Novel Benzothiazole-Based Highly Selective Ratiometric Fluorescent Turn-On Sensors for Zn²⁺ and Colorimetric Chemosensors for Zn²⁺, Cu²⁺, and Ni²⁺ Ions

Abstract

Metal ions play a very important role in environmental as well as biological fields. The detection of specific metal ions at a minute level caught much attention, and hence, several probes are available in the literature. Even though benzothiazole-based molecules have a special place in the medicinal field, only very few chemosensors are reported based on this moiety. The current work describes the design and synthesis of the benzothiazole-based chemosensor for a highly selective and sensitive detection of biologically important metal ions such as Zn²⁺, Cu²⁺, and Ni²⁺. The sensing studies of compound-1 showed a ratiometric as well as colorimetric response toward Zn²⁺, Cu²⁺, and Ni²⁺ ions and color changes from colorless to yellow and is found to be insensitive toward various metal ions (Cd²⁺, Cr³⁺, Mn²⁺, Pb²⁺, Ba²⁺, Al³⁺, Ca²⁺, Fe²⁺, Fe³⁺, Mg²⁺, K⁺, and Na⁺). Further, compound-1 exhibited ratiometric as well as turn-on-enhanced fluorescence response toward Zn²⁺ ions and turn off response for Cu²⁺ and Ni²⁺ ions. The Job plots revealed that the binding stoichiometry of compound-1 and metal ions is 2:1. The detection limits were found to be 0.25 ppm for Zn²⁺, while it was 0.30 ppm and 0.34 ppm for Ni²⁺ and Cu²⁺, respectively. In addition, density functional theory results strongly support the colorimetric response of metals, and the reversibility studies suggested that compound-1 can be used as a powerful chemosensor for the detection of Zn²⁺, Cu²⁺, and Ni²⁺ ions. The bioimaging data illustrated that compound-1 is a very effective ratiometric sensor for Zn²⁺ ions in live cells.

Introduction

It is not surprising that in the advanced scientific era humans in an effort to ease their lives are unaware that their activities are harming nature and organisms. Importantly, the toxic metal ions, which harm the environment, have made us look back and think about the environmental concerns. Efforts are underway to find such toxic metal ions in the environment. Even though researchers have shown progress in this regard, more work is required to gain an upper hand in finding the toxic metal ions in the environment. The role of chemosensors in detecting the toxic metal ions in water bodies is outstanding.

Novel chemosensors continue to be an exciting area of the present research world. In the past, some traditional highly expensive instrumental methods such as atomic absorption spectroscopy,¹ inductively coupled plasma atomic emission,² anodic stripping voltammetry,^3a potentiometry,^3b spectrophotometry, and so on have been used for the detection of ions. Practically, these methods were considered to be unpropitious in terms of time consumption, need experts for the handling and cost.⁴ Compared with the methods mentioned above, fluorescent chemosensors have drawn remarkable attention from the biological, chemical, and environmental research groups due to their ease of synthesis, high efficiency, selectivity, very fast response, low cost, as well as low detection limits toward metal ions particularly for transition-metal ions.⁵ In addition to fluorescence enhancement or quenching, in some cases, the probe would also act as a colorimetric chemosensor for sensing the metal ions.⁶ The colorimetric method of sensing is quick, equally selective and sensitive and associated with naked eye detection without the aid of any instrument. Colorimetric receptors have gained much attention, and researchers have focused on the synthesis of several artificial dyes for the selective recognition of specific metal ions.^6b,7 On the other hand, the development of highly sensitive ratiometric and “turn-on” fluorescent probes have superior significance due to their capability of detection of metal ions even in biological systems,^5b,6d,8 which was found to display different emission behavior upon metal binding. Due to the great impact on fluorescence turn-on receptors, lately, researchers have been devoted to the synthesis and structural modifications on BODIPY,⁹ anthracene,¹⁰ coumarin,¹¹ rhodamine,¹² and so on. However, several synthetic modifications on fluorescent dyes are found to have difficulties in purification and many of them lack the selectivity toward particular metal ions.

