CN106983517B - A kind of no enzyme nanometer flexibility blood glucose microsensor and preparation method thereof - Google Patents

Abstract

The invention discloses an enzyme-free nanometer flexible blood sugar microsensor and a preparation method thereof, comprising: a flexible substrate, an electrode structure, a sensing layer and an ion-selective hydrophilic protective layer. The flexible substrate has good temperature adaptability (-10-300°C), the electrode structure has excellent electrical conductivity and is resistant to acid and alkali corrosion, and the glucose detection microchannel (channel length less than 10mm, width 10μm-500μm) is composed of nanomaterial texture , good flexibility, high sensitivity, linear correlation in the range of glucose concentration (1-28mmol/L), good reversibility and repeatability, hydrophilic wear-resistant film has good ion permeability and strong water absorption characteristics and protect the sensing layer. The blood glucose sensor of the present invention is non-toxic and harmless, comfortable and non-allergic when in contact with human skin, and its overall structure does not exceed 1cm ² , and can realize in-situ real-time measurement and analysis of human blood glucose (1‑28mmol/L) through current response.

Description

A kind of enzyme-free nano flexible blood glucose microsensor and preparation method thereof

technical field

The invention belongs to the field of preparation of electrochemical sensors, in particular to a nano-flexible blood sugar sensor and a preparation method thereof, which are used to realize in-situ real-time detection and analysis of human blood sugar, and ensure that the sensor is suitable for long-term wear and stable work of the human body, and can Realize non-invasive or minimally invasive blood sugar monitoring, and meet the wide-range blood sugar monitoring of diabetes.

Background technique

Blood sugar is closely related to human health. Long-term high blood sugar can cause various complications (such as uremia, limb necrosis, blindness, etc.), while low blood sugar can cause fainting, palpitation and weakness. Therefore, blood sugar testing is of great significance to human health. However, traditional blood glucose testing equipment is bulky and time-consuming, and cannot achieve real-time and accurate detection, which limits its promotion and use in families. The huge market demand has prompted researchers to develop an efficient, cheap, accurate and portable blood glucose testing device.

The existing non-optical blood glucose sensor research mainly focuses on material modification, surface modification, composite electrodes and so on. Most glucose sensors based on resistive or potentiometric responses require glucose oxidase to convert H ⁺ ions for detection. Enzyme-free blood glucose sensors have attracted people's attention and attention in recent years. In the paper "3D nanoporous goldscaffold supported on graphene paper: Freestanding and flexible electrode with high loading of ultrafine PtCo alloy nanoparticles for electrochemical glucose sensing", Zhao et al. loaded 3D nanoporous gold scaffolds on graphene paper, and then loaded platinum-cobalt ultrafine-grained particles Assembling on a scaffold to prepare a glucose sensor, the preparation process is complicated and the cost is high, and the test needs to be carried out in an alkaline environment. In the paper "Doped graphene/Cu nanocomposite: A high sensitivity non-enzymatic glucose sensor for food", Shabnam et al. used a one-pot method to prepare Cu-NGr materials by mixing copper nanoparticles with nitrogen-doped graphene to detect glucose. Three-electrode method Testing is more complicated. However, the existing blood glucose sensors and sensing materials have the following series of problems: 1. Low utility, complex structure design, large volume, and difficult to realize portable and wearable functions; 2. The process is cumbersome, the preparation cost is too high, the service life is short, High cost; 3. The detection accuracy is low, and it is difficult to realize in-situ real-time monitoring; 4. The blood sugar response range is narrow, and it is difficult to meet the monitoring of a wide range of blood sugar concentrations for diabetics; 5. Due to the stability and accuracy of the enzyme-free blood sugar sensor, the current There are currently no related market products.

Contents of the invention

Aiming at the above-mentioned problems in actual use, the present invention designs a nano-flexible blood glucose sensor and its preparation method. Through the combination of nano-wire sensing micro-channel structure and hydrophilic protective layer, micro-analysis and rapid monitoring can be realized, and It can meet the requirements of high efficiency, low cost, small volume and high integration of the sensor. At the same time, various necessary materials that are friendly to the skin are used to prepare the sensor, which avoids human discomfort and material damage caused by the contact between the active layer and the skin. The test environment is simple, and blood sugar changes are analyzed in real time through current response, the process is simple, and the cost is low. The enzyme-free sensor improves the service life of the sensor and reduces the cost of use.

The present invention achieves the above-mentioned technical purpose through the following technical means.

