Synthesis of quinoline-based fluorescence probe for Zn(II) sensing and its applications in anti-counterfeiting ink and imaging in plants and living cells

2.1. Materials and instruments

8-hydroxyquinoline-2-carbaldehyde and 2-hydrazine pyridine were procured from Aladdin (Shanghai, China). All other analytical-grade chemicals were obtained from Sinopharm Chemical (Shanghai, China) and employed without further treatment. Different metal ions were obtained from chlorates (Na⁺, K⁺, Zn²⁺, Co²⁺, Ca²⁺, Mn²⁺, Hg²⁺, Fe³⁺, and Cr³⁺) or nitrate salts (Mg²⁺, Cu²⁺, Ni²⁺, Cd²⁺, Pb²⁺, Al³⁺, and Ag⁺). Double-distilled water was used throughout the spectroscopic measurements.

High-Resolution Mass Spectrometry (HR-MS) and NMR spectra of the probe were obtained from a UPLC G2-XS mass spectrometer (Waters, USA) and Bruker AV-500 spectrometer (Bruker BioSpin Gmbh, Germany), respectively. An Fourier transform infrared (FT-IR) study was carried out on a Thermo-Nicolet Nexus 670 FT-IR spectrometer by using KBr pellets. Beijing XT4-100x microscopic melting point apparatus (Beijing, China) was used to measure the melting point (m.p.). By using Pgeneral TU-1901 spectrophotometer (Pgeneral Instrument Inc., China), absorption spectra were measured. An Agilent Cary Eclipse spectrometer (Agilent Technologies, USA) was used to analyze the fluorescence behavior. The solid-state emission spectra were collected on an Edinburgh FLS920 fluorescence spectrometer (Edinburgh, UK). The transmission electron microscopy (TEM) images were recorded on a JEM-2100Plus microscope (Jeol, Japan). Cell images were carried out with an Olympus FV3000 confocal laser-scanning microscope (Olympus FV3000, Japan).

2.2. Synthesis of compound QP2 (2-((2-(pyridin-2-yl)hydrazinylidene)methyl)quinolin-8-ol)

Compound QP2 was synthesized by the facile Schiff-base condensation reaction. 2-Hydrazineylpyridine (109 mg, 1 mmol) and 8-hydroxyquinoline-2-carbaldehyde (173 mg, 1 mmol) were dissolved in 10 mL of ethanol and refluxed at 90°C for 2 h. Light yellow acicular crystals were obtained by filtration. Yield: 86%. M.p.:239.2-240.5°C. ¹H NMR (500 MHz, DMSO-d₆, ppm) (Figure S1): 11.44 (s, 1H), 9.72 (s, 1H), 8.27 (d, J = 6.4 Hz, 2H), 8.18 (d, J = 4.3 Hz, 1H), 8.14 (d, J = 8.7 Hz, 1H), 7.72 (t, J = 7.2 Hz, 1H), 7.43-7.36 (m, 3H), 7.11-7.07 (m, 1H), 6.89-6.83 (m, 1H). ¹³C NMR (125 MHz, DMSO-d₆, ppm) (Figure S2): δ 153.61, 148.36, 139.68, 138.58, 136.59, 128.73, 127.89, 118.25m 117.94, 116.30, 112.42, 107.22. HRMS (ESI +) (Figure S3): found for [QP2 + Na⁺], 287.0905 (calculated, 287.0904). FT-IR (KBr Pellet) ν(cm^-1) (Figure S4): 1618 (C=N), 1545 (quinoline C=N),1298 (Ar-N), 1280(Ar-O).

2.3. Spectroscopic measurements

Probe QP2 was dissolved in tetrahydrofuran (THF) to afford a stock solution (1 mM). A series of metal ion salts were separately dissolved in deionized water to obtain stock solutions (5 mM). The testing solutions were prepared by taking QP2 stock solution (100 μL) and an appropriate metal solution and then diluted to 5 mL with DMSO/H₂O (f_w = 95%) solution for determination of the spectroscopic behavior. The pH values were adjusted to designated levels using dilute HCl or NaOH solutions as required.

