Oxygen activated adhesives

Abstract

Herein disclosed is an adhesive composition comprising: an organic macromolecule comprising nucleophilic functional groups, wherein each of the nucleophilic functional groups comprises an amino group, a thiol group, a hydroxyl group, or a carboxyl group; and one or more moieties each comprising a ring structure having (i) one carbon attached with an aldehyde or a ketone, (ii) two carbons each having a hydroxyl group, and (iii) one carbon attached with a substituent group; wherein each of the nucleophilic functional groups grafted with one of the one or more moieties has the aldehyde or the ketone converted into a coupling moiety and each of the nucleophilic functional groups not grafted with the one or more moieties remains as the nucleophilic functional group, wherein each of the one or more moieties oxidizes into a diketone when the adhesive composition is subjected to a stimulus, and wherein the diketone forms one or more crosslinkages with the nucleophilic functional groups. A method for forming the adhesive composition, its uses, and a kit, are disclosed.

Description

OXYGEN ACTIVATED ADHESIVESCross-Reference to Related Application

[0001] This application claims the benefit of priority of Singapore Patent Application No. 10202500333W, filed 6 February 2025, the content of it being hereby incorporated by reference in its entirety for all purposes.Technical Field

[0002] The present disclosure relates to an adhesive composition. The present disclosure also relates to a method for forming the adhesive composition, uses of the adhesive composition, and a kit.Background

[0003] Traditionally, catechol may be used as a chemical cross-linking functional group in chemical curing adhesives and tissue adhesives systems. However, all catechol-based adhesives tend to require heat or two-part mixing strategies to induce cross -linking. For example, oxidizing agents are required (e.g., periodate [IO4]’ ) or metal chelators (e.g., Fe+3, Cu+2) may be traditionally utilized in order to activate / oxidize catechol (by periodate) to quinone or chelate-mediated (by metal cations) cross-linking macromolecules. This tends to prevent widespread application as cumbersome mixing equipment or mixing nozzles are required. This also prevents the independent application (e.g., coating) and delayed cross-linking (adhesive curing) of the catechol-based adhesives, which may be an unmet industrial and clinical need. Also, traditional catechol-grafted macromolecules may hold promise as non-toxic bioadhesives but tends to face challenges of adhesive stability and preparation.

[0004] Moreover, many traditional self-curing chemical cross-linking mechanisms may be known to include those of cyanoacrylates (anionic polymerization), silicone (condensation), and acrylates (free radical polymerization), but these do not allow bioadhesion or any adhesion to hydrated or wet substrates or underwater bio / adhesion.

[0005] There is thus a need to provide for a solution that addresses one or more of the limitations mentioned above.Summary

[0006] In a first aspect, there is provided for an adhesive composition comprising: an organic macromolecule comprising nucleophilic functional groups, wherein each of the nucleophilic functional groups comprises an amino group, a thiol group, a hydroxyl group, or a carboxyl group; andone or more moieties each comprising a ring structure having (i) one carbon attached with an aldehyde or a ketone, (ii) two carbons each having a hydroxyl group, and (iii) one carbon attached with a substituent group;wherein each of the nucleophilic functional groups grafted with one of the one or more moieties has the aldehyde or the ketone converted into a coupling moiety and each of the nucleophilic functional groups not grafted with the one or more moieties remains as the nucleophilic functional group,wherein each of the one or more moieties oxidizes into a diketone when the adhesive composition is subjected to a stimulus, and wherein the diketone forms one or more crosslinkages with the nucleophilic functional groups.

[0007] In another aspect, there is provided for a method for forming the adhesive composition of various embodiments of the first aspect, the method comprising:providing the organic macromolecule; andmixing the organic macromolecule with a precursor of the one or more moieties.

[0008] In another aspect, there is provided for a method for adhering or sealing biological tissues or surfaces, the method comprising:applying the adhesive composition of various embodiments of the first aspect to one or more biological tissues or surfaces;subjecting the adhesive composition to the stimulus; andcontacting the biological tissues or surfaces together to adhere or seal the biological tissues or surfaces, respectively.

[0009] In another aspect, there is provided for a kit comprising:the organic macromolecule defined in various embodiments of the first aspect; andthe precursor of the one or more moieties defined in one of the aspect mentioned above related to a method for forming the adhesive composition,wherein the kit contains the organic macromolecule in a separate compartment from the precursor.Brief Description of the Drawings

[0010] The drawings are not necessarily to scale, emphasis instead generally being placed upon illustrating the principles of the present disclosure. In the following description, various embodiments of the present disclosure are described with reference to the following drawings, in which:

[0011] FIG. 1 shows the one step synthesis of the adhesive composition in an anaerobic environment.

[0012] FIG. 2 is a plot on the dynamic mechanical analysis via parallel plate rheometry, wherein a sample PEI-(5-OMe-3,4-DBA)2o, an adhesive composition of the present disclosure, displayed immediate gelation and crosslinking upon exposure to oxygen atmosphere.

[0013] FIG. 3 is a plot on the dynamic mechanical analysis via parallel plate rheometry, wherein a sample PEI-(PhCHO)20 shows no gelation and crosslinking.

[0014] FIG. 4A shows the ’H-NMR of model reaction- 1 (^H NMR in DMSO-d ) as mentioned with respect to FIG. 9.

[0015] FIG. 4B shows the1H-NMR model reaction-2 (^H NMR in CD3OD, except PA which was recorded in DMSO-^J as mentioned with respect to FIG. 9.

[0016] FIG. 4C shows the chemical structures of plausible products under model reaction-2.

[0017] FIG. 4D shows oxygen triggered polymerisation of PEI-(5-OMe-3,4-DBA)20- The polymerization reaction follows the same mechanistic pathway as shown for various products formation in MR-2 (also see FIG. 4B and FIG. 4C).

[0018] FIG. 4E shows theNMR spectra of PEI-(5-OMe-3,4-DBA)2o before and after O2 bubbling (15 min) in CD3OD. TheNMR spectra of PEI-(3,4,5-TBA)2o in CD3OD.

[0019] FIG. 5A shows UV-vis spectra of PEI-(3-Br-4,5-DBA)2o under oxygen bubbling.

[0020] FIG. 5B shows the decay of 334.5 nm absorption peak of PEI-(3-Br-4,5-DB A)2o under oxygen bubbling.

[0021] FIG. 5C shows the UV-vis spectra of PEI-(5-OMe-3,4-DBA)2o under oxygen bubbling.

[0022] FIG. 5D shows the decay of 342.5 nm absorption peak of PEI-(5-OMe-3,4-DBA)2O under oxygen bubbling.

[0023] FIG. 5E shows the UV-vis spectra of PEI-(5-OMe-3,4-DBA)2o under air bubbling.

[0024] FIG. 5F shows the decay of 342.5 nm absorption peak of PEI-(5-OMe-3,4-DBA)2O under air bubbling.

[0025] FIG. 6 is a plot of the storage modulus of PEI-(3,4-DBA)2o, PEI-(5-NO2-3,4-DBA)2O, PEI-(6-N02-3,4-DBA)2O, PEI-(3-Br-4,5-DBA)20, PEI-(2-Br-4,5-DBA)2o,PEI-(5-OMe-3,4-DBA)2o, and PEI-(2-OMe-4,5-DBA)20in PBS (30 wt%, w / v) at the end of 30 min study. Data presented as mean ± standard deviation, n = 3, p -values are calculated using one-way ANOVA with Tukey test, / ? < 0.05, * = statistical difference.

[0026] FIG. 7 is a bar graph plot of the mechanical and adhesive properties of the adhesive compositions. FIG. 7 shows a comparison of 180-degree peel adhesion strength of PEI-(3,4-DBA)20, PEI-(5-N02-3,4-DBA)2o, PEI-(6-N02-3,4-DBA)2o, PEI-(3-Br-4,5-DBA)2o, PEI-(2-Br-4,5-DBA)2o, PEI-(5-OMe-3,4-DBA)2o, and PEI-(2-OMe-4,5-DBA)2O (30 wt% in EtOH, w / v). Data presented as mean ± standard deviation, n = 3, p -values are calculated using one-way ANOVA with Tukey test, p < 0.05, * = statistical difference.

[0027] FIG. 8 shows oxygen promoted polymerization of catechol grafted PEI.

[0028] FIG. 9 compares the two model reaction schemes for a Schiff-base of catechol, PEI-(5-OMe-3,4-DBA)2o, which reacts to produce various products in presence of oxygen under model reaction-2. Particularly, FIG. 9 shows catechol methyl ester (MDMB) remains unreacted towards propyl amine (MR-1: model Reaction 1) but the corresponding Schiff-base of catechol, PA-(5-OMe-3,4-DBA)2o, reacts with propylamine (PA) (MR-2: model Reaction 2) to produce various products in presence of oxygen.

[0029] FIG. 10 is a plot of the oxygen triggered crosslinking kinetics of three different catechols grafted to polyamine dendrimer - PEI-(3,4-DBA)2o, PEI-(5-OMe-3,4-DBA)2O, and PEI-(5-N02-3,4-DBA)2o.

[0030] FIG. 11 shows the cyclic voltammogram of PEI-(BA)2o, PEI-(MDMB)2o, and PEI-(5-OMe-3,4-DBA)2o in PBS (1.0 mM, pH ~ 7.2) using 3 mm diameter GC working electrode vs Ag / AgCl at 298 ± 2 K in a Faraday cage. Scan rate -0.1 V.s-1.

[0031] FIG. 12A is a plot of the shelf stability of PEI-(5-OMe-3,4-DBA)20in MeOD solution at ambient temperature (24 °C), 4 °C, and at - 20 °C. In the present disclosure, MeOD denotes deuterated methanol.

[0032] FIG. 12B is a plot showing the effect of air and oxygen bubble into the MeOD solution of PEI-(5-OMe-3,4-DBA)2o at room temperature.

[0033] FIG. 12C is a table comparing theXH-NMR shelf stability study of PEI-(5-OMe-3,4-DBA)2O under argon atmosphere at ambient temperature (24°C), 4°C and -20°C. Note: NMR tube caps used in the current study (Norell, TC-5-EVA-W-100) are made of ethylene vinyl acetate (EVA) polymer, through which O2 can permeate at a slow rate. Though there is no report on the rate of oxygen permeation through EVA polymer, however the rate of oxygen permeation reportedly drops at low temperature on polypropylene polymer.

[0034] FIG. 12D is a plot of the effect of air / oxygen bubble into the MeOD solution of PEI-(5-N02-3,4-DBA)2O at room temperature.

[0035] FIG. 12E is a plot of the effect of air / oxygen bubble into the MeOD solution of PEI-(3-Br-4,5-DBA)2o at room temperature.

[0036] FIG. 13A is an apparent viscosity representative graph of PEI-(BA)2o in PBS (30 wt%, w / v).

[0037] FIG. 13B is an apparent viscosity representative graph of PEI-(MDMB)2o in PBS (30 wt%, w / v).

[0038] FIG. 14A is a plot of an apparent viscosity of PEI-(B A)20, PEI-(MDMB)2o, and PEI-(5-OMe-3,4-DBA)2o in PBS (30 wt%, w / v). Data presented as mean ± standard deviation, n = 3, p -values are calculated using one-way ANOVA with Tukey test, p < 0.05, * = statistical difference.

[0039] FIG. 14B is a plot of storage modulus of PEI-(BA)20, PEI-(MDMB)20, and PEI-(5-OMe-3,4-DBA)2o in PBS (30 wt%, w / v) at the end of 30 min study. Data presented as mean ± standard deviation, n = 3, p -values are calculated using one-way ANOVA with Tukey test, p < 0.05, * = statistical difference.

[0040] FIG. 14C is a plot of shear strain versus modulus graph of PEI-(BA)2o, PEI-(MDMB)2O, and PEI-(5-OMe-3,4-DBA)2o in PBS after 30 min curing. Data presented as mean ± standard deviation, n = 3, p -values are calculated using one-way ANOVA with Tukey test, p < 0.05, * = statistical difference.

[0041] FIG. 15A is a plot of storage and loss modulus (G’ and G”) representative graphs of PEI-(BA)20-

[0042] FIG. 15B is a plot of storage and loss modulus (G’ and G”) representative graphs of PEI-(MDMB)2O-

[0043] FIG. 15C is a plot of storage and loss modulus (G’ and G”) representative graphs of PEI-(5-OMe-3,4-DBA)2o-

[0044] FIG. 16A is a plot of lap shear adhesion strength representative graph of PEI-(BA)20.

[0045] FIG. 16B is a plot of lap shear adhesion strength representative graph of PEI-(MDMB)2O-

[0046] FIG. 17A is a comparison of lap shear adhesion strength between PEI-(5-OMe-3,4-DBA)2O and PEI-(MDMB)2o (30 wt% in PBS, w / v). Data presented as mean ± standard deviation, n = 3,p -values are calculated using one-way ANOVA with Tukey test, / ? < 0.05, * = statistical difference.

[0047] FIG. 17B is a comparison of lap shear modulus of toughness between PEI-(5-OMe-3,4-DBA)2o and PEI-(MDMB)2o (30 wt% in PBS, w / v). Data presented as mean ± standard deviation, n = 3,p -values are calculated using one-way ANOVA with Tukey test, / ? < 0.05, * = statistical difference.

[0048] FIG. 17C is a comparison of 180° peel adhesion strength between PEI-(5-OMe-3,4-DBA)2O and PEI-(MDMB)2o (30 wt% in PBS, w / v). Data presented as mean ± standard deviation, n = 3,p -values are calculated using one-way ANOVA with Tukey test, / ? < 0.05, * = statistical difference.

[0049] FIG. 17D is a comparison of 180° peel adhesion modulus of toughness between PEI-(5-OMe-3,4-DBA)2o and PEI-(MDMB)2o (30 wt% in PBS, w / v). Data presented as mean ± standard deviation, n = 3,p -values are calculated using one-way ANOVA with Tukey test, / ? < 0.05, * = statistical difference.

[0050] FIG. 18A shows the 180° peel adhesion strength representative graph of PEI-(BA)20.

[0051] FIG. 18B shows the 180° peel adhesion strength representative graph of PEI-(MDMB)2O-

[0052] FIG. 18C shows the 180° peel adhesion strength representative graph of PEI-(5-OMe-3,4-DBA)2o.

[0053] FIG. 19 shows the ’H NMR of PEI, PEI-(5-N02-3,4-DBA)2o, PEI-(6-NO2-3,4-DBA)2O, PEI-(3-Br-4,5-DBA)2o, PEI-(2-Br-4,5-DBA)20, PEI-(5-OMe-3,4-DBA)20, and PEI-(2-OMe-4,5-DBA)2o in MeOD at room temperature.

[0054] FIG. 20 shows the UV-vis spectra of PEI-(5-N02-3,4-DBA)2o, PEI-(3-Br-4,5-DBA)2O, PEI-(5-OMe-3,4-DBA)2o, and PEI-(2-OMe-4,5-DBA)20in methanol (MeOH) at room temperature.

[0055] FIG. 21A is a cyclic voltammogram of PEI-(5-NO2-3,4-DBA)20 and PEI-(3,4-DBA)2O, in PBS (pH ~ 7.2) using 3 mm diameter GC working electrode vs Ag / AgCl at 298 ± 2 K in a Faraday cage. Scan rate -0.1 V.s-1.

[0056] FIG. 21B is a cyclic voltammogram of PEI-(3-Br-4,5-DBA)2o and PEI-(3,4-DBA)2O in PBS (pH ~ 7.2) using 3 mm diameter GC working electrode vs Ag / AgCl at 298 ± 2 K in a Faraday cage. Scan rate -0.1 V.s-1.

[0057] FIG. 21C is a cyclic voltammogram of PEI-(5-OMe-3,4-DBA)2o and PEI-(3,4-DBA)2O in PBS (pH ~ 7.2) using 3 mm diameter GC working electrode vs Ag / AgCl at 298 ± 2 K in a Faraday cage. Scan rate -0.1 V.s-1.

[0058] FIG. 21D is a cyclic voltammogram of PEI-(6-NO2-3,4-DBA)20 and PEI-(3,4-DBA)2O in PBS (pH ~ 7.2) using 3 mm diameter GC working electrode vs Ag / AgCl at 298 ± 2 K in a Faraday cage. Scan rate -0.1 V.s-1.

[0059] FIG. 21E is a cyclic voltammogram of PEI-(2-Br-4,5-DBA)2o and PEI-(3,4-DBA)2O in PBS (pH ~ 7.2) using 3 mm diameter GC working electrode vs Ag / AgCl at 298 ± 2 K in a Faraday cage. Scan rate -0.1 V.s-1.

[0060] FIG. 21F is a cyclic voltammogram of PEI-(2-OMe-4,5-DBA)2o and PEI-(3,4-DBA)2O in PBS (pH ~ 7.2) using 3 mm diameter GC working electrode vs Ag / AgCl at 298 ± 2 K in a Faraday cage. Scan rate -0.1 V.s-1.

[0061] FIG. 21G is a cyclic voltammogram of PEI-(B A)2o and PEI-(3,4-DB A)2o in PBS (pH ~ 7.2) using 3 mm diameter GC working electrode vs Ag / AgCl at 298 ± 2 K in a Faraday cage. Scan rate -0.1 V.s-1.

[0062] FIG. 22 is a plot of oxidation potential (V) Vs peak current (I) graph of PEI-(sCat)2o compounds (0.13 mM) in PBS (1.0 mM, pH ~ 7.2) using 3 mm diameter GC working electrode vs Ag / AgCl at 298 ± 2 K in a Faraday cage. Scan rate -0.1 V.s-1.

[0063] FIG. 23A shows UV-vis of PEI-(2-OMe-3,4-DBA)2o under oxygen bubble.

[0064] FIG. 23B shows decay of 388.5 nm absorption peak of PEI-(2-OMe-3,4-DBA)2o under oxygen bubble.

[0065] FIG. 24A shows the shelf stability of PEI-(5-N02-3,4-DBA)2o in MeOD solution at ambient temperature (24°C) and at 4°C.

[0066] FIG. 24B shows the shelf stability of PEI-(3-Br-4,5-DBA)2o in MeOD solution at ambient temperature (24 °C) and at 4°C.

[0067] FIG. 24C shows the shelf stability of PEI-(5-OMe-3,4-DBA)20in MeOD solution at ambient temperature (24°C) and at 4°C.

[0068] FIG. 25 is a bar plot of the apparent viscosity of PEI-(3,4-DBA)2o, PEI-(5-NO2-3,4-DBA)2O, PEI-(6-N02-3,4-DBA)2O, PEI-(3-Br-4,5-DBA)20, PEI-(2-Br-4,5-DBA)2o, PEI-(5-OMe-3,4-DBA)2o, and PEI-(2-OMe-4,5-DBA)20in PBS (30 wt%, w / v). Data presented as mean ± standard deviation, n = 3, p -values are calculated using one-way ANOVA with Tukey test, / ? < 0.05, * = statistical difference.

[0069] FIG. 26A is a plot of the representative apparent viscosity of PEI-(5-NO2-3,4-DBA)2O in PBS (30 % w / v).

[0070] FIG. 26B is a plot of the representative apparent viscosity of PEI-(3-Br-4,5-DBA)2O in PBS (30 % w / v).

[0071] FIG. 26C is a plot of the representative apparent viscosity of PEI-(5-OMe-3,4-DBA)2O in PBS (30 % w / v). Inset shows a magnified version of the same plot of FIG.26C.

[0072] FIG. 26D is a plot of the representative apparent viscosity of PEI-(6-NO2-3,4-DBA)2O in PBS (30 % w / v).

[0073] FIG. 26E is a plot of the representative apparent viscosity of PEI-(2-Br-4,5-DBA)2O in PBS (30 % w / v).

[0074] FIG. 26F is a plot of the representative apparent viscosity of PEI-(2-OMe-4,5-DBA)2O in PBS (30 % w / v).

[0075] FIG. 27A shows the representative storage modulus of PEI-(5-NO2-3,4-DBA)20 in PBS (30 % w / v) over 30 mins curing period.

[0076] FIG. 27B shows the representative storage modulus of PEI-(3-Br-4,5-DBA)2o in PBS (30 % w / v) over 30 mins curing period.

[0077] FIG. 27C shows the representative storage modulus of PEI-(5-OMe-3,4-DBA)2o in PBS (30 % w / v) over 30 mins curing period.

[0078] FIG. 27D shows the representative storage modulus of PEI-(6-NO2-3,4-DBA)20 in PBS (30 % w / v) over 30 mins curing period.

[0079] FIG. 27E shows the representative storage modulus of PEI-(2-Br-4,5-DBA)2o in PBS (30 % w / v) over 30 mins curing period.

[0080] FIG. 27F shows the representative storage modulus of PEI-(2-OMe-4,5-DBA)2o in PBS (30 % w / v) over 30 mins curing period.

[0081] FIG. 28A shows the representative shear strain versus moduli graph of PEI-(5-N02-3,4-DBA)2O.

[0082] FIG. 28B shows the representative shear strain versus moduli graph of PEI-(3-Br-4,5-DBA)20.

[0083] FIG. 28C shows the representative shear strain versus moduli graph of PEI-(5-OMe-3,4-DBA)2o.

[0084] FIG. 28D shows the representative shear strain versus moduli graph of PEI-(6-N02-3,4-DBA)2O.

[0085] FIG. 28E shows the representative shear strain versus moduli graph of PEI-(2-Br-4,5-DBA)20.

[0086] FIG. 28F shows the representative shear strain versus moduli graph of PEI-(2-OMe-4,5-DBA)2o.

[0087] FIG. 29A shows the representative lap shear adhesion strength of PEI-(5-NO2- 3.4-DBA)2O in ethanol (30 % w / v) over 30 mins on collagen film.

[0088] FIG. 29B shows the representative lap shear adhesion strength of PEI-(3-Br- 4.5-DBA)2O in ethanol (30 % w / v) over 30 mins on collagen film.

[0089] FIG. 29C shows the representative lap shear adhesion strength of PEI-(5-OMe-3,4-DBA)2O in ethanol (30 % w / v) over 30 mins on collagen film.

[0090] FIG. 29D shows the representative lap shear adhesion strength of PEI-(6-NO2-3,4-DBA)2O in ethanol (30 % w / v) over 30 mins on collagen film.

[0091] FIG. 29E shows the representative lap shear adhesion strength of PEI-(2-Br-4,5-DBA)2O in ethanol (30 % w / v) over 30 mins on collagen film.

[0092] FIG. 29F shows the representative lap shear adhesion strength of PEI-(2-OMe- 4.5-DBA)2O in ethanol (30 % w / v) over 30 mins on collagen film.

[0093] FIG. 30A is a bar graph plot of the mechanical and adhesive properties of the adhesive compositions. FIG. 30A shows a comparison of lap shear adhesion strength of PEI-(3,4-DBA)2O, PEI-(5-N02-3,4-DBA)2O, PEI-(6-N02-3,4-DBA)2O, PEI-(3-Br- 4.5-DBA)2O, PEI-(2-Br-4,5-DBA)2o, PEI-(5-OMe-3,4-DBA)2o, and PEI-(2-OMe-4,5-DBA)2O (30 wt% in EtOH, w / v). Data presented as mean ± standard deviation, n = 3, p -values are calculated using one-way ANOVA with Tukey test, p < 0.05, * = statistical difference.

[0094] FIG. 30B is a comparison of lap shear adhesion modulus of toughness of PEI-(3,4-DBA)2O, PEI-(5-N02-3,4-DBA)2O, PEI-(6-N02-3,4-DBA)2O, PEI-(3-Br-4,5-DBA)2O, PEI-(2-Br-4,5-DBA)2o, PEI-(5-OMe-3,4-DBA)2o, and PEI-(2-OMe-4,5-DBA)2O (30 wt% in EtOH, w / v). Data presented as mean ± standard deviation, n = 3, p -values are calculated using one-way ANOVA with Tukey test, p < 0.05, * = statistical difference.

[0095] FIG. 31A shows the representative 1800peel adhesion strength of PEI-(5-N02-3,4-DBA)2O in ethanol (EtOH) (30 % w / v) over 30 mins on collagen film.

[0096] FIG. 3 IB shows the representative 1800peel adhesion strength of PEI-(3-Br- 4.5-DBA)2O in EtOH (30 % w / v) over 30 mins on collagen film.

[0097] FIG. 31C shows the representative 1800peel adhesion strength of PEI-(5-OMe-3,4-DBA)2o in EtOH (30 % w / v) over 30 mins on collagen film.

[0098] FIG. 3 ID shows the representative 1800peel adhesion strength of PEI-(6-N02-3,4-DBA)2O in EtOH (30 % w / v) over 30 mins on collagen film.

[0099] FIG. 3 IE shows the representative 1800peel adhesion strength of PEI-(2-Br- 4,5-DBA)2O in EtOH (30 % w / v) over 30 mins on collagen film.

[0100] FIG. 3 IF shows the representative 1800peel adhesion strength of PEI-(2-OMe-4,5-DBA)2o, in EtOH (30 % w / v) over 30 mins on collagen film.

[0101] FIG. 32A shows a cyclic voltammogram of 2-OMe-4,5-DBA in PBS (1.0 mM, pH - 7.2) using 3 mm diameter GC working electrode vs Ag / AgCl at 298 ± 2 K in a Faraday cage. Scan rate -0.1 V.s-1.

[0102] FIG. 32B shows a cyclic voltammogram of 5-OMe-3,4-DBA in PBS (1.0 mM, pH ~ 7.2) using 3 mm diameter glassy carbon (GC) working electrode vs Ag / AgCl at 298 ± 2 K in a Faraday cage. Scan rate -0.1 V.s-1.

[0103] FIG. 33 is a graphical abstract showing the oxygen (or oxygen in air) trigger polymerization (crosslinking) of the PEI-(5-OMe-3,4-DBA)2o monomer.

[0104] FIG. 34A shows the skin sensitisation results of in chemico DPRA analysis (OECD 442C) of Cysteine and Lysine peptide depletion with PEI-(3,4-DBA)2o and PEI-(5-OMe-3,4-DBA)2o adhesives leachates (20 mg polymeric mixture / mL leachate) using cinnamic aldehyde and CH3CN as positive and negative control, respectively.

[0105] FIG. 34B shows the in vitro bacterial reverse mutation (AMES OECD 471) assay of PEI-(3,4-DBA)2o and PEI-(5-OMe-3,4-DBA)2o adhesives at 37°C at a treatment concentration of 5mg / plate, in the presence of a metabolic activation system. Data presented as mean ± standard deviation, n = 3 for graph a, n=100 for graph b and c, p -values are calculated using one-way ANOVA with Tukey test, p < 0.05, * = statistical difference.

[0106] FIG. 34C shows the in vitro bacterial reverse mutation (AMES OECD 471) assay of PEI-(3,4-DBA)2o and PEI-(5-OMe-3,4-DBA)2o adhesives at 37°C at a treatment concentration of 5mg / plate, in the absence of a metabolic activation system. Data presented as mean ± standard deviation, n = 3 for graph a, n=100 for graph b and c, p -values are calculated using one-way ANOVA with Tukey test, p < 0.05, * = statistical difference.

[0107] FIG. 35 shows the reparation of disk samples: bacteria (E.coli and S. aureus) were cultured in tube until optical density of 0.1 was achieved, then applied on the agar plate, after which coated filter paper disks (d=6.5 mm) were placed. Plate was incubated for 24 hrs with hourly monitoring. Paper coated with ampicillin was used as positive control.

[0108] FIG. 36 is a table of the list of compounds and the surface concentration of coating on the surface of the filter paper (diameter - 55 mm, mass - 220 mg).

