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Journal of Fluorescence

, Volume 28, Issue 6, pp 1281–1294 | Cite as

Synthesis and Evaluation of a Schiff-Based Fluorescent Chemosensors for the Selective and Sensitive Detection of Cu2+ in Aqueous Media with Fluorescence Off-On Responses

  • Maria SadiaEmail author
  • Robina Naz
  • Jehangir Khan
  • Rizwan Khan
ORIGINAL ARTICLE
  • 195 Downloads

Abstract

Copper being an essential nutrient; also pose a risk for human health in excessive amount. A simple and convenient method for the detection of trace amount of copper was employed using an optical probe R1 based on Schiff base. The probe was synthesized by Schiff base condensation of benzyl amine and 2-hydroxy-1-napthaldehyde and characterized by single X-ray diffraction, 1H NMR and FTIR. By screening its fluorescence response in a mixture of DMSO and H2O (20:80, v/v) R1 displayed a pronounced enhancement in fluorescence only upon treatment with copper. Other examined metal ions such as alkali, alkaline and transition had no influence. Within a wide pH range 5–12 R1 could selectively detect copper by interrupting ICT mechanism that results in CHEF. From Job’s plot analysis a 2:1 binding stoichiometry was revealed. The fluorescence response was linear in the range 1–10 × 10−9 M with detection limit 30 × 10−9 M. Association constant was determined as 1 × 1011 M−2 by Benesi-Hilderbrand plot. As a fast responsive probe it possesses good reproducibility and was employed for detection of copper in different water samples.

Keywords

Fluorescent chemosensor Schiff base Copper CHEF Real-time detection 

Introduction

In the last few years, the development of fast, cheap and efficient fluorescent chemosensors with high sensitivity and selectivity for the detection of heavy and transition metal ions such as Cu2+, Cd2+, Pb2+, and Hg2+ has attracted wide-spread interests of chemists, environmentalists, clinical biochemists and biologists because of their fundamental role in biological, environmental and medicinal application [1, 2, 3, 4, 5, 6].

Fluorescent chemosensors typically consist of two parts: ionophore (metal chelator) and fluorophore (signaling unit). The two parts are interconnected through a proper spacer. Ionophore is required for analyte binding and as a result of metal binding, photophysical properties of the fluorophore such as fluorescence intensity, lifetime of the excited state or absorption wavelength changes, achieving the purpose of identification [7, 8, 9].

Among the metal ions, after zinc and iron, copper is the third most abundant vital trace element for life that plays an important role in various fundamental physiological and biological processes in organisms [10]. It is also a toxic chemical that is not biodegradable and bioaccumulative and can accumulate in food chain or human through uptake or consumption and may be hazardous to human health or the environment [11]. Copper is present in sediments, natural water, and in the other medium such as air and soil. Industrial effluent are the potential source of copper contamination such as waste water from industries manufacturing fungicides, fertilizers, bactericides, algaecides, electronic goods, copper plumbing, as well as the use of copper as a mordant for textile dyes, as an activator in froth floatation of sulfide ores and in plating by products. Natural sources also contribute to the contamination of copper that include sea spray, forest fires, decaying vegetation, volcanoes and windblown. [12, 13, 14]. Due to these extensive industrial applications copper can be a significant environmental pollutant of worldwide concern [15].

Copper is also an essential micronutrient in the human body. Because of its redox-active nature it acts as an essential cofactor of many enzymes such as tyrosinase, cytochrome c oxidase and superoxide dismutase [16]. It is necessary for the development of bone, nerve coverings, and connective tissue [17].

Copper exist in three common oxidation states Cu2+ (cupric ion), Cu+ (cuprous ion) and Cu0 (metal). Among the three species the most commonly occurring and toxic to living organism is the cupric ion. Copper possess high affinity for the ligand containing sulfur and nitrogen donors thus it effects enzymes whose activities depend on amino and sulhydryl groups [18]. Due to its toxicity at cellular level, its distribution and homeostasis is highly controlled by various factors including chaperones and copper transport proteins. Abnormal level of copper can cause various problems and disorders. For example, under overloading condition copper can cause disorders such as allergies, migraine headaches, anxiety, depression, anorexia, premenstrual syndrome, fatigue, kidneys and blood problems, Wilson’s, Parkinson’s, Alzheimer’s and prion diseases in humans and its deficiency is associated with myelopathy. In living cells uncontrolled reaction of copper ion with oxygen result in the production of ROS (reactive oxygen species) that can damage proteins, nucleic acid and lipids Excessive absorption of Cu2+ also effects the growth of plants including fewer leaves, shortening of root length and decrease in the plants biomass [19, 20, 21, 22]. According to World Health Organization (WHO), the permissible limit of copper in drinking water is 2 ppm (30 μM) or 1.5 mg/L [23, 24]. In view of such toxic effects of copper, efficient detection of trace amount of copper in environmental water samples is very necessary. For these reasons, great effort has been dedicated to design fluorescent chemosensors for specific recognition and detection of copper.

