Glucose sensor based on Wulff type boric acid

Abstract

The present invention relates to a biosensor for measuring glucose concentration and its use in glucose sensing, wherein the biosensor comprises a polymer comprising a moiety of formula (I). The biosensor of the invention is particularly useful in glucose monitoring of individuals under intensive care, as well as in cases where glucose monitoring is performed on unconscious individuals.

Description

[0001] This application claims priority to European patent application EP 23187710.1, filed on 25 July 2023, which is incorporated herein by reference in its entirety.

[0002] This invention relates to biosensors for measuring glucose concentration and their use in glucose sensing. The biosensors of this invention comprise polymers (particularly polymer hydrogels) containing a boric acid-based glucose-binding moiety of formula (I). The biosensors of this invention are particularly useful for glucose monitoring in individuals under intensive care and in cases where glucose monitoring is performed on unconscious individuals. Therefore, this invention relates to the development of glucose sensors based on osmotic pressure measurements, incorporating the boric acid-glucose-binding moiety (GBM) of formula (I), thereby producing glucose-responsive materials with specific sensing properties.

[0003] Beyond glucose sensing applications, there is a growing demand for polymers capable of delivering active agents in a glucose concentration-dependent manner, particularly for treating conditions characterized by pathological glucose concentrations. For example, diabetes is a glucose regulation disorder characterized by the accumulation of glucose in the blood. Glucose regulation disorders can be attributed to the endocrine pancreas's inability to secrete insulin or the body's inability to properly utilize insulin. For type 1 diabetes, the usual treatment involves multiple daily glycemic controls and subcutaneous injections. However, better control of excessive glycemia can be achieved if insulin doses can be continuously adjusted to adapt to blood glucose levels; therefore, glucose levels below or above the normal range that could lead to harmful complications should be avoided. In this context, closed-loop insulin delivery is highly desirable. This can therefore be achieved by using a glucose concentration-sensing polymer delivery system carrying insulin.

[0004] However, the application of glucose sensors based on polymers containing glucose-binding moieties (GBMs) may be limited due to the limited sensitivity of such measurements. Furthermore, since the pKa of boronic acid typically falls in the 7–9 range, such polymer sensors have been shown to be highly pH-dependent. (Skjaervold et al., ANESTHESIOLOGY 2011; 114:18–20, Worsley et al., J Diabetes Sci Technol Vol. 2, No. 2, March 2008, Strasma et al., DOI: 10.1177 / 1932296815585872).

[0005] The paper "The Development of a Continuous Intravascular Glucose Monitoring Sensor" published by Crane, BC, et al. in the Journal of Diabetes Science and Technology (https: / / doi.org / 10.1177 / 1932296815587937) discloses a fluorescent hydrogel containing Wulff-type boronic acid for glucose sensing. This paper discloses the application of Wulff-type boronic acid in glucose sensing and discusses the stoichiometry of boronic acid binding to glucose. A key difference between this disclosure and the present invention is that the sensor is based on fluorescence measurements rather than changes in hydrogel volume.

[0006] The article “A Wulff-type boronate for boronate affinity capture of cis-diol compounds at medium acidic pH condition” (Li et al., Chem. Commun., 2011, 47, 8169-8171) discloses a borate affinity column containing a boric acid moiety for capturing cis-diols. However, this article does not disclose the use of the provided polymer for glucose sensing.

[0007] Kim et al. (“Polymeric Monosaccharide Receptors Responsive at NeutralpH”) described the synthesis of styrene-ADAM monomers and corresponding linear polymers via RAFT. Glucose sensitivity was determined by changes in the turbidity of the polymer solution (anhydrous gel) (DOI: 10.1021 / ja905652w).

[0008] Document US 2016 / 109370 discloses a sensor comprising a polymer structure (HEAA-BIS) containing 5-acryloylamino-2-((dimethylamino)methyl)phenylboronic acid units.

[0009] Document US 2008 / 214912 discloses a biosensor for detecting glucose, which contains a 4-aminomethyl-2-N,N'-dimethylaminomethylphenylboronic acid unit in its polymer structure.

[0010] Document CN 102 219 800 B discloses the copolymerization of 4-amino-2-(dimethylaminomethyl)phenylboronic acid with glycidyl methacrylate and polyethylene glycol diacrylate.

[0011] This invention addresses the problem of providing a glucose biosensor with improved performance. In particular, the biosensor of this invention overcomes the significant pH interference observed in existing boric acid-based biosensors. Furthermore, the biosensor of this invention, especially in a particular embodiment, is characterized by improved selectivity of glucose relative to fructose, and / or suppression of mannitol interference and other common interferences.

[0012] Therefore, the present invention provides formulations for polymer sensors, particularly polymer hydrogel sensors that linearly shrink upon exposure to increased glucose concentrations. It is presumed that the pKa of the boronic acid moiety is reduced due to the presence of an ortho-amino group in the portion of formula (I), thereby allowing glucose to coordinate at physiological pH (i.e., pH 7.35–7.45) and exhibiting extremely low pH dependence within said physiological range.

[0013] Therefore, this problem is solved by the embodiments disclosed herein and the features that constitute the claims.

[0014] The invention is summarized in the following embodiments.

[0015] In a first embodiment, the present invention relates to a biosensor for measuring glucose concentration, the biosensor comprising a polymer comprising a portion of formula (I):

[0016]

[0017] The portion of formula (I) is fixed to the polymer.

[0018] Where R is independently C 1-5 Alkyl, C 2-5 alkenyl or C 2-5 alkynyl group;

[0019] Where R S Each is independently selected from C 1-5 Alkyl, C 2-5 alkenyl, C 2-5 alkynyl group, -(C 0-3 alkylene)-OH, -(C 0-3 alkylene)-O(C 1-5 alkyl), -(C 0-3 alkylene)-O(C 1-5 alkylene)-OH, -(C 0-3 alkylene)-O(C 1-5 alkylene)-O(C 1-5 alkyl), -(C 0-3 alkylene)-SH, -(C 0-3 alkylene)-S(C 1-5alkyl), -(C 0-3 alkylene)-S(C 1-5 alkylene)-SH, -(C 0-3 alkylene)-S(C 1-5 alkylene)-S(C 1-5 alkyl), -(C 0-3 alkylene)-NH2, -(C 0-3 alkylene)-NH(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-OH, -(C 0-3 alkylene)-N(C 1-5 alkyl)-OH, -(C 0-3 alkylene)-NH-O(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-O(C 1-5 alkyl), -(C 0-3 alkylene)-halogen, -(C 0-3 alkylene)-(C 1-5 Halogenated alkyl), -(C 0-3 alkylene)-O-(C 1-5 Halogenated alkyl), -(C 0-3 alkylene)-CN, -(C 0-3 alkylene)-NO2, -(C 0-3 alkylene)-CHO, -(C 0-3 alkylene)-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-COOH, -(C 0-3 alkylene)-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-O-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-CO-NH2, -(C 0-3 alkylene)-CO-NH(C 1-5 alkyl), -(C 0-3 alkylene)-CO-N(C 1-5 Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-CO-(C 1-5alkyl), -(C 0-3 alkylene)-NH-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-O-CO-NH-(C 1-5 alkyl), -(C 0-3 alkylene)-O-CO-N(C 1-5 alkyl)-(C 1-5 alkyl), -(C 0-3 alkylene)-SO2-NH2, -(C 0-3 alkylene)-SO2-NH(C 1-5 alkyl), -(C 0-3 alkylene)-SO2-N(C 1-5 Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-SO2-(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-SO2-(C 1-5 alkyl), -(C 0-3 alkylene)-SO2-(C 1-5 alkyl), -(C 0-3 alkylene)-SO-(C 1-5 alkyl), -(C 0-3 alkylene)-carbocyclic and -(C 0-3 alkylene)-heterocyclic group, wherein -(C 0-3 The carbocyclic moiety in the alkylene group and the -(C 0-3 In each of the alkylene-heterocyclic groups, the heterocyclic moiety is optionally substituted by one or more groups, said groups being independently selected from C10. 1-4 Alkyl, halogen, -CN, -NO2, -OH, -O-(C 1-4 Alkyl), -SH, -S-(C 1-4 Alkyl groups, -NH2, -NH(C) 1-4 alkyl), -N(C) 1-4 Alkyl)(C 1-4 Alkyl groups, -COOH, -COO (C 1-4 Alkyl group), -CONH2, -CONH(C 1-4 Alkyl), -CON(C) 1-4 Alkyl)(C 1-4 alkyl), -NHCO(C 1-4 alkyl) and -N(C) 1-4alkyl)-CO(C 1-4 alkyl);

[0020] And n can be 0, 1, 2 or 3.

[0021] In a second embodiment, the present invention relates to a polymer contained in a biosensor as described above in the first embodiment.

[0022] In a third embodiment, the present invention relates to a biosensor for measuring glucose concentration, as described in the first embodiment of the invention, for use in in vivo diagnostic methods.

[0023] In a fourth embodiment, the present invention relates to a biosensor for measuring glucose concentration, as described in a first embodiment of the invention, for use in an in vivo glucose monitoring method.

[0024] In a fifth embodiment, the present invention relates to a biosensor for measuring glucose concentration, as described in a first embodiment of the invention, for use in vivo in diagnosing hyperglycemia or hypoglycemia.

[0025] In a sixth embodiment, the present invention relates to the use of a biosensor according to the first embodiment of the invention in an in vitro diagnostic method.

[0026] In a seventh embodiment, the present invention relates to the use of a biosensor according to a first embodiment of the invention in measuring glucose concentration in a sample.

[0027] In the eighth embodiment, the present invention relates to the use of the biosensor of the first embodiment of the invention in an in vitro method for diagnosing hyperglycemia or hypoglycemia.

[0028] In a ninth embodiment, the present invention relates to the use of polymers as described in a second embodiment of the invention in the preparation of reagents or biosensors for monitoring glucose levels in individuals.

[0029] In a tenth embodiment, the present invention relates to a glucose-concentration-sensitive release formulation comprising a polymer as described in a second embodiment.

[0030] In the eleventh embodiment, the present invention relates to a biosensor or glucose concentration-sensitive release agent for the treatment of glucose concentration-dependent conditions (e.g., diabetes).

[0031] The invention is further illustrated by the accompanying drawings; however, the drawings should not be construed as limiting.

[0032] Figure 1This shows the response of the ADAM-3%-TMPAPA sensor length to changes in the pH of different concentrations of glucose (indicator) PBS at 37°C.

[0033] Figure 2 This shows a comparison of pH interference between the 3APB sensor (Group A) and the ADAM sensor (Group B).

[0034] Figure 3 shows the response of the ADAM sensor to glucose with increasing mol% TMAPAA (A) and the interference of the ADAM sensor at 2.2 mM glucose with increasing mol% TMAPAA: B: 1 mM fructose, C: 5 mM mannitol, D: 38 mM NaCl, E: 10 mM lactate, F: 2 mM citrate, G: pH 6.9, H: pH 7.6. ±0.85 mM indicates the acceptable limit of interference. PBS pH 7.4, 37°C.

[0035] Figure 4 The response curves of the ADAM-3%-TMAPAA sensor (1-5) to glucose (in mM) and various interfering substances are shown. The interfering substance concentrations tested were significantly higher than the expected concentrations in vivo.

[0036] Figure 5 This shows the effect of 2APB copolymerization with ADAM on interference with 5 mM mannitol in all cases.

[0037] Figure 6 Display (A) Fabry-Pérot interferometer model. The Fabry-Pérot cavity is constructed of hydrogel. Refractive index n eff n g n f These represent the effective refractive index of the fiber, the refractive index of the hydrogel, and the refractive index of the fluid, respectively. The distance between the two boundaries is denoted as L. g (A) Also known as the hydrogel length; (B) Typical interferogram from a low-resolution Fabry-Pérot cavity. The wavelength of light is converted into frequency on the x-axis. The observed 0.35 phase shift corresponds to a 100 nm change in the hydrogel cavity length.

[0038] Figure 7 shows the change in sensor length of the ADAM-HEAA sensor (dashed line) and ADAM-acrylamide sensor (solid line) as glucose concentration increases (Figures A and B show hydrogels containing TMAPAA and hydrogels without TMAPAA, respectively).