[0109] FIG. 37A shows pictures of coated disks (d=6.5 mm) planted on agar bed with E.coli bacterial species planted on the agar bed, compared against time.

[0110] FIG. 37B shows pictures of coated disks (d=6.5 mm) planted on agar bed with S. aureus bacterial species planted on the agar bed, compared against time.

[0111] FIG. 37C shows pictures of coated disks (d=6.5 mm) after dip washing in deionized water, planted on agar bed with E.coli bacterial species planted on the agar bed, compared against time.

[0112] FIG. 37D shows pictures of coated disks (d=6.5 mm) after dip washing in deionized water, planted on agar bed with S. aureus bacterial species planted on the agar bed, compared against time.

[0113] FIG. 38 shows the optical results of inhibition area for E.coli and S. ureus bacteria after 24 h of incubation - comparison of neat (no washing) and washed samples (in deionized water (DI) and EtOH).

[0114] FIG. 39A is a table containing label explanation on the samples tested.

[0115] FIG. 39B is a bar plot of inhibition area measurements, with area of disk (0.33 cm2) as reference against E.coli, and S. aureus, without washing. (n=3, p<0.05) Error bars for normalized samples is a sum of standard deviations of all variables in equation.

[0116] FIG. 39C is a bar plot of zone of inhibition against E.coli bacteria, with samples washed by dipping 3 times in 1 mL of DI water and samples washed in EtOH using the same method, normalized against the inhibition area of unwashed samples. (n=3, p<0.05) Error bars for normalized samples is a sum of standard deviations of all variables in equation.

[0117] FIG. 39D is a bar plot analogous as in FIG. 39C, but for S. aureus bacteria. (n=3, p<0.05) Error bars for normalized samples is a sum of standard deviations of all variables in equation.

[0118] FIG. 40A shows the procedure of preparing leachates from soaking disk (d=0.65 cm) for 3+ hours in DI water and mixing it 1: 1 with bacteria in broth;

[0119] FIG. 40B shows the optical density at 600 nm from prepared mixture after 24 h incubation in 37 °C (n=4).

[0120] FIG. 41 showcase differences between catechol isomers. Several catechol isomers were investigated on their mechanical performance as bioadhesives by Schiff-base graft to amine-rich branched polymer, resulting in varying degree of performance in both aqueous an anaerobic media, showcasing possible pathways over traditional DOPA mimics.

[0121] FIG. 42A shows grafting the catechol-aldehyde isomer on the amine macromolecule via Schiff base. Resulting compound can undergo oxidation in aqueousformulations to quinone intermediate which crosslinks further with amines via Michael addition, or form Schiff base with the aryl ring.

[0122] FIG. 42B shows the catechol isomers investigated for their stability, mechanical performance and reactivity, using traditional isomers as controls.

[0123] FIG. 43 is a table for catechol isomers chosen for screening. Shelf-life stability based on proton NMR integration loss of formed Schiff base.

[0124] FIG. 44 shows possible reaction of grafted catechol isomer.

[0125] FIG. 45 shows the proton NMR of bPEI-g-(2,5-DBA)20, bPEI and 2,5-DBA in MeOD. The disappearance of aldehyde peak paired with presence of Schiff base in the final product is taken as indicator of finished reaction.

[0126] FIG. 46 shows various UV-vis data of bPEI-g-(DBA)2o during grafting, showcasing spectra of aldehyde (No PEI) and Schiff base after grafting on bPEI.

[0127] FIG. 47 shows various UV-vis data of grafted catechol isomers after addition of 10 vol.% PBS.

[0128] FIG. 48 A shows the UV-vis data of bPEI-g-(DBA)2o control (before Air, before O2) with gas bubbles through solution at a rate of - 2 cm3. min1, and wherein bPEI-g-(2,5-DBA)2o when bubbled with air.

[0129] FIG. 48B shows the UV-vis data of bPEI-g-(DBA)2o control (before Air, before O2) with gas bubbles through solution at a rate of - 2 cm3. min1, and wherein bPEI-g-(2,5-DBA)2o when bubbled with oxygen.

[0130] FIG. 48C shows the UV-vis data of bPEI-g-(DBA)2o control (before Air, before O2) with gas bubbles through solution at a rate of - 2 cm3. min1, and wherein bPEI-g-(3,5-DBA)2o when bubbled with air.

[0131] FIG. 48D shows the UV-vis data of bPEI-g-(DBA)2o control (before Air, before O2) with gas bubbles through solution at a rate of - 2 cm3. min1, and wherein bPEI-g-(3,5-DBA)2o when bubbled with oxygen.

[0132] FIG. 49 is a table comparison of oxygen concentration.

[0133] FIG. 50A shows example of cyclic voltammetry curve of NPA-g-(3,4-DBA), showing oxidation potential in PBS against Ag / AgCl as RE, using glassy carbon and Pt as WE and CE, scan speed 100 mV-s-1, vertex potentials <-1.2 V; 1.2 V>.

[0134] FIG. 50B shows a scheme highlighting the energy change in between states of catechol isomer, calculated through Density Functional Theory simulation in order to predict the most reactive catechol isomers.

[0135] FIG. 50C shows compiled CV results of catechol isomers grafted on linear propylamine, highlighting the oxidation potential Eox, sorted in order of value of complex modulus G*.

[0136] FIG. 50D shows DFT calculations showing change of energy between base and oxidized states for aromatic ring and Cl atom.

[0137] FIG. 51A is a map of first oxidation peaks the normalized oxidation currents. In PBS; <-1.2;1.2> V, start voltage at 0 V, scan speed 100 mV-s’l, Ag / AgCl as a reference electrode, glassy carbon as a working electrode and Pt as a counter electrode.

[0138] FIG. 51B shows cyclic voltammetry curves of n-MPA-g-(2,3-DBA)2o in PBS; <-1.2;1.2> V, start voltage at 0 V, scan speed 100 mV-s'l, Ag / AgCl as a reference electrode, glassy carbon as a working electrode and Pt as a counter electrode.

[0139] FIG. 51C shows cyclic voltammetry curves of n-MPA-g-(2,4-DBA)2o in PBS; <-1.2;1.2> V, start voltage at 0 V, scan speed 100 mV-s'l, Ag / AgCl as a reference electrode, glassy carbon as a working electrode and Pt as a counter electrode.

[0140] FIG. 51D shows cyclic voltammetry curves of n-MPA-g-(3,4-DBA)2o in PBS; <-1.2;1.2> V, start voltage at 0 V, scan speed 100 mV-s'l, Ag / AgCl as a reference electrode, glassy carbon as a working electrode and Pt as a counter electrode.

[0141] FIG. 51E shows cyclic voltammetry curves of n-MPA-g-(3,5-DBA)2o in PBS; <-1.2;1.2> V, start voltage at 0 V, scan speed 100 mV-s'l, Ag / AgCl as a reference electrode, glassy carbon as a working electrode and Pt as a counter electrode.

[0142] FIG. 51F shows cyclic voltammetry curves of n-MPA-g-(2,5-DBA)2oin PBS; <-1.2;1.2> V, start voltage at 0 V, scan speed 100 mV-s'l, Ag / AgCl as a reference electrode, glassy carbon as a working electrode and Pt as a counter electrode.

[0143] FIG. 52A shows H1NMR Spectra of bPEI-g-(3,4-DBA)20in CD3OD, showing peak from Schiff base in range 7.75-8.25 ppm, and aryl proton peaks at 6.25-7.5 ppm range.

[0144] FIG. 52B shows peaks (b+c+d) of grafted catechol isomers (referred to in FIG.52A) in CD3OD, exposed to 10 vol. % deuterated saline. Error peaks obtained fromstandard deviation of ratio of MeOD:bPEI peaks (3.33-3.29 ppm):(2.95-2.20 ppm), relative to mean value (n=9).

[0145] FIG. 52C shows integrated Schiff base peaks (a) of grafted catechol isomers (referred to in FIG. 52A) in CD3OD, subjected to 10 vol. % of D2O, normalized against integrated values before the addition of D2O. Error peaks obtained from standard deviation of ratio of MeOD:bPEI peaks (3.33-3.29 ppm):(2.95-2.20 ppm), relative to mean value (n=9).

[0146] FIG. 53 is a table ofNMR (AUC) Schiff base before and after addition of D2O.

[0147] FIG. 54 is a table ofNMR (AUC) Schiff base before and after addition of PBS.

[0148] FIG. 55A shows proton NMR of bPEI-g-(2,5-DBA)20and bPEI-g-(3,5-DBA)2O in MeOD after treating the compound with free oxygen gas for Ih.

[0149] FIG. 55B shows comparison of aryl ring integration between control (synthesized compound without any additions), compound after 3.5h of adding 10 vol.% PBS, 10 vol.% D2O and compound treated with free oxygen for Ih.

[0150] FIG. 55C shows Schiff base integration ranges between control (synthesized compound without any additions), compound after 3.5h of adding 10 vol.% PBS, 10 vol.% D2O and compound treated with free oxygen for Ih.

[0151] FIG. 56A relates to parallel plate rheometry results from investigation of mechanical performance of grafted catechol isomers. FIG. 56A shows example of curing 30 wt. % bPEI-g-(3,4-DBA)2o in PBS, oscillation (10 Hz), 6.28 rad-s1for 30 min. All samples were 30 wt. % bPEI-g-(DBA)2o in PBS, with*ANOVA statistical analysis with Tukey correction (n = 300; p < 0.05).

[0152] FIG. 56B shows example of amplitude sweep of 30 wt. % bPEI-g-(3,4-DBA)2o in PBS, from which the toughness modulus is determined (integration of highlighted area), 1-1000 % shear strain. All samples were 30 wt. % bPEI-g-(DBA)2o in PBS, with*ANOVA statistical analysis with Tukey correction (n = 300; p < 0.05).

[0153] FIG. 56C shows comparison of complex modulus recorded at 30 min time, with samples ranked with respect to gelation time. All samples were 30 wt. % bPEI-g-(DBA)2O in PBS, with*ANOVA statistical analysis with Tukey correction (n = 300; p < 0.05).

[0154] FIG. 56D shows max shear stress obtained from 1-100% amplitude sweep. All samples were 30 wt. % bPEI-g-(DBA)2o in PBS, with*ANOVA statistical analysis with Tukey correction (n = 300; p < 0.05).

[0155] FIG. 57A is a representative graph showing the oscillation rheometry results of one of the grafted catechol isomer (30 wt.% in PBS).

[0156] FIG. 57B is a representative graph showing the oscillation rheometry results of one of the grafted catechol isomer (30 wt.% in PBS).

[0157] FIG. 57C is a representative graph showing the oscillation rheometry results of one of the grafted catechol isomer (30 wt.% in PBS).

[0158] FIG. 57D is a representative graph showing the oscillation rheometry results of one of the grafted catechol isomer (30 wt.% in PBS).

[0159] FIG. 57E is a representative graph showing the oscillation rheometry results of one of the grafted catechol isomer (30 wt.% in PBS).

[0160] FIG. 57F is a representative graph showing the oscillation rheometry results of one of the grafted catechol isomer (30 wt.% in PBS).

[0161] FIG. 58 A is a representative graph showing the decrease of moduli of one of the grafted catechol isomers (30 wt.% in PBS) under amplitude sweep.

[0162] FIG. 58B is a representative graph showing the decrease of moduli of one of the grafted catechol isomers (30 wt.% in PBS) under amplitude sweep.

[0163] FIG. 58C is a representative graph showing the decrease of moduli of one of the grafted catechol isomers (30 wt.% in PBS) under amplitude sweep.

[0164] FIG. 58D is a representative graph showing the decrease of moduli of one of the grafted catechol isomers (30 wt.% in PBS) under amplitude sweep.

[0165] FIG. 58E is a representative graph showing the decrease of moduli of one of the grafted catechol isomers (30 wt.% in PBS) under amplitude sweep.

[0166] FIG. 58F is a representative graph showing the decrease of moduli of one of the grafted catechol isomers (30 wt.% in PBS) under amplitude sweep.

[0167] FIG. 58G shows a stress-strain plot of one of the grafted catechol isomers under amplitude sweep.

[0168] FIG. 58H shows a stress-strain plot of one of the grafted catechol isomers under amplitude sweep.

[0169] FIG. 581 shows a stress-strain plot of one of the grafted catechol isomers under amplitude sweep.

[0170] FIG. 58 J shows a stress-strain plot of one of the grafted catechol isomers under amplitude sweep.

[0171] FIG. 58K shows a stress-strain plot of one of the grafted catechol isomers under amplitude sweep.

[0172] FIG. 58L shows a stress-strain plot of one of the grafted catechol isomers under amplitude sweep.

[0173] FIG. 59 is a Bar graph showcasing mean gelation time of grafted catechol isomers (30 wt.% in PBS) during oscillation rheometry (n=5). All other tested grafted catechol aldehydes either did not exhibit gelation time (bPEI-g-(2,3-DBA)2o, bPEI-g-(2,4-DBA)2O), or the gelation time was lower than the time of first measurement (t< 15 s for bPEI-g-(3,4,5-TBA)2o).

[0174] FIG. 60A shows representative storage and loss moduli of bPEI-g-(2,3-DBA)2O during rheometry when subjected to constant voltage of -IV - no gelation. ANOVA statistical analysis with Tukey correction (n = 5; p = 0.05);

[0175] FIG. 60B shows representative storage and loss moduli of bPEI-g-(2,3-DBA)2o during rheometry when subjected to constant voltage of -2V - gelation occurs. ANOVA statistical analysis with Tukey correction (n = 5; p = 0.05).

[0176] FIG. 60C shows representative storage and loss moduli of bPEI-g-(2,3-DBA)2o during rheometry when subjected to constant voltage of -3V - gelation occurs. ANOVA statistical analysis with Tukey correction (n = 5; p = 0.05).

[0177] FIG. 60D is a comparison of complex modulus at 30 min mark for bPEI-g-(2,3-DBA)2O, using bPEI-g-(3,4-DBA)2o as control, showing activation at lower voltages. ANOVA statistical analysis with Tukey correction (n = 5; p = 0.05);

[0178] FIG. 61 is a bar graph showcasing mean gelation time of grafted catechol isomers (30 wt.% in PBS) during oscillation rheometry (n=5) under constant voltage. bPEI-g-(2,3-DBA)2o did not exhibit gelation at 0, -1 and 3 V.

[0179] FIG. 62A shows lap shear performance of the bPEI-g-(DBA)2o that showed gelation during parallel plate rheometry: example of lap shear test result of 30 wt. % bPEI-g-(DBA)2o in phosphate-buffered saline, showing the elastic region and shear strength of the sample when cured for 30 min on hydrated collagen strips.

[0180] FIG. 62B shows lap shear performance of the bPEI-g-(DBA)2o that showed gelation during parallel plate rheometry: example of lap shear under the same conditions but using ethanol as solvent.

[0181] FIG. 62C shows yielding point of different bPEI-g-(DBA)2o against Evicell™ reference when dissolved in PBS (white bar) and EtOH (grey bar) with applied ANOVA statistical analysis with Tukey correction (n = 5, n = 4 for bPEI-g-(2,5-DBA)2o ; p = 0.05) - no significant difference seen.

[0182] FIG. 63 A shows, in EtOH medium, lap shear results of collagen samples glued using one of the grafted catechol isomers in (30 wt.% in alcohol) on 2x2 cm area and sheared with speed of 10 mm / min.

[0183] FIG. 63B shows, in EtOH medium, lap shear results of collagen samples glued using one of the grafted catechol isomers in (30 wt.% in alcohol) on 2x2 cm area and sheared with speed of 10 mm / min.

[0184] FIG. 63C shows, in EtOH medium, lap shear results of collagen samples glued using one of the grafted catechol isomers in (30 wt.% in alcohol) on 2x2 cm area and sheared with speed of 10 mm / min.

[0185] FIG. 63D shows, in EtOH medium, lap shear results of collagen samples glued using one of the grafted catechol isomers in (30 wt.% in alcohol) on 2x2 cm area and sheared with speed of 10 mm / min.

[0186] FIG. 63E shows, in EtOH medium, lap shear results of collagen samples glued using one of the grafted catechol isomers in (30 wt.% in alcohol) on 2x2 cm area and sheared with speed of 10 mm / min.

[0187] FIG. 63F shows, in EtOH medium, lap shear results of collagen samples glued using one of the grafted catechol isomers in (30 wt.% in alcohol) on 2x2 cm area and sheared with speed of 10 mm / min.

[0188] FIG. 64A shows, in PBS medium, lap shear results of collagen samples glued using grafted catechol isomers (30% w / w in PBS) on 2x2 cm area and sheared with speed of 10 mm / min.

[0189] FIG. 64B shows, in PBS medium, lap shear results of collagen samples glued using grafted catechol isomers (30% w / w in PBS) on 2x2 cm area and sheared with speed of 10 mm / min.

[0190] FIG. 64C shows, in PBS medium, lap shear results of collagen samples glued using grafted catechol isomers (30% w / w in PBS) on 2x2 cm area and sheared with speed of 10 mm / min.

[0191] FIG. 64D shows, in PBS medium, lap shear results of collagen samples glued using grafted catechol isomers (30% w / w in PBS) on 2x2 cm area and sheared with speed of 10 mm / min.

[0192] FIG. 64E shows, in PBS medium, lap shear results of collagen samples glued using grafted catechol isomers (30% w / w in PBS) on 2x2 cm area and sheared with speed of 10 mm / min.

[0193] FIG. 64F shows, in PBS medium, lap shear results of collagen samples glued using grafted catechol isomers (30% w / w in PBS) on 2x2 cm area and sheared with speed of 10 mm / min.

[0194] FIG. 65 shows Pictures of 30 wt. % bPEI-g-(2,5-DBA)2o (top) and bPEI-g-(3,5-DBA)2O (bottom) in ethanol on hydrated collagen strips after a lap shear test, with observed cohesive failures with singular mixed (adherend and cohesive) failure.

[0195] FIG. 66 shows the measurement of layer thickness via cross section: image via Olympus BX5, 50x0.75 lens, focus stacking via Infinity Analyze software.

[0196] FIG. 67 is a bar plot of Toughness of grafted catechol isomers when assuming uniform layer of adhesive with thickness 367 pm. Significance determined via ANOVA with Tukey correction (p<0.05, n=5).

[0197] FIG. 68A shows absorbance over time of grafted bPEI-g-(DB A)2o in methanol when subjected to 10 vol. % of PBS to determine the oxidation kinetics: Absorbance at peak for grafted catechol isomers on bPEI, normalized against absorbance at 1 min, showing changes when subjected to 10 vol. % of PBS, Wavelengths: 336 nm (bPEI-g-(2,4-DBA)20), 350 nm (bPEI-g-(2,5-DBA)20), 343 nm (bPEI-g-(3,4-DBA)20), 323 nm (bPEI-g-(3,5-DBA)20).

[0198] FIG. 68B shows the oxidation kinetics of bPEI-g-(2,5-DBA)2o after contact with oxidative agents - air / oxygen / 10 vol. % PBS vs. control (no contact) to determine the impact of aqueous medium compared to oxidation through gas for, peak at 350 nm.

[0199] FIG. 68C shows the oxidation kinetics of bPEI-g-(3,5-DBA)2o with same method, peak at 324 nm for oxygen, 323 nm for air, 10 vol. % PBS and control.

[0200] FIG. 68D is a comparison of zero-order kinetics of linear fits at forced intercept b=l.

[0201] FIG. 69 is a table of values of zero-order kinetics from UV-vis absorbance (normalized), forced intercept=l.

[0202] FIG. 70A shows range of chemical interactions for grafted triol aldehyde, showcasing versatility of phenolic adhesive compositions and adhesive of the present disclosure.

[0203] FIG. 70B shows example of Schiff base reaction, resulting in polyethyleneimine grafted phenol, with varying range of gelation time and modulus after 30 min when prepared in aqueous buffer.

[0204] FIG. 70C shows structures of tested triol aldehydes in the present disclosure.

[0205] FIG. 71A is a comparison of apparent viscosity values obtained through 30 s of steady state parallel plate rheometry with shear rate of 10 s’1(n=120). All statistical difference done via one way ANOVA with Tukey correction (p<0.05).

[0206] FIG. 7 IB is a comparison of storage modulus values at the end of 30 min constant oscillation (6.28 rad • s’1, n=5). All statistical difference done via one way ANOVA with Tukey correction (p<0.05).

[0207] FIG. 71C is a comparison of maximum lap shear strength obtained on hydrated collagen samples (6 x 2 cm strips, 2 x 2 cm area, n=5) in regards to adhesive concentration in PBS. All statistical difference done via one way ANOVA with Tukey correction (p<0.05).

[0208] FIG. 7 ID is a comparison of lap shear strength values on various substrates (n=5, with n=3 for wooden samples due to samples exceeding machine limit). All statistical difference done via one way ANOVA with Tukey correction (p<0.05).

[0209] FIG. 72A is a representative graph showing curing of bPEI-g-(3,4,5-TBA)2o 30 wt.% in PBS under constant oscillation.

[0210] FIG. 72B is a comparison of apparent viscosity values for grafted pyrogallol with and without trimethyl borate protective group, obtained through 30 s of steady state parallel plate rheometry with shear rate of 10 s’1(n=120); all statistical analysis performed via one way ANOVA with Tukey correction (p<0.05).

[0211] FIG. 72C is a comparison of storage modulus at the end of the curing stage for not protected and trimethyl borate protected samples (n=5); all statistical analysis performed via one way ANOVA with Tukey correction (p<0.05).

[0212] FIG. 72D is a comparison of toughness values obtained through 1-1000% amplitude sweep for not protected and trimethyl borate protected samples (n=5); all statistical analysis performed via one way ANOVA with Tukey correction (p<0.05).

[0213] FIG. 73 is a comparison of lap shear strength on wooden coupons using 500N load cell, with variety in solvent and whether the pyrogallol group was protected by trimethyl borate group, with one way ANOVA statistical analysis with Tukey correction (n=5, p<0.05).

[0214] FIG. 74 is a table showing the masses for components used for preparing the spraying solutions.

[0215] FIG. 75 is a photo of solutions of trihydroxy aldehyde and bPEI undergoing a reaction in methanol.

[0216] FIG. 76 is a photo of wooden clips of grafted 3,4,5-TBA after lap shear.

[0217] FIG. 77 is a photo of wooden clips of grafted 2,3,4-TBA after lap shear.

[0218] FIG. 78 is a photo of wooden clips of grafted 2,4,5-TBA after lap shear.

[0219] FIG. 79 is a plot of the lap shear results of the grafted 2,3,4-TBA samples on wooden coupons.

[0220] FIG. 80 is a plot of the lap shear of the grafted 3,4,5-TBA samples on wooden coupons.

[0221] FIG. 81 is a plot of the lap shear of the grafted 2,4,5-TBA samples on wooden coupons.

[0222] FIG. 82 is a plot of the lap shear strength for all tested samples - ANOVA statistical difference noted only betweedn3,4,5-TBA and 2,4,5-TBA (n=5, p<0.05)

[0223] FIG. 83 shows aluminum strips with applied grafted 2,4,5-TBA. Poor adhesion to the surface and low concentration led to spillage even after 30 min.

[0224] FIG. 84 is a plot comparing previous lap shear tests on wooden coupons using 500N load cell and speed of 10 mm / min.

[0225] FIG. 85 is a table showing amounts of aldehyde and TMB added for protection of hydroxyl groups.

[0226] FIG. 86 is a table showing amounts of aldehyde and bPEI added to achieve the final adhesive with 20% grafting, with noted leftover methanol

[0227] FIG. 87 is a table showing amounts of adhesive added to achieve the 30 wt.% in PBS. MeOH was considered when measuring the amounts but was not included in final weight calculation.

[0228] FIG. 88 is a plot of the mean apparent viscosity of both TMB protected adhesives in relation to their neat, unprotected variants.

[0229] FIG. 89 shows representative G’ and G” curves for grafted and TMB protected 3,4,5-TBA (left) and 2,3,4-TBA (right), show in respect to their unprotected variants. There can be seen slight delay in the gelation time, albeit it is still not captured for grafted 2,3,4-TBA adhesive composition.

[0230] FIG. 90 is a plot of the mean storage modulus value at 30 min mark for both protected adhesives, shown in relation to their unprotected variants, with ANOVA statistical difference (n=5, n=4 for unprotected 2,3,4-TBA, p<0.05).

[0231] FIG. 91 shows the stress-strain curves for all samples tested, in comparison to their unprotected variants.

[0232] FIG. 92 is a plot of the mean toughness comparison, with ANOVA statistical difference (n=5, n=4 for unprotected 2,3,4-TBA, p<0.05).

[0233] FIG. 93 is a table showing masses used to prepare solutions for lap shear adhesion. Residual methanol was taken into account while weighing the adhesive amount, but was not added to final weight calculation.

[0234] FIG. 94 is a photo of wooden clips of grafted 2,3,4-TBA protected with TMB in MeOH after lap shear.

[0235] FIG. 95 is a photo of wooden clips of grafted 2,3,4-TBA protected with TMB in PBS after lap shear.

[0236] FIG. 96 is a photo of wooden clips of grafted 3,4,5-TBA protected with TMB in MeOH after lap shear.

[0237] FIG. 97 is a photo of wooden clips of grafted 3,4,5-TBA protected with TMB in PBS after lap shear.

[0238] FIG. 98 is a plot of the lap shear of the grafted 2,3,4-TBA protected with TMB samples on wooden coupons - PBS (blue) vs MeOH (red).

[0239] FIG. 98 is a plot of lap shear of the grafted 3,4,5-TBA protected with TMB samples on wooden coupons - PBS (blue) vs MeOH (red).

[0240] FIG. 100 is a plot of the lap shear strength for all tested samples with ANOVA statistical analysis. (n=5, p<0.05). Failed samples were noted as 0.Detailed Description

[0241] The following detailed description refers to the accompanying drawings that show, by way of illustration, specific details and embodiments in which the present disclosure may be practiced.

[0242] Features that are described in the context of an embodiment may correspondingly be applicable to the same or similar features in the other embodiments. Features that are described in the context of an embodiment may correspondingly be applicable to the other embodiments, even if not explicitly described in these other embodiments. Furthermore, additions and / or combinations and / or alternatives as described for a feature in the context of an embodiment may correspondingly be applicable to the same or similar feature in the other embodiments.

[0243] The present disclosure relates to an adhesive composition. Advantageously, the adhesive composition can be used as a stable bio-adhesive that not only has shelf stability, but also allows self-curing when exposed to oxygen atmosphere to induce cross-linking into a viscoelastic solid matrix. The adhesive composition is advantageously non-toxic, responsive to external or internal stimulus configurable with or without delay, able to rapidly crosslink (in 60 seconds or less) with or without a tunable time lag, thus allowing for adjustments during application.

[0244] The adhesive composition can be activated, for example, using oxygen as a stimulus, hence termed herein “oxygen activated adhesive composition”. The adhesive composition can be synthesized using a one-pot reaction scheme that is easily scaled through batch reactors or continuous flow chemistry protocols. The synthesis creates a Schiff base that can covalently link the two primary key intermediates: (1) a branched macromolecule containing end-chain amines and (2) a crosslinking group consisting of catechol aldehyde that may have been electronically substituted.

[0245] The adhesives formed from the adhesive composition, and the adhesive composition itself, are stable in anaerobic organic solvents (such as ethanol / methanol)and reacts upon exposure to gases or substrates containing oxygen gas or dissolved oxygen. For example, the oxygen activated adhesives can be stored in anaerobic, anhydrous conditions in either liquid or solid form, activated by diluting it in phosphate buffered saline solution with concentration of 10-50 % (w / w mg / mg). In such state, the adhesive can be applied onto a substate for binding purposes within a tunable lag-time.