Chelation enhanced fluorescent type chemosensor are more sensitive to metal ions as compared to Chelation enhanced quenching chemosensors. Cu2+ is known as a fluorescence quencher and most of the fluorescent chemosensors reported for copper so far, detection of copper occur through a fluorescence quenching process that undergoes a charge or energy transfer mechanism and “turn-on” fluorescent sensors for recognition of copper are still rare. Thus, there is a great demand to develop “turn-on” fluorescent chemosensors for specific and sensitive detection of copper [25, 26, 27].

Schiff base derivatives have attracted much attention as an optical chemosensor for the detection of the metal ions because of their structural flexibility, excellent applicability, high selectivity, high efficiency and simplicity of synthetic mode. Schiff base compounds form strong complexes with transition metal ions. For the Schiff base to be used as a fluorescent sensor, the presence of a strong fluorophore is required. Schiff base compounds incorporating π- conjugated fluorescent fragment are extensively used as fluorescent chemosensor for the metal ions [28, 29, 30, 31].

It was within our interests to design a CHEF type Schiff-base fluorescent sensor for selective detection of copper ion. Therefore we designed and develop a new fluorescent probe for copper ion named as (E)-1 ((benzylimino)methyl)naphthalen-2-ol (R1). The synthesis was simple, single step the product was highly selective towards copper ion. R1 exhibited very weak fluorescence due to ICT mechanism which was inhibited upon interaction with copper ion and enhancement of fluorescence was observed. The proposed fluorescent chemosensors contain two parts naphthalene group which is the signaling moiety acting as a fluorophore and imine and hydroxyl group are the binding sites acting as a receptor. We select 2-Hydroxynaphthalene as a fluorophore because of its cheap cost, good binding sites, competitive stability in the environment and characteristic photophysical properties such as low fluorescence quantum yield and short fluorescence lifetime [32, 33].

Experimental

Materials and Instrumentation

2-hydroxy-1-napthaldehyde, Benzyl amine, Dimethyl sulfoxide (DMSO), Hydrochloric acid 37%, Sodium hydroxide, Acetone, and salts of Cu2+, Na+, Mg2+, K+, Zn2+, Mn2+, Hg2+, Ba2+, Pb2+, Ni2+, Cd2+, Ni2+, Co2+, As3+, Fe3+ and Ce3+ were purchased from commercial sources and were used without further purification. Throughout all experiments distilled water was used. pH adjustments were made by using HCl or NaOH. To minimize the release of cations in the solution and their sorption, the glassware’s were first washed with acid and then rinsed with distilled water. For fluorometric determination of Cu2+ optically four sides’ clear quartz cuvettes were used and were washed with acetone before use.

The 1H NMR spectra of the samples were obtained on Bruker Avance 400 MHz. spectrometer. For FTIR spectra samples were prepared as KBr pallets and the spectrum was obtained on FTIR spectrophotometer Pretige 21 Shimadzu Japan in the region 400–4000 cm−1. X-ray structure analysis of the sample was conducted using Bruker kappa APEXIICCD diffractometer. For the measurement of melting points a Bicote Stuart-SMP 10 Japan was used. UV–vis absorption spectroscopy measurements were conducted on UV-visible 1800 spectrophotometer. The emission spectra were monitored using fluorescence spectrophotometer RF 5301 PC Shimadzu Japan equipped with fluorescence free quartz cuvettes of 1 cm path length. Slit width were 5 nm both for excitation and emission and a steady state 150 W Xenon lamp was used as an excitation source. The pH of the test solution was monitored by a BANTE instrument pH meter. All measurements were conducted at room temperature.

