Biocompatible materials and methods of making and using same

Abstract

To provide a composition containing at least one or more polymers capable of forming hydrogel, as well as methods of manufacturing and using the hydrogel.SOLUTION: For example, a composition can include one or more hydrogel-forming polymers having an intrinsic viscosity [η] (e.g., as measured by an Ubbelohde viscometer) of at least 3 dL / g in the composition. A concentration CT of the hydrogel-forming polymer in the composition can be at most about 5 mg / ml. More specifically, the present disclosure provides compositions that include polymers (e.g., biocompatible polymers) capable of forming soft hydrogels, and methods of making and using the same.SELECTED DRAWING: None

Description

[Background technology]

[0001] Many polymers used in biomedical applications are water-soluble polymers with hydroxyl groups. Examples of water-soluble polymers with hydroxyl groups include hyaluronic acid (HA), polyethylene glycol (PEG), polyvinyl alcohol, alginate, cyclodextrin, etc. Water-soluble polymers with hydroxyl groups are generally non-toxic, and the hydroxyl groups generally make these polymers soluble in aqueous environments. However, water-soluble polymers with hydroxyl groups usually do not have functionality in animal cells or tissues by themselves and must be modified to exhibit desired properties.

[0002] For example, hydroxyl-bearing water-soluble polymers can be modified for use as hydrogels. The modification of hydroxyl-bearing water-soluble polymers for use in hydrogels has traditionally been expensive and involves complex chemistry and harsh conditions that may not be suitable for biomedical applications. Summary of the Invention [Problem to be solved by the invention]

[0003] Therefore, there is a need to create hydrogels with desired properties that contain polymers with appropriately modified hydroxyls. [Means for solving the problem]

[0004] The present disclosure provides compositions comprising polymers (e.g., biocompatible polymers) capable of forming hydrogels, as well as methods of making and using the same. For example, the compositions can include one or more hydrogel-forming polymers having an intrinsic viscosity [η] (e.g., as measured by an Ubbelohde viscometer) of at least 3 dL / g in the composition. The concentration C of the hydrogel-forming polymer in the composition is Tmay be at most about 5 mg / ml. More specifically, the present disclosure provides compositions including polymers (e.g., biocompatible polymers) capable of forming soft hydrogels, as well as methods of making and using the same. Hydrogels formed according to the present disclosure can have a relatively low G' (e.g., G' less than about 10.0 Pa), a high G' relative to G'' (e.g., G'' / G' < 1), while having a relatively large yield strain (e.g., 10% or greater). Thus, hydrogels formed according to the present disclosure are suitable for application to surfaces. In addition, hydrogels formed according to the present disclosure can be self-healing. Hydrogels of the present disclosure can have low viscosities at high shear rates (e.g., complex viscosities of about 0.2 Pa·s or less), demonstrating that they can be easily applied to an entire surface with little force. Additionally, it has been shown that the viscosity of hydrogels according to the present disclosure can increase exponentially with low shear rates and stabilize at rest. Furthermore, the hydrogels of the present disclosure can be capable of moving freely in water without dissolving, and can exhibit this property even after being immersed in water for extended periods of time (e.g., 24 hours or more).

[0005] In one aspect, the present disclosure provides a composition comprising a polymer capable of forming a hydrogel, wherein the polymer is present in a concentration of about 0.1 mg / ml to about 5 mg / ml (e.g., about 0.2 mg / ml to about 5 mg / ml, about 0.3 mg / ml to about 5 mg / ml, about 0.4 mg / ml to about 5 mg / ml, about 0.5 mg / ml to about 5 mg / ml, about 0.6 mg / ml to about 5 mg / ml, about 0.7 mg / ml to about 5 mg / ml, about 0.8 mg / ml to about 5 mg / ml, about 0.9 mg / ml to about 5 mg / ml, about 1.0 mg / ml to about 5 mg / ml, about 1.0 mg / ml to about 4 mg / ml, about 1.0 mg / ml to about 3 mg / ml, about 1.0 mg / ml to about 2 mg / ml, about 0.3 mg / ml to about 0.8 mg / ml, about 0.3 mg / ml to about 0.6 ... The composition has a concentration C of about 0.3 mg / ml to about 0.5 mg / ml.

[0006] In another aspect, the present disclosure provides a composition comprising one or more hydrogel-forming polymers having an intrinsic viscosity [η] of at least 3 dL / g (e.g., at least 5 dL / g, at least 8 dL / g, at least 10 dL / g, at least 12 dL / g, at least 15 dL / g, at least 16 dL / g, at least 17 dL / g, at least 18 dL / g, at least 19 dL / g, at least 20 dL / g, at least 25 dL / g, or more) in the composition, wherein the concentration C of such hydrogel-forming polymer in the composition T is at most about 5 mg / ml (e.g., at most about 4 mg / ml, at most about 3 mg / ml, at most about 2 mg / ml, at most about 1.5 mg / ml, at most about 1 mg / ml, at most about 0.9 mg / ml, at most about 0.8 mg / ml, at most about 0.7 mg / ml, at most about 0.6 mg / ml, at most about 0.5 mg / ml, at most about 0.4 mg / ml, at most about 0.3 mg / ml, at most about 0.2 mg / ml, at most about 0.1 mg / ml, or less). The intrinsic viscosity [η] can be as measured by an Ubbelohde viscometer.

[0007] In another aspect, the present disclosure provides a composition comprising one or more polymers capable of forming a hydrogel, wherein the concentration C (e.g., total concentration) of the one or more polymers (e.g., hydrogel-forming polymers) in the composition is at most about 5 mg / ml (e.g., at most about 4 mg / ml, at most about 3 mg / ml, at most about 2 mg / ml, at most about 1.5 mg / ml, at most about 1 mg / ml, at most about 0.9 mg / ml, at most about 0.8 mg / ml, at most about 0.7 mg / ml, at most about 0.6 mg / ml, at most about 0.5 mg / ml, at most about 0.4 mg / ml, at most about 0.5 mg / ml, at most about 0.6 mg / ml, at most about 0.7 mg / ml, at most about 0.8 mg / ml, at most about 0.9 mg / ml, at most about 0.8 mg / ml, at most about 0.7 mg / ml, at most about 0.6 mg / ml, at most about 0.5 mg / ml, at most about 0.4 mg / ml, at most about 0.5 mg / ml, at most about 0.6 ...6 mg / ml, at most about 0.5 mg / ml, at most about 0.6 mg / ml, at most about 0 and wherein at least one of the one or more polymers has an intrinsic viscosity [η] of at least about 3 dL / g (e.g., at least about 5 dL / g, at least about 8 dL / g, at least about 10 dL / g, at least about 12 dL / g, at least about 15 dL / g, at least about 16 dL / g, at least about 17 dL / g, at least about 18 dL / g, at least about 19 dL / g, at least about 20 dL / g, at least about 25 dL / g, or more) in the composition.

[0008] In some embodiments, the one or more polymers each have an intrinsic viscosity [η] in the composition of at least 3 dL / g (e.g., at least about 5 dL / g, at least about 8 dL / g, at least about 10 dL / g, at least about 12 dL / g, at least about 15 dL / g, at least about 16 dL / g, at least about 17 dL / g, at least about 18 dL / g, at least about 19 dL / g, at least about 20 dL / g, at least about 25 dL / g, or more).

[0009] In some embodiments, the concentration C of the polymer in the composition is less than or equal to concentration C * about 0.8 to about 5 times (for example, about 0.8 to about 4.5 times, about 0.8 to about 4.0 times, about 0.8 to about 3.5 times, about 0.8 to about 3.0 times, about 0.8 to about 2.5 times, about 0.8 to about 2.0 times, about 0.8 to about 1.5 times, about 1.0 to about 1.2 times, about 1.8 to about 2.0 times, or about 1.0 to about 2.0 times) of C* = 1 / ([η]), where [η] is the intrinsic viscosity of the polymer.

[0010] In some embodiments, the polymer in the composition has an [η] of at least 3 dL / g (e.g., at least 5 dL / g, at least 8 dL / g, at least 10 dL / g, at least 12 dL / g, at least 15 dL / g, at least 16 dL / g, at least 17 dL / g, at least 18 dL / g, at least 19 dL / g, at least 20 dL / g, at least 25 dL / g, or more).

[0011] In some embodiments, at least some of the hydrogel-forming polymers is contained in the formed hydrogel in the composition.

[0012] The hydrogel may have at least one of the following: 1) a storage modulus G' of about 10.0 Pa or less (e.g., about 8.0 Pa or less, about 7.0 Pa or less, about 6.0 Pa or less, about 5.0 Pa or less, about 4.0 Pa or less, about 3.0 Pa or less, about 2.0 Pa or less, about 1.0 Pa or less, about 0.8 Pa or less, about 0.7 Pa or less, about 0.6 Pa or less, about 0.5 Pa or less, or less), as measured in a dynamic oscillatory shear test; 2) a loss modulus G'' of about 10.0 Pa or less (e.g., about 8.0 Pa or less, about 7.0 Pa or less, about 6.0 Pa or less, about 5.0 Pa or less, about 4.0 Pa or less, about 3.0 Pa or less, about 2.0 Pa or less, about 1.0 Pa or less, about 0.8 Pa or less, about 0.7 Pa or less, about 0.6 Pa or less, about 0.5 Pa or less, or less), as measured in a dynamic oscillatory shear test; 3) A compressive strength of about 0.2 Pa·s or less (e.g., about 0.2 Pa·s or less) measured in a dynamic oscillatory shear test at a frequency of less than about 100 rad / s (e.g., less than about 90 rad / s, less than about 80 rad / s, less than about 70 rad / s, less than about 50 rad / s, less than about 40 rad / s, less than about 30 rad / s, less than about 20 rad / s, less than about 10 rad / s, or less). a complex viscosity of 0.1 Pa·s or less, about 0.08 Pa·s or less, about 0.07 Pa·s or less, about 0.06 Pa·s or less, about 0.05 Pa·s or less, or about 0.04 Pa·s or less; 4) Measured in a dynamic oscillatory strain sweep test A yield strain of at least about 10% (e.g., at least about 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.2%, or 0.1%) is achieved.

[0013] In some embodiments, the hydrogel has a loss modulus G'' that is less than or equal to about 100% of its storage modulus G', as measured in a dynamic oscillatory shear test.

[0014] In some embodiments, the polymer is hydrophilic and / or water-soluble.

[0015] In some embodiments, the polymer is selected from the group consisting of polysaccharides, poly(acrylic acid), poly(hydroxyethyl methacrylate), elastin, collagen, derivatives thereof, and any combination thereof. For example, the polymer is selected from the group consisting of poly(acrylic acid), poly(hydroxyethyl methacrylate), derivatives thereof, and any combination thereof.

[0016] In some embodiments, the polymer is selected from the group consisting of hyaluronic acid, guar gum, starch, chitosan, chondroitin sulfate, alginate, carboxymethylcellulose, derivatives thereof, and any combination thereof. For example, the polymer is selected from the group consisting of hyaluronic acid, guar gum, alginate, carboxymethylcellulose, derivatives thereof, and any combination thereof.

[0017] In some embodiments, the polymer is selected from the group consisting of hyaluronic acid, its derivatives, and any combination thereof.

[0018] In some embodiments, the polymer is modified (e.g., polymer derivative) with one or more modifications selected from the group consisting of acrylate, maleimide, vinyl sulfone, N-hydroxysuccinimide, aldehyde, ketone, carbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, amine, and any combination thereof. For example, the polymer is modified with one or more modifications selected from the group consisting of maleimide, vinyl sulfone, N-hydroxysuccinimide, aldehyde, mercaptonicotinamide, quinone, thiol, amine, and any combination thereof. It is modified by the modification of

[0019] For example, the polymer can include derivatives modified with one or more modifications selected from the group consisting of acrylate, maleimide, vinyl sulfone, N-hydroxysuccinimide, aldehyde, ketone, carbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, amine, and any combination thereof. For example, the polymer can include derivatives modified with one or more modifications selected from the group consisting of maleimide, vinyl sulfone, N-hydroxysuccinimide, aldehyde, mercaptonicotinamide, quinone, thiol, amine, and any combination thereof.

[0020] The derivatives can have an average degree of modification (DM) of about 3% to about 50% (e.g., about 4% to about 45%, about 5% to about 40%, about 6% to about 40%, about 7% to about 40%, about 8% to about 39%, about 8% to about 38%, about 8% to about 35%, about 9% to about 32%, about 8% to about 30%, about 10% to about 30%, about 12% to about 30%, about 13% to about 30%, about 14% to about 30%, about 15% to about 35%, or about 15% to about 30%).

[0021] In some embodiments, the polymers include a first polymer population including polymers modified with at least a first modification and a second polymer population including polymers modified with a second modification, where the first modification is different from the second modification, and the polymers of the first polymer population react with the polymers of the second polymer population to form a hydrogel. In some embodiments, the ratio of polymers in the first polymer population to polymers in the second polymer population is about 10:1 to about 1:10 (e.g., 10:2, 10:3, 10:4, 10:5, 10:6, 10:7, 10:8, 10:9, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, or 1:10). In some embodiments, the ratio of polymers in the first polymer population to polymers in the second polymer population is about 3:1 to about 1:3. For example, the ratio of polymers in the first polymer population to polymers in the second polymer population can be from about 2:1 to about 1:2.

[0022] The ratio can be a mass ratio, a molar ratio, a volume ratio, and / or a DM ratio. For example, the ratio can be about 3:1 to about 1:3, or about 2:1 to about 1:2, e.g., about 1:1. The DM ratio refers to the ratio of the average DM of the polymers in the first polymer population to the average DM of the polymers in the second polymer population.

[0023] In some embodiments, the composition comprises at least a first polymer derivative and a second polymer derivative, wherein the first polymer derivative comprises a first modification and the second polymer derivative comprises a second modification, wherein the first modification is different from the second modification, and wherein the first polymer derivative is capable of reacting with the second polymer derivative to form the hydrogel.

[0024] The mass ratio of the first polymer derivative to the second polymer derivative in the composition can be from about 10:1 to about 1:10 (e.g., 10:2, 10:3, 10:4, 10:5, 10:6, 10:7, 10:8, 10:9, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9 or 1:10), for example, from about 3:1 to about 1:3 or from about 2:1 to about 1:2, for example, about 1:1.

[0025] The molar ratio of the first polymer derivative to the second polymer derivative in the composition can be from about 10:1 to about 1:10 (e.g., 10:2, 10:3, 10:4, 10:5, 10:6, 10:7, 10:8, 10:9, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9 or 1:10), for example, from about 3:1 to about 1:3 or from about 2:1 to about 1:2, for example, about 1:1.