[0246] In another example, the neat oxygen activated adhesive can be diluted using known plasticizers (e.g., anhydrous ethanol at 1-99 % solid wt%) and applied on a wet substrate, allowing for dissolved oxygen with the residual moisture to activate the adhesive for bonding or coating purposes.

[0247] The examples section further below serve to demonstrate (but not to limit) the synthesis, manufacturing, and utility of oxygen activated adhesives with several examples of based on constituents of (1) electronically substituted catechol aldehyde, (2) paired with a polyethyleneimine dendrimer macromolecule, whereby the two key intermediates are (3) covalently grafted via a Schiff base functional group. The combination of the three components yields a benign “prepolymer” in anaerobic environments (see FIG. 1), but switches to initiating and propagation of condensation polymerization within the presence of gaseous or dissolved oxygen. The nature of the networking condensation polymerization gives it utility as an adhesive or coating reagent that is relatively easy to apply and as a platform of one-component adhesive or incorporation within dual component adhesives found in manufacturing.

[0248] Details of various embodiments of the adhesive composition, its method of making and uses, and advantages associated with the various embodiments are now described below and / or with reference to the drawings. Where advantages of the embodiments and features are already demonstrated in one or more examples below and / or in the drawings, they shall not be reiterated for brevity.

[0249] In the present disclosure, there is provided for an adhesive composition. In certain examples, the adhesive composition may be referred to as a “prepolymer”, as the adhesive composition may be a polymer that has not undergone crosslinking yet before being subject to a stimulus. The adhesive composition can be used as an adhesive on various surfaces, including wet substrates. The term “adhesive composition” herein refers to the composition or formulation that is applied to the substrates for adhering the substrates. The term “adhesive” herein is distinguished from “adhesivecomposition”, wherein the adhesive is the resultant cured form of the present adhesive composition and the adhesive has the cross-linkages formed. Advantageously, the adhesive composition is usable as a “one-component” adhesive composition, meaning to say no oxidants or metals need to be present in the adhesive composition which traditional adhesive compositions may need. Also, the adhesive composition can be oxygen-activated to render crosslinking without added oxidants or metal ions to render the adhesive.

[0250] In various embodiments, the adhesive composition may comprise an organic macromolecule comprising nucleophilic functional groups, wherein each of the nucleophilic functional groups may comprise an amino group, a thiol group, a hydroxyl group, or a carboxyl group. For brevity, the organic macromolecule may be referred to as “macromolecule” or “macromer” in the present disclosure. The term “macromolecules”, alternatively termed herein as “unbranched or branched polymers” or “macromers”, refer to any large organic molecules composed of multiple small structural units linked together such that pendent functional groups (e.g., electrochemical donors, acceptors, and nucleophiles) can be confined to intermolecular reactions and constrained from intramolecular reactions. The one or more macromolecules can be a natural macromolecule (not artificially created). The one or more macromolecules can be a synthetic macromolecule, meaning that it is artificially created, such as via chemical synthesis.

[0251] In various embodiments, the adhesive composition may comprise one or more moieties each comprising a ring structure having (i) one carbon attached with an aldehyde or a ketone, (ii) two carbons each having a hydroxyl group, and (iii) one carbon attached with a substituent group.

[0252] In various embodiments, each of the nucleophilic functional groups grafted with one of the one or more moieties may have the aldehyde or the ketone converted into a coupling moiety and each of the nucleophilic functional groups not grafted with the one or more moieties may remain as the nucleophilic functional group.

[0253] In various embodiments, each of the one or more moieties may oxidize into a diketone when the adhesive composition is subjected to a stimulus, and wherein the diketone forms one or more crosslinkages with the nucleophilic functional groups.

[0254] In various embodiments, the organic macromolecule may comprise a polymer or a dendrimer. The polymer may be branched or linear. In the context of the present disclosure, a dendrimer is not identical to a branched polymer. The dendrimer comprises a core from which branches extend radially outwards to render a circular or spherical structure and the dendrimer may be symmetrical in nature in terms of its branching. In contrast, a branched polymer may have a linear core from which branches may extend (e.g., substantially or completely orthogonal to the linear core, but not radially outwards). In various embodiments, the polymer may be a random or statistical polymer.

[0255] In various embodiments, the organic macromolecule may comprise polyethylenimine, poly amine, poly amidoamine, poly lysine, polyester, polycaprolactone, chitosan, or a mixture thereof.

[0256] In various embodiments, each of the one or more moieties may comprise a diol with two non-adjacent carbons each having one hydroxyl group, a catechol, a triol, or a derivative thereof. In various embodiments, the triol may have a substituent group at a position not occupied by a hydroxy group and can be at a meta or an ortho or a para position of the ring structure relative to the aldehye or the ketone. In various embodiments, the triol may have two or three adjacent carbons each having a hydroxy group.

[0257] In various embodiments, each of the one or more moieties may be derived from a compound represented by a chemical formula of (la) or (lb) or (Ic) or (Id):

[0258] wherein R1may be an aldehyde or a ketone; and wherein R2represents the substituent group at a position not occupied by a hydroxy group and at a meta or an ortho or a para position of the ring structure relative to R1, wherein the substituent group may comprise an electron donating group or an electron withdrawing group.

[0259] In various embodiments, the substituent group may comprise an alkoxy group, a halogen, a nitro group, an oxido group, a hydroxy group, an acyloxy group, an acylamido group, a (di)alkylphosphino group, an alkylthio group, a sulfhydryl group, or an alkyl halide.

[0260] Advantageously, the electron donating and electron withdrawing substituents help in configuring or tuning oxidation potential, which in turn allows for an adjustable gelation time, modulus and toughness. In other words, the present adhesive composition may be fast curing or slower in curing, hence versatile and able to cater to various application needs.

[0261] In various embodiments, the diketone may comprise ortho-quinone or paraquinone. In various embodiments, the coupling moiety comprises a Schiff base. Advantageously, a Schiff base formation is stable in absence of O2 and aids in enabling long shelf life and transport.

[0262] In various embodiments, the stimulus may comprise an aqueous medium, an organic medium, or a mixture thereof, containing moisture, oxygen, an oxidant, an electrical voltage, or a combination thereof.

[0263] In various embodiments, 10 molar% to 50 molar%, 20 molar% to 50 molar%, 30 molar% to 50 molar %, 40 molar % to 50 molar %, 10 molar % to 40 molar %, 10 molar % to 30 molar%, 10 molar % to 20 molar%, 20 molar% to 40 molar %, 20 molar% to 30 molar%, etc., of the nucleophilic functional groups are grafted, each with one of the one or more moieties, and 50 molar% to 90 molar%, 50 molar% to 80 molar%, 50 molar % to 70 molar%, 50 molar % to 60 molar%, 60 molar% to 90 molar %, 70 molar%to 90 molar%, 80 molar% to 90 molar%, 60 molar% to 80 molar%, 60 molar% to 70 molar%, etc. of the nucleophilic functional groups remain as the nucleophilic functional group.

[0264] In various embodiments, the adhesive composition may further comprise an additive comprising 1,2-aliphatic diol, catechol, tannic acid, 1,2-furan-diol, ascorbic acid, vicinal diol, polysaccharide-diol, or a mixture thereof.

[0265] In various embodiments, the adhesive composition can be for use as an adhesive to adhere biological tissues; or as a sealant in medical and / or veterinary applications comprising intestinal anastomosis, vascular anastomosis, tissue repair, and / or tissue implantation; or as a general-purpose adhesive or as a resin material; or as an anti-microbial coating or an anti-microbial fabric; or as a cosmetic coating or a dermatological coating.

[0266] The present disclosure also provides for a method for forming the adhesive composition of various embodiments of the first aspect. Embodiments and advantages described for the present adhesive composition of the first aspect can be analogously valid for the present method subsequently described herein, and vice versa. As the various embodiments and advantages have already been described above and in the examples demonstrated herein further below, they shall not be iterated for brevity.

[0267] In various embodiments, the method may comprise providing the organic macromolecule, and mixing the organic macromolecule with a precursor of the one or more moieties.

[0268] In various embodiments, providing the organic macromolecule may comprise providing a stock solution comprising the organic macromolecule in a first anhydrous organic solvent, and removing dissolved oxygen from the stock solution.

[0269] In various embodiments, mixing the organic macromolecule with the precursor may be carried out in an inert environment (e.g., argon, nitrogen).

[0270] In various embodiments, mixing the organic macromolecule with the precursor may comprise adding the precursor to the stock solution having oxygen removed therefrom, or dissolving the precursor in a second anhydrous organic solvent to obtain a precursor solution, and mixing the precursor solution and the stock solution having oxygen removed therefrom.

[0271] In various embodiments, the precursor may comprisee a dihydroxybenzaldehyde having the substituent group. In various embodiments, the precursor may comprise 1,3 -dihydroxybenzaldehyde, 2,3-dihydroxybenzaldehyde, 3,4-dihydroxybenzaldehyde, 4,5-dihydroxybenzaldehyde, or a derivative thereof, having the substituent group. The precursor may also be a triol with the substituent group. The triol has been described in one or more embodiments mentioned above.

[0272] In various embodiments, the precursor may comprise a catechol variant represented by the chemical formula of (la) or (lb) or (Ic) or (Id):

[0273] wherein R1is an aldehyde or a ketone, and wherein R2represents the substituent group at a position not occupied by a hydroxy group and at the meta or the ortho or the para position of the ring structure relative to R1, wherein the substituent group may comprise the electron donating group or the electron withdrawing group.

[0274] In various embodiments, the substituent group may comprise an alkoxy group, a halogen, a nitro group, an oxido group, a hydroxy group, an acyloxy group, an acylamido group, a (di)alkylphosphino group, an alkylthio group, a sulfhydryl group, or an alkyl halide.

[0275] The present disclosure also provides for a method for adhering or sealing biological tissues or surfaces. Embodiments and advantages described for the present adhesive composition of the first aspect and the method described above can be analogously valid for the present method subsequently described herein, and vice versa.As the various embodiments and advantages have already been described above and in the examples demonstrated herein further below, they shall not be iterated for brevity.

[0276] In various embodiments, the method may comprise applying the adhesive composition of various embodiments of the first aspect to one or more biological tissues or surfaces, subjecting the adhesive composition to the stimulus, and contacting the biological tissues or surfaces together to adhere or seal the biological tissues or surfaces, respectively.

[0277] In various embodiments, subjecting the adhesive composition to the stimulus may comprise exposing the adhesive composition the aqueous solution containing oxygen, the oxidant, applying the electrical voltage to the adhesive composition, or a combination thereof.

[0278] The present disclosure also provides for a kit. Embodiments and advantages described for the present adhesive composition of the first aspect and the methods described above can be analogously valid for the kit subsequently described herein, and vice versa. As the various embodiments and advantages have already been described above and in the examples demonstrated herein further below, they shall not be iterated for brevity.

[0279] In various embodiments, the kit may comprise the organic macromolecule defined in various embodiments of the first aspect, and the precursor of the one or more moieties defined in various embodiments of one or more aspects mentioned above, wherein the kit contains the organic macromolecule in a separate compartment from the precursor.

[0280] In the present disclosure, the term “alkoxy” in such embodiments refer to an alkyl group singularly bonded to oxygen. Examples include methoxy, ethoxy, n-propoxy, and isopropoxy. The term “dialkyl” in certain embodiments refer to the presence of two alkyl groups.

[0281] In the present disclosure, the terms “redox donor”, “electrochemical donor”, “electron donor”, “redox donor internal additive” and “electron donating moiety”, are used interchangeably. The term “redox acceptor”, “electrochemical acceptor”, “electron acceptor”, “redox acceptor internal additive”, and “electron accepting moiety”, are used interchangeably.

[0282] In the present disclosure, the term “room temperature” may be a range from 20°C to 30°C.

[0283] The word “substantially” does not exclude “completely” e.g. a composition which is “substantially free” from Y may be completely free from Y. Where necessary, the word “substantially” may be omitted from the definition of the present disclosure.

[0284] In the context of various embodiments, the articles “a”, “an” and “the” as used with regard to a feature or element include a reference to one or more of the features or elements.

[0285] In the context of various embodiments, the tilde symbolthe term “about”, and the term “approximately”, as applied to a numeric value encompasses the exact value and a reasonable variance. The variance may be ±20%, ±10%, ±5%, ±1%, ±0.5%, ±0.1%, etc.

[0286] As used herein, the term “and / or” includes any and all combinations of one or more of the associated listed items.

[0287] As used herein, the term “hour” and “hours” may be denoted interchangeably using “h”, “hr”, and “hrs”.

[0288] Unless specified otherwise, the terms "comprising" and "comprise", and grammatical variants thereof, are intended to represent "open" or "inclusive" language such that they include recited elements but also permit inclusion of additional, unrecited elements.Examples

[0289] The present disclosure relates to an adhesive composition. The adhesive composition of the present disclosure may be an oxygen activated adhesive composition for bonding and coating substrates.

[0290] The adhesive composition can include an unbranched or branched polymer, substituted catechol internal additives, Schiff base graftings, and nucleophilic functional groups, wherein the composition can spontaneously form a cross-linking groups that covalently bonds nucleophilic groups after exposure to oxygen. The composition is inert and inactive in a water-free, anhydrous state, but upon exposure to oxygen via atmosphere, oxygen gas, and / or reconstitution in a dissolved oxygen solvent, cross-linking can be activated within 60 seconds with no energy inputs. Fabricationmethod, working principle, and uses of the adhesive composition, are described in further details, by way of non-limiting examples, as set forth below.

[0291] Example 1A: Introductory / General Discussion of Adhesive Composition

[0292] Various examples herein, including the other examples below, relate to an adhesive composition, such as an oxygen activated adhesive composition, and a method of preparing such adhesive compositions.

[0293] Various examples that demonstrate for an adhesive composition, which can be an oxygen activated adhesive composition, is provided. The adhesive composition can include (i) a macromolecule (e.g. a branched or unbranched polymer), wherein the macromolecule can have 10 or more primary amines per molecule; (ii) can be grafted with electronically modified dihydroxybenzaldehyde which are precursors to crosslinking groups (such as substituted catechols), (iii) can have substituted catechol-aldehyde grafted onto the macromolecule amine, so as to form a grafted substituted catechol- Schiff base, (iv) and that upon exposure to oxygen, turns into a cross-linking functional group, where the cross-linking precursors can include substituted 3,4-aryl-diols or 4,5-aryl-diols (e.g. substituted catechol).

[0294] Optionally, the adhesive composition can include additives including electronically substituted catechol aldehydes, trihydroxy-benzaldehydes, 5-Methoxy-3,4-dihydroxybenzaldehyde, 2-Methoxy-4,5-dihydroxybenzaldehyde, 5-Nitro-3,4-dihydroxybenzaldehyde, 3-Bromo-4,5-dihydroxybenzaldehyde.

[0295] In various examples, the macromolecule can be further grafted with substituted catechols and / or moieties (e.g., at least 10 substituted catechols and / or moieties), that allow oxidation upon exposure to oxygen, wherein the substituted catechols and / or moieties preferably comprise unsaturated functional groups, and more preferably Schiff bases, imines, oxime, ferrocenes, and / or sacrificial anodes.

[0296] In various examples, the macromolecule can include nucleophilic groups (e.g., amino groups on the branched or unbranched polymer) that can spontaneously react with the cross-linking functional groups (e.g. quinone or 1,2-cyclo-diketone), and in certain non-limiting embodiments, the substituted catechol precursors and / or moieties may be grafted on different macromolecules.

[0297] Advantageously, the adhesive composition can have shelf- stability within anaerobic conditions, but upon oxygen exposure or reconstitution in oxygen dissolved solvents, spontaneously cross-links within 60 seconds after exposure to oxygen.

[0298] In various examples, a method of preparing an adhesive using the adhesive composition is demonstrated. The method can include (a) providing the adhesive composition, and (b) adding a rehydration mixture containing dissolved oxygen (greater than 30-90% water w / v) to the adhesive composition so as to activate the cross-linking groups within 60 seconds. The method can include providing atmosphere or oxygen gases to cross-link the cross-linking groups in less than 60 seconds.

[0299] In various examples, an adhesive prepared by the method is described.

[0300] In various examples, a kit for adhering hydrated substrates is described. The kit can include (a) the adhesive composition, (b) a rehydration mixture containing dissolved oxygen (e.g., an isotonic aqueous reconstitution solution).

[0301] In various examples, a method of adhering biological tissues is described. The method can include (a) applying the adhesive composition to a first dry or rehydrated substrate, (b) contacting a second dry or hydrated substrate with the resultant coating (i.e., the applied adhesive composition), and (c) applying pressure to one or both substrates to adhere the first substrate to the second substrate for a time period exceeding the gelation time or more than 60 seconds after application.

[0302] In various examples, there is disclosed a use of the adhesive composition or the adhesive prepared by the method, as a wet substrate adhesive and sealant for underwater and medical and veterinary applications, supplementing or replacing sutures or other mechanical fixation methods.

[0303] Various examples refer to the adhesive composition, which can be an oxygen activated adhesive composition. The oxygen activated adhesive composition disclosed herein can be constructed using an oxygen activation based on incorporation of substituted catechols that can be activated after exposure to oxygen gas or aqueous reconstitution within a solution containing dissolved atmospheric oxygen. A branched polymer comprising one or more macromolecules grafted with at least ten substituted catechol moiety, at least 10 nucleophilic functional groups rapidly cross-links after oxygen exposure or reconstitution in a oxygen containing solvent within 60 secondsafter oxygen exposure stimulus. It has the advantage in that it requires no mixing of chemical components and has long term shelf stability under non-aerobic conditions.

[0304] Substituted catechol moiety consist of dihydroxybenzaldehyde that have been substituted with known (preferably) electron donating group or electron releasing group whereby such atoms or functional groups donate their electron density into conjugated pi systems. Examples include oxido, alkoxy, hydroxy, acyloxy, acylamido groups, (di)alkylphosphino, alkylthio, bromo, chloro, sulfhydryl groups, and alkyl halides.

[0305] Advantageously, preparation of the oxygen activated adhesive composition disclosed herein is inexpensive, as raw materials can be easily synthesized and scaled for manufacturing. Further, catechol-grafted branched polymers may be achieved in a one-step reaction that simultaneously creates the substituted catechols grafted with Schiff bases without requiring special conditions such as elevated temperature, high vacuum, and / or high pressure, or special equipment. Ease of use of the oxygen activated self adhesive composition renders its feasibility for underwater, wet substrate, veterinary, and clinical applications. The adhesive composition disclosed herein also exhibits high adhesive force on wet substrates as demonstrated on hydrated substrates.

[0306] With the above in mind, the term “adhesive composition” as used herein can refer to a formulation that may be used to join or to bond wet substrates, biological tissues, hydrated substrates, or combinations thereof together. The term “wet substrate” refers to a matrix that contains more than 1% water, has a surface of immobilized water layers, or is submerged in an aqueous solvent. Examples of wet substrates include, but are not limited to, submerged metal boat hulls, soft tissues, bones, and hydrated biomaterials.

[0307] The adhesive composition, which can be oxygen activated, can comprise a macromer comprising one or more macromolecules grafted with at least ten substituted catechol moiety (quinone precursor), at least ten nucleophilic functional groups (e.g. amines, thiols, carboxylic acids). The molar equivalents of substituted catechols to macromolecules should have a ratio between 2 to 100 and one non-limiting example is 40.

[0308] With the above in mind, the term “substituted catechol” as used herein refers to a functional group that upon an exposure to oxygen, donates 1 or more electrons (referred to as anodic oxidation) to the surrounding matrix. The substituted catecholmay then be converted into a functional group that allows spontaneous covalent bonding of nucleophiles. Examples of substituted catechol includes, but is not limited to trihydroxy -benzaldehydes, 5-methoxy-3,4-dihydroxybenzaldehyde, 2-methoxy-4,5-dihydroxybenzaldehyde, 5-nitro-3,4-dihydroxybenzaldehyde, 3-bromo-4,5-dihydroxybenzaldehyde.

[0309] With the above in mind, the term “substituted catechol” as used herein may exist as a protected precursor to prevent activation under ambient conditions (e.g anaerobic solvents) for improvement of shelf stability.

[0310] The adhesive composition of the present disclosure is inter alia (1) safe and non-toxic, (2) rapidly cross-links upon air exposure, (3) self-cures within 60 seconds of application, (4) allows for liquid manipulation before adhesive application.

[0311] In various examples disclosed herein, the adhesive composition / adhesive is developed by constituting (1) a macromer comprising one or more macromolecules grafted with (2) at least ten substituted catechol moieties with (3) at least ten nucleophilic functional groups (e.g., amines, thiols, carboxylic acids).

[0312] The adhesive composition can be rapidly cross-linked with exposure to atmosphere or 100% oxygen gas, with or without aqueous reconstitution. The oxygen activated adhesive may also covalently cross-link after aqueous reconstitution with dissolved oxygen present.

[0313] In one or more examples, the adhesive composition may be stored under anaerobic conditions in a liquid or solid form. For example, the adhesive composition in the form of a solid mixture can be activated by dissolving in saline or phosphate buffered saline solution: the concentration is 10%-70% (w / v, g / mL). The activated mixture solution of the adhesive composition may be smeared on the substrate and then another substrate may be adhered after gentle pressure beyond the gelation time.

[0314] In certain examples, the adhesive composition may be stored under anhydrous, anaerobic conditions in a liquid or solid form. For example, the adhesive composition in the form of a solid mixture can be applied directly on a wet substrate, where the wet substrate donates oxygen through dissolved oxygen chemical gradient to the oxygen activated adhesive composition. Then another substrate may be adhered after gentle pressure beyond the gelation time.

[0315] By developing such adhesive composition, problems mentioned above may be addressed and the adhesive composition is able to meet all the requirements for adhesion to wet surfaces.

[0316] Advantageously, the macromer as the backbone may be a natural or synthetic bio / macromolecule which does not contain any toxic components, in particular aldehyde, acrylate, urethane, or cyanoacrylate groups. The oxygen activated adhesive composition can be fabricated using a substituted catechol, where it integrates oxygen activation and Schiff-base grafting for long term effectiveness by covalent crosslinking.

[0317] In various examples, the adhesive composition can be configured as kit. The kit can be used for adhering wet substrates according to various examples that may include the dried adhesive composition and a reconstitution solution (e.g., saline). Such kit may be considerably easy to use. For example, a user may first add the reconstitution solution to the dry oxygen activated adhesive composition and the mixture may be applied on a wet substrate. The liquid oxygen activated adhesive composition can be manipulated easily during the lag-time. Manipulation can be by means of an aerosol spray, brush-based coatings, syringe-delivery, knife casting, or combination thereof.

[0318] Various examples of the kit may include the adhesive composition dissolved in an anhydrous solvent within an anaerobic container. Such kit may be very easy to use. For example, a user may pressure the container and release the mixture on a water permeated substrate exposed to an aerobic atmosphere. Adhesion of a second substrate to the first substrate may be carried out by contacting the first substrate with the second substrate where the adhesive composition is applied and exerting gentle pressure on one or both the first or second substrates.

[0319] Example IB: Introductory / General Synthesis Example of the Adhesive Composition (Oxygen Activated Adhesive)

[0320] A macromer was synthesized by the spontaneous coupling of 5-methoxy-3,4-dihydroxybenzaldehyde with amine-functionalized dendrimers. Briefly, polyethylenimine (PEI) was degassed in a methanol stock solution to remove dissolved oxygen. 5-methoxy-3,4-dihydroxybenzaldehyde (5-OMe-3,4-DBA) was dissolved in anaerobic, anhydrous methanol and subsequently added dropwise to rapidly stirring PEI stock solution to obtain 10-50% grafting (moles 5-OMe-3,4-DBA / moles amine per dendrimer x 100). Reaction was stirred at room temperature (20°C to 30°C) under darkcondition in an inert gas atmosphere overnight. An adhesive composition (oxygen activated adhesive composition) was obtained by precipitating in anhydrous diethyl ether, followed by washing the precipitations with diethyl ether for two times then drying under vacuum. The adhesive composition was stored under anaerobic, anhydrous conditions at -20°C. The resultant oxygen activated adhesive was characterized by!H NMR and size exclusion chromatography. Content of catechol groups and Schiff base group was determined by!H NMR.

[0321] Example 1C: An Introductory / General Example of the Adhesive Composition based on Example IB.

[0322] The steps carried out for the adhesive composition of this example was adopted from example IB above. Oxygen activated adhesive prepolymer Schiff-base was synthesized by spontaneous coupling of dihydroxy benzaldehyde with amine-functionalized dendrimers (FIG. 1). Briefly, the methanolic solution of polyethylenimine was degassed with argon to remove dissolved oxygen. Catechol aldehyde was added in portion over a period of 5 mins while stirring vigorously to obtain 10-50% grafting (moles catechol aldehyde / moles amine per dendrimer x 100). The reaction mixture was stirred for 30 to 60 mins at room temperature. It was concentrated in a rotary evaporator and dried under vacuum for 4 to 5 hrs to afford the oxygen activated adhesive prepolymer Schiff-base which was stored under anaerobic, anhydrous conditions at -20°C. The adhesive prepolymer Schiff-base was characterized by!H NMR. From this one step reaction, it can be seen that the reaction is easily scalable through batch reactors or through flow chemistry setups.

[0323] Example ID: An Introductory / General Example on the Adhesive Gelation and Curing initiated by Oxygen Atmosphere

[0324] The viscoelastic properties of the adhesive composition were recorded on a rheometer (MCR102, Anton Paar, Singapore). The rheology tests were performed with a 10-mm diameter steel rheometer probe having parallel-plate geometry (PP10) at the measuring gap of 0.3 mm. 20 pL sample was casted on the rheometer base plate. Steady state rotational viscosity tests were performed at fixed shear rate of 62.8 rad.s1(10 s1@ %R). Storage and loss modulus (G’ and G”) had a data acquisition rate of 1 Hz using a 10 Hz angular frequency and 10% strain amplitude. Amplitude sweeps of cured adhesives were determined under the strain range of 1 to 1000%.

[0325] The adhesive composition was prepared in phosphate buffered saline (or other aqueous and / or aqueous / solvent liquid) at ambient conditions and 20 p L of the adhesive formulation PEI-(5-OMe-3,4-DBA) was loaded onto the rheometer base plate. Upon exposure to the atmosphere, oxygen mediated cross-linking initiated curing and crosslinking of the sample, where gelation occured in less than 30 seconds and continues to cure over a 30 minute time range. Gelation (G’ > G”) and ongoing curing was seen in the real time rheology evaluation, as shown in FIG. 2A. This demonstrates the three factors for the oxygen-activated curing: (1) catechol aldehyde with a m-methoxy substitution, (2) Schiff-base grafting, (3) presence of oxygen to induce spontaneous reaction of the catechol to quinone formation, where the latter is known to impart crosslinking and adhesion. A control sample that incorporateed only two of the three features (Schiff base grafting and oxygen atmosphere) with the m-methoxy substituted catechol aldehyde removed and replaced with an benzaldehyde displayed no crosslinking or curing behavior as shown in FIG. 3.

