Highly Selective Fluorescence Sensing of Mercury Ions over a Broad

Apr 25, 2012 - Roop Shikha Singh , Rakesh Kumar Gupta , Rajendra Prasad Paitandi , Arvind Misra , Daya Shankar Pandey. New Journal of Chemistry 2015 ...
0 downloads 0 Views 4MB Size
Download PDF
Article pubs.acs.org/Macromolecules

Highly Selective Fluorescence Sensing of Mercury Ions over a Broad Concentration Range Based on Mixed Polymeric Micelles Jinming Hu, Tao Wu, Guoqing Zhang,* and Shiyong Liu* CAS Key Laboratory of Soft Matter Chemistry, Hefei National Laboratory for Physical Sciences at the Microscale, Department of Polymer Science and Engineering, University of Science and Technology of China, 96 Jinzhai Road, Hefei, Anhui Province, 230026 China S Supporting Information *

ABSTRACT: We report on the fabrication of a new type of polymeric fluorescent Hg2+ probe covering a broad effective concentration range from nanomolar to micromolar levels and exhibiting considerably enhanced detection selectivity. Two amphiphilic diblock copolymers colabeled with Hg2+-reactive caged dye (RhBHA) and Hg2+-catalyzed caged fluorophore (HCMA) in the hydrophilic segments, PS-b-P(DMA-co-HCMA) and PS-b-P(DMA-co-RhBHA), were synthesized via sequential reversible addition−fragmentation chain transfer (RAFT) polymerization, where PS, DMA, HCMA, and RhBHA are polystyrene, N,N-dimethylacrylamide, hydrazone-caged coumarin, and rhodamine B (RhB) derivatives, respectively. The two amphiphilic diblock copolymers can spontaneously self-assemble into mixed micelles in aqueous solution possessing hydrophobic PS cores and HCMA and RhBHA moieties colabeled hydrophilic PDMA coronas. Fluorescence emissions of caged RhBHA and HCMA moieties can effectively turn on in the presence of low and high Hg2+ concentrations via Hg2+-induced ring-opening reaction and Hg2+-catalyzed hydrolysis mechanisms, respectively. The drastically different, but self-complementary reaction kinetics and optimum working concentration ranges of RhBHA and HCMA moieties endow the sensing system with high selectivity and broad sensing concentration range (from nanomolar to micromolar). In addition, the Hg2+-sensing platform can be further employed for the fluorescent ratiometric detection of Cu2+ ion via its selective quenching of the emission of acyclic RhBHA moieties. This work presents a new example of ensembling two partially selective chemical reaction-based fluorometric sensing motifs to achieve enhanced metal ion sensing selectivity and broadened working concentration ranges, which can be further generalized for the construction of other highly selective and broad dynamic range sensing systems.

■

INTRODUCTION Mercury ion (Hg2+) poses a huge threat to human beings and the environment due to its notorious biological membrane permeability, bioaccumulation, and long residence in central nervous and endocrine systems.1,2 Thus, it is quite imperative to screen suitable Hg2+ detection systems with high sensitivity and reliability.3−6 In the past decades, numerous fluorometric and colorimetric Hg2+ probes based on small molecule chromophores,7−23 quantum dots,24,25 nanoparticles/nanoclusters,26−33 biomolecules (DNA and proteins),34−42 synthetic polymers,43−46 and polymeric assemblies47−54 have been developed, mainly utilizing specific supramolecular recognition or Hg2+-induced chemical reaction of caged chromophores. Current efforts in this area have focused on developing watersoluble, cell-permeable and ratiometric fluorescent Hg2+ sensing ensembles with enhanced selectivity and sensitivity.3,4 In addition to further improving detection limits by introducing new designing motifs, the screening of fluorometric Hg2+ probes covering an effective broad concentration range is also in high demand aiming to gather reliable information © 2012 American Chemical Society

concerning the spatiotemporal biodistribution and bioaccumulation of Hg2+ ions within living organisms.1 Though there exist a few examples of Zn2+ ion probes working over broad concentration ranges in previous literature reports, which are exclusively based on supramolecular recognition by ensembling multiple sensing moieties of varying binding affinity,55−59 the capability of integrating the highly selective reaction-based detection strategy and fluorometric sensing of Hg2+ ions over a broad concentration range has, to the best of our knowledge, not been achieved. Compared to the supramolecular recognition-based approach, chemical reaction-based one represents a more sophisticated and robust type which has drawn intensive attention in recent years due to its enhanced selectivity and detection specificity.60 We herein propose that if two types of fluorophore decaging reactions of varying mechanisms and Received: March 30, 2012 Revised: April 17, 2012 Published: April 25, 2012 3939

dx.doi.org/10.1021/ma3006558 | Macromolecules 2012, 45, 3939−3947

Macromolecules

Article

Scheme 1. Schematic Illustration of the Fabrication of Highly Selective Fluorescent Hg2+ Probes Covering a Broad Concentration Range Based on Hybrid Micelles Self-Assembled from PS-b-P(DMA-co-RhBHA) and PS-b-P(DMA-co-HCMA) Diblock Copolymers by Integrating Two Types of Decaging Reactions, Namely, Hg2+-Induced Ring-Opening Reaction of RhBHA Residues and Hg2+-Catalyzed Hydrolysis of Caged HCMA Moietiesa