A method for preparing an enzyme-free nano-flexible blood glucose microsensor, characterized in that it comprises the following steps:

Step 1: Carbon nanotube film formation

Weigh carbon nanotubes according to the concentration of 0.1-0.3 mg/mL, place them in a mixed acid with a volume ratio of 3:1 mass fraction of 95-98% concentrated sulfuric acid and 65-68% mass fraction of concentrated nitric acid, and stir at 110°C 45-90min, centrifuge to remove the supernatant, adjust the pH to neutral; lay flat at 30-60°C and evaporate to dryness to obtain carboxylated carbon nanotube film;

Step 2: Preparation of glucose sensing material

Mix Ga ₂ O ₃ powder and carbon powder at a molar ratio of 1:3, grind for more than 5 minutes to obtain a precursor powder, use the film obtained in step 1 as a substrate, and plate a 5nm-30nm metal catalyst film; the precursor powder and The substrate is prepared in a plasma-enhanced chemical vapor deposition system PECVD: the mass ratio of the precursor powder to the film is 3:1-15:1, N ₂ and H ₂ are used as reaction gases, the pressure is 30-120Pa; the substrate temperature is 500 ℃-1200℃; RF power supply 40-100W; N ₂ and H ₂ gas mixture, N ₂ and H ₂ gas flow rate is 10 cm ³ /min-100 cm ³ /min, the reaction time is 45-120min;

Step 3: Flexible substrate pretreatment

Process the flexible substrate material in a plasma-enhanced chemical vapor deposition system: use hydrogen or oxygen reaction pressure of 20-120Pa, radio frequency power of 20-80w, and treat the flexible substrate at room temperature so that the contact angle with the liquid is less than 5°; The final substrate is hang-coated with a thickness of 1-3 μm epoxy resin;

Step 4: Electrode and sensing microchannel design

Electrode templates are made on the pretreated flexible substrate through ultraviolet exposure, and metal electrodes with a distance of 1-10 μm are deposited by electron beam evaporation, and the electrode leads at both ends are drawn out by pressure welding or silver glue; the microchannel electrode layer is prepared , design the length of the microchannel to be less than 1-6cm, and the width to be 3-100μm, and introduce the sensing material into the microchannel;

Step 5: Hydrophilic protective layer

Treat the vinylidene fluoride PVDF film with a pore size of 0.01-0.5 microns in a plasma-enhanced chemical vapor deposition system: use hydrogen or oxygen reaction pressure of 20-120Pa, radio frequency power of 20-80w, and treat at room temperature to make the contact angle with the liquid Less than 5°, as an ion-selective hydrophilic protective layer, pasted on the electrode and sensing material to protect the electrode material and sensing material while ensuring the penetration of glucose molecules.

The order of step 2 and step 3 can be interchanged successively.

Further, magnetron sputtering and evaporation, the film thickness is controlled at 100-250nm.

Further, the electrode adopts Ti/Au electrode, UV exposure parameters: AZ5214 adhesive is pre-baked at 110°C for 1-15min, exposed for 0.1-20s, post-baked at 120°C for 1-5min, exposed for 10-15s, developed for 0.1-5min; electron beam evaporation Parameters: vacuum degree 5×10 ^{- 6} mbar, Ti/Au thickness 10/100nm respectively.

The prepared nano-flexible blood glucose microsensor is used for linear response to the concentration of 1-28mmol/L blood glucose.

An enzyme-free nano-flexible blood glucose sensor and a preparation method thereof, comprising: a flexible substrate, an electrode structure, a glucose detection microchannel, and an ion-penetrating hydrophilic protective layer. Using anti-bending flexible polymer as the substrate, the electrode micro-channel structure is designed by the template method, the nanowire texture material is transferred into the micro-channel as the sensitive layer, and the electrode and the sensitive layer are covered by ions through the hydrophilic wear-resistant layer. Glucose absorbs quickly and avoids direct contact with the human body.

The flexible substrate is made of non-toxic, harmless and anti-bending polymers such as polyimide (PI), etc., and the hydrophilicity (contact angle<5°) is enhanced through plasma treatment.

The thickness of the flexible substrate is 50-200 μm, and the overall area is less than 1 cm ² .

The material of the sensitive layer is selected to introduce the texture of carboxyl-functionalized carbon nanotubes grown with gallium nitride nanowires into the microchannel by using micro-nano-fabrication in the textured nanowire microchannel as the in-situ sensing region material for glucose molecules.

The micro-channel structure of the electrode is made by template method, the length of the micro-channel is less than 5 mm, and the width is 3-100 μm.

