curves were collected, and optical hashes were obtained by feature extraction.

[0124] Methods for obtaining the optical hash Hl include:

[0125] The time-domain decay signal l(t) is obtained and discretized to obtain the discrete signal l[n].

[0126] DWT is performed on l[n] using the Morlet wavelet basis. At F preset scales, the E wavelet coefficients W with the largest amplitudes are selected at each scale, for a total of 2K W. For each selected W, the phase is calculated, and the 2K phases are normalized to [0, 2]. K / 2 The range is -1], and 2K K / 2-bit integers are obtained;

[0127] 2K K / 2-bit integers are concatenated into a binary string, and then compressed into a 256-bit hash value using the SHA-256 hash function; this yields the optical hash.

[0128] By acquiring and discretizing the time-domain decay signal of CdTe quantum dots, a multi-scale DWT transform is performed on the discrete signal using the Morlet wavelet basis. Wavelet coefficients with the largest amplitude are selected at a preset scale, and the phase is calculated. After normalizing and quantizing the phase values, they are concatenated into a binary string, and then a 256-bit optical hash is generated using the SHA-256 hash function. This method leverages the time-frequency localization characteristics of wavelet transform to solve the technical problems of incomplete decay curve feature extraction and weak noise resistance in traditional fluorescence detection. By fusing multi-scale analysis and phase features, subtle differences in quantum dot fluorescence decay are captured, achieving accurate characterization of the optical signal after the interaction of medicinal material DNA and origami structure. Simultaneously, the collision resistance and irreversibility of the hash algorithm ensure the integrity and security of the optical feature data. This optical hash, in conjunction with acoustic hash and other multi-modal features, constructs a multi-dimensional identity identifier for medicinal materials, effectively solving the industry pain points of insufficient accuracy in single optical signal identification and easy tampering of data storage. It provides a high-precision and high-reliability optical fingerprint for the identification of genuine and counterfeit medicinal materials, helping to achieve a closed-loop technology process from molecular recognition to blockchain-based trusted storage.

[0129] After applying chaotic encryption to acoustic and optical hashes, the results are combined with identification tags for on-chain evidence storage.

[0130] Methods for chaotic encryption of acoustic hashes and optical hashes include:

[0131] Select the Lorentz parameter, such as using the real-time resonant frequency of the SAW-PUF chip as the initial value;

[0132] The Lorentz parameters are calculated in M ​​steps iteratively, the x-values ​​of the chaotic trajectory are extracted, and the trajectory is mapped to the [0,1] interval through the sigmoid function to generate an M-bit chaotic key stream.

[0133] The acoustic hash and optical hash are converted into 256-bit hexadecimal strings to obtain preprocessed data. The identification tag and preprocessed data are XORed byte by byte with the chaotic key stream to obtain encrypted data.

[0134] For the chaotically encrypted data, the SHA-256 algorithm is used to calculate the acoustically enhanced hash; the SHA-3 algorithm is used to calculate the optically enhanced hash.

[0135] The SHA-256 algorithm is then used to compute and encrypt the acoustic enhanced hash, optical enhanced hash, and chaotic key stream.

[0136] A verification contract is deployed on the blockchain for data integrity verification and timestamp consensus. During data integrity verification, the result L (obtained through chaotic encryption) is acquired. The contract then calls a SAW-PUF chip to generate a PUF key in real time, reproducing the chaotic system calculation to verify the legitimacy of H. This is further enhanced by the blockchain's timestamp service to ensure the immutability of the time data is uploaded to the blockchain.

[0137] The blockchain only writes H to the blockchain, the original encrypted data is stored in IPFS, and only the IPFS hash value is recorded on the chain.

[0138] If the tag is identified as genuine, the blockchain node only needs one SAW-PUF resonance verification; if the tag is identified as fake, five SAW-PUF verifications (which can be adaptively adjusted according to the actual situation) plus chaotic system recalculation are triggered.