The obtained dually emitting hybrid micelles can be further utilized for the ratiometric fluorescent sensing of Cu2+ ions due to selective quenching of the emission of acyclic RhBHA moieties. a

caged fluorophore (HCMA) which is prone to be subjected to Hg2+-catalyzed decaging reaction, respectively. These two types of reactions can transform initially nonfluorescent RhBHA and HCMA moieties into orange-emitting and blue-emitting ones, respectively. Notably, they possess drastically different, but selfcomplementary reaction kinetics and optimum working concentration ranges. The RhBHA moiety was chosen to be the faster and more quantitative reaction-based caged fluorophore compared to that exhibited by HCMA, and the decaging of HCMA is of catalytic origin modulated by Hg2+

detection sensitivity can exhibit partial selectivity toward a specific metal ion, the integration of them within one ensemble, e.g., diblock copolymer micelles in the current work, might endow the probing system with improved detection selectivity, enhanced reliability, and most importantly, extended effective working concentration ranges.49,61 A proof-of-concept design attempting to confirm the above hypothesis is shown in Scheme 1, which involves two amphiphilic diblock copolymers labeled with one type of Hg2+-reactive caged dye (RhBHA)43,51 and another type of 3940

dx.doi.org/10.1021/ma3006558 | Macromolecules 2012, 45, 3939−3947

Macromolecules

Article

concentrations. Furthermore, the diblock copolymer micelle scaffold provides the sensing ensemble with excellent water dispersibility and structural stability. In addition, the fluorescence emissions of decaged RhBHA and HCMA are quite well-resolved with ∼170 nm difference in their emission maxima (590 and 420 nm, respectively). Thus, the high-fidelity, ultraselective, and highly sensitive fluorometric detection of Hg2+ ions over a broad concentration ranging from nM to mM levels can be facilely achieved (Scheme 1).

■

Scheme 2. Schematic Illustration for the Synthesis of PS-bP(DMA-co-RhBHA) and PS-b-P(DMA-co-HCMA) Amphiphilic Diblock Copolymers

EXPERIMENTAL SECTION

Materials. N,N-Dimethylacrylamide (DMA, 98%, TCI) was dried with CaH2 and distilled at reduced pressure prior to use. Styrene (St, 99.5%, Sinopharm Chemical Reagent Co.) was successively washed with aqueous NaOH (5.0 wt %) and saturated NaCl solution, and then distilled over CaH2 at reduced pressure. 2,2′-Azoisobutyronitrile (AIBN) was recrystallized from 95% ethanol. 4-Methoxyphenylhydrazine, 2,4-dihydroxybenzaldehyde, methyl acetoacetate, glacial acetic acid (CH3COOH), potassium thiocyanate (KSCN), and hydrazine hydrate (Sinopharm Chemical Reagent Co.) were used as received. Tetrahydrofuran (THF), ethanol, and all the other solvents were used as received. Nitrate salts (K+, Na+, Li+, Co2+, Cd2+, Pb2+, Zn2+, Fe2+, Fe3+, Ca2+, Ag+, Cu2+, and Hg2+) were used for all sensing experiments. Water was deionized with a Milli-Q SP reagent water system (Millipore) to a specific resistivity of 18.2 MΩcm. 3-Acetyl-7-(2methacroyloxy-n- ethyloxy) coumarin (CMA) monomer,62 RhBHA,43 and S-1-propyl-S′-(α,α′-dimethyl-α″-acetic acid)- trithiocarbonate (PDMAT)63 were synthesized according to previously reported literature procedures. Sample Synthesis. Synthetic routes employed for the preparation of PS-b-P(DMA-co-RhBHA), PS-b-P(DMA-co-CMA), and PS-bP(DMA-co-HCMA) amphiphilic diblock copolymers are shown in Scheme 2. The structural parameters of PS macroRAFT agent and amphiphilic diblock copolymers are summarized in Table S1, Supporting Information. Synthesis of PS macroRAFT agent (Scheme 2). Typical procedures employed for the synthesis of PS macroRAFT agent are as follows. Into a reaction tube equipped with a magnetic stirring bar, St (5.21 g, 50 mmol), PDMAT (0.238 g, 1 mmol), AIBN (16.4 mg, 0.1 mmol), and 1,4-dioxane (5.21 g) were charged. The tube was carefully degassed by three freeze−pump−thaw cycles and then sealed under vacuum. After thermostating at 80 °C in an oil bath for 10 h under magnetic stirring, the reaction tube was quenched into liquid nitrogen, opened, and diluted with THF; the mixture was then precipitated into an excess of methanol. The above dissolution−precipitation cycle was repeated for three times. After drying in a vacuum oven overnight at room temperature, PS macroRAFT agent was obtained as a yellowish powder (0.53 g, yield: ∼10%). The DP of PS block was determined to be 10 by 1H NMR analysis in CDCl3 (Figure S1a, Supporting Information). GPC analysis gives an Mn of 1.3 kDa and an Mw/Mn of 1.10 (Figure S2, Supporting Information). Thus, the obtained PS macroRAFT agent was denoted as PS10. Synthesis of PS-b-P(DMA-co-RhBHA) Amphiphilic Diblock Copolymer (Scheme 2). Typical procedures employed for synthesis of amphiphilic diblock copolymer, PS-b-P(DMA-co-RhBHA), are as follows. Into a reaction tube equipped with a magnetic stirring bar, PS10 (0.13 g, 0.1 mmol trithiocarbonate moieties), DMA (0.99 g, 10 mmol), RhBHA (28.5 mg, 0.05 mmol), AIBN (2 mg, 12 μmol), and THF (2.0 mL) were charged. The tube was carefully degassed by three freeze−pump−thaw cycles and then sealed under vacuum. After thermostating at 80 °C in an oil bath under magnetic stirring for 4 h, the reaction tube was quenched into liquid nitrogen, opened, and diluted with THF; the mixture was then precipitated into an excess of dry diethyl ether. The above dissolution−precipitation cycle was repeated for three times. The obtained diblock copolymer was further treated with an excess of AIBN (20 equiv relative to trithiocarbonate moieties) in THF. After being stirred at 80 °C for 4 h, the mixture was precipitated into an excess of diethyl ether, and the dissolution− precipitation cycle was repeated for three times. The product was then

