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Talanta 80 (2009) 996–1000

Contents lists available at ScienceDirect

Talanta

journa l homepage: www.e lsev ier .com/ locate / ta lanta

novel fluorescent and chromogenic probe for cyanide detection in water basedn the nucleophilic addition of cyanide to imine group

ue Sun, Yunlong Liu, Maliang Chen, Wei Guo ∗

chool of Chemistry and Chemical Engineering, Shanxi University, Wucheng Road, Taiyuan 030006, China

r t i c l e i n f o

rticle history:eceived 17 June 2009eceived in revised form 16 August 2009ccepted 21 August 2009

a b s t r a c t

A fluorescent and colorimetric probe bearing salicylaldehyde hydrazone functionality has been preparedfor cyanide sensing. The detection of cyanide was performed via the nucleophilic attack of cyanide anionon the imine group of the probe with a 1:1 binding stoichiometry, which could be confirmed by 1H NMRand MS studies. The specific reaction results in a prominent fluorescence enhancement and a color change

vailable online 28 August 2009

eywords:hemosensoryanide sensing

from colorless to yellow.© 2009 Elsevier B.V. All rights reserved.

ydrazone functionality-Hydroxy-1-naphthaldehyde-(N,N-Dimethylamino)benzoylhydrazine

. Introduction

Anion recognition is an area of growing interest in supramolec-lar chemistry due to its important role in a wide range ofnvironmental, clinical, chemical, and biological applications, andonsiderable attention has been focused on the design of arti-cial receptors that are able to selectively recognize and sensenion species [1,2]. Among various anions, cyanide is one of theost concerned anions because it is being widely used in syn-

hetic fibers, resins, herbicide, and the gold-extraction process [3].nfortunately, cyanide anion is extremely detrimental, and coulde absorbed through lungs, gastrointestinal track and skin, lead-

ng to vomiting, convulsion, loss of consciousness, and eventualeath [4–6]. Thus, there is a need for an efficient sensing systemor cyanide to monitor cyanide concentration from contaminantources.

Many of the cyanide anion receptors reported to date have reliedn hydrogen-bonding motifs and, as a consequence, have generallyisplayed poor selectivity relative to other anions [7–9]. To over-ome this limitation, reaction-based receptors, rationally designedyanide anion indicators, have been developed recently [10–21].his reaction-based recognition mode takes advantage of the par-

icular feature of the cyanide ion: its nucleophilic character, andnables the recognition system with some characteristic featuresuch as analyte-specific response and little competition from thequeous media, which are highly desirable features for an efficient

∗ Corresponding author. Tel.: +86 351 7011600.E-mail address: guowsxu@yahoo.com.cn (W. Guo).

039-9140/$ – see front matter © 2009 Elsevier B.V. All rights reserved.oi:10.1016/j.talanta.2009.08.026

recognition and detection system. Based on this idea, nucleophilicaddition of cyanide to oxazine [10,11], pyrylium [12], squarane [13],trifluoroacetophenone [14,15], acyltrazene [16], acridinium [17],salicylaldehyde [18,19] and carboxamide [20,21] has been reportedin the past few years, in which the interference by other anions,such F− and AcO−, can be efficiently minimized.

Recently, we have developed a series of nitroaniline-basedbenzamide compounds for the ‘naked-eye’ detection of cyanidein aqueous environment with high selectivity [22]. These com-pounds react with CN− in a 1:1 stoichiometric manner, a processwhich induces a large enhancement in absorption intensity anda marked color changes from colorless to yellow in DMSO–H2O(1:1, v/v) at room temperature. In continuation of our work, inthis paper, we wish to report a new type of probe molecule(1) bearing hydrazone functionality for highly selective cyanidedetection in water (Scheme 1). 1 was designed on the basis of 4-(N,N-dimethylamino)benzamide fluorophore [22] and 2-naphtholfluorophore, which were connected by a rigid hydrazone function-ality. Cyanide is expected to be detectable by nucleophilic attacktoward an imine functional group of 1, which is activated by anintramolecular hydrogen bond, and fast proton transfer of the phe-nol hydrogen to the developing nitrogen anion would then bringabout a spectroscopic change (Scheme 1).