[0026] The volume ratio of the first polymer derivative to the second polymer derivative in the composition can be about 10:1 to about 1:10 (e.g., 10:2, 10:3, 10:4, 10:5, 10:6, 10:7, 10:8, 10:9, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9 or 1:10), for example, about 3:1 to about 1:3 or about 2:1 to about 1:2, for example, about 1:1.

[0027] In some embodiments, the first polymer derivative comprises a first DM (DM1) and the second polymer derivative comprises a second DM (DM2), and the ratio of the first DM (DM1) to the second DM (DM2) is about 10:1 to about 1:10 (e.g., 10:2, 10:3, 10:4, 10:5, 10:6, 10:7, 10:8, 10:9, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9, or 1:10), such as about 3:1 to about 1:3, or about 2:1 to about 1:2, such as about 1:1.

[0028] In some embodiments, the first modification and the second modification are independently selected from the group consisting of acrylate, maleimide, vinyl sulfone, N-hydroxysuccinimide, aldehyde, ketone, carbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, amine, and any combination thereof.

[0029] In some embodiments, the first modification and the second modification are independently selected from the group consisting of maleimide, vinyl sulfone, N-hydroxysuccinimide, aldehyde, mercaptonicotinamide, quinone, thiol, amine, and any combination thereof.

[0030] In some embodiments, the polymer is hyaluronic acid, the first polymer population comprises hyaluronic acid modified with one or more vinyl sulfone groups, and the second polymer population comprises hyaluronic acid modified with one or more thiol groups, and the polymers of the first polymer population are capable of reacting with the polymers of the second polymer population to form a hydrogel.

[0031] For example, one or more polymers may be hyaluronic acid derivatives, where a first polymer derivative may be hyaluronic acid modified with one or more vinyl sulfone groups and a second polymer derivative may be hyaluronic acid modified with one or more thiol groups, and the first polymer derivative may react with the second polymer derivative to form a hydrogel.

[0032] In some embodiments, the polymer has a molecular weight of about 100,000 Daltons to about 5,000,000 Daltons (e.g., about 120,000 Daltons to about 5,000,000 Daltons, about 200,000 Daltons to about 5,000,000 Daltons, about 300,000 Daltons to about 5,000,000 Daltons, about 400,000 Daltons to about 5,000,000 Daltons, about 500,000 Daltons to about 5,000,000 Daltons). About 5,000,000 daltons, about 600,000 daltons to about 5,000,000 daltons, about 670,000 daltons to about 5,000,000 daltons, about 1,000,000 daltons to about 5,000,000 daltons, about 1,500,000 daltons to about 5,000,000 daltons, about 2,000,000 daltons to about 5,000,000 daltons, about 2,500,000 daltons to about 50,000 00 daltons, about 2,600,000 daltons to about 5,000,000 daltons, about 3,000,000 daltons to about 5,000,000 daltons, about 3,500,000 daltons to about 5,000,000 daltons, about 3,600,000 daltons to about 5,000,000 daltons, about 2,000,000 daltons to about 4,000,000 daltons, about 2,500,000 daltons to about 3,500,000 daltons ton, about 2,600,000 daltons to about 3,600,000 daltons, about 1,000,000 daltons to about 2,600,000 daltons, about 800,000 daltons to about 2,600,000 daltons, about 700,000 daltons to about 2,500,000 daltons, about 670,000 daltons to about 2,600,000 daltons, or about 600,000 daltons to about 2,500,000 daltons) It has a molecular weight.

[0033] In some embodiments, the polymer includes a modified polymer (polymer derivative) that can have an average degree of modification of about 3% to about 50% (about 4% to about 45%, about 5% to about 40%, about 6% to about 40%, about 7% to about 40%, about 8% to about 39%, about 8% to about 38%, about 8% to about 35%, about 9% to about 32%, about 8% to about 30%, about 10% to about 30%, about 12% to about 30%, about 13% to about 30%, about 14% to about 30%, about 15% to about 35%, or about 15% to about 30%).

[0034] In some embodiments, the composition further comprises a crosslinker (e.g., a crosslinker different from the polymer in the composition). The crosslinker can be a small molecule crosslinker, a polymeric crosslinker, or any combination thereof.

[0035] For example, the crosslinker can be a small molecule crosslinker including molecules having groups selected from acrylate, maleimide, vinyl sulfone, hydroxysuccinimide, aldehyde, ketone, multi-carbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, amine, and any combination thereof.

[0036] In some embodiments, the crosslinker is a polymeric crosslinker comprising a polymer having groups selected from acrylate, maleimide, vinyl sulfone, hydroxysuccinimide, aldehyde, ketone, multicarbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, amine, and combinations thereof.

[0037] In some embodiments, the cross-linking agent is selected from the group consisting of dithiothreitol, dicysteine, PEG-dithiol, 4- or 8-arm PEG thiol, divinyl sulfone, bis(vinylsulfonyl)methane, PEG-VS, and 4- or 8-arm PEGVS.

[0038] In some embodiments, the polymer is modified with one or more vinyl sulfone groups and the crosslinker is a thiol-containing molecule, e.g., the crosslinker can be selected from the group consisting of dithiothreitol, dicysteine, PEG-dithiol, and 4- or 8-arm PEG thiol.

[0039] In some embodiments, the polymer is modified with one or more thiol groups and the crosslinker is a vinyl sulfone-containing molecule, e.g., the crosslinker is selected from the group consisting of divinyl sulfone, bis(vinylsulfonyl)methane, PEG-VS, and 4- or 8-arm PEGVS.

[0040] In some embodiments, the composition does not include any crosslinkers that are different from the polymers in the composition, for example, the composition does not include small molecule crosslinkers or PEG-based crosslinkers.

[0041] In some embodiments, the composition does not include any additional crosslinking agent other than the polymer of the present disclosure.

[0042] In some embodiments, the composition comprises a buffer, which may be a phosphate buffer.

[0043] In some embodiments, the composition has a pH of about 3.5 to about 9.0 (e.g., about 4.0 to about 9.0, about 4.5 to about 9.0, about 5.0 to about 9.0, about 5.5 to about 9.0, about 6.0 to about 9.0, about 6.5 to about 9.0, about 7.0 to about 9.0, about 7.5 to about 9.0, about 8.0 to about 9.0, about 8.5 to about 9.0, about 7.0 to about 7.8, or about 7.4).

[0044] In another aspect, the present disclosure relates to hydrogels formed by the compositions of the present disclosure.

[0045] In some embodiments, the hydrogel is biocompatible.

[0046] In some embodiments, the hydrogel has a storage modulus G' of about 10.0 Pa or less (e.g., about 8.0 Pa or less, about 7.0 Pa or less, about 6.0 Pa or less, about 5.0 Pa or less, about 4.0 Pa or less, about 3.0 Pa or less, about 2.0 Pa or less, about 1.0 Pa or less, about 0.8 Pa or less, about 0.7 Pa or less, about 0.6 Pa or less, about 0.5 Pa or less, or less), as measured in a dynamic oscillatory shear test.

[0047] In some embodiments, the hydrogel has a loss modulus G'' that is about 100% or less (e.g., about 90% or less, about 80% or less, about 70% or less, about 60% or less, about 55% or less, about 50% or less, about 45% or less, about 40% or less, about 35% or less, about 30% or less, about 25% or less, or about 20% or less) of its storage modulus G', as measured in a dynamic oscillatory shear test.

[0048] In some embodiments, the hydrogel has a complex viscosity of about 0.2 Pa·s or less (e.g., about 0.1 Pa·s or less, about 0.08 Pa·s or less, about 0.07 Pa·s or less, about 0.06 Pa·s or less, about 0.05 Pa·s or less, or about 0.04 Pa·s or less), measured in a dynamic oscillatory shear test at a frequency greater than about 100 rad / s.

[0049] In some embodiments, the hydrogel has a yield strain of at least about 10% (e.g., at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, at least about 100%, at least about 110%, or more) as measured in a dynamic oscillatory strain sweep test.

[0050] In another aspect, the present disclosure relates to methods of making hydrogels (such as the hydrogels of the present disclosure), which may include: a) providing a composition of the present disclosure and exposing the composition to conditions that allow for the formation of a hydrogel.

[0051] In some embodiments, exposing comprises incubating the composition at about 30°C to about 45°C (eg, about 32°C to about 40°C, about 35°C to about 40°C, eg, about 37°C).

[0052] In some embodiments, the method can include: 1) preparing a first polymer population and a second polymer population (e.g., the polymers in the first polymer population can comprise hyaluronic acid modified with one or more vinyl sulfone groups, having a molecular weight of about 2,600,000 daltons and a degree of modification of about 30%, and the polymers in the second polymer population can comprise hyaluronic acid modified with one or more thiol groups, having a molecular weight of about 2,600,000 daltons and a degree of modification of about 30%) in water and adjusting the pH to about 7.4 (e.g., by adding a buffer solution); 2) mixing the polymers of the first polymer population with the polymers of the second polymer population in a ratio (e.g., by mass) of about 1:1 (wherein the concentration C (e.g., total concentration) of the polymers is about 0.3 mg / ml to 0.5 mg / ml); and 3) incubating the mixture at about 37° C. for about 24 hours.

[0053] In some embodiments, the method includes: 1) preparing a first polymer population and a second polymer population (e.g., the polymers in the first polymer population comprise hyaluronic acid modified with one or more vinyl sulfone groups, having a molecular weight of about 2,600,000 daltons and a degree of modification of about 8%); The method may include: 1) preparing a first polymer population and a second polymer population (wherein the polymers in the second polymer population may comprise hyaluronic acid modified with one or more thiol groups, having a molecular weight of about 2,600,000 daltons and a degree of modification of about 8%) in water and adjusting the pH to about 7.4 (e.g., by adding a buffer solution); 2) mixing the polymers of the first polymer population with the polymers of the second polymer population in a ratio (e.g., by mass) of about 1:1 (wherein the concentration (e.g., total concentration) C of the polymers is about 0.3 mg / ml to 0.5 mg / ml); and 3) incubating the mixture at about 37° C. for about 24 hours.

[0054] Additional aspects and advantages of the present disclosure will become readily apparent to those skilled in the art from the following detailed description, which shows and describes only exemplary embodiments of the present disclosure. As will be recognized, the present disclosure is capable of other and different embodiments, and its several details are capable of modifications in various obvious respects, all without departing from the present disclosure. Accordingly, the drawings and description are to be regarded as illustrative in nature, and not as restrictive.

[0055] Incorporation by Reference All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.

[0056] The novel features of the invention are set forth with particularity in the appended claims. Further features and advantages of the invention will become apparent from the following detailed description and accompanying drawings (also referred to herein as "figure" and "FIG.") that set forth illustrative embodiments in which the principles of the invention are utilized. This is better understood by [Brief explanation of the drawings]

[0057] [Figure 1] FIG. 1 shows the synthesis of HA-VS polymer. [Figure 2] FIG. 1 shows the synthesis of HA-SH polymer. [Figure 3] FIG. 1 illustrates the synthesis of hydrogels of the present disclosure. [Figure 4] FIG. 1 shows the elastic modulus of hydrogels of the present disclosure. [Figure 5] FIG. 1 shows the elastic modulus of hydrogels of the present disclosure. [Figures 6A-6B] FIG. 1 shows the elastic modulus and complex viscosity of hydrogels of the present disclosure. [Figures 7A-7B] FIG. 1 shows strain sweep and frequency sweep tests of hydrogels of the present disclosure. [Figure 8A-8B]FIG. 1 shows strain sweep and frequency sweep tests of hydrogels of the present disclosure. [Figures 9A-9D] FIG. 1 shows examples illustrating the mechanical properties of hydrogels. DETAILED DESCRIPTION OF THE INVENTION

[0058] While various embodiments of the present invention have been shown and described herein, it will be apparent to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions will occur to those skilled in the art without departing from the invention. It will be understood that various alternatives to the embodiments of the invention described herein may be used.

[0059] The term "biocompatible" or "biocompatibility," as used herein, generally refers to the condition of being compatible with living tissue or systems in that it is not toxic, injurious, or physiologically reactive and / or does not cause immunological rejection. vinegar.

[0060] The term "polymer," as used herein, generally refers to a chemical compound or mixture of compounds formed by polymerization and consisting essentially of repeating structural units.

[0061] The term "hydrogel," as used herein, generally refers to a gel or gel-like structure that includes one or more polymers suspended in an aqueous solution (eg, water).

[0062] The term "viscosity," as used herein, generally refers to the property of resistance to flow in a fluid or semifluid.

[0063] The term "intrinsic viscosity," as used herein, generally refers to a measure of the contribution of a substance (e.g., a polymer) to the viscosity of a system (e.g., a solution). In the present disclosure, intrinsic viscosity [η] can be measured by an Ubbelohde viscometer or a differential viscometer. Alternatively, intrinsic viscosity [η] can be calculated from the Mark-Hwink equation, which follows the established relationship between intrinsic viscosity and molecular weight.

[0064] The term "hydrogel-forming polymer," as used herein, generally refers to a polymer that participates in the formation of a hydrogel. A hydrogel-forming polymer can be a natural or synthetic polymer capable of forming a hydrogel. A hydrogel-forming polymer can include a polymer(s) that contribute to hydrogel formation. In some embodiments, a hydrogel-forming polymer does not include a polymer that is not capable of participating in hydrogel formation and / or is incapable of forming a hydrogel, even if present in a composition of the present disclosure. In some instances, a hydrogel-forming polymer may also be referred to as a "backbone polymer."

[0065] "C T The term "as used herein generally refers to the total concentration of polymer(s) in a composition. For example, the C of a hydrogel-forming polymer T may refer to the total concentration of polymers that form and / or have formed the hydrogel of the present disclosure. For example, C T may refer to the total concentration of hydrogel-forming polymer present in the composition of the present disclosure. In some cases, the composition of the present disclosure may contain hydrogel-forming polymers that have already formed a hydrogel and / or that have not yet been incorporated into a hydrogel, and the C of the hydrogel-forming polymer may be T can refer to the total concentration of hydrogel-forming polymers present in the composition (e.g., including both polymers already incorporated into a hydrogel and polymers not yet incorporated into a hydrogel). In another example, the C of a hydrogel-forming polymer having an intrinsic viscosity [η] of at least 3 dL / g can be used. Tmay refer to the total concentration of hydrogel-forming polymers having an intrinsic viscosity [η] of at least 3 dL / g.

[0066] The term "storage modulus" as used herein generally refers to the measured stored energy and represents the elastic portion.

[0067] The term "substantial," as used herein, generally refers to more than a minimal or insignificant amount, and "substantially" generally refers to more than a minimal or insignificant amount. The term "substantial portion of," as used herein, generally refers to an amount, quantity, sequence, length, concentration, etc., of a portion of a corresponding object that is at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, or at least about 99% of the total amount, quantity, sequence, length, concentration, etc., of the corresponding object.