dried in a vacuum oven overnight at room temperature, affording a slightly pink powder (0.84 g, yield: ∼72%; Mn = 9.2 kDa, Mw/Mn = 1.12, Figure S2, Supporting Information). The DP of P(DMA-coRhBHA) block was determined to be 80 by 1H NMR analysis in CDCl3 (Figure S1b, Supporting Information). Thus, the obtained amphiphilic diblock copolymer was denoted as PS10-b-P(DMA-coRhBHA)80. The molar content of RhBHA moieties (relative to the DMA block) was determined to be ∼0.45 mol % based on standard UV absorbance calibration curves. Synthesis of PS-b-P(DMA-co-CMA) Amphiphilic Diblock Copolymer (Scheme 2). PS-b-P(DMA-co-CMA) was synthesized by utilizing PS10 as the macroRAFT agent following similar procedures employed for the synthesis of PS-b-P(DMA-co-RhBHA) amphiphilic diblock copolymer (Scheme 2). The feed molar ratio of macroRAFT agent, DMA, CMA, and AIBN was 1/100/0.8/0.1. The reaction mixture was stirred at 80 °C for 4 h before quenching into liquid nitrogen. The product was further treated with an excess of AIBN (20 equiv relative to trithiocarbonate moieties) in THF. After stirring at 80 °C for 4 h, the amphiphilic diblock copolymer was precipitated into an excess of diethyl ether, and the dissolution−precipitation cycle was repeated for three times. The product was dried in a vacuum oven overnight at room temperature, affording a slightly yellowish powder (0.96 g, yield: ∼84%; Mn = 10.2 kDa, Mw/Mn = 1.12, Figure S2, Supporting Information). The DP of P(DMA-co-CMA) block was determined to be 90 by 1H NMR analysis in CDCl3 (Figure S1c, Supporting Information). Thus, the obtained amphiphilic diblock copolymer was denoted as PS10-b-P(DMA-co-CMA)90. The molar content of CMA moieties (relative to the DMA block) was determined to be ∼0.74 mol % based on standard UV absorbance calibration curves. Synthesis of PS10-b-P(DMA-co-HCMA)90 (Scheme 2). PS10-bP(DMA-co-CMA)90 (0.6 g, [CMA] = 0.4 mM) was dissolved in ethanol (20 mL). 4-Methoxyphenyl-hydrazine (0.11 g, 0.8 mM) and glacial acetic acid (∼100 μL) were then added. The reaction mixture was stirred at reflux under N2 atmosphere for 30 min, cooled to room 3941

dx.doi.org/10.1021/ma3006558 | Macromolecules 2012, 45, 3939−3947

Macromolecules

Article

temperature, and then precipitated into an excess of diethyl ether. The above dissolution−precipitation cycle was repeated for three times. After drying in a vacuum oven overnight at room temperature, the product was obtained as a yellowish powder (0.46 g, yield: ∼70.2%). CMA moieties in PS10-b-P(DMA- co-CMA)90 amphiphilic diblock copolymer were confirmed to be quantitatively converted into hydrazone-caged coumarin moieties (HCMA) as evidenced by 1H NMR characterization (Figure S1d, Supporting Information). Preparation of Mixed Micellar Solutions. Typical procedures employed for the preparation of mixed micellar solutions are as follows. PS10-b-P(DMA-co-HCMA)90 (30.0 mg) and PS10-b-P(DMAco-RhBHA)80 (5.0 mg) amphiphilic diblock copolymers were dissolved in 5.0 mL of THF at first. Under vigorous stirring, the solution was injected into 45 mL deionized water. After a further stirring for 30 min, THF was removed under reduced pressure. The final concentrations of PS10-b-P(DMA-co-HCMA)90 and PS10-b-P(DMA-co-RhBHA)80 in the mixed micellar solution were adjusted to 0.3 g/L and 0.05 g/L by diluting with pH 5.0 acetate buffer just prior to sensing experiments. Following similar procedures, pure PS10-b-P(DMA-co-HCMA)90 and PS10-b-P(DMA-co-RhBHA)80 micelles were also fabricated. Characterization. All 1H NMR spectra were recorded on a Bruker AV300 NMR spectrometer (resonance frequency of 300 MHz for 1H) operated in the Fourier transform mode. CDCl3 was used as the solvent. Molecular weights and molecular weight distributions were determined by gel permeation chromatography (GPC) equipped with Waters 1515 pump and Waters 2414 differential refractive index detector (set at 30 °C). The detection components used a series of two linear Styragel columns (HR2 and HR4) at an oven temperature of 45 °C. The eluent was THF at a flow rate of 1.0 mL/min. A series of low polydispersity polystyrene standards were employed for calibration. Dynamic LLS measurements were conducted on a commercial spectrometer (ALV/DLS/SLS-5022F) equipped with a multitau digital time correlator (ALV5000) and a cylindrical 22 mW UNIPHASE He−Ne laser (λ0 = 632 nm) as the light source. Scattered light was collected at a fixed angle of 90° for duration of ∼5 min. Distribution averages and particle size distributions were computed using cumulants analysis and CONTIN routines. All data were averaged over three measurements. AFM measurement was performed on a Digital Instrument Multimode Nanoscope IIID operating in the tapping mode under ambient conditions. Silicon cantilever (RFESP) with resonance frequency of ∼80 kHz and spring constant of ∼3 N/m was used. The set-point amplitude ratio was maintained at 0.7 to minimize sample deformation induced by the tip. The sample was prepared by dip coating aqueous micellar solutions onto freshly cleaved mica surfaces. Fluorescence spectra were recorded using an F4600 (Hitachi) spectrofluorometer at room temperature. The slit widths were both set at 5 nm for excitation and emission. All the fluorescence spectra were recorded after 30 min incubation at 25 °C upon addition of varying amount of metal ions unless otherwise stated. All UV−vis spectra were acquired on a Unico UV/vis 2802PCS spectrophotometer. All absorption spectra were recorded after 30 min incubation at 25 °C upon addition of varying amount of metal ions unless otherwise stated.