Zinc is the second most abundant metal in the human body and plays an important role in the biological reactions such as regulatory functions in enzyme catalysis, neurophysiology, gene expression, and DNA binding.¹³ However, the imbalance of these metal ions can ultimately lead to neuron dysfunctions causing Alzheimer’s, Parkinson’s disease, amyotrophic lateral sclerosis, Creutzfeldt–Jakob disease, multisystem atrophy, diabetes, and prostate cancer.¹⁴ Moreover, notably an excess of Zn leads to environmental pollution via the reduction of soil microbial activity causing phytotoxic effects.¹⁵ Accordingly, due to the great relevance of Zn, it is very important to monitor its level in the biological system. Owing to the advantages and disadvantages of Zn, the design and synthesis of novel receptors which can detect and monitor its concentration have been a concern for chemists and are necessary. Moreover, recently, there have been several fluorescent receptors developed for the selective recognition of Zn²⁺ ions such as chelating peptides, proteins, and macrocyclic compounds.¹⁶

Copper is the third most abundant transition-metal ion found in all body tissues after Fe³⁺ and Zn²⁺ owing to several essential biological processes. Moreover, copper is present in everything including soil, water, and air and is also an essential element for the all living beings such as human, animals, and plants.^17a Several copper-containing pesticides and herbicides are extensively used for the treatment of plant diseases.^17b,17c Additionally, it plays an important role in the human body, for example, iron absorption, maintaining nerves, blood vessels, and active catalytical cofactor in several metalloenzymes, for instance, superoxide dismutase, tyrosinase, and cytochrome c oxidase.¹⁸ However, the high amount of copper intake also becomes toxic to the body; destructive consequences causing irritation of the nose and throat, nausea, vomiting, and diarrhea. Further, a very high concentration of copper leads to the annihilation of organs in infants, whereas the deficiency of copper leads to abnormal growth of bones. The abnormal levels of copper lead to the disorder of the cellular homeostasis, which causes oxidative stress accompanied by several neurodegenerative syndromes, including Menkes, Wilson, familial amyotrophic lateral sclerosis, and Alzheimer’s.¹⁹

Nickel is also an important trace element for a living system in respiration, biosynthesis, and metabolism.^20a The deficiency of the nickel(II) ion leads to harmfulness to prokaryotic and eukaryotic organisms.²⁰ However, the accumulation of excessive nickel can cause lung cancer, asthma, sinus, pneumonitis, acute pneumonitis, syndromes of the central nervous system, and abnormal increment of blood cells.²¹ In addition, Ni²⁺ ions are also an environmental pollutant along with other heavy metal ions. Consequently, there is a very high demand for novel receptors of nickel in various chemical and biological samples.²²

Even though numerous different types of chemosensors are available for the recognition of these biologically important metal ions (Zn²⁺, Cu²⁺, and Ni²⁺), ratiometric, fluorescence turn-on, as well as colorimetric “naked eye” chemosensors have gained great attention due to their easy sample preparation, naked eye detection, effortlessness, high sensitivity, easy way of functioning, and usefulness of identifying metal ions in biological media via cellular imaging.²³ Additionally, the development of the single receptor, which could target multiple analytes, has been anomalously increasing in the past decade due to several advantages, such as potential cost and analytical time reduction. Recently, various research groups around the world focused on receptors which respond to multiple metal ions simultaneously, including Cr³⁺/Al³⁺,²⁴ Cu²⁺/Hg²⁺,²⁵ Cu²⁺/Zn²⁺,²⁶ Zn²⁺/Cd²⁺,²⁷ Ag⁺/Mn²⁺,²⁸ Al³⁺/Fe³⁺,²⁹ and Cr³⁺/Fe³⁺.³⁰

