Fluorescent Chemosensing Properties of New Isoindoline Based-Receptors towards F − and Cu 2 + Ions

Two simple colorimetric and fluorescent anion chemosensors based on amide moieties, 4-nitro-N-[(1,3-dioxoisoindolin-2yl)benzamide (1), 3,5-dinitro-N-[(1,3-dioxoisoindolin-2yl)benzamide (2) have been synthesized and characterized. Comparing with other anions studied, the UV-visible absorption spectrum in DMSO shows significant response toward F ion with high selectivity, and meanwhile color change is observed from colorless to pink and violet for 1 and 2 respectively in the presence of tetrabutylammonium fluoride (TBAF) (5 × 10 M). Moreover, F-induced color changes remain the same even in the large excess of Cl, Br and I. In addition, the H NMR spectrum titration in DMSO-d6 shows deprotonation of the receptors 1 and 2 in the presence of basic F ion. In particular, addition of F to the receptors 1 and 2 resulted in an enhancement in fluorescence intensity at 770 nm. Both receptors 1 and 2 exhibit a fluorescent emission enhancement when addition of Zn, Ni, Co, Cu and Mn metal ions. The fluorescent enhancement values for the receptor 1 are 2.29 (Cu), 1.85 (Ni), 1.37 (Co), 1.27 (Mn), 1.25 (Zn) and for 2, 2.57 (Cu), 1.66 (Ni), 1.36 (Co), 1.15 (Mn), 1.09 (Zn). The selectivity of Cu is higher for 1 and 2 than other metal ions such as Zn, Ni, Co and Mn.


Introduction
The development of new molecular systems for the colorimetric detection of anions [1][2][3], and cations [4,5] or neutral molecules [6] have attained prime significance due to their biological and environmental applications.The incorporation of fluorescent chromophores into the receptors that are sensitive interactions between the host and guest molecules has recently gained considerable attention because of their highly sensitivity and low detection limit [7,8].The appeal of fluorescent sensors originnates from the high sensitivity of fluorescent detection compared to other spectroscopic methods.When a guest species binds to the fluorescent receptor, a certain property such as fluorescent intensity, wavelength, lifetime, etc changes, and such a change serve as an indicator of guest binding.
Among the anions, the detection of fluoride (F − ) is of particular interests [9,10] as it plays an essential role in a broad range of biological, medical and chemical processes and applications such as dental care [11], treatment of osteoporosis [12], association with hydrolysis of the nerve gas sarin [13], or even chemical and nuclear warfare agents [14].Being small and highly electronegative, fluoride has unique chemical properties and can form strong hydrogen bonds with hydrogen-bond donors.The majority of reported fluoride sensors are based on colorimetric changes [15] or fluorescence quenching [16], few of them experience fluorescence enhancement [17].
The development of fluorescent molecular sensors for metal ions, especially for cations with biological interest, has always been of particular interest.More specifically, fluorescent sensors for the detection and measurement of copper ions are actively investigated, as this metal ion is a significant environmental pollutant and an essential trace element in biological systems [18].For most of the reported Cu 2+ fluorescent sensors, the binding of the metal ion causes a quenching of the fluorescence emission [19].Only a few sensors in which the binding of Cu 2+ ion causes an increase in the fluorescence have been reported [20].A good case in point is Kumar's sensor displaying fluorescence enhancement with Cu 2+ [21].Samanta and co-workers have also achieved fluorescence enhancement with Cu 2+ using an electron an electron-deficient fluorophore component to reduce the M-F interaction [22,23].
We have recently embarked upon designing systems that can sense both anions and metal ions [24].Commonly, a typical chemosensor is the compound coupling of at least two units: a fluorophore and a binding unit.In this paper, with the conception of designing fluorescent receptors for F − and Cu(II), two receptors were designed and synthesized by the combination of naphthalimide moiety (as the fluorophore) and amide group (as the anion binding unit) (see Scheme 1).

Chemicals
The solvent DMSO was used as HPLC grade and purchased from Qualigens (Chennai, India).All anions, in the form of tetrabutylammonium salts, were purchased from Sigma-Aldrich Chemical Co.(Bangalore, India).

Physical Measurements
The 1 H and 13 C NMR spectra were recorded on a JEOL 400 MHz spectrometer in DMSO-d 6 .The IR spectra were recorded on a Perkin-Elmer FT-IR LX-185256 spectrometer.UV-visible experiments were performed on a Perkin-Elmer FT UV-Visible λ-35 spectrometer.Elemental analysis of the receptors was obtained in Heraeus CHN rapid analyzer.The EI mass spectra were recorded using a JEOL DX 303 mass spectrometer.Emission spectrum was recorded using a Perkin-Elmer Fluorescence λ-45 spectrometer.