S1d, Supporting Information). In aqueous solution, PS10-bP(DMA-co-RhBHA)80 and PS10-b-P(DMA-co-HCMA)90 amphiphilic diblock copolymers spontaneously self-assemble into mixed micelles consisting of hydrophobic PS cores and hydrophilic PDMA coronas colabeled with RhBHA and HCMA moieties (Scheme 1). Spherical mixed micelles possess an intensity-average hydrodynamic radius, ⟨Rh⟩, of ∼59.5 nm and a size polydispersity, μ2/Γ2, of 0.12, as revealed by dynamic laser light scattering (LLS) and atomic force microscopy (AFM) characterization (Figure 1). Note that the close

Figure 1. (a) Hydrodynamic radius distribution, f(Rh), and (b) AFM height image recorded for mixed micelles self-assembled from PS10-bP(DMA-co-RhBHA)80 (0.05 g/L) and PS10-b-P(DMA- co-HCMA)90 (0.3 g/L) diblock copolymers.

proximity between RhBHA and HCMA moieties within micellar coronas allows for the occurrence of fluorescence resonance energy transfer (FRET) process once they are decaged via Hg2+-induced and Hg2+-catalyzed reactions (Scheme 1).64 In the next step, we investigated the time- and Hg2+ concentration-dependent decaging reaction of pure PS10-bP(DMA-co-HCMA)90 micelles in aqueous solution. It was found that initially the micellar solution is almost nonfluorescent due to the presence of hydrazone caging moiety.65 In the present work, by serendipity we discovered that for PS10b-P(DMA-co-HCMA)90 micelles, Hg2+ ions are strongly potent in catalyzing the decaging reaction of HCMA moieties, transforming them into highly fluorescent CMA moieties with an emission maximum at ∼420 nm (Scheme 1), as compared to those exhibited by other metal ions (Figures S3− S5, Supporting Information). As shown in Figure S3a,Supporting Information, upon addition of varying amount of Hg2+ ions (0−2.0 mM) into the PS10-b-P(DMA-co-HCMA)90 micellar solution (0.3 g/L), we can observe the time-dependent increase of emission intensity at 420 nm, which can be ascribed to Hg2+catalyzed decaging reaction of HCMA moieties. Higher Hg2+ concentration leads to more dramatic emission enhancement. Typically, the addition of 2.0 mM Hg2+ ions into the micellar

■

RESULTS AND DISCUSSION At first, well-defined amphiphilic diblock copolymers labeled with two types of fluorophores in the hydrophilic poly(N,Ndimethylacrylamide) (PDMA) block, PS-b-P(DMA-coRhBHA) and PS-b-P(DMA-co-CMA), were synthesized via reversible addition−fragmentation chain transfer (RAFT) copolymerization of DMA with RhBHA and CMA, respectively (Scheme 2). Table S1 (Supporting Information) summarizes structural parameters of diblock copolymers, together with the 1 H NMR and GPC characterization data (Figures S1 and S2, Supporting Information). Highly fluorescent coumarin (CMA) moieties in PS-b-P(DMA-co-CMA) were then quantitatively caged by reacting with 4-methoxyphenylhydrazine, affording nonfluorescent P(DMA-co-HCMA) diblock copolymer (Figure 3942

dx.doi.org/10.1021/ma3006558 | Macromolecules 2012, 45, 3939−3947

Macromolecules

Article

solution results in ∼33-fold cumulative increase in emission intensity after 30 min incubation. We are also aware of the fact that hydrazone caging functionality in HCMA is prone to spontaneous hydrolysis under mildly acidic media; however, control experiment (Figure S3b, Supporting Information) revealed that it takes ∼13 h for complete decaging reaction and the emission intensity only increased ∼1.7-fold within 30 min incubation. Thus, the above observed emission enhancement is predominantly due to Hg2+-catalyzed decaging reaction rather than acid-catalyzed hydrazone hydrolysis. To avoid possible interference from spontaneous hydrazone hydrolysis and considering that Hg2+-catalyzed decaging reaction is highly time-dependent, changes in fluorescence emission intensity in subsequent sections were mainly monitored for 30 min after addition of varying concentrations of Hg2+ ions or other metal ions. Further fluorescence titration experiments were conducted upon addition of varying amount of Hg2+ ions (0−2.0 mM) into the micellar solution of PS10-b-P(DMA-co-HCMA)90. As shown in Figure S4, Supporting Information, the emission intensity of micellar solution gradually increases with Hg2+ concentrations and an almost linear correlation can be established in the Hg2+ concentration range of 0−0.2 mM (Figure S4c, Supporting Information). Note that upon addition of 20 μM Hg2+ ions (1.0 equiv relative to HCMA moieties), the fluorescence emission was relatively weak after 30 min incubation (only ∼1.8-fold emission increase); whereas the presence of 2.0 mM Hg2+ ions (100.0 equiv) dramatically leads to the turn-on of intense blue emission, which can be facilely discerned by naked eyes (Figure 2a). Previously, Han et al.65 reported that Cu2+ ions can catalyze the decaging reaction of hydrazone-caged coumarin 343, which renders possible the construction of fluorescence turn-on Cu2+ probes. In the current case, we found that for micellar solution of PS10-b-