[0139] By selecting the real-time resonant frequency of the SAW-PUF chip as the initial value of the Lorentz system and iteratively generating a chaotic key stream, the acoustic hash and optical hash are converted into hexadecimal strings and then XORed with the identification tag. Then, a quantum-resistant enhanced hash is generated through dual reinforcement using SHA-256 and SHA-3 algorithms. Finally, the hash value is written to the blockchain and the original encrypted data is stored in IPFS. Combined with a differentiated SAW-PUF verification mechanism based on the authenticity of the identification tag (e.g., one verification for genuine tags / five verifications for counterfeit tags), the above method utilizes the dynamic key characteristics of chaotic encryption to solve the problem of fixed and easily cracked keys in traditional hash encryption. It leverages the physical non-cloning technology of SAW-PUF to ensure the uniqueness of the initial key, and achieves immutability and efficient storage of data on the chain through blockchain timestamps and IPFS distributed storage. The differentiated verification mechanism improves the efficiency of verifying genuine medicinal materials while increasing the cost of tampering with counterfeit products. Ultimately, it constructs a full-link anti-counterfeiting system from multimodal feature encryption to hardware-level trusted evidence storage, effectively solving the technical challenge of balancing data security, evidence credibility, and verification efficiency in medicinal material identification.

[0140] This embodiment solves the technical problems existing in the prior art by integrating deep DNA enrichment using topologically protected acoustic waves, molecular computation using optically controlled CRISPR-DNA origami logic gates, and multi-dimensional verification using AFM force spectrum-acoustic-optical dual-modal hashing. Therefore, the inventors have broken through the disciplinary barriers of acoustic topological protection, molecular logic computation, and nanomechanical detection to address the technical problems existing in the prior art. They have proposed a scheme that uses the anti-interference properties of topological acoustic wave boundary states for DNA directional enrichment, uses optically controlled origami structures to replace nucleic acid amplification to achieve single-molecule computation, and combines SAW-PUF physical keys to construct quantum-resistant hashing. This scheme requires the inventors to make creative contributions in the intersection of topological acoustics and biological detection. The final technical solution has significant substantive features and progress.

[0141] Example 2

[0142] The method for obtaining the target medicinal material in this embodiment is the same as in Embodiment 1, except that the duration of the topological protection acoustic waves emitted to the medicinal material is 40ms, 60ms, 80ms, 100ms, 120ms, 140ms, and 160ms, and multiple identical experiments are set up for each group of experiments, and the average value is calculated; the results of the corresponding DNA enrichment efficiency are shown in Table 1.

[0143] Table 1. Effect of Topological Protection Acoustic Wave Time on DNA Enrichment Efficiency

[0144]

[0145]

[0146] As shown in the table, the DNA enrichment efficiency is highest when the duration of the topological protection acoustic wave emitted to the medicinal material is 100 ms.

[0147] Example 3

[0148] The method for obtaining the target medicinal material in this embodiment is the same as in Embodiment 1, except that the frequencies of the topological protection acoustic waves emitted to the medicinal material are 4MHz, 6MHz, 8MHz, 10MHz, 12MHz, 14MHz, and 16MHz. Multiple sets of the same experiment are set up for each group, and the average value is calculated. The results of the corresponding DNA enrichment efficiency are shown in Table 2.

[0149] Table 2. Data on the Influence of Topological Protection Acoustic Wave Frequency on DNA Enrichment Efficiency

[0150]

[0151]

[0152] As shown in the table, the DNA enrichment efficiency is highest when the frequency of the topological protection acoustic wave emitted to the medicinal material is 10MHz.

[0153] Example 4

[0154] The method for obtaining the target medicinal material in this embodiment is the same as in Example 1, except that the BSA solution concentrations are 0.5%, 1%, 3%, 5%, 6%, 7%, and 8%, and multiple identical experiments are set up for each group, with the average value calculated. The results of the corresponding non-specific binding rates are shown in Table 3.

[0155] Table 33 shows the effect of BSA solution concentration on nonspecific binding rate.

[0156] Experimental group (BSA concentration / %) Influencing indicator: Non-specific binding rate (%) 0.5 18.76 1 8.32 3 3.15 5 3.28 6 4.59 7 6.81 8 10.24

[0157] As shown in the table, the nonspecific binding rate was lowest when the BSA solution concentration was 3%.

[0158] Example 5

[0159] The method for obtaining the target medicinal material in this embodiment is the same as in Example 1, except that the molar ratio of quantum dots to antibodies is 1:5, 1:1, 5:1, 10:1, 30:1, 50:1, and 70:1, and multiple identical experiments are set up for each group of experiments, and the average value is calculated; the results of the corresponding probe labeling rates are shown in Table 4.

