Device for quantitative assessment of glucose content in body fluids

Abstract

The invention relates to the field of medicine, namely to biosensor analytical devices, and can be used for quantitative assessment of glucose content in body fluids, namely blood, plasma and serum The device for quantitative assessment of glucose content in body fluids containing a graphite screen printed electrode (1), a surface of which is modified by a hybrid composite (2) having inclusions of glucose oxidase enzyme (4), and the hybrid composite (2) contains an organosilicon matrix (3) made of tetraethoxysilane, and placed in said matrix (3) inclusions of single-walled carbon nanotubes (5), inclusions of a redox-active polymer (6) based on chitosan covalently bound to a Nile blue mediator. The technical effect of the invention is an increase in the operating time of the device for quantitative assessment of glucose content in body fluids without replacing the hybrid composite while maintaining high sensitivity to glucose content in body fluids.

Description

[0001] DEVICE FOR QUANTITATIVE ASSESSMENT OF GEUCOSE CONTENT IN BODY

[0002] FEUIDS

[0003] DESCRIPTION

[0004] TECHNICAE FIELD

[0005] The invention relates to the medicine, namely to biosensor analytical devices (biosensors) and can be used for quantitative assessment of glucose content in body fluids, namely blood, plasma and serum.

[0006] BRIEF DESCRIPTION OF THE PRIOR ART

[0007] This is a known device for quantitative assessment of glucose content, which is functionalized graphene on a screen-printed electrode coated with gold sputtering and the glucose oxidase enzyme [Akhtar M. A. et al. Functionalized graphene oxide bridging between enzyme and Au- sputtered screen-printed interface for glucose detection / / ACS Applied Nano Materials, 2019 vol. 2, no. 3, pp. 1589-1596]. A characteristic feature of the device in question, which is common with the claimed invention, is an amperometric method for measuring glucose levels using the glucose oxide enzyme on a screen-printed electrode. A major disadvantage of this device is its low sensitivity which is 3.1732 pA.dm3 / mmol.com2.

[0008] The closest prior art of the claimed invention is a device for quantitative assessment of glucose content in liquids, which is disclosed in the Patent RU218094 dated 11.05.2023. The device contains a graphite screen-printed electrode modified with a hybrid composite. The glucose oxidase enzyme is used as a biorecognition element. A conductive polymer based on thermally expanded graphite and an electropolymerized neutral red mediator is used as a hybrid composite. The device provides quantitative assessment of glucose content ranging from 0.006 to 0.5 mM (mmol / dm3) of glucose, which is insufficient in some cases and operates without replacing the hybrid composite for up to 30 days.

[0009] Thus, the claimed invention is aimed at solving the technical problem of insufficient operating time of known devices for quantitative assessment of glucose content in body fluids without replacing the hybrid composite.

[0010] SUMMARY OF THE INVENTION The technical effect of the invention is an increase in the operating time of the device for quantitative assessment of glucose content in body fluids without replacing the hybrid composite while maintaining high sensitivity to glucose content in body fluids.

[0011] The sensitivity indicators of the device should be understood as the sensitivity factor, as well as the range of glucose concentrations that can be detected.

[0012] This technical effect is achieved in a device for quantitative assessment of glucose content in body fluids due to the fact that it contains a graphite screen-printed electrode, a surface of which is modified with a hybrid composite having inclusions of a glucose oxidase enzyme. The hybrid composite material contains an organosilicon matrix made of tetraethoxysilane and placed in said matrix inclusions of single-walled carbon nanotubes and inclusions of a redox-active polymer based on chitosan covalently bound to a Nile blue mediator.

[0013] In a particular embodiment of the invention:

[0014] - the hybrid composite is prepared by mixing tetraethoxysilane, glucose oxidase enzyme, sodium fluoride catalyst (NaF), a solution of chitosan in acetic acid, a suspension of single-walled carbon nanotubes, a saturated solution of the Nile blue mediator and a glutaraldehyde solution, applying the resulting mixture to the surface of the graphite screen-printed electrode and subsequent drying.

[0015] The tetraethoxysilane organosilicon matrix containing inclusions of single-walled carbon nanotubes and inclusions of the redox-active polymer based on chitosan, covalently bound to the Nile blue mediator, used for immobilization of the glucose oxidase enzyme, creates a stable biocompatible environment that maintains the oxidative activity of this enzyme at a high level. Tetraethoxy silane used to form the silicon-containing matrix, is chemically inert, which provides a robust porous rigid structure that prevents the biorecognition component in the form of the glucose oxidase enzyme from being washed out from the electrode surface, thus increasing the operating time of the device without replacing the hybrid composite.

[0016] The Nile blue mediator covalently cross-linked with chitosan, being an efficient electron carrier, allows to transform a non-conductive matrix into a conductive redox-active polymer and provides an increase in the sensitivity of the device.