2. Experimental

2.1. Materials and apparatus

Commercially available compounds were used without furtherpurification. Solvents were dried according to standard proce-

Y. Sun et al. / Talanta 80 (2009) 996–1000 997

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Scheme 1. Proposed cya

ures. All reactions were magnetically stirred and monitored byhin-layer chromatography (TLC) using Huanghai GF254 silica geloated plates. The fluorescence spectra were carried out on Var-an Cary Edipse fluorescence spectrometer. The absorbance spectra

ere recorded with Agilent 8453 UV–vis spectrophotometer. NMRpectra were recorded on a Bruker Avance DRX300 MHz nuclearagnetic resonance spectrometer with tetramethylsilane (TMS)

s internal standards. ESIMS were taken on Fourier transform ionyclotron resonance mass spectrometry (Varian 7.0T).

.2. Preparation of probe molecule 1

A solution of 4-(N,N-dimethylamino)benzoylhydrazine (89 mg,.5 mmol) and 2-hydroxy-1-naphthaldehyde (86 mg, 0.5 mmol) inthanol (20 mL) was heated under reflux for 1 h. After left to cool,he precipitated crystals were filtered and washed with ethanolo give pure 1 as yellow solid (160 mg, 95%). 1H NMR (300 MHz,MSO-d6) ı 12.99 (s, 1H), 11.91 (s, 1H), 9.46 (s, 1H), 8.16 (d, 1H),.86 (m, 4H), 7.61 (s, 1H), 7.40 (s, 1H), 7.23 (d, 1H), 6.80 (d, 2H), 3.02s, 6H); 13C NMR (75 MHz, DMSO-d6) ı 168.1, 163.7, 158.3, 150.8,37.8, 137.4, 134.8, 133.5, 133.1, 129.0, 125.8, 124.8, 124.4, 116.5,14.3, 84.6, 84.2; HRMS (ESI): 356.1368 (M+Na)+.

.3. Titration experiment of 1

Deionized water and a spectroscopic grade of DMSO were useds the solvent for titration experiment. The titrations were carriedut in 10-mm quartz cuvettes at 25 ◦C. 1 was dissolved in DMSO tofford a concentration of 20 mM stock solution, which was diluted

ig. 1. (a) Absorbance changes, (b) fluorescence changes and the corresponding color carious anions (vials from the left: only 1, CN− , F− , Cl− , Br− , I− , AcO− , ClO4

− , NO3− , H2PO

he splits were 5.0 nm.

ensing mechanism of 1.

with DMSO–H2O (1:1, v/v) up to 20 �M. The sodium cyanide stocksolution of 1.0 × 10−1 M was diluted to 1.0 × 10−2 M with deionizedwater. The absorbance was measured from 250 to 550 nm, againsta blank DMSO–H2O (1:1, v/v) solution, and different cyanide solu-tions were added to the 20 �M host solution (2 mL) in portions(total volume: 4, 8, 12, 16, 20, 24, 32, 40, 48, 60 �L, of 1.0 × 10−2 M,and 62, 64, 66, 70, 74, 78, 82 �L, of 1.0 × 10−1 M). The emission wasmeasured from 400 to 600 nm, and different cyanide solutions wereadded to the 20 �M host solution (2 mL) in portions (total volume:4, 8, 12, 16, 20, 28, 36, 44, 52, 60 �L, of 1.0 × 10−2 M, and 62, 64,66, 68, 70, 74, 78, 82, 86, 94, 102 �L, of 1.0 × 10−1 M). The resultingsolution was shaken well and the absorption and emission spec-tra were recorded immediately. For detection limit measurements,1.0 × 10−4 and 1.0 × 10−3 M solutions of sodium cyanide were used.Unless otherwise noted, for all measurements, the excitation wave-length was at 334 nm, and both the excitation and emission slitwidths were 5 nm.

3. Results and discussion

3.1. Absorption and fluorescence spectral characteristics of 1

The ability of 1 to complex with anions was explored withUV–vis absorption and fluorescence spectrometry. Among the

eleven anions tested in solution (DMSO–H2O, 1:1, v/v), namely,CN−, F−, Cl−, Br−, I−, AcO−, ClO4

−, NO3−, H2PO4

−, N3−, and HSO4

−

as their sodium salts, 1 responded to only CN− resulting in a colorchange from colorless to yellow and a bright green fluorescence(Fig. 1), indicating that probe 1 can serve as a “naked-eye” indicator

hanges of 1 (20 �M, 25 ◦C) in DMSO–H2O (1:1, v/v) upon addition of 50 equiv. of4

− , N3− and HSO4

−). The excitation wavelength for the complex was 334 nm, and

998 Y. Sun et al. / Talanta 80 (2009) 996–1000

Fig. 2. Absorption spectra of 1 (20 �M) in DMSO–H2O (1:1, v/v) upon additionoc

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to nonradiative decay through rotatory motion of the amide group[34–36]. In addition, the proton transfer of the phenol hydrogen tothe developing nitrogen anion upon cyanide attack can also leadto the opening of another fluorescence emission channel relativeto naphtholate anion, resulting in the emission enhancement [37].

f cyanide anion (0–70 equiv.). (Inset) Plots of absorption 443 nm vs. the cyanideoncentration.

or CN− in DMSO aqueous media. The absorption and fluorescencerofiles of the sensor showed a remarkably higher selectivity foryanide over the other anions in water.