[0326] Example IE: An Introductory / General Example on Oxygen Atmosphere and Schiff Base Grafting for Oxygen Mediated Crosslinking

[0327] Oxygen induced activation involves the utilization of the Schiff base grafting to link the crosslinker (e.g., 5-OMe-3,4-DBA) to the macromolecule. For example, a traditional method in the art of synthesizing adhesives grafts crosslinkers to macromolecules via ester bonds. However, such a traditional grafting procedure may render the present adhesive composition inoperable. Two model reactions demonstrate such feature by grafting a macromolecule substitute, propyl amine (PA), with Model Reaction 1 (MR-1) via a catechol ester defined as methyl-3,4-dihydroxy-5-methoxybenzoate (MDMB), Model Reaction 2 (MR-2) compares the present feature of Schiff-base grafting with PA and catechol aldehyde, defined as (5-OMe-3,4-DBA) (MR-2) (FIG. 9). O2 was bubbled into both the reaction mixture (MR-1 and MR-2) for 90 min. ’H-NMR investigation exhibited MR-1 remained unreacted in presence of O2, as no new peaks were observed that should be present if crosslinking occured (see FIG.4A). The1H NMR of MR-2 exhibited many new peaks when the reaction carried out in presence of O2, suggesting formation of crosslinking groups and subsequent quinone related products known to occur by oxidation of catechol groups (see FIG. 4B). The reaction mixture of MR-2 exhibited no new peaks when the reaction carried out underan inert argon atmosphere. This experiment demonstrates the three factors for the oxygen-activated curing: (1) catechol aldehyde with a m-methoxy substitution, (2) Schiff-base grafting of the substituted catechol and macromolecule, and (3) presence of oxygen to induce spontaneous reaction of the catechol to quinone formation.

[0328] Example IF : An Introductory / General Example on the Reaction Kinetics Modulated Through Electronic Substitution of the Catechol Aldehyde

[0329] Adhesive curing kinetics, often referred to as adhesive pot-life, may be one of the parameters to look at in manufacturing. Adhesives and adhesive compositions that have a tunable pot-life and stable shelf- stability are advantageous for many industrial and manufacturing entities.

[0330] The features of the adhesive composition herein incorporating electron donating (ED) substituted catechol (OMe) on PEI dendrimers show 2 months stability in methanolic solution at 4 °C, while Schiff-bases prepared from electron withdrawing (EW) substituted catechol (NO2) possess poor shelf stability at either room temperature as well as at 4°C due to their increased kinetics and oxygen sensitivity. The rate of decay of catechol functionality in various catechol grafted Schiff-bases were evaluated by UV-vis absorption kinetics. UV-vis spectra were recorded at every 15 min interval for a period of 3 hrs. O2 was bubbled into MeOH solution of Schiff-base (3.4 x 10-6M) for a period of 5 seconds before recording the UV-vis spectrum. Before O2 bubbling, the Schiff-base containing EW (electron withdrawing) bromo catechol functionality displayed an intense UV-vis absorption peak at 334.5 nm and a weak UV-vis absorption peak at - 420 nm (FIG. 5A), which were attributed to catechol and quinone respectively. There was no significant decay of catechol absorption peak (334.5 nm) over a 3 h period of UV-vis study (FIG. 5B). A slight reduction of the quinone peak intensity indicated relatively slow crosslinking reaction.

[0331] The Schiff-base containing ED (electron donating) methoxy catechol functionality (OMe) displayed strong catechol and weak quinone absorption peaks at 342.5 nm and at 440 nm respectively in the UV-vis spectra before O2 bubbling into its solution (FIG. 5C). The O2 bubbling over 3 h resulted in significant decay of both catechol (1storder decay) and quinone absorption peaks, alongside a formation of new broad peak at 480-600 nm (FIG. 5C and FIG. 5D). Occurrence of these spectral changes are due to the crosslinking of quinone intermediate with other catechol to form aryl-aryl coupling product and / or with amine by various Michael type reactions. These results demonstrate that curing kinetics are tunable through EW substituent (e.g., -Br) that retard curing and and ED substituent (OMe) that accelerates curing via catechol to quinone crosslinking reaction when combined with Schiff base grafting.

[0332] Rheological measurements further reveal the impact of ED (OMe) and EW (NO2) catechol substituents on the rate of crosslinking and ensuing mechanical properties of the adhesive. 20 p L Schiff-base formulation was loaded on the rheometer base plate to perform rheology investigations at ambient condition. Changes in storage modulus (G’) and loss modulus (G”) of the adhesives were recorded for 30 mins using 10 Hz angular frequency and 10% amplitude strain (FIG. 6). The observed storage moduli of the adhesives at the end of 30 min are PEI-(5-NO2-3,4-DBA)20 < 1.0 Pa, PEI-(6-N02-3,4-DBA)2O < 1.0 Pa, PEI-(5-OMe-3,4-DBA)2o 5458.6 ± 290.6 Pa, and PEI-(2-OMe-4,5-DBA)2o 6394.1 ± 222.3 Pa respectively. Thus, Schiff-base adhesive compositions with ED substituents (5-OMe and 2-OMe) exhibit much higher G’ than with strong EW NO2 substituent. This demonstrates the kinetics and material properties of the oxygen mediated adhesives are tunable through ED catechol substituents that exhibit greater adhesive curing and crosslinking (e.g. 5-OMe) compared to EW catechol substituents (5-NO2).

[0333] Example 1G: An Introductory / General Example of Utilization of the Adhesive Composition on Bonding Wet Substrates

[0334] Adhesive compositions to bond wet surfaces that cannot be dried due to their environment, material makeup (e.g. hydrogels, animal or human tissues), or releated factors, may be industrially relevant for cosmetic, medical, and husbandry industries. Adhesive compositions of the present disclosure, which are oxygen activated catechol derived Schiff-base adhesive compositions, demonstrate bonding of wet collagen substrates via industry standard lap-shear and 1800peel adhesion assays. The lap shear adhesion strengths were calculated by dividing the maximum load at failure (N) by the bonding area (m2). The adhesive formulation (in ethanol) was applied on the 2 cm x 2 cm area of a collagen substrate which was glued with another collagen substrate. The glued collagen-collagen structure was allowed to cure for 30 mins by sandwiching between two glass slides with paper clips. The average lap shear adhesion strength calculated at the yielding point of force-displacement graphs are PEI-(5-NO2-3,4-DBA)2O 14.6 ± 1.3 kPa, PEI-(6-N02-3,4-DBA)2o 10.7 ± 2.3 kPa, PEI-(3-Br-4,5-DBA)2019.9 ± 0.8 kPa, PEI-(2-Br-4,5-DBA)2018.7 ± 1.2 kPa, PEI-(5-OMe-3,4-DBA)2026.2 ± 5.7 kPa, and PEI-(2-OMe-4,5-DBA)2011.1 ± 0.9 kPa respectively (FIG. 30A). The PEI-(5-OMe-3,4-DBA)2o and PEI-(3-Br-4,5-DBA)2o Schiff-base adhesives displayed significantly higher lap shear adhesion strength than rest of the Schiff-base adhesives as compared to control PEI-(3,4-DBA)2o

[0335] 180° peel adhesion were performed to evaluate the interfacial toughness of oxygen activated Schiff-base adhesives. The peel adhesion strengths were calculated by dividing the maximum load at failure (N) by width of the collagen strips (m). The adhesive formulation and curing procedure used for the peel adhesion tests was similar to that of lap-shear adhesion measurements. The average value of 1800peel adhesion strength calculated were PEI-(5-N02-3,4-DBA)2o 29.2 ± 4.8 J.nr2, PEI-(6-NO2-3,4-DBA)2022.1 ± 1.8 J.nr2, PEI-(3-Br-4,5-DBA)2023.3 ± 3.5 J.nr2, PEI-(2-Br-4,5-DBA)2022.9 ± 3.5 J.nr2, PEI-(5-OMe-3,4-DBA)2028.7 ± 3.0 J.nr2. and PEI-(2-OMe-4,5-DBA)2O 19.2 ± 1.7 J.nr2respectively (FIG. 7).

[0336] Example 1H: Summary Discussion based on Examples 1A to 1G

[0337] In summary, adhesive compositions of the present disclosure can include organic macromolecules each comprising nucleophilic functional groups, wherein each of the organic macromolecules can be a linear macromolecule or a branched macromolecule, wherein each of the nucleophilic functional groups can comprise an amino group, a thiol group, a hydroxyl group, or a carboxyl group, and (ii) electronically substituted catechol aldehyde or ketone, (iii) a schiff-base functional group that covalently links (i) and (ii).

[0338] The adhesive composition of the present disclosure can be a one-component (1C) adhesive composition as described above, wherein the formulation can be shelfstable within anaerobic environments. The one-component (1C) adhesive composition can be activated via a stimulus, wherein the stimulus can comprise gaseous oxygen, solvents containing dissolved oxygen, or combination thereof. The curing kinetics of the adhesive compositions can be modulated through electronic withdrawing and electronic donating groups via the electronically substituted catechol aldehyde. The adhesive compositions can bond two substrates when either the adhesive composition, or the substrates, or both, are exposed to gaseous oxygen, solvents containing dissolvedoxygen, or combination thereof. The adhesive composition can bond one substrate to from a 5 nm to 1 mm thick coating when either the adhesive composition, or the substrates, or both, are exposed to gaseous oxygen, solvents containing dissolved oxygen, or combination thereof.

[0339] Example 2A: Another Example of the Present Adhesive Composition -Oxygen Initiated Polymerization of Catechol-Schiff Base Dendrimers

[0340] Catechol oxidation process using greener means can circumvent the need for toxic additive (oxidant or chelator) that hinders catechol grafted dendrimers for use in clinical needs. Examples 2A to 2L show that adhesive compositions of the present disclosure, e.g., PEI-(5-OMe-3,4-DBA)2o, a catechol grafted dendrimer Schiff base, can be oxidised by air / Ch to initiate crosslinking reaction though the corresponding catechol ester remains stable. Air / Ch initiated crosslinking of PEI-(5-OMe-3,4-DBA)2o was confirmed by lower values of aromatic CH integration (XH-NMR) and by first order UV-vis absorption decay of catechol functional group. In addition, PEI-(5-OMe-3,4-DBA)2O formulation gelled in < 1 min, displayed steady increase in storage modulus, and exhibited 26.2 ± 5.7 kPa lap shear adhesion strength on collagen film substrates.

[0341] Example 2B: Discussion for Example 2A

[0342] The catechol functionality, 3,4-dihydroxyphenylalanine (DOPA), in Mussel adhesive proteins (MAPs) may be a key to adhere to both organic (biological tissue) and inorganic surfaces (rock, metal, sand).

[0343] The MAPs cohesion and adhesion to surfaces involves two mechanisms -polyphenol oxidase mediated oxidation of DOPA to quinone, and metal ion chelation of the catechol group of DOPA. The oxidized quinone intermediate crosslinks with nucleophiles (in MAPs) and with biological substrates endowing adhesion ability. However, most of such traditionally designed catechol adhesives require an external stimulus (oxidant - NaIO4, H2O2 etc., or chelator - Fe+3, Ca+2, etc.) for adhesion. Two component adhesives (2C) with chemical initiators offer an on-demand adhesion strategy, but traditionally may suffer from tissue incompatibility (toxic oxidizers) and requires complex applicators.

[0344] Adhesive compositions of the present disclosure offer a simpler method, redox-active catechol derivatives for voltage sensitive configurations were prepared. The Schiff base / catechol variants were found to spontaneously activate, presumablydue to exposure to oxygen, water, or both. It is hypothesized that the electronically modified catechols may be induced to spontaneously form quinones faster than control structures. Ideally, conversion rates could be accelerated to within an order of minutes compared to weeks / months. To explore this hypothesis, macromolecules were grafted with 5-Methoxy-3,4-dihydroxybenzaldehyde, yielding the Schiff base PEI-(5-OMe- 3.4-DBA)2O- Crosslinking kinetics and viscoelastic material properties of PEI-(5-OMe- 3.4-DBA)2O before and after exposure to oxygen were mapped. It is noted that traditional reports did not investigate air / Ch initiated kinetics of tailored catechols susceptible to an oxygen stimulus within minutes.

[0345] Spontaneous generation of a crosslinker (e.g., quinone) with oxygen stimulus can provide an easier method of exploiting catechol-based biomaterials. Cyclic voltammetry evaluates catechol structures with a metastable oxidation potential sufficiently low enough for oxygen activation. Shelf stability assessment utilizes1H-NMR to map dynamic conversion of catechol to quinone. Absorbance measurements estimate catechol kinetic after exposure to air and pure oxygen. Further analyses of viscoelastic and adhesive properties demonstrate the oxygen- sensitive resins crosslinking potential on tissue-like wet substrates.

[0346] Example 2C: Materials and Characterizations based on Examples 2A and 2B

[0347] Materials

[0348] Branched polyethylenimine (PEI, average Mw~ 25000), 5-methoxy-3,4-dihydroxybenzaldehyde (5-OMe-3,4-DBA), benzaldehyde (BA), Na2B4O?, methyl gallate, dimethyl sulfate, Sodium hydroxide (NaOH), methanol (MeOH), phosphate buffer (PBS), were purchased from Sigma-Aldrich, Singapore. Collagen film was purchased from Nippi Collagen Inc (Tokyo, Japan).

[0349] General methods

[0350] All experiments were carried out in dried glassware.!H NMR (400 MHz) and13C NMR (100 MHz) spectra were recorded on Bruker AVANCE 400 MHz instruments in the deuterated solvents (CD3OD or DMSO-de). Chemical shifts are given in parts per million (ppm, d) based on the middle peak of the solvent signal (XH NMR, d = 3.31 for CD3OD and d = 2.50 for DMSO-d6;13C NMR, d = 39.51 for DMSO-d6) as an internal standard. The J values are given in Hz. Multiplicities are indicated as ‘s’ (singlet), ‘d’(doublet), ‘t’ (triplet), ‘q’ (quartet), ‘m’ (multiplet) and ‘brd’ (broad doublet). All reactions were carried out under argon. Evaporation of solvents was performed at reduced pressure using a rotary evaporator. All reagents and solvents were used as purchased without further purification unless otherwise indicated.

[0351] Synthesis ofMethyl-3,4-dihydroxy-5-methoxybenzoate (MDMB)

[0352] To a stirred suspension of NaiEUCh (13.6 g, 67.8 mmol) in H2O (330 mL) was added methyl gallate (5.0 g, 27.1 mmol) at room temperature. After 1 hr, dimethyl sulfate (13.3 g, 105.9 mmol) and NaOH (16 .7 mL, 6.5 M aq. solution) were added dropwise simultaneously over 1 h. The reaction mixture was stirred overnight. Aqueous H2SO4 (tkCktkSCU 90:10) was added dropwise until pH became 4-5. The reaction mixture was extracted with EtOAc (200 mL). The organic phase was washed with H2O (2 x 100 mL), aq. saturated NaCl (50 mL), dried (Na2SO4) and concentrated. The residue was purified by column chromatography on silica gel (ethyl acetate / hexane 60:40) to afford Methyl-3,4-dihydroxy-5-methoxybenzoate as white solid (4.2g, 78%).1H NMR (400 MHz, DMSO-d6) 8 10.30-8.30 (brs, 2H), 7.09 (s, 1H), 7.03 (s, 1H), 3.77 (s, 3H), 3.76 (s, 3H);13C NMR (400 MHz, DMSO-d6) 8 166.4, 148.0, 145.4, 139.5, 119.4, 110.6, 104.7, 56.0, 51.9.

[0353] Synthesis of PEI-(5-OMe-3,4-DBA)2o, PEI-(MDMB)20, and PEI-(BA)20

[0354] PEI-(5-OMe-3,4-DBA)2o, PEI-(MDMB)2o, and PEI-(BA)2o were prepared following the same procedure. Briefly, to a methanolic solution of PEI (1.12 g, 0.044 mmol, in 30 mL MeOH), was added 5-OMe-3,4-DBA (293 mg, 1.74 mmol) over a period of 5 min under argon atmosphere. The reaction mixture was stirred 30 mins. It was concentrated in a rotary evaporator and dried in vacuum for 4-5 h to afford PEI-(5-OMe-3,4-DBA)2o in quantitative yield.

[0355] UV-visible spectroscopy study

[0356] The UV-visible spectroscopic study of PEL(5-OMe-3,4-DBA)2o (3.4 x 10-6M, MeOH) was performed on a Shimadzu UV-2700 spectrophotometer in a quartz cuvette. The reference cuvette contained MeOH.

[0357] Cyclic voltametric studies

[0358] Cyclic voltametric (CV) measurements were performed using a Metrohm Autolab PGSTAT302 N potentiostat in a three-electrode setup. A 3 mm diameter planar glassy carboExample 2C, “n disk (Metrohm) is used as a working electrode (WE) inconjunction with a platinum plate counter electrode (CE, Metrohm) and Ag / AgCl reference electrode (RE, filled with 3.0 M KC1 solution). The cyclic voltammetric data of PEI-(sCat)2o compounds are recorded in PBS solution (1.0 mM), at the scan rate 0.1 V / s. All voltametric experiments were conducted under argon atmosphere, at 24°C in a Faraday cage. Prior to each scan, the working electrode is cleaned by polishing with alumina oxide (grain size 0.3 mm) slurry on a Buehler Ultra-pad polishing cloth, rinsing with water and ethanol, and then drying with a lint-free tissue.

[0359] General Preparation of adhesive Compositions

[0360] The 30 wt% formulation of PEI-(5-OMe-3,4-DBA)20in PBS was prepared by adding 90 mg PEI-(5-OMe-3,4-DBA)2oin 210 pL PBS buffer (pH - 7.2) and vortexing the mixture until a clear homogeneous solution is observed. This formulation (20 L) was loaded onto the rheometer base plate for rheology measurements. For adhesion strength measurements study the 30 wt% formulation of PEI-(5-OMe-3,4-DBA)2o adhesive was prepared in a similar way using EtOH in place of PBS. For rheology and adhesion strength measurements of PEI-(MDMB)2o, PEI-(BA)2o, and PEI-(3,4,5-TBA)2O adhesive formulations were prepared similar way as mentioned for PEI-(5-OMe-3,4-DBA)20.

[0361] Rheology studies

[0362] All rheological experiments were performed on a rotational / oscillating rheometer (MCR102, Anton Paar, Singapore) in a plate-plate geometry with a 10 mm diameter stainless steel probe (PP10). Liquid sample (20 pL) was loaded onto the lower plate with a 0.3 mm probe / plate gap. Steady state rotational conditions: shear rate of 62.8 rad.s-1(10 s-1@ % R) for 30 s. Dynamic mechanical analysis conditions: storage modulus (G’) and loss modulus (G”) at 10 Hz angular frequency and 10% amplitude strain, with a data sampling rate of 1 Hz. Amplitude sweep: shear strain range of 0-1000%, 10 Hz angular frequency. All samples were evaluated in triplicate with a minimum torque requirement of 20 Nm.

[0363] Lap shear adhesion test

[0364] Lap shear adhesion was measured according to ASTM standard F2255-05 using a tensile tester (Chatillon Force Measurement Products, USA) equipped with a 50 N load cell. The collagen strips were cut to 6 cm (long) x 2 cm (wide) and dipped 5 min in PBS prior to testing. 50 pL of the adhesive formulation (in PBS) was appliedover 2 cm x 2 cm area of these hydrated collagen strips. The glued adherends were sandwiched in between two glass slides with paper clips (~ 1 N compressive force) and cured 30 min at ambient temperature. Reported adhesion strengths were calculated dividing the maximum load at failure (N) by the bonding area (m2). All adhesion strength measurements were performed in triplicate.

[0365] Peel adhesion test

[0366] 1800peel adhesion was measured according to ASTM standard F2255-05 using a tensile tester (Chatillon Force Measurement Products, USA) equipped with a 50 N load cell. The collagen strips were cut to 6 cm (long) x 2 cm (wide) and dipped 5 min in PBS prior to testing. 60 |1L of the adhesive formulation (in PBS) was applied over 3 cm (long) x 2 cm (wide) area of these hydrated collagen strips. The glued adherends were sandwiched in between two glass slides with paper clips (~ 1 N compressive force) and cured 30 min at ambient temperature. Reported peel adhesion strengths were calculated dividing the maximum load at failure (N) by width of the specimen(m). All adhesion strength measurements were performed in triplicate.

[0367] Statistical analysis

[0368] For statistics, data were assessed with one-way analysis of variance (ANOVA) followed by Tukey’s test. Specifically, the statistical data were presented as mean ± standard deviation of at least three experiments. One way analysis of variance (ANOVA) with Tukey’s post-hoc test was used to analyse the significant difference between the groups of experimental results. A p-value of < 0.05 was considered statistically significant (n > 3).

[0369] Example 2D: Detailed Discussion on Grafting with Schiff base for Oxygen Activation

[0370] Two model reactions were set-up to investigate the method of grafting and to evaluate the affects on gas (argon versus oxygen) and grafting chemistry (ester versus Schiff base. Model reaction- 1 (MR-1) contained a mixture of methyl-3,4-dihydroxy-5-methoxybenzoate (MDMB) & propyl amine (PA) while the model reaction-2 (MR-2) contained a mixture of 5-Methoxy-3,4-dihydroxybenzaldehyde (5-OMe-3,4-DBA) and PA (FIG. 9). In MR-1, the two components MDMB and PA remained unreacted while in MR-2, 5-OMe-3,4-DBA reacted with PA to form the Schiff base PA-(5-OMe-3,4-DBA) (see FIG. 4A and FIG. 4B). After 30 min, when Schiff-base formation completedin MR-2, O2 was bubbled into both the reaction mixture (MR-1 and MR-2). ’H-NMR investigation exhibited many new peaks revealing the formation of several new products in MR-2 (see FIG. 4C), while in MR-1 the components (MDMB and PA) remained unchanged (see FIG. 4 A and FIG. 4B). Several factors — including the ambient reaction temperature (24 °C), the relatively short reaction time (1 h 30 min), the use of only 1.0 equiv of amine, and the possible formation of catecholate species in the presence of a basic amine — may have collectively suppressed aminolysis in MR-1. Formation of several new products in MR-2 was attributed to the oxidation of catechol functionality in PA-(5-OMe-3,4-DBA) Schiff-base and subsequent reaction with PA, which is consistent with an earlier observation that reported catechol oxidation (NaIO4) in presence of amine produces more than 60 new products. This finding suggested that air / Oi can initiate crosslinking reaction of 5-OMe-3,4-DBA grafted to polyamine [PEI-(5-OMe-3,4-DBA)2o].

[0371] Example 2E: Example Selection of Catechols that are Oxygen Activated within Minutes

[0372] After finding that PA_(5-OMe-3,4-DBA) can undergo various O2 triggered crosslinking reaction with PA, the O2 triggered crosslinking kinetics of 5-OMe-3,4-DBA grafted to the polyamine dendrimer was evaluated and compared with that of 3,4-DBA and 5-NO2-3,4-DBA grafted polyamine dendrimer. The catechol functional group of 3,4-DBA has a hydrogen atom in place of OMe group and is grafted to polymers to prepare adhesives while the catechol moiety of 5-NO2-3,4-DBA contains an electron withdrawing NO2 group in the place of OMe in 5-OMe-3,4-DBA. The rate of crosslinking was measured by calculating aromatic proton integration onNMR spectra. NMR study demonstrated significant reduction in aromatic proton integration of PEI-(5-OMe-3,4-DBA)2o while the aromatic proton integration values of PEI-(3,4-DBA)2O and PEI-(5-NO2-3,4-DBA)20 remained almost unchanged after 10 minutes of O2 bubbling (FIG. 10). This observation resulted in the selection of PEI-(5-OMe-3,4-DBA)2o for further studies.

[0373] From certain studies, 20% grafting to macromolecule (PEI) was identified to have provided suitable viscoelastic material properties, such as viscosity < 1 Pa.s and gelation within minutes after stimulus. Formation of the Schiff-bases occured through a simple one step synthesis. Calculations for 20% grafting are described as follows.

[0374] The general formula of PEI: (CH2CH2NH)n= 43 x n.

[0375] The molecular weight of PEI is 25000.

[0376] Number of (CH2CH2NH) units present =25000= 58143

[0377] So, the number of nitrogens present in PEI = 581

[0378] Since primary:(secondary + tertiary amines) are present in a ratio of 1:2 (Sigma- Aldrich product description)581

[0379] Number of primary amines present = — = 194

[0380] Thus, for 20% grafting on PEI surface amount of 5-OMe-3,4-DBA needed =194 38.8 equivalent.

[0381] At the end of reaction,1H-NMR (FIG. 12B) showed absence of any aldehyde peak, suggesting complete consumption of 5-OMe-3,4-DBA for Schiff-base formation.

[0382] Grafting % calculation from1H NMR before O2 bubbling

[0383] Aromatic CH integration (FIG. 12B) = 75.647 64

[0384] So, the number of 5-OMe-3,4-DBA grafted = — — = 37.82

[0385] There are a total of 194 primary amines present on PEI surface. Therefore, the 3787% of primary amines grafted = x 100 = 19.5%.

[0386] Example 2F: Example of Catechol Functional Group Displaying Low Oxidation Potential

[0387] Cyclic voltammogram displayed the oxidation potential of all the three adhesives. The catechol functionality of both PEI-(MDMB)2o and PEI-(5-OMe-3,4-DBA)2O displayed an intense oxidation peak at low oxidation potential of Eoxi= 0.18 V (vs Ag / AgCl) and Eoxi= 0.22 V (vs Ag / AgCl) respectively (FIG. 11). Thus, catechol functional groups in both the compounds can be taken to have similar oxidation propensity.

[0388] Example 2G: Example of PEI-(5-OMe-3,4-DBA)2o stable for 2 months at 4°C

[0389] The shelf stability of PEI-(5-OMe-3,4-DBA)2o in MeOD solution was investigated at three different temperatures - ambient temperature (24°C), 4°C, and at -20°C. The MeOD solutions of PEI-(5-OMe-3,4-DBA)2o (2.2 x 10-3M) were stored at these temperatures and 'H-NMR recorded for a period of 2 months. The PEI-(5-OMe-3,4-DBA)2O was found to be stable at 4 °C and -20 °C while it decomposed significantly at ambient temperature (FIG. 12A and FIG. 12C).

[0390] Example 2H: Exampe of O2 Bubbling for Higher Crosslinking of PEI-(5-OMe-3,4-DBA)2o

[0391] After finding that the solution of PEI-(5-OMe-3,4-DBA)2o could be stored at 4°C without significant decomposition, its crosslinking kinetics upon exposure to air / O2 was evaluated. Accordingly, air / O2 was bubbled through the MeOD solution of PEI-(5-OMe-Cat)2o for a period of 15 mins. The progress of crosslinking kinetics was monitored by recording the ’H-NMR at 5 mins, 10 mins, and 15 mins intervals. Amine crosslinking to catechol aromatic ring (Michael type reaction), aryl-aryl coupling reaction, etc., result the reduction of aromatic CH integration value. FIG. 12B showed that air / O2 bubble resulted in significant decrease of aromatic CH integration value within 15 min, though O2 caused much higher rate of aromatic CH decrease as compared to air. This suggests that air / O2 initiated crosslinking reaction in PEI-(5-OMe-3,4-DBA)2o.

[0392] Example 21: UV-vis Study: PEI-(5-OMe-3,4-DBA)2o display first order decay in presence of oxygen

[0393] The effect of O2 bubbling on the crosslinking kinetics of PEI-(5-OMe-3,4-DBA)2O Schiff base in MeOH solution was monitored for a period of 3 h using UV-vis spectroscopy. The concentration of PEI-(5-OMe-3,4-DBA)2o Schiff base was 3.4 x 10-6M and 02 / air bubbled from a balloon for 5 s at every 15 min interval. Before bubbling O2, the PEI-(5-OMe-3,4-DBA)2o solution exhibited an intense UV-vis absorption peak at 342.5 nm which originated from the catechol functional group (FIG. 5C and FIG.5E).