[0160] Table 4. Effect of quantum dot to antibody molar ratio on probe labeling rate.

[0161] Experimental group (molar ratio) Influencing metric: Probe labeling rate (%) 1:5 55.12 1:1 59.87 5:1 62.58 10:1 79.36 30:1 91.74 50:1 88.62 70:1 68.23

[0162] As shown in the table, the probe labeling rate is highest when the molar ratio of quantum dots to antibodies is 30:1.

[0163] Example 6

[0164] The method for obtaining the target medicinal material in this embodiment is the same as in Embodiment 1, except that the centrifugation speeds are 8000 rpm, 10000 rpm, 12000 rpm, 15000 rpm, 16000 rpm, 17000 rpm and 18000 rpm respectively. Multiple sets of the same experiment are set for each group of experiments, and the average value is calculated. The results of the corresponding precipitate purity are shown in Table 5.

[0165] Table 5. Data on the effect of centrifugation speed on precipitation purity

[0166] Experimental group (speed / rpm) Influencing indicator: Precipitate purity (%) 8000 75.32 10000 86.59 12000 92.17 15000 91.83 16000 89.46 17000 85.72 18000 81.55

[0167] As shown in the table, the precipitate purity is highest when the centrifugation speed is 12000 rpm.

[0168] Example 7

[0169] The method for obtaining the target medicinal material in this embodiment is the same as in Embodiment 1, except that the shaking incubation time is 0.5h, 1h, 2h, 3h, 4h, 5h and 6h respectively. Multiple sets of the same experiment are set for each group of experiments, and the average value is calculated. The results of the corresponding probe binding efficiency are shown in Table 6.

[0170] Table 6. Effect of Oscillation Incubation Time on Probe Binding Efficiency

[0171]

[0172]

[0173] As shown in the table, the probe binding efficiency is highest when the oscillation incubation time is 2 hours.

[0174] Example 8

[0175] The method for obtaining the light-controlled origami probe solution in this embodiment is the same as in Example 1, except that the molar ratio of gold nanoparticles to His-tag is 1:2, 1:1, 2:1, 3:1, 4:1, 5:1 and 6:1, and multiple identical experiments are set up for each group of experiments, and the average value is calculated. The results of the corresponding coupling efficiency and non-specific binding rate are shown in Table 7.

[0176] Table 7. Effect of the molar ratio of gold nanoparticles to His-tag on coupling efficiency.

[0177] Experimental group (molar ratio) Influencing indicator: Coupling efficiency (%) Non-specific binding rate (%) 1:2 49.54 18.45 1:1 57.34 11.48 2:1 65.32 12.78 3:1 82.47 8.31 4:1 91.63 4.29 5:1 89.75 5.16 6:1 83.24 7.43

[0178] As shown in the table, the coupling efficiency is the highest and the non-specific binding rate is the lowest when the molar ratio of gold nanoparticles to His-tag is 4:1.

[0179] Example 9

[0180] The method for obtaining the target medicinal material in this embodiment is the same as in Example 1, except that the molar ratio of biotinylated DNA to purified structure is 1:2, 1:1, 2:1, 3:1, 4:1, 5:1, 6:1 and 7:1, respectively. Multiple sets of the same experiment are set up for each group of experiments, and the average value is calculated. The results of comparing the corresponding fluorescence signal intensity and steric retardation are shown in Table 8.

[0181] Table 8. Effect of the molar ratio of biotinylated DNA to purified structure on signal intensity.

[0182] Experimental group (molar ratio) Influencing indicator: Fluorescence signal intensity (au) Spatial resistivity (%) 1:2 869.17 15.6 1:1 928.37 16.7 2:1 1168.41 21.5 3:1 1247.32 25.6 4:1 1863.59 18.2 5:1 2135.87 12.9 6:1 2017.43 15.7 7:1 1782.65 21.4

[0183] As shown in the table, the highest fluorescence signal intensity and the lowest steric hindrance were observed when the molar ratio of biotinylated DNA to purified structure was 5:1.