In this context, we have proposed functionalized thiazole derivative compound 1 as a superior chemosensor for the recognition of Zn²⁺, Cu²⁺, and Ni²⁺ with multiple binding sites such as N–O–N or S–O–S chelation. Heterocyclic compounds containing nitrogen and sulfur such as quinoline³¹ and benzothiazole³² correspondents were extensively studied in the field of the metal ion sensing chemistry as well as biological aspects. However, benzothiazole-based ratiometric and fluorescent turn-on chemosensors for Zn²⁺ ions are scarce in the literature.³⁶ Herein, we have designed and synthesized biphenyl-based benzothiazole (1) via a condensation reaction between the 4-phenyl 2,6-diformylphenol and 2-aminobenzenethiol. The receptor was found to show a colorimetric response toward Zn²⁺, Cu²⁺, and Ni²⁺ with a significant color change from colorless to yellow in DMSO/CHCl₃ (50:50, v/v) and no change upon the addition of other metal ions such as Al³⁺, Ca²⁺, Cd²⁺, Co²⁺, Fe²⁺, Fe³⁺, Hg²⁺, K⁺, Mg²⁺, Mn²⁺, Na⁺, Cd²⁺, and Pb²⁺. Further, fluorescence titration of 1 was found to be ratiometric and fluorescence turn-on for Zn²⁺, whereas turn off for Cu²⁺ and Ni²⁺. Moreover, the receptor showed the detection limit for Zn²⁺ as low as 0.25 ppm and was very low as compared to Ni²⁺ and Cu²⁺ ions, which were 0.30 and 0.34 ppm, respectively.

Results and Discussion

The synthetic procedure for the new sensor is described as follows. First, 4-phenylphenol was treated with (hexamethylenetetramine) HMTA/(trifluoroacetic acid) TFA mixture and was refluxed for 48 h. The crude product was purified through column chromatography and the pure compound was obtained, 4-phenyl 2,6-diformylphenol as a yellow solid with 78% yield.³³ Further, the condensation reaction was carried out between 4-phenyl 2,6-diformylphenol and aminothiophenol in EtOH under reflux conditions for 24 h and a white compound was obtained as a target molecule with good yields, which was purified via column chromatography (Scheme 1).^32e The target compound was structurally characterized by ¹H, ¹³C NMR, as well as high-resolution mass spectrometry (HRMS) (see in the Supporting Information).

Scheme 1. Synthetic Scheme for the Target Molecule.

After successfully obtaining the pure form of compound-1, we have evaluated the sensing ability toward diverse cations in a 1:1 ratio of DMSO and CHCl₃ solutions. The absorption spectrum of compound-1 in DMSO displayed two strong bands around 300 and 370 nm, which can be attributed to π–π* and n−π* transitions, respectively. The metal ion sensing studies were carried out using UV–visible and fluorescence spectroscopic studies. 1 μM of compound-1 was titrated with Zn²⁺ ions and found that upon incremental addition, the absorption band at 370 nm was gradually decreased and simultaneously a new absorption band was generated at 440 nm. As a result, the color of the solution changed from colorless to dark yellow. Under similar conditions, Ni²⁺ and Cu²⁺ also showed similar changes in absorption bands, but a drastic change was seen in the case of Ni²⁺ as compared with Zn²⁺ and Cu²⁺. Further, compound-1 exhibited a ratiometric response toward Ni²⁺, Cu²⁺, and Zn²⁺ (see the Supporting Information). The isosbestic point was found to be at 393 nm for Zn²⁺, while for Cu²⁺ and Ni²⁺ were found to be at 392 and 405 nm, respectively, which clearly indicated the smooth molecular conversion of free ligand to metal complex. Therefore, the new bathochromic peaks can be accredited to the metal to ligand charge transfer, which is accountable for the naked eye detection of Zn²⁺, Cu²⁺, and Ni²⁺ ions (Figures 1 and 2). Furthermore, we have checked the colorimetric response toward the other competitive metal ions such as Cd²⁺, Cr³⁺, Mn²⁺, Pb²⁺, Ba²⁺, Al³⁺, Ca²⁺, Fe²⁺, Fe³⁺, Mg²⁺, K⁺, and Na⁺, which caused no change or very slight change to the absorption spectra even in the presence of excess amounts of these metal ions.

Figure 1.

Change in absorption spectra of compound-1 (1 μM) upon the addition of Zn²⁺ (left) and Cu²⁺ (right) in a 1:1 ratio of DMSO and CHCl₃ (inset photographs were taken in the presence of normal light).