[0394] As the 02 / air bubbling progressed into the PEI-(5-OMe-3,4-DBA)2o solution, the intensity of 342.5 nm peak decreased and it displayed a gradual bathochromic shift with the appearance of a low intense broad peak at 341-455 nm (FIG. 5C and FIG. 5E). The reaction mixture also exhibited low intensity UV-vis absorption at 450-600 nm. The decay of 342.5 nm peak followed first order kinetics, with ti / 2 = 16.2 min under O2 (FIG. 5D) and ti / 2 = 17.4 min under air (FIG. 5F), consistent with a faster decay rate in pure O2 compared to air. Crosslinking kinetics of PEI-(5-OMe-3,4-DBA)2o with air bubble gave similar result (ti / 2 = 17.4 min, FIG. 5F).

[0395] O2 bubbling through the reaction mixture oxidised part of the catechol functional groups to reactive quinone. Many amine nucleophiles present in close proximity reacted with the quinone immediately rearomatizing the benzene ring. The decreasing intensity of catechol UV-vis absorption peak (342.5 nm), its gradual transformation into a broad peak at 341-455 nm, and appearance of a low intensity peak at 450-600 nm suggesting the formation of various crosslinking products (FIG. 4D and FIG. 4E).

[0396] Example 2J: Example of Rheological study - PEI-(5-OMe-3,4-DBA)2o Adhesive Composition Gels in < 1 min and Displays Higher Storage Modulus (G’l

[0397] Rheological measurements of PEI-(5-OMe-3,4-DBA)2o and PEI-(MDMB)2o were performed in PBS solution (adhesive:PBS 30:70, w / v) and the results were compared with those of PEI-(B A)2o (control). For all the rheological measurements, 20 |iL of the adhesive solution was loaded on rheometer base plate, which was then subjected to three successive rheology study - steady state viscosity, moduli (G’ & G”), and shear strain measurements. The adhesive was sheared 30 s at the fixed rate of 62.8 rad.s-1for steady state rotational viscosity measurement. At the end of 30 s shearing, the apparent viscosity of all the formulations were found to be stable and in the syringable range (64 - 417 mPa.s) (FIG. 13A to FIG. 13B, and FIG. 26C). The higher value of steady state viscosity of PEI-(5-OMe-3,4-DBA)2o as compared to PEI-(BA)2o (control) (FIG. 14A), suggest the initiation of crosslinking in the formulation. Next, storage and loss modulus (G’ and G”) of the adhesive was measured as a function of angular frequency (FIG. 14B). The PEI-(5-OMe-3,4-DBA)2o formulation gelled in < 1 min (G’ exceeded G”), suggesting its liquid to elastic transition. The PEI-(MDMB)2o and PEI-(BA)2O formulations did not gel within 30 min of study (FIG. 15A to FIG.15C). The storage modulus of PEI-(5-OMe-3,4-DBA)2o formulation continued to increase while that of PEI-(MDMB)2o and PEI-(BA)2o displayed no observable increase and thus these materials remained liquid during the 30 min evaluation period.

[0398] The 30 min cured PEI-(5-OMe-3,4-DBA)2o adhesive displayed linear viscoelastic region (LVR) under small amplitude of applied strain (1-31%, angular frequency = 10 rad.s-1) (FIG. 14C). Within this limit of applied strain G’ and G” remained unchanged and the material displayed elastic behaviour. Further increase in amplitude strain (> 31%), resulted in strain hardening followed by yielding (> 42%)when both G’ and G” decreased and at one point the adhesive acquired quasi-liquid character as the value of G’ becomes lower than G”.

[0399] Example 2K: Example of PEI-(5-OMe-3,4-DBA)2o Display Significantly Adhesion

[0400] Lap shear adhesion

[0401] The adhesion strength of the three adhesives were compared on hydrated collagen film substrates. The adhesive formulation (in ethanol) was applied on the 2 cm x 2 cm area and allowed to cure for 30 min sandwiching between two glass slides. FIG.16A to FIG. 16B, and FIG. 29C displays the representative load displacement graphs of three formulations. As shown in FIG. 17A and FIG. 17B, PEI-(5-OMe-3,4-DBA)2o formulation displayed much higher lap shear adhesion strength (26.2 ± 5.7 kPa) as compared to that of PEI-(MDMB)2o (5.5 ± 0.7 kPa). Both the adhesives displayed cohesive failure, suggesting the difference in adhesion is due to different cohesive force at the interface. The significantly high lap shear adhesion of PEI-(5-OMe-3,4-DBA)2o was attributed to the catechol oxidation and subsequent crosslinking of reactive quinone intermediate with the NFh groups, while the poor lap shear adhesion strength of PEI-(MDMB)2O and PEI-(BA)2o was ascribed to the absence of crosslinking in these materials.

[0402] Peel adhesion

[0403] 1800peel test was used to evaluate the adhesion performance of the adhesives on wet collagen. FIG. 18A to FIG. 18C displays the representative peel adhesion graphs of three formulations. The peel adhesion strengths of PEI-(MDMB)2o and PEI-(2-OMe-3,4-DBA)2O were 4.4 ± 0.1 kPa, and 28.7 ± 3.0 kPa respectively (FIG. 17C). Like lap shear adhesion, PEI-(2-OMe-3,4-DBA)2o displayed significantly higher peel adhesion strength than PEI-(MDMB)20- The pel adhesion strength of PEI-(MDMB)2o correlated with that of control PEI-(BA)2o (7.0 ± 3.0 kPa). Cohesive failure was observed in all three adhesive formulations.

[0404] For both lap shear and peel adhesion, the modulus of toughness of both the adhesives correlated with the adhesion strength (FIG. 17B and FIG. 17D).

[0405] Example 2L: Discussion of Examples 2A to 2K

[0406] In green chemistry, each chemical process is designed to eliminate the use or generation of hazardous materials. Traditional catechol grafted dendrimers result inhigh material property, but toxic chemicals (oxidant or metal chelator) are often required to attain crosslinking. Aiming to look for a tissue friendly method for catechol oxidation, attempts were made to oxidise catechol esters with O2. However, the reaction did not produce positive results. It was then surmised that extended conjugation of catechol might facilitate its oxidation. Preliminary investigation revealed that a catechol Schiff base in presence of excess amine produced several new peaks (Model reaction-2, FIG. 4B) upon O2 exposure, suggestive to the formation of several new products. It was reasoned that catechol grafted polyamine dendrimer Schiff base PEI-(5-OMe-3,4-DBA)2O, which contains excess free amine, would undergo crosslinking upon exposure to air / O2. The material properties of PEI-(5-OMe-3,4-DBA)2o was compared with PEI-(MDMB)2O which is the mixture of catechol ester (MDMB) and PEI. CV measurements revealed that the catechol functionality of both PEI-(5-OMe-3,4-DBA)2o and PEI-(MDMB)2O possessed similar oxidation potential (FIG. 11). The Schiff-base PEI-(B A)2o which did not have a catechol functional group was used as a control. All the three adhesives were prepared in a one step synthesis procedure. Model reaction showed that the catechol group of MDMB was not oxidised to quinone upon O2 exposure and therefore did not react with amine present in the reaction system (MR-1, FIG. 9, FIG.4A). PEI-(5-OMe-3,4-DBA)2o upon exposure to air / O2 crosslinked by Michael addition or aryl-aryl coupling reaction, resulting in a significant decrease in aromatic CH integration value (FIG. 12B). O2 initiated crosslinking of PEI-(5-OMe-3,4-DBA)2o was further evidenced by the first order decay of catechol absorption peak (342.5 nm) and its transformation to a broad peak at 341-455 nm (FIG. 5C to FIG. 5F). At 30 wt% PEI-(5-OMe-3,4-DBA) 20 formulation, the polyamine createed a pH of 11-12, which may form anionic catechols. However, in the absence of complexing metal ions in the formulation, the observed polymerization was attributed solely to catechol oxidation.

[0407] The crosslinking in PEI-(5-OMe-3,4-DBA)2o was further reflected in its material property. The higher value of steady state viscosity of PEI-(5-OMe-3,4-DBA)2O as compared to PEI-(MDMB)2o and PEI-(BA)2o (control) at the end of 30 s searing was due to the initiation of crosslinking while preparing the PBS formulation at ambient conditions. The O2 dissolved in PBS oxidised catechol in PEI-(5-OMe-3,4-DBA)2O Schiff base to reactive quinone, which subsequently crosslinked via various reactions. Although PEI-(MDMB)2o contains catechol, it did not oxidize to quinonebecause the catechol was not in Schiff-base form, while PEI-(BA)2o lacked catechol entirely. As a result, the PEI-(5-OMe-3,4-DBA)2o formulation thus gelled in < Imin and exhibited a steady increase in storage modulus (G’) over a 30 min evaluation period. The PEI-(MDMB)2o like the control PEI-(BA)2o did not gel or exhibit increase in its storage modulus (G’) (FIG. 14B, FIG. 15A to FIG. 15C).

[0408] The alcoholic solution of PEI-(5-OMe-3,4-DBA)2o was found to be stable at 4 °C if stored under inert atmosphere (argon). However, the solution slowly decomposed at ambient temperature (FIG. 12A). Thus, for application on substrates, the alcoholic solution stored in cold condition can be used repeatedly, without any need for preparing fresh adhesive formulation for every use. After applying on substrate, the alcohol evaporates and simultaneously exposes the catechol-Schiff base to air, which initiates crosslinking. As expected, PEI-(5-OMe-3,4-DBA)2o exhibited significantly higher lap shear and 1800peel adhesion strength than PEI-(MDMB)2o (FIG. 17 A, FIG. 17C), though all the adhesives displayed cohesive failure. Higher cohesive strength is suggestive to the presence of higher crosslinking density, which in turn increases the intermolecular forces (H-bond, Tt-Tt stacking, Van der Walls interactions, etc., and physical entangling between polymeric chains resulting in higher cohesive force in PEI-(MDMB)2O adhesive.

[0409] The present examples showed that air / O2 can initiate catechol oxidation when the catechol group is grafted as Schiff base.

[0410] Example 3A: Examples on Electron Donating Groups Enhancing Crosslinking Kinetics of Substituted Catechol-Schiff base Adhesive Compositions

[0411] Examples 3A to 3K shows a library of catechol grafted PEI dendrimers, PEI-(sCat)2o, bearing catechol substituents of different electronic nature at the o- and m-position of aromatic moiety that were synthesized employing a green process of Schiff base formation. 5-OMe substituted catechol Schiff base displayed the most favourable oxidation potential and peak current for the catechol to quinone oxidation. Crosslinking of PEI-(sCat)2o prepolymers was triggered with nontoxic and environmentally friendly molecular O2. ’H NMR and UV-vis study revealed that EW (NO2, Br) catechol substituted PEI-(sCat)2o displayed slower crosslinking kinetics while ED (OMe) catechol substituted PEI-(sCat)2o displayed higher crosslinking kinetics under oxygen gas exposure. Rheological investigation displayed that ED (OMe) catechol substitutedPEI-(sCat)2o gelled in < 1 min and could achieve G’ 6.4 kPa while EW (NO2, Br) catechol substituted PEI-(sCat)2o did not gel in 30 min. ED catechol substituted Schiff base PEI-(5-OMe-3,4-DBA)2o displayed highest lap shear (26.2 ± 5.7 kPa) and 1800peel adhesion strength (28.7 ± 3.0 J.nr2) on wet collagen substrates.

[0412] To reiterate, mussel adhesive proteins (MAPs) tend to be abundant with 3,4-dihydroxyalanine (DOPA) and L-lysine. The adhesion ability of mussels on a variety of surfaces may be ascribed to the oxidation of the catechol group of DOPA to quinone followed by crosslinking with L-lysine. Since the discovery of mussel adhesion chemistry, grafted polymers have been attempted to exploit the catechol chemistry for adhesive designs. A traditional strategy conjugates natural and synthetic polymers with commercially available dopamine or its carboxylic derivative hydrocaffeic acid. However, it seems to remain unknown how catechol substituents affect crosslinking kinetics and the material properties of the final adhesive. The crosslinking kinetics and adhesive strength of catechol adhesives currently rely on external additives for initiation (oxidant / chelator). The oxidant or chelator additive are cytotoxic, which limits their application.

[0413] Aiming to find a greener reagent for catechol oxidation, it was discovered that oxygen (O2) can oxidise a 5-OMe substituted catechol to the corresponding quinone when present within a Schiff base structure (see examples 2A to 2L). Encouraged by this observation, it was hypothesized that catechol oxidation with molecular O2 and the resulting adhesive properties of the catechol derived Schiff base could be tuned using substituents of different electrochemical nature. Generally, an electron donating (ED) substituent may facilitate catechol oxidation by enhancing aromatic ring electron density while an electron withdrawing (EW) substituent may slow it down by reducing aromatic ring electron density. Further investigation is therefore sought to evaluate the effect of different substituents on catechol oxidation by molecular O2. A library of PEI derived catechol Schiff bases, PEI-(sCat)2o, was synthesized using a variety of substituents at the o- and m- position of the catechol ring. A comparative study of these catechol Schiff bases was carried out to investigate their propensity towards oxidation with molecular O2 and ensuing material property of resulting adhesives (FIG. 1).

[0414] Branched polyethylenimine (PEI) was chosen as the dendrimer backbone for these examples, as it presents a surface of 1°, 2°, and 3° amine groups. Catecholaldehydes were spontaneously grafted onto the PEI surface through Schiff base formation. This catechol-g-PEI forms a prepolymer that undergoes crosslinking via condensation polymerization when the catechol converts to quinone. Excess amines of PEI enhance water solubility, create a basic environment suitable for catechol oxidation, and endow antimicrobial, antiviral coatings of the cured adhesive.

[0415] Experimental - Materials

[0416] Polyethylenimine (PEI), 3,4-dihydroxybenzaldehyde (3,4-DBA), Benzaldehyde (BA), aluminium bromide (AIBra), nitrobenzene, phosphate buffer, anhydrous methanol (MeOH), absolute ethanol (EtOH), Phosphate-buffered saline (PBS), were purchased from Sigma-Aldrich, Singapore. 3,4-Dihydroxy-5-nitrobenzaldehyde (5-NO2-3,4-DBA) and 3,4-Dihydroxy-5-Methoxybenzaldehyde (5-OMe-3,4-DBA) were purchased from TCI, Singapore. 3,4-Dihydroxy-6-nitrobenzaldehyde (6-NO2-3,4-DBA), 3-Bromo-4,5-dihydroxybenzaldehyde (3-Br-4,5-DBA), 2-Bromo-4,5-dihydroxybenzaldehyde (2-Br-4,5-DBA), 6-methoxy-l,3-benzodioxole-5-carbaldehyde were purchased from Fluorochem, UK.

[0417] Experimental - General Methods

[0418] All experiments were carried out as described in Example 2C, “General methods”.

[0419] Experimental - Synthesis of4,5-dihydroxy-2-methoxybenzaldehyde (2-OMe-4,5-DBA)<"

[0420]

[0421] To an ice-cold solution of 6-methoxy-l,3-benzodioxole-5-carbaldehyde (1.0 g, 5.5 mmol) in nitrobenzene (50 mL) was added Al Bra (2.9 g, 11.1 mmol) in portions under argon atmosphere. The reaction mixture was stirred at room temperature for 7 h.200 mL aqueous HC1 (0.3 N) was added and stirred for 1 h. The reaction mixture was extracted with ether (2 x 100 mL). The organic extracts were combined, dried (NaiSCU), filtered and concentrated. Petroleum ether (200 mL) added into the nitrobenzene residue while stirring vigorously. The resulting precipitate was filtered, washed with petroleumether to afford 4,5-dihydroxy-2-methoxybenzaldehyde (2-OMe-4,5-DBA) as a brown solid (337 mg, 36%). ’H NMR (400 MHz, CD3OD) 8 10.11 (s, 1H), 7.16 (s, 1H), 6.53 (s, 1H), 3.84 (s, 3H).

[0422] Experimental - General procedure for the synthesis of PEI-(sCat)2o prepolymers

[0423] To a methanolic solution of PEI (0.044 mmol, in 30 mL MeOH), was added substituted catechol aldehyde (1.74 mmol) in portions over a period of 5 min under argon atmosphere. The reaction mixture was stirred 30 min. It was concentrated in a rotary evaporator and dried in vacuum for 4-5 h to afford PEI-(sCat)2o in quantitative yield.

[0424] Experimental - Cyclic voltammetry ( CV)

[0425] The cyclic voltametric measurements were recorded as described in Example 2C, “Cyclic voltametric studies”. In this instance, the cyclic voltammetric data of PEI- (sCat)2o compounds are recorded in PBS solution (0.13 mM), at the scan rate 0.1 V / s.

[0426] Experimental - Preparation of PEI-(sCat)2o adhesive formulation

[0427] The 30 wt% formulation in PBS was prepared by adding 90 mg PEI-(sCat)2o in 210 p L PBS buffer (pH - 7.2) and vortexing the mixture until a clear homogeneous solution is observed. This formulation (20 pL) was loaded onto the rheometer base plate for rheology measurements. For adhesion strength measurements study the 30 wt% formulation of PEI-(sCat)2o adhesives were prepared in a similar way using EtOH in place of PBS.

[0428] Experimental - Rheology Measurements

[0429] The viscoelastic properties of PEI-(sCat)2o adhesives were recorded as described in Example 2C, “Rheological studies”. Storage and loss modulus (G’ and G”) had a data acquisition rate of 1 Hz using a 10 Hz angular frequency and 10% strain amplitude. Amplitude sweeps determine the shear strain at break, yield stress, linear viscoelastic range, or combination thereof of the cured PEI-DBA20 adhesive.

[0430] Experimental - Lap shear and 180° Peel adhesion on collagen substrates

[0431] Lap Shear and peel adhesion studies are performed as described in example 2C, “Lap shear adhesion test” and “Peel adhesion test”. Collagen strips (2 cm x 5 cm) were used as the adherend. The collagen strips were dipped 5 min into PBS buffer, removed and the superficial water is absorbed with tissue paper. PEL(sCat)2o adhesive(50 p L) was applied at one end of a collagen strip (2cm x 2 cm) and sandwiched with another collagen strip. This structure was placed in between two glass slides secured with paper clips (~ IN compressive force). After 30 minutes curing at room temperature lap shear adhesion test is performed. Adhesive shear strength at failure determined by maximum load divided by the adhered surface area. At least three lap shear adhesion tests performed for each adhesive.

[0432] The interfacial toughness of PEI-(sCat)2o adhesives were evaluated with 180-degree peel adhesion strength measured on a tensile tester (Chatillon Force Measurement Products, USA) pulling the sample at the speed of 10mm min-1using a 50N load cell. The collagen strips adhered with PEI-(sCat)2o adhesives (60 pL, length 3cm x width 2 cm) were prepared the same way as lap shear adhesion test. Peel adhesion strength is determined following previously established guidelines. At least three peel adhesion tests performed for each adhesive.

[0433] Experimental - Leachate assessments sample preparation

[0434] The PEI-(3,4-DBA)20and PEI-(5-OMe-3,4-DBA)20adhesives (1.66 g each) were cured in a sample vial at 37 °C for 48 h in an oven. ISO 109935 guidelines suggest to extract adhesive leachates at temperatures to which the adhesive may be exposed to during use. Accordingly, PBS (5 mL, pH - 7.2) was added onto the adhesive and the vial placed at 37 °C. After 24 h, the leachates were filtered, topped up to 10 mL with PBS and stored at -18 °C prior to use for in vitro skin sensitisation direct peptide reactivity assay (DPRA).

[0435] Experimental - In vitro Skin Sensitization Assay Analysis

[0436] An in vitro skin sensitization assay was conducted on leachates of PEI-(3,4-DBA)2O and PEI-(5-OMe-3,4-DBA)2o adhesive samples based on OECD TG 442C Guideline for in chemico skin sensitisation: direct peptide reactivity assay (DPRA) with slight modification. Briefly, the stock solutions of cystine (0.667 mM, 0.501 mg / mL in phosphate buffer at pH = 7.5 ± 0.05) and lysine were freshly prepared with a purity concentration of above 85%. lOOmM of cinnamic aldehyde (in acetonitrile) was used as the positive control. The test substances derived from leachates were used for the assay directly (composition and their concentration unknown). Peptide standards were diluted from the stock solution in the range of 0.534mM to 0.0167mM. Peptidedepletion was measured by HPLC peak area using UV detector (220 nm). Co-elution control of test substances did not absorb at 220 nm suggesting no interference.

[0437] Experimental - AMES Assay Analysis

[0438] An Ames test was conducted on 3,4-DBA and 5-0me-DBA samples to determine if it could cause DNA mutations based on the protocol by the Organization for Economic Cooperation and Development (OECD) 471 Guideline for Testing of Chemicals: Bacterial Reverse Mutation Test. Briefly, the positive controls used in the absence activation system are sodium azide, 2-nitrofluorene and mitomycin-C for bacterial strain TA 100, TA 98 and TA 102 respectively. Positive control used for the presence activation system is benzo(a)pyrene for all 3 bacteria strains (TA 100, TA 98 and TA 102). DI water was used as the negative control for all experimental runs.

[0439] Experimental - Statistical method

[0440] The statistical method was carried out as described in Example 2C, “Statistical analysis”.

[0441] Example 3B: Scope of PEI-(sCat)2o Adhesive Compositions

[0442] All the catechol grafted polyethylenimine (catechol-g-PEI) adhesives were synthesized following a one step simple procedure. To compare rheological property and adhesion strength, substituted catechol (sCat) grafting percentage was maintained at 20% mol / mol of catechol to primary amine, represented as PEI-(sCat)20- The synthesized catechol Schiff bases were characterized byNMR and UV-vis spectroscopy. Cyclic voltammetry evaluates the oxidation potential of the substituted catechol within a Schiff base structure. Crosslinking kinetics of PEI-(sCat)2o was evaluated byNMR and UV-vis as a function of oxygen exposure. Adhesive formulations were prepared in PBS / EtOH (30% w / v) to compare rheological and adhesion properties. Viscosity measurement revealed liquid behaviour and wettability on substrates. Adhesive viscoelasticity parameters were evaluated as a function of time and catechol structure. Adhesive strength demonstration on a wet collagen film display potential applications on wet substrates and similar soft surfaces.

[0443] Example 3C: 'II NMR and UV-vis Spectra Confirms PEI-(sCat)2o Schiff Base Prepolymers

[0444] Green synthesis of Schiff base formation was carried out using a simple one-step mixing procedure.1H NMR spectra showed that the Schiff base formation wascompleted within 1 hr regardless of the type of electron donating / withdrawing group catechol (FIG. 19). The catechol grafting was confirmed byNMR and UV-vis spectra analysis. TheNMR spectra of all the compounds show characteristic imine CH peak (shaded yellow) and aromatic CH peaks (6.0-7.8 ppm).

[0445] The UV-vis spectra of Schiff base prepolymers (i.e., the adhesive compositions) exhibit strong absorption peaks at 301 nm, 334.5 nm, 341.5 nm, and 388.5 nm for PEI-(5-N02-3,4-DBA)2o, PEI-(3-Br-4,5-DBA)20, PEI-(5-OMe-3,4-DBA)2O, and PEI-(2-OMe-4,5-DBA)2o respectively (FIG. 20) which are attributed to the catechol functionality in the Schiff base. The analysis of ’H NMR and UV-vis spectra together demonstrated the successful grafting of catechol to PEI surface amines.

[0446] Example 3D: Example of PEI-(5-OMe-3,4-DBA)2o Schiff Base Redox Potential

[0447] Cyclic voltammetry measurement evaluates the oxidation potential of PEI-(sCat)2o compounds. PEI-(sCat)2o compounds were measured in PBS (0.13 mM) at 0.1 V.s-1scan rate. As shown in FIG. 21A to FIG. 21G, oxidation potential of 5-NO2substituted catechol Schiff base was the highest (Eoxi= 0.31 V) suggesting it had lower oxidation kinetics (i.e., higher oxidation activation energy), while the oxidation potential of 2-OMe substituted catechol Schiff base was the lowest (Eoxi= 0.11 V), suggesting it had higher oxidation kinetics (i.e., lower oxidation activiation energy) to quinone. Rest of the substituents displayed moderate oxidation potential of 0.21-0.23 V. Measuring CV of PEI-(sCat)2o prepolymers under the same concentration (0.13 mM) and same scan rate (0.1 V.s-1), oxidation peak current indicated the proportion of catechol groups was oxidised to quinone as the catechol to quinone oxidation is a two electron irreversible process. FIG. 22 and FIG. 21A to FIG. 21G showed that 5-OMe substituted catechol in PEI-(5-OMe-3,4-DBA)2o had moderate oxidation potential and highest peak current and further exhibited one of the most favourable O2triggered oxidation.

[0448] Example 3E: 'll NMR of Electron Donating Substituents Demonstrate Higher Tendency towards O2Oxidation

[0449] ’ll NMR and UV-vis studies were performed to compare the crosslinking kinetics of various PEI-(sCat)2o prepolymers. Crosslinking of PEI-(sCat)2o prepolymers involves the oxidation of catechol to reactive quinone intermediate. Condensationpolymerization proceeds via aryl-aryl coupling reaction or via reactions with nucleophiles (NH2, SH, OH, etc.) via Michael type addition or Schiff base formation to produce more than 60 different products. In presence of excess amines, Michael type additions form the major crosslinking mechanism. Thus, most of the reaction products are formed by replacing an aromatic proton with carbon or a heteroatom (N, S, etc.) which can be quantitatively measured by the decrease in aromatic proton integration on ’H NMR. Air or oxygen exposure tests were performed to evaluate the tendency of substituted catechols towards O2 initiated oxidation in Schiff base structure. The PEI-(sCat)2o structures within the gas-bubbled reaction mixture were recorded via1H NMR over a 15 min period. FIG. 12D, FIG. 12E and FIG. 12B demonstrate no significant reduction in aromatic proton integration, suggesting that catechols bearing strong (NO2) and weak (Br) electron withdrawing substituent did not crosslink significantly during 15 min of air / O2 bubbling. However, air / O2 bubbling (15 min) into catechol Schiff base bearing an electron donating substituent (OMe) resulted in a significant reduction in aromatic proton integration (FIG. 12B), suggesting their proneness towards O2 triggered oxidation and subsequent crosslinking reaction.

[0450] Example 3F: Example of Electron Donating Catechol Substituents Display 1storder Catechol Group Decay on UV-Vis

[0451] UV-vis studies provided further insight into O2 triggered crosslinking kinetics of PEI-(sCat)2o prepolymers. O2 bubbling UV-vis study was carried out over a 3 h period. Every time before recording the UV-vis spectra, O2 was bubbled into a MeOH solution (3.4 x 10-6M) of PEI-(sCat)2o for a period of 5 s at every 15 min interval. Before O2 bubbling, PEI-(3-Br-4,5-DBA)2o solution displayed an intense UV-vis absorption peak at 334.5 nm and a weak UV-vis absorption peak at - 420 nm (FIG.5A), which were attributed to catechol and quinone respectively. FIG. 5B showed no significant decay of catechol absorption peak (334.5 nm) over 3 hrs period. A slight reduction of the quinone peak intensity, indicateed less than 5% conversion to quinone after 180 mins.

[0452] Before O2 bubbling, the catechol peak (342.5 nm) predominated in the UV-vis spectra of PEI-(5-OMe-3,4-DBA)2o, with only a weak peak of quinone observed at at 440 nm, as seen in FIG. 5C. The O2 bubbling over 3 h resulted in significant decay of both catechol (1storder decay) and quinone absorption peaks, besides formation of newbroad peak at 480-600 run (FIG. 5C to FIG. 5D). The latter broad peaks were evidence of quinone crosslinking intermediates. Similar UV-vis spectra were also observed for ED OMe substituent at the o-position (FIG. 23A and FIG. 23B). BothNMR and UV-vis results suggest that an EW substituent (Br) slowed down and an ED substituent (OMe) accelerated the catechol crosslinking reaction in their Schiff base structure.