Synthesis of Chemosensor R1

R1 was prepared according to the published method [34, 35]. The synthetic route of R1 is shown in the Scheme 1.2-hydroxy-1- napthaldehyde 10 mmol (1.7218 g) was dissolved in 10 mL methanol and after adding few drop of acetic acid, it was boiled for a while. After 5 min methanolic solution of benzylamine 10 mmol (1.09 mL) was slowly added to it drop wise. The mixture was refluxed for about 12 h at 80 °C to give a clear yellow solution. To get the crystal, solvent was evaporated slowly at room temperature by keeping the solution in air for several days. After evaporation of the solvent rod shaped yellow color single crystal were obtained which were recrystallized from ethanol. Yield 80%, mp 120–122 °C, IR (KBr, cm−1): νC-OH, 1296, 1207; ν C=N, 1614, νaromaticC=C, 1492, 1435; νaromatic-C-H, 3035. 1HNMR (400 MHz, DMSO): δ14.5 s (NHO), 9–10s (-N=CH, 1H), 7.7–8.2d (H-naphtha, 2H), 6.0–8.0 m (ArH, 9H), 4.5–5.0 s (ArCH2, 2H).
Scheme 1

Synthetic route of chemosensor R1

General Information

Melting point was determined in open mouth capillary and uncorrected. For spectroscopic investigation analytical reagent grade DMSO and distilled water were used. Fluorescence and absorption spectra of the chemosensor R1 was recorded in DMSO:H20 (20:80) at room temperature. Fluorescence-sensing property of the chemosensor R1 towards metal ions was conducted using sulphate salt of Cu2+, Mg2+, Zn2+, Mn2+; nitrate salt of Pb2+, Ba2+, Cd2+, K+, Ce3+, Hg2+; bromide salt of Co2+; chloride salt of Na+, Ni2+, Fe3+ and oxide of As3+. For pH adjustment HCL and NaOH were used. From the fluorescence intensity data association constant was determined by Benesi-Hilderbrand Plot. The excitation and emission wavelength were 320 and 645 nm respectively. All the measurements were conducted at room temperature.

Results and Discussion

Synthesis and Characterization

Chemosensor R1 was obtained with good yield by condensing 2-hydroxy-1- napthaldehyde with benzylamine in methanol as shown in scheme 2 and it was well characterized by FTIR, 1H NMR and single X-ray crystallography and spectroscopic tools.
Scheme 2

Chemical structure and synthesis of (E)-1-((benzylimino)methyl)naphthalen-2-ol

Characterization by FTIR

In order to determine the functional group in the synthesized Schiff-based fluorescent chemosensor R1 FTIR spectroscopy was conducted and the data is given in the Table 1.
Table 1

FTIR spectral frequencies for chemosensor R1

IR-band (C=N) cm−1

IR-band (C-OH) cm−1

IR-band aliphatic (-C-O) cm−1

IR-band aromatic (-C-H) cm−1

IR-band aromatic (C=C) cm−1

1614

1296, 1207

1138, 1038

3035

1492, 1435

In the region 400–4000 cm−1 several absorption bands were observed in the FTIR spectrum of the chemosensor R1. Assignments of characteristics bands correspond to various functional groups present in the chemosensor were made by comparison method. The FTIR spectrum confirms the formation of imine bond (-C=N) and no band assigned to (C=O) was detected. Thus, the disappearance of carbonyl (-C=O) peak in the region of 1700 cm−1 and appearance of the (-C=N) peak in the region of 1614 cm−1 confirms the formation of chemosensor R1. The band at 3035 cm−1 corresponds to the stretching vibration of aromatic-C-H. The spectrum exhibit absorption band at 1492, 1435 and at 1296, 1207 cm−1 typically of aromatic-C=C and phenolic-O-H stretching vibration. The bands at 1138 and 1038 cm−1 correspond to aliphatic-C-O [36, 37, 38].

Characterization by Single Crystal X-Ray Diffraction

The synthesis of chemosensor R1 was a single-step straight forward reaction and no side products were observed or isolated during the course of reaction. From reaction mixtures, by slow evaporation of the solvent crystals were separated, Suitable for single X-ray diffraction analysis. Crystallographic measurement were carried outs with Bruker kappa APEXIICCD diffractometer, with graphite-monochromator (Mo-kα radiation (λ = 0.71073o A) at ambient temperature. For data collection ωscan and multi-scan correction were applied. The crystal structure solution and refinements of chemosensor R1 were handled with SHELXL-97 [39] and publ CIF [40]. Full-matrix least-squares techniques were done for final refinement on F2. The molecular view of chemosensor R1 with partial numbering scheme is shown in Figs. 1 and 2. The crystallographic data reveals to crystal structure of chemosensor R1 and relevant refinement parameters are given in Table 2 and the selected bond angles and bond lengths are listed in Table 2.
Fig. 1