[0184] Example 10

[0185] The method for obtaining the target medicinal material in this embodiment is the same as in Example 1, except that the backbone chain / staple chain concentrations are 6nM / 6μM, 8nM / 8μM, 10nM / 10μM, 12nM / 12μM and 14nM / 14μM, respectively. Multiple sets of the same experiment are set up for each group of experiments, and the average value is calculated. The results of the corresponding correct assembly rate and polymer formation rate are shown in Table 9.

[0186] Table 9. Data on the effect of skeleton chain and staple chain concentration on assembly yield.

[0187] Experimental group (nM / μM) Influencing indicator: Correct assembly rate (%) Polymer formation rate (%) 6 / 6 58.32 28.7 8 / 8 76.49 15.3 10 / 10 89.27 7.6 12 / 12 85.63 10.4 14 / 14 78.15 16.2

[0188] As shown in the table, the correct assembly rate is the highest and the polymer formation rate is the lowest when the backbone chain / staple chain concentration is 10 nM / 10 μM.

[0189] Example 11

[0190] The method for obtaining the target medicinal material in this embodiment is the same as in Example 1, except that the annealing temperatures are 50℃, 55℃, 60℃, 65℃ and 70℃, and multiple identical experiments are set up for each group of experiments, and the average value is calculated; the results of comparing the corresponding Tm values ​​and unfolding rates are shown in Table 10.

[0191] Table 10. Data on the Influence of Annealing Temperature on Structural Stability

[0192]

[0193]

[0194] As shown in the table, the Tm value is the highest and the defolding rate is the lowest when the annealing temperature is 60℃.

[0195] Example 12

[0196] The method for obtaining the target medicinal material in this embodiment is the same as in Example 1, except that the ultraviolet irradiation time is 6s, 8s, 10s, 12s and 14s respectively. Multiple sets of the same experiment are set for each group of experiments, and the average value is calculated. The results of the corresponding conformational conversion rate and irreversible damage rate are shown in Table 11.

[0197] Table 11. Effect of UV irradiation time on origami conformation conversion rate.

[0198] Experimental group(s) Influencing indicator: Conformation conversion rate (%) Irreversible damage rate (%) 6 42.63 2.1 8 78.49 3.5 10 91.27 4.8 12 89.63 7.2 14 76.35 12.4

[0199] As shown in the table, the conformational conversion rate is highest when the ultraviolet light irradiation time is 10s, and the irreversible damage rate is lowest when the ultraviolet light irradiation time is 5s.

[0200] Example 13

[0201] The method for obtaining the target medicinal material in this embodiment is the same as in Example 1, except that the concentrations of Phi29 polymerase are 6U, 8U, 10U, 12U and 14U, and multiple identical experiments are set up for each group of experiments, and the average value is calculated; the results of the corresponding amplification yield and product length uniformity are shown in Table 12.

[0202] Table 12 shows the effect of Phi 29 polymerase concentration on amplification efficiency.

[0203] Experimental group (U) Influencing indicator: Amplification yield (ng / μL) Product length uniformity (%) 6 127.3 68.5 8 215.6 79.3 10 289.4 87.6 12 273.8 85.2 14 241.5 78.9

[0204] As shown in the table, the amplification yield and product length uniformity were highest when the Phi29 polymerase concentration was 10U.

[0205] Example 14

[0206] The method for obtaining the target medicinal material in this embodiment is the same as in Example 1, except that the Cas12a / gRNA concentration ratios are 60nM / 16μM, 80nM / 180μM, 100nM / 200μM, 120nM / 220μM and 140nM / 240μM, respectively. Multiple sets of the same experiment are set up for each group, and the average value is calculated. The results of the corresponding cleavage efficiency and non-specific cleavage rate are shown in Table 13.

[0207] Table 13. Effect of Cas12a / gRNA concentration ratio on enzyme digestion efficiency.

[0208] Experimental group (nM / μM) Influencing indicator: Cutting efficiency (%) Non-specific cutting rate (%) 60 / 160 62.7 18.4 80 / 180 78.3 12.6 100 / 200 89.5 7.3 120 / 220 87.2 9.1 140 / 240 82.6 13.7

[0209] As shown in the table, the cleavage efficiency is the highest and the nonspecific cleavage rate is the lowest when the Cas12a / gRNA concentration ratio is 100nM / 200μM.