[0039] Figure 8 The interference effect of 2 mM citrate was shown when detecting 6 mM glucose concentration.

[0040] Figure 9 shows the glucose response (in mM) of the modified ADAM sensor in PBS at pH 7.4 and 37°C in Figure A; and the glucose response (in mM) of the directly polymerized ADAM sensor containing 28 mol% ADAM in PBS at pH 7.4 and 37°C in Figure B.

[0041] As described above, the present invention relates to a biosensor for measuring glucose concentration, the biosensor comprising a polymer (preferably a polymer hydrogel) comprising a portion of formula (I):

[0042]

[0043] The portion of formula (I) is fixed onto the polymer.

[0044] In equation (I), R is independently C. 1-5 Alkyl, C 2-5 alkenyl or C 2-5 Alkyne group. Preferably, each group is independently C10. 1-5 Alkyl group. More preferably, each of R is C10. 1-2 Alkyl group. Even more preferably, R is methyl group.

[0045] In equation (I), R S Each is independently selected from C 1-5 Alkyl, C 2-5 alkenyl, C 2-5 alkynyl group, -(C 0-3 alkylene)-OH, -(C 0-3 alkylene)-O(C 1-5 alkyl), -(C 0-3 alkylene)-O(C 1-5 alkylene)-OH, -(C 0-3 alkylene)-O(C 1-5 alkylene)-O(C 1-5 alkyl), -(C 0-3 alkylene)-SH, -(C 0-3 alkylene)-S(C 1-5 alkyl), -(C 0-3 alkylene)-S(C 1-5 alkylene)-SH, -(C 0-3 alkylene)-S(C 1-5 alkylene)-S(C 1-5 alkyl), -(C 0-3 alkylene)-NH2, -(C 0-3alkylene)-NH(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-OH, -(C 0-3 alkylene)-N(C 1-5 alkyl)-OH, -(C 0-3 alkylene)-NH-O(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-O(C 1-5 alkyl), -(C 0-3 alkylene)-halogen, -(C 0-3 alkylene)-(C 1-5 Halogenated alkyl), -(C 0-3 alkylene)-O-(C 1-5 Halogenated alkyl), -(C 0-3 alkylene)-CN, -(C 0-3 alkylene)-NO2, -(C 0-3 alkylene)-CHO, -(C 0-3 alkylene)-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-COOH, -(C 0-3 alkylene)-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-O-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-CO-NH2, -(C 0-3 alkylene)-CO-NH(C 1-5 alkyl), -(C 0-3 alkylene)-CO-N(C 1-5 Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-NH-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-O-CO-NH-(C1-5 alkyl), -(C 0-3 alkylene)-O-CO-N(C 1-5 alkyl)-(C 1-5 alkyl), -(C 0-3 alkylene)-SO2-NH2, -(C 0-3 alkylene)-SO2-NH(C 1-5 alkyl), -(C 0-3 alkylene)-SO2-N(C 1-5 Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-SO2-(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-SO2-(C 1-5 alkyl), -(C 0-3 alkylene)-SO2-(C 1-5 alkyl), -(C 0-3 alkylene)-SO-(C 1-5 alkyl), -(C 0-3 alkylene)-carbocyclic and -(C 0-3 alkylene)-heterocyclic group, wherein -(C 0-3 The carbocyclic moiety in the alkylene group and the -(C 0-3 In each of the alkylene-heterocyclic groups, the heterocyclic moiety is optionally substituted by one or more groups, said groups being independently selected from C10. 1-4 Alkyl, halogen, -CN, -NO2, -OH, -O-(C 1-4 Alkyl), -SH, -S-(C 1-4 Alkyl groups, -NH2, -NH(C) 1-4 alkyl), -N(C) 1-4 Alkyl)(C 1-4 Alkyl groups, -COOH, -COO (C 1-4 Alkyl group), -CONH2, -CONH(C 1-4 Alkyl), -CON(C) 1-4 Alkyl)(C 1-4 alkyl), -NHCO(C 1-4 alkyl) and -N(C) 1-4 alkyl)-CO(C 1-4 alkyl).

[0046] Preferably, R S Each is independently selected from C 1-5 Alkyl, C 2-5 alkenyl, C 2-5 alkynyl group, -(C 0-3alkylene)-OH, -(C 0-3 alkylene)-O(C 1-5 alkyl), -(C 0-3 alkylene)-O(C 1-5 alkylene)-OH, -(C 0-3 alkylene)-O(C 1-5 alkylene)-O(C 1-5 alkyl), -(C 0-3 alkylene)-SH, -(C 0-3 alkylene)-S(C 1-5 alkyl), -(C 0-3 alkylene)-S(C 1-5 alkylene)-SH, -(C 0-3 alkylene)-S(C 1-5 alkylene)-S(C 1-5 alkyl), -(C 0-3 alkylene)-NH2, -(C 0-3 alkylene)-NH(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-OH, -(C 0-3 alkylene)-N(C 1-5 alkyl)-OH, -(C 0-3 alkylene)-NH-O(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-O(C 1-5 alkyl), -(C 0-3 alkylene)-halogen, -(C 0-3 alkylene)-(C 1-5 Halogenated alkyl), -(C 0-3 alkylene)-O-(C 1-5 Halogenated alkyl), -(C 0-3 alkylene)-CN, -(C 0-3 alkylene)-NO2, -(C 0-3 alkylene)-CHO, -(C 0-3 alkylene)-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-COOH, -(C 0-3 alkylene)-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-O-CO-(C 1-5 alkyl), -(C 0-3alkylene)-CO-NH2, -(C 0-3 alkylene)-CO-NH(C 1-5 alkyl), -(C 0-3 alkylene)-CO-N(C 1-5 Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-NH-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-O-CO-NH-(C 1-5 alkyl), -(C 0-3 alkylene)-O-CO-N(C 1-5 alkyl)-(C 1-5 alkyl), -(C 0-3 alkylene)-SO2-NH2, -(C 0-3 alkylene)-SO2-NH(C 1-5 alkyl), -(C 0-3 alkylene)-SO2-N(C 1-5 Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-SO2-(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-SO2-(C 1-5 alkyl), -(C 0-3 alkylene)-SO2-(C 1-5 alkyl) and -(C 0-3 alkylene)-SO-(C 1-5 alkyl).

[0047] More preferably, R S Each is independently selected from C 1-5 Alkyl, C 2-5 alkenyl, C 2-5 alkynyl group, -(C 0-3 alkylene)-OH, -(C 0-3 alkylene)-O(C 1-5 alkyl), -(C 0-3 alkylene)-O(C 1-5alkylene)-OH, -(C 0-3 alkylene)-O(C 1-5 alkylene)-O(C 1-5 alkyl), -(C 0-3 alkylene)-SH, -(C 0-3 alkylene)-S(C 1-5 alkyl), -(C 0-3 alkylene)-S(C 1-5 alkylene)-SH, -(C 0-3 alkylene)-S(C 1-5 alkylene)-S(C 1-5 alkyl), -(C 0-3 alkylene)-NH2, -(C 0-3 alkylene)-NH(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-OH, -(C 0-3 alkylene)-N(C 1-5 alkyl)-OH, -(C 0-3 alkylene)-NH-O(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-O(C 1-5 alkyl), -(C 0-3 alkylene)-halogen, -(C 0-3 alkylene)-(C 1-5 Halogenated alkyl), -(C 0-3 alkylene)-O-(C 1-5 Halogenated alkyl), -(C 0-3 alkylene)-CN, -(C 0-3 alkylene)-NO2, -(C 0-3 alkylene)-CHO, -(C 0-3 alkylene)-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-COOH, -(C 0-3 alkylene)-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-O-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-CO-NH2, -(C 0-3 alkylene)-CO-NH(C 1-5 alkyl), -(C 0-3 alkylene)-CO-N(C 1-5Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-NH-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-O-CO-NH-(C 1-5 alkyl), -(C 0-3 alkylene)-O-CO-N(C 1-5 alkyl)-(C 1-5 alkyl).

[0048] Even more preferably, R S Each is independently selected from C 1-5 Alkyl, C 2-5 alkenyl, C 2-5 alkynyl group, -(C 0-3 alkylene)-OH, -(C 0-3 alkylene)-O(C 1-5 alkyl), -(C 0-3 alkylene)-O(C 1-5 alkylene)-OH, -(C 0-3 alkylene)-O(C 1-5 alkylene)-O(C 1-5 alkyl), -(C 0-3 alkylene)-SH, -(C 0-3 alkylene)-S(C 1-5 alkyl), -(C 0-3 alkylene)-S(C 1-5 alkylene)-SH, -(C 0-3 alkylene)-S(C 1-5 alkylene)-S(C 1-5 alkyl), -(C 0-3 alkylene)-NH2, -(C 0-3 alkylene)-NH(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-OH, -(C 0-3 alkylene)-N(C 1-5alkyl)-OH, -(C 0-3 alkylene)-NH-O(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-O(C 1-5 alkyl), -(C 0-3 alkylene)-halogen, -(C 0-3 alkylene)-(C 1-5 Halogenated alkyl), -(C 0-3 alkylene)-O-(C 1-5 Halogenated alkyl), -(C 0-3 alkylene)-CN and -(C 0-3 (alkylene)-NO2.

[0049] Still more preferably, R S Each is independently selected from C 1-5 Alkyl, C 2-5 alkenyl, C 2-5 Alkyne group, -OH, -O(C) 1-5 Alkyl), -O(C) 1-5 alkylene)-OH, -O(C 1-5 alkylene)-O(C 1-5 Alkyl), -SH, -S(C 1-5 alkyl), -S(C 1-5 alkylene)-SH, -S(C 1-5 alkylene)-S(C 1-5 Alkyl groups, -NH2, -NH(C) 1-5 alkyl), -N(C) 1-5 Alkyl)(C 1-5 Alkyl), -NH-OH, -N(C 1-5 alkyl)-OH, -NH-O(C 1-5 alkyl), -N(C) 1-5 alkyl)-O(C 1-5 Alkyl), -halogen, C 1-5 Haloalkyl, -O-(C 1-5 Halogenated alkyl groups), -CN, and -NO2.

[0050] Even more preferably, R S Each is independently selected from -OH, -SH, -NH2, -NH-OH, -halogen, -CN, and -NO2.

[0051] In formula (I), n is 0, 1, 2, or 3. Preferably, n is 0 or 1. More preferably, n is 0. It should be understood that the variable n represents the substituent R connected to the corresponding phenyl moiety. S The number of substituents. If n is 0, then there is no substituent R. SThis makes the corresponding benzene ring unsubstituted (i.e., carrying hydrogen substitution R). S Therefore, the benzene ring in the preferred formula (I) is not R S replace.

[0052] It should be further understood that the portion of formula (I) can be attached to (or fixed to) the polymer by any ring carbon atom of the corresponding benzene ring, as reflected by the bonds extending into the benzene ring, where the wavy line (at one end of the bond) indicates the connection point of the portion of formula (I).

[0053] There are no particular limitations on the manner in which the portion of formula (I) is connected to the remainder of the polymer, and any chemically feasible connection is included in this invention. For example, any of these portions can be connected by an amide bond or a reverse amide bond, such as by a -NH-CO- or -CO-NH- group. Particularly preferred connections are exemplified in formula (I-1) below; another possible connection point is shown in formula (Ib) below. Other examples of connection points are apparent from the disclosure of methods for preparing the polymers and monomers used, including any of those described in the Examples section.

[0054]

[0055] The preferred polymer inclusion portion of the present invention is (Ia):

[0056]

[0057] Among them, R, R S And n is as described in equation (I).

[0058] However, in one embodiment of the invention, the polymer of the invention includes the portion of formula (Ib):

[0059]

[0060] Among them, R, R S And n as described in (I).

