1

Supplementary Information

A red-emitting fluorescent probe for detecting Hg2+ in aqueous medium, living cells and organism with a large Stokes shiftLei Yang,a‡ Yuanan Su,a‡ Yani Geng,a Haiqing Xiong,a Jinliang Han,a Qian Fanga and Xiangzhi Songa*

a College of Chemistry & Chemical Engineering, Central South University, Changsha, Hunan Province, P. R. China, 410083.

* To whom correspondence should be addressed. Fax: +86-731-88836954; Tel: +86-731-88836954; E-mail: song@rowland.harvard.edu

‡ These authors contributed equally to this work.

Table S1 ....................................................................................................................................................................2Table S2 ....................................................................................................................................................................3Fig. S1 .......................................................................................................................................................................4Fig. S2 .......................................................................................................................................................................4Fig. S3 .......................................................................................................................................................................5Fig. S4 .......................................................................................................................................................................5Fig. S5 .......................................................................................................................................................................6Fig. S6 .......................................................................................................................................................................6Fig. S7 .......................................................................................................................................................................7Fig. S8 .......................................................................................................................................................................7Fig. S9 .......................................................................................................................................................................8Fig. S10 .....................................................................................................................................................................8Fig. S11 .....................................................................................................................................................................9Fig. S12 .....................................................................................................................................................................9Fig. S13 ...................................................................................................................................................................10Fig. S14 ...................................................................................................................................................................10Fig. S15 ...................................................................................................................................................................11Fig. S16 ...................................................................................................................................................................11

Electronic Supplementary Material (ESI) for Organic & Biomolecular Chemistry.This journal is © The Royal Society of Chemistry 2018

2

Table S1 Comparison of Fluorescent Probes for Hg2+.

Probeλex/λem (nm)

Stokes shift （nm）

Detection limit (nM)

Analytical applications

Literature

O

N

O

NH2

N N

HOOC

520/573 53 97River water,

pond water and living cells

Analytica Chimica Acta,

2016, 934, 218-225

N O S

N

S 487/511 24 1700 NoChem.

Commun., 2009, 3560-3562

O

N

S

N

S

HO OH

509/529 20 39 Living cellsTalanta, 2017, 170, 103–110

S

SS

S O 370/418 48 50 NoChem.

Commun., 2005, 2161–2163

O

N

O

N N

N

N

NHS

S

HN

NH

SNH

520/583 63 304Pond water and

tap water

Dyes and Pigments, 2016,

127, 94-99

S

NO

HNO

365/500 135 100 NoOrg. Lett., 2011, 13, 3422-3425

S SN N

N N

380/458 78 80 NoOrg. Lett., 2012, 14, 6084-6087

O O

NCCN

NC

560/613 53 10River water and

living cells

Sensors and Actuators B,

2014, 191, 605–611

O

N

S

NH

NH

534/566 32 39 Living cellsChem.

Commun., 2010, 46, 3529–3531

3

NO O

NH2

420/480 66 50 NoOrg. Lett., 2010, 22, 5310-5313

N

N O

CN

CN 475/625 150 7.1Water samples, living cells and

organismThis work

Table S2 Determination of Hg2+ in water samples.Sample HgCl2 spiked (μM) HgCl2 recovered (μM) Recovery (%)

0 not detected2 2.07 ± 0.03 1034 3.82 ± 0.12 956 5.78 ± 0.13 968 7.78 ± 0.09 9710 10.50 ± 0.07 10512 12.61 ± 0.21 10514 13.48 ± 0.19 9616 16.51 ± 0.14 10318 17.74 ± 0.19 99

Tap water

20 19.48 ± 0.03 970 not detected2 2.24 ± 0.09 1094 3.82 ± 0.05 966 5.69 ± 0.12 958 7.76 ± 0.20 9710 10.18 ± 0.16 10212 12.64 ± 0.19 10514 13.95 ± 0.22 10016 16.37 ± 0.06 10218 17.47 ± 0.08 97