Figure 2.

Change in absorption spectra of compound-1 (1 μM) upon the addition of Ni²⁺ (left) and different cations (right) in a 1:1 ratio of DMSO and CHCl₃ (inset photographs were taken in the presence of normal light).

The complete saturation of compound-1 was obtained after the addition of 2.5 equiv of metal ions (Ni²⁺, Cu²⁺, and Zn²⁺). Moreover, the binding stoichiometry of the receptor with metals (Ni²⁺, Cu²⁺, and Zn²⁺) was determined using a Job's plot. This is an unceasing process as well as well known for the determination of the stoichiometric ratio of complexes. Compound-1 displayed an absorption maximum around 440 nm upon the addition of metal ions, which was monitored for Job’s plot to determine the ratios of compound-1 and metal ions. Various mole fractions of metal ions and compound-1 were mixed in different proportions from 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, and 0.9. The maximum absorbance for the complex was observed at 0.3 mole fraction corresponding to a 2:1 stoichiometry of the metal–compound-1 complex (Figures 3 and 4). Additionally, we have determined the association constants (k_a) of compound-1 with Zn²⁺, Cu²⁺, and Ni²⁺ ions which were found to be 4.9 × 10¹¹, 6.0 × 10¹⁰, and 1.6 × 10¹¹M^-1, respectively. These results strongly suggested that the binding nature of Zn²⁺ ions to the compound-1 is much stronger as compared to Cu²⁺ and Ni²⁺ ions.

Figure 3.

Job’s plot according to the method of continuous variations, indicating the 1:2 stoichiometry for compound-1 + Zn²⁺ (left) and compound-1 + Cu²⁺ (right).

Figure 4.

Job’ plot according to the method of continuous variations, indicating the 1:2 stoichiometry for compound-1 + Ni²⁺ ions.

The fluorescence emission of compound-1 exhibited a very bright yellow color (575 nm) with 0.55 quantum yields, upon excitation at 375 nm in a 1:1 ratio of DMSO and CHCl₃ solutions. To better understand the sensing ability of compound-1, we have carried out metal binding studies using fluorescence spectroscopy. During UV–visible titrations, we observed a color change in the solution as well as different fluorescence upon the addition of Zn²⁺ ions. For better investigation, we have carried out the fluorescence titration of compound-1 with the subsequent addition of Zn²⁺ and found that the emission intensity at 575 nm was gradually decreased and consequently, a new emission band emerged at 520 nm. The ratiometric and fluorescence enhancement of compound-1 witnessed the visual detection of Zn²⁺ ions by the naked eye, when the samples were exposed to a UV lamp. The quantum yields of compound-1 increased from 0.55 to 0.69 upon the binding of Zn²⁺. Accordingly, compound-1 could be utilized as an excellent turn-on fluorescence chemo receptor for Zn²⁺ ions (Figure 5). On the other hand, the titration of Cu²⁺ and Ni²⁺ ions completely quenched the fluorescence intensity (Figures 5 and 6). The fluorescence quenching effect by Cu²⁺ and Ni²⁺ might be due to the presence of unpaired electrons and the energy-transfer process from the probe to the open shell d–orbitals of metal ions, which allows the non-radiation attenuation of the excited states to be more rapid and effective. The detection limit of compound-1 toward Zn²⁺, Cu²⁺, and Ni²⁺ was obtained using the calibration curve of emission versus composition, which is an important factor for a better sensor and should be less than the limit set by the U.S. EPA (∼20 μM or less) regulations.³⁴ The detection limits of compound-1 toward metal ions, for example, Zn²⁺, Cu²⁺, and Ni²⁺ were found to be as low as 0.25, 0.30, and 0.34 ppm, respectively. The quenching constant was calculated from the Stern–Volmer equation and found to be 2.0 × 10⁵, 2.4 × 10⁵, and 5.6 × 10⁵ for Zn²⁺, Cu²⁺, and Ni²⁺, respectively (Table S1). Further, the quenching efficiency of compound-1 with Zn²⁺, Cu²⁺, and Ni²⁺ was obtained to be 69, 96, and 98%, respectively, and further, the quantum yields of Zn²⁺, Cu²⁺, and Ni²⁺ strongly supported the quenching efficiency (see the Supporting Information). From these results, one can conclude that the quenching efficiency and quenching constant for Ni²⁺ is high as compared with that of Zn²⁺ and Cu²⁺ ions.