[0210] Example 15

[0211] This embodiment provides a sorting method for origami structures used in Embodiment 1, comprising the following steps:

[0212] The AFM image is used as input to the U-Net network to obtain the segmentation results, and the free energy of the corresponding origami structure is calculated based on the segmentation results.

[0213] Training methods for the U-Net network include:

[0214] W sets of image training data were collected in advance, including AFM images and segmentation results;

[0215] Image training data is used as input to the U-Net network, and the segmentation results are used as output. With the goal of minimizing the error between the output segmentation result and the actual segmentation result, the network parameters of the U-Net network are optimized using a natural heuristic optimization algorithm. The network parameters that minimize the error between the output segmentation result and the actual segmentation result are obtained. The U-Net network constructed with the corresponding network parameters is used as the trained U-Net network.

[0216] Example 16

[0217] Please see Figure 4 As shown, this embodiment provides a medicinal herb identification system, including:

[0218] Enrichment sampling module: Emits topologically protected acoustic waves to the medicinal material, and modifies the medicinal material DNA with CdTe quantum dots-nucleic acid probes in the microcavity of the medicinal material to obtain a medicinal material sample;

[0219] Sample processing module: scrape the medicinal sample from the microcavity of the medicinal material, extract DNA from the medicinal sample by CTAB method, amplify the DNA target gene by PCR, design the logic gate staple chain based on the DNA target gene sequence, and prepare the light-controlled origami probe solution;

[0220] Authenticity identification module: The optically controlled origami probe solution is injected into the nanopore array of the PZT chip and photoactivated to obtain an activated origami structure. An axial force is applied to the activated origami structure through an AFM probe, and the free energy of the activated origami probe is analyzed. Based on the free energy and a preset database, the medicinal materials are identified to obtain an identification label.

[0221] First extraction module: Emits topology protection acoustic waves to the PZT chip, collects the acoustic energy spectrum, performs feature extraction, and obtains acoustic hash;

[0222] The second extraction module: excites CdTe quantum dots, collects fluorescence decay curves, and performs feature extraction to obtain optical hashes;

[0223] Encrypted Evidence Preservation Module: After performing chaotic encryption on acoustic hash and optical hash, it combines the identification tag to perform on-chain evidence preservation.

[0224] The above description is merely a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any variations or substitutions that can be easily conceived by those skilled in the art within the technical scope disclosed in the present invention should be included within the scope of protection of the present invention. Therefore, the scope of protection of the present invention should be determined by the scope of the claims.

[0225] In conclusion, the above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for identifying medicinal materials, characterized in that, Includes the following steps: Topologically protected acoustic waves are emitted into the medicinal material, and the DNA of the medicinal material is labeled by CdTe quantum dots-nucleic acid probes in the microcavity of the medicinal material to obtain the target medicinal material; A sample of the target medicinal material was scraped from the microcavity, and DNA was extracted from the sample using the CTAB method. The DNA target gene was amplified by PCR, and a logic gate staple chain was designed based on the DNA target gene sequence to prepare a light-controlled origami probe solution. The photo-controlled origami probe solution is injected into the nanopore array of the PZT chip and photoactivated to obtain an activated origami structure. An axial force is applied to the activated origami structure using an AFM probe, and the free energy of the activated origami probe is analyzed. Based on the free energy and a preset database, medicinal materials are identified to obtain identification labels. Methods for obtaining identification tags include: An axial force is applied to activate the origami structure using an AFM probe, and the AFM force curve is recorded. The stretching curve corresponds to the molecular unfolding process, and the retraction curve corresponds to the molecular folding process. When the molecules in the folded region are not stretched, the conformational length corresponding to the displacement of the molecule is taken as the folding displacement; in the completely unfolded region, the conformational length when the force change is lower than a preset threshold, corresponding to the length of the stretched molecule, is taken as the completely unfolding displacement. The free energy of the medicinal material is obtained by integrating the AFM force curve by combining the folded displacement and the completely defolded displacement. The system detects whether the free energy of medicinal materials is within the range of the free energy of genuine medicinal materials in the preset database. If it is, the material is identified as genuine and marked with an identification label of 1; otherwise, it is identified as counterfeit and marked with an identification label of 0. A topology protection acoustic wave is emitted to the PZT chip, the acoustic energy spectrum is collected, features are extracted, and an acoustic hash is obtained. CdTe quantum dots were excited, fluorescence decay curves were collected, and optical hashes were obtained by feature extraction. After applying chaotic encryption to acoustic and optical hashes, the results are combined with identification tags for on-chain evidence storage.