An enzyme-free nano-flexible blood glucose microsensor and a preparation method thereof, comprising the following steps:

(1) Selection and surface treatment of flexible substrates: The selection of flexible substrate materials includes PI, etc., which have good mechanical and temperature adaptability, and the surface treatment enhances the adhesion properties of electrodes, sensing channels and hydrophilic films. A flexible substrate 1 is produced.

(2) Electrode micro-channel structure design and production: The structural size can be designed according to the device testing and integration requirements. In order to ensure good electrical conductivity of the electrode and save costs, through the most structural design, Au, Pt, etc. are selected to resist acid and alkali corrosion. Metal material with good chemical stability. The electrode microchannel 2 is fabricated by photolithography.

(3) Fabrication of nano-textured glucose sensitive layer. The carboxylated carbon nanotube texture with gallium nitride nanowires grown by PECVD is introduced into the microchannel as the in-situ sensing region material for glucose molecules, and the glucose molecule sensitive layer 3 is prepared.

(4) The ions are treated and packaged through the hydrophilic protective layer, and the treated hydrophilic wear-resistant layer is surface-mounted. The overall area is larger than the size of the substrate, and the electrode and the sensitive layer are completely covered to avoid friction, ensuring that the electrode layer is in contact with the sensitive layer. The material will not fall off after repeated use for a long time. Ions permeate through the hydrophilic and wear-resistant layer to form a porous hydrophilic polymer film, which is treated by oxygen plasma, with a pore size of 0.1-1 μm and a thickness of no more than 0.1 mm.

Beneficial effects of the present invention:

(1) An enzyme-free nano-flexible glucose sensor and its preparation method provided by the present invention, the combination of nanowire texture sensing microchannel and ion-selective hydrophilic wear-resistant coating is designed by template method to realize the small volume of the device It is highly integrated and fully protects the active layer of the device to prevent peeling and failure due to repeated use. GaN forms a heterojunction with the carboxylated carbon tube film, which produces a linear current response to glucose molecules,

(2) An enzyme-free nano-flexible glucose sensor and its preparation method according to the present invention avoid direct contact between sensitive materials and electrode materials and the measured body through the use of ion permeation hydrophilic wear-resistant layer, and can be applied to Wearing the bio-glucose sensor can eliminate the allergic and uncomfortable reactions caused by the direct contact between the material and the human body, and at the same time protect the sensitive layer from damage and work accurately for a long time. At the same time, the whole material does not contain glucose oxidase, which avoids the test error caused by the activity of the enzyme being changed due to the influence of temperature, acidity and alkalinity.

(3) The test principle of the present invention is based on the current response caused by resistance changes, and the test environment requires low requirements. The sensor responds linearly to blood sugar with a concentration of 1-28mmol/L (the normal blood sugar concentration of the human body is 3.61-6.11mmol/L).

Description of drawings

Figure 1 is the structure diagram of the sensor (1 is the flexible substrate, 2 is the electrode layer, 3 is the position of the interdigital electrode, that is, the position of the sensing material, and 4 is the protective layer)

Figure 2 is an enlarged schematic diagram of 3 in the sensor structure diagram, and a schematic diagram of the electrode microchannel structure made by the mask plate

Figure 3 shows the growth of GaN nanowires on carbon tube films

Fig. 4 is embodiment 1 response curve

Fig. 5 is embodiment 2 response curves

Figure 6 The influence of bending test on the sensitivity

Detailed ways

Example 1

(1) The mixture of analytically pure concentrated nitric acid (68% by mass fraction) and analytically pure concentrated sulfuric acid (98% by mass) with a volume ratio of 3:1 is used as the acidifying liquid, and carboxyl groups are introduced into single-walled carbon nanotubes, Stir at 110°C for 90 minutes, centrifuge in a high-speed centrifuge at 9,500 rad/min, filter off the supernatant, adjust the pH of the carbon tubes to neutral, and evaporate the water to dryness at 50°C to obtain a single-walled carboxyl-functionalized carbon tube film; Mix Ga ₂ O ₃ powder with carbon powder (molar ratio 1:3), and grind for 5 minutes to obtain a precursor powder: the carboxylated carbon tube film is used as a substrate, and 10nm Au is plated as a catalyst film; the precursor powder and the substrate The substrate is prepared in a plasma-enhanced chemical vapor deposition system: the mass ratio of the precursor powder to the film is 10:1, the mixed gas of N ₂ and H ₂ is used as the reaction gas, the pressure is 90Pa; the substrate temperature is 700°C; the RF power supply is 80W ; The N ₂ gas flow rate is 80 cm ³ /min, the H ₂ gas flow rate is 50 cm ³ /min, and the reaction is 65 min. A sensing material is produced that is the same size as the microchannel.