[0068] The term "fluorescer" or "fluorescent moiety," as used herein, generally refers to a substance or portion thereof that is capable of exhibiting fluorescence in the detectable range. Exemplary detectable moieties suitable for use as detectable labels can include, for example, affinity tags and fluorescent proteins.

[0069] The term "consisting essentially of," as used herein, generally refers to being made up in substantial part of the specified components or ingredients.

[0070] The term "loss modulus G''" as used herein generally refers to the measured loss energy and represents the viscous portion.

[0071] The term "yield strain," as used herein, generally refers to the strain at which a material begins to deform plastically; the yield point is the point at which nonlinear (elastic and plastic) deformation sets in. Prior to the yield point, the material deforms elastically and returns to its original shape when the applied stress is removed.

[0072] The term "hydrophilic," as used herein, generally refers to having an affinity for water and being able to absorb or be wetted by water. Hydrophilic molecules or portions of molecules are those in which interactions with water and other polar substances are thermodynamically more favorable than interactions with oils or other hydrophobic solvents.

[0073] The term "average degree of modification (DM)," as used herein, generally refers to the percentage of repeating units in a polymer that have pendant groups. DM can reflect the degree of modification of a hydrogel-forming polymer derivative.

[0074] The term "crosslinker," as used herein, generally refers to an agent that links one polymer chain to another by a bond. Crosslinkers can achieve crosslinking via covalent or non-covalent bonds. A "polymer chain" can refer to a synthetic polymer or a natural polymer (such as a protein). In polymer chemistry, when a synthetic polymer is "crosslinked," it usually means that the entire bulk of the polymer has been exposed to a crosslinking process. The resulting change in mechanical properties is highly dependent on the crosslink density. Crosslinks can be formed by chemical reactions initiated by heat, pressure, pH changes, or radiation.

[0075] The term "about," when used in connection with a numerical value, generally refers to a value that is 1% to less than 15% (e.g., less than 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, or 15%) above or below the specified value.

[0076] The term "soft hydrogel," as used herein, generally refers to a hydrogel that has a relatively low G' (e.g., less than or about 1 Pa) and / or a high G' compared to G'' (e.g., G'' / G'<1), while having a relatively large yield strain (e.g., 10% or more, 20% or more, 50% or more, 70% or more, or even 100% or more). For example, soft hydrogels may be particularly suitable for application to surfaces.

[0077] Where a range of values ​​(e.g., a numerical range) is presented, it is understood that each value between the upper and lower limit of that range, to one-tenth of the unit of the lower limit, and any other stated or intervening value within the stated range, is encompassed within the invention, unless the context clearly dictates otherwise. Where a limit in a stated range is specifically excluded, the upper and lower limits of those smaller ranges can independently be included within the smaller ranges and are also encompassed within the invention. Where the stated range includes one or both limits, ranges excluding either or both of those included limits are also included in the invention.

[0078] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Methods and materials similar or equivalent to those described herein may be used in the practice or testing of the present invention. Possible and suitable methods and materials are described below. All publications mentioned herein are incorporated by reference to disclose and describe the methods and / or materials in connection with which the publications are cited.

[0079] As used herein, the singular forms "a," "and," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, a reference to "a particle" includes a plurality of such particles, a reference to "the sequence" includes a reference to one or more such sequences and equivalents thereof known to those skilled in the art, and so forth.

[0080] As will be understood by those skilled in the art upon reading this disclosure, each individual embodiment depicted and illustrated herein has individual components and features which may be readily separated from or combined with the features of any of the other several embodiments without departing from the scope or spirit of the invention. Any described method can be carried out in the order of events described or in any other order which is logically possible. All such combinations are intended to be supported.

[0081] The present disclosure provides compositions comprising polymers capable of forming hydrogels, as well as methods of making and using the same. For example, the compositions can include one or more hydrogel-forming polymers having an intrinsic viscosity [η] (e.g., as measured by an Ubbelohde viscometer) of at least 3 dL / g in the composition. The concentration C of the hydrogel-forming polymer in the composition can be expressed as: T can be at most about 5 mg / ml. More specifically, the present disclosure provides compositions comprising polymers capable of forming soft hydrogels, and methods of making and using the same.

[0082] In one aspect, the present disclosure provides a composition comprising a polymer capable of forming a hydrogel, e.g., a composition comprising one or more polymers capable of forming a hydrogel. The polymer or one or more polymers in the composition can have a concentration C of at most about 5 mg / ml (e.g., at most about 4 mg / ml, at most about 3 mg / ml, at most about 2 mg / ml, at most about 1.5 mg / ml, at most about 1 mg / ml, at most about 0.9 mg / ml, at most about 0.8 mg / ml, at most about 0.7 mg / ml, at most about 0.6 mg / ml, at most about 0.5 mg / ml, at most about 0.4 mg / ml, at most about 0.3 mg / ml, at most about 0.2 mg / ml, at most about 0.1 mg / ml, or less). For example, the polymer or polymers in the composition may be present in a concentration of about 0.05 mg / ml to about 5 mg / ml (e.g., about 0.1 mg / ml to about 5 mg / ml, about 0.2 mg / ml to about 5 mg / ml, about 0.3 mg / ml to about 5 mg / ml, about 0.4 mg / ml to about 5 mg / ml, about 0.5 mg / ml to about 5 mg / ml, about 0.6 mg / ml to about 5 mg / ml, about 0.7 mg / ml to about 5 mg / ml, about 0.8 mg / ml to about 5 mg / ml, about 0.9 mg / ml to about 5 mg / ml, about 10 mg / ml to about 10 mg / ml, about 11 mg / ml to about 11 mg / ml, about 12 mg / ml to about 12 mg / ml, about 13 mg / ml to about 13 mg / ml, about 14 mg / ml to about 14 mg / ml, about 15 mg / ml to about 15 mg / ml, about 16 mg / ml to about 16 mg / ml, about 17 mg / ml to about 17 mg / ml, about 18 mg / ml to about 18 mg / ml, about 19 mg / ml to about 19 mg / ml, about 20 mg / ml to about 20 mg / ml, about 21 mg / ml to about 21 mg / ml, about 22 mg / ml to about 22 mg / ml, about 23 mg / ml to about 23 mg / ml, about 24 mg / ml to about 24 mg / ml, about 25 mg / ml to about 25 mg / ml, about 26 mg / ml to about 26 mg / ml, about 27 mg / ml to about 27 mg / ml, about 28 mg / ml to about 28 mg / ml, about 29 mg / ml to about 29 mg / ml, about 30 mg / ml The concentration C may be from about 0.8 mg / ml to about 5 mg / ml, from about 0.9 mg / ml to about 5 mg / ml, from about 1.0 mg / ml to about 5 mg / ml, from about 1.0 mg / ml to about 4 mg / ml, from about 1.0 mg / ml to about 3 mg / ml, from about 1.0 mg / ml to about 2 mg / ml, from about 0.3 mg / ml to about 0.8 mg / ml, from about 0.3 mg / ml to about 0.6 mg / ml, or from about 0.3 mg / ml to about 0.5 mg / ml.

[0083] In another aspect, the present disclosure provides compositions comprising one or more hydrogel-forming polymers. Such hydrogel-forming polymers can have an intrinsic viscosity [η] (e.g., as measured by an Ubbelohde viscometer) of at least 3 dL / g in the composition (e.g., at least about 5 dL / g, at least about 8 dL / g, at least about 10 dL / g, at least about 12 dL / g, at least about 15 dL / g, at least about 16 dL / g, at least about 17 dL / g, at least about 18 dL / g, at least about 19 dL / g, at least about 20 dL / g, at least about 25 dL / g, or more). In some embodiments, the intrinsic viscosity [η] is measured by a differential viscometer. Alternatively, the intrinsic viscosity [η] can be calculated from the Mark-Hwink equation, which follows the established relationship between intrinsic viscosity and molecular weight.

[0084] The total concentration of the hydrogel-forming polymer(s) in the composition (e.g., hydrogel-forming polymers having an intrinsic viscosity [η] of at least 3 dL / g) is C T C T can be at most about 5 mg / ml (e.g., at most about 4 mg / ml, at most about 3 mg / ml, at most about 2 mg / ml, at most about 1.5 mg / ml, at most about 1 mg / ml, at most about 0.9 mg / ml, at most about 0.8 mg / ml, at most about 0.7 mg / ml, at most about 0.6 mg / ml, at most about 0.5 mg / ml, at most about 0.4 mg / ml, at most about 0.3 mg / ml, at most about 0.2 mg / ml, at most about 0.1 mg / ml, or less). For example, C T is about 0.05 mg / ml to about 5 mg / ml (e.g., about 0.1 mg / ml to about 5 mg / ml, about 0.2 mg / ml to about 5 mg / ml, about 0.3 mg / ml to about 5 mg / ml, about 0.4 mg / ml to about 5 mg / ml, about 0.5 mg / ml to about 5 mg / ml, about 0.6 mg / ml to about 5 mg / ml, about 0.7 mg / ml to about 5 mg / ml, about 0.8 mg / ml to about 5 mg / ml) mg / ml, about 0.9 mg / ml to about 5 mg / ml, about 1.0 mg / ml to about 5 mg / ml, about 1.0 mg / ml to about 4 mg / ml, about 1.0 mg / ml to about 3 mg / ml, about 1.0 mg / ml to about 2 mg / ml, about 0.3 mg / ml to about 0.8 mg / ml, about 0.3 mg / ml to about 0.6 mg / ml, or about 0.3 mg / ml to about 0.5 mg / ml).

[0085] In some cases, the polymer, one or more polymers, or hydrogel-forming polymers in the composition are present in an amount of from about 0.01% (w / v) to about 4% (w / v), e.g., from about 0.02% (w / v) to about 4% (w / v), from about 0.03% (w / v) to about 4% (w / v), from about 0.04% (w / v) to about 4% (w / v), from about 0.05% (w / v) to about 4% (w / v), or from about 0.0 6% (w / v) ~ approx. 4% (w / v), approx. 0.07% (w / v) ~ approx. 4% (w / v), approx. 0.08% (w / v) ~ approx. 4% (w / v), approx. 0.09% (w / v) ~ approx. 4% (w / v) ), about 0.1% (w / v) to about 4% (w / v), about 0.11% (w / v) to about 4% (w / v), about 0.12% (w / v) to about 4% (w / v), about 0.13% (w / v) to about 4% (w / v), about 0.14% (w / v) to about 4% (w / v), about 0.15% (w / v) to about 4% (w / v), about 0.2% (w / v) to about 4% (w / v), about 0.25% (w / v) ) ~ 4% (w / v), approximately 0.3% (w / v) ~ 4% (w / v), approximately 0.4% (w / v) ~ 4% (w / v), approximately 0.5% (w / v) ~ 4% (w / v), approximately 0.6% (w / The concentration C may be from about 0.7% (w / v) to about 4% (w / v), from about 0.7% (w / v) to about 4% (w / v), from about 0.8% (w / v) to about 4% (w / v), from about 0.9% (w / v) to about 4% (w / v), from about 1% (w / v) to about 4% (w / v), from about 2% (w / v) to about 4% (w / v), from about 3% (w / v) to about 4% (w / v), or from about 3.5% (w / v) to about 4% (w / v).

[0086] The polymer, at least one of the one or more polymers, or hydrogel-forming polymer may have an intrinsic viscosity [η] in the composition of at least about 3 dL / g (e.g., at least about 5 dL / g, at least about 8 dL / g, at least about 10 dL / g, at least about 12 dL / g, at least about 15 dL / g, at least about 16 dL / g, at least about 17 dL / g, at least about 18 dL / g, at least about 19 dL / g, at least about 20 dL / g, at least about 25 dL / g, or more).

[0087] In some cases, one or more of the polymers, or each of the hydrogel-forming polymers, in the composition may have an intrinsic viscosity [η] of at least 3 dL / g (e.g., at least about 5 dL / g, at least about 8 dL / g, at least about 10 dL / g, at least about 12 dL / g, at least about 15 dL / g, at least about 16 dL / g, at least about 17 dL / g, at least about 18 dL / g, at least about 19 dL / g, at least about 20 dL / g, at least about 25 dL / g, or more).

[0088] For example, when only one polymer (e.g., a hydrogel-forming polymer) is present in the composition, that polymer may have an intrinsic viscosity [η] in the composition of at least 3 dL / g (e.g., at least about 5 dL / g, at least about 8 dL / g, at least about 10 dL / g, at least about 12 dL / g, at least about 15 dL / g, at least about 16 dL / g, at least about 17 dL / g, at least about 18 dL / g, at least about 19 dL / g, at least about 20 dL / g, at least about 25 dL / g, or more). When two or more polymers (e.g., hydrogel-forming polymers) are present in the composition, one or more of them, or even each of them, can have an intrinsic viscosity [η] in the composition of at least 3 dL / g (e.g., at least about 5 dL / g, at least about 8 dL / g, at least about 10 dL / g, at least about 12 dL / g, at least about 15 dL / g, at least about 16 dL / g, at least about 17 dL / g, at least about 18 dL / g, at least about 19 dL / g, at least about 20 dL / g, at least about 25 dL / g, or more).

[0089] For a polymer (e.g., a hydrogel-forming polymer) in the composition, its concentration C is * may be about 0.8 to about 5 times (for example, about 0.8 to about 4.5 times, about 0.8 to about 4.0 times, about 0.8 to about 3.5 times, about 0.8 to about 3.0 times, about 0.8 to about 2.5 times, about 0.8 to about 2.0 times, about 0.8 to about 1.5 times, or about 1.0 to about 1.2 times) of C * = 1 / ([η]), where [η] is the intrinsic viscosity of the polymer.

[0090] For example, if there are two or more polymers (e.g., hydrogel-forming polymers) in the composition, their total concentration in the composition is C T and each type of polymer may have a concentration C. The concentration C of one type of polymer may be different from the concentration C of another type of polymer. For each type of polymer, its concentration C may be * may be about 0.8 to about 5 times (for example, about 0.8 to about 4.5 times, about 0.8 to about 4.0 times, about 0.8 to about 3.5 times, about 0.8 to about 3.0 times, about 0.8 to about 2.5 times, about 0.8 to about 2.0 times, about 0.8 to about 1.5 times, or about 1.0 to about 1.2 times) of C * = 1 / ([η]), where [η] is the intrinsic viscosity of the polymer.