FTIR spectrum of the chemosensor R1

Fig. 2

Molecular structure of chemosensor R1 with partial numbering scheme

Table 2

Crystallographic data and details of refinements for chemosensor R1

Crystal

R1

Empirical formula, Formula weight

C18H15NO, 261.31

Crystal system, Space group

Monoclinic, P21/c

Temperature (K)

296

a, b, c, (Å)

25.391 (4), 6.5797 (11), 8.1779 (13)

β (o)

92.277 (9)

V3)

1365.2 (4)

Z

4

Radiation type

Mo Kα

μ (mm−1)

0.08

Crystal

Yellow, crystal size (0.42 × 0.30 × 0.26)

Data collection

 Diffractometer, scan mode

Bruker kappa APEXIICCD, Multi-scan and ωscan

 No of measured, independent and observed [I ≥ 2σ(I)] reflections

5051, 2746, 1772

Rint

0.040

  (sin θ/λ)max−1)

0.651

Refinements

R [F2 > 2σ(F2)], wR (F2), S

0.050, 0.126, 0.96

 No. of reflections

2746

 No. of restraints

2

 H-atom treatment

H-atom parameter constrained

 Δ ρmax, Δρmin (e Å−3)

0.16, −0.17

 Absolute structure

Flack x determined using 598 quotients [(I+)-(I-)]/[(I+) + (I-)] (Parsons, Flack and Wagner, Acta Cryst. B69 (2013) 249–259)

 Absolute structure parameter

−0.7 (10)

The crystal of R1 is mono-clinic crystal system with space group P21/n. In the crystal amine moiety through 84.74o is twisted with respect to naphthalene moiety. Around N1 the geometry is trigonal planner, demonstrating that the lone pair of nitrogen is involved in conjugation without disturbing sp2 hybridization. The bond distance between N1-C11 is 1.295 Å which is comparable with the double bond character of C=N bond and bond length of N1-C12 is 1.464 Å and C1-O is 1.267 Å. X-ray diffraction map revealed that imine-N is bonded with H and exists as –C=NH+– and O of phenolic –OH as O which is identical with similar reported structure [33]. Intramolecular hydrogen bonding stabilized the structure as shown in the Fig. 3.
Fig. 3

Part of 3D network of chemosensor R1 having Intramolecular hydrogen bonding

Characterization by 1H NMR

The 1HNMR spectrum of the chemosensor R1 displays a proton at δ14.5 which shows the interaction of imine nitrogen with the OH proton resulting in zwitter ion as shown in the Fig. 4. A singlet proton at δ9–10 attributed to -N=CH represents the imine group (Schiff base). Similarly two doublet protons exhibited at δ7.7–8.2 are assigned for the naphthalene. Further aromatic group protons at δ6.0–8.0 represent the benzene and naphthalene rings. A pair of singlet protons at δ4.5–5.0 are assigned for the ArCH2.
Fig. 4

1H NMR spectrum of chemosensor R1

Preparation of Stock and Working Solutions of Metals and Chemosensor R1

For practical utilization and for detection of toxic metal ions in water samples it is very necessary for a chemosensor to work under aqueous condition. Chemosensor R1 was not soluble in pure water. Therefore 0.001 M stock solution of chemosensor R1 was prepared in pure DMSO. The chemosensor R1 was stable in DMSO. Working solutions were prepared by dilution of a stock solution in DMSO with a mixture of DMSO:H2O (20:80). Stock solutions of metal ions (0.01 M) were prepared in distilled water. Before spectroscopic analysis, test solutions were freshly prepared by appropriate dilution of stock solution to the corresponding desired concentration.