Figure 5.

Change in emission spectra of compound-1 (1 μM) upon the addition of Zn²⁺ (left) and Cu²⁺ (right) in a 1:1 ratio of DMSO and CHCl₃ (inset photographs were taken in the presence of UV light).

Figure 6.

Change in emission spectra of compound-1 (1 μM) upon the addition of Ni²⁺ ions in a 1:1 ratio of DMSO and CHCl₃ (inset photographs were taken in the presence of UV light).

Furthermore, we explored the sensitivity and selectivity of compound-1 toward several metal ions and was compared with Zn²⁺. A 1 μM stock solution of compound-1 in a 1:1 ratio of DMSO and CHCl₃ solutions was treated with excess amounts of diverse metal ions such as Cd²⁺, Cr³⁺, Mn²⁺, Pb²⁺, Ba²⁺, Al³⁺, Ca²⁺, Fe²⁺, Fe³⁺, Mg²⁺, K⁺, and Na⁺ for the investigation of selectivity. Compound-1 exhibited high selectivity and ratiometric as well as fluorescence turn-on response toward Zn²⁺, while Cu²⁺ and Ni²⁺ metal ions displayed fluorescence turn-off, other metal ions do not influence the emission spectra (Figure 7).

Figure 7.

Examination of selectivity (left) and competitive (right) studies of compound-1 (1 μM) toward various metal ions. The competitive binding ability of compound-1 (1 μM) were observed toward Zn²⁺ in the presence of other interfering metal ions and the emission band was monitored at 520 nm. Inset photographs were taken in UV light.

In addition, competitive selectivity of compound-1 with respect to Zn²⁺ in the presence of other interfering candidates such as Cu²⁺, Ni²⁺, Cd²⁺, Cr³⁺, Mn²⁺, Pb²⁺, Ba²⁺, Al³⁺, Ca²⁺, Fe²⁺, Fe³⁺, Mg²⁺, K⁺, and Na⁺ was studied. The initial emission band of compound-1 with excess amounts of various other metal ions exhibited very low and constant intensity which upon the addition of 5 equiv of Zn²⁺ led to efficient enhancement of green emission with good quantum yields (Figure 7). These results strongly suggest the excellent selectivity of compound-1 for Zn²⁺ ions in the presence of other mentioned interfering metal ions.

To get an insight into the photophysical responses of the compound-1 with metal ions, we carried out the quantum mechanical calculations of the free probe and its metal coordinated complexes with the help of density functional theory (DFT) method using a Gaussian 09 program³⁵ at the level of B3LYP/631G/LANL2DZ (d,p) for C, H, N, S, O, and metal ions (Zn²⁺, Cu²⁺, and Ni²⁺). The energy minimized structure of compound-1 and compound-1–Zn²⁺ complexes is shown in Figure 8, whereas for complexes of Cu²⁺ and Ni²⁺ are given in the Supporting Information.

Figure 8.

Selected MOs of compound-1 and compound-1 + Zn²⁺ (not to scale; isovalue = 0.02).

In addition, the frontier molecular orbitals reveal that compound-1 as well as metal complexes of Zn²⁺, Cu²⁺, and Ni²⁺ will provide the valuable information about electronic transitions. Using the optimized structures, time-dependent DFT (TDDFT) calculations were carried out and created a checkpoint file which was used to generate frontier molecular orbital cubes. The highest occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO) of compound-1 were mostly concentrated on the whole π-moiety of benzothiazole as well as biphenyl units with a HOMO–LUMO gap of 4.02 eV, which indicated the transition of π to π* of the benzothiazole unit. As evident from UV–visible and fluorescence titration spectra, the stoichiometry of compound-1 and Zn²⁺, Cu²⁺, and Ni²⁺ was confirmed strongly as 1:2 and hence, the optimization of the metal complex was carried out with 1:2 coordinated complexes taking acetate as an auxiliary ligand and optimized using LAN2DZ basis sets. In the compound-1–Zn²⁺ complex, the HOMO is situated completely on the whole biphenyl–benzothiazole moiety, whereas LUMO is spread over Zn²⁺ ions with a band gap of 3.88 eV (Figure 6). These results clearly revealed the interruption of internal charge transfer after the appendage of Zn²⁺ ions to compound-1. Similarly, the calculated HOMO–LUMO gap of other metal complexes (Cu²⁺ and Ni²⁺) were small compared to that of compound-1. These findings intensely supported the reason for the colorimetric response upon the addition of metal ions (Zn²⁺, Cu²⁺, and Ni²⁺) to compound-1.