[0091] In some cases, the total concentration C of hydrogel-forming polymers (e.g., hydrogel-forming polymers having an intrinsic viscosity [η] of at least 3 dL / g in the composition) T can be at most about 5 mg / ml (e.g., at most about 4 mg / ml, at most about 3 mg / ml, at most about 2 mg / ml, at most about 1.5 mg / ml, at most about 1 mg / ml, at most about 0.9 mg / ml, at most about 0.8 mg / ml, at most about 0.7 mg / ml, at most about 0.6 mg / ml, at most about 0.5 mg / ml, at most about 0.4 mg / ml, at most about 0.3 mg / ml, at most about 0.2 mg / ml, at most about 0.1 mg / ml, or less). For example, the total concentration C of hydrogel-forming polymers having an intrinsic viscosity [η] of at least 3 dL / g Tis about 0.05 mg / ml to about 5 mg / ml (e.g., about 0.1 mg / ml to about 5 mg / ml, about 0.2 mg / ml to about 5 mg / ml, about 0.3 mg / ml to about 5 mg / ml, about 0.4 mg / ml to about 5 mg / ml, about 0.5 mg / ml to about 5 mg / ml, about 0.6 mg / ml to about 5 mg / ml, about 0.7 mg / ml to about 5 mg / ml, about 0.8 mg / ml to about 5 mg / ml) mg / ml, about 0.9 mg / ml to about 5 mg / ml, about 1.0 mg / ml to about 5 mg / ml, about 1.0 mg / ml to about 4 mg / ml, about 1.0 mg / ml to about 3 mg / ml, about 1.0 mg / ml to about 2 mg / ml, about 0.3 mg / ml to about 0.8 mg / ml, about 0.3 mg / ml to about 0.6 mg / ml, or about 0.3 mg / ml to about 0.5 mg / ml).

[0092] The composition has a viscosity of about 3.5 to about 9.0, for example, about 4.0 to about 9.0, about 4.5 to about 9.0, about 5.0 to about 9.0, about 5.5 to about 9.0, about 6.0 to about 9.0, about 6.5 to about 9.0, about 7.0 to about 9.0, about 7.5 to about 9.0, about 8.0 to about 9.0, about 8.5 to about 9.0, It may have a pH of about 7.0 to about 7.8, or about 7.4.

[0093] In another aspect, the present disclosure provides a hydrogel formed by the composition of the present disclosure. The hydrogel can be biocompatible.

[0094] In some cases, the hydrogel-forming polymers in the composition may initially be present in a non-crosslinked form (e.g., no hydrogel has yet formed), and after certain treatments (e.g., after incubation for a certain period of time, e.g., 12 hours or more, e.g., at 37°C), some of the hydrogel-forming polymers may chemically and / or physically crosslink to form a hydrogel. In certain embodiments, substantially all of the hydrogel-forming polymers may chemically and / or physically crosslink to form a hydrogel.

[0095] Thus, in some cases, the composition may contain a formed hydrogel. In such cases, at least a portion of the hydrogel-forming polymers are contained in the formed hydrogel in the composition. For example, the hydrogel-forming polymers in the composition may include both polymers that have already formed a hydrogel and polymers that have not yet been incorporated into a hydrogel but are capable of forming a hydrogel under certain reaction conditions (e.g., after incubation at 37°C for a certain period of time, e.g., 12 hours). In such cases, the total concentration C of both the hydrogel-forming polymers already contained in the hydrogel and the hydrogel-forming polymers not yet incorporated into the hydrogel may be expressed as T can be at most about 5 mg / ml.

[0096] In some cases, substantially all of the hydrogel-forming polymers in the composition may form a hydrogel (eg, by chemical and / or physical crosslinking).

[0097] Hydrogels according to the present disclosure may have one or more particular characteristics / properties.

[0098] For example, hydrogels of the present disclosure may have a storage modulus G' of about 10.0 Pa or less (e.g., about 8.0 Pa or less, about 7.0 Pa or less, about 6.0 Pa or less, about 5.0 Pa or less, about 4.0 Pa or less, about 3.0 Pa or less, about 2.0 Pa or less, about 1.0 Pa or less, about 0.8 Pa or less, about 0.7 Pa or less, about 0.6 Pa or less, about 0.5 Pa or less, or less), as measured in a dynamic oscillatory shear test.

[0099] Hydrogels of the present disclosure may have a loss modulus G'' of about 100% or less (e.g., about 90% or less, about 80% or less, about 70% or less, about 60% or less, about 55% or less, about 50% or less, about 45% or less, about 40% or less, about 35% or less, about 30% or less, about 25% or less, or about 20% or less) of their storage modulus G' as measured in a dynamic oscillatory shear test.

[0100] Hydrogels of the present disclosure may have a complex viscosity of about 0.2 Pa·s or less (e.g., about 0.1 Pa·s or less, about 0.08 Pa·s or less, about 0.07 Pa·s or less, about 0.06 Pa·s or less, about 0.05 Pa·s or less, or about 0.04 Pa·s or less), measured in a dynamic oscillatory shear test at a frequency of less than about 100 rad / s (e.g., less than about 90 rad / s, less than about 80 rad / s, less than about 70 rad / s, less than about 50 rad / s, less than about 40 rad / s, less than about 30 rad / s, less than about 20 rad / s, less than about 10 rad / s, or less).

[0101] The hydrogels of the present disclosure have a modulus of elasticity of at least about 10% (e.g., at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, at least about 100%, at least about 110%, or The yield strain may be greater than or equal to 1000 MPa.

[0102] For example, in dynamic oscillatory shear testing, a sinusoidal force (e.g., stress) can be applied to a material and the resulting displacement (strain) can be measured. For a perfectly elastic solid, the resulting strain and stress can be perfectly in phase. For a purely viscous fluid, there can be a 90 degree phase lag of strain relative to stress. Viscoelastic polymers with intermediate characteristics may have a phase lag during testing, and the storage modulus can be calculated accordingly.

[0103] Dynamic mechanical analyzers (DMA) can be used to measure storage modulus and can be used for dynamic oscillatory shear testing. In another example, a DMA analyzer can include a displacement sensor (such as a linear variable differential transformer) that can measure the change in voltage caused by the passage of an instrument probe through a magnetic core. The DMA analyzer can also include a temperature control system or furnace, a drive motor (e.g., a linear motor for probe loading that can effect the loading of an applied force), a drive shaft support, and a guidance system that acts as a force guide from the motor to the sample, and one or more sample clamps for holding the sample being tested.

[0104] Various types of DMA analyzers can be used. For example, forced resonance analyzers or free resonance analyzers can be used. Free resonance analyzers measure the damping free vibration of the sample being tested by suspending and oscillating the sample. Forced resonance analyzers allow the sample to oscillate at a specific frequency, enabling temperature sweeps to be performed reliably. The analyzer may be for both stress (force) and strain (displacement) control. For example, in strain control, the probe can be moved and the resulting sample stress can be measured by implementing a force balance transducer that can use different axes. In stress control, a constant force can be applied and some other experimental condition (temperature, frequency, or time) can be varied. Stress and strain can be applied by torsional or axial analyzers. In torsional analyzers, force is applied in a torsional motion. Axial analyzers can be used for bending, tension, and / or compression tests.

[0105] Various test modes, such as temperature sweep, frequency sweep, dynamic stress-strain, or a combination thereof, can be used to probe the viscoelastic properties of polymers. For example, in dynamic stress-strain tests, dynamic stress-strain measurements can be performed by gradually increasing the oscillation amplitude. The variation of the storage and loss moduli with increasing stress can be used to characterize the material and to determine the upper limit of the linear stress-strain regime of the material.

[0106] Various mechanical properties can be determined by DMA. These properties include storage modulus (G'), loss modulus (G''), complex modulus (G * ), loss angle (tan(δ)), complex viscosity (η * ), its in-phase component (η') and out-of-phase component (η''), complex compliance (J * ), storage compliance (J'), loss compliance (J''), etc.

[0107] In another aspect, the present disclosure provides methods of making hydrogels (e.g., hydrogels of the present disclosure). The methods can include providing a composition (e.g., a composition including one or more polymers of the present disclosure) and exposing the composition to conditions that allow for the formation of a hydrogel (e.g., allow for crosslinking of the polymers that form the hydrogel). For example, the conditions can include incubating the composition at about 30°C to about 45°C (e.g., about 32°C to about 40°C, about 35°C to about 40°C, e.g., about 37°C).

[0108] The method may further include preparing a polymer as defined in the present disclosure in a buffer to form a polymer solution. The solution may include one or more polymers. The polymer (e.g., hydrogel-forming polymer) in the polymer solution has a total concentration C of at most about 5 mg / ml (e.g., at most about 4 mg / ml, at most about 3 mg / ml, at most about 2 mg / ml, at most about 1.5 mg / ml, at most about 1 mg / ml, at most about 0.9 mg / ml, at most about 0.8 mg / ml, at most about 0.7 mg / ml, at most about 0.6 mg / ml, at most about 0.5 mg / ml, at most about 0.4 mg / ml, at most about 0.3 mg / ml, at most about 0.2 mg / ml, at most about 0.1 mg / ml, or less). T may have:

[0109] For example, the hydrogel-forming polymer in the polymer solution may be present in a concentration of about 0.1 mg / ml to about 5 mg / ml (e.g., about 0.2 mg / ml to about 5 mg / ml, about 0.3 mg / ml to about 5 mg / ml, about 0.4 mg / ml to about 5 mg / ml, about 0.5 mg / ml to about 5 mg / ml, about 0.6 mg / ml to about 5 mg / ml, about 0.7 mg / ml to about 5 mg / ml, about 0.8 mg / ml to about 5 mg / ml). about 5 mg / ml, about 0.9 mg / ml to about 5 mg / ml, about 1.0 mg / ml to about 5 mg / ml, about 1.0 mg / ml to about 4 mg / ml, about 1.0 mg / ml to about 3 mg / ml, about 1.0 mg / ml to about 2 mg / ml, about 0.3 mg / ml to about 0.8 mg / ml, about 0.3 mg / ml to about 0.6 mg / ml, or about 0.3 mg / ml to about 0.5 mg / ml) T may have:

[0110] In some cases, the total concentration of hydrogel-forming polymers, C T can be about 0.3 mg / ml to 0.5 mg / ml, or less.

[0111] In some cases, the method may include crosslinking the polymer in solution to produce a hydrogel, e.g., conditions that allow for the formation of a hydrogel may also allow for crosslinking of the polymer in solution.

[0112] The buffer may be an aqueous solution and may comprise water and a suitable salt useful for adjusting the pH or buffering capacity of the aqueous solution, hi some embodiments, the buffer comprises a phosphate buffer, such as phosphate buffered saline (PBS).

[0113] To this end, compositions according to the present disclosure may further comprise a buffer such as a phosphate buffer, for example phosphate buffered saline (PBS).

[0114] In the methods of the present disclosure, the polymer solution can have a pH of about 3.5 to about 9.0, e.g., about 4.0 to about 9.0, about 4.5 to about 9.0, about 5.0 to about 9.0, about 5.5 to about 9.0, about 6.0 to about 9.0, about 6.5 to about 9.0, about 7.0 to about 9.0, about 7.5 to about 9.0, about 8.0 to about 9.0, about 8.5 to about 9.0, about 7.0 to about 7.8, or about 7.4.

[0115] In some cases, the polymer solution can have a pH of about 6 to 10, e.g., about 6.5 to 10, about 7 to 10, about 7.1 to 10, about 7.2 to 10, about 7.3 to 10, about 7.4 to 10, about 7.5 to 10, about 7.6 to 10, about 7.7 to 10, about 7.8 to 10, about 7.9 to 10, about 8.0 to 10, about 8.5 to 10, about 9 to 10, or about 9.5 to 10. In some embodiments, the polymer solution has a pH of about 7 to 8, e.g., about 7.4.

[0116] In the methods of the present disclosure, crosslinking can include incubating the polymer solution at about 20°C to about 50°C, about 25°C to about 50°C, about 30°C to about 50°C, about 31°C to about 50°C, about 32°C to about 50°C, about 33°C to about 50°C, about 34°C to about 50°C, about 35°C to about 50°C, about 36°C to about 50°C, about 37°C to about 50°C, about 38°C to about 50°C, about 39°C to about 50°C, about 40°C to about 50°C, or about 45°C to about 50°C. In some embodiments, crosslinking can include incubating the polymer solution at about 20°C to about 50°C, about 25°C to about 50°C, about 30°C to about 50°C, about 31°C to about 50°C, about 32°C to about 50°C, about 33°C to about 50°C, about 34°C to about 50°C, about 35°C to about 50°C, about 36°C to about 50°C, about 37°C to about 50°C, about 38°C to about 50°C, about 39°C to about 50°C, about 40°C to about 50°C, or about 45°C to about 50°C. at a temperature of about 30°C to about 40°C, for example, about 35°C to about 39°C, for example, about 37°C.

[0117] In the methods of the present disclosure, crosslinking can include incubating the polymer solution for at least about 1 hour, e.g., at least about 2 hours, at least about 3 hours, at least about 4 hours, at least about 5 hours, at least about 6 hours, at least about 7 hours, at least about 8 hours, at least about 9 hours, at least about 9.5 hours, at least about 10 hours, at least about 10.5 hours, at least about 11 hours, at least about 12 hours, at least about 13 hours, at least about 14 hours, at least about 15 hours, at least about 16 hours, at least about 17 hours, at least about 18 hours, at least about 19 hours, at least about 20 hours, at least about 24 hours, or more.

[0118] For example, in the methods of the present disclosure, crosslinking can include incubating the polymer solution at a temperature of about 30°C to about 40°C (e.g., about 35°C to about 39°C, e.g., about 37°C) for at least 5 hours (e.g., at least about 10 hours or more, e.g., about 24 hours or more).

[0119] For example, the method may include: 1) preparing a first polymer population (or a first polymer derivative) and a second polymer population (or a second polymer derivative) (e.g., the polymers in the first polymer population may comprise hyaluronic acid modified with one or more vinyl sulfone groups, and the polymers in the second polymer population may comprise hyaluronic acid modified with one or more thiol groups) in water and adjusting the pH (e.g., by adding a buffer solution); 2) mixing the polymers of the first polymer population (or the first polymer derivative) with the polymers of the second polymer population (or the second polymer derivative) in a predetermined ratio (the concentrations of the polymers in the composition are as defined in the present disclosure); and 3) incubating the mixture under conditions that allow for the formation of a hydrogel according to the present disclosure.

[0120] According to any aspect of the present disclosure, the polymers of the present disclosure can be hydrophilic and / or water-soluble.

[0121] According to any aspect of the present disclosure, the polymer (e.g., hydrogel-forming polymer) can be selected from the group consisting of polysaccharides, poly(acrylic acid), poly(hydroxyethyl methacrylate), elastin, collagen, derivatives thereof, and any combination thereof. For example, the polymer (e.g., hydrogel-forming polymer) in the composition can include one or more of the following: polysaccharides, one or more polysaccharide derivatives, poly(acrylic acid), one or more poly(acrylic acid) derivatives, poly(hydroxyethyl methacrylate), one or more poly(hydroxyethyl methacrylate) derivatives, elastin, one or more elastin derivatives, collagen, and one or more collagen derivatives.