[0061] Those skilled in the art will understand that the portion of formula (I) can be interconverted with a form comprising a five-membered ring, which is formed through the bonding interaction between nitrogen and boron atoms in the portion of formula (I). Both forms can exist in equilibrium, depending on external conditions, such as the surrounding medium. In particular, in an aqueous medium, these forms can be interconverted by undergoing a ring-closing reaction or a corresponding ring-opening reaction, as illustrated in the following examples:

[0062]

[0063] There are no particular limitations on the polymers provided by the present invention. Preferably, the polymer is hydrophilic, which is advantageous when the polymer comes into contact with individual blood. Preferably, the polymer forms a polymer hydrogel, i.e., the polymer is preferably a polymer hydrogel. A hydrogel is generally understood as a biphase material comprising a solid polymer (preferably forming a porous and permeable structure) and an interstitial fluid contained within and in contact with said solid polymer. In the hydrogel, the fluid is preferably water or an aqueous solution (e.g., a liquid composition containing at least 90% (v / v) water). It is understood that polymer hydrogels typically comprise crosslinks between the individual polymer molecules constituting the hydrogel. Such crosslinks can be covalent or non-covalent (and thus can also be dynamic, i.e., undergoing formation and dissociation).

[0064] The term polymer preferably also includes dendritic polymers and non-crosslinked polymers.

[0065] Preferably, the term polymer refers to a polymer comprising crosslinks (covalent or non-covalent) between individual polymer molecules.

[0066] Preferred examples of polymers (or polymer hydrogels) used in the biosensors of the present invention are described below. Typically, the preferred polymers do not contain hyaluronic acid. Therefore, it is preferred that the polymer hydrogels included in the biosensors of the present invention do not contain hyaluronic acid.

[0067] Preferably, the polymer of the present invention is a polyacrylamide-based polymer. The term "polyacrylamide-based polymer" preferably refers to a polymer composed of (or consisting of) more than 50% (w / w) of acrylamide monomer, more preferably at least 60% (w / w) of acrylamide monomer, even more preferably at least 70% (w / w) of acrylamide monomer, even more preferably at least 80% (w / w) of acrylamide monomer, or even more preferably at least 90% (w / w) of acrylamide monomer. Therefore, the polyacrylamide-based polymer is preferably obtained by polymerizing a monomer composition wherein more than 50% (w / w) (or more preferably, at least 60% (w / w), at least 70% (w / w), at least 80% (w / w), or at least 90% (w / w)) of the monomer is acrylamide monomer.

[0068] It should be understood that "acrylamide monomer" is a compound containing an acrylamide moiety, for example, as shown in the following example:

[0069]

[0070] These types of acrylamide monomers can also be called N-substituted acrylamide monomers.

[0071] Typically, the alkenyl moiety in the aforementioned acrylamide monomer can be further substituted, but it is preferred that the alkenyl moiety is unsubstituted. Therefore, reference can be made to polymers based on unsubstituted polyacrylamide, preferably polymers consisting of (or composed of) more than 50% (w / w) of acrylamide monomers, wherein the alkenyl moiety is unsubstituted; more preferably at least 60% (w / w) of acrylamide monomers, wherein the alkenyl moiety is unsubstituted; even more preferably at least 70% (w / w) of acrylamide monomers, wherein the alkenyl moiety is unsubstituted; even more preferably at least 80% (w / w) of acrylamide monomers, wherein the alkenyl moiety is unsubstituted; and even more preferably at least 90% (w / w) of acrylamide monomers, wherein the alkenyl moiety is unsubstituted. Therefore, polymers based on unsubstituted polyacrylamide are preferably obtained by polymerization of monomer compositions, wherein more than 50% (w / w) (or more preferably at least 60% (w / w), at least 70% (w / w), at least 80% (w / w), or at least 90% (w / w)) of the monomers are acrylamide monomers, wherein the alkenyl moiety is unsubstituted.

[0072] The use of acrylamide monomers is advantageous because these monomers incorporate the desired hydrophilicity, which facilitates the formation of hydrogels and is readily functionalized, as required when constructing the polymers of this invention.

[0073] The polymers of the present invention can also be polymethacrylamide-based polymers. The term "polymethacrylamide-based polymer" preferably refers to a polymer composed of (or consisting of) more than 50% (w / w) of methacrylamide monomer, more preferably at least 60% (w / w) of methacrylamide monomer, even more preferably at least 70% (w / w) of methacrylamide monomer, even more preferably at least 80% (w / w) of methacrylamide monomer, or even more preferably at least 90% (w / w) of methacrylamide monomer. Therefore, polymethacrylamide-based polymers are preferably obtained by polymerization of a monomer composition wherein more than 50% (w / w) (or more preferably, at least 60% (w / w), at least 70% (w / w), at least 80% (w / w), or at least 90% (w / w)) of the monomer is a methacrylamide monomer.

[0074] It can be understood that "methacrylamide monomer" refers to a compound containing a methacrylamide moiety, for example, as shown in the following example:

[0075]

[0076] These types of methacrylamide monomers can also be called N-substituted methacrylamide monomers.

[0077] The polymers of the present invention can also be polyacrylate-based polymers. The term "polyacrylate-based polymer" preferably refers to a polymer composed of (or consisting of) more than 50% (w / w) of acrylate monomers, more preferably at least 60% (w / w) of acrylate monomers, even more preferably at least 70% (w / w) of acrylate monomers, even more preferably at least 80% (w / w) of acrylate monomers, or even more preferably at least 90% (w / w) of acrylate monomers. Therefore, polyacrylate-based polymers are preferably obtained by polymerization of a monomer composition wherein more than 50% (w / w) (or more preferably, at least 60% (w / w), at least 70% (w / w), at least 80% (w / w), or at least 90% (w / w)) of the monomers are acrylate monomers.

[0078] It should be understood that "acrylate monomer" is a compound containing an acrylate moiety, for example, as shown in the following example:

[0079]

[0080] The polymers of the present invention can also be polymethacrylate-based polymers. The term "polymethacrylate-based polymer" preferably refers to a polymer composed of (or consisting of) more than 50% (w / w) of methacrylate monomers, more preferably at least 60% (w / w) of methacrylate monomers, even more preferably at least 70% (w / w) of methacrylate monomers, even more preferably at least 80% (w / w) of methacrylate monomers, or even more preferably at least 90% (w / w) of methacrylate monomers. Therefore, polymethacrylate-based polymers are preferably obtained by polymerization of a monomer composition wherein more than 50% (w / w) (or more preferably, at least 60% (w / w), at least 70% (w / w), at least 80% (w / w), or at least 90% (w / w)) of the monomers are methacrylate monomers.

[0081] It should be understood that "methacrylate monomer" refers to a compound containing a methacrylate moiety, for example, as shown in the following example:

[0082]

[0083] It has been demonstrated that, compared with other polymers, particularly polymers based on substituted acrylamides, polymers based on polyacrylamide (where the acrylamide is unsubstituted acrylamide) are better suited for the biosensors of the present invention. In particular, it has been demonstrated that sensors based on unsubstituted polyacrylamides are superior to those based on substituted polyacrylamides, such as HEAA, which is N-(2-hydroxyethyl)acrylamide, at least in terms of sensitivity. Therefore, it has been demonstrated that, for the same monomer ratio, the length change of the ADAM-acrylamide sensor at 20 mM glucose is approximately 1.6 times that of the ADAM-HEAA sensor (see Example 2 for further details).

[0084] Therefore, the preferred polymer is a polymer based on unsubstituted polyacrylamide.

[0085] As used herein, “measuring glucose concentration” can include measuring the absolute value of glucose concentration in a liquid (e.g., blood) in contact with a biosensor. However, measuring glucose concentration also refers to determining glucose concentration relative to a threshold. For example, measuring glucose concentration can also involve determining whether the glucose concentration is below or above a threshold, or whether the concentration falls within a reference concentration range. This is particularly useful in clinical settings, such as when blood glucose concentration drops below a certain critical threshold requiring clinician intervention.

[0086] Measuring glucose concentration using the biosensor of this invention predicts the change in volume of a polymer upon contact with a glucose-containing liquid, in other words, upon contact with glucose. This invention is based, at least in part, on the surprising discovery that the polymer of this invention reduces its volume upon contact with glucose. Therefore, when the polymer (or, preferably, the polymer hydrogel) is contacted with a glucose-free liquid (also referred to as a liquid containing glucose below a minimum detectable concentration or below a certain threshold), in other words, in the absence of glucose, the boric acid-based glucose-binding portion does not form any new interaction with the polymer (or polymer hydrogel), and the volume of the polymer (or polymer hydrogel) remains unchanged. Conversely, when exposed to a glucose-containing liquid (i.e., containing a detectable concentration of glucose or containing glucose at a concentration exceeding a certain threshold), in other words, in the presence of glucose, the boric acid-based glucose-binding portion binds to the glucose, resulting in a change in polymer volume. Upon transition from a glucose-free liquid to a glucose-containing liquid, in other words, from the absence of glucose to the presence of glucose, a change in the volume of the polymer (or polymer hydrogel) can be observed; this is a reduction in the volume of the polymer (or polymer hydrogel), i.e., shrinkage. Unbound by theory, it is presumed that the shrinkage of the polymer upon contact with glucose is attributed to the formation of a stoichiometric 2:1 complex between the portion of formula (I) and glucose, as shown in the following scheme:

[0087]

[0088] Therefore, preferably, in the biosensor of the present invention for measuring glucose concentration, the polymer contracts upon contact with glucose.

[0089] Preferably, the degree of shrinkage is substantially linearly proportional to the glucose concentration. In this document, the degree of shrinkage is preferably understood as |ΔL / L|, i.e., the absolute value of ΔL / L, where ΔL is the change in the linear dimension of the polymer (or polymer hydrogel), and L is the total linear dimension of the polymer (or polymer hydrogel). Therefore, biosensors comprising polymers as described herein preferably exhibit a substantially linear correlation, more preferably a linear correlation between glucose concentration and ΔL / L. As used herein, the term "substantially linear" preferably means within the expected value of the linear correlation, particularly with a tolerance of 10% (i.e., within ±10% of the linear correlation), more preferably with a tolerance of 5%, and even more preferably with a tolerance of 2%.

[0090] As will be apparent to those skilled in the art, the dependence of the degree of shrinkage on glucose concentration allows for the detection of changes in glucose concentration. For example, reference curves of swelling / shrinkage (ΔL / L) measured at a variety of known glucose concentrations can be prepared; by comparing the measured values ​​of swelling / shrinkage with the reference curves, the glucose concentration can be determined.

[0091] As used herein, whenever the term "about" is used in conjunction with a numerical value, it preferably refers to ±10% of the indicated value, more preferably ±5%, even more preferably ±2%, even more preferably ±1%, and most preferably the exact indicated value. If the term "about" is used to describe the endpoints of a range, it preferably refers to the range from -10% of the lower endpoint of the indicated value to +10% of the upper endpoint of the indicated value; more preferably, it refers to the range from -5% of the lower endpoint to +5% of the upper endpoint; even more preferably, it refers to the range from -2% of the lower endpoint to +2% of the upper endpoint; more preferably, it refers to the range from -1% of the lower endpoint to +1% of the upper endpoint; and most preferably, it refers to the range defined by the exact values ​​of the lower and upper endpoints. It should be noted that the numerical value can also be a ratio, for example, a ratio used to represent the composition of the polymers of the present invention, as described above. While a ratio can be described as a division of two numerical values, it can also be expressed as a single number obtained by said division. For example, a ratio of 1:2 can also be expressed as a ratio of 0.5. The aforementioned ±x% deviation can be applied to the corresponding individual numbers (generated by the division operation), and the endpoints of the resulting range can each be converted back to the corresponding ratio. For example, a ratio of 1:2 ±10% can also be expressed as 0.5 ±10% or a ratio of 0.45-0.55, which is equivalent to a ratio of 1:1.82-1:2.22.

[0092] Furthermore, the determination / detection of polymer volume changes can also be achieved by measuring another relevant value, particularly a proxy (or substitute) value that depends on the polymer volume, such as by measuring osmotic pressure (or osmotic pressure change). Therefore, the present invention also relates to a biosensor (as described herein) for measuring glucose concentration, wherein the measurement of glucose concentration is based on the sensitivity of glucose concentration-osmotic pressure changes within a polymer (or, as preferred in the present invention, a polymer hydrogel) contained in the biosensor.

