Adjacent cationic–aromatic sequences yield strong electrostatic adhesion of hydrogels in seawater

Electrostatic interaction is strong but usually diminishes in high ionic-strength environments. Biosystems can use this interaction through adjacent cationic–aromatic amino acids sequence of proteins even in a saline medium. Application of such specific sequence to the development of cationic polymer materials adhesive to negatively charged surfaces in saline environments is challenging due to the difficulty in controlling the copolymer sequences. Here, we discover that copolymers with adjacent cation–aromatic sequences can be synthesized through cation–π complex-aided free-radical polymerization. Sequence controlled hydrogels from diverse cation/aromatic monomers exhibit fast, strong but reversible adhesion to negatively charged surfaces in seawater. Aromatics on copolymers are found to enhance the electrostatic interactions of their adjacent cationic residues to the counter surfaces, even in a high ionic-strength medium that screens the electrostatic interaction for common polyelectrolytes. This work opens a pathway to develop adhesives using saline water.


Characterization of polymerization kinetics
The monomer conversion for the free radical polymerization in DMSO was analyzed using with 1 H-NMR reaction kinetics study. The mixed solution with different total monomer concentration of 1:1 monomer ratio and 0.25 mol% UV initiator (2-oxoglutaric acid, in a concentration relative to the total monomer concentration) in DMSO were polymerized in the glass vials under the irradiation of UV light (3.9 mW cm -2 ) in the glove box. To study the monomer conversion during the copolymerization, 150 L of the sample was taken from the reaction sample tube at different reaction times and transferred from the glove box to air immediately to quench the reaction, and then added into 300 L DMSO-d6 solution. The concentration of unreacted monomers remaining in solution was determined from the integral area ratio of 1 H-NMR signals.

Synthesis of polymer hydrogels
The formulation of hydrogels is shown in Supplementary Table 1. All hydrogels were synthesized using the one-step random copolymerization of the prescribed monomers in DMSO.
The monomers (molar ratio is 1:1) with the prescribed total monomer concentration (Cm), 0.25-mol% UV initiator (2-oxoglutaric acid, relative to the total monomer molar concentration) and 0-0.15 mol% 3 chemical crosslinker (MBAA, relative to the total monomer molar concentration) were first dissolved in DMSO, and then the resulting mixture was poured into a reaction cell consisting of a pair of glass plates with a 1-mm spacing and irradiated with a 365-nm UV light for 11 h. After the polymerization, the as-prepared gel was immersed in a large amount of 0.7 M NaCl (aqueous) solution to wash away the DMSO and residual chemicals. The saline water was exchanged every 12 h for over 1 week, after that the samples reached equilibrium. Before the test, the hydrogels were stored in 0.7 M NaCl solutions.
The water content (CW) of hydrogels was measured using Moisture Balance (SHIMADZU, MOC-120H). We assumed that the salt concentration (Csalt) in the gel is equal to that in the outer solution. The polymer content (CP) was calculated from CW as: where H2O is the density of water, Csalt is the molar concentration of salt in water, and Msalt is the molar weight of salt.

Tensile test
The tensile stress-strain measurements were performed using a universal testing machine (UTM, INSTRON 5965) at a steady velocity of 100 mm min -1 in air. The samples were cut into a dumbbell-shape with the standard JIS-K6251-7 size (12 mm (L) × 2 mm (d) × 1~2 mm (w)). The elastic modulus (E) was calculated from the slope over 4-8% of the strain of the stress-strain curve.
The tensile stress was calculated from the tensile force divided by the cross-section area of the virgin sample. The tensile strain was calculated from the displacement of cross-head of the testing machine divided by the initial gauge length (12 mm). The initial strain rate was calculated from the deformation velocity divided by the initial gauge length.

Rheological test
Rheological tests were performed using an ARES-G2 rheometer (TA Instruments). A rheological angular frequency sweep from 0.01 to 100 rad s -1 was performed with a shear strain of 0.1% in the parallel-plate geometry at 24°C. The disc-shaped samples with thicknesses of approximately 1.5 mm and diameters of 15 mm were placed on the plates and surrounded by 0.7 M NaCl saline solution.

Static polymer absorption on SiO2
The amount of polymer adsorbed on SiO2 surface was measured by Quartz Crystal Microbalance (QCM, AFFINIX QN Pro, QCM2008-PRKIT, with frequency 27 MHz). The SiO2-4 coated sensor was first washed in UV. The 450-L 0.7 M NaCl solution was added into the chamber and left undisturbed until the frequency stabilized. After that, a 4.5-L polymer solution (10 mg mL -1 in 0.7 M NaCl) was added into the chamber and left undisturbed until equilibrium (final polymer concentration 0.1 mg mL -1 ).