Furthermore, the TDDFT of compound-1 and metal complexes (Zn²⁺, Cu²⁺, and Ni²⁺) were carried out in the LAN2DZ basis set to investigate the electronic transition in absorption spectra in vacuum. Further, the compared oscillator strength of compound-1 and its metal complexes provided the valuable evidence for the colorimetric response. It was noticed that the absorption spectrum of compound-1 consists of two major peaks at 295 and 375 nm with energies 3.8894904 and 4.480113 eV for the key electron transitions, respectively. However, TDDFT data showed two major strong bands at 389.37 and 279.24 nm with energies of 3.1842 and 4.4401 eV correspondingly. As evident from TDDFT data, the absorption bands of the Zn²⁺ complex displayed at 411.32 and 309.69 nm and the strong absorption band in the visible region are attributed to intraligand charge-transfer bands, which also matched with the experimental data. These outcomes are strongly supported by the oscillator strength of compound-1 before and after metal complexes with a change from 0.2110 to 1.8996. The formation of the metal ion complex with compound-1 had gradually enhanced the absorption band in the visible region, which might be due to the change in electronic transition and marginal sensitivity of the metal ions. For example, compound-1 and its complexation with Zn²⁺ was found to give a bathochromic shift with 40 nm as noted in UV–visible spectra. On the other hand, the resultant Ni²⁺ complex led to a huge bathochromic shift in the absorption of (Δλ) 52 nm and Cu²⁺ prompts instead similar red shifts (Δλ ≈ 42 nm) as compared with Zn²⁺. Further, these arguments are completely supported by the change in energy gap values, which are observed from DFT studies. In addition, we carried out DFT/TDDFT calculations of compound-1–Cu²⁺ and compound-1–Ni²⁺ complexes under similar conditions and found that the oscillator strength values were relatively unchanged compared with compound-1–Zn²⁺. Also, these data provided further evidence for the observed selectivity of the compound-1 with Zn²⁺.

The strong affinity of compound-1 toward Zn²⁺ and the ratiometric response of compound-1 encouraged us to explore the applicability of compound-1 for the ratiometric fluorescence imaging of Zn²⁺ ions in live cells. Initially, the cytotoxicity of compound-1 toward HeLa cells was studied by the standard MTT assay, explaining the least cytotoxic nature to the cells. Further, HeLa cells were cultured in Dulbecco’s modified Eagle medium (DMEM) and the cells were incubated with compound-1 for 30 min at 25 °C. Later, the cells were imaged through a fluorescence microscope, a high red fluorescence was observed only inside the cellular regions and was clearly supported by the ICT mechanism in biological samples also. Furthermore, 10 μM of Zn²⁺ solution (DMEM) was added prior to the incubation and was kept for 10 min. Later, the cells were washed with PBS buffer solutions to remove an excess amount of Zn²⁺ ions and were subjected to fluorescence microscopy imaging and the bright green emission was observed inside the cells (Figure 9). From these results, one can conclude that compound-1 could be a promising candidate for the practical protocol in ratiometric imaging of living cells in biology.

Figure 9.

Cellular images of Zn²⁺ ions in HeLa cell lines at 37 °C (a) bright field image of compound-1 + Zn²⁺ ion-treated HeLa cells; (b) fluorescence imaging of HeLa cells with compound-1 after 10 min of treatment of 10 μM Zn²⁺ ions; and (c) HeLa cells incubated with compound-1 for 30 min.