(2) Treat the 100μm flexible PI substrate in oxygen plasma, specific parameters: vacuum degree 0.1Pa, working pressure 20Pa, working gas O ₂ , flow rate 20sccm, RF power 60w, room temperature treatment for 5 minutes so that the contact angle between the flexible substrate and the liquid is less than 5 °, hang-coat 1 μm epoxy resin on the pre-treated substrate; then cut it into 1×1cm ² pieces to make a flexible substrate.

(3) Design the photolithography template as shown in Figure 1, make electrode templates on the treated PI sheet through UV exposure, and deposit Ti/Au electrodes with a spacing of 3 μm by electron beam evaporation. , Expose for 1s, bake at 120°C for 2min, expose for 20s, develop for 1min. Electron beam evaporation parameters: vacuum degree 5×10 ^-6 mbar, Ti/Au thicknesses respectively 10/100nm. The electrodes at both ends are led out by pressure welding or silver glue. The length of the micro-channel is designed to be 1 cm, and the width is 10 μm to prepare a micro-channel electrode layer with a thickness of 150 nm. The sensing material prepared in step (1) is introduced into the channel.

(4) The PVDF microporous wear-resistant layer with a thickness of 100 μm and a pore diameter of 0.47 μm is subjected to surface oxygen plasma treatment to increase hydrophilicity. Specific parameters: vacuum degree 0.1Pa, working pressure 20Pa, working gas O ₂ , flow rate 20sccm, radio frequency power 60w, 5min at room temperature. Then it is cut into 1×1cm ² pieces, and mounted on the electrode and the sensing material, so that ions can pass through the hydrophilic and wear-resistant layer, so that the contact angle with the liquid is less than 5°;

(5) The PI substrate can be used below 400°C. The sensor is sensitive to glucose solution, and the response curve is close to linear. Take 3 μL of glucose solution with a concentration of 3mmol/L, 4mmol/L, 5mmol/L, and 6mmol/L respectively. Test its sensitive properties. The test data is shown in Figure 4, the response time is less than 1s, and the recovery of the device is tested, and the recovery rate is greater than 90%.

Example 2

(1) The mixture of analytically pure concentrated nitric acid (68% by mass fraction) and analytically pure concentrated sulfuric acid (98% by mass) with a volume ratio of 3:1 is used as the acidifying liquid, and carboxyl groups are introduced into single-walled carbon nanotubes, Stir at 110°C for 90 minutes, centrifuge in a high-speed centrifuge at 9,500 rad/min, filter off the supernatant, adjust the pH of the carbon tubes to neutral, and evaporate the water to dryness at 50°C to obtain a single-walled carboxyl-functionalized carbon tube film; Mix Ga ₂ O ₃ powder with carbon powder (molar ratio 1:3), and grind for 4 minutes to obtain a precursor powder: the carboxylated carbon tube film is used as a substrate, and 20nm Au is plated as a catalyst film; the precursor powder and the substrate The substrate was prepared in a plasma-enhanced chemical vapor deposition system: the mass ratio of the precursor powder to the film was 5:1, N ₂ and H ₂ were used as reaction gases, and the pressure was 60Pa; the substrate temperature was 500°C; the RF power supply was 80W; N ₂ The gas flow rate is 60 cm ³ /min, the H ₂ gas flow rate is 50 cm ³ /min, and the reaction time is 75 min. A sensing material is produced that is the same size as the microchannel.

(2) Treat the 100μm flexible PI substrate in oxygen plasma, specific parameters: vacuum degree 0.1Pa, working pressure 20Pa, working gas O ₂ , flow rate 20sccm, RF power 60w, room temperature treatment for 5 minutes so that the contact angle between the flexible substrate and the liquid is less than 5 °, hang-coat 1 μm epoxy resin on the pretreated substrate; then cut it into 1×1cm ² square pieces to make flexible substrates.

(3) Design the photolithography template as shown in Figure 1, make electrode templates on the processed PI sheet through ultraviolet exposure, and deposit Ti/Au electrodes with a spacing of 6 μm by electron beam evaporation. , Expose for 1s, bake at 120°C for 2min, expose for 20s, develop for 1min. Electron beam evaporation parameters: vacuum degree 5×10 ^-6 mbar, Ti/Au thicknesses respectively 10/100nm. The electrodes at both ends are led out by pressure welding or silver glue. A microchannel electrode layer was prepared, the thickness of the electrode layer was 200 nm, the length was 2 cm, and the width was 10 μm. The sensing material prepared in step (1) is introduced into the channel.