[0453] Example 3G: Example of Electron Donating Catechol Substituted Schiff Base has 2 Months Stability at 4°C

[0454] Stability of MeOD solutions of 5-NOi, 3-Br, and 5-OMe substituted PEI-(sCat)2o prepolymers (2.2 x 10-3M) at ambient temperature (24°C), and at 4°C was investigated for 2 months byNMR analysis. As mentioned earlier, decreases in aromatic proton integration suggests the transformation of catechol to quinone and subsequent crosslinking reactions. FIG. 24A to FIG. 24C demonstrated that 5-NOi substituted catechol Schiff base was unstable both at ambient temperature as well as at 4°C and that this monomer could not be stored even at cold condition. The aromatic protons integration for 3-Br and 5-OMe substituted catechol Schiff base reduced significantly at ambient temperature but instead displayed only a small reduction at 4°C. Thus, the methanolic solutions of PEI-(3-Br-4,5-DBA)2o and PEI-(5-OMe-3,4-DBA)2o could be stored for months at 4°C without significant decomposition.

[0455] Example 3H: Example of Electron Donating Catechol Substituted Schiff Base Display Higher Modulus

[0456] The viscoelastic and rheological properties of PEI-(sCat)2o adhesives were investigated using an oscillatory rheometer. The unsubstituted catechol Schiff base PEI-(3,4-DBA)2O was used as a control for this study. The adhesive formulations of PEI-(sCat)2o prepolymers were prepared by vortexing 20-30 min in PBS (30% w / v). 20 p L PEI-(sCat)2o of each formulation was loaded on the rheometer base plate and three successive rheology investigations were performed, namely apparent viscosity, moduli change, and amplitude sweep. Apparent viscosity data obtained by shearing the formulations for 30 s at the fixed rate of 62.8 rad.s"1at ambient temperature showed liquid and syringe-able behaviour of all the adhesive formulations (FIG. 25, FIG. 26A to FIG. 26F). After 30 seconds viscosity measurements, storage modulus (G’) and loss modulus (G”) of the adhesives were recorded for 30 min using 10 Hz angular frequency and 10% amplitude strain. Increase in G’ over time suggests the increase in crosslinkingdensity until crosses the gelation point (defined as the timepoint were G’(t) > G”(t)). After this timepoint, crosslinking has reached a viscoelastic solid nature. Only the -OMe formulation had storage moduli greater than the control; PEI-(5-OMe-3,4-DB A)io @ 5500+ 300 Pa, and PEI-(2-OMe-4,5-DBA)206400 + 200 Pa respectively (FIG. 6). PEI-(sCat)2o adhesives with ED substituents (5-OMe and 2-OMe) exhibited much faster gelation and higher G’ than with EW substituents of 3-Br or NO2(FIG. 27A to FIG.27F). At the end of 30 min moduli measurement, the cured material was subjected to amplitude sweep (1 to 1000% strain) to evaluate the yield stress and strain. EW substituted PEI-(sCat)2o adhesives (NO2, Br) displayed predominately liquid behaviour (G” > G’) (FIG. 28 A to FIG. 28F). Electron donating OMe substituted PEI-(sCat)2o adhesives displayed a linear viscoelastic regime up to 40 % amplitude strain.

[0457] Example 31: Example of Electron Donating Catechol Substituted Schiff Base Display Higher Adhesion Strength

[0458] Lap shear and 1800peel adhesion strength of different PEI-(sCat)2o adhesives were compared on hydrated collagen substrates. The latter serves as a wet substrate mimic of soft tissues that that remains an unmet clinical challenge within bioadhesives. The PEI-(sCat)2o adhesive formulations were prepared by vortexing 10-15 min in EtOH (30% w / v). The adhesive formulation was applied in between two wet collagen strips (2 cm x 2 cm) which were then allowed to cure 30 min. Representative lap shear adhesion strength graphs of PEI-(sCat)2o adhesives are shown in FIG. 29A to FIG. 29F. Lap shear adhesion strength calculated at the yielding point of force-displacement graphs are PEI-(5-NO2-3,4-DBA)20 14.6 + 1.3 kPa, PEI-(6-NO2-3,4-DBA)20 10.7 + 2.3 kPa, PEI-(3-Br-4,5-DBA)2o 19.9 + 0.8 kPa, PEI-(2-Br-4,5-DBA)2o 18.7 + 1.2 kPa, PEI-(5-OMe-3,4-DBA)2o 26.2 + 5.7 kPa, and PEI-(2-OMe-4,5-DBA)2o 11.1 + 0.9 kPa respectively (FIG. 30A). The PEI-(5-OMe-3,4-DBA)2o and PEI-(3-Br-4,5-DBA)2o adhesives displayed significantly higher lap shear adhesion strength than rest of the PEL(sCat)2o adhesives as compared to control. The lap shear adhesion modulus of toughness of PEL(5-OMe-3,4-DBA)2o adhesive was significantly higher than all other PEL(sCat)2o adhesives (FIG. 30B).

[0459] Preparation of PEL(sCat)2o adhesives formulation, application on wet collagen substrates (length = 3 cm, breadth = 2 cm), and curing procedure for 1800peel adhesion measurement followed the procedure of lap shear adhesion. FIG. 31 A to FIG. 3 IF showthe representative peel adhesion test graphs. The average value of 1800peel adhesion strength calculated are PEI-(5-N02-3,4-DBA)2o 29.2 ± 4.8 J.nr2, PEI-(6-NO2-3,4-DBA)2O 22.1 ± 1.8 J.nr2, PEI-(3-Br-4,5-DBA)2023.3 ± 3.5 J.nr2, PEI-(2-Br-4,5-DBA)2022.9 ± 3.5 J.nr2, PEI-(5-OMe-3,4-DBA)2028.7 ± 3.0 J.nr2. and PEI-(2-OMe-4,5-DBA)2O 19.2 ± 1.7 J.nr2respectively (FIG. 7). Thus, PEI-(5-N02-3,4-DBA)2o and PEI-(5-OMe-3,4-DBA)2o displayed significantly high peel adhesion strength than other PEI-(sCat)2o adhesives. Cohesive failure was observed for all the specimens.

[0460] Example 3J: Examples of Adhesive Composition, PEI-(3,4-DBA)2o and PEI-(5-OMe-3,4-DBA)2o, and such adhesives have low skin sensitization reactivity and are non-mutagenic

[0461] Repeated exposure to allergenic substance may provoke adverse health effects. Skin sensitization assays identify chemicals that can elicit allergic responses. Direct peptide reactivity assay (DPRA) is a nonanimal in vitro method which evaluates skinsensitizing ability of a chemical by measuring its reactivity with a peptide. DPRA quantitatively measures the depletion of synthetic Cysteine and Lysine peptides with the leachates of PEI-(3,4-DBA)2o and PEI-(5-OMe-3,4-DBA)2o adhesives that mimics the chemical reactions involved in skin sensitization. The degradation products of both PEI-(3,4-DBA)2O and PEI-(5-OMe-3,4-DBA)2o adhesives exhibit low reactivity towards both Cysteine and Lysine peptide binding in DPRA assay (FIG. 34A).

[0462] The Ames assay was employed to evaluate the mutagenic potential of PEI-(3,4-DBA)2O and PEL(5-OMe-3,4-DBA)2o adhesives in bacterial systems, with the objective of assessing their possible carcinogenic properties. As shown in FIG. 34B and FIG. 34C, neither compound induced a statistically significant increase in revertant colony formation at a concentration of 5 mg / plate in the presence or absence of metabolic activation, relative to the negative control. Only the positive control yielded a marked increase in colony numbers, confirming the assay's sensitivity. Furthermore, data presented in FIG. 34B and FIG. 34C demonstrated no dose-dependent increase in revertant colonies at lower treatment concentrations (2.5, 1.25, 0.625, and 0.3125 mg / plate) for both PEL(3,4-DBA)2o and PEL(5-OMe-3,4-DBA)2o adhesives. These findings suggested that under the conditions tested, neither compound possessed mutagenic activity in this bacterial model.

[0463] Example 3K: Discussion for Examples 3 A to 3J

[0464] Catechol Schiff base prepolymers (the adhesive compositions) were synthesized with the catechol aromatic moieties carrying EW and ED substituents at o-and m- positions to investigate the structure property relationships of PEI-(3-Br-4,5-DBA)2O having catechol substituents of diverse electronic nature. Generally, an electron donating substituent (OMe) enhances electron density of the catechol aromatic ring and an electron withdrawing substituent (-NO2, Br, etc.) has the opposite effect. Therefore, ED substituents may facilitate the oxidation process by lowering the oxidation potential value, and an EW substituent may slow down the oxidation process by increasing the value oxidation potential of catechol group in PEI-(sCat)2o compounds. The overall preparation of adhesives reported here is a two-step process - first step is the grafting of catechol onto PEI, and the second step is crosslinking reaction of catechol grafted PEI dendrimer. Green chemistry aims to design chemical process that reduces or eliminates the use of toxic or hazardous chemicals. Development of an efficient green technology reduces environmental pollution, create a safe work environment and manufactures safe products to the end user. In developing a green and sustainable grafting reaction, Schiff base prepolymers appeared more appropriate as no reagent was needed to synthesise them. With the Schiff base prepolymers in hand, the next step was to crosslink them to attain adhesive properties. The formation of quinone intermediate leads to crosslinking of the catechol derived Schiff base prepolymers. The successful grafting of substituted catechol aldehydes to PEI was characterized byNMR and UV-vis spectra analysis of PEI-(sCat)2o prepolymers (mentioned in examples 3A to 31), which was further confirmed by the changes in redox behavior of substituted catechol aldehydes after Schiff base formation (FIG. 32A and FIG. 32B). While oxidation potential reveals the oxidation tendency of a molecule, the peak current states the proportion of the molecules in solution may undergo oxidation in that potential. Weak peak current in the CV spectra of 3-Br, 2-Br, and 2-OMe substituted catechol Schiff bases suggest that only a minor proportion of catechol groups of these Schiff bases undergo catechol to quinone oxidation despite having lower oxidation potential. The Oxidation 5-OMe substituted catechol had the highest peak current with a moderate oxidation potential, suggesting this is the optimial candidate for O2 oxidation (FIG. 22 and FIG. 21A to FIG. 21F).NMR investigation of methanolic solution of PEI-(5-N02-3,4-DBA)2O monomer bearing an EW NO2 group reveals that it is unstable in bothambient temperature (24°C) and cold condition (4°C). The Schiff base prepolymers PEI-(3-Br-4,5-DBA)2o and PEI-(5-OMe-3,4-DBA)2o which have a weak electron withdrawing Br and a strong electron donating 5-OMe catechol substituent respectively are stable for 2 months at 4°C but they crosslink significantly at 24°C and thus cannot be stored at ambient temperature.

[0465] O2 bubbling into methanolic solution of PEI-(sCat)2o prepolymers and concurrent ’H NMR and UV-vis investigation evaluated their crosslinking kinetics. There was no significant drop of aromatic CH integration of PEI-(sCat)2o compounds bearing EW substituents (NO2 and Br) after 15 min of O2 bubbling (FIG. 12D and FIG.12E), while the catechol Schiff base with ED substituent (OMe) showed significant reduction of CH integration in ’H NMR spectra (FIG. 12B). The UV-vis absorption peak of catechol group at 334.5 nm of weekly EW 3-Br substituted Schiff base display no significant decay (FIG. 5B) while UV-vis absorption peak at 342.5 nm of ED 5-OMe substituted Schiff base display 1storder decay (FIG. 5D). Thus, both the ’H NMR and UV-vis investigation suggests that ED catechol substituents facilitate while EW substituents slow down catechol oxidation and ensuing crosslinking reactions.

[0466] The rheological behavior of PEI-(sCat)2o prepolymers measured under ambient condition aligns with the UV / NMR kinetics evaluations. Upon dissolution in PBS, the catechol group in PEI-(sCat)2o prepolymers are oxidized to quinone at rates dependent on the electronic nature of the substituent. The reactive quinone intermediate covalently crosslinks with excess amine nucleophiles on PEI surface via Michael type addition, Schiff base formation or couples with another catechol group forming a chemically bonded polymeric chain. Various nucleophilic functional groups (e.g., OH, NH2) present on substrates and natural polymers undergo a combination of mechanical, physical, and chemical bonding. The chemical and physical bonding together build up the 3-D supramolecular network in PEI-(sCat)2o adhesives that give rise to their cohesive strength. The measurement of rheological properties provides quantitative measurement about the changes in cohesive force of the adhesive over time. Apparent viscosity of EW substituted PEI-(sCat)2o adhesives (NO2, Br) below 0.2 Pa.s suggests little crosslinking reaction on the minute timeframe. In contrast, ED substituted PEI-(sCat)2o adhesives (OMe) above 0.2 Pa.s suggest faster reaction kinetics (FIG. 25). The rate of increase in crosslinking density is reflected on achieving gelation times withinminutes of dissolution. EW substituted PEI-(sCat)2o adhesives (NO2, Br) did not observe gelation within the 30 min evaluation period (FIG. 25, FIG. 27A to FIG. 27F). The ED substituted PEI-(sCat)2o adhesives (OMe) displayed an evolving G’ value of 6 kPa at the end of 30 min. This further supports that ED catechol substituents facilitate catechol to quinone oxidation.

[0467] The degradation products of PEI-(3,4-DBA)2o and PEI-(5-OMe-3,4-DBA)2o adhesives show a little reactivity to in vitro skin sensitization test (DPRA assay). This is because the phenolic functional groups of catechol or its Schiff base can undergo covalent interaction with the sulfhydryl (-SH) and amine (-NH2) groups of cysteine and lysine peptides.

[0468] Example 4A: Adhesive Compositions as Antibacterial, Wash-Resistant Coatings based on Imine-Grafted Catechol Isomers

[0469] Examples 4A to 4D demonstrate adhesive composition of the present disclosure as antibacterial and / or wash-resistant coatings.

[0470] With constant need of surface sterilization to prevent bacterial contamination, easily applicable, non-leaching, wash resistant and contact-killing coatings are the ideal solution to the problem of inhibiting bacterial growth. Herein catechol coatings utilizing Schiff-base grafting to polyethyleneimine matrix are evaluated based on their contact inhibition against S. aureus and E.coli bacteria and their performance after water and ethanol wash, comparing zone of inhibition on agar plates as well as toxicity of leachates from water submerged coatings

[0471] In both gastronomical and biomedical fields, bacterial contamination is still a prevalent problem, despite rising sanitation standards. Bacteria such as S. ureus and E.coli can live from up to 12 days on dry inanimate surfaces such as plastics, depending on the conditions. Usage of metals shortens the bacterial lifespan, but even metal tools the lifespan of bacterial contamination can reach up to 72 hrs on stainless steel, with reduction up 3-6 hrs on copper. Heat, disinfectants and UVA-sterilisation are commonly used as sanitation vectors, but when coming in contact with food containers and food directly, the number of methods is further limited. Application of thin, antibacterial coatings alleviates the need for sterilization, using either drug-releasing materials, surface-killing films, such as nanoparticle-releasing gels and growth preventing surfaces through topological modifications (plasma treatment, chaingrafting). There appears however no easy to apply a coating that shows contact-killing behaviour.

[0472] Drug-releasing coatings utilize metal ions (such as silver, copper, zinc.), enzymes for targeting gram positive / negative strains (such as lysozyme for Grampositive, and acylase for Gram-negative), or antibiotics such as penicilins. Such surface treatments are prepared using hydrogel matrices and are often paired with stimuli-responsive polymers for controlling the dosage of antibacterial leachates, but their applications are limited to biomedical and clinical fields due to costs and controlled release of active agent.

[0473] Surface-killing coatings require physical contact with the bacterial wall to degrade and rupture the outer layer, killing the bacteria in process, often utilizing positively charged ammonium groups (R4N+). While effective, the accumulation of dead bacterial residue inhibits the coating’s effectiveness, while functionalization of the surfaces impedes scalability of the method. Therefore, easy-to-apply surface coatings attaining drug-releasing properties and exhibiting contact-killing characteristics allows for fine-tuning the coating to the necessary applications. In the examples below, we further evaluate the adhesive composition as coatings (e.g., polyethyleneimine coatings) for their bactericidal performance and leaching properties.

[0474] Example 4B: Materials and Methods

[0475] Synthesis & coating

[0476] Polyethyleneimine (bPEI, branched, Mw= 25 kDa), anhydrous ethanol, 2,5-dihydroxybenzaldehyde (2,5-DBA), 3,4-dihydroxybenzaldehyde (3,4-DBA), 3,5-dihydroxybenzaldehyde (3,5-DBA) and 3,4,5-trihydroxybenzaldehyde monohydrate (3,4,5-TBA-H2O) were procured from Sigma Aldrich.

[0477] 3,4-Dihydroxy-5-Methoxybenzaldehyde (5-OMe-3,4-DBA) purchased from TCI, Singapore. 4,5-dihydroxy-2-methoxybenzaldehyde (2-OMe-4,5-DBA) was synthesized from 6-methoxy-l,3-benzodioxole-5-carbaldehyde (purchased from Fluorochem, UK).NMR was done using 400 MHz in CD3OD). Characterized peaks: 5 10.11 (s, 1H), 7.16 (s, 1H), 6.53 (s, 1H), 3.84 (s, 3H).

[0478] Reaction was performed in anhydrous methanol, aiming for 100 mg / mL of the final product in MeOH: branched polyethyleneimine (bPEI) (18.5 pmol) was added to catechol aldehyde (726 pmol) dropwise over 5 min under stirring. After 1 hr at 25°C,the filter paper with diameter of 55 mm was dipped into the solution and dried in oven for 30 min at 37 °C. Molar ratio of bPEI : catechol aldehyde was kept constant with a grafting ratio of ~ 38.8 catechols per bPEI.

[0479] Agar plate preparation

[0480] Agar powder was procured from BD Bacto Agar (214010) and LB powder was from BD Difco LB broth, Miller (244620). Agar powder (1.8 g) and LB powder (2.5 g) was added into 500ml DI water in flask and mixed. Flask was sealed with aluminum foil and autoclaved until hot, then poured onto petri dish until bottom was fully covered. Dish was left to cool down and solidify, then sealed.

[0481] Preparation of ampicillin control disk

[0482] Ampicillin was dissolved to concentration of 1 pg / mL, then 10 L of diluted ampicillin was added onto the filter paper disk (d=0.65 cm) to achieve surface concentration 10 pg / disk, then the disk was dried in room temperature.

[0483] Measurement of inhibition area

[0484] LB broth (10 mL) was added to falcon tube with 10 pL of bacteria stock, put into shaking incubator and incubated overnight. The culture was set to 0.1 OD, measured using nanodrop, then 30 pL was added onto prepared agar plates, spreading evenly using L-shape rod. Sample disks were placed onto the plates and incubated for 24 h at 37 °C, with hourly monitoring via photographing (full data available in FIG.37A to FIG. 37D). Then the inhibition area was measured using ImageJ, using prepared disks as scale reference.

[0485] Washing procedure

[0486] For samples washed in DI water and EtOH, disks were dipped 3 times into 1 mL of solvent, then transferred onto the agar plate with present bacterial strains. Samples were monitored hourly for DI water washed set and EtOH washed samples were checked after 24 hours only. Inhibition area was measured using ImageJ, using diameter of the disk as scale reference.

[0487] Soaking procedure to determine leachate strength

[0488] Sample disks (d=0.65 cm) were soaked in 1 mL of DI water for 3 h and 24 h, After which the solvent was mixed 1 : 1 by volume with LB broth containing bacteria at 0.2 OD, measured using nanodrop resulting in mixture with initial OD of 0.1, transferred to 96 well plate and incubated for 24 h at 37 °C. After the incubation period,the well plates OD was measured using BioTek Synergy Hl Microplate reader at 600 nm.

[0489] Example 4C: Results Discussion for Examples 4A and 4B

[0490] Six formulations were tested alongside the branched polymer as control: two reference catechol aldehydes mimicking commonly used DOPA and gallic acid adhesives (bPEI-g-(3,4-DBA) and bPEI-g-(3,4,5-TBA)), two methoxy-modified catechol isomers geared toward electronic stabilization of aryl ring (bPEI-g-(2OMe-4,5-DBA) and bPEI-g-(5OMe-3,4-DBA)), and two catechol isomers with varied hydroxyl position reported previously as suitable adhesives (bPEI-g-(2,5-DBA) and bPEI-g-(3,5-DBA))[ll]. Ampicilin (10 pg) as positive control, empty filter paper disk as negative control. Branched polyethyleneimine was used to measure impact of amine groups on bactericidal effect.

[0491] Inhibition area present within 24 h

[0492] All coated disks show contact killing behaviour, preventing bacterial growth on the disk itself (FIG. 38) for both Gram-positive and Gram- negative bacteria, with smaller inhibition zones present for samples subjected to E.coli propagation. Inhibition areas present in samples are attributed to catechol aldehyde leachates debonding due to reversible Schiff base grafting from polyethyleneimine, prominent in samples V (bPEI-g-(2,5-DBA)) and VI (bPEI-g-(3,5-DBA)) for both bacterial species. Smaller zones of inhibition present for samples I (bPEI-g-(3,4,5-TBA)) and II (bPEI-g-(3,4-DBA)) in S. aureus with no inhibition zones in E.coli plates. Pure bPEI coating (sample X) exhibit slight diffusion which seems to positively impact the bacterial growth in early stages of growth.

[0493] Measured inhibition area showed water-resistant, non-leaching coatings

[0494] To determine the short-term wash resistance, the samples were dipped in distilled water before the placement on the agar bed and compared against coatings without dip-washing (FIG. 39C and FIG. 39D).

[0495] Empirical measurements confirm the presence of inhibition zone for bPEI-g-(2,5-DBA) and bPEI-g-(3,5-DBA) in both S. aureus and E. coli sets, with E.coli showing weaker response to the leachate. Other samples exhibit contact-killing behaviour, with slight leaching seen in unwashed samples.

[0496] Water treatment shows diminishing in inhibition zone, most prominent in ampicillin control, where no additional adhesive was added to prevent the removal of the active component. For the remaining catechol samples, they retain their contactkilling effect, with only notable decrease with inhibition zones for bPEI-g-(2,5-DBA) and bPEI-g-(3,5-DBA).

[0497] Ethanol wash reduced leaching, preserving antibacterial surface

[0498] As ethanol was the main solvent during the synthesis, dip-wash in it should dissolve any non-binded bioadhesive macromolecules and impede the amount of leachates observed.

[0499] All samples showing at least contact-killing behavior, with inhibition zones in samples where previously inhibition zone was perceived (FIG. 38). Comparing the inhibition areas against unwashed samples (FIG. 39C and FIG. 39D). dipping the coating in either water or ethanol diminish the inhibition area, most prominent in samples without any binding macromolecules (Ampicillin, sample C). Due to present moisture in the disk slight dilution of bacterial concentration occurred, which can be seen impacting the inhibition zone in samples I (bPEI-g-(3,4,5-TBA)), and II (bPEI-g-(3,4-DBA)) in EtOH washed samples, and III (bPEI-g-(2OMe-4,5-DBA)) and VI (bPEI-g-(3,5-DBA) in DI washed sample for S. aureus, with only VI (bPEI-g-(3,5-DBA) shown to exceed the range when washed in DI for E.coli.

[0500] Soaking shows slight inhibition of bacterial growth

[0501] To compare the strength of the possible leachates from the coated filter disks, the disks were soaked in deionised water for extended period of time, then mixed with bacteria in 1:1 ratio. Possible leachates in DI water would be then detected via the inhibition of bacterial growth.

[0502] The resulting optical density change shows inhibited growth in S. ureus in samples containing C (Ampicillin), V (bPEI-g-(2,5-DBA), VI (bPEI-g-(3,5-DBA) and X (bPEI), with no growth inhibition for other samples. Leachate data from bPEI coated sample remain inconclusive - there should be minimal leachate present due to molecular size of the polymer. For E. coli, all samples show bacterial growth, albeit growth is lower for samples that exhibit leaching on agar plates.

[0503] Example 4D: Summary Discussion for Examples 4A to 4C

[0504] In these examples, it was showed that an amine-rich polymer matrix crosslinked via Schiff-base grafted catechol aldehyde. Aspects of the crosslinking reaction could be modifed with a range on electronic densities of said catechol, where electronics were modifed through through hydroxyl position or additional functional groups. All adhesives were previously tested and showed notable lap shear performance exceeding 10 kPa in hydrated collagen strips. These examples presented the bactericidal and contact-killing performance of the coating from bPEI-g-DB A adhesives, which was retained after the wash in deionized water and anhydrous ethanol - both known as good solvents for the adhesive in its non-crosslinked state. While shown to be wash resistant after initial dipping, long-term washing cycle investigation should be performed, preferably while introducing commonly used surfactants to determine the practicality of presented coatings on textiles.

[0505] The leachates were observed only in grafted catechol isomers containing only hydroxyl groups on the aryl ring, with methoxy modified formulations (bPEI-g-(2OMe-4,5-DBA), bPEI-g-(5OMe-3,4-DBA))) showing only contact-killing performance. As the only leachates that could be prevalent in the system could be from reversal of the imine bond used to graft the catechol isomer (on account of polymer matrix being substantially of higher molar mass and not bactericidal), the resulting lack of inhibition zone for methoxy modified isomers can be attributed to the stabilization of aryl ring via electron donating nature of methoxy groups, with catechol triol showing similar, albeit weaker behaviour. The reversibility of the Schiff base in turn allows for stronger inhibition zone without notable sacrifice in coating mechanical strength. The data obtained from bPEI coated samples remains inconclusive - as coating contains exclusively macromolecule, prominent leaching is unlikely - the coating shows a slight impact on the bacterial growth, changing the local density of the growth and showing gradient unprecedented in other samples. Current working hypothesis suggests localized alkaline pH change promoting formation of reactive oxygen species on agar plates, allowing, albeit not fully inhibiting bacterial growth.

[0506] Several structures containing catechol groups in their structure and being a possible active element in their antibiotic properties. In general, polydopamine (PDA) as well as polymerized tannic acid hydrogels exhibit exhibit bactericidal behaviour, inhibiting bacterial growth. Such matrices can be further functionalized via addition ofmetal nanoparticles, or used as bed for creating repellent surfaces via chain graft. Reactive oxygen species are identified as main agent in bactericidal behaviour of dopamine-related coatings, showing practical applications as antibacterial membranes with 95% efficiency within 24h of S. aureus incubation.

[0507] A key 2,5-dihydroxybenzaldehyde derivative with antimicrobial potential is gentisaldehyde (which is another name for 2,5-dihydroxybenzaldehyde itself) and its Schiff base derivatives. Gentisaldehyde and its related compounds have shown antibacterial activity against various bacteria, including Staphylococcus aureus. Schiff base derivatives, synthesized by reacting gentisaldehyde with amines, and their metal complexes are also being explored for antibiotic development due to their broadspectrum antimicrobial and other biological activities. 2,5-dihydroxybenzaldehyde derivatives show antimicrobial activity against various pathogens, including bacteria like Staphylococcus aureus and Mycobacterium avium, and have demonstrated nematicidal effects. Derivatives, such as Schiff bases, can also exhibit promising antimicrobial and antifungal properties, though their effectiveness needs further in vivo study.

[0508] 5-viniferin contain active meta- positioned hydroxyls also present in 3,5-dihydroxybenzaldehyde. 5-viniferin and its derivatives exhibit antibiotic activity against Gram-positive bacteria, by damaging the cell membrane. Dehydro- 5-viniferin, a promising derivative, has shown significant activity against bacteria like Listeria monocytogenes, causing membrane depolarization, loss of integrity, and morphological changes.

