Multi-Stimuli Responsive FRET Processes of Bifluorophoric AIEgens in an Amphiphilic Copolymer and Its Application to Cyanide Detection in Aqueous Media

Abstract

A novel amphiphilic aggregation-induced emission (AIE) copolymer, that is, poly(NIPAM-co-TPE-SP), consisting of N-isopropylacrylamide (NIPAM) as a hydrophilic unit and a tetraphenylethylene-spiropyran monomer (TPE-SP) as a bifluorophoric unit is reported. Upon UV exposure, the close form of non-emissive spiropyran (SP) in poly(NIPAM-co-TPE-SP) can be photo-switched to the open form of emissive merocyanine (MC) in poly(NIPAM-co-TPE-MC) in an aqueous solution, leading to ratiometric fluorescence of AIEgens between green TPE and red MC emissions at 517 and 627 nm, respectively, via Förster resonance energy transfer (FRET). Distinct FRET processes of poly(NIPAM-co-TPE-MC) can be observed under various UV and visible light irradiations, acid-base conditions, thermal treatments, and cyanide ion interactions, which are also confirmed by theoretical studies. The subtle perturbations of environmental factors, such as UV exposure, pH value, temperature, and cyanide ion, can be detected in aqueous media by distinct ratiometric fluorescence changes of the FRET behavior in the amphiphilic poly(NIPAM-co-TPE-MC). Moreover, the first FRET sensor polymer poly(NIPAM-co-TPE-MC) based on dual AIEgens of TPE and MC units is developed to show a very high selectivity and sensitivity with a low detection limit (LOD = 0.26 μM) toward the cyanide ion in water, which only contain an approximately 1% molar ratio of the bifluorophoric content and can be utilized in cellular bioimaging applications for cyanide detections.

Keywords: Förster resonance energy transfer (FRET); aggregation-induced emission (AIE); cyanide; spiropyran; tetraphenylethylene.

Figure 1.

(a) PL spectra and (b) relative fluorescence intensity at 517 nm of poly(NIPAM-co-TPE-SP) and (c) PL spectra and (d) relative fluorescence intensity at 627 nm of poly(NIPAM-co-TPE-MC) with different H₂O fractions in THF/H₂O solutions. Inset: PL photo-images of panel (b) poly(NIPAM-co-TPE-SP) and panel (d) poly(NIPAM-co-TPE-MC) in THF (left) and water (right). Concentration: 1 g/L, λ_ex: 365 nm.

Figure 2.

Time-dependent (a) absorption and (b) PL spectra (λ_ex = 365 nm) of poly(NIPAM-co-TPE-SP) and poly(NIPAM-co-TPE-MC) (1 g/L) in water upon UV exposure (0−90 s). Insets: Photo-images of naked-eye observation and photoluminescence color changes. (c) Schematic illustration of the energy transfer from TPE to MC unit via FRET process after UV exposure and (d) spectral overlap of absorption spectra of poly(NIPAM-coTPE-MC) and emission spectra of poly(NIPAM-co-TPE-SP).

Figure 3.

(a) PL spectra of poly(NIPAM-co-TPE-MC) and (b) relative fluorescence intensities of poly(NIPAM-co-TPE-MC) at 517 and 627 nm with different pH values. Insets: PL photo-images of polymer solutions at different pH values. Concentration: 1 g/L, λ_ex: 365 nm.

Figure 4.

(a) PL spectra of poly(NIPAM-co-TPE-MC) and (b) relative fluorescence intensities of poly(NIPAM-co-TPE-MC) at 517 and 627 nm under UV exposure upon heating (20−60 °C). Insets: PL photo-images of polymer solutions at different temperatures. Concentration: 1 g/L, λ_ex: 365 nm.

Figure 5.

(a) Titration profile change and hypsochromic shift of poly(NIPAM-co-TPE-MC) after addition of different CN⁻ concentrations, (b) proposed schematic illustration of the CN⁻ detection mechanism, (c) PL spectra (insets: PL photo-images of polymer solution with/without cyanide ion), and (d) fluorescence intensity ratios response of poly(NIPAM-co-TPE-MC) upon addition of various analytes (200 μM). Inset: PL photo-images of polymer solutions in the presence of various analytes. Concentration: 1 g/L, λ_ex: 365 nm, MOPS buffer solution, *poly(NIPAM-co-TPEMC.

Figure 6.

(a) Optimized structures of TPE-SP, TPE-MC, and TPE-MC-CN⁻ at B3LYP/6–31G(d). Only MC and MC-CN⁻ fragments of the latter two structures are shown, since the TPE fragment of all three compounds is similar (the optimized coordinates are included in the Supporting Information). Electronic transitions and corresponding molecular orbitals for (b) TPE-SP, (c) TPE-MC, and (d) TPE-MC-CN⁻. Orbital energies were computed at IEF-PCM-B3LYP/6–31G(d).

Figure 7.

Optimized structures of (a) protonated and (b) deprotonated TPE-MC at B3LYP/6–31G(d). Protonated and deprotonated positions are circled in red. Molecular orbital diagrams of (c) protonated and (d) deprotonated TPE-MC, with orbital energies computed at IEF-PCM-B3LYP/631G(d).

Figure 8.

Confocal fluorescence images of HeLa. (a−e) Control experiments of HeLa. (f−j) Cells were incubated with poly(NIPAM-co-TPE-SP) for 30 min. (k−o) Cells were treated with poly(NIPAM-co-TPE-SP) for 30 min followed by incubation with CN⁻ for 30 min. (p−t) Cells were incubated with poly(NIPAM-co-TPE-MC) for 30 min. (u−y) Cells were treated with poly(NIPAM-co-TPE-MC) followed by incubation with CN⁻ for 30 min. The polymeric fluorophores and cyanide concentration = 10 μM. Green and red emissions were collected at 500−550 and 600−650 nm, respectively. Scale bars = 10 μm, λ_ex: 365 nm.

Scheme 1.

Synthetic Routes of Monomer TPE-SP and Copolymers Poly(NIPAM-co-TPE-SP) and Poly(NIPAM-co-TPE-MC)