Metal Ion Binding Study by Fluorescence Spectroscopy

First the fluorescence of chemosensor R1 10 μM was investigated in mixed aqueous medium DMSO:H2O (20:80) by taking 3 mL solution of it in a fluorometric cell. The fluorescence emission spectrum of this solution was taken upon excitation at 320 nm. Due to internal charge transfer its fluorescence emission was quenched and very weak fluorescence was observed at 645 nm. To investigate cation sensing ability of R1, 2 equiv. of Cu2+ and 20 equiv. of various alkali (Na+, K+), alkaline (Mg2+, Ba2+) transition metal ions (Ce3+, Fe3+, As3+, Hg2+, Zn2+, Cd2+, Pb2+, Mn2+, Co2+, Ni2+) were added separately to the solution of R1 (2 mL). The mixture were allowed to equilibrate for 2 min at room temperature and then transferred separately into quartz cuvette and the fluorescence emission spectrum was monitored in the range 250–800 nm with an excitation at 320 nm and emission at 645 nm. The spectrum of all the examined metal ions was compared with the spectrum of R1. It was found that the non-fluorescence behavior become highly fluorescent only in the presence of Cu2+ ions and no detectable change in the fluorescence spectra was observed by other metal ions except a significant increase in the initial fluorescence intensity of R1 only upon addition of Cu2+ as shown in Figs. 5 and 6. So among the tested metal ions only Cu2+ was the one that readily binds with R1. The increase in the fluorescence emission of R1 can be explained on the basis of restriction of internal charge transfer (ICT) mechanisms. In the absence of Cu2+ ions, due to ICT mechanism, the fluorescence emission of the naphthalene fluorophore is greatly quenched. But in the presence of Cu2+ ions ICT phenomenon is restricted and fluorescence emission was considerably enhanced as result of rigid chelated complex formation [33].
Fig. 5

Fluorescence emission spectrum of R1 and relative fluorescence intensity changes upon the addition of monovalent (Na+, K+), divalent (Hg2+, Zn2+, Cd2+, Pb2+, Mn2+, Co2+, Ni2+, Cu2+) and trivalent cations (Ce3+, Fe3+, As3+) as their aqueous solutions with an excitation at 320 nm, temp 25 °C

Fig. 6

Turn-On fluorescence behaviour of R1 and the proposed binding mode and mechanism

UV-Visible Study

Since chemosensor R1 was not soluble in 100% aqueous media therefore DMSO was used as a cosolvent. UV-vis spectra of chemosensor R1 was taken in a mixed aqueous media in the absence and presence of different concentration of Cu2+ ions in distilled water. The UV-vis spectrum of chemosensor R1 exhibited characteristic main absorption band in the range 260–380 nm with a λmax of 302 nm as shown in Fig. 7a. Upon addition of Cu2+ ions to chemosensor R1 λmax shifts to 308 nm. The red-shift of the λmax is indicative of the coordination involving the hydroxyl and imine as a receptor and Cu2+ as an analyte. When various concentration Cu2+ ions were added to the solution of chemosensor R1 the absorbance at 308 nm was enhanced as shown in the Fig. 7c. Test samples were prepared by using 10 μM solution of chemosensor R1 and various concentrations of Cu2+ ions. All the measurements were made at room temperature.
Fig. 7

a absorbance of 10 μM chemosensor R1. b Shifting of wavelength upon addition of Cu2+ ions, concentration of chemosensor R1 and Cu2+ ions is 10 μM. c Enhancement of absorbance at 308 nm upon addition of Cu2+ ions in the range 1–10 μM

Fluorogenic Detection of Cu2+

To investigate the fluorescence-sensing behavior of chemosensor R1, a quantitative analysis of the binding affinity of chemosensor R1 with Cu2+ ion was studied by fluorescence emission spectroscopy. To address the sensitivity, the fluorescence spectral properties of chemosensor R1 were examined in DMSO:H2O (20:80) as a function of concentration of Cu2+ ion added at room temperature. When excited at 320 nm the fluorescence emission spectrum of chemosensor R1 displayed was very weak emission at 645 nm due to ICT mechanism in naphthalene moiety. However, the addition of Cu2+ ion in the range 1–10 μM led to a rapid increase in the fluorescence emission intensity at 645 nm as shown in Fig. 8 while no significant change in position of λex and λem was observed. The enhancement of fluorescence was attributed to the restriction of ICT mechanism. The increase in fluorescence constitutes the basis for the detection of Cu2+ ion with the chemosensor R1 proposed in this work. We also determined the linearity by plotting the emission intensity of chemosensor R1 at 645 nm as a function of the Cu2+ ion concentration in the range 1–10 μM. Chemosensor R1 was sensitive to Cu2+ ion and the increase in fluorescence had a linear behaviour with correlation coefficient of R2 = 0.9978 as shown in Fig. 9.
Fig. 8

