Magnetically-enhanced light-driven hydrogel micro-robot and application thereof in detecting biological fluid
The hydrogel microrobot driven by magneto-enhanced light recognizes and determines pollutants in biological fluids, solving the detection problems caused by sensor contamination and high viscosity, achieving high sensitivity and rapid analysis, and is suitable for the detection of complex biological fluids.
Patent Information
- Application Number
- CN202510596473.0
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-05-09
- Publication Date
- 2025-08-15
AI Technical Summary
There are problems in biological fluid detection such as sensor contamination and passivation, high viscosity leading to long analysis time, low mass transfer efficiency and low sensitivity.
A hydrogel microrobot driven by magneto-enhanced light, including hydrogel droplets and calcium alginate gel layer, contains magnetic materials, light-driven materials and molecular imprinting materials, identify target pollutants in biological fluids through light-driven and magnetic movement, and their content is determined by electrochemical methods.
It improves the anti-pollution ability and sensitivity of the sensor, shortens the analysis time, enhances the mass transfer efficiency, and has good biocompatibility, and is suitable for efficient detection of trace targets in complex biological fluids.
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Abstract
Description
Technical Field
[0001] The present invention belongs to the field of electrochemical sensing, and in particular relates to a magnetically enhanced light-driven hydrogel microrobot and a preparation method thereof, as well as applications in detecting biological fluids. Background Art
[0002] With the advancement of industrial technology, artificial chemical products are widely used around the world. When discharged into the environment, they form pollutants. After pollutants enter the human body through various contact routes, they can be transported to different organs through the blood circulation. Therefore, rapid and highly sensitive biomonitoring of pollutants in various biological fluids can not only determine the accumulation level of pollutants in the body, but also investigate the metabolic processes and potential health risks associated with exposure to these pollutants. Biological fluids include blood, urine, bile, breast milk, cerebrospinal fluid, saliva, and sweat, which play a vital role in maintaining life activities and physiological functions. Electrochemical sensors have the advantages of simple and fast operation, high sensitivity, low instrumentation, and easy miniaturization.
[0003] However, the detection of biofluids faces many difficulties: 1) The presence of numerous biomacromolecules and biomicromolecules in complex biofluids can easily cause biofouling of electrochemical sensors, resulting in sensor passivation and reduced analytical sensitivity; 2) The high viscosity of biofluids can lead to low mass transfer efficiency, hindering the analyte from reaching the detection sensor, resulting in low detection signals; 3) Toxic materials modified on the sensor surface can produce biotoxicity in direct contact with biofluids and poor biocompatibility; 4) When using the solution dilution method (100 to 1000 times) for biofluid detection, trace amounts of the target are diluted, sample cross-contamination is easily caused, and the method is time-consuming. Summary of the Invention
[0004] The technical problem to be solved by the present invention is to address the deficiencies of the above-mentioned existing technologies and provide a magnetically enhanced light-driven hydrogel microrobot, which solves the problems faced in biological fluid detection such as sensor contamination and passivation caused by complex solution systems, long analysis time, low mass transfer efficiency and low sensitivity caused by high viscosity fluids.
[0005] The technical solution adopted by the present invention to solve the above-mentioned technical problems is: A magnetically enhanced light-driven hydrogel microrobot comprises a hydrogel droplet and a hydrogel coating; wherein the hydrogel droplet is a hydrogel containing a magnetic material, a light-driven material and a molecularly imprinted material; and the hydrogel coating is a calcium alginate gel layer.
[0006] According to the above scheme, the light-driven material uses bismuth vanadate-carbon nanotubes (BiVO4-MWCNT), and its preparation method is: using bismuth nitrate and ammonium metavanadate as precursors, adding carboxylated carbon nanotubes during the hydrothermal synthesis of bismuth vanadate in nitric acid solution, thereby modifying carbon nanotubes on the synthesized bismuth vanadate to obtain bismuth vanadate-carbon nanotubes.
[0007] According to the above scheme, the water content of the hydrogel droplets is between 85% and 95%. The mass ratio of the magnetic material, bismuth vanadate-carbon nanotubes, and molecularly imprinted material in the hydrogel droplets is 1:(2-4):(2-4). The molecularly imprinted material can be directly a molecularly imprinted polymer (MIP), or the MIP can be formed by synthesizing a MIP on bismuth vanadate-carbon nanotubes.