P(DMA-co-HCMA)90 containing hydrazone-caged coumarin moieties (HCMA), Hg2+ ions are more potent in catalyzing the decaging reaction compared to that exhibited by Cu2+ ions (Figure S5, Supporting Information). This is presumably due to the difference in fluorophore chemical structures and variation of solvent conditions (pure aqueous media in this work versus acetonitrile/aqueous buffer mixture). Upon incubating the micellar solution with 100.0 equiv (2.0 mM) of various metal ions for 30 min, Hg2+ and Cu2+ ions exhibit ∼33-fold and ∼9.6fold emission enhancement, respectively, whereas all other metal ions exhibit negligible effects (Figure S5, Supporting Information). The above results thus established that PS10-bP(DMA- co-HCMA)90 micelles can serve as a partially selective catalytic reaction-based fluorescence turn-on Hg2+ probe. On the other hand, RhBHA moieties within PS-b-P(DMAco-RhBHA) micelles are expected to exhibit much faster Hg2+induced ring-opening reaction, transforming the initially nonfluorescent RhBHA moieties in the spirolactam form into the highly fluorescent acyclic form. In agreement with our previous report concerning RhBHA-labeled amphiphilic and double hydrophilic diblock copolymer micelles,43,51 it was found that for 0.05 g/L PS10-b-P(DMA-co-RhBHA)80 micelles ([RhBHA] = 2 μM), the presence of as low as 10 μM Hg2+ ions (5.0 equiv) can lead to the fast decaging of RhBHA moieties and the process is essentially complete within ∼4−5 min (Figures S6 and S7, Supporting Information). The emission intensity at ∼590 nm characteristic band which are exclusively based on supramolecular recognition of decaged RhBHA moieties (acyclic form; Scheme 1) exhibits an initial dramatic increase and then quick stabilizes out, resulting in a cumulative ∼15.6-fold increase (Figures S6 and S7, Supporting Information). On the other hand, the presence of 10 μM Hg2+ ions only leads to negligible changes for PS10-b-P(DMA-coHCMA)90 micellar solution after 30 min incubation (Figures S3 and S6, Supporting Information). Macroscopic images taken under UV irradiation clearly revealed the turn-on of orange emission for PS10-b-P(DMA-co-RhBHA)80 micellar solution upon addition of 20 μM Hg2+ ions (Figure 2b). In contrary, the decaging of HCMA moieties of PS10-b-P(DMA-co-HCMA)90 micelles is not prominent at the same Hg2+ concentration at least for the first 30 min incubation duration (Figure 2a). The above results prompted us to further explore and design a novel type of fluorescent turn-on Hg2+ probe capable of working in a broad concentration range by integrating the two established Hg2+ sensing motifs with complementary working concentration ranges (Scheme 1). Mixed micelles coassembled from PS10 -b-P(DMA-coHCMA)90 and PS10-b-P(DMA-co-RhBHA)80 diblocks thus provide to be an excellent platform, in which RhBHA and HCMA moieties are both located within micellar coronas. The HCMA decaging kinetics for mixed micelles upon addition of varying amount of Hg2+ ions was then examined (Figure S8, Supporting Information) and found to be quite comparable to that of pure PS10-b-P(DMA-co-HCMA)90 micelles (Figure S3a, Supporting Information). Negligible changes of emission intensity at 420 nm were observed at low Hg2+ concentration ranges (5.0 equiv of Hg2+ ions. We tentatively ascribe this to the FRET process between decaged HCMA and decaged RhBHA moieties at higher Hg 2+ concentrations (Scheme 1), which can enhance the emission of decaged RhBHA (FRET acceptor).64 Figure S9, Supporting Information, shows the plot of emission intensity ratio, I420/I590, as a function of Hg2+ concentrations. In the low concentration range (0−10 μM), the ratio decreases from 1.2 to 0.07; however, upon further addition of Hg2+ ions (10 μM-2 mM), the trend of intensity ratio change is conversed, exhibiting an adverse increase from 0.07 to 0.81. We can thus tell that depending on the Hg2+ concentration range, which can be judged from the relative intensity of emission bands characteristic of decaged HCMA and RhBHA (Figure 3a,b), the fluorescent ratiometric detection of Hg2+ ions can be also achieved.

on features (Figure 3). In the Hg2+ concentration range of 0− 14 μM (0−7.0 equiv relative to RhBHA), we can only observe the dramatic increase of emission band at ∼590 nm characteristic of decaged RhBHA, whereas the emission band of decaged HCMA does not exhibit any appreciable changes (Figure 3, parts a, c, and e). At higher Hg2+ concentration ranges (20 μM to 2 mM), we can observe the concomitant increase of two emission bands at ∼420 and ∼590 nm characteristic of decaged HCMA and RhBHA moieties, respectively (Figure 3, parts b, d, and f). This suggests that HCMA moieties only start to be decaged at high Hg2+ concentrations (Figure S8, Supporting Information). UV absorption characterization further gives complementary results (Figure 4). If we arbitrarily define the Hg2+ detection limit as the concentration at which a 10% increase in the fluorescence emission intensity at 590 nm or at 420 nm can be measured, the Hg2+ detection limits were then determined to be ∼40 nM or ∼14 μM, respectively, based on the two characteristic emission bands (Figures 3e and 3d). In addition, fluorometric transitions of mixed micellar solution in the presence of varying amount of Hg2+ ions can be clearly discerned by the naked eye (Figure 2c). In the range of Hg2+ concentrations of 0−2.0 mM, the micellar solution exhibits sequential transitions from nonfluorescent to orange and to pink emissions. Thus, mixed micelles coassembled from the two diblock copolymers are capable of effective broad concentration range fluorometric 3944

dx.doi.org/10.1021/ma3006558 | Macromolecules 2012, 45, 3939−3947

Macromolecules

Article

Figure 5. (a) Fluorescence emission spectra and (b) relative emission intensities at 420 nm (blue bar) and 590 nm (red bar) (λex = 370 nm; slit widths: ex. 5 nm; em. 5 nm) recorded for the aqueous solution (pH 5.0 acetate buffer, 20 °C) of mixed micelles self-assembled from PS10-b-P(DMA-co-RhBHA)80 (0.05 g/L; [RhBHA] = 2 μM) and PS10b-P(DMA-co- HCMA)90 (0.3 g/L; [HCMA] = 20 μM) diblock copolymers after incubation for 30 min upon addition of 1000.0 equiv (relative to RhBHA moieties; 2.0 mM) Li+, Na+, K+, Ag+, Mg2+, Cd2+, Co2+, Ca2+, Cu2+, Zn2+, Pb2+, Fe2+, and Hg2+ ions, respectively.