[0122] For example, the polymer (e.g., hydrogel-forming polymer) can be selected from the group consisting of hyaluronic acid, guar gum, starch, chitosan, chondroitin sulfate, alginate, carboxymethylcellulose, derivatives thereof, and any combination thereof. For example, the polymer (e.g., hydrogel-forming polymer) in the composition can include one or more of the following: hyaluronic acid, one or more hyaluronic acid derivatives, guar gum, one or more guar gum derivatives, starch, one or more starch derivatives, chitosan, one or more chitosan derivatives, chondroitin sulfate, one or more chondroitin sulfate derivatives, alginate, one or more alginate derivatives, carboxymethylcellulose, and one or more carboxymethylcellulose derivatives. In some embodiments, the polymer (e.g., hydrogel-forming polymer) in the composition includes one or more of the following: hyaluronic acid and one or more hyaluronic acid derivatives.

[0123] According to any aspect of the present disclosure, a polymer (e.g., a hydrogel-forming polymer) can be modified with one or more modifications, e.g., to provide a polymer derivative of the present disclosure. In one example, a polymer (e.g., a hydrogel-forming polymer) of the present disclosure can be modified with one or more vinyl sulfone groups (or molecules containing one or more vinyl sulfone groups). In another example, a polymer (e.g., a hydrogel-forming polymer) of the present disclosure can be modified with one or more thiol groups (or molecules containing one or more thiol groups).

[0124] For example, the polymers of the present disclosure can be modified with one or more modifications selected from the group consisting of acrylate, maleimide, vinyl sulfone, N-hydroxysuccinimide, aldehyde, ketone, carbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, amine, and any combination thereof.

[0125] For example, the polymer may comprise one or more polysaccharide derivatives, which may be polysaccharides modified with acrylates, maleimides, vinyl sulfones, N-hydroxysuccinimides, aldehydes, ketones, carbodiimides, carbonates, iodoacetyl, mercaptonicotinamides, quinones, thiols, and / or amines. In some cases, the polymer may comprise one or more poly(acrylic acid) derivatives, which may be poly(acrylic acid)s modified with acrylates, maleimides, vinyl sulfones, N-hydroxysuccinimides, aldehydes, ketones, carbodiimides, carbonates, iodoacetyl, mercaptonicotinamides, quinones, thiols, and / or amines. In some cases, the polymer may comprise one or more poly(hydroxyethyl methacrylate) derivatives, which may be poly(hydroxyethyl methacrylate) modified with acrylate, maleimide, vinyl sulfone, N-hydroxysuccinimide, aldehyde, ketone, carbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, and / or amine. In some cases, the polymer may comprise one or more elastin derivatives, which may be elastin modified with acrylate, maleimide, vinyl sulfone, N-hydroxysuccinimide, aldehyde, ketone, carbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, and / or amine. In some cases, the polymer may include one or more collagen derivatives, which may be acrylate, maleimide, vinyl sulfone, N-hydroxysuccinimide, aldehyde, ketone, carbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, and / or amine modified collagen.

[0126] In one embodiment, the polymer comprises a derivative of hyaluronic acid modified with one or more thiol groups (HA-SH), and optionally, the HA-SH can form a HA-SH-based polymer-polymer hydrogel under appropriate conditions. In another embodiment, the polymer comprises a derivative of hyaluronic acid modified with one or more vinyl sulfone groups (HA-VS), and optionally, the HA-VS can form a HA-VS-based polymer-polymer hydrogel under appropriate conditions. In another embodiment, the polymer comprises a derivative of hyaluronic acid modified with one or more thiol groups and a derivative of hyaluronic acid modified with one or more vinyl sulfone groups, and optionally, the HA-SH and HA-VS can react with each other to form a polymer-polymer hydrogel under appropriate conditions.

[0127] The derivative may have an average degree of modification (DM) of about 3% to about 50% (e.g., about 4% to about 45%, about 5% to about 40%, about 6% to about 40%, about 7% to about 40%, about 8% to about 39%, about 8% to about 38%, about 8% to about 35%, about 9% to about 32%, about 8% to about 30%, about 10% to about 30%, about 12% to about 30%, about 13% to about 30%, about 14% to about 30%, about 15% to about 35%, or about 15% to about 30%).

[0128] According to any aspect of the present disclosure, the polymers (e.g., hydrogel-forming polymers) of the present disclosure can include a first polymer population including polymers modified with at least a first modification and a second polymer population including polymers modified with a second modification. The first modification can be different from the second modification. The polymers of the first polymer population can react with the polymers of the second polymer population to form a hydrogel. The ratio (e.g., mass ratio, volume ratio, molar ratio, and / or DM ratio) of the polymer contained in the first polymer population to the polymer contained in the second polymer population is about 10:1 to about 1:10, for example, about 8:1 to about 1:10, about 6:1 to about 1:10, about 5:1 to about 1:10, about 4:1 to about 1:10, about 3:1 to about 1:10, about 2:1 to about 1:10, about 1.75:1 to about 1:10, about 1.5:1 to about 1:10, about 1.25:1 to about 1:10, about 1:1 to about 1:10, The ratio may be about 1:1.25 to about 1:10, about 1:1.5 to about 1:10, about 1:1.75 to about 1:10, about 1:2 to about 1:10, about 1:3 to about 1:10, about 1:4 to about 1:10, about 1:5 to about 1:10, about 6:1 to about 1:6, about 5:1 to about 1:5, about 4:1 to about 1:4, about 3:1 to about 1:3, about 2:1 to about 1:2, about 1.75:1 to about 1:1.75, about 1.5:1 to about 1:1.5, about 1.25:1 to about 1:1.25, or about 1.1:1 to about 1:1.1.

[0129] For example, the ratio (e.g., mass ratio, volume ratio, molar ratio, and / or DM ratio) of the polymers contained in the first polymer population to the polymers contained in the second polymer population can be from about 3:1 to about 1:3. For example, the ratio can be from about 2:1 to about 1:2 (e.g., about 1:1).

[0130] For example, the composition may include at least a first polymer derivative and a second polymer derivative. The first polymer derivative may include a first modification, and the second polymer derivative may include a second modification. The first modification may be different from the second modification. The first polymer derivative may be capable of reacting with the second polymer derivative to form a hydrogel. For example, the first polymer derivative may be a polymer (e.g., hyaluronic acid) modified with one or more vinyl sulfone groups. The second polymer derivative may be a polymer (e.g., hyaluronic acid) modified with one or more thiol groups.

[0131] The mass ratio of the first polymer derivative to the second polymer derivative in the composition can be from about 10:1 to about 1:10 (e.g., 10:2, 10:3, 10:4, 10:5, 10:6, 10:7, 10:8, 10:9, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9 or 1:10), for example, from about 3:1 to about 1:3 or from about 2:1 to about 1:2, for example, about 1:1.

[0132] The molar ratio of the first polymer derivative to the second polymer derivative in the composition can be from about 10:1 to about 1:10 (e.g., 10:2, 10:3, 10:4, 10:5, 10:6, 10:7, 10:8, 10:9, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9 or 1:10), for example, from about 3:1 to about 1:3 or from about 2:1 to about 1:2, for example, about 1:1.

[0133] The volume ratio of the first polymer derivative to the second polymer derivative in the composition can be about 10:1 to about 1:10 (e.g., 10:2, 10:3, 10:4, 10:5, 10:6, 10:7, 10:8, 10:9, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9 or 1:10), for example, about 3:1 to about 1:3 or about 2:1 to about 1:2, for example, about 1:1.

[0134] In some cases, the first polymer derivative has a first DM (DM1) and the second polymer derivative has a second DM (DM2), and the ratio of the first DM (DM1) to the second DM (DM2) is about 10:1 to about 1:10 (e.g., 10:2, 10:3, 10:4, 10:5, etc.). , 10:6, 10:7, 10:8, 10:9, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:9 or 1:10), for example from about 3:1 to about 1:3, or from about 2:1 to about 1:2, for example about 1:1.

[0135] In some embodiments, the first polymer derivative and the second polymer derivative in the composition may have a mass ratio defined in this disclosure, a molar ratio defined in this disclosure, a volume ratio defined in this disclosure, and / or a DM ratio defined in this disclosure.

[0136] According to any embodiment of the present disclosure, the first modification and the second modification may each independently be selected from the group consisting of acrylate, maleimide, vinyl sulfone, N-hydroxysuccinimide, aldehyde, ketone, carbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, amine, and any combination thereof. The first modification and the second modification may be different from each other.

[0137] Thus, the first polymer derivative (or polymer in the first polymer population) may be a polymer of the present disclosure modified with one or more vinyl sulfone groups (or molecules comprising one or more vinyl sulfone groups), and the second polymer derivative (or polymer in the second polymer population) may be a polymer of the present disclosure modified with one or more thiol groups (or molecules comprising one or more thiol groups). The first polymer derivative (or polymer in the first polymer population) may be capable of reacting with the second polymer derivative (or polymer in the second polymer population) to form a hydrogel.

[0138] In another example, a first polymer derivative (or a polymer in a first polymer population) may be a polymer of the present disclosure modified with one or more thiol groups (or molecules comprising one or more thiol groups), and a second polymer derivative (or a polymer in a second polymer population) may be a polymer of the present disclosure modified with one or more vinyl sulfone groups (or molecules comprising one or more vinyl sulfone groups). The first polymer derivative (or a polymer in a first polymer population) may be capable of reacting with the second polymer derivative (or a polymer in a second polymer population) to form a hydrogel.

[0139] In some embodiments, the polymer is hyaluronic acid, and the first polymer population (or first polymer derivative) comprises hyaluronic acid modified with one or more vinyl sulfone groups. The second polymer population (or second polymer derivative) comprises hyaluronic acid modified with one or more thiol groups. The polymers (or first polymer derivatives) of the first polymer population can react with the polymers (or second polymer derivatives) of the second polymer population to form a hydrogel.

[0140] In another example, the polymer is hyaluronic acid, and the first polymer population (or first polymer derivative) comprises hyaluronic acid modified with one or more thiol groups. The second polymer population (or second polymer derivative) comprises hyaluronic acid modified with one or more vinyl sulfone groups. The polymers (or first polymer derivatives) of the first polymer population can react with the polymers (or second polymer derivatives) of the second polymer population to form a hydrogel.

[0141] According to any embodiment of the present disclosure, the polymer in the composition may have a molecular weight of about 100,000 Daltons to about 5,000,000 Daltons, e.g., about 120,000 Daltons to about 5,000,000 Daltons, about 200,000 Daltons to about 5,000,000 Daltons, about 300,000 Daltons to about 5,000,000 Daltons, about 400,000 Daltons to about 5,000,000 Daltons, about 500,000 Daltons to about 5,000,000 Daltons, about 600,000 Daltons to about 5,000,000 Daltons, about 670,000 Daltons to about 5,000,000 Daltons, about 1,000,000 Daltons, Daltons to about 5,000,000 Daltons, about 1,500,000 Daltons to about 5,000,000 Daltons, about 2,000,000 Daltons to about 5,000,000 Daltons, about 2,500,000 Daltons to about 5,000,000 Daltons, about 2,600,000 Daltons to about 5,000,000 Daltons, about 3,000,000 Daltons to about 5,000,000 Daltons, about 3,500,000 Daltons to about 5,000,000 Daltons, about 3,600,000 Daltons to about 5,000,000 Daltons, about 2,000,000 The polymer may have an average molecular weight of from about 4,000,000 daltons, from about 2,500,000 daltons to about 3,500,000 daltons, from about 2,600,000 daltons to about 3,600,000 daltons, from about 1,000,000 daltons to about 2,600,000 daltons, from about 800,000 daltons to about 2,600,000 daltons, from about 700,000 daltons to about 2,500,000 daltons, from about 670,000 daltons to about 2,600,000 daltons, or from about 600,000 daltons to about 2,500,000 daltons.

[0142] The polymers of the present disclosure may contain one or more modifications (also referred to in the present disclosure as polymer derivatives), and may have a molecular weight of about 3% to about 50%, e.g., about 4% to about 50%, about 5% to about 50%, about 6% to about 50%, about 7% to about 50%, about 8% to about 50%, about 9% to about 50%, about 10% to about 50%, about 11% to about 50%, about 12% to about 50%, about 13% to about 50%, about 14% to about 50%, about 15% to about 50%, about 16% to about 50%, about 17% to about 50%, about 18% to about 50%, about 19% to about 50%, about 20% to about 50%, about 21% to about 50%, about 22% to about 50%, about 23% to about 50%, about 24% to about 50%, about 25% to about 50%, about 26% to about 50%, about 27% to about 50%, about 28% to about 50%, about 29% to about 50%, about 30% to about 50%, about 31% to about 50%, about 32% to about 50%, about 33% to about 50%, about 34% to about 50%, about 35% to about 50%, about 36% to about 50%, about 37% to about 50%, about 38% to about 50%, about 39% to about 50%, about 40% to about 50%, about 41% to about 50%, about 42% to about 50%, about 43% to about 50%, about 44% to about 50%, about 45% to about 50%, about 46% to %, about 21% to about 50%, about 22% to about 50%, about 23% to about 50%, about 24% to about 50%, about 25% to about 50%, about 26% to about 50%, about 27% to about 50%, about 28% to about 50%, about 29% to about 50%, about 30% to about 50%, about 31% to about 50%, about 32% to about 50%, about 33% to about 50%, about 34% to about 50%, about 35% to about 50%, about 36% to about 50%, about 37% to about 50%, about 38% to about 50%, about 39% to about 50%, about 40% to about 50%, or about 40% to about 50%.