[0509] (-)-trans-s-Viniferin exhibits promising antimicrobial and antifungal activity. It has shown potential against both Gram-positive bacteria, like methicillin-resistant Staphylococcus aureus (in combination with vancomycin), and fungi, such as the pathogens affecting grapevines. The compound works by damaging bacterial cell membranes and has also demonstrated antiviral effects against some viruses.

[0510] 2-hydroxy-5-methoxybenzaldehyde and 2-hydroxy-3 -methoxybenzaldehyde, exhibit antibiotic and anti-pathogen activity. Compounds with mixed OH and OCH3 groups exhibited variable results, i.e., in some cases OCH3 groups enhanced activity. 2-hydroxy-5-methoxybenzaldehyde exhibit antibiotic and anti-pathogen activity.

[0511] Derivatives of 3-bromo-4,5-dihydroxybenzaldehyde show antibiotic and other pathogen-related activities, including antibacterial, antifungal, and antiviral properties. These compounds, known as bromophenols, can be found in red algae and are being researched for their potential uses in medicine due to their diverse biological properties.

[0512] Six grafted catechols isomers were evaluated on their antibacterial properties against Gram-positive and Gram-negative bacteria. Two formulations showed significant leaching, with all other formulation exhibiting minimal leaching to exclusively contact-killing behaviour. The samples showed to retain its bactericidal properties after dip-washing in water and alcohol solvent, with coatings showing stronger response against S. aureus bacteria. Leaching correlates to the electronic density on the aryl ring of the catechol group. Overall, it shows a promise for imine-based catechol grafted adhesives as suitable wash-resistant antibacterial coatings, with future work expanding on the extent of long-term wash resistance with or without additions of surfactants.

[0513] Example 5A: Examples on Catechol isomers used for Moisture Initiated Adhesive Compositions and Bioadhesives of the Present Disclosure

[0514] Examples 5A to 5E demonstrate for adhesive compositions that can be moisture activiated and used as bioadhesives.

[0515] While catechol chemistry may be known, the selection of catechols applied for resins and adhesive purposes has relied almost exclusively on 1-dopamine variants. Herein five catechol isomers evaluate ortho, meta, and para dihydroxybenzene (DHB) structures on adhesion related mechanical properties, including organic / aqueous stability, gelation time, and adhesion strength on soft substrates. A model system evaluates the catechol-aldehyde isomers through Schiff base grafting to an amine rich macromolecule, branched polyethylenimine. This work evaluates how grafted-catechol isomers can be exploited to tune reactivity to both solvent and external stimuli. The formulations allow a range of sensitivity, from designs that observe gelation time within minutes of water exposure, to water-stable formulations that can be cured through via voltage stimulation.

[0516] Catechol-based bioadhesives require oxidation to yield the quinone intermediates, which crosslinks with amines and thiols to form covalent bonds. Oxidation occurs through alkaline pH, two-component external oxidants (such asNaIO4, H2O2, etc.) or other redox additives that may have toxic profiles. These limit their application as tissue adhesives. The most common quinone precursors exploit 1-dopamine (L-DOPA). Investigations mapping catechol isomers, such as ortho, meta, and para hydroxyl groups remain a research gap.

[0517] Traditionally, catechol variants allow facile grafting on amine terminated polymers. The click-chemistry approach exploits aldehydes to rapidly graft catechols through a Schiff base. A one-component glue results in the form of a catechol-g-macromolecule. Placement in aqueous medium provides the reactive quinone intermediate, although the mechanism remains speculative. One advantage of this: organic formulations remain stable, while curing ensues upon contact with aqueous substrates or environment. Amine rich macromolecules, e.g. branched polyethyleneimine (bPEI), accelerates formation of the quinone intermediate.

[0518] Addressing the two research gaps above, expanded catechol structures need assessment. Hydroxyl-rich catechols such as gallic acid enhance crosslinking kinetics and related material properties. Electron withdrawing nitro group in dopamine accelerates the curing rate and mechanical properties of PEG-DOPA adhesive. This design allows for electronically modified catechols with improved curing rates and adhesive properties. This work investigates a range of bPEI-g-catechol isomers and correlates the electronics to adhesive properties, including gelation time, shear modulus, yield stress, and lap-shear adhesion strength on hydrated collagen substrates. FIG. 44 summarizes known catechol crosslinking mechanisms relevant for adhesives. FIG. 43 displays the selected catechol isomers.

[0519] Example 5B: Materials and Methods

[0520] Polyethyleneamine (branched, Mn= 105, Mw= 25 kDa), solvents (anhydrous methanol, anhydrous ethanol), n-propylamine, 2,3 -dihydroxybenzaldehyde (2,3-DBA), 2,4-dihydroxybenzaldehyde (2,4-DBA), 2,5-dihydroxybenzaldehyde (2,5-DBA), 3,4-dihydroxybenzaldehyde (3,4-DBA), 3,5-dihydroxybenzaldehyde (3,5-DBA) and 3,4,5-trihydroxybenzaldehyde monohydrate (3,4,5-TBA H2O) were procured from Sigma Aldrich, Singapore. Phosphate buffered saline (PBS) used was procured from Gibco, Singapore. Disposable Zensor® electrodes used as support for parallel plate rheometry were procured from Zensor R&D Company, Taipei, Taiwan.

[0521] Synthesis of catechol-grafted polyethyleneimine

[0522] Branched polyethyleneimine (bPEI) (400 mg, 0.016 mmol in MeOH) was added to catechol aldehyde (0.621 mmol in MeOH) dropwise over 5 min under stirring. After 1 h at 25°C, residue was dried in vacuo. For 3,4,5-TBA H2O, the compound was dissolved in anhydrous methanol with molecular sieves. To achieve the grafting ratio of 20% to the primary amines, the molar concentrations of bPEI : DBA were set to 1 : 38.8. The final product was evaluated via NMR spectroscopy proton spectra (Bruker 400MHz dissolved in CD3OD), characterized peaks: 8 10.11 (s, 1H), 7.16 (s, 1H), 6.53 (s, 1H), 3.84 (s, 3H).

[0523] Cyclic Voltammetry & DFT calculations

[0524] Catechol-Schiff base was prepared with n-propylamine in place of bPEI (NPA-g-DBA) to prevent non-specific fouling of electrode surfaces (1:1.05 eq. of n-propylamine: catechol). The catechol-n-propylamine Schiff-base (366.5 mg) was dissolved in PBS (20 mL, cone. 0.1 M) and degassed with argon. The CV measurements were performed at the scan rate of 100 mV-s1with scans from 0 to -1.2V, and then 1.2 V. Ag / AgCl electrode served as reference electrode, glassy carbon as a working electrode and Pt as a counter electrode.

[0525] DFT simulation had structures of n-ethylamine (NEA-g-(DBA)) and brought to oxidized state with the b31yp / 6-311g(d,p) with GB3BJ method. Additionally, to test the previously stated Schiff-base / quinone redox hypothesis, the Cl atom energy charge was monitored.

[0526] NMR studies of catechol grafting and oxidation

[0527] Grafting reaction was performed in situ by mixing catechol aldehyde and bPEI in NMR tube before the measurement of 1H spectra (Bruker 400MHz dissolved in CD3OD), example seen on FIG. 45. The reaction was considered complete when no aldehyde peak was first observed, but due to catechol reactivity no grafting efficiency could be calculated. The effect of PBS / D2O addition on crosslinking kinetics of bPEI-g-(DBA)2o was evaluated by adding 10 vol. % of PBS / D2O into the NMR tube containing the reaction mixture, final solvent being 10% PBS 90% CD3OD. The first NMR was recorded immediately after PBS / D2O addition and subsequently at every 30 min interval up to 3.5 h. For comparison between the compounds, sum integration of protons on the aryl ring (6.25-7.50 ppm) was taken alongside with integration of one proton on the Schiff base (7.75-8.25 ppm).

[0528] UV-vis studies of catechol grafting and oxidation

[0529] UV-vis studies were performed using the Shimadzu UV-2700 spectrometer using thin layer quartz Helma cuvette with a pathlength of 0.5 mm, using methanol as a solvent. bPEI-g-(DBA)2o (2.28- 10’2mmol / L) was dissolved in 10 ml of MeOH within 2 min of, then -150 pL of this solution was transferred into the cuvette. UV-vis spectra (225-450 nm, with focus at peak range 320-350 nm) were recorded at every 10 min interval for a period of 2 h (FIG. 46).

[0530] To measure the effect of PBS on the crosslinking kinetics of bPEI-g-(DB A)io, 10 vol. % PBS was added into the 9 ml of solution (final concentration 2.28- 10’2mmol / L) after 15 min and UV-vis spectra were recorded as mentioned above (FIG. 47).

[0531] The effect of oxygen / air on the crosslinking kinetics of bPEI-g-(DBA)2o was evaluated by bubbling air and oxygen into the solution using a Helma cuvette with pathlength of 10 mm. The solution concentration used for this UV-vis study was around 1 / 20 as compared to the previous (standing at 9.8- 10’4mmol / L) to ensure comparable exposure to UV-vis light for all the studies. The gases (oxygen / air) were bubbled for -5 s at every 15 min interval for a period of 2 h (FIG. 48). Referential data for oxygen concentration was measured using Metrohm 914 pH / DO / Conductometer with DO probe, measuring oxygen concentration in air and in before and after 1 h of degassing using argon (FIG. 49).

[0532] Parallel plate rheometry of grafted catechols in aqueous environment

[0533] Parallel plate rheometry was done using Anton Paar MC102 Rheometer using ceramic probe and carbon disposable Zensor® electrode (as base), using 30 pL of adhesive at a gap of 0.3 mm; The bPELg-(DBA)2o in PBS (30 wt. %) was pipetted onto the Zensor® electrode, then rheological studies were performed: apparent viscosity measurement at the steady shear rate of 10 s-1for 30 s; storage and loss modulus measurement at the angular frequency of 6.28 rad-s-1over 30 min; amplitude sweep from 1-1000% to determine the shear stress at failure.

[0534] For samples subjected to constant voltage, current was applied via Iviumsoft potentiostat by connecting Zensor® electrode using crocodile clips, attaching working and counter electrode. Voltage was applied from 2 min mark of the constant oscillation experiment for 28 min (until the end of constant oscillation).

[0535] Lap shear adhesion of bPEI-g-(DBA)2o on hydrated collagen substrates

[0536] Lap shear was performed on hydrated collagen substrates made from 36 mm collagen casings (Shandong Haios Biotechnology Co., Ltd., China). Collagen strips were cut into 2 cm x 6 cm, hydrated for 1 min in PBS on each side and surface dried with tissue paper. Strips were glued over 2 x 2 cm area by applying 35 p L of bPELg-(DBA)io in solvent (PBS, EtOH) (30 wt. %). After 30 min air exposure, lap shear test was performed. The tensile tester (Chatillon Force Measurement Products, USA) was equipped with a 50N load cell with an elongation rate of 10 mm / min.

[0537] Measurements for toughness estimation

[0538] To determine the thickness of the applied catechol grafted layer, a (bPEI-g-(2,5-DBA)2O) solution was applied onto collagen strip, analogous to lap shear sample preparation. The sample was slow dried by pressing the sample in between microscope slides and kept in vacuum sealed desiccator for a month. The sample was then opened, sliced with a razor to stripes of 2mm thickness, then the cross section was examined via Olympus BX5 optical microscope with 50x0.75 lens. Due to height differences in the sample, focus stacking of pictures was done using via Infinity Analyze software, after which three measurements of the thickness of the adhesive layer was done.

[0539] Example 5C: Characterisation Observations

[0540] UV-vis spectra

[0541] During the formation of Schiff-base (bPELg-(DBA)2o) and subsequent polymerization catalyzed by the moisture and oxygen in air the changes in the UV-vis spectra of catechol aldehydes observed; the UV-vis peaks at 280 nm (BA), 320 nm (3,4-DBA), 350 nm (2,3-DBA), and 370 nm (2,5-DBA) are attributed to aromatic. As the grafting reaction progresses, the aldehyde peak disappears and new peak appears at 270-280 nm (bPEI-g-(BA)20), 350 nm (bPEI-g-(3,4-DBA)20), 320-340 nm (bPEI-g-(2,3-DBA)2O), and 350 nm (bPELg-(2,5-DBA)2o), indicating the formation of Schiff-base. Appearance of peak at 400-440 nm, is attributed to the formation of reactive quinone / semiquinone intermediate. However, the quinone quickly crosslinks by reacting with the nearby nucleophilic amine, rearomatizing the catechol ring, evidenced by the gradual decrease of 400-440 nm peak intensity and increase in peak intensity at 260 nm.

[0542] Cyclic Voltammetry & Density Functional Theory

[0543] The lowest oxidation energy change for the aromatic ring as well as the Cl atom of n-MEA-g-(2,3-DB A) is explained by its similar tautomeric structures as that of n-MEA-g-(2,5-DBA) andn-MEA-g-(3,4-DBA). The tautomeric structure of n-MEA-g-(2,4-DBA) shows that only one -OH can oxidise to ketone easily while the oxidation of the other -OH demands higher energy output, as observed in DFT calculation. There is no tautomeric structure possible for n-MEA-g-(3,5-DBA) for -OH oxidation to the corresponding ketone, which is reflected in the highest oxidation energy change in its DFT calculation.

[0544] Example 5D: Results Discussion

[0545] Five catechol isomers (see FIG. 43) evaluate if diol / triol benzaldehydes and their electronic density affect redox behavior, kinetics (e.g. gelation time) and adhesive performance (e.g., lap shear strength). For application with bioadhesion and tissue fixation, we seek to identify formulations stable in anhydrous solvents but have gelation times of < 20 min when exposed to aqueous substrates, with lap shear adhesion strength exceeding 1 kg.f.cm’2— like commercial skin tapes. This investigation exploits a model system of bPEI-g-catechols, where catechol grafting occurs through a one-pot click chemistry reaction between quinone precursors of benzaldehyde (key adhesive intermediate) and amine-functionalized macromolecule. bPEI (Mw~ 25 kDa) serves as a model hyperbranched macromolecule. Gelation time (where G’7G’ = 1) compares liquid-to-solid kinetics and intermolecular crosslinking, as measured via oscillatory shear rheometry. Two additional aldehydes benchmark minimum and maximum performance that all samples should fall in between; non-reactive benzaldehyde (BA) and instable gallic acid (TBA, reactive triol). BA serves as a negative control with nocrosslinking capability (no quinone formation) and TBA displays as a positive crosslinking formulation with instant quinone formation, as gelation occurs before rheometer / probe positioning. Cyclic voltammetry (CV), UV-VIS absorption correlates chemical kinetics to the lap-shear adhesion. Density-functional theory (DFT) calculates initial partial charge electronic densities and allows hypothesis testing to empirical oxidation potential. Previous investigations attributed spontaneous crosslinking to tautomeric structures, but an updated hypothesis suggests oxygen stimuli may be responsible for quinone formation, with water presenting suitable reaction environment.Support for this hypothesis exploits absorbance-based chemical kinetics with respect to anhydrous, anaerobic samples exposed to water, oxygen, and combination thereof.

[0546] Oxidation potential rankings display the 3,4-catechol isomer near the highest.

[0547] Oxidation potential compares the electrochemical energy needed for oxidation of the catechol to quinone (presumed). Both empirical and simulation methods give quantitative assessments for structure property correlations. Cyclic voltammetry (CV) evaluates the catechol to quinone lability after Schiff-base formation with n-propylamine (NPA). NPA replaces bPEI to prevent artefacts from dendrimer mobility or cationic surface fouling. Oxidation potentials may predict gelation time and shelf stability. Higher Eoxwould prevent spontaneous transition to quinones by atmospheric gases but may lead to longer gelation times (FIG. 50A). FIG. 50B and FIG. 50C summarizes the oxidation potentials, Eox, determined by cyclic voltammetry (CV) and DFT calculation, respectively. For analysis, Eoxis ordered via gelation time. Representative CV data can be found in FIG. 51A to FIG. 5 IF.

[0548] DFT calculations estimate Eoxwith good agreement of the CV empirical data. The DFT-derived AE correlates to the oxidation potential from grafted catechol to quinone (FIG. 50B). Charge differences on Cl near the Schiff-base suggest electron deficiency, where electron attracting aldehyde substituents display the opposite behaviour. This may explain the stability of the Schiff base (see next section) and prevention of reduction in aqueous environments, but does not support the original tautomer theory proposed in earlier investigations.

[0549] All DBA samples display chemically irreversible oxidation peaks, suggesting the formation of quinone like intermediates. Isomers with the lowest oxidation potentials had para hydroxy -Schiff base positions. Meta isomers display a larger Eoxcompared to ortho diols and appear to be more reactive. Ordering the catechol isomers by complex modulus (G*) or faster gelation time (see rheometry data below) show no clear dependence on Eoxor maximum current.

[0550] Catechol stability in anhydrous MeOH but depletes upon aqueous exposure while Schiff base remains stable under same conditions.

[0551] Proton NMR evaluates the dynamics of Schiff base formation and stability via integration of the Schiff base proton and non-exchangeable aryl protons. Dynamicscaused by grafting, oxidation, or both are evaluated with signature Schiff-base or aromatic peaks as shown in FIG. 52A. These peaks are further analysed before and after addition of deuterated saline. Numerical values of proton integration available in FIG.53 and FIG. 54.

[0552] Up to 3.5 h, no Schiff base reduction is observed. Aromatic protons diminish only in aqueous samples of in bPEI-g-(3,5-DBA)2o and bPEI-g-(3,4,5-TBA)2o, displayed in FIG. 52B. This suggests meta-hydroxy / Schiff base positions appear more reactive. bPEI-g-(3,5-DBA)2o and bPEI-g-(2,5-DBA)2o formulations is subjected to pure oxygen bubbling for 1 h followed by proton NMR analysis. Schiff base and aryl ring protons diminish, suggesting accelerated reactivity with oxygen (FIG. 55A to FIG.55C). Depletion of aryl rings implies Michael addition and inter-catechol crosslinking.

[0553] bPEI-g-(2,5-DBA)2o and bPEI-g-(3,5-DBA)2o display gelation within 10 minutes

[0554] Rheometry evaluates time-dependent mechanical properties of viscosity, modulus, gelation time, and shear strength to correlate with chemical data above. Benzaldehyde serves as a non-reactive negative control. The lack of quinone isolates reversible cohesive forces such as hydrogen bonding, van der Waals, dipole, electrostatic, or combination thereof. A reduced form of gallic acid, gallic aldehyde, serves as a positive control known for spontaneous oxidation and crosslinking. The five catechol isomers are benchmarked to these controls with respect to apparent viscosity, complex modulus, gelation time, and shear strength (FIG. 56A to 56D). Representative graphs from constant oscillation measurements are present in FIG. 57A to FIG. 57F, while data from amplitude sweep is collated in FIG. 58 A to FIG. 58L.

[0555] The triol positive control (gallic aldehyde) uncontrolled reactivity displays an instant rise in viscosity > 10 Pa.s. Catechol isomers compounds display a range of moduli (G*) after 30 minutes exposure to PBS. Gelation times, ranked from highest to lowest: bPEI-g-(3,4-DBA)2o, bPEI-g-(3,5-DBA)20and bPEI-g-(2,5-DBA)20is 20, 9, and 0.5 min (FIG. 56C). bPEI-g-(BA)20, bPEI-g-(2,3-DBA)20and bPEI-g-(2,4-DBA)20has no measurable cross-overpoints within 30 min. Positive control (triol) sees gelation time faster than time take for placement on rheometer. Visual observations display a non-flowable gel within 15 sec of water addition. Shear strength is highest in bPEI-g-(3,5-DBA)2O (excluding positive control) out of all catechol isomers, which paired withlower moduli suggests gel-like matrix with lower crosslinking density (FIG. 56D). Meanwhile, bPEI-g-(2,5-DBA)2o exhibits similar shear strength while retaining higher storage modulus and shorter gelation time. Gelation time comparison is available in FIG. 59.

[0556] The best performing samples are selectred for lap shear adhesion on hydrated collagen substrates. Non-reactive compounds are screened for voltage-dependent crosslinking via electrorheometry in FIG. 60A to FIG. 60D.

[0557] bPEI-g-(2,3-DB A)2O shows higher complex modulus values when subjected to negative voltage (FIG. 60D). The gelation time is shown to be over 18 and 12 min respectively for -2 and -3 V, while no gelation occurred for - IV and 3V. This rules out the possible water electrolysis impact, which starts to occur at around 1.23 V. Lack of recorded reactivity may be due to placement of screen-printed carbon electrodes on the electrode itself, as higher shears are observed alongside probe radius. Comparison between applied voltage and time of gelation can be found in FIG. 61.

[0558] Samples in anhydrous solvent yield better shear strength on hydrated collagen substrates

[0559] Mechanical performance is evaluated under two dispersants; neat sample immediately dissolved in PBS buffer and shelf-stable samples stored in anhydrous ethanol (FIG. 62C). Ethanol serves as a more biocompatible candidate versus methanol, as the latter’s metabolites are known to cause blindness. We hypothesize substrate moisture, air exposure, or combination thereof is sufficient for quinone formation when applied in a water miscible solvent. Raw data is available in Supplementary Information (FIG. 63A to FIG. 63F, FIG. 64A to FIG. 64F).

[0560] Failures observed are the following: cohesive failure (bPEI-g-(DBA)2o, adherend failure (collagen film fracture) or mixture of both (when collagen starts tearing, but cohesive ccurs first). The adherend failure occurs from 7.8 N of load alongside cohesive failures, with highest loads for it being 9.3 N and 10.6 N respectively for bPEI-g-(2,4-DBA)2o, translating to 23.3 kPa and 26.7 kPa of shear strength, respectively. One replicate of bPEI-g-(2,5-DBA)2o is not included due to clamp damage (n=4). Example failures are provided in FIG. 65.

[0561] Usage of anaerobic solvent shows that bPEI-g-(DBA)2o formulations can be locally activated by air and moisture present on applied area, as there is no significantdifference between anaerobic, anhydrous solvent, and aqueous solvent. EtOH additionally mixing with moisture creating azeotrope and penetrating the collagen sample increases the local concentration of bPEI-g-(DBA)2o, allowing for better adhesion to the sample, as highest shear strength results were exclusively in more volatile solvent. That behaviour does explain the unusually high performance of bPEI-g-(2,4-DBA)2o, which did not show gelation during parallel plate rheometry. Overall, bPEI-g-(2,4-DBA)2o, bPEI-g-(3,4-DBA)2o, bPEI-g-(3,5-DBA)2o, bPEI-g-(2,5-DBA)20and bPEI-g-(3,4,5-TBA)2o samples are consistently higher when utilizing alcohol solvent for the adhesive formulations, beating Evicell™ performance under the same conditions. The specific differences between the catechol isomers can be attributed to formation of covalent bonds such as Michael adducts and new Schiff base formations through oxidized catechol groups, both which depend on the amine and thiol surface density.

[0562] Assuming uniform layer thickness of 368 pm (estimated via optical microscope measurement - FIG. 66), toughness was calculated via integration of area under the curve, showing significant improvement for bPEI-g-(2,4-DBA)2o and bPEI-g-(3,4,5-TBA)2o (data is available in FIG. 67).

[0563] Absorbance kinetics correlates to oxygen exposure.

[0564] Determining the rate of impact on the grafted catechol isomer oxidation, addition of PBS to anhydrous solution causes absorbance increase over time. Addition of PBS accelerates this crosslinking process, manifesting as rise in absorbance in 320-350 nm range. Quinone being highly reactive intermediate is expected to react instantly with amines as soon as it forms, not appearing on most of the spectra in its predicted range (~ 440 nm) showing broad banding instead.

[0565] Spectra were normalized against spectra before contact with 10 vol. % PBS as oxidant, showing rise in relative absorbance of grafted catechol isomers, showcased in FIG. 68 A to FIG. 68D. Out of four tested compounds, only bPEI-g-(2,5-DBA)2o and bPEI-g-(3,5-DBA)2o shows to be constantly rising over the period of the experiment, while bPEI-g-(2,4-DBA)2o shows no increase and bPEI-g-(3,4-DBA)2o exhibit saturation after 1.5 h of adding the PBS. The compounds shown to have constant increase of absorbance were moved to tests with air and oxygen bubbling, shown in FIG. 68B and FIG. 68C for bPEI-g-(2,5-DBA)20and bPEI-g-(3,5-DBA)20respectively.Whole UV-vis spectra are present in FIG. 48 A to FIG. 48D, whereas zero-order kinetic values are present in FIG. 69.

[0566] For bPEI-g-(2,5-DBA)2o, oxidation rate by PBS is on par with air oxidation -measuring the oxygen content of the PBS solution showed the concentration like that of air (FIG. 49). Highest oxidation rate was achieved with pure oxygen gas, which overall show trend between oxygen concentration in medium and oxidation kinetics of this grafted catechol isomer. In contrast, bPEI-g-(3,5-DBA)2o shows the highest oxidation rate for air bubbled formulation. No correlation can be seen between oxygen concentration and the oxidation kinetics.

[0567] Example 5E: Discussion for Examples 5 A to 5D

[0568] Herein five catechol isomers were screened towards developing one-component bioadhesives. Two formulations observed spontaneous conversion (< 10 min) to quinones upon exposure to aqueous solvents, which were based on 3,5-DBA and 3,4,5-DBA. The 2,3-DBA isomer was inert to both aqueous and oxygen exposure but could be stimulated by voltage. The development found 5 out of 6 hydroxybenzene tested activates upon aqueous exposure without the addition of synthetic oxidants. The five dihydroxybenzene aldehydes (DBA) have unique electronic densities and allow facile coupling to macromolecules via Schiff-base click chemistry. Grafting is thus simplified to a one-step mixing reaction with numerous advantages; energy efficient (no heat required), stoichiometric conversion within 30 min, and absence of purification procedures that plague carbodiimide protocols required for 1-dopamine. This accelerates hypothesis testing, exemplified herein with seven aryl-aldehyde structures. Simplifying organic chemistry allows higher throughput of assessing precursors hypothesized to form benzoquinones upon oxidation.

[0569] Evaluation of oxidation potential and related DFT calculations validate that each catechol isomer had unique value and electronic density, supporting the hypothesis that the isomer can tune reactivity in the formation of the benzoquinone. A connection between the oxidation potential may explain the degrees of stimuli response. By mapping oxidation potential of catechols, novel designs of oxygen-activated glues can then be designed with a range of curing profiles. Benzoquinone serves as the active crosslinker for cohesive covalent crosslinking — which converts the liquid glues into networks of solid resins. NMR analysis found all samples were relatively stable inmethanol under anaerobic conditions. Two formulations observed spontaneous conversion (< 10 min) to quinones upon exposure to aqueous solvents or oxygen, which were based on 3,5-DBA and 3,4,5-DBA. Stability of the Schiff base grafting was observed in both organic and aqueous solvents, but saturated oxygen observed some oxidation over a 1 hr timeframe. Combined observations of Schiff base stability and aromatic oxidation (e.g. quinone formation) does not support the tautomer mechanism previously published. The current results suggest catechol oxidation via dissolved oxygen as the reductant (O2 + 2H+2H2O). Measurements of dissolved oxygen were found to be ~5x more in saline, than degassed methanol (FIG. 49). This explains the stability in degassed organic alcohols compared to water exposed formulations.

