Accepted Manuscript A Coumarin-indole Based Colorimetric and “Turn on” Fluorescent Probe for Cyanide Yu Xu, Xi Dai, Bao-Xiang Zhao PII: DOI: Reference:

S1386-1425(14)01631-X http://dx.doi.org/10.1016/j.saa.2014.11.013 SAA 12949

To appear in:

Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy

Received Date: Revised Date: Accepted Date:

6 September 2014 28 October 2014 5 November 2014

Please cite this article as: Y. Xu, X. Dai, B-X. Zhao, A Coumarin-indole Based Colorimetric and “Turn on” Fluorescent Probe for Cyanide, Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy (2014), doi: http://dx.doi.org/10.1016/j.saa.2014.11.013

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A Coumarin-indole Based Colorimetric and “Turn on” Fluorescent Probe for Cyanide Yu Xua,b, Xi Daia , Bao-Xiang Zhao a,∗

a

School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100, PR

China. b

Taishan College, Shandong University, Jinan 250100, PR China

* Corresponding Author: Bao-Xiang Zhao Tel.: +86 531 88366425; fax: +86 531 88564464; E-mail: [email protected]

1

Abstract A novel coumarin-indole based chemodosimeter with a simple structure was designed and prepared via a condensation reaction in high yield. The probe exhibited very high selectivity towards cyanide on both fluorescence and UV-vis spectra, which allowed it to quantitatively detect and imaging cyanide ions in organic-aqueous solution by either fluorescence enhancement or colorimetric changes. Confirmed by 1 H NMR and HRMS spectra, the detection mechanism was proved to be related with the Michael addition reaction induced by cyanide ions, which blocked the intramolecular charge transfer (ICT) of the probe. Moreover, the probe was able to be utilized efficiently in a wide pH range (7.5-10) with negligible interference from other anions and a low detection limit of 0.51 µM. Application in 5 kinds of natural water source and accurate detection of cyanide in tap water solvent system also indicated the high practical significance of the probe. Keywords: Coumarin; Colorimetric probe; Fluorescent chemodosimeter; Cyanide; Tap water detection.

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1. Introduction

Cyanide is known as one of the most toxic species in nature. When cyanide enters body by oral, inhalation or dermal exposure, it exerts its acute effects by complexing with ferric iron atoms in metalloenzymes, resulting in histotoxic anoxia through inhibition of cytochrome c oxidase [1]. It has been reported that 0.5-3.5 mg of cyanide in per kg of body weight is lethal for human beings [2, 3]. In the “Guidelines for Drinking-water Quality”, published in 1993, the maximum limit of cyanide in drinking water was suggested to be 0.07 mg/L [4]. As for fish, cyanide has even more deadly effects. More than 200 tons of fish died in Tisza River because of cyanide pollution in an industrial accident in 2000. Other animals such as otters and eagles that ate the tainted fish were also threatened endangered [5]. Although being highly toxic, cyanide is still in widespread use in industry, particularly in electroplating, metallurgy and gold mining [5, 6]. Moreover, cyanide ions can also be produced by some organisms including bacteria, algae and fungi, which produce cyanide anions as part of their nitrogen metabolic pathways [4, 7]. Thus, developing modern methods of cyanide detection has attracted much interest of scientists in recent years and various kinds of fluorescent sensors for cyanide with good effect have been reported [7-17]. However, some drawbacks existing in previous probes still need to be improved. For instance, many cyanide sensors required pure organic solution and did not work well in aqueous systems [18-21]; some probes were 3

influenced greatly by changes of pH values [22-24]; some had comparatively high detection limit that cannot satisfy detection requirement [25, 26]. Moreover, there were limited number of probes that exhibited good selectivity towards cyanide on both fluorescence and UV-vis spectra [27-30]. Herein, we reported a novel coumarin-indole based colorimetric and “turn on” fluorescent chemodosimeter with a simple molecular structure for cyanide detection and applied it to natural water source. The probe had very high selectivity for cyanide ions in organic-aqueous solution (DMSO: H 2O=95:5, pH=8.5) with the limit of detection (LOD) of 0.51 µM. Based on the ICT mechanism induced by Michael addition reaction, the probe was able to quantitatively monitor cyanide via either UV-vis or fluorescence spectra.