[0143] For example, the composition can include at least a first polymer population including polymers modified with a first modification (i.e., a first polymer derivative) and a second polymer population including polymers modified with a second modification (i.e., a second polymer derivative). The total concentration C of polymers (e.g., hydrogel-forming polymers) in the composition is Tcan be at most about 5 mg / ml (e.g., at most about 4 mg / ml, at most about 3 mg / ml, at most about 2 mg / ml, at most about 1.5 mg / ml, at most about 1 mg / ml, at most about 0.9 mg / ml, at most about 0.8 mg / ml, at most about 0.7 mg / ml, at most about 0.6 mg / ml, at most about 0.5 mg / ml, at most about 0.4 mg / ml, at most about 0.3 mg / ml, at most about 0.2 mg / ml, at most about 0.1 mg / ml, or less). The first polymer derivative can have an average DM of about 3% to about 50% (e.g., about 5% to about 35%, about 8% to about 35%, about 10% to about 35%, about 12% to about 35%, about 15% to about 35%, about 20% to about 35%, about 25% to about 35%, or about 30% to about 35%) and an average molecular weight of about 100,000 Daltons to about 5,000,000 Daltons (e.g., about 1,200 00 Daltons to about 5,000,000 Daltons, about 200,000 Daltons to about 5,000,000 Daltons, about 300,000 Daltons to about 5,000,000 Daltons, about 400,000 Daltons to about 5,000,000 Daltons, about 500,000 Daltons to about 5,000,000 Daltons, about 600,000 Daltons to about 5,000,000 Daltons, about 670,000 Daltons to about 50 00000 Daltons, about 1 million to about 5 million, about 1.5 million to about 5 million, about 2 million to about 5 million, about 2.5 million to about 5 million, about 2.6 million to about 5 million, about 3 million to about 50 million 00 Daltons, about 3,500,000 Daltons to about 5,000,000 Daltons, about 3,600,000 Daltons to about 5,000,000 Daltons, about 2,000,000 Daltons to about 4,000,000 Daltons, about 2,500,000 Daltons to about 3,500,000 Daltons, about 2,600,000 Daltons to about 3,600,000 Daltons, about 1,000,000 Daltons to about 2,600,000 Daltons The second polymer derivative can have an average DM of about 3% to about 50% (e.g., about 5% to about 35%, about 8% to about 35%, about 10% to about 35%, about 12% to about 35%, about 15% to about 35%, about 20% to about 35%, about 25% to about 35%, or about 30% to about 35%) and an average molecular weight of about 100,000 to about 5,000,000 daltons (e.g., about 120,000 to about 5,000,000 daltons, about 200,000 to about 35%). Daltons to about 5,000,000 Daltons, about 300,000 Daltons to about 5,000,000 Daltons, about 400,000 Daltons to about 5,000,000 Daltons, about 500,000 Daltons to about 5,000,000 Daltons, about 600,000 Daltons to about 5,000,000 Daltons, about 670,000 Daltons to about 5,000,000 Daltons, about 1,000,000 Daltons to about 5,000,000 Daltons, about 1,500,000 Daltons to about 5,000 000 Daltons, about 2,000,000 Daltons to about 5,000,000 Daltons, about 2,500,000 Daltons to about 5,000,000 Daltons, about 2,600,000 Daltons to about 5,000,000 Daltons, about 3,000,000 Daltons to about 5,000,000 Daltons, about 3,500,000 Daltons to about 5,000,000 Daltons, about 3,600,000 Daltons to about 5,000,000 Daltons, about 2,000,000 Daltons to about 4,000,000 Daltons daltons, about 2,500,000 to about 3,500,000 daltons, about 2,600,000 to about 3,600,000 daltons, about 1,000,000 to about 2,600,000 daltons, about 800,000 to about 2,600,000 daltons, about 700,000 to about 2,500,000 daltons, about 670,000 to about 2,600,000 daltons, or about 600,000 to about 2,500,000 daltons). The ratio (e.g., mass ratio, volume ratio, molar ratio, and / or DM ratio) of the first polymer derivative to the second polymer derivative in the composition can be about 10:1 to about 1:10 (e.g., about 3:1 to about 1:3, or about 2:1 to about 1:2, e.g., about 1:1).The first polymer derivative may be capable of reacting with the second polymer derivative to form a hydrogel. In addition, the first polymer derivative may have an intrinsic viscosity [η] in the composition of at least 3 dL / g (e.g., at least about 5 dL / g, at least about 8 dL / g, at least about 10 dL / g, at least about 12 dL / g, at least about 15 dL / g, at least about 16 dL / g, at least about 17 dL / g, at least about 18 dL / g, at least about 19 dL / g, at least about 20 dL / g, at least about 25 dL / g, or more). The second polymer derivative may have an intrinsic viscosity [η] in the composition of at least 3 dL / g (e.g., at least about 5 dL / g, at least about 8 dL / g, at least about 10 dL / g, at least about 12 dL / g, at least about 15 dL / g, at least about 16 dL / g, at least about 17 dL / g, at least about 18 dL / g, at least about 19 dL / g, at least about 20 dL / g, at least about 25 dL / g, or more).

[0144] The concentration C of the first polymer derivative is * may be about 0.8 to about 5 times (for example, about 0.8 to about 4.5 times, about 0.8 to about 4.0 times, about 0.8 to about 3.5 times, about 0.8 to about 3.0 times, about 0.8 to about 2.5 times, about 0.8 to about 2.0 times, about 0.8 to about 1.5 times, or about 1.0 to about 1.2 times) of C * = 1 / ([η]), where [η] is the intrinsic viscosity of the first polymer derivative.

[0145] The concentration C of the second polymer derivative is * may be about 0.8 to about 5 times (for example, about 0.8 to about 4.5 times, about 0.8 to about 4.0 times, about 0.8 to about 3.5 times, about 0.8 to about 3.0 times, about 0.8 to about 2.5 times, about 0.8 to about 2.0 times, about 0.8 to about 1.5 times, or about 1.0 to about 1.2 times) of C * = 1 / ([η]), where [η] is the intrinsic viscosity of the second polymer derivative.

[0146] In some cases, the composition may comprise a polymer (e.g., a first polymer derivative or The polymer does not contain any cross-linking agent different from the second polymer derivative.

[0147] In some embodiments, the composition does not include any small molecule crosslinkers or PEG-based crosslinkers.

[0148] In a particular example, the first polymer derivative is hyaluronic acid modified with one or more vinyl sulfone groups (e.g., HA-VS), and the second polymer derivative is hyaluronic acid modified with one or more thiol groups (e.g., HA-SH).

[0149] In some cases, the compositions or polymer solutions of the present disclosure may include a crosslinker (e.g., a crosslinker different from the polymer in the composition).

[0150] The crosslinker can be a small molecule crosslinker, a polymeric crosslinker, or a combination thereof. In some embodiments, the crosslinker is a small molecule crosslinker and can include molecules having groups selected from acrylate, maleimide, vinyl sulfone, hydroxysuccinimide, aldehyde, ketone, multicarbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, amine, and / or combinations thereof.

[0151] In some embodiments, the crosslinker is a polymeric crosslinker and may include a polymer having groups selected from acrylate, maleimide, vinyl sulfone, hydroxysuccinimide, aldehyde, ketone, multicarbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, amine, and / or combinations thereof.

[0152] For example, a composition may include a polymer of the present disclosure (e.g., a polymer derivative of the present disclosure) modified with one or more thiol groups and a crosslinker, where the crosslinker may be a vinyl sulfone-containing molecule, e.g., the crosslinker may be selected from the group consisting of divinyl sulfone, bis(vinylsulfonyl)methane, PEG-VS, and 4- or 8-arm-PEGVS.

[0153] In another example, a composition can include a polymer of the present disclosure (e.g., a polymer derivative of the present disclosure) modified with one or more vinyl sulfone groups and a crosslinker, where the crosslinker can be a thiol-containing molecule, for example, the crosslinker can be selected from the group consisting of dithiothreitol, dicysteine, PEG-dithiol, and 4- or 8-arm PEG thiol.

[0154] The hydrogels of the present disclosure can enhance the resistance of materials to drainage at surfaces (e.g., mucosal surfaces). The hydrogels of the present disclosure can have excellent fluid properties similar to those of aqueous solutions. The hydrogels of the present disclosure can have excellent properties against drainage.

[0155] Therefore, the present disclosure also relates to the following embodiments: 1. A composition comprising a polymer capable of forming a hydrogel, the polymer having a concentration C * has a concentration C of about 0.8 to about 5 times that of * = 1 / ([η]), where [η] is the intrinsic viscosity of the polymer. 2. The above polymer has a concentration of C * 2. The composition according to embodiment 1, having a concentration C of from about 0.8 to about 1.5 times that of 3. The above polymer has a concentration of C * 2. The composition according to embodiment 1, wherein the concentration C is about 1.0 to about 1.2 times the concentration C of the composition according to embodiment 1. 4. The above polymer has a concentration of C * 2. The composition according to embodiment 1, wherein the concentration C is about 1.8 to about 2.0 times that of the composition according to embodiment 1. 5. A composition according to any of the preceding embodiments, wherein said polymer has an [η] of at least 3 dL / g. 6. A composition according to any of the previous embodiments, wherein the polymer has an [η] of at least 5 dL / g. 7. A composition according to any of the previous embodiments, wherein the polymer has an [η] of at least 20 dL / g. 8. A composition according to any of the previous embodiments, wherein said polymer is hydrophilic and / or water-soluble. 9. The composition according to any of the previous embodiments, wherein the polymer is selected from the group consisting of polysaccharides, poly(acrylic acid), poly(hydroxyethyl methacrylate), elastin, collagen, and combinations thereof. 10. The composition according to any of the previous embodiments, wherein the polymer is selected from the group consisting of poly(acrylic acid), poly(hydroxyethyl methacrylate), and combinations thereof. 11. The composition according to any of the previous embodiments, wherein the polymer is selected from the group consisting of hyaluronic acid, guar gum, starch, chitosan, chondroitin sulfate, alginate, carboxymethylcellulose, and combinations thereof. 12. The composition according to any of the previous embodiments, wherein the polymer is selected from the group consisting of hyaluronic acid, guar gum, alginate, carboxymethylcellulose, and combinations thereof. 13. A composition according to any of the previous embodiments, wherein said polymer is hyaluronic acid. 14. The composition according to any of the preceding embodiments, wherein the polymer is modified with one or more modifications selected from the group consisting of acrylates, maleimides, vinyl sulfones, N-hydroxysuccinimides, aldehydes, ketones, carbodiimides, carbonates, iodoacetyl, mercaptonicotinamide, quinones, thiols, amines, and combinations thereof. 15. The composition according to any of the preceding embodiments, wherein the polymer is modified with one or more modifications selected from the group consisting of maleimides, vinyl sulfones, N-hydroxysuccinimides, aldehydes, mercaptonicotinamide, quinones, thiols, amines, and combinations thereof. 16. The composition according to any of the preceding embodiments, wherein the polymer comprises a first polymer population comprising the polymer modified with at least a first modification, and a second polymer population comprising the polymer modified with a second modification, wherein the first modification is different from the second modification, and wherein the polymers of the first polymer population react with the polymers of the second polymer population to form a hydrogel. 17. The composition according to embodiment 16, wherein the ratio of polymers in the first polymer population to polymers in the second polymer population is from about 10:1 to about 1:10. 18. The composition according to embodiment 17, wherein the ratio of polymers in the first polymer population to polymers in the second polymer population is from about 3:1 to about 1:3. 19. The composition according to embodiment 18, wherein the ratio of polymers in the first polymer population to polymers in the second polymer population is from about 2:1 to about 1:2. 20. The composition according to any of embodiments 16-19, wherein the first modification and the second modification are independently selected from the group consisting of acrylates, maleimides, vinyl sulfones, N-hydroxysuccinimides, aldehydes, ketones, carbodiimides, carbonates, iodoacetyl, mercaptonicotinamides, quinones, thiols, amines, and combinations thereof. 21. The composition according to any of embodiments 16-19, wherein the first modification and the second modification are independently selected from the group consisting of maleimide, vinyl sulfone, N-hydroxysuccinimide, aldehyde, mercaptonicotinamide, quinone, thiol, amine, and combinations thereof. 22. The polymer is hyaluronic acid, the first polymer population comprises hyaluronic acid modified with one or more vinyl sulfone groups, and the second polymer population comprises one or more chitosan groups. 22. The composition according to embodiment 21, comprising hyaluronic acid modified with an ol group, wherein the polymers of said first polymer population react with the polymers of said second polymer population to form a hydrogel. 23. A composition according to any of the previous embodiments, wherein said polymer has an average molecular weight of about 100,000 Daltons to about 5,000,000 Daltons. 24. A composition according to any of the previous embodiments, wherein said polymer has an average molecular weight of about 120,000 Daltons to about 5,000,000 Daltons. 25. A composition according to any of the previous embodiments, wherein said polymer has an average molecular weight of about 670,000 Daltons to about 5,000,000 Daltons. 26. A composition according to any of the previous embodiments, wherein the polymer has an average molecular weight of about 2,600,000 Daltons to about 5,000,000 Daltons. 27. A composition according to any of the previous embodiments, wherein said polymer has an average molecular weight of about 3,600,000 Daltons to about 5,000,000 Daltons. 28. A composition according to any of the previous embodiments, wherein the polymer has an average molecular weight of about 2,600,000 Daltons to about 3,600,000 Daltons. 29. A composition according to any of the previous embodiments, wherein said polymer has an average molecular weight of about 670,000 Daltons to about 2,600,000 Daltons. 30. A composition according to any of the previous embodiments, wherein the polymer comprises a polymer with modifications, the polymer having an average degree of modification of about 3% to about 50%. 31. A composition according to any of the previous embodiments, wherein the polymer comprises a polymer with modifications, the polymer having an average degree of modification of about 8% to about 35%. 32. A composition according to any of the previous embodiments, wherein the polymer comprises a polymer with modifications, the polymer having an average degree of modification of about 15% to about 35%. 33. A composition according to any of the previous embodiments, wherein the polymer comprises a polymer with modifications, the polymer having an average degree of modification of about 15% to about 30%. 34. A composition according to any of the previous embodiments, wherein the polymer has a concentration C of from about 0.1 mg / ml to about 5 mg / ml. 35. A composition according to any of the previous embodiments, wherein the polymer has a concentration C of about 0.3 mg / ml to about 0.6 mg / ml. 36. A composition according to any of the previous embodiments, further comprising a crosslinker, wherein the crosslinker is a small molecule crosslinker, a polymeric crosslinker, or a combination thereof. 37. The composition according to embodiment 36, wherein the crosslinker is a small molecule crosslinker comprising molecules having groups selected from acrylate, maleimide, vinyl sulfone, hydroxysuccinimide, aldehyde, ketone, multicarbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, amine, and / or combinations thereof. 38. The composition according to embodiment 36, wherein the crosslinker is a polymeric crosslinker comprising a polymer having groups selected from acrylate, maleimide, vinyl sulfone, hydroxysuccinimide, aldehyde, ketone, multicarbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, amine, and / or combinations thereof. 39. The composition according to embodiment 36, wherein the polymer is modified with one or more vinyl sulfone groups and the crosslinker is a thiol-containing molecule. 40. The composition according to embodiment 39, wherein the cross-linking agent is selected from the group consisting of dithiothreitol, dicysteine, PEG-dithiol, and 4- or 8-arm PEG thiol. 41. The composition according to embodiment 36, wherein the polymer is modified with one or more thiol groups and the crosslinker is a vinyl sulfone-containing molecule. 42. The composition according to embodiment 41, wherein the crosslinker is selected from the group consisting of divinyl sulfone, bis(vinylsulfonyl)methane, PEG-VS, and 4- or 8-arm PEGVS. 43. The composition according to any of embodiments 1-35, wherein the composition does not include a crosslinker different from the polymer. 44. A composition according to any of the preceding embodiments, comprising a buffer. 45. The composition according to embodiment 44, wherein the buffer comprises a phosphate buffer. 46. ​​A composition according to any of the preceding embodiments, having a pH of about 3.5 to about 9.0. 47. A composition according to any of the preceding embodiments, having a pH of about 7.0 to about 7.8. 48. A composition according to any of the preceding embodiments, having a pH of about 7.4. 49. A hydrogel formed by a composition according to any one of embodiments 1 to 48. 50. The hydrogel according to embodiment 49, wherein the hydrogel is biocompatible. 51. The hydrogel according to embodiment 49 or 50, having a storage modulus of about 10.0 Pa or less, as measured by dynamic oscillatory shear testing. 52. A hydrogel according to any of embodiments 49-51, having a storage modulus of about 5.0 Pa or less, as measured by dynamic oscillatory shear testing. 53. A hydrogel according to any of embodiments 49-52, having a storage modulus of about 1.0 Pa or less, as measured by dynamic oscillatory shear testing. 54. A hydrogel according to any of embodiments 49-53, having a storage modulus of about 0.6 Pa or less, as measured by dynamic oscillatory shear testing. 55. A hydrogel according to any of embodiments 49-54, having a loss modulus of less than or equal to about 100% of the storage modulus, as measured by dynamic oscillatory shear testing. 56. A hydrogel according to any of embodiments 49-55, having a loss modulus of about 50% or less of the storage modulus, as measured by dynamic oscillatory shear testing. 57. A hydrogel according to any of embodiments 49-56, having a loss modulus of about 25% or less of the storage modulus, as measured by dynamic oscillatory shear testing. 58. The hydrogel according to any of embodiments 49-57, having a complex viscosity of about 0.2 Pa s or less, as measured by dynamic oscillatory shear testing at a frequency greater than about 100 rad / s. 59. The hydrogel according to any of embodiments 49-58, having a complex viscosity of about 0.1 Pa s or less, as measured by dynamic oscillatory shear testing at a frequency greater than about 100 rad / s. 60. The hydrogel according to any of embodiments 49-59, having a complex viscosity of about 0.05 Pa s or less, as measured by dynamic oscillatory shear testing at a frequency greater than about 100 rad / s. 61. A method of making a hydrogel, comprising: providing a composition comprising a polymer according to any one of embodiments 1-48; and exposing the composition to conditions that allow crosslinking of the polymer to form a hydrogel. 62. The method according to embodiment 61, wherein said exposing comprises incubating said polymer solution at about 30°C to about 45°C. 63. The method according to embodiment 62, wherein the exposing comprises incubating the polymer solution at about 37°C. 64. The method according to any of embodiments 61-63, wherein the resulting hydrogel is according to any of embodiments 49-60. [Example]