Changes in the fluorescence emission spectra of chemosensor R1 10 μM with increasing amount of Cu2+ ion at 25 °C, λex 320 nm, [Cu2+] from left to right (1, 2, 3, 4, 5, 6, 7, 8, 9, 10 μM

Fig. 9

Fluorescence emission profile of chemosensor R1 versus concentration of Cu2+ ion at 645 nm

Detection Limit

The detection limit of chemosensor R1 as a fluorescent sensor for the recognition of Cu2+ ion was determined from the plot of fluorescence emission intensity as a function of Cu2+ ion concentration. The detection limit was calculated based on 3σ/k [41], where σ is the standard deviation of blank measurement and, and k is the slope of the plot. To determine σ, the fluorescence emission intensity of chemosensor R1 was measured six times independently in the absence of copper ions. The detection limit calculated was found to be 30 × 10−9 M which is far below the WHO acceptable limit (30 μM or 1.5 mg/L of Cu2+) in drinking water. The detection limit of the proposed chemosensor R1 is lower than the fluorescent chemosensors of ref. [11, 42, 43, 44, 45]. The detection limit was sufficiently low to recognize Cu2+ in nano molar range. In term of the detection limit and linear range it can be seen that the proposed sensor displays more sensitivity and selectivity for Cu2+ ion by fluorescence spectra.

Determination of Binding Stoichiometry and Association Constant

For determining the stoichiometry of chemosensor R1 with Cu2+ ion the Job’s plot experiment was undertaken in solution state using fluorescence spectroscopy. For spectroscopic analysis equimolar solutions of chemosensor R1 and Cu2+ ion were prepared and mixed in different ratios by continuous increase of one constituent with the similar decrease of second constituent keeping total concentration constant. The sum of concentration of chemosensor R1 and Cu2+ ion was kept constant at 10 μM according to the continuous variations, changing the mole fraction of chemosensor R1 from 0.1 to 0.9 in a solution of [R1] + [Cu2+]. Binding stoichiometry was determined by plotting the graph between the molar fraction of chemosensor R1 and respective fluorescence emission intensity. When molar fraction reached to 0.6, maximum fluorescence emission was achieved at 645 nm, suggesting that the chemosensor R1 and Cu2+ formed a 2:1 stoichiometry complex as shown in Fig. 10. It means one Cu2+ ion bind with two chemosensor R1 molecules.
Fig. 10

Job’s plot of R1-Cu2+ complex at 645 nm, [R1] + [Cu2+] = 10 μM, λex is 320 nm, showing a binding stoichiometry of 2:1

On the basis of 2:1 stoichiometry as determined by Job’s plot the association constant was calculated with the help Benesi–Hildebrand plot from the increasing fluorescence emission intensity of chemosensor R1 as a function of Cu2+ concentration. The association constant of the complex was calculated through the Benesi-Hildebrand equation stated below
$$ \frac{{\mathrm{F}}_{\max -}{\mathrm{F}}_{\mathrm{o}}}{\mathrm{F}-{\mathrm{F}}_{\mathrm{o}}}=1+\frac{1}{\mathrm{K}{\left[{\mathrm{Cu}}^{2+}\right]}^2\ } $$
(1)
Where Ka (M−2) is an association constant. F0 is the fluorescence emission intensity of chemosensor R1 in the absence of Cu2+, Fmax is the fluorescence emission intensity of chemosensor R1 at [Cu2+] in large excess and F is the fluorescence emission intensity obtained at different [Cu2+] (λex = 320 nm and λem = 645 nm. The association constant Ka was determined graphically by plotting Fmax-F0/F-F0 versus 1/[Cu2+] as shown in Fig. 11. According to the Eq. (1) data were linearly fitted showing a good liner relationship with slope (1 × 10−11), and intercept = (1.4677) and confirming 2:1 complexation. The value of Ka value was determined from the slope and intercept of the line.
Fig. 11

Benesi-Hilderbrand plot for determination association constant of Cu2+ (1–10 μM) with chemosensor R1.The excitation wavelength is 320 nm and the emission wavelength is 645 nm

Effect of Chemosensor R1 Concentration on Fluorescence Response

The effect of an increasing concentration of chemosensor R1 on fluorescence spectra of R1-Cu2+ complex was also investigated. Working solution for analysis were prepared in presence of Cu2+ ion (10 μM) by addition of various concentration of chemosensor R1 solution in an incremental fashion in the range 1–15 μM. The fluorescence emission intensity of each sample was monitored separately at 645 nm and was plotted versus concentration of chemosensor R1 as shown in Fig. 12. The fluorescence emission intensity enhanced gradually with increasing concentration of chemosensor R1 but at higher concentration the fluorescence remain steady. Further replicate analyses were conducted to figure out reproducibility using constant concentration of chemosensor R1 and Cu2+ ion (10 μM). Relative standard deviation (RSD) was 0.24% for 10 repeated fluorescence measurements.
Fig. 12

