Development of Spiropyran Immobilization and Characterization Protocols for Reversible Photo-Patterning of SiO₂ Surfaces

Spiropyran, a type of photochromic compound, is distinguished by its reversible transformation between its colorless, hydrophobic spiropyran (SP) form and the purple, hydrophilic merocyanine (MC) form. This transformation can be triggered by UV light and reversed by either visible light or heat. Spiropyran functionalized surface, therefore, present reversible contrast in properties such as color, fluorescence, and reactivity, rendering it optimal for a range of applications in chemical sensor, biosensor, drug delivery, and heavy metal extraction.<br/><br/>Despite the successful application of spiropyran attachment in various substrates: non-covalently in metal organic frameworks (MOFs), poly (ethylene glycol) (PEG) lipids, polydimethylsiloxane (PDMS), and covalently in poly (methyl methacrylate) (PMMA), polystyrene microbeads, silica microbeads, cellulose nanofibers, cellulose nanocrystals - its implementation on plain surfaces has been limited due to surface area constraints and insufficient characterization.<br/><br/>Our research focuses on a 3-step spiropyran immobilization protocol tailored for thermally grown SiO₂surfaces. This process involves surface activation, amination using (3-aminopropyl) triethoxysilane (APTES) and spiropyran functionalization. We have also developed a robust characterization method utilizing techniques such as ellipsometry, X-ray photoelectron spectroscopy (XPS), Fourier transform infrared spectroscopy (FTIR), and fluorometric detections.<br/><br/>Our main results include comparing the effectiveness of various activation methods, such as H₂SO₄ acid, oxygen plasma, and piranha solution treatments. XPS and contact angle measurements show that the surface is clean and properly activated for all cases. We then monitored the uniformity and thickness of the APTES layer. By varying the reaction temperature, time, and concentration, we achieved a uniform and thin layer immobilization of APTES, verified by ellipsometry and XPS. A 1% APTES incubation for 5 minutes at room temperature yielded the best results. The functionalization step was verified by fluorescence microscopy, showing a clear contrast between SP and MC forms. FTIR monitors the change of functional groups after each step. The developed protocol demonstrated exceptional repeatability, with photo-patterning results showing reversibility over more than ten cycles and feature sizes of less than 20 µm.<br/><br/>We also discussed the intrinsic limitations of immobilizing spiropyran onto a plain and rigid substrate, such as steric hindrance and effective activated surface sites, which greatly reduced the number of spiropyran immobilized. To address these challenges and expand the potential applications of our protocol, we propose several innovative directions for future research. One approach involves immobilizing spiropyran to a porous media that provides a larger surface area or finding a type of surface with more effective activated sites. Another strategy is to further modify the MC form to enhance the contrast between SP and MC forms.<br/><br/>In conclusion, our study highlights the potential of spiropyran in surface patterning applications and offers a comprehensive protocol for its immobilization on SiO₂ surfaces, which should also be applicable to other surfaces. These findings pave the way for the development of on-chip sensing and other applications.