New insights in polydopamine formation via surface adsorption

The study addresses the unresolved mechanism of polydopamine (PDA) film formation and the identity of key intermediates during dopamine polymerization. PDA is a mussel-inspired, broadly adherent and biocompatible coating formed by auto-oxidation of dopamine in alkaline media, yet its insolubility and concurrent particle/film growth complicate structural elucidation. Prior reports propose diverse structural models (polymeric, supramolecular, and trimer-based), with conflicting roles for 5,6-dihydroxyindole (DHI) and uncertainty due to technique limitations (e.g., mass spectrometry ambiguity for isobaric species). The purpose here is to slow and capture intermediate species by exploiting adsorption on negatively charged, high-surface-area halloysite nanotubes (HNTs), enabling time-resolved analysis of functional groups and intermediates during PDA formation and clarifying the polymerization pathway.

The paper surveys competing PDA structural models: polymeric eumelanin-like DHI-based chains; open-chain poly(catechol/quinone); mixed uncycled/cyclized units with TRIS incorporation; high-molecular-weight covalent polymers; versus supramolecular aggregates stabilized by noncovalent interactions. Some studies identify noncovalent dopamine/DHI assemblies and dopamine-dopaminechrome (DAC) complexes; others propose covalent trimers like (DHI)2/pyrrole dicarboxylic acid linked noncovalently. Time-resolved XPS studies on flat substrates reported rapid stabilization of surface chemistry within minutes. Limitations of mass spectrometry to distinguish isomers with identical masses are highlighted, motivating functional-group-resolved approaches (XPS, ssNMR) to track chemical evolution during polymerization.

• Systems studied: Dopamine auto-oxidation at pH 8.5 (10 mM TRIS buffer) with and without halloysite nanotubes (HNTs). HNTs provide high surface area and negative charge for intermediate adsorption.
• Sample preparation:
  • PDA aggregates (PDA-A): 0.4 g dopamine in 200 ml 10 mM TRIS (pH 8.5), stirred 24 h; precipitate isolated by centrifugation and dried.
  • HNT-PDA coatings: 0.8 g HNTs dispersed in 200 ml 10 mM TRIS (pH 8.5), sonicated 30 min; add 0.4 g dopamine; stir at room temperature for 5, 15, 30, 60, 120, 240 min; isolate by centrifugation, wash, dry.
  • Control for buffer effects: carbonate buffer (10 mM, pH 8.5) replacing TRIS; samples at 5 and 15 min.
  • Macroscopic substrate comparison: Si wafers with native oxide (1.5 × 1.5 cm2) immersed in 2 mg/ml dopamine + 10 mM TRIS; withdrawn at set times, rinsed, dried (SiO2-PDA).
• Characterization and time-resolved tracking:
  • Dynamic light scattering (DLS): Particle size distributions of reaction mixtures with/without HNTs; reaction stopped by acidification to pH 2 with 4 M HCl before measurement. Also DLS for dopamine adsorption at pH ~5 without TRIS.
  • Nitrogen adsorption/desorption porosimetry (BJH analysis): Pore size distributions to assess pore filling and aggregation evolution.
  • Thermogravimetric analysis (TGA): Mass fraction of organics (PDA) on HNTs vs time (25–700 °C at 5 °C/min under N2).
  • STEM-EDS mapping: Spatial distribution of C, Si, Al, O on individual nanotubes (1, 4, 24 h polymerization).
  • Solid-state 13C CP/MAS NMR: Short (80 µs) and long (2 ms) CP contact times for PDA-A and HNT-PDA (24 h), and time series for HNT-PDA; assignment of aliphatic and aromatic carbons to DA, DAQ, DAC, TS, PDCA, DHI, and TRIS.
  • X-ray photoelectron spectroscopy (XPS): Survey and high-resolution C 1s and N 1s spectra as a function of deposition time for HNT-PDA and SiO2-PDA; peak fitting (C–C/C=C, C–O/C–N, C=O/C=N, O=C–O; N 1s components for R–NH3+, R–NH–R, imine). Elemental atomic percentages and N/C ratios used to evaluate film growth and chemistry.
• Numerical modeling: Empirical model using XPS peak components to estimate relative percentages of key building blocks (DA, DAQ, DAC, PDCA, TS, TRIS) vs time; details in Supplementary Information.