(4) The PVDF microporous wear-resistant layer with a thickness of 100 μm and a pore diameter of 0.47 μm is subjected to surface oxygen plasma treatment to increase hydrophilicity. Specific parameters: vacuum degree 0.1Pa, working pressure 20Pa, working gas O ₂ , flow rate 20sccm, radio frequency power 60w, 5min at room temperature. Then it is cut into 1×1cm ² pieces, and mounted on the electrode and the sensing material, so that ions can pass through the hydrophilic and wear-resistant layer, so that the contact angle with the liquid is less than 5°;

(5) The PI substrate can be used below 400°C. The sensor is sensitive to glucose solution, and the response curve is close to linear. Take 3 μL of glucose solution with a concentration of 2mmol/L, 4mmol/L, 6mmol/L, and 8mmol/L respectively. Test its sensitive properties. The test data is shown in Figure 5, the response time is less than 1s, and the recovery of the device is tested, and the recovery rate is greater than 90%.

Claims (5)

1. A method for preparing an enzyme-free nanometer flexible blood glucose microsensor, characterized in that, comprising the following steps: Step 1: Carbon nanotube film formation Weigh carbon nanotubes according to the concentration of 0.1-0.3 mg/mL, place them in a mixed acid with a volume ratio of 3:1 mass fraction of 95-98% concentrated sulfuric acid and 65-68% mass fraction of concentrated nitric acid, and stir at 110°C 45-90min, centrifuge to remove the supernatant, adjust the pH to neutral; lay flat at 30-60°C and evaporate to dryness to obtain carboxylated carbon nanotube film; Step 2: Preparation of glucose sensing material Mix Ga ₂ O ₃ powder and carbon powder at a molar ratio of 1:3, grind for more than 5 minutes to obtain a precursor powder, use the film obtained in step 1 as a substrate, and plate a 5nm-30nm metal catalyst film; the precursor powder and The substrate is prepared in a plasma-enhanced chemical vapor deposition system PECVD: the mass ratio of the precursor powder to the film is 3:1-15:1, N ₂ and H ₂ are used as reaction gases, the pressure is 30-120Pa; the substrate temperature is 500 ℃-1200℃; RF power supply 40-100W; N ₂ and H ₂ gas mixture, N ₂ and H ₂ gas flow rate is 10 cm ³ /min-100 cm ³ /min, the reaction time is 45-120min; Step 3: Flexible substrate pretreatment Process the flexible substrate material in a plasma-enhanced chemical vapor deposition system: use hydrogen or oxygen to react, the air pressure is 20-120Pa, the radio frequency power is 20-80w, and the flexible substrate is processed at room temperature so that the contact angle with the liquid is less than 5°; Hang-coat epoxy resin with a thickness of 1-3 μm on the treated substrate; Step 4: Electrode and sensing microchannel design Electrode templates are made on the pretreated flexible substrate by ultraviolet exposure, and metal electrodes with an interval of 1-10 μm are plated by magnetron sputtering or evaporation, and the electrodes at both ends are led out by pressure welding or silver glue; Microchannel electrode layer, the designed microchannel length is less than 6cm, width 3-100μm, and the sensing material is introduced into the microchannel; Step 5: Hydrophilic protective layer Process the vinylidene fluoride PVDF film with a pore size of 0.01-0.5 microns in a plasma-enhanced chemical vapor deposition system: use hydrogen or oxygen to react, the pressure is 20-120Pa, the radio frequency power is 20-80w, and it is treated at room temperature to make it contact with the liquid The angle is less than 5°. As an ion-selective hydrophilic protective layer, it is pasted on the electrode and sensing material to protect the electrode material and sensing material while ensuring the penetration of glucose molecules. 2. The preparation method according to claim 1, in step 4, the thickness of the metal electrode is controlled at 100-250nm. 3. The preparation method according to claim 1, the order of step 2 and step 3 can be interchanged successively. 4. The preparation method according to claim 1, characterized in that the electrodes are Ti/Au electrodes, and the UV exposure parameters: AZ5214 glue is pre-baked at 110°C for 1-15min, exposed for 0.1-20s, and then baked at 120°C for 1-5min. Expose for 10-15s, develop for 0.1-5min; the vapor deposition is electron beam vapor deposition, electron beam vapor deposition parameters: vacuum degree 5×10 ^-6 mbar, Ti/Au thickness 10/100nm respectively. 5. The nano-flexible blood glucose microsensor prepared by the preparation method according to any one of claims 1-4, which is used for linearly responding to a concentration of 1-28mmol/L blood glucose.