A good sensor should exhibit better selectivity as well as reversibility nature; thus, we have examined the reversibility of compound-1 with Zn²⁺ in a 1:1 ratio of DMSO and CHCl₃ solutions. Upon the addition of 5 equiv of the EDTA solution to the Zn²⁺ + compound-1 solution; the original emission of compound-1 was regenerated (Figure 10), which indicated that the EDTA was involved in captivating the Zn²⁺ ion. Cu²⁺ and Ni²⁺ complexes with compound-1 also furnished the same results (Figures 10 and 11). These experiments strongly confirm that compound-1 has very good reversibility and thus, it can be employed as a useful and better chemosensor probe selectively for Zn²⁺ ions.

Figure 10.

Reversibility studies of compound-1 upon the addition of EDTA to compound-1 + Zn²⁺ (left) and compound-1 + Cu²⁺ (right).

Figure 11.

Reversibility studies of compound-1 upon the addition of EDTA to compound-1 + Ni²⁺.

Conclusions

In summary, we have designed and synthesized a biphenyl–benzothiazole-based chemosensor and was characterized by ¹H, ¹³C NMR, as well as HRMS. The novel sensor exhibited ratiometric and colorimetric responses toward biologically important metal ions such as Zn²⁺, Cu²⁺, and Ni²⁺ with significant color changes from colorless to yellow in DMSO/CHCl₃ (50:50, v/v) over other metal cations (Cd²⁺, Cr³⁺, Mn²⁺, Pb²⁺, Ba²⁺, Al³⁺, Ca²⁺, Fe²⁺, Fe³⁺, Mg²⁺, K⁺, and Na⁺). Further, fluorescence studies revealed that there was an enhanced fluorescence in the emission spectra, upon the addition of Zn²⁺ ions and that make compound-1 as a potential ratiometric as well as fluorescence turn-on chemosensor, which was strongly supported by the elevating quantum yields, whereas a turn-off chemosensor for Cu²⁺ and Ni²⁺ ions. The selectivity and competitive studies also strongly suggested that compound-1 was excellent for Zn²⁺ ions in the presence of other mentioned interfering metal ions. Moreover, compound-1 showed the detection limit for Zn²⁺ ions as 0.25 ppm which was lower compared to that of Ni²⁺ and Cu²⁺ ions which were 0.30 and 0.34 ppm, respectively. The binding stoichiometry of the metal complex was determined using a Job's plot and was found to be 1:2 for compound-1 with all the three metal ions. The reversibility of compound-1 was studied upon the addition of EDTA and found that the green color emission of compound-1 was changed to yellow, restoring the emission of compound-1, proving the reversibility of the chemosensor. Similarly, metal complexes of compound-1 with Ni²⁺ and Cu²⁺ ions follow similar trends of Zn²⁺ ions. The ratiometric response of compound-1 toward Zn²⁺ ions extend its applicability in the fluorescence cellular imaging of Zn²⁺ ions in live cells and found a color change from red to green fluorescence in cells. Due to the presence of these novel properties, compound-1 can act as an excellent chemosensor for the detection of biologically important metal ions such as Zn²⁺, Cu²⁺, and Ni²⁺ in the environment as well as bioimaging (Zn²⁺ ions) of live cells.