[0093] The volume change of the polymer (polymer hydrogel), i.e., the swelling / shrinkage response in response to glucose, is reversible and can be measured (and thus monitored in real time) by a variety of signal transduction mechanisms, including electrochemical, mechanical, and optical techniques (e.g., via Fabry-Pérot interferometry or refractive indexing). The refractive index of the polymer can change simply due to volume change (i.e., if the polymer swells, it becomes more rarefied, and thus its refractive index decreases) and / or be attributed to the binding of analyte molecules to the polymer chains.

[0094] Polymers, preferably polymer hydrogels, have been described above. According to established practice in the art, polymers can be defined according to their preparation methods, particularly according to the monomers being polymerized. Therefore, preferred polymers (or polymer hydrogels) can be obtained in a polymerization reaction of a composition comprising the following components:

[0095] (i) Acrylamide monomer, which includes the boric acid-based glucose-binding moiety of formula (I).

[0096] It should be understood that the boric acid-based glucose-binding moiety in the acrylamide monomer (i) is connected to the remainder of the corresponding monomer via a linker, as shown above. It should be noted that some formulas showing preferred embodiments of the boric acid-based glucose-binding moiety include a -NH-CO- portion; in the corresponding acrylamide monomer (i) or (ii), this -NH-CO- portion preferably forms part of the acrylamide moiety, i.e., the -CO- in the -NH-CO- portion is preferably linked to a vinyl group (-CH=CH2), thereby forming the acrylamide moiety -NH-CO-CH=CH2.

[0097] The preferred option, (i), is:

[0098]

[0099] Among them, R, R S And n is as described in equation (I).

[0100] However, in one implementation, (i) is:

[0101]

[0102] Among them, R, R S And n as described in formula (I). Such compounds are known from the literature (Chem. Commun., 2011, 47, 8169-8171, DOI: 10.1039 / c1cc11096a).

[0103] Preferably, the composition for the polymerization reaction further comprises:

[0104] (ii) Acrylamide monomers without the boric acid moiety, and

[0105] (iii) Crosslinking agent acrylamide monomer.

[0106] Acrylamide monomers that do not contain the boric acid portion are monomers that include the following components:

[0107] ,

[0108] Preferably, it does contain a copy of this moiety but does not contain any boric acid moiety. There are no particular limitations on such monomers, and they preferably include unsubstituted acrylamide monomers (where the aforementioned moiety is linked to hydrogen) or substituted acrylamide monomers, where the aforementioned moiety is linked, for example, to a hydroxyalkyl or ethylene glycol oligomer. Exemplary preferred monomers (ii) are selected from the following monomers:

[0109] and , wherein q is an integer from 0 to 10; preferably, q is an integer from 2 to 5, more preferably, q is 2 or 5. One type of monomer (ii) or more types (e.g., two or three) of monomer (ii) may be present in the composition which is polymerized to obtain the polymer of the present invention.

[0110] The crosslinking agent acrylamide monomer (iii) preferably comprises at least two copies of the following moiety:

[0111] ,

[0112] Preferably, it contains two copies of the aforementioned portion. Furthermore, it is preferable that it does not contain the boric acid portion.

[0113] Preferably, the crosslinking agent acrylamide monomer (iii) comprises (or is) a monomer selected from:

[0114] and ,

[0115] Where q is an integer from 0 to 10, preferably an integer from 2 to 5, and more preferably q is 2 or 5.

[0116] Preferably, for the polymer of the present invention (more preferably the polymer hydrogel), the acrylamide monomer (i) accounts for 6-30 mol% of the acrylamide-based component in the composition used for the polymerization reaction. More preferably, the acrylamide monomer (i) accounts for 10-14 mol% of the acrylamide-based component in the composition used for the polymerization reaction. However, the amount of acrylamide monomer (i) may also be higher, and accounts for 15-24 mol% of the acrylamide-based component in the composition used for the polymerization reaction, preferably 16-20 mol% of the acrylamide-based component in the composition used for the polymerization.

[0117] Preferably, in the polymer of the present invention, the monomer (ii) accounts for 60-94 mol of the acrylamide-based component in the composition used for the polymerization reaction, more preferably 66-90 mol of the acrylamide-based component in the composition used for the polymerization reaction, and more preferably 74-84 mol of the acrylamide-based component in the composition used for the polymerization reaction.

[0118] Preferably, in the polymer of the present invention, the crosslinking agent acrylamide monomer (iii) accounts for 0.1-4 mol% of the acrylamide-based component in the composition used for the polymerization reaction. More preferably, in the polymer of the present invention, the crosslinking agent acrylamide monomer (iii) accounts for 1-4 mol% of the acrylamide-based component in the composition used for the polymerization reaction.

[0119] In an alternative embodiment of the invention, the polymer (or polymer hydrogel) may be obtained in the polymerization reaction of the composition, the composition comprising:

[0120] (ia) Non-acrylamide monomers comprising the boric acid-based glucose-binding moiety of formula (I). In this document, non-acrylamide monomers should be understood as monomers that do not contain the -NH-CO-CH=CH2 moiety.

[0121] Therefore, in an alternative embodiment of the invention, the polymer (or polymer hydrogel) can be obtained in a polymerization reaction as described above, wherein (ia) is used instead of (i).

[0122] In this alternative embodiment of the invention, it is preferred that (ia) be:

[0123]

[0124] Among them, R, R SAnd n as described in equation (I). Such monomers are known from the literature (J. Am. Chem. Soc. 2009, 131, 13908-13909, doi: 10.1021 / ja905652w).

[0125] In another alternative embodiment, (ia) is an acrylate monomer.

[0126] The polymers and polymer hydrogels of the present invention (if applicable) can be prepared according to the methods described above, and as exemplified in the Examples section.

[0127] Therefore, it is preferable to incorporate the portion of formula (I) into the monomer and then copolymerize it into the polymer of the present invention (preferably a polymer hydrogel) containing said portion. However, the present invention also includes embodiments in which the portion of formula (I) is incorporated into a pre-prepared polymer by modifying said polymer. An exemplary manner of such modification is a Michael addition reaction of the -SH group present as a thiolactone in the formed polymer with the acrylamide moiety.

[0128] The polymers of the present invention, particularly the polymeric hydrogels of the present invention, exhibit the advantageous properties shown above and in the examples, and are therefore particularly suitable for the biosensors of the present invention for measuring (or sensing) glucose concentration.

[0129] Particularly preferably, the degree of shrinkage of the polymer (or polymer hydrogel) in the presence of glucose is substantially independent of pH within a range of approximately pH 6.9 to approximately pH 7.6. This pH range is particularly important in intensive care unit (ICU) applications, where not only the normal blood pH range of approximately 7.4 to approximately 7.6 can be observed in the treated patients, but also a wider range of blood pH, including pathological pH values.

[0130] As used herein, the expression "substantially independent of pH value" in a specific pH range preferably refers to a maximum change / variation of 15% in swelling / shrinkage observed within said pH range, understood as 15% ΔL / L (equivalent to sensitivity (0-20 mM glucose)), more preferably 10%, and even more preferably 5%. The term "substantially independent of" also includes the narrower meaning of "independent of" specifically referred to.

[0131] The biosensors and polymers of the present invention may include other components and monomers, which may provide additional advantages. These descriptions are given in particular embodiments of the biosensors of the present invention disclosed below.

[0132] In a first particular embodiment of the biosensor of the present invention, the polymer further comprises a portion that is positively charged at pH=7.4. As understood herein, a positively charged portion is the portion whose net charge is greater than 0 at pH=7.4. Therefore, the zwitterionic portion containing both positive and negative charges is not considered positively charged because its net charge is neutral.

[0133] Preferably, in this first particular embodiment, the polymer also contains a portion comprising an amino (or ammonium) group that is positively charged at pH=7.4.

[0134] Examples of such groups include primary amino groups -NH2 (which have -NH3 in their ammonium form). + (structure), secondary amino-NH(C) 1-5 Alkyl groups, such as -NHCH3 (which in its ammonium form has -(NH2(C 1-5 alkyl)) + (structure), for example -(NH2CH3) + ), or tertiary amino-N(C 1-5 Alkyl)2, for example -N(CH3)2 (which in its ammonium form has -(NH(C)2) 1-5 Alkyl)2) + (structure).

[0135] An example of a quaternary ammonium group is -N(C 1-5 Alkyl)3 + For example, it can be -N(CH3)3 + .

[0136] Therefore, preferably, in this first particular embodiment, the polymer further comprises a moiety containing groups selected from: -NH2, -NH3 + -NH(C 1-5 alkyl), -(NH2(C 1-5 alkyl)) + -N(C 1-5 alkyl)2、-(NH(C) 1-5 Alkyl)2) + and -N(C 1-5 Alkyl)3 + .

[0137] It should be understood that the positively charged portion is preferably placed within the monomer used in the polymerization reaction (or, in other words, the positively charged portion is preferably obtained / available by using a suitable monomer in the polymerization reaction, for example, as described below). Preferably, in this particular embodiment, the monomer is selected from:

[0138]

[0139] and .

[0140] However, the present invention also includes embodiments in which the aforementioned positively charged portion is introduced into a reaction occurring after the polymerization reaction. In other words, the positively charged portion can also be introduced into the polymer through modification of the polymer.

[0141] Preferably, in this first particular embodiment of the biosensor of the present invention, the polymer further comprises a portion containing a quaternary ammonium group. The portion containing the quaternary ammonium group may be a portion of formula (II):

[0142]

[0143] The portion containing the quaternary ammonium group, particularly the portion of formula (II), is attached to the polymer of the present invention in the same manner as described above for the portion of formula (I). Therefore, in this first particular embodiment, the portion containing the quaternary ammonium group is preferably the portion of formula (IIa):

[0144]

[0145] It has been confirmed that the biosensors of the present invention, comprising a portion containing a quaternary ammonium group, possess advantageous properties in glucose sensing. Therefore, a first particular embodiment of the invention is based, at least in part, on the finding that if the polymer of the invention further comprises a portion containing a quaternary ammonium group, such as the portion of formula (II), the biosensors of the invention exhibit increased selectivity for glucose relative to common interfering agents (including, but not limited to, fructose, mannitol, lactate, and citrate). In the context of the invention, it has been surprisingly found that a ratio of monomer containing a quaternary ammonium group to monomer containing a boric acid moiety of 1:2 to 1:7, preferably 1:3 to 1:6, more preferably 1:4 to 1:5, is particularly advantageous, as it is sufficient to substantially suppress interference from fructose, mannitol, lactate, and citrate. A further advantageous ratio of monomer containing a quaternary ammonium group to monomer containing a boric acid moiety is 1:5 to 1:7. This is further confirmed in Figure 3.

[0146] Therefore, it is particularly advantageous that the ratio of the monomer containing the positively charged portion at pH=7.4 (including any particularly and preferred such portions disclosed herein) to the monomer containing the boric acid portion is 1:2 to 1:7, preferably 1:3 to 1:6, and more preferably 1:4 to 1:5, as this is sufficient to substantially suppress interference from fructose, mannitol, lactate, and citrate. An additional advantageous ratio of the monomer containing the positively charged portion at pH=7.4 (including any particularly and preferred such portions disclosed herein) to the monomer containing the boric acid portion is 1:5 to 1:7.

[0147] As used herein, when interference from a particular substance is substantially suppressed, the reading is substantially independent of the concentration of such substance. As used herein, the expression "substantially independent of concentration" for a particular substance within a specific concentration range preferably means a maximum change / variation of 15% in swelling / shrinkage observed within said concentration range, understood as 15% ΔL / L (equivalent to sensitivity (0-20 mM glucose)), more preferably a maximum change / variation of 10%, and even more preferably a maximum change / variation of 5%. The term "substantially independent of" also includes, specifically, the narrower meaning of "independent of".

[0148] Preferably, in this first particular embodiment of the invention, the portion of formula (II) is introduced into the polymer by polymerization of the reaction mixture, the reaction mixture comprising, in addition to the components described above, (iv) a monomer containing the portion of formula (II). Preferably, (iv) is:

[0149]

[0150] Preferably, in this first particular embodiment of the invention, for the polymer of the invention (more preferably polymer hydrogel), the acrylamide monomer (iv) accounts for 0.1-6 mol% of the acrylamide-based component in the composition / reaction mixture used for the polymerization reaction. More preferably, the acrylamide monomer (iv) accounts for 2-4 mol% of the acrylamide-based component in the composition used for the polymerization reaction.