Fluorescence emission intensity at 645 nm as a function of concentration of chemosensor R1. Copper concentration was kept constant (10 × 10−6 M)

Effect of Time

Response time is very important parameter of a fluorescence chemosensor therefore the kinetic studies of chemosensor R1 (20 μM) both in the absence and presence of Cu2+ (2 μM) was investigated by fluorescence spectra. For this purpose the fluorescence emission intensities were measured at different interval of time. The time was varied in the range 1–15 min. In the absence of Cu2+ at 645 nm almost no changes in the fluorescence emission intensity of chemosensor R1 was found as displayed in Fig. 13, demonstrating stability of chemosensor R1 in the assay condition. In fluorescent spectroscopy long response time is undesired. The response of R1 was almost instantaneous and fluorescence emission intensity turn on remarkably at 645 nm upon addition of aqueous solution of Cu2+ to solution of chemosensor R1, suggesting the rapid reaction of chemosensor R1 with Cu2+. For higher concentration of Cu2+ chemosensor R1 has a fast response time of less than 30 s. This result indicates that the recognition was completed immediately without any detectable time delay after addition of Cu2+. The chemosensor R1 is highly suitable for real-time detection of Cu2+ in aqueous solutions in practical analysis.
Fig. 13

Time dependent fluorescence emission intensity of chemosensor R1 in the absence and presence of Cu2+ at 645 nm, λex 320 nm

Effect of pH

For practical utilization the effect of pH on the fluorescence emission intensity was investigated to determine a suitable pH range for sensing of chemosensor R1 to Cu2+. The fluorescence emission intensity measurements were performed in the absence and presence of 10 μM Cu2+. The pH of the solutions was adjusted using HCl and NaOH and the alteration in fluorescence emission intensity at 645 nm was monitored in the pH range 2–12. The fluorescence emission intensity versus pH plot for the chemosensor R1 and R1-Cu2+ complex is shown in Fig. 14. As shown in Fig, the fluorescence emission intensities of the metal free chemosensor R1 remain unaffected in the pH range 2–12. This result indicate that the chemosensor R1, which is organic solvent-stable and also pH-stable, can be employed for recognition of Cu2+ as a useful potential fluorescent sensing material. After the addition of Cu2+ to the solution of chemosensor R1 the fluorescence emission intensity at 645 nm rapidly enhanced in the pH range 5–12. As it is cleared from the Fig. 14, in the section of lower pH value (pH < 5) the emission intensity remain in the quenched state indicating poor stability of the R1-Cu2+ complexes at low pH. This is due to protonation on the chemosensor R1 that prevents the formation of the R1-Cu2+ complex. The fluorescence emission intensity was in the turn-on state for pH > 5 indicating maximum complexation at high pH values and good fluorescence detection ability to Cu2+.
Fig. 14

Fluorescence response of chemosensor R1 in the presence and absence of Cu2+ as a function of different pH values at room temperature, λex is 320 nm and λem is 645 nm

Metal Ion Competition Experiment

Achieving high selectivity for a specific metal ion over a complex background of competitive metals is an important feature to evaluate the performance of a chemosensor. Therefore the suitability and practical applicability of chemosensor R1 as a selective fluorescent probe for Cu2+ ion was checked by competition experiments in the presence of various metal ions. To investigate the interference from a number of cations the change in fluorescence emission intensity at 645 nm was recorded with solution of R1 (20 μM) containing Na+, K+, Mg2+, Ba2+, Ce3+, Fe3+, As3+, Hg2+, Zn2+, Cd2+, Pb2+, Mn2+, Co2+, Ni2+ (200 μM) and Hg2+, Fe3+ (20 μM) followed by the addition of Cu2+ (20 μM). In recognition of Cu2+ with chemosensor R1 no significant interference was observed in the presence of most competitive metal ions except for the Hg2+ and Fe3+ ion as shown in the Fig. 15. This indicates that only Hg2+ and Fe3+ compete with Cu2+ for binding with chemosensor R1. The fluorescence enhancement caused by the Cu2+ ion remained unaffected with most examined metal ions. These observations suggests that R1 can be applied as an outstanding selective and sensitive fluorescent chemosensors for estimation of trace amount of Cu2+ ion in real samples in the presence of these metals ions.
Fig. 15