Measurement of adhesiveness
The tack test was used to characterize the adhesiveness. The hard substrates used were commercially available glass (Matsunami Glass, Osaka, Japan, S2112), positively charged glass (MAS-coated Superfrost, Matsunami Glass, Osaka, Japan, S9441), polyethylene terephthalate (PET), and polymethyl methacrylate (PMMA). The hydrogels, P(NaSS) gel, P(ATAC-adj-PEA)-0.1 gel, and P(AAm) gel were used as soft substrates (see synthesis method below). The hard substrates were rinsed with ethanol, and then with deionized water before use. The soft substrates were swollen in the 0.7 M NaCl saline water to reach equilibrium before use.
The test was performed on the SHIMADZU tester (autograph AG-X) with Trapezium X software. To perform the experiment, the hydrogel with a diameter of 15 mm and thickness of approximately 1.2-2.0 mm was first glued to the probe using cyanoacrylate (super glue), and then the gel (on the probe) was immersed into the test solution for 5 min, so that it can reach equilibrium before the test. The probe approached the substrate surface at a speed of 10 m s -1 , held by the applied pressure for 10 s (value equals to the elastic modulus of the tested sample), and then retracted at a rate of 100 m s -1 . Except stated otherwise, all the tests were performed under 0.7 M NaCl solution.

Synthesis of soft substrates for adhesion test
All hydrogels were synthesized by polymerization of a 10-mL mixed solution with the prescribed chemicals (Supplementary Table 2) in a reaction cell with 1-mm spacing under UV light for 11 h. After polymerization, the as-prepared gels were immersed in a large amount of 0.7 M NaCl solution to wash the residual chemicals. Before the test, the hydrogels were stored in 0.7 M NaCl solutions. The thicknesses of these hydrogels were in the range of 1.5-2.0 mm.

Supplementary Note 2: Characterization of monomer sequence of copolymers
The sequence of aromatic residues on copolymer chains was characterized by 1 H-NMR.
Supplementary Figure 5a shows the partial (aromatic protons) 1 H-NMR spectra of P(ATAC-co-PEA) with different PEA fractions, f, in DMSO. Firstly, we compare the NMR spectra of PEA monomer and homopolymer poly(PEA) (f 1.0). The aromatic proton signals of poly(PEA) shift to the higher field and broaden due to different chemical environments on the polymer chains. For copolymers, when the fraction of PEA is low (f ≤ 0.5), the aromatic signals broaden slightly, but maintain almost the same position with the monomers, indicating no apparent aromatic-rich sequence in the copolymer. However, for copolymers bearing high PEA molar fraction (f > 0.5), the signals show a broad shoulder at the higher field (indicated by arrows), and the shoulders shift to the higher chemical field by increasing the PEA fraction, indicating the formation of aromatic-rich segments in the copolymer chains. This broad shoulder at higher field is a signature to identify the existence of aromatic-rich segment in copolymers.
Accordingly, we compared the NMR signals of copolymers from four pairs of monomers (monomer ratio 1:1). Supplementary Figure 5b shows the partial (aromatic protons) 1 H-NMR spectra of copolymers in DMSO. For copolymers from a pair of monomers having the same vinyl group (P(ATAC-adj-PEA) and P(MATAC-adj-PEMA)), the signals show symmetric broadening around the peak of phenyl protons of the aromatic monomer, but no obvious shift. However, the signal of the other two copolymers (P(ATAC-r-PEMA) and P(MATAC-r-PEA)) have broad shoulders at the higher field (indicated by arrows). The NMR results shown above indicate that at the equimolar ratio, the pairs of cationic and aromatic monomers having the same vinyl groups (R1 = R2) can form a copolymer with adjacently dispersed cationic and aromatic residues. Otherwise (R1 ≠ R2), the obtained copolymers have inhomogeneously dispersed residues containing both cationic-rich and aromatic-rich segments.

Supplementary Note 3: Characterization of cation-π interaction of copolymers
Poly(cation-adj-π) is soluble in water because the strong electrostatic repulsion of cationic residues prevent the adjacent hydrophobic residues from aggregating; in salt water (0.7 M NaCl), the salt ions screen the long-range electrostatic repulsion and strengthen the effective attraction between cationic and aromatic groups, forming strong intra-chain/inter-chain cation-π interactions, which causes the formation of coacervate 1 . Raman spectroscopy (RENISHAW, inVia Reflex, with a 532-nm laser light as an excitation source and a power of 10 mW) was performed to probe cation-π interactions in the system. In the Raman spectra, the ring breathing mode, centered at 998 cm -1 , appears as a very strong band compared to the adjacent 1,028 cm -1 band (out-of-plane bending of C-Hring) of polymer in water. Upon the addition of salts, the intensity ratio, I998/I1028, changed from 2.92 in water to 2.19 in saline water ( Supplementary Figure 7), indicating the strengthening of cation-π interactions in saline solution. The reduction of intensity of the ring breathing mode observed in the systems is a result of the formation of cation-π interactions between quaternary-N in the cationic monomers and phenyl group in the aromatic monomers, because an interaction of this type reduces the polarizability of the π-electron density in the ring 2 .

Supplementary Figures
Supplementary Figure 1  to the elastic modulus E of the gels. All the tests were performed on a glass substrate.