Experimental Section

All reagents were of analytical grade and used without further purification, otherwise noted the purification process. Glassware was oven dried at 140 °C for at least 2 h prior to use and allowed to cool under vacuum. 4-Phenylphenol, aminothiophenol, (Alfa Aesar), TFA, EtOH, MeOH, diethylether, and DMSO (Spectrochem). HMTA and all the metal salts were used as such without purification (Merck India). Flash chromatography was carried out on Merck Kieselgel 60 (230–400 mesh) as the stationary phase under a positive pressure using AR grade solvents; the procedure includes the removal of solvents under reduced pressure. Thin-layer chromatography was performed on precoated Merck Kieselgel silica gel plates (60 F254, Merck, Germany) monitored by UV light and iodine. Electrospray ionization time-of-flight mass spectrometry (TOF MS ES+) and HRMS were recorded on a Waters QTOFMS Xevo G2 spectrometer in the positive ion mode. The ¹H, ¹³C{¹H} NMR were recorded on a Bruker AVANCE III 400 MHz NMR spectrometer. All solution ¹H and ¹³C spectra were referenced internally to the solvent signal (TMS). In rare instances where other NMR solvents have been used, an appropriate mention has been made in the text. Chemical shifts are expressed in parts per million (ppm) and coupling constants (J) are in Hz. The ¹H and ¹³C{¹H} NMR spectra were referenced using residual H impurities in the deuterated solvents. Deuterated solvents (CDCl₃) were purchased from Cambridge Isotope Laboratories, dried over calcium hydride, degassed by three freeze–pump–thaw cycles, and vacuum-transferred prior to use. Electronic absorption spectra were recorded on a PerkinElmer LAMBDA 750 UV/visible spectrophotometer. The solutions were prepared using a microbalance (±0.1 mg) and volumetric glassware and then charged in quartz cuvettes with sealing screw caps. Fluorescence emission studies were carried out on a Horiba JOBIN YVON Fluoromax-4 spectrometer. All aqueous solutions were prepared with double distilled deionized water. The standard stock solution of Ag⁺, Co²⁺, Pb²⁺, Zn²⁺, Cd²⁺, Cr³⁺, Mn²⁺, Pb²⁺, Ba²⁺, Al³⁺, Ca²⁺, Fe²⁺, Fe³⁺, Mg²⁺, K⁺, Cu²⁺, Na⁺, Cd²⁺, and Hg²⁺ was prepared by dissolving their nitrate salts.

Synthesis of 4-Phenyl 2,6-Diformylphenol (A)

4-Phenylphenol (5 g, 29 mmol) was dissolved in TFA (100 mL). Hexamethylene tetramine (32 g, 232 mmol) was added to the solution and refluxed at 160 °C for 48 h (Scheme 1). The reaction mixture was stirred in 6 N HCl (100 mL) for 30 min. The crude reaction mixture was extraction with DCM and purified through silica gel column chromatography (2:8 ratios of ethyl acetate and hexanes). A yellow colored powder was obtained with yield 78%. ¹H NMR (400 MHz, CDCl₃) δ 11.63 (s, 1H), 10.32 (s, 2H), 8.19 (s, 2H), 7.59 (d, J = 7.3 Hz, 2H), 7.48 (t, J = 7.5 Hz, 2H), 7.41 (d, J = 7.3 Hz, 1H) (the ¹H NMR data was matching with literature reports). HRMS (EI): calcd for C₁₄H₁₀NaO₃ [M + Na]⁺m/z, 249.0528; found m/z, 249.0532.

Synthesis of Compound-1

A stirred mixture of 4-phenyl 2,6-diformylphenol (0.5 g, 2.2 mmol) and aminothiophenol (0.7 g, 5.5 mmol) in 60 mL of EtOH was refluxed for 24 h. The crude product was purified by column chromatography using silica gel and solvent was dried under vacuum. A dirty white product was obtained with 71% yield. ¹H NMR (400 MHz, CDCl₃): δ 14.24 (s, 1H), 8.48 (s, 2H), 8.10 (d, J = 8.0 Hz, 2H), 7.98 (d, J = 7.5 Hz, 2H), 7.80–7.71 (m, 2H), 7.61–7.48 (m, 4H), 7.49–7.39 (m, 3H). ¹³C NMR (101 MHz, CDCl₃): δ 155.6, 151.8, 139.6, 133.2, 128.9, 127.5, 127.1, 126.6, 125.5, 122.6, 121.6, 116.7. HRMS (EI): calcd for C₂₆H₁₇N₂OS₂ [M + H]⁺m/z, 437.0782; found m/z, 437.0780.

Supporting Information Available

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acsomega.1c02855.

• ¹H NMR, ¹³C NMR, HRMS, and characterization data for all the compounds, photophysical data, association constants, detection limits, and DFT results (PDF)

The authors declare no competing financial interest.