[0017] Inclusions of single-walled carbon nanotubes considerably increase the electrical conductivity of the tetraethoxysilane organosilicon matrix due to their large surface area and serve as a "conductive bridge" between the glucose oxidase enzyme molecules and the Nile blue mediator in possible places of spatial remoteness of the molecules from each other, which also contributes to increased sensitivity to glucose content.

[0018] Brief description of drawings

[0019] The invention is explained by the figures, where:

[0020] Figure 1 schematically shows the general view of the claimed device,

[0021] Figure 2 shows a general cross-sectional view of the device,

[0022] Figure 3 shows the recorded signal from the device as current strength (nA) vs. time (s),

[0023] Figure 4 shows the device calibration curve, and

[0024] Figure 5 shows a linear section of the calibration curve.

[0025] The elements are indicated in the figures by the following positions:

[0026] 1. graphite printed electrode,

[0027] 2. hybrid composite,

[0028] 3. organosilicon matrix,

[0029] 4. glucose oxidase enzyme inclusions,

[0030] 5. inclusions of single-walled carbon nanotubes,

[0031] 6. inclusions of a chitosan-based redox-active polymer covalently bound to the Nile blue mediator.

[0032] Embodiment of the invention

[0033] The claimed device includes a graphite screen -printed electrode (1), an operating surface of which is modified (coated) with a hybrid composite material (2). The hybrid composite material (2) contains an organosilicon matrix (3) made of tetraethoxysilane, and placed in said matrix (3) inclusions of a biorecognition component in a form of glucose oxidase enzyme (4), inclusion of single-walled carbon nanotubes (5) and inclusions of a redox-active polymer (6) based on chitosan, covalently bound to a Nile blue mediator.

[0034] An example of obtaining the claimed device is shown below. 50 pL of tetraethoxysilane, 25 pL of glucose oxidase enzyme (titer 25 mg / ml, activity 100 E / mg), 2.5 pl of 1.2M sodium fluoride (NaF) catalyst solution are mixed in an Eppendorf test tube. The obtained mixture is stirred for 5 minutes, then 10 pl of 1% solution of chitosan in 1% acetic acid, 10 pl of 1% suspension of single-walled carbon nanotubes, 5 pl of saturated Nile blue mediator solution, 7.5 pl of 25% glutaraldehyde solution are added and stirred. Then 3 pl of the obtained mixture is applied to the operating surface of the graphite screen-printed electrode (1) and left until completely dry.

[0035] The operating principle of the device for assessment of glucose content in body fluids is as follows.

[0036] A potentiostat, a 5 mL cell, a magnetic stirrer and a portable computer are used for measurements. The device is placed in the cell, 5 mL of sodium-potassium phosphate buffer solution (pH 7.4) is added thereto, the magnetic stirrer is turned on and the background current in the measuring cell is recorded. The measurement results can be recorded with a CS150 potentiostat (Contest, China) connected to graphite screen-printed electrodes and a personal computer. Then the analyzed sample of body fluid is introduced. The measurements are carried out at room temperature and an operating potential of -500 mV due to the redox properties of the hybrid material. After each measurement, the measuring cuvette is rinsed with 5 mL of phosphate buffer solution (pH 7.4).

[0037] The signal of the graphite printed electrode (1) is the cunent (nA) vs. time (s) (see Fig. 3). Next, the amplitude of the change in cunent after introducing the sample into the measuring cuvette is calculated (biosensor response, Al, nA). The glucose content in the sample is determined using the pre-built calibration curve shown in Fig. 4. The glucose content in the cell is calculated using the equation describing the linear section of the calibration curve shown in Fig. 5: y=694-x-27, where x is the concentration of glucose in the cell, mmol / dm3, y is the biosensor response, nA.

[0038] Then the glucose content is determined directly in the sample, taking into consideration the dilution factor. An example of calculations is given in Table 1.

[0039] Table 1. Results of calculating the concentration of glucose in the blood using a device for quantitative assessment of glucose content in body fluids.

[0040] The main features of the claimed device for quantitative assessment of glucose content in body fluids and the closest prior art device are shown in Table 2.

[0041] Table 2. Comparative features of devices for quantitative assessment of glucose content in body fluids

[0042] Thus, the claimed device for quantitative assessment of glucose content in body fluids makes it possible to increase the duration of device operation without replacement of hybrid composite up to 50 days, while featuring a high sensitivity factor, as well as a wide range of detectable glucose concentrations.

Claims

CLAIMS1. A device for quantitative assessment of glucose content in body fluids, containing a graphite screen-printed electrode (1), a surface of which is modified with a hybrid composite (2) having glucose oxidase enzyme inclusions (4), characterized in that the hybrid composite (2) contains an organosilicon matrix (3) made of tetraethoxysilane and placed in said matrix (3) inclusions of single-walled carbon nanotubes (5), inclusions of a redox-active polymer (6) based on chitosan covalently bound to a Nile blue mediator.

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