[0156] The following examples are set forth to provide those of ordinary skill in the art with a complete disclosure and description of how to make and use the present invention, and are not intended to limit the scope of what the inventors regard as their invention, nor are they intended to represent that the following experiments are all or the only experiments performed. Efforts have been made to ensure accuracy with respect to numbers used (e.g., amounts, temperatures, etc.), but some experimental error and deviation must be accounted for. Unless otherwise specified, parts are parts by weight, molecular weight is weight average molecular weight, temperature is in degrees Celsius, and pressure is at or near atmospheric. Standard abbreviations, e.g., bp, base pair(s); kb, kilobase(s); pl, picoliter(s); s or sec, second(s); min, minute(s); Sometimes terms such as h or hr, time(s); aa, amino acid(s); nt, nucleotide(s); im, intramuscular (into the muscle); ip, intraperitoneal (into the abdominal cavity); sc, subcutaneous (under the skin), etc. may be used.

[0157] Example 1 Measurement of HA samples 1.1. Measurement of [η] [η] can be directly measured using a capillary viscometer. For example, three HA samples, namely, Sample A, Sample B, and Sample C, were measured using an Ubbelohde viscometer. These HA samples were purchased from Bloomage Freda Biopharm Co. Ltd. (Shandong, China). The polymers were purchased from ). The polymers were measured by the supplier according to the European Pharmacopoeia. Table 1 shows the intrinsic viscosities ([η]) of Samples A, B, and C measured by an Ubbelohde viscometer.

[0158] [Table 1]

[0159] 1.2 Estimation of polymer molecular weight The molecular weight (MW) of the polymer was calculated from the intrinsic viscosity measured by an Ubbelohde viscometer using the Mark-Hwink-Sakurada equation: [η]=K M (MW) a (In the formula, K M and a (the Mark-Hwink-Sakurada index) is a constant for a given polymer-solvent pair).

[0160] The MW of samples A, B and C were then calculated to be 2.6 MDa, 670 kDa and 120 kDa for samples A, B and C.

[0161] Example 2 Preparation of polymer derivatives 2.1 Preparation of HA-VS Hyaluronic acid (HA) was modified with pendant VS as described by Yu and Chau (Biomacromolecules 2015, 16(1), 56-65). Briefly, HA was dissolved in deionized water (DI water). The concentration ranged from 1 mg / ml to 40 mg / ml depending on the molecular weight (MW) of the HA. For high-MW HA (e.g., MW > 500 kDa), the concentration was lower (e.g., 1 mg / ml to 5 mg / ml), and for low-MW HA (e.g., MW < 500 kDa), the concentration was higher (e.g., 5 mg / ml to 40 mg / ml).

[0162] After complete dissolution, 5 M NaOH was added dropwise to the polymer solution to a final concentration of 0.1 M. Divinyl sulfone (DVS) was immediately added with vigorous mixing. Alternatively, DVS was first dissolved in DI water at 1 mg (DVS) / 8 ml (water) and immediately added to the polymer with vigorous mixing. The molar ratio of DVS to the hydroxyl groups (OH) of HA was at least 1.25:1. At low concentrations of HA, the molar ratio was 6:1 or greater. The reaction time was selected depending on the target degree of modification (DM). For a given reaction time, the degree of modification also depended on the concentrations of both HA and DVS, the temperature, and the final NaOH concentration.

[0163] In HA sample C (molecular weight 120 kDa), the HA concentration was 20 mg / ml, and the molar ratio of DVS to OH of HA was 1.5:1. In HA sample B (molecular weight 670 kDa), In HA sample A, the HA concentration was 10 mg / ml and the molar ratio of DVS to OH of HA was approximately 3:1, and in HA sample B (molecular weight 2.6 MDa), the HA concentration was 2.5 mg / ml and the molar ratio of DVS to OH of HA was approximately 6:1.

[0164] The reaction was stopped by adding 6 M HCl. The polymer was purified by dialysis using a dialysis bag or by tangential flow filtration against DI water (pH 5.5) or acidic DI water adjusted to pH 4 with HCl. The purified polymer was stored as a solution at 4°C. For measurement of the degree of modification (DM), HA-VS was freeze-dried and1 1H NMR was used to measure the

[0165] 2.2 Preparation of HA-SH Hyaluronic acid (HA) was modified with pendant SH groups as described by Yu and Chau (Biomacromolecules 2015, 16(1), 56-65). Briefly, HA was first modified to HA-VS (as described in Example 2.1). The HA-VS solution was purged with N for at least 20 minutes. A 10-fold molar excess of dithiothreitol (DTT) over VS groups or the amount required to make a 0.05 M DTT solution (depending on which DTT concentration is higher) was dissolved in water (pH approximately 5.5) at approximately 400 mg / ml, purged with N for at least 5 minutes, and added to the HA-VS solution. The pH of the HA-VS / DTT solution was approximately 4, and the system was kept purged with N. Then, 1 / 10 of the volume of HA-VS was added to 0.5 M DTT solution. Phosphate buffer (PB) was purged with N2 for at least 5 minutes and added to the HA-VS / DTT solution. The reaction was allowed to proceed for at least 25 minutes. The reaction was stopped by adding 1 M HCl to lower the pH to 3.5-4.5. The polymer was purified by dialysis using a dialysis bag or by tangential flow filtration against DI water or DI water adjusted to pH 4 with HCl. The purified polymer was stored as a solution at 4°C. The degree of modification (DM) was determined by the following procedure: 1 HA-SH was determined by 1 H NMR and Ellmans' assay.

[0166] The synthesis of vinylsulfonated hyaluronic acid (HA-VS) is shown in Figure 1. Thiolated hyaluronic acid (HA-SH) was also synthesized according to Figure 2.

[0167] Example 3 Hydrogel Formation 3.1. Formation of HA-VS / HA-SH polymer-polymer hydrogels The concentrations of HA-VS and HA-SH were first determined. Known volumes of polymer solution were lyophilized, and the dry weight of the polymer was measured. The dry polymer was at least 4 mg to ensure accurate measurements. Alternatively, polymer concentrations were measured by CTAB assay as previously described (Oueslati et al., CTAB turbidimetric method for assaying hyaluronic acid in complex environments and under cross-linked form, Carbohydrate Polymers, 2014). Known concentrations of HA-VS and HA-SH were then adjusted to pH 7.4 by adding 0.5 M PB. The final PB concentration was approximately 0.02 M–0.05 M. Osmolality was then adjusted using 25% NaCl. The polymers were then mixed at various target volume and mass ratios and adjusted to the target concentration by adding phosphate-buffered saline (PBS).

[0168] The polymer was incubated at 37°C for at least 10 hours for hydrogel formation. The hydrogel formation reaction is shown in Figure 3.

[0169] For an HA polymer having an average molecular weight of about 2.6 MDa and an intrinsic viscosity [η] (measured by Ubbelohde viscometer) of about 39.2 dL / g, the concentration of the HA polymer (including both HA-SH and HA-VS) for forming the HA-VS / HA-SH-based polymer-polymer hydrogel of the present disclosure can be as low as about 0.4 mg / ml (i.e., the water content in the hydrogel is greater than about 99.96%).

[0170] In this disclosure, HA-SH / HA-VS polymer-polymer type hydrogel samples A1, A2, A3, A4, A5, A6, A7, A8, and A9 were prepared based on the HA polymer of Sample A in Example 1.1 (having an average molecular weight of about 2.6 MDa and an intrinsic viscosity [η] of about 39.2 dL / g). For Samples A1, A2, A3, and A4, the C of the hydrogel-forming polymer was used. T(e.g., total concentrations of HA-VS and HA-SH) were about 1.2 mg / ml, 0.8 mg / ml, 0.5 mg / ml, and 0.27 mg / ml, respectively; the DM of HA-VS and HA-SH were each about 10%, and the molar and mass ratios of HA-VS to HA-SH were 1:1. For Sample A5, a hydrogel was formed by mixing HA-VS and HA-SH in a DM ratio of 1:1 and a mass ratio of 1:1, and the total concentration C of the hydrogel-forming polymers in the composition was T The DM of HA-VS and HA-SH was about 20%. For sample A6, a hydrogel was formed by mixing HA-VS (8% DM) and HA-SH (8% DM) in a 1:1 DM ratio and a 1:1 mass ratio, and the total concentration of hydrogel-forming polymers in the composition, C T For sample A7, a hydrogel was formed by mixing HA-VS (30% DM) and HA-SH (30% DM) in a 1:1 DM ratio and a 1:1 mass ratio, and the total concentration of hydrogel-forming polymers in the composition, C T The concentration of HA-VS (approximately 10% DM and 0.6 mg / ml) and HA-SH (approximately 5% DM and 0.3 mg / ml) were mixed to form a hydrogel. T For sample A9, the hydrogel was formed by mixing HA-VS (15% DM and 0.6 mg / ml) and HA-SH (10% DM and 0.3 mg / ml). T is approximately 0.9 mg / ml.

[0171] HA-SH / HA-VS polymer-polymer type hydrogel samples B1 and B2 were prepared based on the HA polymer of sample B in Example 1.1 (having an average molecular weight of about 670 kDa and an intrinsic viscosity [η] of about 14 dL / g). For samples B1 and B2, the C of the HA polymer (containing both HA-SH and HA-VS) was Twere approximately 1.8 mg / ml and 1.08 mg / ml, respectively. The DM of HA-SH and HA-VS was 30%, and the molar and mass ratios of HA-SH to HA-VS were 1:1.

[0172] Sample C1 of the HA-SH / HA-VS polymer-polymer type hydrogel was prepared based on the HA polymer of Sample C in Example 1.1 (having an average molecular weight of about 120 kDa and an intrinsic viscosity [η] of about 3.7 dL / g). For Sample C1, the C of the HA polymer (containing both HA-SH and HA-VS) was prepared. T The DM of HA-SH and HA-VS was 30%, and the molar and mass ratios of HA-SH to HA-VS were 1:1.

[0173] 3.2 Formation of HA-SH polymer-polymer hydrogel Sample A of HA polymer (having a weight-average molecular weight of about 2.6 MDa and an intrinsic viscosity [η] of about 39.2 dL / g) was used. HA-SH was prepared according to Example 2.2. HA-SH was adjusted to pH 7.4 by adding 0.5 M PB. The final concentration of PB was about 0.02 M to 0.05 M. The DM of the polymer was about 10%. The polymer was then adjusted to the target concentration by adding phosphate-buffered saline (PBS). Sample F of the HA-SH polymer-polymer hydrogel was prepared by adding 0.5 M PB to a C of about 1.7 mg / ml. T It was made with.

[0174] The polymer was incubated at 37°C for at least 10 hours for hydrogel formation.

[0175] 3.3. Polymer-Small Molecule Crosslinker Hydrogel Formation DTT was mixed with HA-VS (HA-VS was prepared based on the HA polymer of Sample A) at a molar ratio of 3:1 (DTT:VS). A hydrogel was formed after 30 minutes. The hydrogel was dialyzed against double deionized water for 2 days and then recovered. HA-VS had a weight-average molecular weight of approximately 1.9 MDa, an intrinsic viscosity [η] of approximately 24.5 dL / g, and a total polymer (e.g., hydrogel-forming polymer) concentration C of approximately 3.8 mg / ml. T The DM of HA-VS was 20%. The resulting HA-SH / DTT hydrogel was named HA-SH / DTT hydrogel sample H.