[0151] It has been confirmed that the effectiveness of suppressing interferences as described herein depends on the presence of a positive charge in the monomer. The addition of TMAPAA has a beneficial effect on the resulting glucose sensor by reducing interference from fructose, mannitol, and citrate. This effect is further confirmed to originate from the positive charge on the molecule, and similar cationic monomers can thus provide similar benefits to TMAPAA.

[0152] In a second particular embodiment of the invention, the polymer further comprises a portion of formula (III):

[0153]

[0154] Therefore, this second particular embodiment of the invention is based at least in part on a surprising discovery that the biosensor of the invention exhibits suppressed sensitivity to mannitol if a portion of formula (II) is present in addition to the portion of formula (I). Preferably, the ratio of the portion of formula (III) to the portion of formula (I) is 1:10 to 1:5, more preferably 1:8 to 1:6.

[0155] Preferably, in this second particular embodiment, the portion of formula (III) is introduced into the polymer by polymerization of the reaction mixture, which, in addition to the components described above, also contains (v) a monomer containing the portion of formula (III). Preferably, monomer (v) is a compound of the following formula:

[0156]

[0157] Preferably, in this second particular embodiment of the invention, for the polymer of the invention (more preferably the polymer hydrogel), the acrylamide monomer (v) accounts for 1-5 mol% of the acrylamide-based component in the composition used for the polymerization reaction. More preferably, the acrylamide monomer (v) accounts for 2-4 mol% of the acrylamidoyl component in the composition used for the polymerization reaction. Even more preferably, the acrylamide monomer (v) accounts for about 3 mol% of the acrylamide-based component in the composition used for the polymerization reaction. In this second particular embodiment of the invention, preferably, acrylamide monomers (i) and (v) account for about 25 mol% and about 3 mol%, respectively.

[0158] Unless otherwise specified, the following definitions apply throughout this specification and the claims.

[0159] The term "hydrocarbon group" refers to a group composed of carbon and hydrogen atoms.

[0160] The term "alicyclic group" is used in conjunction with cyclic groups and indicates that the corresponding cyclic group is a non-aromatic group.

[0161] As used herein, the term "alkyl" refers to a monovalent saturated acyclic (i.e., non-cyclic) hydrocarbon group, which can be straight-chain or branched. Therefore, "alkyl" does not contain any carbon-carbon double bonds or any carbon-carbon triple bonds. 1-5 "alkyl" refers to an alkyl group having 1-5 carbon atoms. Preferred exemplary alkyl groups are methyl, ethyl, propyl (e.g., n-propyl or isopropyl), or butyl (e.g., n-butyl, isobutyl, sec-butyl, or tert-butyl). Unless otherwise defined, the term "alkyl" preferably refers to C14. 1-4 Alkyl, more preferably methyl or ethyl, and even more preferably methyl.

[0162] As used herein, the term "alkenyl" refers to a monovalent unsaturated acyclic hydrocarbon group, which may be straight-chain or branched, and contains one or more (e.g., one or two) carbon-carbon double bonds, but no carbon-carbon triple bonds. The term "C" 2-5"Alkenyl" refers to an alkenyl group having 2-5 carbon atoms. Preferred exemplary alkenyl groups are vinyl, propenyl (e.g., prop-1-en-1-yl, prop-1-en-2-yl, or prop-2-en-1-yl), butenyl, butadienyl (e.g., but-1,3-dien-1-yl or but-1,3-dien-2-yl), pentenyl, or pentadienyl (e.g., isoprene). Unless otherwise defined, the term "alkenyl" preferably refers to C 2-4 Alkenyl group.

[0163] As used herein, the term "alkynyl" refers to a monovalent unsaturated acyclic hydrocarbon group, which may be straight-chain or branched, and contains one or more (e.g., one or two) carbon-carbon triple bonds and optionally one or more (e.g., one or two) carbon-carbon double bonds. The term "C..." 2-5 "Alynyl" refers to an alkynyl group having 2-5 carbon atoms. Preferred exemplary alkynyl groups are ethynyl, propynyl (e.g., propynyl), or butynyl. Unless otherwise defined, the term "alkynyl" preferably refers to C... 2-4 Alkyne group.

[0164] As used herein, the term "alkylene" refers to an alkyl dienyllium, i.e., a divalent saturated acyclic hydrocarbon group, which can be straight-chain or branched. "C" 1-5 "alkylene" refers to an alkylene group having 1-5 carbon atoms, and the term "C" is used to indicate that the alkylene group has 1-5 carbon atoms. 0-3 "alkylene" indicates the presence of a covalent bond (equivalent to selecting "C0 alkylene") or C 1-3 Alkylene. Preferred exemplary alkylenes are methylene (-CH2-), ethylene (e.g., -CH2-CH2- or -CH(-CH3)-), propylene (e.g., -CH2-CH2-CH2-, -CH(-CH2-CH3)-, -CH2-CH(-CH3)- or -CH(-CH3)-CH2-), or butylene (e.g., -CH2-CH2-CH2-CH2-). Unless otherwise defined, the term "alkylene" preferably refers to C10. 1-4 Alkylenes (especially including straight-chain C) 1-4 Alkylene), more preferably methylene or ethylene, and even more preferably methylene.

[0165] As used herein, the term "carbocyclic" refers to a hydrocarbon cyclic group, including monocyclic, bridged, spirocyclic, and / or fused ring systems (which may consist of, for example, two or three rings), wherein the cyclic group may be saturated, partially unsaturated (i.e., unsaturated but not aromatic), or aromatic. Unless otherwise defined, "carbocyclic" preferably refers to an aryl, cycloalkyl, or cycloalkenyl group.

[0166] As used herein, the term "heterocyclic group" refers to a cyclic group, including monocyclic rings and bridged rings, spirocyclic rings, and / or fused ring systems (which may consist of, for example, two or three rings), wherein the cyclic group contains one or more (e.g., one, two, three, or four) cyclic heteroatoms independently selected from O, S, and N, and the remaining ring atoms are carbon atoms, wherein one or more S ring atoms (if present) and / or one or more N ring atoms (if present) may optionally be oxidized, wherein one or more carbon ring atoms may optionally be oxidized (i.e., forming an oxo group), and further wherein the cyclic group may be saturated, partially unsaturated (i.e., unsaturated but not aromatic), or aromatic. For example, each heteroatom-containing ring contained in the cyclic group may contain one or two O atoms and / or one or two S atoms (which may optionally be oxidized) and / or one, two, three, or four N atoms (which may optionally be oxidized), provided that the total number of heteroatoms on the respective heteroatom-containing ring is 1-4, and at least one carbon ring atom (which may optionally be oxidized) is present on the respective heteroatom-containing ring. Unless otherwise defined, "heterocyclic" preferably refers to heteroaryl, heterocyclic alkyl, or heterocyclic alkenyl.

[0167] As used herein, the term "aryl" refers to an aromatic hydrocarbon cyclic group, including monocyclic aromatic rings and bridged and / or fused ring systems containing at least one aromatic ring (e.g., a ring system consisting of two or three fused rings, wherein at least one of these fused rings is an aromatic ring; or a bridged ring system consisting of two or three rings, wherein at least one of these bridged rings is an aromatic ring). If the aryl group is a bridged and / or fused ring system, and in addition to one or more aromatic rings, it contains at least one non-aromatic ring (e.g., a saturated ring or an unsaturated alicyclic ring), then one or more carbon atoms on each non-aromatic ring may optionally be oxidized (i.e., forming an oxo group). "Aryl" may, for example, refer to phenyl, naphthyl, dihydronaphthyl (i.e., 1,2-dihydronaphthyl), tetrahydronaphthyl (i.e., 1,2,3,4-tetrahydronaphthyl), indenyl, indenyl (e.g., 1H-indenyl), anthracene, phenanthrene, 9H-fluorenyl, or azulel. Unless otherwise defined, "aryl" preferably has 6-14 ring atoms, more preferably 6-10 ring atoms, even more preferably refers to phenyl or naphthyl, and most preferably refers to phenyl.

[0168] As used herein, the term "heteroaryl" refers to an aromatic ring group, including monocyclic aromatic rings and bridged rings and / or fused ring systems containing at least one aromatic ring (e.g., a ring system consisting of two or three fused rings, wherein at least one of these fused rings is an aromatic ring; or a bridged ring system consisting of two or three rings, wherein at least one of these bridged rings is an aromatic ring), wherein the aromatic ring group comprises one or more (e.g., one, two, three or four) cyclic heteroatoms independently selected from O, S and N, and the remaining ring atoms are carbon atoms, wherein one or more S ring atoms (if present) and / or one or more N ring atoms (if present) may optionally be oxidized, and further wherein one or more carbon ring atoms may optionally be oxidized (i.e., forming an oxo group). For example, each heteroatom-containing ring in the aromatic ring group may contain one or two O atoms and / or one or two S atoms (which may be optionally oxidized) and / or one, two, three or four N atoms (which may be optionally oxidized), provided that the total number of heteroatoms on the corresponding heteroatom-containing ring is 1-4, and at least one carbon ring atom (which may be optionally oxidized) is present on the corresponding heteroatom-containing ring."Heteroaryl" can refer to, for example, thienyl (i.e., thiophenyl), benzo[b]thienyl, naphtho[2,3-b]thienyl, thianyl, furyl (i.e., furanyl), benzofuranyl, isobenzofuranyl, chromanyl, chromenyl (e.g., 2H-1-benzopyranyl or 4H-1-benzothienyl), isochromenyl (e.g., 1H-2-benzopyranyl), chromone, xanthonyl, phenoxathiinyl, pyrroleyl (e.g., 1H-pyrroleyl), imidazolyl, pyrazolyl, pyridyl (i.e., pyridinyl); for example, 2 -pyridyl, 3-pyridyl or 4-pyridyl), pyrazinyl, pyrimidinyl, pyridazinyl, indoleyl (e.g., 3H-indoleyl), isoindoleyl, indazoleyl, indazinyl, purineyl, quinolinyl, isoquinolinyl, phthalazinyl, naphthidyl, quinoxalinyl, cenolinyl, pteridinyl, carbazoyl, β-carolinyl, phenanthridineyl, acridineyl, perimidinyl, phenanthroxolinyl (e.g., [1,10]phenanthroxolinyl, [1,7]phenanthroxolinyl or [4,7]phenanthroxolinyl), phenazinyl, thiazolyl, iso... Thiazolyl, phenothiazyl, oxazolyl, isoxazolyl, oxadiazolyl (e.g., 1,2,4-oxadiazolyl, 1,2,5-oxadiazolyl (i.e., furazonyl) or 1,3,4-oxadiazolyl), thiadiazolyl (e.g., 1,2,4-thiadiazolyl, 1,2,5-thiadiazolyl or 1,3,4-thiadiazolyl), phenothiazyl, pyrazolo[1,5-a]pyrimidinyl (e.g., pyrazolo[1,5-a]pyrimidin-3-yl), 1,2-benzisoxazol-3-yl, benzothiazolyl, benzothiadiazolyl, benzoxazolyl Benzisoxazolyl, benzimidazolyl, benzo[b]thiophene (i.e., benzothiophene), triazolyl (e.g., 1H-1,2,3-triazolyl, 2H-1,2,3-triazolyl, 1H-1,2,4-triazolyl or 4H-1,2,4-triazolyl), benzotriazolyl, 1H-tetrazolyl, 2H-tetrazolyl, triazinyl (e.g., 1,2,3-triazinyl, 1,2,4-triazinyl or 1,3,5-triazinyl), furano[2,3-c]pyridyl, dihydrofuranopyridyl (e.g., 2,3-dihydrofuranopyridyl) , ...Unless otherwise defined, the term "heteroaryl" preferably refers to a 5-14 member (more preferably 5-10 member) monocyclic or fused-ring system comprising one or more (e.g., one, two, three or four) cyclic heteroatoms independently selected from O, S and N, wherein one or more S ring atoms (if present) and / or one or more N ring atoms (if present) are optionally oxidized, and wherein one or more carbocyclic atoms are optionally oxidized; even more preferably, "heteroaryl" refers to a 5 or 6 member monocyclic ring comprising one or more (e.g., one, two or three) cyclic heteroatoms independently selected from O, S and N, wherein one or more S ring atoms (if present) and / or one or more N ring atoms (if present) are optionally oxidized, and wherein one or more carbocyclic atoms are optionally oxidized.