[0570] Catechol isomers had a range of reactivity, with the 2,3-DBA isomer formulation inert to both aqueous and oxygen exposure. This formulation was susceptible to voltage-activated crosslinking with similar moduli as the previously tested 3,4-DBA. Isomers 2,5-DBA, 3,5-DBA and 3,4,5-TBA display reactivity and shear strength surpassing the 3,4-DBA dopamine variant, suggesting that the biological catechol variant may not be optimized for synthetic bioadhesives.

[0571] Additionally, bPEI-g-(3,5-DBA)2o was one of the better performing catechol when grafted, despite the double -meta position of hydroxyls preventing the tautomerization of the structure - suggesting another, oxygen reacting mechanism, contradicting previous Schiff base-quinone redox pairing hypothesis. Oxygen being the driving factor of catechol oxidation is often cited, but the exact mechanism was not investigated further. Additionally, reversible bonds (such as o-bonds or hydrogen bonding between -OH groups on the aryl ring) may be one of the driving forces for this specific compound.

[0572] Lap shear performance indicates that substrate moisture may be enough to activate catechol-grafted formulations. In all tested formulations, shear strengths for anaerobic ethanol exceeded saline formulations. Some samples exceeded the tensile strength of the substrate (shear stress > 20 kPa), thus this may not give a true reflection of shear strength. Moisture and inherent dissolved oxygen serves as a chemical stimulus while the miscible alcohol solvent may assist entanglement on the collagen surface, explaining why shear strength was better with EtOH. Modulus of toughness hadconsistent values across both linear lap shear vs. rotational amplitude sweep rheometry despite different substrates.

[0573] Absorbance dynamics, measured under the forced exposure to air and oxygen provide further chemical support for oxygen as the primary stimulus in catechol oxidation. Further factors are likely involved in stimulation, which will need further evaluation with respect to oxygen diffusion. Recent studies on dopamine variants support catechols isomers may allow optimization and better design of biomimetic adhesives.

[0574] Recent research investigations have evaluated similar structures of bPEI-g-(3,4-DBA) for adhesion on both natural and synthetic materials of wood, aluminum, steel, and epoxy. However, this requires processing at 80°C for 48 hours, unlike the ambient activation described herein. Aiming for environmentally friendly construction adhesive, they achieved MPa shear strengths through solid resins by adjusting parameters of DBA ratios and curing time. Others have modified cellulose with 3,4-DBA and observed adhesion strength of 140 kPa on porcine skin and over 4 MPa on wood, but also require extended curing times to even 72 h. Similar macromolecules of chitosan and hyaluronic acid modified DBA show up to 0.9 kPa after 4h of curing. The lower strength may be attributed to pH differences between formulations.

[0575] In summary, five catechol isomers evaluate structure dependence on adhesive properties. Two formulations observed spontaneous conversion (< 10 min) to quinones upon exposure to aqueous solvents, with gelation times within minutes. The isomer was inert to both aqueous and oxygen exposure and could be exploited for voltage-stimulated activation. Most of the isomers tested activated upon water exposure, attributed to dissolved oxygen. This suggests that catechols designed through L-DOPA mimics may be tuned for stability through alternatives isomer structures. Reactivity ranges from reactive catechols that crosslinked in minutes, versus others that were stable to water, alkanol, ambient oxygen, or combination thereof. Overall, adhesive performance can be improved and suggests electron donating and electron withdrawing groups may allow addition control of adhesive properties such as shear strength, potlife, gelation time and shelf stability.

[0576] Example 6A: Examples of Adhesive Compositions based on Triols (e.u.. Triol Benzaldehydes)

[0577] Traditional examples may focus on diol benzaldehydes grafted onto amine-rich branched polymer via spontaneously forming benzyl-imine linkages. Spontaneous grafting occurs in both aqueous solvents, such as saline, or anaerobic, anhydrious methanol. Imine grafting in anaerobic, anhydrous solvents yield shelf stable formulation that cure within minutes when subjected to oxygen and moisture.

[0578] Gallic acid derived compounds are phenolic compound found tea leaves, tree bark, and grapes, and other plant derived foodstuffs. Gallic acid is known for its application as an antioxidant, anti-inflammatory, and antimicrobial properties. Currently it is being explored for food preservation, cosmetics, and medicine. Chemically, it's a trihydroxybenzoic acid (C7H6O5) and is generally recognized as safe (GRAS) by the FDA, appearing as a white solid that often browns upon exposure to air. The latter properties prevent shelf stability as a key intermediate due to spontaneous formation of polyphenols like ellagic acid. A more stable variant are the four gallic acid derivatives of triol benzaldehydes (four isomers possible), which can also spontaneously graft to amine rich macromolecules and polymers. The macromolecules with grafted triol benzyl-imine have higher crosslinking reactivity (compared to diol benzyl-imine) that yields a higher degree of crosslinking. However, they have lower shelf stability, even in anaerobic, anhydrous solvents. A method has been found to impart shelf stability in anaerobic, anhydrous solvents. A oxygen-resistant, water susceptible protective group prevents spontaneous crosslinking until exposure to air and moisture.

[0579] Example 6B: Preparation of Adhesive Composition based on Example 6A using on triol benzaldehydes

[0580] Branched polyethyleneimine (bPEI, 0.032 mmol) was dissolved in degassed methanol (15 ml) in round bottom flask, while triol aldehyde (1.30 mmol) was dissolved in another vial of degassed methanol (8 ml). The two compounds were mixed by slowly dropping the triol solution into magnetically stirred bPEI solution. After fully mixing, the mixture was left stirring for 1 h, after which the magnetic stirrer was taken out, washed, and the solvent was evaporated in vacuo on RotaVap for 2 h. Amount of methanol residue was registered, dependent on the compound in question, but within 10-30 wt. % MeOH.

[0581] Example 6C: Rheometry Observes Increase in Complex Modulus (G*) upon Air and / or Water Exposure for Example 6B

[0582] To perform the parallel plate rheometry, grafted compound was mixed with PBS, achieving concentration of 10-30 wt. %. Resulting mixtures above 20% however were paste-like and resembled Bingham Plastics, but could not be pipetted. Therefore, samples were applied using spatula, and excess was removed plate. Rheometry was performed by applying 28 pL on the Zensor electrode with ceramic PP10_C probe, with 0.3 mm gap, after 30 s pre-shearing at steady rate of 10 s’1, with angular frequency of oscillating test being 6.28 rad / s over 30 min to observe the changes in G’ and G”. Results show crosslinking of macromolecules upon exposure to ambient air and controlled humidity 40-50% relative humidity. As shown in FIG. 71A and FIG. 7 IB.

[0583] Example 6D: Adhesion on Wet Surfaces of Natural and Synthetic Materials for Example 6B

[0584] Lap shear adhesion evaluated adhesives diluted in anhydrous, anaerobic MeOH (10-30 wt% solute) and applying it on hydrated substrates of collagen, glass, or wood strips of 2 x 6 cm. Substrates were pretreated by submerging in saline for 1 min, then excess removed by via absorbent tissue. Adhesive (35 pL or 30 mg) was applied on 2 x 2 cm area, then two strips were pressed together for 30 min using IN paper clips. Some samples were applied by spatula in FIG. 71C. In next experiment (FIG. 7 ID), the sample was applied on substrates via spray coatings badhesive mixture dissolved in MeOH (10 wt%).

[0585] Based on FIG. 7 ID, bPEI-(2,3,4-TBA) performed significantly better than bPEI-g-(3,4,5-TBA) on glass and wood. Glass and wood have different surface characteristics; glass is smooth and inorganic, whereas wood is porous and has a complex organic structure. Compared to bPEI-g-(3,4,5-TBA), the particular configuration of functional groups in bPEI-g-(2,3,4-TBA) may improve bonding or coating performance on certain surfaces.

[0586] It can be noted that bPEI-(2,3,4-TBA) performed well on wood, with a shear strength at break up to almost 300 kPa, which is close to the 100N force cell limit. This can be due to its unique chemical interactions and mechanical properties. The adhesive forms multiple hydrogen bonds with the hydroxyl groups present in wood, creating a robust 3D network that enhances both adhesion and cohesion.

[0588] To improve shelf stability but retain water activation, 1:1 molar ratio of triol benzaldehyde to the trimethyl borate (TMB) allows formation of a borate ester. The latter prevent spontaneous oxidation, but is easily deprotected upon exposure to water. Triol benzaldehyde was dissolved in anhydrous methanol (4-5 mL, depending on sample), then TMB was added dropwise until the mass of the desired molar ratio was reached. Solution was stirred and left for 24h in the 4 deg C fridge. As the TMB deprotects in water, 3,4,5-TBA monohydrate was dissolved pre-emptively and kept with 3 molecular sieves (20A, for water capture) beforehand, kept in fridge for the same amount of time. After protection, the compounds were added to round flask containing branched polyethyleneimine (with molar ratio of 38.8 : 1 to achieve 20% primary amine graft) dissolved in 6 ml methanol under constant magnetic stir. The reaction took Ih, after which the methanol was removed in vacuo. Resulting compounds were stored under nitrogen until needed.

[0589] Example 6F: Rheometry Observes Lower Visosity and Increase in Complex modulus (G*) upon Air and / or Water Exposure of TMB Borate Esters

[0590] To test the impact of the TMB protection on the adhesive, synthesized adhesives were dissolved in PBS to achieve 30 wt.% concentration, then tested on the parallel plate rheometer with Zensor electrode and 10mm ceramic probe as testing surfaces. Samples were dissolved within 10-20 min, then tested by applying 28 pL on the electrode, then pre-shearing for 30 s at steady rate of 10 s’1, with angular frequency of oscillating test being 6.28 rad / s over 30 min to observe the changes in G’ and G”, after which 5 min of amplitude sweep 1-100% was performed.

[0591] Protection of the hydroxyl groups lowered apparent viscosity, allowing for easier pumping and handling. Borate ester of the bPEI grafted 2,3,4-TBA improved crosslinking performance and shelf stability, shown by the increase in gelation time. Shelf stability of grafted 3,4,5-TBA improved but with a retarded storage modulus over time (FIG. 72).

[0592] Example 6G: Adhesion on Wet Surfaces of Borate Ester Protected Adhesive Compositions

[0593] To test the strength of the borate ester protected adhesives, the synthesized compounds were dissolved in MeOH and in PBS to 10 wt.% concentration. The resulting adhesives (105 pl) was then applied on the wooden couponsthen pressed together with IN paper clips for 30 minutes. After 30 minutes, the samples were tested on MTS42 tensile tester with speed of 10 mm / min and mechanical grips. TMB protection did not interfere with the grafted 2,3,4-TBA adhesive performance. Protected borate grafted 3,4,5-TBA remains stable at ambient temperature and requires long adhesive, heated substrates, or both.

[0594] Example 7A: Water Activated Adhesive Composition and Antiviral

[0595] Examples 7A to 7D demonstrate the present adhesive composition as a water activated version, and / or usable as an antiviral coating.

[0596] Synthesis of bPEI-g-(3,4,5-TBA)20, bPEI-g-(2,4,5-TBA)20and bPEI-g- (2,3,4-TBA)2ofor lap shear on wooden coupons - Lap Shear sample preparation

[0597] Lap shear of prepared samples on Mechanical Tester MTS C42 with 500N load cell, with mechanical grips to avoid slip.

[0598] Both trihydroxylbenzaldehyde (TBA) and branched polyethyleneamine (bPEI) were dissolved in methanol (analytical grade) in separate vials, then mixed to achieve concentration of 10 wt.% allowing for 1 h reaction to form grafted compound (see FIG. 74 and FIG. 75). In the mixing process, it should be noted the aldehyde may clump together at this concentration, resulting in lower grafting ratio.

[0599] The resulting compound was then applied on the wooden coupons: using 105 pl (3 times the volume used for collagen samples, on account of lower concentration) was applied onto 17 x 20 mm area of the wooden coupon, then pressed together with clips to cured the adhesive for 30 minutes.

[0600] After 30 minutes, the samples were tested on MTS42 tensile tester with speed of 10 mm / min and mechanical grips - previous attempts used pneumatic grips with max strength of 55 psi, on which slips of sample occurred.

[0601] Additionally, same procedure was applied to 5 sets of cut and filed aluminium strips: 105 pl of bPEI-g-(2,4,5-TBA) was applied to 20 x 20 mm area, and the sides were fastened with clips.

[0602] Example 7B: Discussion for Result based on Example 7A

[0603] Resulting samples were correctly tested - no slips occurred for all 15 wooden samples (see FIG. 76 to FIG. 82 and FIG. 84).

[0604] The adhesive cured poorly on the aluminum strips, with leakages observed instead compared to the ones on wooden samples (see FIG. 83).

[0605] Usage of mechanical grips prevented problems with slippage. When compared against the data obtained, the tested triol aldehydes showed mechanical performance within range of 0.5-1 MPa, with no statistical difference between grafted 3,4,5-TBA and 2,3,4-TBA. Tests also included glass substrate, on which grafted 2,3,4-TBA showed to have equally good or better performance. Accordingly, the wooden and glass samples demonstrate the adhesives’ effectiveness on organic and ceramic (inorganic and / or non-metal) substrates, respectively. For applications that require a different extent of adhesiveness as demonstrated via the present aluminum sample (a nonlimiting example of inorganic metal substrate), the present adhesive compositions may still be suitable.

[0606] Example 7C: Another Example of Water Activated Adhesive Composition and Antiviral Coatings

[0607] In this example, TMB protected bPEI-g-(3,4,5-TBA)2o, and bPEI-g-(2,3,4-TBA)2o were synthesized for rheology and lap shear on wooden coupons. Rheology of TMB protected bPEI-g-(3,4,5-TBA)20, and bPEI-g-(2,3,4-TBA)2oin 30 wt. % PBS. Lap shear of prepared samples on Mechanical Tester MTS C42 with 500N load cell, with mechanical grips to avoid slip.

[0608] Synthesis of TMB protected adhesives

[0609] To achieve the protection with trimethyl borate, 1:1 molar ratio of aldehyde to the protecting agent was used. Aldehyde was dissolved in anhydrous methanol (4-5 ml, depending on sample), then TMB was added dropwise until desired mass was added. Solution was stirred on vortex stirrer and left for 24 h in the fridge. There was no available kinetics of the synthesis, therefore long period in low temperature was deemed suitable for this test. As the TMB deprotects in water, 3,4,5-TBA monohydrate was dissolved pre-emptively and kept with 3 molecular sieves (20A, for water capture) beforehand, kept in fridge for the same amount of time (see FIG. 85).

[0610] After protection, the compounds were added to round flask containing branched polyethyleneimine (with molar ratio of 38.8 : 1 to achieve 20% amine graft)dissolved in 6 ml methanol under constant magnetic stir. The reaction took Ih, after which the methanol was removed in vacuo. Resulting compounds were weighted to estimate the amount of leftover solvent (see FIG. 86).

[0611] Resulting compounds showed minimal clumping and were homogeneous despite lower volume of solvent used (previous attempt of unprotected synthesis required over 20 ml of MeOH to avoid clumping).

[0612] Rheology of TMB protected adhesives

[0613] To test the impact of the TMB protection on the adhesive, synthesized adhesives were dissolved in PBS to achieve 30 wt.% concentration (see FIG. 87), then tested on the parallel plate rheometer with Zensor electrode and 10 mm ceramic probe as testing surfaces. Samples were dissolved within 10-20 min, then tested by applying 28 pF on the electrode, then pre-shearing for 30 s at steady rate of 10 s’1, with angular frequency of oscillating test being 6.28 rad / s over 30 min to observe the changes in G’ and G”, after which 5 min of amplitude sweep 1-100% was performed.

[0614] Example 7D: Discussion for Result based on Example 7 A

[0615] Protection of the hydroxyl groups seem to positively impact the viscosity, allowing for easier preparation of the solvent and handling. Out of two tested formulations, protection seems to impact them differently - with grafted 2,3,4-TBA the overall performance and stability was improved, while for grafted 3,4,5-TBA the storage modulus plummeted. There may be a possibility of stronger protection for 3,4,5-TBA, resulting in lower crosslinking density (explaining the similar toughness, but lower peak values) and lower modulus due to that.

[0616] Lap Shear sample preparation

[0617] To test the strength of the adhesive, the synthesized compounds were dissolved in MeOh and in PBS to 10 wt.% concentration (see FIG. 93).

[0618] The resulting compound was then applied on the wooden coupons: using 105 pl (3 times the volume used for collagen samples, on account of lower concentration) was applied onto 17 x 20 mm area of the wooden coupon, then pressed together with clips to cured the adhesive for 30 minutes. After 30 minutes, the samples were tested on MTS42 tensile tester with speed of 10 mm / min and mechanical grips.

[0619] Resulting samples were correctly tested - no slips occurred for all 20 wooden samples. However, some samples failed before the test (see FIG. 94 to FIG. 100).

[0620] The resulting data seem to point that TMB protection is not impacting the grafted 2,3,4-TBA adhesive performance, while severely hindering the performance of grafted 3,4,5-TBA. Using PBS instead of MeOH in hopes of speeding up deprotection yielded inconclusive results - for 2,3,4-TBA, the excess of water inhibits the performance, while there is not much difference for grafted 3,4,5-TBA.

[0621] Example 8: Advantages and Commercial Applications

[0622] In summary, adhesive compositions, such as an oxygen (and / or water) activated macromer containing substituted catechol and Schiff-bases, by amine / aldehyde condensation, are successfully developed as a self-curing adhesive when reconstituted in an aqueous solution with a predictable gelation time. The nature of the redox donor / acceptor groups allows the additional capability of voltage-based activation. The novel oxygen activated adhesive composition when dried and stored under anaerobic and anhydrous conditions is not an adhesive and can be stored for long periods at -20°C.

[0623] There are several advantages in the adhesive composition disclosed herein. Firstly the preparation is inexpensive and feasible for scaling up. Raw materials such as branched dendrimers are in abundance and cheap. The substituted catechol grafted dendrimer may be achieved in a single step without using special processing conditions such as high temperature, high pressure, and / or specific catalysts and special equipment. Secondly, use of the oxygen activated adhesive is very easy and convenient, rendering its feasibility for structural, veterinary, and clinical use.

[0624] Thirdly, the adhesives exhibited relatively high adhesion on wet substrates and allows many parameters for tuning the adhesion by substituted catechol, grafting degree, Fourthly, the components are non-toxic, and may be stored at -20°C for several months.

[0625] Fourthly, precursor oxygen activated adhesive exhibited relatively shelf stability under ambient conditions.

[0626] Fifthly, the components are non-toxic, and may be stored at -20°C for several months.

[0627] In various examples, there is demonstrated an adhesive composition, comprising (i) a macromolecule (e.g., a branched or unbranched polymer), wherein the macromolecule (i.e., macromer) can be grafted with substituteddihydroxybenzaldehyde precursors (e.g., at least 10 precursors) which are precursors to cross-linking groups (hereafter substituted catechol), that upon 1 or 2-electron oxidation per precursor by oxygen exposure, turns into a cross-linking functional group; the macromolecule comprises nucleophilic groups (e.g. amino groups on the branched or unbranched polymer) that spontaneously react with the cross-linking functional groups; and optionally (ii) additives comprising 1,2-aliphatic diols, catechol, tannic acid, 1,2-furan-diols, ascorbic acid, vicinal diols, and polysaccharide-diols; wherein the adhesive composition has shelf- stability within anaerobic conditions, but upon aqueous reconstitution, spontaneously cross-links within 60 seconds.

[0628] The adhesive composition according to one or more examples described above, wherein the macromolecule may not contain an aldehyde, acrylate, diazo, diazirine, genipin, and / or a cyan group.

[0629] The adhesive composition according to one or more examples described above, wherein the macromolecule can be selected from the group consisting of an amino group functionalized synthetic polymer, branched polysaccharide, a polyamino acid, and combinations thereof.

[0630] The adhesive composition according to one or more examples described above, wherein the macromolecule is selected from the group consisting of polyamidoamine, polyethylenimine, Boltom polyesters, polycaprolactone, chitosan, and combinations thereof

[0631] The adhesive composition according to one or more examples described above, wherein the macromolecule comprises or consists of polyethylenimine.

[0632] The adhesive composition according to one or more examples described above, wherein the redox donor precursors comprise 1,2-aryl-diols (e.g. catechol).

[0633] The adhesive composition according to one or more examples described above, wherein amount of substituted catechol grafted on the macromolecule is in the range of about 1% (w / w) to about 50% (w / w).

[0634] The adhesive composition according to one or more examples described above, wherein the cross-linking functional group is quinone, 1,2-cyclo-diketone, arylketone, or combinations thereof.

[0635] The adhesive composition according to one or more examples described above, wherein the nucleophilic groups are selected from the group consisting of an amino group, a thiol group, hydroxyl group, and combinations thereof.

[0636] The adhesive composition according to one or more examples described above, wherein the nucleophilic groups comprise or consist of an amino group.

[0637] Various examples also describes for a method of preparing an adhesive, the method comprising (a) providing an adhesive composition according to one or more examples described above; and (b) providing a rehydration mixture that comprises 30-90% water IIN%, mixing the adhesive composition and the rehydration mixture and oxygen activation of the adhesive composition is activated;

[0638] The method according to one or more examples described above, wherein concentration of the adhesive composition in the aqueous solution is in the range of about 10% w / v to about 50% w / v.

[0639] The method according to one or more examples described above, wherein the step of providing the adhesive composition comprises applying the adhesive composition on a wet substrate to form a coating.

[0640] An adhesive prepared by a method according to one or more examples described above.

[0641] An antimicrobial coating prepared by a method according to one or more examples described above.

[0642] While the present disclosure has been particularly shown and described with reference to specific embodiments, it should be understood by those skilled in the art that various changes in form and detail may be made therein without departing from the spirit and scope of the present disclosure as defined by the appended claims. The scope of the present disclosure is thus indicated by the appended claims and all changes which come within the meaning and range of equivalency of the claims are therefore intended to be embraced.

Claims

CLAIMS1. An adhesive composition comprising:an organic macromolecule comprising nucleophilic functional groups, wherein each of the nucleophilic functional groups comprises an amino group, a thiol group, a hydroxyl group, or a carboxyl group; andone or more moieties each comprising a ring structure having (i) one carbon attached with an aldehyde or a ketone, (ii) two carbons each having a hydroxyl group, and (iii) one carbon attached with a substituent group;wherein each of the nucleophilic functional groups grafted with one of the one or more moieties has the aldehyde or the ketone converted into a coupling moiety and each of the nucleophilic functional groups not grafted with the one or more moieties remains as the nucleophilic functional group,wherein each of the one or more moieties oxidizes into a diketone when the adhesive composition is subjected to a stimulus, and wherein the diketone forms one or more crosslinkages with the nucleophilic functional groups.

2. The adhesive composition of claim 1, wherein the organic macromolecule comprises a polymer or a dendrimer.

3. The adhesive composition of claim 1 or 2, wherein the organic macromolecule comprises polyethylenimine, polyamine, polyamidoamine, polylysine, polyester, polycaprolactone, chitosan, or a mixture thereof.

4. The adhesive composition of any one of claims 1 to 3, wherein each of the one or more moieties comprises a diol with two non-adjacent carbons each having one hydroxyl group, a catechol, a triol, or a derivative thereof.

5. The adhesive composition of any one of claims 1 to 4, wherein each of the one or more moieties is derived from a compound represented by a chemical formula of (la) or (lb) or (Ic) or (Id):wherein R1is the aldehyde or the ketone; andwherein R2represents the substituent group at a position not occupied by a hydroxy group and at a meta or an ortho or a para position of the ring structure relative to R1, wherein the substituent group comprises an electron donating group or an electron withdrawing group.

6. The adhesive composition of claim 5, wherein the substituent group comprises an alkoxy group, a halogen, a nitro group, an oxido group, a hydroxy group, an acyloxy group, an acylamido group, a (di)alkylphosphino group, an alkylthio group, a sulfhydryl group, or an alkyl halide.

7. The adhesive composition of any one of claims 1 to 6, wherein the diketone comprises ortho-quinone or para-quinone.

8. The adhesive composition of any one of claims 1 to 7, wherein the coupling moiety comprises a Schiff base.

9. The adhesive composition of any one of claims 1 to 8, wherein the stimulus comprises an aqueous medium, an organic medium, or a mixture thereof, containing moisture, oxygen, an oxidant, an electrical voltage, or a combination thereof.

10. The adhesive composition of any one of claims 1 to 9, wherein 10 molar% to 50 molar% of the nucleophilic functional groups are grafted, each with one of the one or more moieties, and 50 molar% to 90 molar% of the nucleophilic functional groups remain as the nucleophilic functional groups.

11. The adhesive composition of any one of claims 1 to 10, further comprising an additive comprising 1,2-aliphatic diol, catechol, tannic acid, 1,2-furan-diol, ascorbic acid, vicinal diol, polysaccharide-diol, or a mixture thereof.

12. The adhesive composition of any one of claims 1 to 11 for useas an adhesive to adhere biological tissues; oras a sealant in medical and / or veterinary applications comprising intestinal anastomosis, vascular anastomosis, tissue repair, and / or tissue implantation; oras a general-purpose adhesive or as a resin material; oras an anti-microbial coating or an anti-microbial fabric; oras a cosmetic coating or a dermatological coating.

13. A method for forming the adhesive composition of any one of claims 1 to 12, the method comprising:providing the organic macromolecule; andmixing the organic macromolecule with a precursor of the one or more moieties.

14. The method of claim 13, wherein providing the organic macromolecule comprises: providing a stock solution comprising the organic macromolecule in a first anhydrous organic solvent, andremoving dissolved oxygen from the stock solution.

15. The method of claim 13, wherein mixing the organic macromolecule with the precursor is carried out in an inert environment.

16. The method of claim 14, wherein mixing the organic macromolecule with the precursor comprises:adding the precursor to the stock solution having oxygen removed therefrom; ordissolving the precursor in a second anhydrous organic solvent to obtain a precursor solution, and mixing the precursor solution and the stock solution having oxygen removed therefrom.

17. The method of any one of claims 13 to 16, wherein the precursor comprises a dihydroxybenzaldehyde having the substituent group.

18. The method of any one of claims 13 to 17, wherein the precursor comprises 1,3-dihydroxybenzaldehyde, 2,3 -dihydroxybenzaldehyde, 3 ,4-dihydroxybenzaldehyde, 4,5-dihydroxybenzaldehyde, or a derivative thereof, having the substituent group.

19. The method of any one of claims 13 to 18, wherein the precursor comprises a catechol variant represented by the chemical formula of (la) or (lb) or (Ic) or (Id):wherein R1is the aldehyde or the ketone; andwherein R2represents the substituent group at a position not occupied by a hydroxy group and at the meta or the ortho or the para position of the ring structurerelative to R1, wherein the substituent group comprises the electron donating group or the electron withdrawing group.

20. The method of any one of claims 17 to 19, wherein the substituent group comprises an alkoxy group, a halogen, a nitro group, an oxido group, a hydroxy group, an acyloxy group, an acylamido group, a (di)alkylphosphino group, an alkylthio group, a sulfhydryl group, or an alkyl halide.

21. A method for adhering or sealing biological tissues or surfaces, the method comprising:applying the adhesive composition of any one of claims 1 to 12 to one or more biological tissues or surfaces;subjecting the adhesive composition to the stimulus; andcontacting the biological tissues or surfaces together to adhere or seal the biological tissues or surfaces, respectively.

22. The method of claim 21, wherein subjecting the adhesive composition to the stimulus comprises exposing the adhesive composition the aqueous solution containing oxygen, the oxidant, applying the electrical voltage to the adhesive composition, or a combination thereof.

23. A kit comprising:the organic macromolecule defined in claim 1 ; andthe precursor of the one or more moieties defined in claim 13,wherein the kit contains the organic macromolecule in a separate compartment from the precursor.

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