2. The method for identifying medicinal materials according to claim 1, characterized in that, Methods for obtaining the target medicinal material include: Emit 8-12MHz topological protection acoustic waves for 80-120ms to the medicinal materials, then place the medicinal materials in PBS buffer and shake to clean them. Repeat 2-3 times. Add 1%-5% BSA solution to the cleaned medicinal materials and incubate at room temperature for 30-60 minutes; Wash the medicinal materials again with PBS buffer 3-5 times to obtain the medicinal materials to be processed; In a centrifuge tube, add CdTe quantum dot solution and antibody solution in a molar ratio of 10:1-50:1, mix well, and shake on a shaker at room temperature for 1-2 hours. After shaking, place the centrifuge tubes in a centrifuge and centrifuge at 10,000-15,000 rpm for 10-15 minutes. The precipitate was resuspended in PBS buffer to obtain the CdTe quantum dot-antibody probe solution; Place the medicinal material to be treated into a container containing CdTe quantum dot-antibody probe solution; incubate at room temperature for 1-3 hours with shaking at 100-200 rpm on a shaker. After incubation, wash the medicinal material 3-5 times with PBS buffer to obtain the target medicinal material.

3. The method for identifying medicinal materials according to claim 1, characterized in that, Methods for obtaining optically controlled origami probe solutions include: Origami structures were prepared based on medicinal material samples; 3-8 nm gold nanoparticles were prepared, Ni-NTA was modified on the surface of the gold nanoparticles, and His-tag was mixed with Ni-NTA at a molar ratio of 3:1-5:

1. The mixture was reacted at 20-30 °C in the dark for 10-15 h to form directional coupling. The photolysis linker was dissolved in DMSO and mixed with the origami structure at a molar ratio of 1:

1. The mixture was reacted at room temperature in the dark for 1-3 hours, and then purified by magnetic beads to obtain the purified structure. The biotinylated DNA short strand and the vertex sequence of the purified structure are incubated at 35-38℃ for 20-40 min in a molar ratio of 4:1-6:1 to form the biotinylated exposed end and obtain the labeled structure. Streptavidin-modified CdSe@ZnS quantum dots were added to the labeled structure at a molar ratio of 1:

1. The reaction was carried out at 4-6℃ in the dark for 1-2 hours. The mixture was then filtered using a 90-120kDa membrane at a flow rate of 1-3mL / min to obtain the light-controlled origami structure.

4. The method for identifying medicinal materials according to claim 3, characterized in that, Methods for obtaining origami structures include: Design a 110-130nt circular DNA template containing a latch strand sequence, a logic gate targeting sequence, and a complementary sequence to the M13mp18 phage backbone strand; Add 80-150 μL of amplification reaction system and react at 25-35℃ for 15-18 hours to obtain long-chain backbone DNA containing the latching strand; Add 80-120 nM Cas12a and 180-220 nM gRNA to the amplification product, incubate at 35-38℃ for 1-2 h, cleave the long backbone containing the locked strand, and then purify it with streptavidin magnetic beads to obtain the backbone strand. Design a logic gate staple chain, introduce an azobenzene photoresponsive group at the 5' end of the logic gate staple chain, and modify the 3' end with a His-tag; Add a self-assembly system, denature at 90-100℃ for 3-8 minutes, anneal at 55-65℃ for 1-2 hours; slowly cool to 25-30℃ for 48-50 hours; then irradiate with 365nm ultraviolet light for 8-12 seconds to obtain the origami structure.