Fluorescence response of R1 to 200 μM various tested metal ions (grey bar) and to the mixture of 200 μM tested metal ions with 20 μM Cu2+ (black bar) at room temperature, λex = 320 nm

Comparison with Pervious Works

Specific features of the proposed chemosensor R1 towards Cu2+ was compared with some previously reported chemosensor for Cu2+ in Table 3.
Table 3

Comparison of R1 with previous work [19, 20, 24, 42, 46, 47]

Most of the chemosensor required rigorous testing media and most of them displayed quenching of fluorescence upon interaction with Cu2+. Our proposed chemosensor R1 presents a number of attractive analytical features such as one step synthesis, high selectivity for Cu2+, sensitivity, enhancement of fluorescence, good reproducibility, wide pH range and can be used for the rapid detection of Cu2+ in natural water samples.

Reversibility of Chemosensor R1

In the development of fluorescent chemosensor, the main feature to increase their applicability is the detection. The use of a chemosensor for detection of particular metal ion, reversibility is an important aspect in practical applications. To investigate whether the enhancement of fluorescence emission intensity was as a result of indeed Cu2+ ion binding with chemosensor R1 and not photoactivation of chemosensor R1 or a chemical reaction reversibility experiment was conducted to study whether the complexation of chemosensor R1 with Cu2+ is reversible or not. For this purpose we use a strong metal ion chelator EDTA to revert the fluorescence. To the solution containing R1-Cu2+ complex 1 equiv. of EDTA was introduced and its fluorescence emission intensity was monitored at 645 nm after interaction with EDTA. After the addition of EDTA fluorescence emission signal was restored at 645 nm. This analysis suggests that the complexation between copper and chemosensor R1 is really and chemically reversible as shown in Fig. 16.
Fig. 16

Reversibility of binding interaction between chemosensor R1 (10 μM) and copper (20 μM) by the introduction of EDTA (20 μM) to the solution of complex

Application to the Analysis of Natural Water Samples

To investigate the practical applicability and sensitivity of the new chemosensor R1 in complex matrices, we examined the ability of chemosensor R1 to detect trace amount of Cu2+ ions in three natural water samples using the spike method. Chemosensor R1 was sensitive enough to detect Cu2+ ion in natural water samples. For this purpose samples were collected from three significantly different sourcses: the river water from river Swat, tap water from laboratory water pipe located at university of Malakand KPK and well water from Batkhela and Thana. Different concentrations of Cu2+ ion were added into the water samples and assayed within 24 h of the collection. The fluorescence emission intensity of each sample was monitored at 645 nm and the results are shown in the Fig. 17. As it is cleared from the Figure that fluorescence emission intensity of chemosensor R1 was proportional to the amount of Cu2+ ion added. These results displayed that relative to distilled water chemosensor R1 can also detect Cu2+ ion in the complex media, pointing out its potential applicability in the field of toxicology and environment sciences.
Fig. 17

Respective fluorescence responses of chemosensor R1 to Cu2+ ion added to the natural water samples at room temperature, [Cu2+] 2–10 μM, λex is 320 nm and λem is 645 nm, [R1] 10 μM

Conclusion

In conclusion, we have developed a Schiff based fluorescent chemosensor for Cu2+ ion and it was characterized by FTIR and single X-ray diffraction and 1H NMR. Significant fluorescence enhancement was observed by this sensor only in the presence of Cu2+ ion. The enhancement was due to the restriction of ICT phenomenon. The optimal pH range for detection of Cu2+ ion was 5–12. Our proposed chemosensor R1 possess high selectivity, sensitivity, and fast response time towards Cu2+ ion over a large number of competing metal ions. The excellent low limit of detection of this chemosensor R1 towards Cu2+ ion can be useful in detection of trace amount of Cu2+ ion in environmental samples. The response was reversible and the given sensor was successfully applied for the detection of Cu2+ in natural water sample.

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Maria Sadia
    • 1
    Email author
  • Robina Naz
    • 1
  • Jehangir Khan
    • 1
  • Rizwan Khan
    • 1
  1. 1.Department of ChemistryUniversity of MalakandLower DirPakistan

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