[0176] Example 4 Characterization of Hydrogels 4.1 Elastic modulus of the prepared hydrogel samples (1) Preparation of hydrogel samples for measurement After mixing the hydrogel-forming polymers as described in Example 3, the mixed polymers were uniformly applied onto a 50 mm (D) x 1 mm (H) mold. The mold was attached to the metal plate by covering the plate with parafilm. The mold with the polymer was placed in a humidified chamber to allow hydrogel formation. For hydrogel formation, the polymer was maintained at 37°C for 24 hours. After the formation of the hydrogel (i.e., the HA-VS / HA-SH-based polymer-polymer hydrogel), the entire parafilm-mold-gel assembly was separated from the metal plate and mounted on a 50 mm plate fixture. Next, The mold was then removed, leaving the Parafilm and the formed hydrogel on the fixture. The Parafilm was then tightly wrapped around the lower plate. This assembly was then placed in a DMA apparatus (ARES rheometer, TA Instruments, New Castle, DE) to measure the mechanical properties using parallel plates with a diameter of 50 mm.

[0177] Alternatively, the hydrogel-forming polymers were mixed as described in Example 3 and then incubated in a centrifuge tube at 37° C. for at least 10 hours (e.g., 24 hours unless otherwise specified). The formed hydrogels (i.e., HA-VS / HA-SH-based polymer-polymer hydrogels) were then introduced into the lower plate or lower bucket of a cone-plate, double-gap, or concentric cylinder fixture, and the mechanical properties were measured using an Anton Paar rheometer.

[0178] (2) Elastic modulus of HA-VS / HA-SH polymer-polymer hydrogels Three samples of HA polymer (i.e., Samples A, B, and C described in Examples 1.1 and 1.2 above) were modified according to Examples 2.1 and 2.2, and the modified polymers were used to form hydrogels of the present disclosure according to Example 3.1. The formed hydrogels were measured according to Example 4.1. DMA measurements and / or direct observation (to determine whether the sample flows like water and cannot be attached to the DMA instrument) were used to determine whether the polymer formed a hydrogel rather than remaining as a polymer solution. For DMA measurements, a high G' value relative to the G" value in the linear viscoelastic region (LVR) (e.g., G" / G' < 1) was used as an indication of hydrogel formation.

[0179] The results are summarized in Table 2 below. Mechanical properties were measured in the LVR region of the gels. The inventors surprisingly found that the intrinsic viscosity [η] of the hydrogel-forming polymers in the composition was relatively high (e.g., at least about 3 dL / g, as measured by an Ubbelohde viscometer) and the total concentration C of the hydrogel-forming polymers (e.g., HA-VS and HA-SH) in the composition was 0.01. TWe found that when the ρ is relatively low (e.g., less than about 5 mg / ml, e.g., 1.2 mg / ml, 0.8 mg / ml, 0.5 mg / ml, 1.8 mg / ml, 1.08 mg / ml, and 3.96 mg / ml, respectively), very soft hydrogels can be formed, as indicated by the G′ and G″ of the hydrogels. G′ and G″ were 10.0 Pa or less, indicating the formation of a hydrogel.

[0180] [Table 2]

[0181] Figure 4 shows the elastic modulus (using a frequency sweep test) of HA-VS / HA-SH polymer-polymer hydrogel samples A1, A2, and A3 prepared according to Example 3.1. The elastic modulus of the hydrogels was measured using an Anton Paar MCR502 rheometer with a concentric double-gap fixture. The strain was 1% for all tests.

[0182] Figure 5 shows the elastic modulus (using strain sweep testing) of the HA-VS / HA-SH polymer-polymer hydrogel sample A8 prepared according to Example 3.1 using a concentric double gap fixture with an Anton Paar MCR502. The frequency was 1 rad / s. The C of the HA-VS / HA-SH polymer-polymer hydrogel sample A8 T was 0.9 mg / ml, which was lower than 5 mg / ml. Both G' and G'' were below 10.0 Pa. Also, G'' was less than G', indicating the formation of a hydrogel.

[0183] 6A and 6B show the elastic modulus and complex viscosity (using strain and frequency sweep tests) of Sample A9, a HA-VS / HA-SH-based polymer-polymer hydrogel prepared according to Example 3.1, using an Anton Paar MCR302 with a 50 mm cone-plate fixture. The frequency for the strain sweep test was 5 rad / s, and the strain for the frequency sweep test was 5%. The C of Sample A9, a HA-VS / HA-SH-based polymer-polymer hydrogel, was measured. T was 0.9 mg / ml, which was lower than 5 mg / ml. Both G' and G'' were below 10.0 Pa. Also, G'' was less than G', indicating the formation of a hydrogel.

[0184] (3) Elastic modulus of HA-SH polymer-polymer hydrogel 7A and 7B show strain and frequency sweep tests of Sample F, a 10% DM HA-SH polymer-polymer hydrogel prepared according to Example 3.2. In FIG. 7, G′, G″, and Eta * are the storage modulus, loss modulus, and complex viscosity, respectively. The hydrogels were measured using an Anton Paar MCR 302 rheometer with a 50 mm cone-plate fixture. The strain sweep test was performed at 5 rad / s, and the frequency sweep test was performed at 5% strain. C for sample F of the HA-SH polymer-polymer hydrogel T was 1.7 mg / ml, which was lower than 5 mg / ml. Both G' and G'' were below 10.0 Pa. Also, G'' was less than G', indicating the formation of a hydrogel.

[0185] (4) Elastic modulus of HA-SH polymer-small molecule crosslinker hydrogel Figures 8A and 8B show the strain sweep and frequency sweep tests of Sample H, an HA-SH / DTT hydrogel prepared according to Example 3.3. The strain sweep test was performed at 1 rad / s, and the frequency sweep test was performed at 5% strain. Here, G', G'', and Eta *refers to the storage modulus, loss modulus, and complex viscosity, respectively. C of sample H of HA-SH / DTT hydrogel T was approximately 3.8 mg / ml, lower than 5 mg / ml. Both G' and G'' were below 10.0 Pa. Also, G'' was less than G', indicating the formation of a hydrogel.

[0186] The above results show that for all hydrogels tested, the storage modulus (G') is higher than its corresponding loss modulus (G'') in dynamic oscillatory shear tests at frequencies between approximately 0.1 rad / s and approximately 10 rad / s, and the G' values ​​remain relatively constant, indicating that the materials are indeed gels (i.e., very soft, solid-like crosslinked networks) rather than viscous solutions.

[0187] 4.2 Yield strain of hydrogels The yield strain can be evaluated by strain sweep tests (Figures 5 to 8). For example, the results in Figures 5 and 7 show that the hydrogel has a low yield stress (about 0.1 Pa to about 1 Pa) and a relatively high yield strain (about 30%).

[0188] 4.3 Recovery of hydrogels from yielding by strain sweep tests The low yield point predicted that the hydrogels of the present disclosure would be mechanically unstable. Surprisingly, the mechanical properties of the hydrogels of the present disclosure can be similar before and after fracture. In this example, the HA-VS / HA-SH-based polymer-polymer hydrogel sample A3 prepared according to Example 3.1 was used to test the recovery of the hydrogel from yield using a strain sweep test.

[0189] Sample A3, a HA-VS / HA-SH polymer-polymer hydrogel, was placed in a concentric double-gap fixture on an Anton Paar MCR502. Strain sweep tests were performed from 0% to 5000%, and time sweep tests (1% strain and 1 rad / s) were performed before and after the hydrogel fracture to evaluate the hydrogel's elastic modulus. The hydrogel was allowed to equilibrate for approximately 5 minutes before each measurement. G' and G'' were approximately 0.11 Pa and 0.02 Pa before fracture, and 0.11 Pa and 0.05 Pa after fracture. The results show that the elastic modulus was nearly identical before and after fracture.

[0190] 4.4 Complex viscosity of hydrogels From the frequency sweep test of the hydrogel shown in Figures 6 to 8, it is clear that the complex viscosity of the hydrogel becomes smaller at relatively slow frequencies (for example, Eta at frequencies of about 10 rad / s to 100 rad / s). * <0.01 Pa). These results demonstrate that the hydrogel of the present disclosure can be easily applied to an entire surface with little force. In addition, the viscosity of the hydrogel of the present disclosure increases exponentially with low shear rates and remains stable at rest.

[0191] Example 5 Preparation of fluorescent dye conjugation 5.1. Tagging polymers with fluorescent probes The hydrogel of the present disclosure was made fluorescent by tagging the polymer with a fluorescent dye. Thiol-modified fluorescein was used as an example dye. The dye was produced by the reaction of aminofluorescein with N-acetyl-S-trityl-L-cysteine ​​in DMF catalyzed by HOBT / HBTU. The product was precipitated in ether three times and dried under vacuum. The dried product was deprotected with a mixture of TFA:HO:TIPS (18:1:1) for 1 hour. The final product was precipitated in ether, dried under vacuum, and stored at 4°C. The progress of the reaction was monitored by mass spectrometry.

[0192] 5.2. Fluorescent dye conjugation In one example, HA-VS was adjusted to pH 8.8 by adding 1.5 M Tris buffer at pH 8.8 to a final buffer concentration of 0.15 M. The solution was purged with N for at least 30 minutes. Then, a vinyl sulfone reactive dye (the amino group of the lysine side chain is fluorescent) prepared according to Example 5.1 or purchased from GL Biochem (Shanghai) Ltd. was added. A cysteine-lysine dipeptide conjugated with a dye and capped at the amino end with an amide, i.e., AC-Cys-Lys (FAM), was dissolved in Tris buffer and added to the polymer for 12 hours. Because the conjugation efficiency was not high and not all VS groups were covered by the reaction, the molar ratio of dye to VS groups could be up to 1:1 for low DM (e.g., DM = 8%) polymers. Ideally, at least 80% of the VS groups should not be covered by the dye after the reaction. The polymer was then dialyzed against DI water adjusted to a pH of approximately 4 with HCl. The storage conditions for the purified fluorescently labeled polymer were the same as for HA-VS. The fluorescently labeled HA-VS was then mixed with HA-SH to form a hydrogel (as described in Example 3).

[0193] Formed hydrogels (e.g., Sample A6 formed by mixing HA-VS (8% DM) and HA-SH (8% DM) in a 1:1 DM ratio and a 1:1 mass ratio, where the total concentration of hydrogel-forming polymers in the composition was C T The hydrogel (concentration of which was approximately 0.45 mg / ml) was observed to be resistant to dilution in water due to its special mechanical properties (e.g., properties discussed in Example 4 above). As shown in Figures 9A-9D, the hydrogel of the present disclosure was able to move freely in water without dissolving (Figures 9A and 9B), and after 24 hours, the hydrogel had not dissolved and was still able to move freely in water without dissolving (Figures 9C and 9D).

[0194] While preferred embodiments of the present invention have been shown and described herein, it will be apparent to those skilled in the art that such embodiments are provided by way of example only. It is not intended that the present invention be limited by the specific examples provided herein. While the present invention has been described with reference to the above specification, the descriptions and examples of the embodiments herein are not intended to be construed in a limiting sense. Numerous variations, changes, and substitutions will occur to those skilled in the art without departing from the invention. Furthermore, it should be understood that all aspects of the present invention are not limited to the specific depictions, configurations, or relative proportions set forth herein, which depend upon a variety of conditions and variables. It should be understood that various alternatives to the embodiments of the present invention described herein may be utilized in practicing the present invention. It is therefore intended that the present invention also cover any such alternatives, modifications, variations, or equivalents. The appended claims define the scope of the invention, and it is intended that methods and structures within the scope of the claims, and their equivalents, be covered thereby.

Claims

1. 1. A composition comprising one or more hydrogel-forming polymers, each said hydrogel-forming polymer has an intrinsic viscosity [η] in the composition of at least 3 dL / g as measured by an Ubbelohde viscometer; The total concentration C of the hydrogel-forming polymer in the composition Ｔ is 0.3 mg / ml to 5 mg / ml, The composition is the following a) or b): a) a composition capable of forming a hydrogel in the presence of a crosslinker, the composition comprising one or more hydrogel-forming polymers and a crosslinker different from the one or more hydrogel-forming polymers, wherein the hydrogel-forming polymer is a derivative of hyaluronic acid modified with a thiol group or a vinyl sulfone group; b) a composition that contains one or more hydrogel-forming polymers, but does not contain a crosslinker different from the one or more hydrogel-forming polymers, and that is capable of forming a hydrogel in the absence of a crosslinker, wherein the hydrogel-forming polymer is a derivative of hyaluronic acid modified with a thiol group or a vinyl sulfone group; The hydrogel has at least one of the following properties 1) to 3): 1) The storage modulus G' measured by a dynamic oscillatory shear test is 10.0 Pa or less; 2) A complex viscosity of 0.2 Pa s or less as measured by a dynamic oscillatory shear test at a frequency greater than 100 rad / s; 3) A yield strain of at least 10% as measured by dynamic oscillatory strain sweep testing.

2. The composition of claim 1 , wherein at least a portion of the hydrogel-forming polymer is contained in the formed hydrogel in the composition.

3. 3. The composition of claim 1, wherein the hydrogel-forming polymer is hydrophilic and / or water-soluble.

4. 10. The composition of claim 1, wherein the hydrogel-forming polymer has an average molecular weight of from 100,000 Daltons to 5,000,000 Daltons.

5. The composition of claim 1 , wherein the crosslinker is a small molecule crosslinker, a polymeric crosslinker, or any combination thereof.

6. 6. The composition of claim 5, wherein the crosslinker is a small molecule crosslinker comprising a molecule having groups selected from acrylate, maleimide, vinyl sulfone, hydroxysuccinimide, aldehyde, ketone, multicarbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, amine, and any combination thereof.

7. 6. The composition of claim 5, wherein the crosslinker is a polymeric crosslinker comprising a polymer having groups selected from acrylate, maleimide, vinyl sulfone, hydroxysuccinimide, aldehyde, ketone, multicarbodiimide, carbonate, iodoacetyl, mercaptonicotinamide, quinone, thiol, amine, and any combination thereof.

8. 6. The composition of claim 5, wherein the cross-linking agent is selected from the group consisting of dithiothreitol, dicysteine, PEG-dithiol, 4- or 8-arm PEG thiol, divinyl sulfone, bis(vinylsulfonyl)methane, PEG-VS, and 4- or 8-arm PEGVS.

9. The composition according to any one of claims 1 to 8, comprising a phosphate buffer.

10. The composition of any one of claims 1 to 9, having a pH of from 3.5 to 9.

0.

11. A hydrogel formed by the composition of any one of claims 1 to 10.

12. 12. The hydrogel of claim 11, which is biocompatible.

13. 13. The hydrogel of claim 11 or 12, wherein the hydrogel has a loss modulus G'' that is less than or equal to 100% of its storage modulus G' as measured in a dynamic oscillatory shear test.

14. 1. A method of making a hydrogel, comprising: a) providing a composition according to any one of claims 1 to 10; b) exposing the composition to conditions that allow the formation of the hydrogel; A method comprising:

15. 15. The method of claim 14, wherein said exposing comprises incubating said composition at 30°C to 45°C.

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