[0008] According to the above scheme, the molecularly imprinted material is a molecularly imprinted polymer (MIP) of the target, capable of specific recognition of the target. Appropriate functional monomers are selected based on the properties of the target. Using the target as a template molecule, the functional monomers polymerize under the action of a crosslinker and initiator to produce a MIP that specifically recognizes the target. During the synthesis of the MIP, an appropriate amount of bismuth vanadate-carbon nanotubes can be added along with the crosslinker and initiator to produce a MIP formed on the bismuth vanadate-carbon nanotubes. This MIP can form stable non-covalent bonds (such as hydrogen bonds) or electrostatic interactions with the target, thereby achieving specific recognition. For example, for targets containing acidic groups, such as perfluorooctane sulfonic acid, functional monomers containing basic functional groups, such as amino groups, can be selected.
[0009] According to the above scheme, the hydrogel droplets are preferably in the shape of water droplets, and the radial size of the water droplets is generally in the range of 450 μm ~ 1200 μm, preferably 900 μm ~ 1100 μm, and the axial size is generally in the range of 300 μm ~ 900 μm, preferably 600 μm ~ 800 μm, and the radial size is larger than the axial size; the thickness of the hydrogel coating is generally only 4 ~ 12 μm, so the size of the magnetically enhanced light-driven hydrogel microrobot can be considered to be basically equivalent to the hydrogel droplets.
[0010] According to the above solution, the magnetic material is preferably one of magnetic ferric oxide (γ-Fe2O3), ferrosoferric oxide (Fe3O4), etc., or a mixture of two or more.
[0011] The present invention also provides a method for preparing the above-mentioned magnetically enhanced light-driven hydrogel microrobot, comprising the following steps: (1) Mixing the magnetic material, the light-driven material, and the molecularly imprinted material in a sodium alginate solution and uniformly dispersing them to obtain a precursor solution; (2) The precursor solution is injected into the calcium chloride solution using a microfluidic syringe to form water droplet-shaped hydrogel droplets; (3) The above-mentioned hydrogel droplets are placed on a nylon mesh, and sodium alginate solution and calcium chloride solution are added dropwise in sequence to form a calcium alginate gel layer on the surface of the hydrogel droplets, thereby obtaining a magnetically enhanced light-driven hydrogel microrobot.
[0012] According to the above scheme, in step (1), the concentration of the sodium alginate solution is 1.5 ~ 2.5 wt%; the concentrations of the magnetic material and the light-driven material in the sodium alginate solution are 2 ~ 4 mg / mL and 6 ~ 12 mg / mL, respectively; and the concentration of the molecular imprinting material in the sodium alginate solution is 6 ~ 12 mg / mL.
[0013] According to the above scheme, in step (2), the concentration of the calcium chloride solution is 0.2 to 0.8 wt%; the vertical height of the microfluidic syringe from the surface of the calcium chloride solution is 0.3 to 0.9 mm, the flow rate of the microfluidic injection is 0.01 to 0.1 mL / min, and the diameter of the microfluidic injection needle is 0.2 to 0.8 mm. The calcium chloride solution is contained in a culture dish.
[0014] According to the above scheme, in step (2), the radial size of the droplet-shaped hydrogel is 450 μm to 1200 μm, and the axial size is 300 μm to 900 μm. The asymmetric structure of the hydrogel droplet causes the hydrogel microrobot to produce non-uniform light absorption and photocatalytic effects under light irradiation, thereby generating an asymmetric driving force, which is conducive to its light-driven motion.
[0015] According to the above scheme, in step (3), the pore size of the nylon mesh is 60 to 100 mesh, the concentration of the sodium alginate solution is 0.2% to 1.2 wt%, and the concentration of the calcium chloride solution is 0.3% to 2 wt%.
[0016] The application of the magnetically enhanced light-driven hydrogel microrobot of the present invention to detect pollutants in biological fluids is specifically as follows: After the magnetically enhanced light-driven hydrogel microrobot is placed in a culture dish containing biological fluid, fuel is added, and then a permanent magnet is fixed above the culture dish. Under the irradiation of an excitation light source, the magnetically enhanced light-driven hydrogel microrobot undergoes magnetically enhanced light-driven movement in the biological fluid. At this time, the target object specifically binds to the molecular imprinting material in the magnetically enhanced light-driven hydrogel microrobot through hydrogen bonds, thereby identifying the target object (i.e., the pollutant to be detected) in the biological fluid; then a magnetic glassy carbon electrode is used to adsorb the magnetically enhanced light-driven hydrogel microrobot from the biological fluid, and then the robot is placed in a redox probe electrolyte solution and the differential pulse voltammetry curve is measured using an electrochemical workstation. The content of the target pollutant to be analyzed is determined using a standard curve method.
