Development and applications of fluorogenic probes for mercury(II) based on vinyl ether oxymercuration

Abstract

Mercury is a major threat to the environment and to human health. It is highly desirable to develop a user-friendly kit for on-site mercury detection. Such a method must be able to detect mercury below the threshold levels for drinking water, 1-2 ppb. We developed a fluorescence method based on the oxymercuration of vinyl ethers to detect mercury in dental and environmental samples. Chloride ions interfered with the oxymercuration reaction, but the addition of AgNO(3) solved this problem. Fine electronic and structural tuning led to the development of a more responsive probe that was less sensitive to chloride ion interference. This second-generation probe could detect 1 ppb mercury ions in water.

Figure 1

Conversion of 3 to 1 at 25 °C. [3] = 1.0 μM for all of the experiments. Except for (c), all of the reactions were performed for 1 h. Fluorescence intensities were measured after addition of 1.23 M phosphate pH 7 buffer. (a) pH-dependence. (b) Metal selectivity in a 50 mM phthalate pH 4 buffer. Metal reagents: AgNO₃, AuCl₃, BaCl₂, CaCl₂, CdCl₂•2.5H₂O, CoCl₂, CrCl₃, CuCl₂•2H₂O, FeCl₃, HgCl₂, KCl, LiCl, MgCl₂, MnCl₂•4H₂O, NaCl, NiCl₂, Pb(NO₃)₂, Pd(NO₃)₂, PtCl₂, Rh(PPh₃)₃, RuCl₃, and ZnCl₂. (c) Time-course of the oxymercuration reaction. [Hg(II)] = 0.30 μM, 50 mM phthalate pH 4 buffer. (d) Correlation between fluorescence intensities and [Hg(II)] in the presence and absence of N-chlorosuccinimide (NCS). The experiments were performed in a 50 mM phthalate pH 4 buffer in triplicate. The graph shows the mean values and standard deviations.

Figure 2

Compatibility of our method to detect Hg(II) in the presence of inorganic ions. These experiments were performed in pH 4 buffer. (a) Interference by inorganic materials. [HgCl₂] = 2.5 μM, [reagent shown] = 25 μM. (b) Interference by NaCl but not by NaNO₃.

Figure 3

Applications of probe 3 for the detection of mercury species in environmental and dental samples. All detection was performed after adjustment of pH of samples to pH 4. (a) Comparison of river water and commercial pH 4 buffer. [HgCl₂] = 2.2 μM (440 ppb). In the absence of AgNO₃, Hg(II) cannot be detected by the method (left). In the presence of AgNO₃, Hg(II) can be detected (middle). (b) Mercury detection in river water. [HgCl₂] = 0–256 ppb, [AgNO₃] = 2.0 mM. (c) Mercury detection in river water in the presence of organic compound. [AgNO₃] = 2.0 mM, [compound] = 100 ppb. (d) Dental samples. [NCS] = 500 μM.

Figure 4

(a) pH titration of compound 13. Fluorescence intensity at 515 nm was monitored at ~0.3 pH intervals. (b) UV-Vis absorption spectra of 12 and 13 in 1% DMSO/pH 8 buffer. (c) Emission spectra of 12 and 13 in 0.1% DMSO/pH 8 buffer.

Figure 5

Comparison of reaction conditions for the conversion of 12 to 13. (a) pH-dependence. Fluorescence intensities were measured after addition of 1.3 M pH 7 buffer and 500 mM pH 10 buffer. (b) Metal selectivity. All the metals were tested at 5 μM in pH 4 buffer. (c) Hg detection in the presence of various metal ions. All of the reactions were performed in pH 4 buffer. Metal reagents: AgNO₃, AuCl₃, BaCl₂, CaCl₂, CdCl₂•2.5H₂O, CoCl₂, CrCl₃, CuCl₂•2H₂O, FeCl₃, HgCl₂, KCl, LiCl, MgCl₂, MnCl₂•4H₂O, NaCl, NiCl₂, Pb(NO₃)₂, Pd(NO₃)₂, PtCl₂, Rh(PPh₃)₃, RuCl₃, and ZnCl₂. (d) Correlation between fluorescence intensities at 515 nm and [Hg(II)]. The experiments were performed at pH 4 in triplicate. (e) The effect of anion. All detection was performed in pH 4 buffer.

Figure 6

Comparisons of reactivity between probes developed by our group. [probe] = 1.0 μM, [Hg(II)] = 0.3 μM. (a) Time-course of the oxymercuration reaction at 25 °C in 50 mM phthalate pH 4.0 buffer. The comparison between probe 12 and 14 was shown. (b) Time-course of the oxymercuration reaction at 25 °C in 50 mM phthalate pH 4.0 buffer. The comparison between probe 3 and 12 was shown. To focus on their initial rate, fluorescence measurements were carried out every 1 min for 18 min. (c) Time-course of the oxymercuration reaction in 50 mM phosphate pH 7 buffer. The comparison between probe 3 and 12 was shown.

Figure 7

Application of 12 for the detection of Hg(II) in river water. Titration of Hg(II) in river water was performed. [12] = 1.0 μM; [Hg(II)] = 0, 0.25, 0.5, 1, 2, and 4 ppb; [AgNO₃] = 2.0 mM; 0.5% DMSO in 50 mM phthalate pH 4 water; 1 h; 25 °C.

Scheme 1

(a) Two-step sequence to cleave an allyl ether. (b) Platform for a fluorescence off-on switch. (c) Preparation of 3 and its reaction with HgCl₂ to form 1.

Scheme 2

Preparation of 12 and its reaction with HgCl₂ to form 13.

Scheme 3

Summary of this work.