2.4. Cell imaging experiment

The cell cytotoxicity assays were employed by the methylthiazolyldiphenyl-tetrazolium (MTT) assay. HepG2 cells were grown in Dulbecco’s modified Eagle’s medium (37°C, 5% CO₂) supplemented with 10% fetal bovine serum for 24 h. The cells were subsequently incubated for another 24 h in the presence of QP2 with different concentrations (0-30 μM). Then MTT was added and incubated for 4 h. The number of living cells was quantitatively determined by measuring the absorbance intensity at 570 nm. The entire experiment was independently repeated three times. The imaging of Zn²⁺ using QP2 was performed on HepG2 cells. The HepG2 cells were treated with 20 μM QP2 in culture medium for 30 mins. The cells were washed with phosphate-buffered saline three times and then exposed to Zn²⁺ (10 μM) for 30 mins, after which fluorescence images were obtained under a confocal laser fluorescence microscope.

3.1. Synthesis and characterization of the probe QP2

The probe QP2 was synthesized through a condensation reaction between 8-hydroxyquinoline-2-carbaldehyde and 2-hydrazineylpyridine, followed by comprehensive characterization using NMR, HR-MS, and FT-IR spectra (Figures S1-S4). The ¹H NMR spectrum revealed distinct downfield-shifted signals at δ 11.44, and 9.72 ppm, corresponding to the amine (-NH-) and hydroxyl (-OH) protons, respectively (Figure S1), suggesting the presence of intramolecular/intermolecular hydrogen bonds [27]. FT-IR analysis confirmed the structural features through characteristic absorption bands: an azomethine (C=N) stretching vibration at 1616 cm^-1, along with a separate pyridinic C=N stretching mode at 1554 cm^-1 and quinolinyl C=N stretching vibration at 1545 cm^-1 [28].

3.2. Spectroscopic recognition of Zn^{2 +}

The sensing behavior of probe QP2 toward various metal ions was primarily explored via UV-vis absorption spectroscopy in a DMSO/H₂O (f_w = 95%) solution. As shown in Figure 1(a), the as-synthesized sensor QP2 exhibited two bands at 350 nm and 270 nm, which were attributed to the n-π* electronic transition. After the addition of Zn²⁺, Ni²⁺, Hg²⁺, Cd²⁺, Pb²⁺, and Fe³⁺ ions, the n-π* transition bands shifted significantly. Notably, with the addition of Zn²⁺, the band at 350 nm shifted to 342 nm, and a new shoulder peak appeared at 375 nm (Figure S5), suggesting the formation of the corresponding complex. However, for other metal ions, i.e., Na⁺, K⁺, Ag⁺, Mg²⁺, Ca²⁺, Cu²⁺, Co²⁺, Mn²⁺, Al³⁺, and Cr³⁺, only negligible changes were observed in the absorption spectra.

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Figure 1.

(a) UV-vis spectra of QP2 (20 µM) with different metal ions (10 µM) in DMSO/H₂O (f_w = 95%) solution. (b) Fluorescence spectra of QP2 (20 µM) with various metal ions (10 µM) in DMSO/H₂O (f_w = 95%) solution (Inset: Visual outputs of QP2 before and after treatment with Zn²⁺ under UV_{365 nm} lamp). (c) Normalized UV-vis absorption and emission of QP2 in the presence of Zn²⁺. (d) Fluorescence spectra of QP2 (20 µM) upon Zn²⁺ addition (0-30 µM) in DMSO/H₂O (f_w = 95%) solution. The two absorption peaks on the red curve are located at 342 nm and 375 nm. The red curve represents UV-vis absorption, while the blue curve shows fluorescence emission.