Figure 4. UV−vis absorbance spectra recorded for the aqueous solution (pH 5.0 acetate buffer, 20 °C) of mixed micelles selfassembled from PS10-b-P(DMA-co-RhBHA)80 (0.05 g/L; [RhBHA] = 2 μM) and PS10-b-P(DMA-co-HCMA)90 (0.3 g/L; [HCMA] = 20 μM) diblock copolymers after incubation for 30 min upon addition of (a) 0−7.0 (0−14 μM) and (b) 10.0−1000.0 equiv of Hg2+ ions (20.0 μM to 2.0 mM) relative to RhBHA moieties.

the fluorometric detection of Hg2+ ions can be conducted over a broad dynamic concentration ranging from nM to mM levels (Figure S11, Supporting Information). Upon addition of 2.0 mM Cu2+ ions (1000.0 equiv relative to the RhBHA) into mixed micellar solution, no emission band of decaged RhBHA at ∼590 nm can be discerned. This is due to that Cu2+ ions can selectively quench the emission of acyclic RhBHA moieties.43,51 We then attempted to utilize this feature for the fluorescent ratiometric detection of Cu2+ ions based on mixed micelles pretreated with an excess of Hg2+ ions, in which both emission bands characteristic of decaged HCMA and RhBHA at 420 and 590 nm were switched on (Figure 3b). After incubating for 30 min upon addition of 0.4 mM various metal ions, emission spectra were then recorded (Figure 6a). It was found that only Cu2+ ions exhibit prominent quenching of the acyclic RhBHA band at ∼590 nm, whereas that of decaged HCMA at 420 nm remains essentially unaltered, and the latter can thus be utilized as an internal calibration standard for the ratiometric sensing of Cu2+ ions. All other metal ions lead to nondiscernible effects on both characteristic emission bands (Figure 6b). Finally, we examined the Cu2+ ion sensing capability of mixed micelles. Upon addition of varying amount of Cu2+ ions (0−0.4 mM, 0−200.0 equiv relative to RhBHA) into mixed micellar solution pretreated with Hg2+ ions, we can clearly observe the gradual quenching of emission band of decaged RhBHA (Figure 6c). The fluorescence intensity ratio, I420/I590, increased from 0.63 to 16.6 (∼26.3-fold) over the whole concentration range and the detection limit was determined to be ∼1.2 μM (Figure 6d).

We further examined the metal ion sensing selectivity of mixed micelles (Figures 5 and S10, Supporting Information). Upon addition of 2.0 mM of various metal ions (1000.0 equiv relative to RhBHA), it was found that after 30 min incubation only Hg2+ ions can simultaneously induce the enhancement of emission bands characteristics of decaged HCMA and RhBHA, exhibiting ∼29-fold and ∼45-fold emission intensity increase at ∼420 nm and ∼590 nm, respectively (Figure 5). Cu2+ ion solely leads to the turn-on of decaged HCMA band at 420 nm (∼8.5-fold), whereas Ag+ ions only leads to relatively weak turn-on of decaged RhBHA emission (∼3.6-fold). All other metal ions do not cause any appreciable changes on the two characteristic emission bands. Also note that from Figure S10, Supporting Information, we can tell that the decaging of HCMA moieties of mixed micelles by Hg2+ and Cu2+ ions follows comparable kinetics to those observed for pure PS10-bP(DMA-co-HCMA)90 micelles (Figure S5, Supporting Information), with Hg2+ being the most potent catalyst for HCMA decaging reaction. Macroscopic images taken under UV irradiation for mixed micellar solution after incubating with various metal ions are shown in Figure 2d. The presence of 2.0 mM Cu2+ and Hg2+ ions leads to the turn-on of blue and pink emissions, respectively. The above results suggest that mixed micelles exhibit excellent selectivity toward Hg2+ ions and quantitative differentiation between Hg2+, Cu2+, Ag+, and other metal cations can be facilely accomplished; most importantly, 3945

dx.doi.org/10.1021/ma3006558 | Macromolecules 2012, 45, 3939−3947

Macromolecules

Article

Figure 6. (a and c) Fluorescence emission spectra and (b and d) emission intensity ratios, I420/I590, (λex = 370 nm; slit widths: ex. 5 nm, em. 5 nm) recorded for the aqueous solution (pH 5.0 acetate buffer, 20 °C) of mixed micelles self-assembled from PS10-b-P(DMA-co- RhBHA)80 (0.05 g/L; [RhBHA] = 2 μM) and PS10-b-P(DMA-co-HCMA)90 (0.3 g/L; [HCMA] = 20 μM) diblock copolymers (a and b) upon addition of 200.0 equiv (relative to RhBHA moieties) Li+, Na+, K+, Ca2+, Zn2+, Cd2+, Ag+, Mg2+, Pb2+, Fe2+, Hg2+, Co2+, and Cu2+, respectively, and (c and d) upon gradual addition of Cu2+ ions (0−200.0 equiv relative to RhBHA moieties; 0−0.4 mM). The mixed micellar solution was pretreated with an excess of Hg2+ (2.0 mM) for 30 min before the addition of metal ions.

types of kinetically resolved fluorophore decaging reactions, namely, Hg2+-induced ring-opening reaction of RhBHA residues and Hg2+-catalyzed hydrolysis of caged HCMA moieties, and utilizes mixed diblock copolymer micelles as the ensembling scaffold (Scheme 1). The two types of decaging reactions possess distinctly different, but self-complementary reaction kinetics and optimum working concentration ranges. In addition, the Hg2+-sensing platform can be further employed for the fluorescent ratiometric detection of Cu2+ ions. We trust that the reported strategy of ensembling two partially selective chemical reaction-based fluorometric sensing motifs to achieve enhanced metal ion sensing selectivity and broadened working concentration range can be further generalized for the construction of other ultraselective and broad dynamic range sensing systems.

To further investigate the detection selectivity of mixed micelles-based Hg2+ ion sensing platform in the presence of various competing metal ions, fluorescence emission intensities of mixed micellar solution upon addition of 0.4 mM Li+, Na+, K+, Ag+, Mg2+, Cd2+, Co2+, Ca2+, Fe2+, Zn2+, Pb2+, and Cu2+ ions in the absence and presence of 2.0 mM Hg2+ ions were conducted (Figure S12, Supporting Information). In the absence of Hg2+ ions (Figure S12, Supporting Information, blue bar), Ag+ ions and Cu2+ ions can only slightly turn on the fluorescence emission at 590 and 420 nm, respectively, for a incubation duration of 30 min. On the other hand, the coexistence of Hg2+ ions with metal ions such as Li+, Na+, K+, Ag+, Mg2+, Cd2+, Co2+, Ca2+, Fe2+, Zn2+, and Pb2+ can simultaneously turn on the emission of HCMA and RhBHA moieties (Figure S12, Supporting Information, red bar), confirming the detection selectivity toward Hg2+ ions. However, the coexistence of Hg2+ and Cu2+ ions represented an intriguing exception. In this case, the emission at 420 nm was further enhanced compared to that in the absence of Hg2+ ions, whereas the emission at 590 nm was completely quenched (Figure 6),49,57 and this feature can be employed to qualitatively tell the coexistence of Hg2+ and Cu2+ ions. Thus, the further accurate quantification of Hg2+ ions in the presence of Cu2+ ions shall rely on the removal of Cu2+ ions at first by utilizing strong chelating ligands such as EDTA (ethylenediaminetetraacetic acid)-immobilized resins.