[0169] As used herein, the term "cycloalkyl" refers to a saturated hydrocarbon cyclic group, including monocyclic, bridged, spirocyclic, and / or fused ring systems (which may consist of, for example, two or three rings; for example, fused ring systems consisting of two or three fused rings). "Cycloalkyl" may, for example, refer to cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, decalinyl (i.e., decahydronaphthyl) or adamantyl. Unless otherwise defined, "cycloalkyl" preferably refers to C14. 3-11 Cycloalkyl, and more preferably C 3-7 Cycloalkyl. Particularly preferred “cycloalkyl” is a monocyclic saturated hydrocarbon ring having 3-7 ring members (e.g., cyclopropyl or cyclohexyl).

[0170] As used herein, the term "heterocyclic alkyl" refers to a saturated cyclic group, including monocyclic and bridged, spirocyclic, and / or fused-ring systems (which may consist of, for example, two or three rings; e.g., fused-ring systems consisting of two or three fused rings), wherein the cyclic group comprises one or more (e.g., one, two, three, or four) cyclic heteroatoms independently selected from O, S, and N, and the remaining ring atoms are carbon atoms, wherein one or more S ring atoms (if present) and / or one or more N ring atoms (if present) may optionally be oxidized, and further wherein one or more carbon ring atoms may optionally be oxidized (i.e., forming an oxo group). For example, each heteroatom-containing ring in the saturated cyclic group may contain one or two O atoms and / or one or two S atoms (which may optionally be oxidized) and / or one, two, three, or four N atoms (which may optionally be oxidized), provided that the total number of heteroatoms on the corresponding heteroatom-containing ring is 1-4, and at least one carbon ring atom (which may optionally be oxidized) is present on the corresponding heteroatom-containing ring. "Heterocyclic alkyl" can refer to, for example, aziridinyl, aziridine, pyrrolidinyl, imidazoalkyl, pyrazolyl, piperidinyl, piperazinyl, aziridine-heptyl, diazacycloheptyl (e.g., 1,4-diazacycloheptyl), oxazolyl, isoxazolyl, thiazoalkyl, isothiazolyl, morpholinyl (e.g., morpholin-4-yl), thiomorpholinyl (e.g., thiomorpholin-4-yl), oxazacycloheptyl, ethylene oxide, etc. Oxycyclic butyl, tetrahydrofuranyl, 1,3-dioxolane, tetrahydropyranyl, 1,4-dioxyl, oxycyclic heptyl, thiocyclic propane, thiocyclic butyl, tetrahydrothiophene (i.e., thiocyclic pentyl), 1,3-dithiocyclic pentyl, thiaalkyl, 1,1-dioxothiaalkyl, thiocyclic heptyl, decahydroquinolinyl, decahydroisoquinolinyl or 2-oxa-5-aza-bicyclo[2.2.1]hept-5-yl. Unless otherwise defined, "heterocyclic alkyl" preferably refers to a 3-11 member saturated cyclic group that is a monocyclic or fused-ring system (e.g., a fused-ring system consisting of two fused rings), wherein the cyclic group comprises one or more (e.g., one, two, three, or four) cyclic heteroatoms independently selected from O, S, and N, wherein one or more S ring atoms (if present) and / or one or more N ring atoms (if present) are optionally oxidized, and wherein one or more carbocyclic atoms are optionally oxidized; more preferably, "heterocyclic alkyl" refers to a 5-7 member saturated monocyclic group that comprises one or more (e.g., one, two, or three) cyclic heteroatoms independently selected from O, S, and N, wherein one or more S ring atoms (if present) and / or one or more N ring atoms (if present) are optionally oxidized, and wherein one or more carbocyclic atoms are optionally oxidized.

[0171] As used herein, the term "halogen" refers to fluorine (-F), chlorine (-Cl), bromine (-Br), or iodine (-I). The terms "halogen" and "halogenated" are used interchangeably.

[0172] As used herein, the term "haloalkyl" refers to an alkyl group substituted with one or more (preferably 1-6, more preferably 1-3) halogen atoms, said halogen atoms being independently selected from fluorine, chlorine, bromine, and iodine, and preferably all of them being fluorine atoms. It should be understood that the maximum number of halogen atoms is limited by the number of available linking sites, and thus depends on the number of carbon atoms contained in the alkyl portion of the haloalkyl group. "Haloalkyl" can refer, for example, to -CF3, -CHF2, -CH2F, -CF2-CH3, -CH2-CF3, -CH2-CHF2, -CH2-CF2-CH3, -CH2-CF2-CF3, or -CH(CF3)2. A particularly preferred "haloalkyl" is -CF3.

[0173] The terms “valence bond” and “covalent bond” may be used as synonyms in this document, unless otherwise explicitly indicated or contradicted in the context.

[0174] As used herein, the terms “optional,” “optionally,” and “may” indicate that the indicated feature may be present or may not be present. Whenever the terms “optional,” “optionally,” or “may” are used, the invention specifically relates to both possibilities: the presence or absence of the corresponding feature. For example, the statement “X may be optionally substituted by Y” (or “X may be substituted by Y”) means that X is substituted by Y or not substituted. Similarly, if a component of a composition is indicated as “optional,” the invention specifically relates to both possibilities: the corresponding component is present (included in the composition) or the corresponding component is not present in the composition.

[0175] In this specification, multiple groups are referred to as "optionally substituted". Typically, these groups may carry one or more substituents, such as one, two, three, or four substituents. It should be understood that the maximum number of substituents is limited by the number of available linking sites on the substituted moiety. Unless otherwise defined, "optionally substituted" groups as referred to in this specification preferably carry no more than two substituents, and in particular may carry only one substituent. Furthermore, unless otherwise defined, it is preferable that there are no optional substituents, i.e., the corresponding groups are unsubstituted.

[0176] Those skilled in the art will understand that the substituents contained in the compounds of the present invention can be linked to the remainder of the corresponding compound at multiple different positions of the respective specific substituent. Unless otherwise defined, preferred connection positions of the different specific substituents are as exemplified in the examples.

[0177] As used herein, unless otherwise expressly stated or contradicted by the context, the terms “a,” “an,” and “the” may be used interchangeably with “one or more” and “at least one.”

[0178] It should be understood that all values ​​and subranges included within the numerical ranges provided / disclosed herein are included within the scope of this invention. Therefore, this invention specifically and exclusively relates to values ​​that fall within the numerical ranges disclosed herein, particularly including individual integer values ​​falling within the respective numerical ranges, and each subrange included within the numerical ranges disclosed herein.

[0179] As used herein, the term "comprising" (or "including" or "containing") has the meaning of "particularly comprising," i.e., "in addition to other optional elements," unless otherwise expressly indicated or contradicted in the context. In addition, the term also includes the narrower meanings of "substantially consisting of" and "consisting of." For example, the term "A comprises B and C" means "A specifically comprises B and C," where A may contain other optional elements (e.g., it may also include "A comprises B, C, and D"), but the term also includes the meanings of "A mainly consists of B and C" and "A consists of B and C" (i.e., A does not contain any elements other than B and C).

[0180] As used herein, the terms "individual" or "patient" refer to an animal, preferably a mammal (e.g., a human or a non-human mammal). Most preferably, the "individual" or "patient" refers to a human (e.g., a male or female human).

[0181] Measuring glucose concentration using the biosensor of the present invention typically requires measuring a polymer (or polymer hydrogel), particularly measuring volume changes detected by measuring changes in the length of the polymer / polymer hydrogel. Such measurements can be performed using the method described in US 7,602,498 B2, the entire contents of which are incorporated herein by reference. Specifically, to measure the length of the polymer hydrogel, interference spectra reflected from the hydrogel can be used to monitor changes in hydrogel length induced by different glucose concentrations. The hydrogel is located at the tip of a diced monomolecular fiber. The length L is relative to the initial absolute length. g (t=t1) Measurements are continuously monitored to monitor the relative length change of the hydrogel. The length of the hydrogel is measured from the cutting center along a straight path perpendicular to the cutting surface and up to the tip of the hydrogel.

[0182] The reflection spectrum of the hydrogel approximates a sine wave with DC components, amplitude, period, and phase terms. The hierarchical change in refractive index between the silica fibers, hydrogel, and fluid can be modeled as two weakly reflecting mirrors, represented by the boundary r1 between the fibers and the hydrogel and the boundary r2 between the hydrogel and the surrounding fluid. This system represents a low-resolution Fabry-Pérot (FP) cavity. The model is as follows: Figure 6 As shown in the image.

[0183] The reflection coefficients are r1 and r2. The secondary and weakest reflections at boundary r2 interfere with the reflected light at boundary r1. This interaction generates a sinusoidal interferogram in the reflection intensity as a function of wavelength. The FP interferogram is described below:

[0184]

[0185] in Let be the wavenumber of the light within the hydrogel cavity, and λ be the wavelength of the light. This originates from a light source with a wavelength of 39 nm and n... g The FP reflectance spectrum of the hydrogel cavity with a density of 1.35 is as follows: Figure 6 As shown in the figure. The absolute length of the hydrogel is determined by finding the period of the sinus (free spectral range), and the length change is determined by finding the phase shift between the spectra in a timely manner.

[0186] This invention relates to the specific use of biosensors in measuring glucose concentration. Therefore, the biosensors of this invention are suitable for in vivo diagnostic methods. These methods preferably include the steps of determining the glucose concentration in the blood of an individual (e.g., a human individual) using the biosensors of this invention as described above. Therefore, it should be understood that the biosensors of this invention are suitable for methods of determining the glucose concentration in the blood of an individual.

[0187] Preferably, the measurement can be performed continuously or repeatedly at specific time intervals, so that not only a single measurement point with limited information about the individual / patient's condition can be provided, but also a trend of blood glucose concentration over time or a series of measurements can be obtained, which are important for monitoring the patient's condition over a longer period of time, for example, when monitoring a patient's postoperative recovery or monitoring the patient in an intensive care unit. Such repeated or continuous measurements can also be referred to as glucose monitoring. Therefore, the present invention also relates to a method for glucose monitoring in an individual, the method comprising repeatedly measuring the glucose concentration in the individual's blood using the biosensor of the present invention. Thus, the present invention provides a biosensor for measuring glucose concentration in a method for glucose monitoring in vivo.

[0188] Measurement of blood glucose concentration provides information on diseases, conditions, and disorders characterized by pathological blood glucose concentrations, i.e., concentrations that differ from what is considered a normal range. If blood glucose concentration is below normal, typically below 70 mg / dL (or below 3.9 mmol / L), this can be termed hypoglycemia. If blood glucose concentration is above normal, typically above 200 mg / dL (or above 11.1 mmol / L), this can be termed hyperglycemia. Such conditions can be identified or diagnosed by directly measuring the glucose concentration in the blood. Therefore, the present invention also relates to a method for diagnosing hyperglycemia or hypoglycemia in an individual, the method comprising the step of measuring blood glucose concentration using the biosensor of the present invention. Thus, the present invention provides a biosensor for measuring glucose concentration according to a first embodiment of the present invention for use in an in vivo method for diagnosing hyperglycemia or hypoglycemia. Furthermore, the present invention also relates to the use of the biosensor of the present invention in an in vitro method for diagnosing hyperglycemia or hypoglycemia, for example by measuring glucose concentration in a blood sample obtained from an individual.

[0189] Therefore, the present invention also provides diagnostic methods (particularly in vitro diagnostic methods) using the biosensor of the present invention, the method comprising the step of measuring the blood glucose concentration in a blood sample from an individual using said biosensor. Thus, the biosensor of the present invention can also be used in vitro to measure the glucose concentration in a sample obtained from a patient. The method can also be referred to as an in vitro diagnostic method. In other words, the present invention provides the biosensor of the present invention for use in in vitro diagnostic methods. There are no particular limitations on the method itself, as long as it includes the necessary step of determining the glucose concentration in a blood sample. The measurement can be performed as described above, for example by measuring polymer volume changes that occur in a glucose concentration-dependent manner.