UV-Visible Spectroscopic Methods
The solutions of receptors 1 and 2 (5 × 10 −5 M) were made up with HPLC grade DMSO solvent.UV-visible and fluorescence titrations were carried out by adding 10 μl aliquots of the titrant as tetrabutylammonium salts (5 × 10 −3 M) and cations as its perchlorate salts (5 × 10 −3 M).The UV-visible and fluorescence spectra were recorded after each addition.

1 H NMR Spectral Studies
The increase of electron density in the phenyl ring causes a shielding effect and should promote an upfield shift, which is expected to come from the formed hydrogen bond [29].

Anion Sensing
The recognition properties of the receptors 1 and 2 toward different anions were studied by the several methods such as the naked-eye experiment (Figures 2(a

Absorption Titration Studies
In the naked-eye experiments, receptors 1 and 2 (5 × 10 −5 M in DMSO) showed color changes from colorless to pink and violet respectively in the presence of 2 equivalents of TBAF (5 × 10 −3 M in DMSO).Both the receptors were found to be insensitive even to the addition of large excess of Cl − , Br − and I − ions (5 × 10 −1 M).Peaks at 258 and 287 nm of receptor 1 decreased gradually while two new peaks formed and increased gradually at 383 and 535 nm simultaneously with increasing amounts of F − ions (0.01 ml (0.33 equiv) of 5 × 10 −3 M).The colourless solution of the receptor 1 turned pink at the same time (Figures 2(a) and (b)).These results indicate that the strong electron-withdrawing effect of NO 2 substituents makes the charge delocalization more easily upon interaction with fluoride [28].This could be explained based on the intramolecular electron-transfer interactions [30] between the electron rich amide bond anion and the electron deficient phthalimide ring.In addition there is one isobestic point at approximately 310 nm, which indicated that equilibrium between 1 and 1. F − during the course of the titration.While for the receptor 2, the peak around 260 nm decreased and two new peaks were observed at 405 and 534 nm with an isobestic point at 270 nm, but no detectable spectral changes were observed even in the presence of larger excess of hundred equivalents of the corresponding anions such as Cl − , Br − and I − , which made it clear that 1 and 2 could sense F − over other anions.These observation could be attributed to the small size and higher electro negativity of the F − compared to the other halide ions [25,31].The binding constant of the receptor 1 and 2 were calculated from the fluoride-induced absorption changes using the following Equation ( 1) [32].
Where, A 0 and A are the intensities of absorption of the receptor and complexes (receptor with guest ions) respectively,  L is a molar absorption co-efficient of the receptor,  ML is a molar absorption co-efficient of the receptor with guest ions and K s is the binding constant.The binding constant can be obtained by the ratio of intercept and slope.The binding constant for F − complexation for the receptors 1 and 2 were estimated as (1.7 ± 0.20) × 10 3 M −1 (Figure 3(b) and Table 1) and ( 2  c Changes in the UV-visible spectra were not enough to calculate the binding constant.
The higher binding constant was achieved for the receptor 2 which might be due to the introduction of another nitro group in 2, which increases the acidity of amide-NH group and which enhance the hydrogen bond donating character of this receptor.[33] Restoration of the original spectrum of the sensor system from this receptor-F − adduct upon addition of a trace amount of water/methanol not only suggests that the complexation between F − and 1 or 2 are reversible in nature but also leads further support to the proposition that hydrogen bonding exists between the receptor and F − [34].

Anion Sensing
To corroborate well with those obtained during the UVvisible titrations described above, the fluorescence titration experiment was carried out with TBAF.These observations were illustrated in Figures 4(a) and (b), which showed the fluorescence spectra in different concentration of the receptors 1 and 2 respectively.The fluorescent spectral properties of the receptors 1 and 2 (5 × 10 −5 M) were determined in DMSO showing fluorescence intensity at 770 nm (λ exc = 380 nm).Upon the addition of F − (0.01 ml (0.33 equiv.) of 5 × 10 −3 M) to DMSO solutions of 1 and 2, the fluorescence emission intensity of the receptors 1 and 2 enhanced at 770 nm and the enhancement efficiency increases along with the increases in concentration of F − up to 2 equiv.However no detectable changes were observed even in the presence of large excess of hundred equivalents of the corresponding anions such as Br − , Cl − , and I − which made it clear that 1 and 2 could sense F − over other anions.These results suggested that the receptors 1 and 2 have a higher selectivity for F − compared to other halide anions.The selectivity for F − can be attributed to Brønsted basicity of the receptor.The F − anions have the strongest basicity, and also exhibit the most effective hydrogen bonding or deprotonation with the binding site, amide group.F − coordination to the binding site of the receptors 1 and 2 results in an enhancement of the phthalimide emission.The enhancement of the fluorescence intensities may due to the result of an increase of the rigidity of the receptor molecules upon receptor: F − complexation [35].