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The emission responses of QP2 upon the separate addition of different metal ions were also evaluated (Figure 1b). The metal-ion-free QP2 solution exhibited an extremely weak fluorescence peak, centered at 524 nm. Upon excitation, the proton from the hydroxyl group was transferred to the quinoline N atom (Scheme 1), resulting in ESIPT [12]. Simultaneously, the C=N isomerization process, a non-radiative deactivation pathway, occurred [29,30]. Thus, the weak fluorescence of QP2 was attributed to the combined effect of ESIPT and -C=N isomerization processes [31]. When Zn²⁺ was added to the QP2 solution, a significant enhancement in fluorescence (558 nm) was observed. Notably, a change in the fluorescence emission color, from colorless to golden yellow, was observed with the naked eye under 365 nm UV light (Figure 1b, insets). This was attributed to the formation of the QP2-Zn²⁺ complex. However, the other metal ions, including those that caused changes in the absorption spectrum, produced no emission response. Notably, in the presence of Zn²⁺, a large Stokes shift of 183 nm between the normalized absorption and emission spectra was observed (Figure 1c). This “turn-on” fluorescence emission behavior of QP2 confirmed its potential to effectively detect Zn²⁺ in the DMSO/H₂O (f_w = 95%) solution.

Fluorescence titration experiments were performed to evaluate the quantitative sensing properties of QP2 toward Zn²⁺ (Figure 1d). With increasing Zn²⁺ concentration, the fluorescence intensity at 558 nm continuously increased. At concentrations above 0.5 equivalents, a near-saturation state was reached, corresponding to nearly 75-fold fluorescence enhancement. These results indicated the formation of a 2:1 binding stoichiometric complex between QP2 and Zn²⁺. The plot of emission intensity (I_{558 nm}) versus Zn²⁺ concentrations showed an excellent linear relationship in the range of 0-0.5 equivalents of Zn²⁺ (Figure S6). The limit of detection (LOD) of QP2 for Zn²⁺ was 17.7 Nm. It was determined using the formula 3σ/k, where σ is the standard deviation of the blank measurement and k is the slope of the linear fit obtained from the fluorescence titration experiments. The LOD was significantly below the guideline value (76 µM) for drinking water set by the World Health Organization [8]. These results confirmed the high sensitivity of QP2 toward Zn²⁺.

To study the time-dependent fluorescence of QP2 during Zn²⁺ detection, the response time was measured. As presented in Figure 2(a), the fluorescence intensity of QP2 after treatment with Zn²⁺ reached a maximum value within 3 mins, which remained stable even after a prolonged period of 120 mins. In addition, the effect of pH on the detection of Zn²⁺ by QP2 was studied in a pH range of 3-11 (Figure 2b). Free probe QP2 exhibited weak fluorescence, which remained nearly unchanged in the whole pH range. Notably, in the presence of Zn²⁺, the fluorescence intensity of QP2 was considerably enhanced over a broad pH range 5-10. However, the fluorescence signal intensity decreased at both strong acidic and basic pH values. This was attributed to the protonation of QP2 in strong acidic media and the competition between QP2 and hydroxide ions under strong basic conditions [32]. These results confirmed the ability of QP2 to quickly detect Zn²⁺ in complex environmental and biological samples, without the need to strictly control the pH value of the sample solution. Compared with recently reported AIE-active fluorescent probes for Zn²⁺ detection, QP2 demonstrated applicability over a broad pH range and lower LOD in aqueous systems with a high-water content (Table 1).

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Figure 2.

(a) Time-dependent fluorescence intensity of QP2 in the presence of 0.5 equiv. amount of Zn²⁺ in DMSO/H₂O (f_w = 95%) solution. (b) pH-dependent fluorescence intensity of QP2 (20 µM) before and after treatment with 0.5 equiv. of Zn²⁺ in DMSO/H₂O (f_w = 95%) solution.

