Elsevier

Talanta

Volume 125, 1 July 2014, Pages 418-424
Talanta

“Turn-on” fluorescent dipodal chemosensor for nano-molar detection of Zn2+: Application in living cells imaging

https://doi.org/10.1016/j.talanta.2014.03.002Get rights and content

Highlights

  • A malonohydrazide derivative bearing an imine and phenolic group was synthesized and had a high affinity and selectivity towards Zn2+.

  • The recognition properties of receptor 1 were evaluated using absorbance and fluorescence spectroscopy.

  • The 1:1 stoichiometry of 1.Zn2+ was confirmed from Job׳s plot, mass spectra and density functional theory (DFT).

Abstract

A malonohydrazide derivative bearing an imine and phenolic group was synthesized and had a high affinity and selectivity towards Zn2+. The recognition properties of receptor 1 were evaluated using absorbance and fluorescence spectroscopy. The 1:1 stoichiometry of 1.Zn2+ was confirmed from job׳s plot, mass spectra and density functional theory (DFT). The detection of Zn2+ in intracellular environment of HeLa cell through confocal microscopy was successfully applied for live cell imaging.

Introduction

In recent years, fluorophores gained a considerable space in the field of supramolecular chemistry due to score of applications like sensing and detection of transition metal ions, cell imaging, environmental and clinical roles [1], [2], [3], [4], [5], [6]. Among transition metal ions, Zn2+ is the second most-abundant metal ion in our body and led several biological functions like influencing DNA synthesis, gene expression, enzyme catalysis, apoptosis, immune system function and neuronal signal transmission [7]. A disturbance in level of Zn2+ in our body cause several diseases like amyotrophic lateral sclerosis, epilepsy, Alzheimer׳s disease, loss of taste and hypoxia ischemia [8]. To date, several attempts have been made for the detection of Zn2+; due to its electronic configuration (3d10 4s0) only possible technique to detect Zn2+ in biological system is fluorescence spectroscopy [9]. Generally, available probes sense the analyte through an increase or decrease of the emission intensity [10]. However, the emission intensity is also dependent on other factors, such as the sensor concentration, bleaching, optical path length, and illumination intensity [11]. It is therefore desirable to eliminate the effects of these factors by using a ratiometric sensor that exhibits a spectral shift upon binding to the analyte of interest, so that the ratio between the two emission intensities can be used to evaluate the analyte concentration [12], [13], [14], [15], [16], [17], [18], [19].

Here we are paying attention for the development of ratiometric fluorescent sensor that utilized photo-induced electron-transfer (PET) and intramolecular charge-transfer (ICT) effect. The combination of PET and ICT produced the visual change for the detection of Zn2+ because PET produced enhancement or quenching in the intensity and ICT give shift in the absorption and emission wavelength. Generally these types of molecule are used as an anion sensor due to the presence of amine and hydroxyl group [17]. However, we are reporting PET and ICT based ratiometric fluorescent sensor 1 for the detection of Zn2+. Coordination sites in the receptor 1 show that there are two amine groups and imine linkages with hydroxyl groups (Scheme 1). The binding ability of receptor 1 with various cations such as Cr3+, Mn2+, Fe3+, Co2+, Ni2+, Cu2+, Zn2+, Cd2+, Hg2+, Pb2+ and Bi3+ was evaluated in DMSO/H2O (50:50, v/v); computed the binding constant for 1.Zn2+ and optimized its geometry through using Gaussian09 program.

Section snippets

Reagents and apparatus

All commercial grade chemicals and solvents were purchased from Sigma-Aldrich Chemicals Ltd and used without further purification.1H and 13C-NMR spectra were recorded on a Varian NMR mercury System 300 spectrometer operating at 300 and 75 MHz in DMSO-d6. The fluorescence spectra were recorded on Fluoromax-4 spectrofluorometer and UV–visible spectra on Shimadzu UV–24500 UV–visible spectrophotometer at room temperature using 1 cm cell. The whole set of experiments were performed using DMSO/H2O

Synthesis of receptor 1

Receptor 1 was synthesized by reaction of diethyl malonate with hydrazine hydrate to obtain malonohydrazide and further it was reacted with salicyladehyde in ethanol with stirring for 1.5 h (Scheme 1). The receptor 1 was obtained with quantitative yield and having white appearance. The synthesized receptor was characterized by various techniques such as melting point, IR, 1H-NMR, 13C-NMR and mass spectroscopic methods. The spectral investigation gave consistent data of structure of receptors 1.

UV–visible studies of receptor 1

Cellular imaging study

Inspired from the above results like ON-SITE detection, visual color change and high selectivity for Zn2+, receptor 1 was used for intracellular detection of Zn2+ in HeLa cell Line. To perform this study, HeLa cell was incubated in three different medium (a) enrich with receptor 1, (b) Zn2+ only (c) both receptor 1 and Zn2+ respectively at 37 °C in a humidified atmosphere with 5% CO2 [31]. It is observed that blue fluorescence was seen in case of those cells, which was incubated with both

Conclusion

In conclusion, we have designed and developed a selective and sensitive ratiometric fluorescent receptor 1 for the detection of Zn2+ in semi aqueous medium. The PET and ICT was possible mechanism behind this detection. The binding constant and detection limit revealed that receptor 1 has high selectivity towards Zn2+ and detect the Zn2+ upto 35 nM. Sensing in living cells have been developed by this probe for in vitro imaging. This new Zn2+ selective fluorescent probe could find potential

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