[0190] The measurement of glucose concentration using the biosensor of the present invention is not limited to measurements in blood. Therefore, the present invention generally relates to the use of the biosensor of the present invention in measuring glucose concentration in samples, including non-blood samples such as urine. However, preferably, the sample is a blood sample (e.g., whole blood sample, serum sample, or plasma sample).

[0191] The biosensor of the present invention can be configured to obtain an individual's blood via an indwelling arterial catheter. Because the sensor can be applied without interfering with catheter use, the sensor of the present invention does not require a new catheter or replacement of other devices.

[0192] As further provided herein, the present invention relates to the use of the polymers provided herein in the preparation of reagents or biosensors for monitoring glucose levels in individuals. It should be understood that changes in the properties of the polymer upon contact with different concentrations of glucose allow for measurement.

[0193] As described above, the biosensor of the present invention can also be used for therapeutic purposes, for example, when incorporated into a glucose concentration-dependent release formulation. Therefore, the present invention also provides a glucose concentration-sensitive release formulation comprising the polymer as described herein. The corresponding glucose-sensitive release formulation may also comprise one or more pharmaceutically acceptable carriers and an active substance / therapeutic agent (e.g., insulin) delivered in a glucose concentration-dependent manner.

[0194] This formulation can be prepared using techniques known in the art, such as those disclosed in "Remington: The Science and Practice of Pharmacy," Pharmaceutical Press, 22nd edition. Since it is preferably released into the bloodstream upon measurement of blood glucose concentration, the formulation can be formulated into dosage forms for parenteral administration, such as intramuscular, intravenous, subcutaneous, intra-arterial, or intracardiac administration. Dosage forms for parenteral administration include, for example, solutions, emulsions, suspensions, dispersions, and powders and granules for reconstitution. Emulsions are a preferred dosage form for parenteral administration.

[0195] Therefore, if the formulation is administered parenterally, examples of such administration include one or more of the following: intravenous, intra-arterial, intraperitoneal, intrathecal, intraventricular, intraurethral, ​​intrasternal, intracardiac, intramuscular, or subcutaneous administration, and / or administration via infusion techniques. For parenterally administration, formulations containing the polymers of the present invention are preferably used in the form of a sterile aqueous solution, which may contain other substances, such as sufficient salts or sugars, to make the solution isotonic with blood. If necessary, the aqueous solution should be appropriately buffered (preferably to physiological pH). The preparation of suitable parenterally formulations under sterile conditions is readily accomplished using standard pharmaceutical techniques.

[0196] The formulations of the present invention can also be formulated into sustained-release systems, which may include a semi-permeable polymer matrix in the form of a molded article, such as a film or microcapsule. The sustained-release matrix may include, for example, polylactide, a copolymer of L-glutamic acid and γ-ethyl-L-glutamic acid, poly(2-hydroxyethyl methacrylate), ethylene-vinyl acetate, or poly-D-(-)-3-hydroxybutyric acid.

[0197] Preferred treatments are for conditions / diseases / disorders related to pathological blood glucose concentrations. Therefore, in one embodiment, the present invention relates to the biosensor of the present invention or to a glucose concentration-sensitive release formulation for treating conditions / diseases / disorders dependent on blood glucose concentrations. An example of such a condition is diabetes, such as type 1 diabetes. As is known in the art, diabetes is typically treated by administering insulin to the patient. Therefore, the formulations of the present invention may also contain insulin.

[0198] However, preferably, the biosensor of the present invention is not intended for therapeutic applications and / or the biosensor of the present invention does not contain insulin.

[0199] The invention is illustrated by the following embodiments, which are for illustrative purposes only and should not be construed as limiting the scope of the claims.

[0200] Example

[0201] Names and abbreviations

[0202] 3APB: 3-Acryloylaminophenylboronic acid

[0203] 2APB: 2-Acryloylaminophenylboronic acid

[0204] ADAM: 5-Acryloylamino-2-((dimethylamino)methyl)phenylboronic acid

[0205] TMAPAA: (3-Acryloylaminopropyl)trimethylammonium chloride

[0206] PBS: Phosphate-buffered saline

[0207] GBM: Glucose-binding molecule

[0208] The structural formulas of some of the compounds discussed in the experimental section are provided below.

[0209]

[0210]

[0211] Materials and methods

[0212] chemicals

[0213] Acrylamide, methylenebisacrylamide, N-hydroxyethylacrylamide (HEAA), 1-hydroxycyclohexylphenyl ketone, 3-(trimethoxysilyl)propyl methacrylate, and squalane were purchased from Sigma-Aldrich. 5-Acryloylamino-2-((dimethylamino)methyl)phenylboronic acid (ADAM) and 2-acryloylaminophenylboronic acid (2APB) were purchased from Combi-Blocks Inc. 3-Acryloylaminophenylboronic acid (3APB), (3-acryloylaminopropyl)trimethylammonium chloride (TMAPAA), and N-(3-aminopropyl)methacrylamide hydrochloride (APMA) were purchased from Sigma-Aldrich. N-(3-(dimethylamino)propyl)acrylamide (DMAPAA) was purchased from Tokyo Chemical Industry Co., Ltd. D-(-)-fructose, D-mannitol, sodium L-lactic acid, and trisodium citrate dihydrate were purchased from Sigma-Aldrich. Dimethyl sulfoxide was purchased from Sigma-Aldrich.

[0214] D,L-homocysteine ​​thiolactone acrylamide was purchased from Specific Polymers. Ethanolamine was purchased from Sigma-Aldrich.

[0215] 0.01 M PBS pH 7.4 (0.132 M NaCl) was prepared internally. Disodium hydrogen phosphate was purchased from Alfa Aesar, sodium dihydrogen phosphate monohydrate was purchased from Sigma-Aldrich, and sodium chloride was purchased from Sigma-Aldrich.

[0216] All chemicals are used without further purification.

[0217] Manufacturing of hydrogel sensors

[0218] In typical formulations of hydrogel sensors, the monomers and molar percentages are as follows: acrylamide borate 20%, acrylamide 75%, and methylenebisacrylamide 1-2%. Exemplary compositions are shown in Table 1.

[0219] Dilute the monomer with deionized water (DI), or with 1 M fructose or mannitol in PBS (pH 7.4, 0.132 M NaCl) to a final total monomer concentration of 1.0 or 1.5 M. Add 1-hydroxycyclohexylphenyl ketone (photoinitiator) at a concentration of 1.5 mM.

[0220] Table 1. Exemplary sensor formulations discussed in this article.

[0221]

[0222] The fabrication of the hydrogel sensor follows WO 2007 / 104974 (which is incorporated herein by reference). First, optical glass fibers are stripped and cut to prepare uniform cuts. The fiber cuts are then silanized to covalently attach the hydrogel to the glass surface. Silanization involves first immersing the fiber cuts in hydrochloric acid (1.0 M) for 15 minutes, followed by washing with DI water, and then immersing in ethanol containing 3-(trimethoxysilyl)propyl methacrylate (84 mM) for 10 minutes. Excess 3-(trimethoxysilyl)propyl methacrylate is removed by washing the fiber cuts in flowing ethanol. A single drop of dome-shaped pregel solution is deposited onto the fiber cuts using a pipette, with both the cuts and the pregel contained in a larger drop of squalane oil. The oil is used primarily to maintain the stability of the pregel droplet and secondarily to contain an excess of dissolved photosensitizing initiator 1-hydroxycyclohexylphenyl ketone (132 mM). An excess of photoinitiator in the oil is necessary to enable polymerization to occur without the need for anaerobic conditions, as the resulting free radicals react first with dissolved oxygen in the oil. The higher concentration in the oil compared to pre-gelled droplets also ensures that the photoinitiator does not significantly leach from the pre-gelled droplets into the oil droplets.

[0223] The polymerization of the pregel occurs by irradiating the pregel with a 340 nm light source located directly in front of and inside the pregel droplet (for 5 minutes).

[0224] After polymerization, the sensor was briefly washed with pentane to remove oil, and then washed for 15 minutes with 50% ethanol in DI water to remove unreacted monomers. At this point, the sensor was placed in PBS at pH 6.0 until testing.

[0225] Sensor test setup

[0226] In a typical experiment, a test solution was prepared at 37°C using 0.01 M PBS containing 132 mM NaCl at pH 7.4. The sensor was immersed in the test solution and allowed to equilibrate for at least 10 minutes. After this, the gel chamber signal was locked to obtain the initial absolute length of the hydrogel, and the experiment began. At multiple time points, the glucose fraction from a 1 M stock solution of PBS, which had been in place for at least 6 hours to reach variogram equilibrium, was added to achieve the desired concentration, while continuously monitoring changes in gel length. The length change in response to pH was measured by moving the sensor between two test solutions with constant glucose concentration but different pH values.

[0227] Example 1

[0228] Results and discussion

[0229] Monoboronic acid-based glucose sensors typically exhibit poor selectivity for glucose relative to other monosaccharides (e.g., fructose) and show significant alterations in glucose binding even with small pH changes. In this paper, a sensor has been developed that demonstrates good selectivity for glucose relative to a range of common interfering substances while being minimally affected by pH changes.

[0230] One of the most commonly used monoboronic acids for glucose sensing is 3-acryloylaminophenylboronic acid (3APB), which has been incorporated into materials such as polymers to impart glucose-responsive properties. In this paper, 3APB is used as a reference example.

[0231] The boron center of 3APB has a pKa of approximately 8.8. 3 Furthermore, it exists primarily in a triangular-plane sp2 form at physiological pH, which is unfavorable for glucose binding. 4,5 As a result of the relatively high pKa, small changes from pH 7.4 to higher or lower pH significantly increased or decreased the degree of glucose binding, and thus could significantly affect sensor readings that require complex pH correction. 6,7 This pH sensitivity makes 3APB unsuitable for use in in vivo glucose sensing, where, in extreme cases, blood pH can be as low as 6.9 and as high as 7.6. Tests were conducted at pH 6.9, 7.4, and 7.6 with 3APB and positively charged comonomers (as described in various publications). 7,8,9,10 The glucose response of the sensor () Figure 2 A) and a large deviation was observed between the sensor reading and the reference glucose reading. To address the pH interference issue, the Wulff-type was selected. 2 5-Acryloylamino-2-((dimethylamino)methyl)phenylboronic acid (ADAM) is used as GBM to replace 3APB. Wulff-type boronic acid has an amino group adjacent to boronic acid, which reduces the pKa of boronic acid to approximately 5.2. 2,11 This facilitates glucose binding at physiological pH. Although Wulff-type boric acid has been successfully used in fluorescent glucose sensors... 12,13 However, the exact mechanism of the interaction between amino groups and boron remains controversial. 14,15

[0232] Despite their low pKa, Wulff-type boronic acids have been reported to have a lower affinity for glucose compared to other monoboronic acids. 11,16 This may be a result of spatial steric hindrance. 5In practice, it was found that sensors constructed with ADAM required 2.5–3.5 times the mol% of GBM to produce a glucose response similar to that of 3APB. Nevertheless, by appropriately optimizing the ADAM concentration, a highly linear change (contraction) in the length of the ADAM hydrogel sensor was achieved, which is suitable for glucose detection within the physiological range (Figure 3A). In contrast to the 3APB sensor, the ADAM-based sensor does not require the addition of any cationic monomers to produce a linear contraction due to glucose (Figure 3). 9

[0233] Importantly, the ADAM sensor exhibited significantly lower pH dependence compared to the 3APB sensor. Figure 2 This indicates the suitability of the ADAM sensor for in vivo glucose monitoring.