[0017] According to the above scheme, the concentration of the fuel added to the biological fluid is 55 to 75 mmol / L; the permanent magnet is fixed directly above the culture dish, with a vertical distance of 1 to 4 cm from the liquid surface of the culture dish, and the depth of the biological fluid in the culture dish is 1 to 5 mm; the direction of the light source is preferably at an angle of 30° to 60° with the liquid surface of the culture dish, and its vertical distance from the liquid surface is 5 to 10 cm, and the light source power is greater than 20 mW / mm 2 That's it.
[0018] According to the above scheme, the biological fluid includes complex biological samples such as whole blood, bile, and urine, and the target substances include perfluorooctane sulfonic acid (PFOS), polychlorinated biphenyls, pentachlorophenol, etc.
[0019] According to the above scheme, the fuel can be glucose, uric acid, malic acid, etc.
[0020] According to the above solution, the excitation light source can be a blue laser, a violet laser, etc.
[0021] According to the above scheme, the redox probe electrolyte solution uses a redox couple of potassium ferrocyanide and potassium ferrocyanide, which are dissolved in a potassium chloride electrolyte solution. The concentration of the potassium chloride solution is 0.05 to 0.2 mol / L, and the concentrations of potassium ferrocyanide and potassium ferrocyanide in the potassium chloride solution are both 2 to 10 mmol / L.
[0022] The technical principle of the present invention is as follows: when the magnetically enhanced light-driven hydrogel microrobot detects pollutants in biological fluids (taking perfluorooctane sulfonic acid as an example), under the irradiation of an excitation light source, the light-driven material (BiVO4-MWCNT) inside the hydrogel microrobot generates photogenerated electrons and holes. The fuel glucose in the biological fluid is oxidized by the photogenerated holes, generating an asymmetric chemical reaction on the light-driven material, thereby forming a concentration gradient of redox products (such as gluconic acid) around the hydrogel microrobot; then, the hydrogel microrobot is pushed toward the side facing away from the light by self-diffusion electrophoresis, undergoing magnetically enhanced light-driven motion, and identifying and binding perfluorooctane sulfonic acid in the biological fluid. min; the hydrogel microrobot is then magnetically collected using a magnetic glassy carbon electrode and placed in a redox probe electrolyte solution to measure the differential pulse voltammetry curve. Since perfluorooctane sulfonic acid is a non-electroactive molecule, it will hinder the electron transfer between the redox probe and the glassy carbon electrode described in the present invention. Therefore, the peak current of the differential pulse voltammetry curve will decrease with the increase of perfluorooctane sulfonic acid concentration. Based on this, the standard curve method is used to determine the content of perfluorooctane sulfonic acid in biological fluids.
[0023] Compared with the prior art, the beneficial results of the present invention are: 1. The magnetically enhanced light-driven hydrogel microrobots of the present invention have excellent anti-pollution capabilities, reducing contamination and passivation of sensors by complex biological environments such as biofluids, thereby improving detection sensitivity and accuracy. 2. The magnetically enhanced light-driven hydrogel microrobot described in the present invention has a high movement speed and can play a stirring role in complex biological fluids, thereby enhancing the mass transfer efficiency of the biological fluid and increasing the probability of contact between the sensor (i.e., the hydrogel microrobot) and the target molecule, thereby improving the detection sensitivity and shortening the analysis time; 3. The magnetically enhanced light-driven hydrogel microrobots described in the present invention have good biocompatibility and blood compatibility, which can provide a basis for subsequent in vivo experiments; 4. The magnetically enhanced light-driven hydrogel microrobot prepared by the present invention was successfully applied to the tracking and monitoring of PFOS in whole blood, providing an important research basis for the circulation and distribution of pollutants in the body.
[0024] In summary, the magnetically enhanced light-driven hydrogel microrobot described in the present invention realizes the detection of persistent organic pollutants in complex biological fluids, and has the advantages of high sensitivity, fast detection speed, good anti-pollution performance, and excellent biocompatibility; it is also low-cost and easy to miniaturize, and in the future it can be used for the efficient detection of various trace targets in complex biological fluids. BRIEF DESCRIPTION OF THE DRAWINGS
[0025] Figure 1The figure is a schematic diagram of the preparation process of magnetically enhanced light-driven hydrogel microrobots (HMRs) according to Example 1 of the present invention; wherein, Sodium alginate represents sodium alginate, Nylon mesh represents nylon mesh, Hydrogel microrobot represents HMRs, Magnetically Boosted Light-driven motion represents magnetically enhanced light-driven motion, MGCE represents magnetic glassy carbon electrode, Magnetic collection represents magnetic collection, and MIP for PFOS represents a molecularly imprinted polymer of PFOS or a molecularly imprinted material containing the molecularly imprinted polymer.