Cation Sensing
To investigate the metal-induced fluorescence change the fluorescence spectra of receptors 1 and 2 in complexation with different transition metal ions (Cu 2+ , Zn 2+ , Ni 2+ , Co 2+ and Mn 2+ ) were recorded in DMSO to examine the selectivity.
The emission spectrum of free receptors 1 and 2 display a broad band with an emission maximum at 770 (λ exc = 380 nm).Upon addition different metal ions (0.01 ml (0.03 equiv.) of 5 × 10 −3 M) to a solution of receptors 1 and 2 in DMSO, a significance intensity enhancement of the emission bands a significant increase in the 770 nm emission, which was attributed to the formation of receptors: metal complex was observed (Figures 5(a nature of the added metal cations.The FE = I/I 0 was determined from the ratio of maximum fluorescence intensity I (after addition metal cations) and minimum fluorescence intensity I 0 (before metal cations addition).
It is noticeable that only Cu 2+ from all other metal ion tested is able to produce large change in the emission spectrum indicating that this enhancement for the fluorescence is highly specific for Cu 2+ cations.

Conclusion
In conclusion, we have developed selective and sensitive dual chromo as well as fluorogenic receptors 1 and 2 using phthalimide moiety for the determination of F − and Cu 2+ .These receptors selectively recognize F − ions even in the presence of other halide ions and shows higher selectivity towards Cu 2+ ions than other metal ions studies.

1 H
NMR spectra of the receptors 1 and 2 were recorded using DMSO-d 6 in the presence and absence of fluoride ion to know the interaction between them.1 H NMR spectrum of the receptor 1 showed -NH proton signal at δ 8.5 and the receptor 2 showed NH proton signals at δ 9.7.The signals at δ 7.6 -7.3 and δ 7.8 -7.5 indicated the presence of aromatic group present in phthalimide ring for the receptors 1 and 2 respectively.To investigate anion-binding properties of the receptors 1 and 2, the 1 H NMR spectral change caused by the addition of the fluoride as tetrabutylammonium salts were studied.The 1 H NMR spectrums of receptors 1 and 2 (before and after addition of F − ) are shown in Figures1(a) and (b) respectively.Before discussing the spectroscopic features in detail, two effects, which were responsible for 1 H NMR

Figure 1 .
Figure 1.(a) Partial 1 H NMR (400 MHz) spectra of receptor 1 in DMSO-d 6 in the (a) absence (b) presence of 1 equiv.and (c) 5 equiv. of [nBu 4 N]F.The signals of NH protons are marked with black dots; (b) Partial 1 H NMR (400 MHz) spectra of receptor 2 in DMSO-d 6 in the (a) absence (b) presence of 1 equiv.and (c) 5 equiv. of [nBu 4 N]F.The signals of NH protons are marked with black dots.
) and (b)), the 1 H NMR, UV-visible and fluorescence titrations.The UV-visible titrations were carried out by adding 5 × 10 −3 M solution of tetrabutylammonium salts of anions to the DMSO solution of receptors 1 and 2 (5 × 10 −5 M).

Figure 2 .
Figure 2. (a) Color changes of receptor (R) 1 in DMSO (5.0 × 10 −5 M) before and after the addition of 2 equivalents of representative anions (from the left to right: R, R + F − , R + Cl − , R + Br − , R + I − ); (b) Color changes of receptor (R) 2 in DMSO (5.0 × 10 −5 M) before and after the addition of 2 equivalents of representative anions (from the left to right: R, R + F − , R + Cl − , R + Br − , R + I − ).

Figure 4 .
Figure 4. (a) The changes in the fluorescence emission spectra of receptor 1 (5.0 × 10 −5 M) and (b) for 2 upon titration with solutions of tetrabutylammonium fluoride in DMSO.

Table 1 . Data obtained from the UV-visible spectra upon titration of receptors 1 and 2 with anions a in DMSO.
Countercation was tetrabutylammonium salts for anions.All errors are ±5%; b The error value was obtained by the result of linear fitting.All errors are ±5%; a