2. Experimental section 2.1. Chemicals. All reagents and solvents were purchased from commercial sources and used without further purification. The solutions of cyanide were prepared from n-Bu4N salt; the solutions of fluorine ions were prepared from potassium salt; other anion solutions were prepared from sodium salts. Deionized water was used throughout the process of absorption and fluorescence determination. All samples were prepared at room temperature, shaken for 10 s and rested for 12 h before UV-vis and fluorescence determination. 2.2. Instruments.

4

Melting points were determined on an XD-4 digital micro melting point apparatus. 1H NMR spectra (400 MHz) and

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C NMR spectra (100 MHz) were

recorded on a Bruker Avance 400 spectrometer, using d 6-DMSO as solvent and tetramethylsilane (TMS) as an internal standard. IR spectra were recorded with an IR spectrophotometer VERTEX 70 FT-IR (Bruker Optics). HRMS spectra were

recorded

on

a

Q-TOF6510

spectrograph

(Agilent).

Fluorescent

measurements were recorded on an F-380 luminescence spectrophotometer and UV–vis spectra were recorded on a TU-1901 UV-Vis Spectrometer. The pH measurements were measured by use of a PHS-3C digital pH-meter (YouKe, Shanghai). 2.3.

Synthesis

of

(Z)-3-((7-(diethylamino)-2-oxo-2H-chromen-3-yl)methylene)indolin-2-one (L). 3-Acetyl-7-(diethylamino)-2H-chromen-2-one (1) (245 mg, 1.0 mmol) and indolin-2-one (2) (160 mg, 1.20 mmol) were dissolved in 20 mL ethanol and refluxed for 6 h. Cooled to room temperature, the residue was filtered and washed with ethanol for 6 times to give probe L (325 mg), dark red powder in 90% yield without further purification. mp: 274-276 ◦C. IR (KBr), υ: 3376 (-CONHR), 3066 (-ArH), 2969 (-RH), 2928 (-RH), 1723 (-COOR), 1619 (C=C) cm-1; 1H NMR (DMSO-d 6, 400 MHz), δ (ppm): 1.16 (t, 6H, NCH 2CH3, J = 6.9 Hz), 3.49 (q, 4H, NCH2CH 3, J = 6.9 Hz), 6.60 (s, 1H, Coumarin-H), 6.77 (d, 1H, Coumarin-H, J = 8.9 Hz), 6.83 (d, 1H, Ar-H, J = 7.6 Hz), 6.97 (t, 1H, Ar-H, J = 7.6 Hz), 7.20 (t, 1H, Ar-H, J = 7.6 Hz), 7.48 (d, 1H, 5

Coumarin-H, J = 8.9 Hz), 7.55 (d, 1H, Ar-H, J = 7.6 Hz), 7.71 (s, 1H, Coumarin-H), 9.68 (s, 1H, C=C-H), 10.62 (s, 1H, N-H);

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C NMR (DMSO-d 6,

100 MHz), δ (ppm): 167.95, 161.76, 157.30, 152.37, 145.79, 140.83, 131.67, 129.73, 129.08, 126.09, 125.34, 121.64, 119.56, 112.71, 110.44, 109.87, 108.46, 97.02, 44.83 (2C), 12.85 (2C); HRMS: calcd for C22H 21N 2O 3+ [L+H]+ 361.1552, found: 361.1592.

3. Results and discussion As shown in Scheme 1, the probe was synthesized via the condensation of diethylaminocoumarin-aldehyde (1) and conindolin-2-one (2). Unlike some fluorescent sensors that required multiple steps of synthesis and purification [31-33], pure product of this probe was obtained in a high yield over 90% by a single step of filtration without further purification. The molecular structure of the probe was confirmed by IR, 1H NMR,

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C NMR and HRMS spectra

(Supporting information S7-S10).