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Table 1. Comparison of the properties of some recent AIE-active fluorescent probes for Zn²⁺ detection and this work.

Structure of a chemosensor	Synthesis complexity	Tested media	LOD (nM)	pH range	Ref.
	three steps	DMSO-H2O (1:9)	130	7.0	[33]
	one step	EtOH-H2O (1:9)	718	7-9	[34]
	one step	DMSO-H2O (9:1)	19		[35]
	three steps	DMSO-H2O (1:99)	176		[36]
	four steps	EtOH-H2O (4:1)	21	4.8-12	[37]
	one step	THF-H2O (3:7)	107	6-12	[12]
	one step	DMSO-H2O (5:95)	17.7	5-10	This work

Moreover, to evaluate the selectivity of QP2 toward Zn²⁺, we performed fluorescence competition assays by adding various environmentally and biologically relevant metal ions in a DMSO/H₂O (f_w = 95%) solution. As shown in Figure 3, the QP2-Zn²⁺ system exhibited significant fluorescence enhancement upon the addition of most of the interfering ions. The observed partial quenching of the QP2-Zn²⁺ system’s emission intensity by Cu²⁺ and Fe³⁺ could be attributed to the inherent paramagnetic quenching effect [32] and the oxidative nature of the ions [38,39]. In contrast, other paramagnetic ions (Ni²⁺, Cr³⁺, and Co²⁺) showed negligible interference, consistent with their lack of redox activity under these experimental conditions. Cd²⁺ and Zn²⁺ have been reported to act as competitors because the two metals belong to the same group of the periodic table and exhibit similar photophysical responses toward most probes [8,40]. However, in this study, the coexistence of Cd²⁺ did not lead to any interference with the fluorescence intensity of the QP2-Zn²⁺ system. These results confirmed the potential of QP2 to distinguish Zn²⁺ from most of the other potential competitors, demonstrating the anti-interference feature of the probe.

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Figure 3.

Fluorescence response of QP2 (20 µM) towards Zn²⁺ (0.5 equiv.) in DMSO/H₂O (f_w = 95%) solution after addition of other competitors (1.0 equiv.)

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3.3. Possible sensing mechanism of QP2 toward Zn^{2 +}

The binding stoichiometry of QP2 with Zn²⁺ was explored by the Job’s plot method based on the fluorescence emission spectrum. As shown in Figure 4, the emission intensity of QP2 at 558 nm reached its maximum value when the molar fraction of [Zn²⁺]/[Zn²⁺ + QP2] was about 0.33, indicating a 2:1 binding ratio between QP2 and Zn²⁺. Unfortunately, attempts to acquire the mass spectrum of QP2-Zn²⁺ complex were unsuccessful due to its insufficient solubility in both methanol and acetonitrile.

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Figure 4.

Job’s plot of QP2-Zn²⁺ complex at 558 nm in DMSO/H₂O (f_w = 95%) solution.

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¹H NMR titration was performed to identify the relevant atom for binding with Zn²⁺ and thereby gain insight into the binding behavior of the QP2-Zn²⁺ complex (Figure 5). For QP2, upon the addition of 0.5 equivalents of Zn²⁺, the hydroxyl group proton signal (H_b) at 9.72 ppm disappeared, which was attributed to the deprotonation of the hydroxyl group to form a strong O-Zn bond [41]. Moreover, the imine proton signal (H_c) at 8.26 ppm shifted to 8.34 ppm after Zn²⁺ introduction. Protons (marked by diamonds) in the quinoline ring also exhibited obvious shifts. However, no significant changes were observed in the pyridinium ring proton signals (marked by an asterisk). Owing to the influence of the solution environment, the N-H (H_a) signal became weaker and broader but did not disappear. These results indicated that the deprotonation O atom of the hydroxyl group, and the N atoms of the imine group and quinoline were involved in the coordination with Zn²⁺.

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Figure 5.

