Changes in Optical Properties upon Dye–Clay Interaction: Experimental Evaluation and Applications

The development of hybrid materials with unique optical properties has been a challenge for the creation of high-performance composites. The improved photophysical and photochemical properties observed when fluorophores interact with clay minerals, as well as the accessibility and easy handling of such natural materials, make these nanocomposites attractive for designing novel optical hybrid materials. Here, we present a method of promoting this interaction by conjugating dyes with chitosan. The fluorescent properties of conjugated dye–montmorillonite (MMT) hybrids were similar to those of free dye–MMT hybrids. Moreover, we analyzed the relationship between the changes in optical properties of the dye interacting with clay and its structure and defined the physical and chemical mechanisms that take place upon dye–MMT interactions leading to the optical changes. Conjugation to chitosan additionally ensures stable adsorption on clay nanoplatelets due to the strong electrostatic interaction between chitosan and clay. This work thus provides a method to facilitate the design of solid-state hybrid nanomaterials relevant for potential applications in bioimaging, sensing and optical purposes.

. Summary of the attractive and repulsive interactions based on the partial positive or negative charge of substituent groups in molecules interacting with clay [1]. Attractive  To evaluate the possibility of improving dye adsorption on the MMT and the stability of the molecules here studied, FITC, Rho, E, CCF, CNF and CNH2 were conjugated with chitosan exploiting the presence of primary amino groups in its structure ( Figure S1). Figure S1. Scheme of conjugation. i: Chitosan deacetylated (5 mg) in HCl 1 M (1 mL) stirred for overnight, roomtemperature (RT); filtered and neutralized to 6 pH with NaOH; FITC is added reaching 1.1 mM as final concentration (final volume 5 mL). The reaction is stirred for 24 hours, RT; the excess of FITC is removed by dialysis (24h) and centrifugation 8000 rpm, 10 min (x3).
Free dyes ( Figure S2, Figure S3) and chitosan-conjugated dyes ( Figure S4) were treated with 1C and after 15 minutes absorbance and fluorescent spectra were recorded. The spectra of the dye interacting with MMT were compared with the one measured for the dye in water.
Only relatively small variations were observed in the absorbance spectra of free dyes ( Figure   S2). In particular, when treated with MMT, FITC and Fluo, the spectra showed an increase in the main peak at 490 nm and a decrease in the intensity for the shoulder at 450 nm which is commonly observed for xanthene dyes and is due to the formation of dimers/aggregates between single molecules [2]. In the case of Rho, the main absorbance peak was slightly redshifted from 550 nm to 570 nm. The fluorescence enhancement observed when excited at 490 nm was remarkable, almost 5 times higher for FITC and Fluo, while the fluorescent enhancement observed for Rho was less outstanding (intensity increased from 15 to 25 a.u.). In the case of E, only the intensity of the absorbance peak at 520 nm increased from 0.4 to 0.9 a.u.
while negligible changes in the fluorescent signal at 544 nm were observed. The opposite results were achieved for CCF in which the absorbance peak was comparable between the water and 1C samples at 400 nm, whereas the fluorescent signal measured at 425 nm in the presence of 1C was 2.5 times higher than in water. When treated with MMT, a bathochromic shift was observed for CNH2 in the absorbance spectrum (from 340 nm to 360 nm), while the fluorescent signal measured at 450 nm was found to be around 3 times higher than the one measured for CNH2 in water.   The values of the so calculated E.F. factor are reported in Table S2. The intensity was determined using the following wavelengths as λex-λem respectively: FITC/Chi_FITC 490-520 nm; Rho/Chi_Rho 550-590 nm; CCF/Chi_CCF 500/525 nm; E/Chi_E 520-550 nm; CNH2/Chi_CNH2 340-450 nm. Evaluation of the mechanism involved in the optical changes induced by MMT particles on the studied dyes ( Figure S5 and S6).  Examples of application of hybrid materials composed of dyes and MMT particles.
Once absorbed on the MMT surface, the pH responsiveness of Chi_FITC was lost. This could be of interest in designing materials in which optical behavior should not be altered by pH variation (Figure S7). The E.F. achieved using MMT could be exploited to improve sensitivity, decrease the limit of detection (LOD) and reduce the background noise of fluorescence-based detection systems. As proof of concept, we proved the E.F. which can be achieved with fluorescamine, a compound commonly used for the fluorescent detection of amino acids ( Figure S8). Energy-dispersive X-ray (EDX) analysis was performed using the above mentioned SEM equipped with an INCA X-Sight detector (Oxford Instruments). Three independent specimens were measured and an average value was obtained from the resulting elemental compositions.