[0026] Figure 2 (a) Magnetic characterization of the magnetically enhanced light-driven HMRs prepared in Example 1; (b) Microscope image of the magnetically enhanced light-driven HMRs and calcium alginate gel layer prepared in Example 1 (the inset is a photo of the HMRs); (c) Schematic diagram of the movement principle of the magnetically enhanced light-driven HMRs prepared in Example 1; (d) Time-lapse image of the path change of the magnetically enhanced light-driven HMRs prepared in Example 1 controlled by changing the direction of the light source.
[0027] Figure 3 (a) Cytotoxicity characterization of the magnetically enhanced light-driven HMRs prepared in Example 1; (b) Blood compatibility characterization of the magnetically enhanced light-driven HMRs prepared in Example 1; (c) Red blood cell morphology after the magnetically enhanced light-driven HMRs came into contact with blood.
[0028] Figure 4 The magnetically enhanced light-driven motion speeds of the magnetically enhanced light-driven HMRs in different biological fluids in Example 2.
[0029] Figure 5 This is the linear relationship diagram of Example 3.
[0030] Figure 6 This is a test result diagram showing the changes in PFOS concentration in the blood of rats exposed to different concentrations of PFOS for different days in Example 4.
[0031] Figure 7 This is a comparison chart of the test results of measuring PFOS concentrations in rat blood after exposure to 200 μg / L PFOS for different days using the hydrogel microrobot sensor constructed in Example 4 and high-performance liquid chromatography. DETAILED DESCRIPTION
[0032] In order to better understand the present invention, the content of the present invention is further illustrated below in conjunction with examples, but the present invention is not limited to the following examples.
[0033] In the present invention, the size of the magnetic γ-Fe2O3 is less than 1 μm, and can be purchased or prepared by reference to literature.
[0034] In the present invention, the preparation process of the light-driven material BiVO4-MWCNT is as follows: multi-walled carbon nanotubes (MWCNTs) are placed in a mixed acid (the mixed acid is a mixture of 95% sulfuric acid and 65% nitric acid in a volume ratio of (2 to 4):1) and ultrasonically carboxylated for 20 to 40 minutes at a carboxylation temperature of 40 to 80°C. The solution is filtered through filter paper, thoroughly washed with water, and vacuum dried to obtain carboxylated MWCNTs. 2.6 to 3.2 g of bismuth nitrate pentahydrate and 0.5 to 0.9 g of ammonium metavanadate are added to 40 to 60 mL of nitric acid (nitric acid concentration of 0.5 to 1.5 M) and stirred to obtain a yellow uniform solution. 0 to 0.2 g of the carboxylated MWCNTs and 2 to 3 g of urea are then added and uniformly dispersed. The solution is then heated at 80 to 100°C for reaction for 10 to 14 hours, and a solid product, namely BiVO4-MWCNT, is collected.
[0035] In the following examples, the light-driven BiVO4-MWCNT material was prepared as follows: 1 g of MWCNT was placed in a mixed acid solution (95% sulfuric acid and 65% nitric acid, in a volume ratio of 3:1) at 60°C in a water bath and carboxylated under ultrasonic cleaning for 30 minutes. After dilution with water, the solution was filtered through a 0.22 μm filter paper, washed thoroughly with water, and vacuum-dried at 55°C. The carboxylated MWCNTs were then collected from the filter paper. 2.9 g of bismuth nitrate pentahydrate and 0.7 g of ammonium metavanadate were added to 50 mL of 1.0 M nitric acid and stirred to obtain a yellow, homogeneous solution. 0.15 g of the carboxylated MWCNTs was then added and stirred until dispersed. 2.5 g of urea was then added and, after uniform dispersion, heated to 90°C for 12 hours. The precipitate, BiVO4-MWCNT, was collected by vacuum filtration, washed with copious amounts of water, and dried in an oven for later use.
[0036] Among them, the length of MWCNT is 10 ~ 30 μm and the diameter is 10 ~ 20 nm.