¹H NMR titration of QP2 with Zn²⁺ in DMSO-d₆.

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To further investigate the Zn²⁺ binding site in QP2, the QP2-Zn²⁺ complex was synthesized. For this, a methanolic solution of ZnCl₂ (0.01 mmol) was added to a methanolic solution of QP2 (0.02 mmol) and stirred for 5 mins at room temperature to obtain a yellow solid. The solid was filtered and washed with ethanol to obtain the pure product. The ay-synthesized QP2-Zn²⁺ complex was characterized via FT-IR spectroscopy (Figure S7). The imine C=N stretching band was found to have shifted from 1616 cm^-1 for QP2 to 1605 cm^-1 for the complex, indicating the coordination of the imine N atom with Zn²⁺. Moreover, the characteristic peak of the C=N quinoline ring at 1545 cm^-1 was shifted to 1532 cm^-1, but the pyridinium ring peak at 1554 cm^-1 was unchanged. The displacement of the band assigned to the ν_C=N vibration in the complex confirmed the participation of the quinoline N atom in the coordination [42]. In addition, the hydroxyl group at 3194 cm^-1 was not observed for the complex. The N-H stretching vibration peak was observed in the region of 3450-3415 cm^-1 for both QP2 and the complex. On the basis of the experimental results (Job’s plot, NMR, and IR), we proposed the binding mechanism of QP2 with Zn²⁺, which was confirmed by subsequent density functional theory (DFT) calculations and spectroscopic experiments, in which fluorescence intensity changes were observed with varying water fraction (Scheme 1).

To identify the geometrical structural features of QP2 and the possible QP2-Zn²⁺ complex, DFT calculations were carried out with wB97x-D3/6-311G+ (d,p) level using the ORCA program (Neese, 2022). The polarizable continuum model (PCM) model mimicked aqueous solution in the calculation process. Figure 6(a) shows the optimized structure of QP2 in the ground state, which could be seen to be planar. The distance of OH∙∙∙N (quinoline) was 2.08 Å, indicating the existence of the intramolecular H-bond between the -OH group and quinoline nitrogen atom [43]. The geometric structure of its tautomeric form was also optimized (Figure 6b), and the potential energy curve along the proton-transfer coordinates was schematically described in Figure 6(c). The proton-transfer process from the hydroxyl group to the nitrogen atom of quinoline required overcoming the energy barrier of 17.98 kcal/mol, but the reverse process was easier with an activation energy of only 9.91 kcal/mol. So, the enol form (QP2) was the dominant type in the ground state.

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Figure 6.

(a) Optimized structure of QP2. (b) Optimized structure of the Zwitterionic form QP2’. (c) Energy profile of the hydrogen-transfer reaction of QP2→QP2’. (d) Optimized structures of the complex QP2- Zn²⁺. (e) π-π stacking of the complex QP2- Zn²⁺. (f) Molecular orbitals and energy levels of QP2 and QP2- Zn²⁺.

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Molecular electrostatic potential (MEP) of QP2 was calculated to predict potential binding sites. As could be seen in Figure S8, the blue regions located at O1, N1, and N2 atoms of the compound QP2 were rich in electrons, which implied that QP2 would exhibit good coordination ability to metal ions. Then, the proposed structure of QP2-Zn²⁺ complex was optimized (Figure 6d). In the complex, two QP2 molecules were arranged in head-to-tail and simultaneously coordinated to Zn²⁺ through the two N1 atoms and two O1 atoms of the quinolyl group and the two N2 atoms of the imine group. The hexa-coordinated complex could be described as a distorted octahedron and showed an X-shaped spatial structure (Figure 6e). The Zn-O/N bond lengths were in the range of 2.06-2.62 Å, which were similar to some reported hexa-coordinated complexes [44,45]. Moreover, Figure 6(e) revealed that there existed π-π stacking between the adjacent quinoline ring and the pyridine ring, which could further strengthen the head-to-tail stacking pattern in the QP2-Zn²⁺ complex. So, the QP2-Zn²⁺ system presented a rigid hydrophobic structure, which would be conducive to molecular aggregation [46]. In the aggregated state, the free intramolecular rotation N−N bond could be prevented by the close packing of the molecules, exhibiting AIE.