Scheme 1

This structure was designed on basis of Michael addition reaction considering the strong nucleophilic property of cyanide ions. The conjugated π system between coumarin fluorophore and conindolin-2-one moiety enabled an ICT process to occur, which lead to red shift in the absorption spectra and a fluorescence quenching phenomenon of the probe. Upon the nucleophilic 6

addition of cyanide ions to β-position of the activated Michael acceptor, the ICT process was expected to be blocked and the probe responded to it with fluorescent enhancement and shifts of absorption maximum [22, 34-37]. The above-mentioned sensing mechanism was illustrated in Scheme 2, which was confirmed by 1H NMR and HRMS comparison. As shown in Fig. 1, on one hand, after addition of excess n-Bu4N +CN-, CN- was added to the β-position of the carbonyl group and gave an anionic product. The negative charge on the molecule shielded the electronic field, thus inducing all protons to exhibit upfield shift in 1H NMR spectra. In particular, the chemical shift of the vinylic H (2) shifted to greater extent (1.5 ppm), because its position was closer to the negative charge C. On the other hand, the number of the protons in 1

H NMR spectra did not change, which indicated that only CN- was added to

the probe, rather than HCN, to form a stabilized anionic species. In addition, the L-CN- adduct peaked at 386.1504 (calcd for C23H 20N3O 3- [L-CN]386.1505) in HRMS was clearly observed, which also confirmed this mechanism (Fig. S1).

Scheme 2 Fig. 1

The probe alone exhibited an absorption maximum at 550 nm, which was a typical ICT band on UV-vis spectra. Upon the titration of CN-, the peak 7

decreased at 550 nm while a new peak appeared and increased at 444 nm, resulting in a clear isosbestic point at 460 nm (Fig. 2). In addition, we calculated the ratios of the absorbance at 550 nm to that at 444 nm and drew the relationship between the ratios and the concentration of CN- (Fig. S2). It showed a good linear relationship with R2 = 0.996, which enabled us to quantitatively detect cyanide using UV-vis spectra. Furthermore, as shown in the insert of Fig. 5, the colorimetric response property made L a good cyanide sensor for “naked eyes” detection.

Fig. 2

Unlike some previous reported probes that monitored cyanide via only UV-vis absorption [24, 26], probe L also responded to cyanide with remarkable fluorescence enhancement (Fig. 3). With the addition of CN-, the fluorescence intensity enhanced continuously at 465 nm until CN- reached 20 equiv.. The 20-fold enhancement in fluorescence intensity proved its high sensitivity towards cyanide. Meanwhile, the quantum yield was increased from 0.009 to 0.123, using quinine sulfate dehydrate as standard [38, 39]. Moreover, we studied the relationship between fluorescence intensity and the concentration of CN-, and gave a linear function, y = 508x-552 with R2 = 0.9969 (Fig. S3). Based on LOD = 3σ/K, the limit of detection (LOD) of the probe was calculated to be 0.51 µM, which is much lower than the cyanide standard limit 8

in drinking water (2.7 µM) suggested by World Health Organization (WHO) in 1996 [1].

Fig. 3

To further examine its practical application, we explored the selectivity of the probe towards cyanide ions over other common anions including AcO-, ClO 3-, ClO4-, F-, HCO 3-, I-, IO 3-, NO2-, NO 3-, S 2-, S2O 32-, Br-, SO32- , SO 42-. Only CN- caused obvious fluorescence enhancement at 465 nm while other anions did not influence the fluorescence intensity observably (Fig. 4). The probe also exhibited high selectivity on UV-vis spectra (Fig. 5). Apart from CN-, no other species induced absorption maximum shift. Moreover, the interference study of these anions indicated the probe was able to be applied to complicated conditions with little interference from these species (Fig. S4). Now, it is clear that probe L has very good selectivity towards CN- on both fluorescence and UV-vis spectra.

Fig. 4 Fig. 5

We also studied the pH effect and the response time of probe L to explore its application conditions. As shown in Fig. S5, the fluorescence intensity of L and 9

L-CN remained stable in the pH range from 7.5 to 10.0. The response time recorded in Fig. S6 was 5 h, longer than that of some previous reported probes with two electron-withdrawing groups connected to C=C as doubly activated acceptors [40-42]. However, the comparatively long response time contributed to the α,β-unsaturated carbonyl structure guaranteed the good selectivity of this probe for cyanide. To explore the application of this probe, we applied it to detect cyanide in 5 kinds of natural water source, including Yellow River, Spouting Spring, Daming Lake, Heihu Spring and tap water. As shown in Fig. 6, replacing the deionized water with natural water did not influence the ability of the probe in detecting cyanide. After adding 10 equiv. of CN-, all the five systems exhibited similar remarkable fluorescence enhancement, verifying the probe was able to be used in complicated natural systems. Furthermore, we utilized the probe for quantitatively detecting cyanide in tap water. Different sets of solutions containing CN- were analysed using the developed fluorescence method. Table 1 clearly showed that the values calculated by the work curve (Fig. S3) based on the fluorescence intensity were very close to the real amounts of cyanide added to the samples, indicating the efficiency of the method for the detection of cyanide in tap water.