[0037] In the present invention, the preparation process of the molecular imprinting material is as follows: 80 to 120 mg of the template molecule (perfluorooctane sulfonic acid, PFOS) and 0.12 to 0.16 g of the functional monomer (acrylamide) are dispersed in 20 to 40 mL of acetonitrile and stirred at room temperature for 10 to 14 hours, and then 0.7 to 0.9 g of the crosslinker (ethylene glycol dimethacrylate, EGDMA), 0.03 to 0.05 g of the initiator (azobisisobutyronitrile, AIBN) and 0.05 to 0.15 g of the light-driven material (BiVO4-MWCNT) are added in sequence. After nitrogen is passed through to remove oxygen for 20 to 40 minutes, stirring is continued at a reaction temperature of 40 to 80°C for 22 to 26 hours. The obtained solid is washed with acetonitrile and the solid product is separated. The template molecule 4 to 5 is eluted with a methanol-acetic acid mixed solution. The mixture was washed with methanol until neutral and vacuum dried to obtain a polyacrylamide imprinted polymer formed on the bismuth vanadate-carbon nanotube, which is a molecular imprinting material that can specifically recognize the target PFOS.
[0038] In the following examples, the molecularly imprinted material (MIP) used was prepared by the following specific process: 100 mg of PFOS and 0.1422 g of acrylamide were dispersed in 30 mL of acetonitrile and stirred at room temperature for 12 h. Then, 0.793 g of EGDMA, 0.045 g of AIBN, and 0.1 g of BiVO4-MWCNT were added in sequence, and nitrogen was passed through the mixture to deoxygenate for 30 min. The mixture was then stirred at 60°C for 24 h. The obtained solid was washed with acetonitrile and the solid product was isolated. The solid product was eluted with a methanol-acetic acid mixture with a volume ratio of 9:1 for 7 days to remove PFOS, washed with methanol until neutral, centrifuged, and vacuum dried at 50°C to obtain a polyacrylamide imprinted polymer formed on bismuth vanadate-carbon nanotubes, i.e., a molecularly imprinted material that can specifically recognize PFOS.
[0039] Example 1 A magnetically enhanced light-driven hydrogel microrobot (HMRs) is prepared by a method comprising the following steps: (1) 3 mg of magnetic γ-Fe2O3, 8 mg of BiVO4-MWCNT, and 8 mg of molecular imprinting material were added to 1 mL of 2 wt% sodium alginate solution and dispersed uniformly by ultrasonication to obtain a precursor solution; (2) Add 4 mL of 0.5 wt% calcium chloride solution to the culture dish to a depth of 1.5 cm, then inject the precursor solution into a syringe (needle: 27 G, diameter 0.4 mm), then place the syringe horizontally on the syringe pump, with the needle tip just above the culture dish and the vertical height of the needle tip from the surface of the calcium chloride solution to 0.7 mm. Push the syringe at a constant speed of 0.05 mL / min, and the outflowing precursor solution is divided into multiple small droplets, which fall into the culture dish to form water droplet-shaped calcium alginate hydrogel droplets with a water content of about 90%; (3) The above-mentioned hydrogel droplets were placed on an 80-mesh nylon net, and 0.6 wt% sodium alginate aqueous solution and 0.5 wt% calcium chloride aqueous solution were added dropwise in sequence to form a thin calcium alginate gel layer coated on the surface of the hydrogel droplets, thereby obtaining magnetically enhanced light-driven HMRs.
[0040] Performance Characterization (1) Magnetic and morphological characterization of magnetically enhanced light-driven HMRs like Figure 2 As shown in (a), applying a permanent magnet near the magnetically enhanced light-driven HMRs prepared in Example 1 can control them to move toward the magnet side, indicating that the magnetically enhanced light-driven HMRs of the present invention have good magnetic properties.
[0041] from Figure 2 From the microscope image and photograph in (b), it can be seen that the magnetically enhanced light-driven HMRs prepared in Example 1 are in the shape of tiny water droplets, with a radial size of about 1000 μm and an axial size of about 700 μm. It can also be clearly observed that the HMRs are wrapped with a thin layer of calcium alginate, and the thickness of the calcium alginate layer is 5 to 10 μm.
[0042] (2) Characterization of the motion performance of magnetically enhanced light-driven HMRs The principle of HMRs motion driven by magnetically enhanced light in the present invention is as follows: Figure 2 (c) shows: When one side of the magnetically enhanced light-driven HMRs is irradiated with blue light, the BiVO4-MWCNT therein is excited to generate photogenerated electrons and holes. When the magnetically enhanced light-driven HMRs are placed in a biological fluid, the biocompatible fuel glucose is oxidized by the photogenerated holes, generating an asymmetric chemical reaction on the BiVO4-MWCNT particles, thereby forming a concentration gradient of redox products (such as gluconic acid) around the surface of the HMRs, which then drives the movement of the HMRs through self-diffusion electrophoresis. Figure 2 As can be seen from the motion time-lapse image (d), HMRs show the expected light-driven motion. They can move under blue light irradiation and can achieve motion in different directions by changing the irradiation direction of light.