• Adsorption and aggregation:
  • At pH ~5 (no TRIS), dopamine adsorbs on HNTs, increasing DLS peak from ~320 nm (HNTs) to ~1200 nm.
  • In polymerization medium (10 mM TRIS, pH 8.5): DLS shows monomodal distributions shifting to larger sizes over time.
    • Without HNTs: ~180 nm at 10 min → ~1500 nm at 3 h.
    • With HNTs: ~1500 nm at 10 min → ~3000 nm at 3 h.
  • Monomodal distributions with HNTs indicate predominant growth of PDA on nanotubes rather than coexistence with free particles; intermediates are adsorbed by HNTs.
• Porosity and mass loading:
  • BJH pore distribution: 10–20 nm feature (inner HNT pores) decreases and shifts to smaller widths over time, indicating pore filling by oligomeric/polymeric species; macropore volume (>50 nm) increases, consistent with aggregate formation.
  • TGA: Residual weight decreases up to 700 °C; PDA mass on HNTs increases nonlinearly—faster in first ~2 h, then slower—indicating higher initial deposition rate.
  • STEM-EDS: Carbon signal uniformly covers nanotubes and increases with polymerization time (1, 4, 24 h), evidencing progressive coating.
• ssNMR (13C CP/MAS):
  • Both PDA-A and HNT-PDA exhibit aliphatic signals at ~35 and 44 ppm (sp3 carbons from uncycled units: DA, DAQ; and indoline carbons in DAC/TS/PDCA). A 60 ppm peak (TRIS aliphatic carbons) is pronounced in PDA-A and weaker in HNT-PDA.
  • Aromatic region shows signals near 105, 119, 130, and 146 ppm. The shoulder at 90–110 ppm (characteristic of DHI protonated aromatics) is much less pronounced in HNT-PDA, indicating low DHI content on HNTs (attributed to DHI neutrality and lack of adsorption at pH 8.5).
  • Carbonyl/carboxyl signals (170–190 ppm) are weak/absent at 24 h, suggesting low abundance in mature films, though detectable at earlier times (intermediates).
• XPS:
  • Film growth confirmed by increasing C and N signals and decreasing Si/Al contributions; N/C ~0.10 ± 0.02 matches PDA stoichiometry.
  • C 1s components: C–C/C=C (~284.8 eV), C–O/C–N (~286.1 eV), C=O/C=N (~287.5 eV), O=C–O (~289.2 eV), and shake-up (~291.3 eV).
  • N 1s evolution: Initially dominated by R–NH3+ (~402.5 eV) and R–NH–R (~400.1 eV); with time, emergence and growth of imine component (~398.8 eV). Primary amine prominence early confirms presence of DA/DAQ and intact TRIS units.
  • Early-time C 1s ratio (C–C/C=C):(C–O/C–N) < 1 (at 5–15 min), inconsistent with dopamine-only species, indicating TRIS incorporation; carbonate buffer control eliminates this effect, supporting TRIS inclusion.
  • On SiO2 wafers (macroscopic substrate), minimal temporal evolution of functional groups; compositions comparable to PDA aggregates, attributed to far fewer adsorption sites than HNTs.
• Mechanistic model (from numerical analysis of XPS):
  • DAQ is high at 5 min then decreases; DA increases initially (up to ~60 min) then declines, consistent with intermediate kinetics in consecutive reactions.
  • Proposed sequence: quinone–catechol coupling (reverse dismutation) yields polycatecholamine crosslinks (biphenyl-type dicatechol); subsequent oxidation and intermolecular cyclization of uncycled units generates DAC; tautomerization produces TS in small amounts; oxidative ring opening of DAC forms PDCA units (low abundance). TRIS is incorporated early and decreases as film grows.
  • Overall, oxidative coupling of quinone units to form polycatecholamine is the dominant pathway in the initial stages; cyclization and further transformations follow. DHI plays a minimal role in HNT-deposited films.

By leveraging the high surface area and negative charge of HNTs to adsorb intermediates, the study decelerates PDA formation, enabling time-resolved detection of functional groups and intermediates. The combined DLS, porosimetry, TGA, EDS, ssNMR, and XPS data coherently indicate that early-stage polymerization is dominated by oxidative coupling of quinone-containing species to form polycatecholamine crosslinks, followed by gradual cyclization to dopachrome-related structures (DAC/TS) and minor oxidative ring-opening to PDCA. The clear early incorporation of TRIS, evidenced by both XPS (C–O/C–N excess and primary amines) and ssNMR (~60 ppm), diminishes with time as the PDA network develops. The low DHI contribution in HNT-bound films suggests that DHI is not a primary building block for PDA coatings under these conditions, reconciling discrepancies in prior literature and emphasizing the importance of substrate-mediated intermediate capture. These insights clarify the PDA formation pathway, explain substrate- and time-dependent variations in reported film chemistries, and highlight a practical approach (nanotube adsorption) to interrogate fast polymerization processes.

This work provides a comprehensive, time-resolved picture of polydopamine film formation by exploiting halloysite nanotubes to capture intermediates. The key contributions are: (i) identification of polycatecholamine as a central intermediate formed via oxidative coupling of quinone-containing species; (ii) demonstration that cyclization to dopachrome-related structures occurs subsequently, with minor formation of PDCA; (iii) evidence that DHI plays a limited role in film formation on HNTs; and (iv) verification of early TRIS incorporation that diminishes over time. The methodology reconciles conflicting structural models and offers a pathway to tailor PDA coatings by controlling buffers, substrates, and reaction times. These insights can be used to optimize PDA-based surface modifications for applications in energy, environment, and biomedicine.

Findings are derived from systems where intermediates are captured on negatively charged, high-surface-area halloysite nanotubes; thus, intermediate distributions and kinetics may differ on substrates with fewer adsorption sites or different surface chemistries (e.g., flat SiO2). The low DHI signal on HNTs likely reflects adsorption bias due to DHI neutrality at pH 8.5, which may underrepresent its role in solution-borne aggregates. ssNMR carbonyl signals are weak in mature films, limiting direct quantification of quinone/carboxyl functionalities at late times. The empirical model relies on deconvolution and assignment of XPS components, which carries intrinsic uncertainties when multiple species contribute overlapping signals.