[0234] The selectivity of the ADAM sensor for glucose relative to other common interfering substances was found to increase with the addition of the quaternary ammonium comonomer TMAPAA (Figure 3). Other research groups have reported similar effects when using cationic comonomers with 3APB, where a positive charge is required to achieve an acceptable glucose response with 3APB at physiological pH. 17,18,10

[0235] Compared to other examples, a much smaller ratio of cationic monomer to boric acid is required to almost completely suppress interference from fructose. Figure 2 B). 17,18,10

[0236] The addition of TMAPAA had no significant effect on the sensor's glucose response (Figure 3A), and importantly, the low mol% of cationic groups in the hydrogel was crucial for avoiding unwanted interference from salts (Figure 3D). The 3% TMAPAA and 20% ADAM sensors showed negligible interference from fructose, mannitol, lactate, and citrate, and further reduced interference from pH changes from 6.9 to 7.6; however, the tested concentrations of interfering substances were far higher than the expected concentrations in vivo (Figure 3). Figure 4 This indicates that selectivity is generally not observed for monoboronic acids. The observed selectivity is presumably partly driven by the ability of ADAM hydrogel to form a 2:1 complex with furanose. 1

[0237] 2APB has recently been studied as GBM, and due to the internal coordination of boron with the adjacent carbonyl group, it provides significantly lower pH interference than 3APB. 19,20,21The sensor constructed with 2APB exhibited slow kinetics with glucose, resulting in modest contraction at 6 and 20 mM glucose, but with minimal interference from lactate, citrate, and pH changes. Surprisingly, the 2APB sensor showed large and rapid contraction at 5 mM mannitol. Figure 4 Compared to the ADAM or 3APB sensors, the 2APB sensor exhibited a significantly larger mannitol response relative to glucose or fructose (Table 2). The origin of this response is unclear, but it can be attributed to the crosslinking between 2APB and mannitol. 22 The 2APB sensor shrinks due to mannitol, while the ADAM sensor swells. Therefore, it is proposed that adding a certain mol% of 2APB to the ADAM sensor can selectively suppress mannitol interference. By changing the molar ratio of ADAM to 2APB, it was found that using 3% 2APB and 25% ADAM can suppress mannitol interference at 2.2 mM glucose. Figure 5 ).

[0238] Table 2. Response of sensors based on 3APB, ADAM and 2APB to length changes of glucose, fructose and mannitol.

[0239]

[0240] Example 2

[0241] Following the methods summarized above, this embodiment tested the sensitivity of sensors containing the following polymer hydrogels to glucose, as summarized in Table 3.1 below:

[0242] Table 3.1. Hydrogels obtained in Example 2.

[0243]

[0244] The results are as follows Figure 7A As shown in the figure. The conclusion is that, for the same monomer ratio, the length change of the ADAM-acrylamide sensor at 20 mM glucose is approximately 1.6 times that of the ADAM-HEAA sensor. Therefore, the sensor with acrylamide exhibits higher sensitivity to glucose. Notably, in both cases (acrylamide and HEAA), the sensor shrinks with increasing glucose levels.

[0245] This experiment was repeated for polymer hydrogels that do not contain TMAPAA. These hydrogels were obtained according to the method summarized above and are summarized in Table 3.22 below.

[0246] Table 3.2. Other hydrogels obtained in Example 2.

[0247]

[0248] The results are as follows Figure 7B As shown in the figure. The conclusion is that, for the same monomer ratio, the length change of the ADAM-acrylamide sensor at 20 mM glucose is approximately twice that of the ADAM-HEAA sensor. Therefore, the sensor with acrylamide exhibits higher sensitivity to glucose. Notably, in both cases (acrylamide and HEAA), the sensor shrinks with increasing glucose levels.

[0249] Example 3

[0250] Three different cationic monomers were introduced into the polymer gel formulation, and the resulting biosensor's ability to withstand interference effects derived from the presence of citrate was tested. The introduced monomers are shown below:

[0251]

[0252] The obtained hydrogels are characterized in Table 4 below. The total monomer concentration in all formulations was 1.5 M.

[0253] Table 4. Polymer hydrogels tested in Example 3.

[0254]

[0255] The swelling measurements performed using these hydrogels are summarized below. Figure 8 In this study, sensors lacking cationic comonomers swelled significantly with increasing citrate concentrations, indicating citrate interference, which is undesirable for clinically relevant measurements. The cationic monomers TMAPAA, DMAPAA, and APMA were found to reduce citrate interference without introducing significant alterations in the sensor's response to glucose.

[0256] Example 4

[0257] A polymer hydrogel containing the ADAM moiety was prepared according to the method for obtaining polymer hydrogels, and then modified to obtain the polymer hydrogel of the present invention. 23

[0258] A monomer solution containing 16 or 28 mol% thiolactone acrylamide was photopolymerized to form the corresponding hydrogel. The hydrogel sensor was then incubated in a 1:1 DMSO and PBS solution containing 26 mM olefin (ADAM), and ethanolamine (final concentration 5 M) was added after a short time period to initiate the reaction. The reaction was maintained overnight at room temperature with stirring. Afterward, the sensor was washed with 50% ethanol in DI water for 15 minutes before testing.

[0259]

[0260] The obtained hydrogels are characterized in Table 5 below.

[0261] Table 5. Polymer hydrogels tested in Example 4.

[0262]

[0263] The results are summarized in Figure 9. ADAM sensors can also be constructed through modification after thiol-olefin polymerization. As shown in Figure 9, compared with sensors produced by direct polymerization (… Figure 9B Compared to the modified sensor, a greater contraction effect of glucose was observed. Figure 9A However, it is reiterated that very similar behavior was observed for both types of sensors.

[0264] References

[0265]

[0266] .

Claims

1. A biosensor for measuring glucose concentration, the biosensor comprising a polymer containing a portion of formula (I): The portion of formula (I) is fixed to the polymer. Where R is independently C 1-5 Alkyl, C 2-5 alkenyl or C 2-5 alkynyl group, Where R S Each was independently selected from C 1-5 Alkyl, C 2-5 alkenyl, C 2-5 alkynyl group, -(C 0-3 alkylene)-OH, -(C 0-3 alkylene)-O(C 1-5 alkyl), -(C 0-3 alkylene)-O(C 1-5 alkylene)-OH, -(C 0-3 alkylene)-O(C 1-5 alkylene)-O(C 1-5 alkyl), -(C 0-3 alkylene)-SH, -(C 0-3 alkylene)-S(C 1-5 alkyl), -(C 0-3 alkylene)-S(C 1-5 alkylene)-SH, -(C 0-3 alkylene)-S(C 1-5 alkylene)-S(C 1-5 alkyl), -(C 0-3 alkylene)-NH2, -(C 0-3 alkylene)-NH(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-OH, -(C 0-3 alkylene)-N(C 1-5 alkyl)-OH, -(C 0-3 alkylene)-NH-O(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-O(C 1-5 alkyl), -(C 0-3 alkylene)-halogen, -(C 0-3 alkylene)-(C 1-5 Halogenated alkyl), -(C 0-3 alkylene)-O-(C 1-5 Halogenated alkyl), -(C 0-3 alkylene)-CN, -(C 0-3 alkylene)-NO2, -(C 0-3 alkylene)-CHO, -(C 0-3 alkylene)-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-COOH, -(C 0-3 alkylene)-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-O-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-CO-NH2, -(C 0-3 alkylene)-CO-NH(C 1-5 alkyl), -(C 0-3 alkylene)-CO-N(C 1-5 Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-CO-(C 1-5 alkyl), -(C 0-3 alkylene)-NH-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-CO-O-(C 1-5 alkyl), -(C 0-3 alkylene)-O-CO-NH-(C 1-5 alkyl), -(C 0-3 alkylene)-O-CO-N(C 1-5 alkyl)-(C 1-5 alkyl), -(C 0-3 alkylene)-SO2-NH2, -(C 0-3 alkylene)-SO2-NH(C 1-5 alkyl), -(C 0-3 alkylene)-SO2-N(C 1-5 Alkyl)(C 1-5 alkyl), -(C 0-3 alkylene)-NH-SO2-(C 1-5 alkyl), -(C 0-3 alkylene)-N(C 1-5 alkyl)-SO2-(C 1-5 alkyl), -(C 0-3 alkylene)-SO2-(C 1-5 alkyl), -(C 0-3 alkylene)-SO-(C 1-5 alkyl), -(C 0-3 alkylene)-carbocyclic and -(C 0-3 alkylene)-heterocyclic group, wherein -(C 0-3 The carbocyclic moiety in the alkylene group and the -(C 0-3 In each of the alkylene-heterocyclic groups, the heterocyclic moiety is optionally substituted by one or more groups, said groups being independently selected from C10. 1-4 Alkyl, halogen, -CN, -NO2, -OH, -O-(C 1-4 Alkyl), -SH, -S-(C 1-4 Alkyl groups, -NH2, -NH(C) 1-4 alkyl), -N(C) 1-4 Alkyl)(C 1-4 Alkyl groups, -COOH, -COO (C 1-4 Alkyl group), -CONH2, -CONH(C 1-4 Alkyl), -CON(C) 1-4 Alkyl)(C 1-4 alkyl), -NHCO(C 1-4 alkyl) and -N(C) 1-4 alkyl)-CO(C 1-4 alkyl); And n can be 0, 1, 2 or 3.

2. The biosensor according to claim 1, wherein each of R is a methyl group.

3. The biosensor according to claim 1 or 2, wherein n is 0 or 1, preferably n is 0.

4. The biosensor according to any one of claims 1-3, wherein the polymer comprises the portion of formula (Ia): Among them, R, R S And n as described in any one of claims 1-3.

5. The biosensor according to any one of claims 1-4, wherein the polymer is a polymer hydrogel.

6. The biosensor according to any one of claims 1-5, wherein the polymer is a polymer based on unsubstituted polyacrylamide.

7. The biosensor according to any one of claims 1-6, wherein the polymer further comprises a positively charged portion at pH=7.

4.

8. The biosensor according to any one of claims 1-6, wherein the polymer further comprises a portion containing a group selected from: -NH2, -NH3 + -NH(C 1-5 alkyl), -(NH2(C 1-5 alkyl)) + -N(C 1-5 alkyl)2、-(NH(C) 1-5 Alkyl)2) + and -N(C 1-5 Alkyl)3 + .

9. The biosensor according to any one of claims 1-6, wherein the polymer further comprises a portion of formula (II): 。 10. The biosensor according to any one of claims 1-9, wherein the polymer further comprises a portion of formula (III): 。 11. The biosensor according to any one of claims 1-10, wherein the polymer is obtained in the polymerization reaction of the composition, the composition comprising: (i) an acrylamide monomer comprising a boric acid-based glucose-binding moiety of formula (I), Preferably, (i) is: Among them, R, R S And n as described in any one of claims 1-10.

12. The biosensor of claim 11, wherein the composition for the polymerization reaction further comprises: (ii) Acrylamide monomers without the boric acid moiety, and (iii) Crosslinking agent acrylamide monomer, Preferably, (ii) comprises monomers selected from: and Where q is an integer from 1 to 10; and / or Wherein (iii) contains monomers selected from the following: and , where q is an integer from 1 to 10.

13. The biosensor according to any one of claims 1-12, wherein the polymer shrinks in the presence of glucose, preferably wherein the shrinkage rate is substantially linearly proportional to the glucose concentration.

14. The biosensor of claim 1, wherein each of R is methyl, n is 0, and the polymer is a polymer based on unsubstituted polyacrylamide.

15. The biosensor of claim 1, wherein each of R is methyl, n is 0, and the polymer further comprises a positively charged portion at pH 7.

4.

16. The biosensor of claim 1, wherein each of R is methyl, n is 0, and the polymer further comprises a moiety containing groups selected from -NH2, -NH3. + -NH(C 1-5 alkyl), -(NH2(C 1-5 alkyl)) + -N(C 1-5 alkyl)2、-(NH(C) 1-5 Alkyl)2) + and -N(C 1-5 Alkyl)3 + .

17. The biosensor according to any one of claims 1-16, wherein the glucose-dependent shrinkage rate is substantially independent of pH, preferably, substantially independent of pH for a pH range of 6.9-7.6, more preferably 7.4-7.

6.

18. The biosensor according to any one of claims 1-17, used for diagnostics.

19. The biosensor according to any one of claims 1-17, used for glucose level monitoring.

20. The biosensor for the said use according to claim 19, wherein glucose level monitoring is performed on an individual in intensive care, and / or wherein glucose level monitoring is performed on an unconscious individual.

21. The polymer as defined in any one of claims 1-16.

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