The nucleophilicity of cyanide in water seems to be theajor contributor to the high selectivity of 1 for cyanide. In 1:1MSO–H2O (v/v), anion such as F− and AcO− should interact with

he aqueous medium through H-bonding, and this solvation leadso a decrease in their basicity, thus, resulting in the poor deproto-ation reaction. In contrast, cyanide has much weaker H-bondingbility in comparison with F− and AcO− but has stronger carbonylarbon affinity, which results in the addition reaction of CN− tomine group (Scheme 1). However, in DMSO, not only the nucle-philicity but also the basicity of anions (F− and AcO−) may causearge changes in the UV–vis spectra of 1 and therefore result in lesselectivity for cyanide.

Fig. 2 illustrates the absorption spectral changes for 1 uponddition of CN− in DMSO–H2O (1:1, v/v). 1 shows an absorptionand at 371 nm and two shoulders at 336 and 386 nm. The bandt 336 nm can be assigned to 4-(N,N-dimethylamino)benzamideroup of 1, and the band at 371 nm and a shoulder at 386 nm cane assigned to 2-naphthol group. Upon addition of CN−, the bandt 336 nm kept almost unchanged; however, the bands at 371 and86 nm were attenuated while two new bands appeared at 443 and73 nm, which is similar to the observation on the deprotonationf 2-naphthol by anions where the new absorption was assignedo the naphtholate anion. Two isosbestic points at 344 and 403 nmere observed throughout the titration process. The variation of the

bsorbance at 443 nm was used to evaluate the binding constantsf 1 with CN− by assuming a 1:1 binding stoichiometry. Nonlineareast-squares regression analysis [23–26] of these changes gave ainding constant of 1.17 × 105 M−1. Nice fittings supported the 1:1inding stoichiometry.

Fluorescence monitoring of the cyanide addition reaction waslso performed by using a 20-�M solution of 1 in 1:1 of DMSO–H2Ot room temperature (Fig. 3). 1 shows a weak emission. Upon addi-ion of CN−, the fluorescence emission intensity of 1 was increasednd was saturated at 2.4 mM cyanide (120 equiv.). It is notewor-hy that the cyanide adducts display dual luminescence with twomission bands, 460 and 495 nm, in the visible region. Titration

f 1 with CN− ions gave a simultaneous increase in the intensityf two bands. The former can be assigned to the excited-state CTor the parent CT fluorophore 4-(N,N-dimethylamino)benzamide27–29], and the latter can be assigned to the naphtholate anion

Fig. 3. Fluorescence spectra of 1 (20 �M) in DMSO–H2O (1:1, v/v) upon additionof cyanide anions (0–120 equiv.). The excitation wavelength for the complex was334 nm, and the splits were 5 nm. (Inset) Plots of emission at 495 nm vs. the cyanideconcentration.

[30–32]. The resultant Job plot indicated a 1:1 receptor–cyanidebinding stoichiometry (Fig. 4).

Several factors can be proposed for rationalizing the observedemission enhancement of 1 upon the addition of the cyanide anion.Molecules with lowest excited singlet states of the n�* type aretypically nonemissive in fluid solution [33]. In the presence of CN−,as shown in Scheme 1, an intramolecular hydrogen-bond networkwill be created by nucleophilic attack of CN− at the imine group,and followed by fast proton transfer of the phenol hydrogen to thedeveloping nitrogen anion. With such an intramolecular hydrogen-bond network that interacts with the lone pair of the carbonyloxygen of 4-(N,N-dimethylamino)benzamide group, the energy ofthe n�* state of this group would be raised so that the emissionfrom the �–�* transition of the fluorophore would be restored,leading to a substantial fluorescence enhancement [33]. There wasno change in the UV absorption band at 336 nm on cyanide anionbinding, which suggests that the fluorescence enhancement mayalso be a consequence of a restriction in the conformational flexi-bility of 4-(N,N-dimethylamino)benzamide fluorophore due to theintramolecular hydrogen-bond network that would otherwise lead

Fig. 4. Job’s plot between 1 and cyanide anion in DMSO–H2O (1:1, v/v) at 25 ◦C.