Fig. 6 Table 1 10

4. Conclusion In summary, based on one step of condensation of a coumarin fluorophore and an indole moiety, we developed a new colorimetric and “turn-on” fluorescent sensor for cyanide. The detection mechanism was confirmed by 1H NMR and HRMS analysis to be the Michael addition of CN- to the probe, which blocked its ICT process. Upon addition of cyanide to aqueous-organic solution (DMSO:H 2O=95:5), the probe was able to respond to CN- with a 20-fold fluorescence enhancement at 465 nm and had obvious changes in absorption spectra with decrease at 550 nm and increase at 444 nm, which allowed us to detect cyanide with naked eyes as well. More importantly, the probe exhibited very high selectivity towards CN- over other species on both fluorescence and absorption spectra. On basis of either fluorescence enhancement or the ratio relationship (A 550/A 444) in absorption spectra, we could quantitatively monitor cyanide with a very low detection limit of 0.51 µM, a response time of 5 h in a wide pH range from 7.5 to 10.0. Furthermore, the probe was successfully applied to 5 different kinds of natural water source and exhibited good response to cyanide without interference from these systems. Using tap water as the detection system, the probe also gave reliable concentrations of cyanide with small error. All in all, this work illustrated that the probe could be an ideal sensor for monitoring cyanide in aqueous-organic solution with practical significance. 11

Acknowledgements We thank 973 Program (2010CB933504) and Foundation of Talent Training of Fundamental Subject of China (Grant No: J1103314) for support this work.

Notes and references

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Legend of Figures

Scheme 1. The synthetic route of probe L. Scheme 2. The detection mechanism of probe L towards CN-.

Fig. 1 The 1H NMR spectra comparisons of L and L-CN-. Fig. 2 Absorption spectra of L (10 µM) upon the titration of CN- (0, 2, 3, 4, 5, 7, 8, 9 equiv.) in DMSO/H 2O (pH = 8.5, 95:5, v/v) solution. Fig. 3 Fluorescence emission spectra of L (10 µM) upon the addition of CN- (0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 14, 16, 18, 20 equiv.) in DMSO/H 2O (pH = 8.5, 95:5, v/v) solution. (λex = 385 nm) Fig. 4 Fluorescence emission spectra of probe L (10 µM) in DMSO/H 2O (pH = 8.5, 95:5, v/v) upon additions of various anions (10 equiv.), respectively. (λex = 385 nm) Fig. 5 UV-vis spectral changes of probe L (10 µM) in DMSO/H 2O (pH = 8.5, 95:5, v/v) upon additions of various anions (10 equiv.). Fig. 6 Fluorescence emission spectra of probe L (10 µM) in DMSO/H 2O (pH = 8.6, 95:5, v/v) with or without 10 equiv. CN- in 5 kinds of solvent systems. (the water in these five solvent systems is from Yellow River, Spouting Spring, Daming Lake, Heihu Spring and tap water (From top to bottom), respectively; λex = 385 nm) Table 1. Detection of CN- in tap water samples 16

17

18

19

20

21

22

23

Table 1 Detection of CN- in tap water samples (Concentration of L, 10 µM) -

-

No. of samples

Amount of CN (µM)

CN found (µM)

Error

1

30

29.4

0.020

2

50

50.4

0.008

3

70

70.8

0.011

4

90

89.1

0.010

24

Graphical abstract

25

Highlights

A Coumarin-indole Based Colorimetric and “Turn on” Fluorescent Probe for Cyanide Yu Xua,b, Xi Daia , Bao-Xiang Zhao a,∗

A novel colorimetric and turn-on fluorescent probe for CN- has been designed, synthesized and characterized.

The probe has a low detection limit of 0.50 μM and is able to quantitatively monitor cyanide based on either fluorescence or UV-vis spectra.

The probe has been successfully applied in cyanide detection in tap water with very little error.

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