[0043] (3) Characterization of biocompatibility and hemocompatibility of magnetically enhanced light-driven HMRs The biocompatibility of the magnetically enhanced light-driven HMRs described in the present invention was evaluated by measuring cytotoxicity using a live / dead cell staining method. Human glioma cells were cultured in a 35 mm culture dish for 48 h to allow them to adhere to the wall, and then 20 HMRs were added to the culture medium and cultured for a further 24 h (experimental group). Afterwards, the cells were washed three times with PBS solution, and 1 mL of PBS containing 0.5 μg of calcein (live cells / green cytoplasm stain) and 0.25 μg of PI (dead cells / red nucleic acid stain) was added to the culture dish, and the cells were stained for 10 min (the control group was served without the addition of HMRs). The high cell survival rate in the experimental group was similar to that in the control group ( Figure 3 a), indicating that HMRs have good biocompatibility.
[0044] In addition, the hemolysis rate of HMRs and its effect on red blood cell morphology were evaluated. 2 mL of fresh blood was centrifuged at 1000 rpm for 10 min. After removing the supernatant, the red blood cells were washed several times by repeated centrifugation, and then a 50wt% red blood cell suspension was made with normal saline. Subsequently, 20 HMRs were placed in 1.0 mL of normal saline as the experimental group (normal saline without HMRs was used as the negative control group, and deionized water was used as the positive control group), and then 20 μL of red blood cell suspension was added, and the resulting mixture was rotated and incubated at 37 °C for 10 min. After incubation, the above mixture was centrifuged at 3000 r / min for 10 min, and then 0.2 mL of supernatant was transferred to a 96-well plate, and the absorbance at 545 nm was measured three times with a microplate reader. The hemolysis rate calculation formula is: Hemolysis rate (%) = (A S -A N ) / (A P -A N )×100% (where A S is the absorbance of the experimental sample; A N is the absorbance of the negative control group; A P is the absorbance of the positive control group), such as Figure 3 As shown in (b), the morphology of red blood cells incubated with HMRs in the experimental group had no significant difference from that of normal saline, and the hemolysis rate of HMRs was 0.55%, which was far lower than 5% (the criterion for excellent blood compatibility).
[0045] The morphology of red blood cells in the experimental group and the negative control group before centrifugation was observed under an inverted fluorescence microscope. Figure 3 (c) It can be seen that they hardly cause red blood cell rupture after contact with blood, demonstrating their good blood compatibility.
[0046] Example 2 The magnetically enhanced light-driven HMRs prepared in Example 1 were used to observe the motion of the magnetically enhanced light-driven HMRs in different solutions and record their speed. Using a blue laser as the light source, the HMRs were placed in a 20 mm culture dish containing 300 μL of a 65 mmol / L glucose biofluid (the biofluid was one of water, urine, bile, and whole blood, with a depth of approximately 3 mm). Then, a permanent magnet (2 cm × 5 cm) and a blue laser (38 mW / mm 2 ) were fixed above the culture dish; the permanent magnet was 2.5 cm vertically above the culture dish liquid surface, the blue laser was 7.5 cm above the culture dish liquid surface, and the angle between the light source and the culture dish liquid surface was 45°. Videos of the light-driven movement of HMRs were recorded at 80 frames per second using a Nikon inverted fluorescence microscope and edited using the Jianying software to obtain the movement trajectory of the HMRs and measure their speed (e.g. Figure 4 ), the movement trajectory was plotted using delayed images per second, and its movement speeds in water, urine, bile, and whole blood were 217.1, 203.9, 176.4, and 92.5 μm / s, respectively, demonstrating its excellent movement ability in complex biological fluids.
[0047] Example 3 The magnetically enhanced light-driven HMRs prepared in Example 1 were used to detect PFOS in whole blood. Using a blue laser as the light source, differential pulse voltammetry (DPV) curves were measured using an electrochemical workstation. The magnetically enhanced light-driven HMRs were placed in whole blood containing varying concentrations of PFOS (1 mL of whole blood was added with 1 to 5 μL of a high-concentration PFOS aqueous solution to prepare the concentrations listed in Table 1). PFOS was identified for 5 minutes using the magnetically enhanced light-driven motion. The HMRs were then collected using a magnetic glassy carbon electrode and placed in a 0.1 mol / L potassium chloride solution (containing 5 mmol / L potassium ferrocyanide and 5 mmol / L potassium ferrocyanide) to measure the DPV curves. The response signals of the magnetically enhanced light-driven HMRs to a series of PFOS concentrations are shown in Table 1.