For QP2, the electron density of HOMO was spread across almost the molecule, whereas the electron density of LUMO was mainly located on the quinoline group and imine unit (Figure 6f). For the HOMO → LUMO transition, the electron density of the hydroxyl O atom decreased while that of quinoline N atom increased, which was prone to the ESIPT process [27,41]. After complex formation, the electron density in HOMO was majorly located at imine unit and the deprotonated 8-hydroxyquinoline group, while LUMO was spread across the molecule with Zn²⁺ as the center. Moreover, the HOMO-LUMO energy gap of the QP2-Zn²⁺ (6.95 eV) complex was found to be lower than the free QP2 (7.52 eV). The results indicated that the ESIPT process was inhibited after coordination. Overall, the observed remarkable enhancement of the emission spectra in the QP2-Zn²⁺ system could be ascribed to the concerted effect of the inhibition of the ESIPT and C=N isomerization process and the AIE behavior of the QP2-Zn²⁺ formed complex.

The AIE behavior of the QP2-Zn²⁺ complex was first confirmed by the Tyndall effect, caused by the scattering of light due to aggregation (Figure 7a). As expected, QP2 exhibited negligible Tyndall responses in the DMSO/H₂O (f_w = 95%) solution. However, a strong Tyndall signal was observed upon the addition of Zn²⁺ to the QP2 solution. Next, the fluorescence spectrum of QP2-Zn²⁺ was collected by varying the DMSO/water ratio (0-98%) (Figure 7b). When the fraction of water in the system (f_w) was increased from 0% to 40%, a hypsochromic shift was observed, with a decrease in the orange fluorescence emission intensity. The decrease in the intensity was attributed to the concentration quenching effect [40]. However, as f_w was gradually increased, the fluorescence intensity increased significantly, reaching the maximum at f_w = 95%. This was attributed to the formation of QP2-Zn²⁺ aggregates. Meanwhile, the solution fluorescence changed from orange to a strong golden yellow under 365 nm UV light (Figure 7c). Notably, the quantum yield of QP2-Zn²⁺ increased from 0.06 to 0.33 as the water fraction was increased from 50% to 95% (Table S1). Nonetheless, a slight reduction in the emission intensity was observed when f_w was further increased to 98%, owing to the precipitation of the aggregates. Moreover, the aggregation behavior of the QP2-Zn²⁺ complex was confirmed by TEM. The QP2 probe exhibited sparse particulate distribution (Figure S9a). However, after treatment with 10 µM Zn²⁺, distinct nanoparticles were observed. Notably, with increasing Zn²⁺ concentration, significant aggregation was observed (Figures S9b and c). In addition, the fluorescence of QP2-Zn²⁺ was investigated in methanol-glycerol viscous solvents (Figure S10). The emission intensity increased with increasing glycerol percentage in the mixture, which was attributed to the restriction of intramolecular rotation along the N-N single bonds in high-viscosity media. Furthermore, the emission of the QP2-Zn²⁺ complex was stronger than that of QP2 in the solid state (Figure 7d). The abovementioned phenomenon revealed the AIE characteristic of the QP2-Zn²⁺ system.

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Figure 7.

(a) Tyndall effect of the complex QP2-Zn²⁺. (b) Emission spectra of QP2-Zn²⁺ system in DMSO/H₂O with different water fractions. (c) Fluorescence photographs of QP2-Zn²⁺ system in DMSO/H₂O with different water fractions. (d) Fluorescence emission spectra of QP2 and the complex QP2- Zn²⁺ in the solid state. Inset: photograph of the complex QP2- Zn²⁺ in the solid state under daylight and 365 nm UV lamp.