■

ASSOCIATED CONTENT

S Supporting Information *

Table summarizing the structural parameters and figures showing 1H NMR, GPC, and fluorescence spectra, timedependent changes in fluorescence emission intensities, and schematic interpretation of enhanced metal ion sensing selectivity and broadened effective working concentration range. This material is available free of charge via the Internet at http://pubs.acs.org.

■

■

CONCLUSION In summary, we constructed a new type of polymeric fluorescent Hg2+ probe covering a broad effective concentration range from nM to mM levels and exhibiting considerably enhanced detection selectivity. The probe design integrates two

AUTHOR INFORMATION

Corresponding Author

*E-mail: (S.L.) [email protected]; (G.Z.) [email protected]. Notes

The authors declare no competing financial interest. 3946

dx.doi.org/10.1021/ma3006558 | Macromolecules 2012, 45, 3939−3947

Macromolecules

■

Article

(32) Sau, T. K.; Rogach, A. L.; Jaeckel, F.; Klar, T. A.; Feldmann, J. Adv. Mater. 2010, 22, 1805−1825. (33) Liu, Q.; Peng, J. J.; Sun, L. N.; Li, F. Y. ACS Nano 2011, 5, 8040−8048. (34) Dave, N.; Chan, M. Y.; Huang, P. J. J.; Smith, B. D.; Liu, J. W. J. Am. Chem. Soc. 2010, 132, 12668−12673. (35) Wang, J.; Liu, B. Chem. Commun. 2008, 4759−4761. (36) Miyake, Y.; Togashi, H.; Tashiro, M.; Yamaguchi, H.; Oda, S.; Kudo, M.; Tanaka, Y.; Kondo, Y.; Sawa, R.; Fujimoto, T.; Machinami, T.; Ono, A. J. Am. Chem. Soc. 2006, 128, 2172−2173. (37) Matsushita, M.; Meijler, M. M.; Wirsching, P.; Lerner, R. A.; Janda, K. D. Org. Lett. 2005, 7, 4943−4946. (38) Liu, J.; Lu, Y. Angew. Chem., Int. Ed. 2007, 46, 7587−7590. (39) Ono, A.; Togashi, H. Angew. Chem., Int. Ed. 2004, 43, 4300− 4302. (40) Wen, S.; Zeng, T.; Liu, L.; Zhao, K.; Zhao, Y.; Liu, X.; Wu, H. C. J. Am. Chem. Soc. 2011, 133, 18312−18317. (41) Liu, X.; Tang, Y.; Wang, L.; Zhang, J.; Song, S.; Fan, C.; Wang, S. Adv. Mater. 2007, 19, 1662−1662. (42) Gu, Z.; Zhao, M. X.; Sheng, Y. W.; Bentolila, L. A.; Tang, Y. Anal. Chem. 2011, 83, 2324−2329. (43) Hu, J. M.; Li, C. H.; Liu, S. Y. Langmuir 2010, 26, 724−729. (44) Tang, Y. L.; He, F.; Yu, M. H.; Feng, F. D.; An, L. L.; Sun, H.; Wang, S.; Li, Y. L.; Zhu, D. B. Macromol. Rapid Commun. 2006, 27, 389−392. (45) Fang, Z.; Pu, K. Y.; Liu, B. Macromolecules 2008, 41, 8380− 8387. (46) Murkovica, I.; Wolfbeisb, O. S. Sens. Actuator B-Chem. 1997, 39, 246−251. (47) Ma, C.; Zeng, F.; Huang, L.; Wu, S. J. Phys. Chem. B 2011, 115, 874−882. (48) Kim, I. B.; Phillips, R.; Bunz, U. H. F. Macromolecules 2007, 40, 814−817. (49) Hu, J. M.; Liu, S. Y. Macromolecules 2010, 43, 8315−8330. (50) Wan, X. J.; Liu, T.; Liu, S. Y. Langmuir 2011, 27, 4082−4090. (51) Hu, J. M.; Dai, L.; Liu, S. Y. Macromolecules 2011, 44, 4699− 4710. (52) Lee, J.; Jun, H.; Kim, J. Adv. Mater. 2009, 21, 3674−3677. (53) Li, C. H.; Liu, S. Y. Chem. Commun. 2012, 48, 3262−3278. (54) Li, C. H.; Hu, J. M.; Liu, S. Y. Soft Matter 2012, 8, DOI: 10.1039/C2SM25582K. (55) Zhang, L.; Murphy, C. S.; Kuang, G. C.; Hazelwood, K. L.; Constantino, M. H.; Davidson, M. W.; Zhu, L. Chem. Commun. 2009, 7408−7410. (56) Nolan, E. M.; Jaworski, J.; Okamoto, K.; Hayashi, Y.; Sheng, M.; Lippard, S. J. J. Am. Chem. Soc. 2005, 127, 16812−16823. (57) Goldsmith, C. R.; Lippard, S. J. Inorg. Chem. 2006, 45, 555−561. (58) Shults, M. D.; Pearce, D. A.; Imperiali, B. J. Am. Chem. Soc. 2003, 125, 10591−10597. (59) Komatsu, K.; Kikuchi, K.; Kojima, H.; Urano, Y.; Nagano, T. J. Am. Chem. Soc. 2005, 127, 10197−10204. (60) Cho, D. G.; Sessler, J. L. Chem. Soc. Rev. 2009, 38, 1647−1662. (61) Yoon, J.; Kim, H. N.; Guo, Z. Q.; Zhu, W. H.; Tian, H. Chem. Soc. Rev. 2011, 40, 79−93. (62) Studer, P.; Stossel, R.; Scheifele, P.; Matsumoto, Y. Polymerizable Copolymers for Alignment Layers [P] PCT: WO2005/014677. (63) Lai, J. T.; Filla, D.; Shea, R. Macromolecules 2002, 35, 6754− 6756. (64) Lin, W.; Yuan, L.; Cao, Z.; Feng, Y.; Song, J. Angew. Chem., Int. Ed. 2010, 49, 375−379. (65) Kim, M. H.; Jang, H. H.; Yi, S.; Chang, S. K.; Han, M. S. Chem. Commun. 2009, 4838−4840.