[0048] Analysis of the data in Table 1 shows that the response of the magnetically enhanced light-driven HMRs to the PFOS series concentration increases with the increase of concentration (C), and the generated differential pulse voltammetry peak current signal (I) gradually decreases; linear fitting finds that there is a linear relationship between the peak current change value ΔI (the difference between the I of the target and the I of the blank background) and the logarithm of the PFOS concentration (Appendix Figure 5 ), the regression equation is: ΔI = 36.857 + 9.277 lg C (R 2=0.991)(ΔI: μA; C: pg / mL), with a linear range of 0.1 fg / mL to 1 ng / mL and a detection sensitivity of 0.134 fg / mL, enabling ultrasensitive and rapid detection of PFOS.
[0049] Example 4 The magnetically enhanced light-driven HMRs prepared in Example 1 were used to directly detect PFOS in rat whole blood. The specific process was as follows: Before whole blood testing, 7- to 8-week-old male Sprague-Dawley rats were housed in a controlled environment with a temperature of 20-26°C and a humidity of 35-70%. The rats were randomly divided into four groups. Before oral administration of PFOS, the rats were acclimated for one week and then orally administered 500 μL of PFOS solution daily at concentrations of 0 (control group), 0.5 ng / L, 20 ng / L, and 200 μg / L, respectively. The PFOS solution was dissolved in 2% Tween 20. PFOS solution was administered orally once daily for 7 days. Whole blood was collected 24 hours after the last oral administration of PFOS solution for PFOS analysis. Using a blue laser as the light source, 20 HMRs were placed in a culture dish containing 300 μL of whole blood (containing 65 mM glucose). A permanent magnet was fixed above the HMRs, and the excitation light source was fixed at a 45-degree angle. The light source was turned on to induce magnetically enhanced optical motion of the HMRs for 5 minutes. Finally, the HMRs were collected using a magnetic glassy carbon electrode and detected by differential pulse voltammetry.
[0050] The concentration of PFOS in the whole blood of rats exposed to different concentrations of PFOS for different time periods was monitored (e.g. Figure 6 ), the results showed that the concentrations of PFOS in the whole blood of rats that had been orally administered with 0.5 ng / L PFOS per day were 0.234, 3.413, 13.043 and 46.203 pg / mL, respectively, 1, 3, 5 and 7 days later; the concentrations of PFOS in the whole blood of rats that had been orally administered with 20 ng / L PFOS per day were 0.344, 13.464, 64.071 and 198.738 pg / mL, respectively, 1, 3, 5 and 7 days later; the concentrations of PFOS in the whole blood of rats that had been orally administered with 200 μg / L PFOS per day were 0.253, 84.473, 634.034 and 2487.991 ng / mL, respectively, 1, 3, 5 and 7 days later. This showed that the enrichment of PFOS in rat whole blood was positively correlated with the oral dose and number of days of oral administration, confirming that the sensor constructed by HMRs can quickly track the bioaccumulation level of persistent organic pollutants in living blood.
[0051] Finally, the PFOS concentration in the whole blood of rats that had orally administered 200 μg / L PFOS was determined by high performance liquid chromatography (HPLC) and compared with the results of HMRs. The results of PFOS in the whole blood after 3 and 7 days of oral administration were as follows: 3 days: 88.503 ng / mL (HPLC) and 84.473 ng / mL (HMRs); 7 days: 2370.302 ng / mL (HPLC) and 2487.991 ng / mL (HMRs). The detection results of the two methods were relatively close (e.g. Figure 7 ), demonstrating the reliability of the magnetically enhanced light-driven HMRs described in the present invention in detecting complex biological fluids.
[0052] Based on the disclosure and teachings of the above description, those skilled in the art may also make changes and modifications to the above embodiments. Therefore, the present invention is not limited to the specific embodiments disclosed and described above, and some modifications and variations of the invention should also fall within the scope of protection of the claims of the present invention. In addition, although certain specific terms are used in this description, these terms are only for convenience of description and do not constitute any limitation on the scope of the present invention.
Claims
1. A magnetically enhanced light-driven hydrogel microrobot, characterized in that: It includes hydrogel droplets and a hydrogel coating; wherein the hydrogel droplets are hydrogels containing magnetic material, light-driven material and molecular imprinting material, and the light-driven material is bismuth vanadate-carbon nanotubes; the hydrogel coating is a calcium alginate gel layer.
2. The magnetically enhanced light-driven hydrogel microrobot according to claim 1, characterized in that: The bismuth vanadate-carbon nanotubes are formed by adding carbon nanotubes during the synthesis of bismuth vanadate; the molecularly imprinted material directly uses molecularly imprinted polymers; or the molecularly imprinted material is formed by synthesizing molecularly imprinted polymers using bismuth vanadate-carbon nanotubes as a substrate.