5. The method for identifying medicinal materials according to claim 4, characterized in that, The logic gate targeting sequence is complementary to the DNA of the medicinal sample; The amplification reaction system was prepared by using 8-12 U Phi29 DNA polymerase, 1-3 mM dNTPs containing 10% dUTP, 0.08-0.12 nM circular template, and 0.08-0.12 nM 5'-phosphorylated primers in 18-22 mM Tris-HCl buffer at pH 7.8-8.2 containing 8-12 mM Mg²⁺ to activate Cas12a. The logic gate staple chain includes an 18-22nt target segment complementary to the DNA of the medicinal sample, a 13-18nt CRISPR response segment containing a PAM sequence recognized by Cas12a, and a preset anchor point. The self-assembled system includes an 8-12 nM backbone chain, an 8-12 μM logic gate stapled chain pool, 18-22 mM Tris-HCl with a pH of 7.8-8.2, 10 mM MgCl2, and 50 mM NaCl.

6. The method for identifying medicinal materials according to claim 1, characterized in that, Also includes: Methods for obtaining acoustic hashes include: The time-domain pressure signal is acquired, discretized, and a discrete signal s[n] is obtained. Perform a discrete Fourier transform on s[n] to convert it into a frequency domain amplitude spectrum S[k]; From S[k], obtain the K frequency points with the largest amplitude, corresponding to the spectral line indices, and calculate the K frequency values; obtain the K frequency points with the second largest amplitude, corresponding to the spectral line indices, and calculate the K frequency values. Normalize the 2K frequency values ​​to [0, 2]. K / 2 The range is -1], and 2K K / 2-bit integers are obtained; 2K K / 2-bit integers are concatenated into a binary string, and then compressed into a 256-bit hash value using the SHA-256 hash function; this yields the acoustic hash.

7. The method for identifying medicinal materials according to claim 6, characterized in that, Methods for obtaining the optical hash Hl include: The time-domain decay signal is acquired and discretized to obtain the discrete signal l[n]. DWT is performed on l[n] using the Morlet wavelet basis. At F preset scales, the E wavelet coefficients W with the largest amplitudes are selected at each scale, for a total of 2K W. For each selected W, the phase is calculated, and the 2K phases are normalized to [0, 2]. K / 2 The range is -1], and 2K K / 2-bit integers are obtained; 2K K / 2-bit integers are concatenated into a binary string, and then compressed into a 256-bit hash value using the SHA-256 hash function; this yields the optical hash.

8. The method for identifying medicinal materials according to claim 7, characterized in that, Methods for chaotic encryption of acoustic hashes and optical hashes include: Select the Lorentz parameter, iteratively calculate the Lorentz parameter in M ​​steps, extract the x value of the chaotic trajectory, map it to the [0,1] interval through the sigmoid function, and generate an M-bit chaotic key stream; The acoustic hash and optical hash are converted into 256-bit hexadecimal strings to obtain preprocessed data. The identification tag and preprocessed data are XORed byte by byte with the chaotic key stream to obtain encrypted data. For encrypted data, the SHA-256 algorithm is used to calculate the acoustically enhanced hash; the SHA-3 algorithm is used to calculate the optically enhanced hash. The SHA-256 algorithm is then used to compute and encrypt the acoustic enhanced hash, optical enhanced hash, and chaotic key stream.

9. A medicinal herb identification system, implementing the medicinal herb identification method according to any one of claims 1-8, characterized in that, include: Enrichment sampling module: emits topologically protected acoustic waves to the medicinal material, and modifies the medicinal material DNA with CdTe quantum dots-nucleic acid probes in the microcavity of the medicinal material to obtain a medicinal material sample; Sample processing module: scrape the medicinal sample from the microcavity of the medicinal material, extract DNA from the medicinal sample by CTAB method, amplify the DNA target gene by PCR, design the logic gate staple chain based on the DNA target gene sequence, and prepare the light-controlled origami probe solution; Authenticity identification module: The optically controlled origami probe solution is injected into the nanopore array of the PZT chip and photoactivated to obtain an activated origami structure. An axial force is applied to the activated origami structure through an AFM probe, and the free energy of the activated origami probe is analyzed. Based on the free energy and a preset database, the medicinal materials are identified to obtain an identification label. First extraction module: Emits topology protection acoustic waves to the PZT chip, collects the acoustic energy spectrum, performs feature extraction, and obtains acoustic hash; The second extraction module: excites CdTe quantum dots, collects fluorescence decay curves, and performs feature extraction to obtain optical hashes; Encrypted evidence storage module: After performing chaotic encryption on acoustic hash and optical hash, it combines identification tags for on-chain evidence storage.

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