Y. Sun et al. / Talanta 80 (2009) 996–1000 999

Fig. 5. 1H NMR spectral change of 1 in DMSO-d6 upon addition of cyanide anions:1 only (bottom) and 1 and 20 equiv. of Bu4NCN (top). Note: the broad peak at 3.8 iswater.

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Fig. 6. The ESIMS for the cyanide adduct of 1 [(1+NaCN+H+): 383.1478].

he detailed mechanism, however, remains to be further clarified.etailed work is now underway in this laboratory.

.2. 1H NMR and ESIMS investigation

An analysis on the 1H NMR spectra of 1 before and after theyanide addition proved informative (Fig. 5). The imine proton of(Ha) and the phenol proton (Hb) at around ı 11.9 and 13.0 ppmas dramatically shifted upfield to ı 3.7 and 1.1 ppm, respectively,pon cyanide addition at room temperature, indicating that theyanide anion functions as a nucleophile in water. The formationf cyanide adduct was further confirmed by mass spectrometry.he electrospray ionization mass spectrum of the cyanide adductsor 1 showed a molecular mass of 383.1, which correspond to theormula of [1+NaCN+H+] (Fig. 6).

.3. Fast response

As known, chemosensors always have a problem of longesponse time. In our case, the binding process of CN− to 1 wasound to be very fast (Fig. 7). After adding the sodium cyanide, twoew absorbance bands at 443 and 473 nm appeared and reached

ig. 7. Absorbance changes at 443 nm for 1 (20 �M, 25 ◦C) in a mixture ofMSO–H2O (1:1, v/v) after addition of NaCN (1 mM).

Fig. 8. Fluorescence intensity change at 495 nm for 1 (20 �M) in DMSO–H2O (1:1,v/v) as a function of the concentration of cyanide anions. The excitation wavelengthfor the complex was 334 nm, and the splits were 10 nm.

the plateau region less than 2–3 s, and remains quite stable from3 s to 10 min, suggesting that the addition reaction of CN− to iminegroup of 1 might be completed instantly and the chemosensor hasrapid detection ability for cyanide anion.

3.4. Detection limits

The detection limit was calculated based on the fluorescencetitration. To determine the S/N ratio, the emission intensity of 1without cyanide anion was measured by 10 times and the stan-dard deviation of blank measurements was determined. Under thepresent conditions, a good linear relationship between the fluores-cence intensity and the cyanide concentration could be obtainedin the 2 × 10−7 to 2 × 10−6 M (R2 = 0.9935), as shown in Fig. 8.The detection limit is then calculated with the equation: detec-tion limit = 3�bi/m, where �bi is the standard deviation of blankmeasurements, m is the slope between intensity versus sample con-centration. The detection limit was measured to be 7.4 × 10−8 M.According to the World Health Organization (WHO), cyanide con-centrations lower than 1.9 �M are acceptable in drinking water.This means that our proposed fluorescent method based on probemolecule 1 is sensitive enough to monitor cyanide concentrationsin drinking water.

3.5. Applications

In order to assess the utility of the proposed method, it wasapplied to the quantitative determination of cyanide anion in drink-ing water sample (drinking water from commence). The watersample was found to be free from cyanide and so the sample wasprepared by adding known amounts of cyanide to sample. Theapplications were performed using 2 mL of sample volume, andafter addition of cyanide with the concentration of 1.9 × 10−6 M,which was within the linear calibration range, the sample wasinjected in triplicate. The average content of cyanide anion wasfound to be 1.88 × 10−6 M for drinking water. Then, the recoverypercentage performed well with relative standard deviation lowerthan 2%.

4. Conclusion

In summary, an anion probe bearing 2-hydroxy-1-naphthaldehyde hydrazone moiety for cyanide anion recognition

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as synthesized, and its absorption and fluorescence propertiesn the presence of anions were evaluated. The probe displayedn obvious color change from colorless to yellow and a dramatichange in fluorescence intensity selectively for cyanide anionsver other anions in aqueous solution. Such selectivity results fromhe nucleophilicity of the cyanide anion and the imine activationy the neighboring phenol proton through an intramolecularydrogen bond. The cyanide detection method described herehould have potential application as a new family of probes foretecting cyanide in aqueous solution.

cknowledgements

We would like to thank the Natural Science Foundation of ChinaNSFC no. 20772073) and the Natural Science Foundation of Shanxirovince (2008011015) for financial support.

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