3. The magnetically enhanced light-driven hydrogel microrobot according to claim 1, characterized in that: The water content of the hydrogel droplets is 85% to 95%; in the hydrogel droplets, the mass ratio of the magnetic material, the light-driven material and the molecular imprinting material is 1: (2 to 4): (2 to 4).
4. The magnetically enhanced light-driven hydrogel microrobot according to claim 1, characterized in that: The hydrogel droplets are in the shape of water droplets, with radial dimensions ranging from 450 μm to 1200 μm and axial dimensions ranging from 300 μm to 900 μm; the thickness of the hydrogel coating is 4 to 12 μm.
5. The method for preparing a magnetically enhanced light-driven hydrogel microrobot according to claim 1, characterized in that: The steps include: (1) Mixing the magnetic material, the light-driven material, and the molecularly imprinted material in a sodium alginate solution and dispersing them uniformly to obtain a precursor solution; (2) The precursor solution is injected into the calcium chloride solution using a microfluidic syringe to form water droplet-shaped hydrogel droplets; (3) The above-mentioned hydrogel droplets are placed on a nylon mesh, and sodium alginate solution and calcium chloride solution are added dropwise in sequence to form a calcium alginate gel layer on the surface of the hydrogel droplets, thereby obtaining a magnetically enhanced light-driven hydrogel microrobot.
6. The method for preparing a magnetically enhanced light-driven hydrogel microrobot according to claim 5, characterized in that: In step (1), the concentration of the sodium alginate solution is 1.5 to 2.5 wt%; the concentrations of the magnetic material, the light-driven material, and the molecular imprinting material in the sodium alginate solution are 2 to 4 mg / mL, 6 to 12 mg / mL, and 6 to 12 mg / mL, respectively; the magnetic material is one of magnetic ferric oxide and ferroferric oxide, or a mixture of two or more thereof; In step (3), the pore size of the nylon mesh is 60 to 100 mesh, the concentration of the sodium alginate solution is 0.2% to 1.2 wt%, and the concentration of the calcium chloride solution is 0.3% to 2 wt%.
7. The method for preparing a magnetically enhanced light-driven hydrogel microrobot according to claim 5, characterized in that: In step (2), the concentration of the calcium chloride solution is 0.2 ~ 0.8 wt%; the vertical height of the microfluidic syringe from the liquid surface of the calcium chloride solution is 0.3 ~ 0.9 mm, the flow rate of the microfluidic injection is 0.01 ~ 0.1 mL / min, and the diameter of the microfluidic injection needle is 0.2 ~ 0.8 mm; the radial size of the hydrogel droplet is 450 μm ~ 1200 μm, and the axial size is 300 μm ~ 900 μm.
8. Use of the magnetically enhanced light-driven hydrogel microrobot according to claim 1 in detecting pollutants in biological fluids, characterized in that: The specific application method is as follows: the magnetically enhanced light-driven hydrogel microrobot is mixed with the biological fluid to be detected, fuel is added, a magnet is fixed above the biological fluid, and an excitation light source is provided to irradiate the hydrogel microrobot, so that the hydrogel microrobot undergoes magnetically enhanced light-driven movement in the biological fluid and specifically binds to the target pollutant; a magnetic glassy carbon electrode is then used to adsorb the hydrogel microrobot from the biological fluid, and then the hydrogel microrobot is placed in a redox probe electrolyte solution to measure the differential pulse voltammetry curve, and then the content of the target pollutant in the biological fluid to be detected is determined using a standard curve method; wherein, the molecular imprinting material in the magnetically enhanced light-driven hydrogel microrobot can specifically identify the target pollutant.
9. The use of the magnetically enhanced light-driven hydrogel microrobot in detecting pollutants in biological fluids according to claim 8, characterized in that: The fuel is added to the biological fluid at a concentration of 55 to 75 mmol / L, and the fuel includes one or a mixture of glucose, uric acid, and malic acid in any proportion; The magnet is fixed just above the surface of the biological fluid, with the vertical distance between the magnet and the liquid surface being 1 to 4 cm, and the depth of the biological fluid being 1 to 5 mm; the vertical distance between the excitation light source and the liquid surface is 5 to 10 cm.
10. The use of the magnetically enhanced light-driven hydrogel microrobot in detecting pollutants in biological fluids according to claim 8, characterized in that: The biological fluids include, but are not limited to, whole blood, bile, and urine, and the target pollutants include, but are not limited to, perfluorooctane sulfonic acid, polychlorinated biphenyls, and pentachlorophenol.