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3.4. Application of chemosensor QP2

3.4.1. Anti-counterfeiting ink

Fluorescent ink is widely used as an anti-counterfeiting material [47,48]. Considering the rapid visible and fluorescent response of QP2 on exposure to Zn²⁺, we investigated the practical application of QP2 in anti-counterfeiting. The QP2 stock solution (1 mM) was used as an ink to write the letters “ZKNU” on cellulose paper. Then, the tested paper samples were exposed to Zn²⁺ solution (1 mM). As illustrated in Figure 8(a), the letters obtained using QP2 were invisible in the daylight, but they quickly turned yellow when exposed to Zn²⁺ solution. Moreover, under 365 nm UV light, they appeared bright gold yellow. The patterns were clearly visible under daylight and UV light, even after the test samples were soaked in water for 1 h. However, letters written with a watercolor paint of the same color exhibited no fluorescence (Figure 8b). To further evaluate the anti-counterfeiting performance of QP2-Zn²⁺ complex, we employed an invisible yet stable Zn²⁺ solution as printing ink to imprint a school badge on QP2-coated filter paper (prepared from a 1 mM stock solution) (Figure 8c). The printed pattern also exhibited a yellow coloration under daylight and a bright gold-yellow emission under 365 nm UV light. Remarkably, the fluorescent signature remained stable without noticeable fading even after 10 days of environmental storage, confirming the good luminescence stability of QP2-Zn²⁺. These results indicated that the QP2-Zn²⁺ complex could be potentially used as an anti-counterfeiting ink.

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Figure 8.

(a) Photographs of the letters treated by QP2 upon exposure to Zn²⁺ solution. (b) Photographs of the letters and pattern printing treated by watercolor paint. (c) Photographs of school badge imprinted by Zn²⁺ solution on QP2-coated filter paper. (under daylight and 365 nm UV lamp).

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3.4.2. Determination of Zn^{2 +} in test strips

To evaluate the practical application of the sensor QP2 in on-site Zn²⁺ detection, test strip experiments were performed. First, the prepared filter paper strips were immersed in a THF solution of QP2 (200 μM) for 30 seconds, and dried in air. Later, the dried QP2-coated paper strips were soaked in different metal-ion aqueous solutions (100 μM) for 10 seconds. As presented in Figure 9(a), only Zn²⁺ emitted a yellow fluorescence under 365 nm UV light among all the tested metal ions. Additionally, the fluorescence color changed with increasing Zn²⁺ concentration (Figure S11). These results indicated that QP2 could be conveniently utilized to fabricate paper strips for Zn²⁺ detection.

Figure S11

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Figure 9.

(a) Fluorescence photographs of QP2 pre-stained strip for sensing different metal ions. (b) Images of QP2 and QP2 + Zn²⁺ in mung bean sprouts under daylight and UV_{365 nm} light. Each image from left to right: sprouts treated with QP2; Zn²⁺-pretreated sprouts treated with QP2.

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3.4.4. Cell imaging

To verify the bioimaging application of QP2, cytotoxicity experiments were performed on HepG2 cells via MTT assays with different QP2 concentrations (0, 10, 20, and 30 μM). The viability of cells was found to be above 85% (Figure S12). The results indicated that QP2 had a low cytotoxicity and could be safely used for cell imaging. Moreover, the fluorescence imaging of living HepG2 cells was performed. Probe QP2 alone did not exhibit any fluorescence in HepG2 cells. However, HepG2 cells incubated successively with QP2 and Zn²⁺, exhibited a notable green fluorescence signal (Figure 10). These results indicated that probe QP2 could be potentially used for imaging exogenous Zn²⁺ in living cells.

Figure S12

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Figure 10.

Fluorescence images of HepG2 cells incubated with QP2, and QP2 + Zn²⁺.

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