ACKNOWLEDGMENTS The financial support from the National Natural Scientific Foundation of China (NNSFC) Project (91027026 and 51033005) and the Fundamental Research Funds for the Central Universities is gratefully acknowledged.

■

REFERENCES

(1) Boening, D. W. Chemosphere 2000, 40, 1335−1351. (2) Zheng, W.; Aschner, M.; Ghersi-Egea, J. F. Toxicol. Appl. Pharmacol. 2003, 192, 1−11. (3) Nolan, E. M.; Lippard, S. J. Chem. Rev. 2008, 108, 3443−3480. (4) Selid, P. D.; Xu, H.; Collins, E. M.; Face-Collins, M. S.; Zhao, J. X. Sensors 2009, 9, 5446−5459. (5) Song, Y.; Wei, W.; Qu, X. Adv. Mater. 2011, 23, 4215−4236. (6) Zhao, Q. A.; Li, F. Y.; Huang, C. H. Chem. Soc. Rev. 2010, 39, 3007−3030. (7) Zhang, X. L.; Xiao, Y.; Qian, X. H. Angew. Chem., Int. Ed. 2008, 47, 8025−8029. (8) Chae, M. Y.; Czarnik, A. W. J. Am. Chem. Soc. 1992, 114, 9704− 9705. (9) Yang, Y. K.; Yook, K. J.; Tae, J. J. Am. Chem. Soc. 2005, 127, 16760−16761. (10) Chen, X. Q.; Nam, S. W.; Jou, M. J.; Kim, Y.; Kim, S. J.; Park, S.; Yoon, J. Org. Lett. 2008, 10, 5235−5238. (11) Liu, B.; Tian, H. Chem. Commun. 2005, 3156−3158. (12) Lee, M. H.; Cho, B. K.; Yoon, J.; Kim, J. S. Org. Lett. 2007, 9, 4515−4518. (13) Ros-Lis, J. V.; Marcos, M. D.; Martinez-Manez, R.; Rurack, K.; Soto, J. Angew. Chem., Int. Ed. 2005, 44, 4405−4407. (14) Nolan, E. M.; Lippard, S. J. J. Mater. Chem. 2005, 15, 2778− 2783. (15) Climent, E.; Marcos, M. D.; Martinez-Manez, R.; Sancenon, F.; Soto, J.; Rurack, K.; Amoros, P. Angew. Chem., Int. Ed. 2009, 48, 8519− 8522. (16) Diez-Gil, C.; Martinez, R.; Ratera, I.; Hirsh, T.; Espinosa, A.; Tarraga, A.; Molina, P.; Wolfbeis, O. S.; Veciana, J. Chem. Commun. 2011, 47, 1842−1844. (17) Yang, H.; Zhou, Z.; Huang, K.; Yu, M.; Li, F.; Yi, T.; Huang, C. Org. Lett. 2007, 9, 4729−4732. (18) Lu, H.; Xiong, L.; Liu, H.; Yu, M.; Shen, Z.; Li, F.; You, X. Org. Biomol. Chem. 2009, 7, 2554−2558. (19) Yoon, S.; Albers, A. E.; Wong, A. P.; Chang, C. J. J. Am. Chem. Soc. 2005, 127, 16030−16031. (20) Descalzo, A. B.; Martinez-Manez, R.; Radeglia, R.; Rurack, K.; Soto, J. J. Am. Chem. Soc. 2003, 125, 3418−3419. (21) Mayr, T.; Liebsch, G.; Klimant, I.; Wolfbeis, O. S. Analyst 2002, 127, 201−203. (22) Wu, Y. Q.; Jing, H.; Dong, Z. S.; Zhao, Q.; Wu, H. Z.; Li, F. Y. Inorg. Chem. 2011, 50, 7412−7420. (23) Loe-Mie, F.; Marchand, G.; Berthier, J.; Sarrut, N.; Pucheault, M.; Blanchard-Desce, M.; Vinet, F.; Vaultier, M. Angew. Chem., Int. Ed. 2010, 49, 424−427. (24) Freeman, R.; Finder, T.; Willner, I. Angew. Chem., Int. Ed. 2009, 48, 7818−7821. (25) Li, M.; Wang, Q.; Shi, X.; Hornak, L. A.; Wu, N. Anal. Chem. 2011, 83, 7061−7065. (26) Xie, J. P.; Zheng, Y. G.; Ying, J. Y. Chem. Commun. 2010, 46, 961−963. (27) Huang, C. C.; Chang, H. T. Anal. Chem. 2006, 78, 8332−8338. (28) Rex, M.; Hernandez, F. E.; Campiglia, A. D. Anal. Chem. 2006, 78, 445−451. (29) Lee, J. S.; Han, M. S.; Mirkin, C. A. Angew. Chem., Int. Ed. 2007, 46, 4093−4096. (30) Huang, C. C.; Yang, Z.; Lee, K. H.; Chang, H. T. Angew. Chem., Int. Ed. 2007, 46, 6824−6828. (31) Saleh, S. M.; Ali, R.; Wolfbeis, O. S. Chem.Eur. J. 2011, 17, 14611−14617. 3947

dx.doi.org/10.1021/ma3006558 | Macromolecules 2012, 45, 3939−3947