Yangtze River water

20 19.90 ± 0.15 990 not detected2 1.82 ± 0.13 914 3.81 ± 0.21 956 5.48 ± 0.19 918 7.69 ± 0.07 9610 10.23 ± 0.06 10212 12.36 ± 0.08 10314 13.75 ± 0.11 9816 16.34 ± 0.12 10218 17.92 ± 0.07 99

Xiang River water

20 19.86 ± 0.20 99

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Fig. S1 Absorption spectra of Probe 1 (10.0 µM) in the presence/absence of Hg2+ (5.0 equiv.) in HEPES buffer (20 mM, pH=7.4, containing 30% EtOH) incubation at room temperature for 60 min. Inset: photographs of Probe 1 without (left) and with (right) 5.0 equiv. of Hg2+.

Fig. S2 Absorption (black) and emission spectra (red) of dye 5 (10.0 µM) in HEPES buffer (20 mM, pH=7.4, containing 30% EtOH).

5

Fig. S3 Absorbance at 495 nm of Probe 1 (10.0 µM) versus the concentration of Hg2+ in HEPES buffer (20 mM, pH=7.4, containing 30% EtOH). Inset: the linear relationship between the absorbance of the solution of Probe 1 and Hg2+ concentration (0.0-25.0 µM).

Fig. S4 1H NMR spectrum of compound 3 in CDCl3.

N

N

CHO

OBr

6

Fig. S5 13C NMR spectrum of compound 3 in CDCl3.

301.1410 340.0672

363.05671+

381.0297

mercury\mercury 1: +MS, 0.1-0.2min #3-10

0.0

0.5

1.0

1.5

2.0

2.5

6x10Intens.

300 325 350 375 400 425 450 475 500 m/z

Fig. S6 HRMS spectrum of compound 3.

N

N

CHO

OBr

N

N

CHO

OBr

7

Fig. S7 1H NMR spectrum of compound 4 in CDCl3.

Fig. S8 13C NMR spectrum of compound 4 in CDCl3.

N

N

CHO

O

N

N

CHO

O

8

283.12811+

296.2421

340.26831+

360.30961+

384.2944

mercury\mercury 2: +MS, 0.1-0.3min #5-17

0.00

0.25

0.50

0.75

1.00

1.25

1.50

7x10Intens.

200 225 250 275 300 325 350 375 400 m/z

Fig. S9 HRMS spectrum of compound 4.

Fig. S10 1H NMR spectrum of Probe 1 in CDCl3.

N

N O

CN

CN

N

N

CHO

O

9

Fig. S11 13C NMR spectrum of Probe 1 in CDCl3.

306.23851+

308.1309

309.1344

310.1378

316.1922

mercury\mercury probe: +MS, 0.1-0.2min #4-13

0

2

4

6

5x10Intens.

306 308 310 312 314 316 m/z

Fig. S12 HRMS spectrum of compound Probe 1.

N

N O

CN

CN

N

N O

CN

CN

10

Fig. S13 Plot of the natural log of fluorescence intensity change for Probe 1 (10.0 µM) with Hg2+ (5.0 equiv.) as a function of the reaction time in HEPES buffer (20 mM, pH=7.4, containing 30% EtOH). kobs = 0.04579 min-1.

Fig. S14 1H NMR spectrum of the reaction product between Probe 1 and Hg2+ in CDCl3.

N

N

O

CN

CNN

N

O

CN

Probe 1 Dye 5

HgCl2NH

11

17:36:45 11-Jul-20154-1

m/z220 230 240 250 260 270 280 290 300 310 320 330 340

%

0

100LZX-88 5 (0.094) AM (Cen,4, 80.00, Ht,5000.0,0.00,1.00); Sm (Mn, 2x3.00); Cm (1:31) TOF MS ES+

1.49e3283.1557

274.2751

246.2435218.2111

284.1557

318.3021284.3288302.3060

285.1549 330.3371

Fig. S15 HRMS spectrum of the reaction product between Probe 1 and Hg2+.

Fig. S16 Cytotoxicity assay of Probe 1 at different concentration for HeLa cells.

N

N

O

CN

CNN

N

O

CN

Probe 1 Dye 5

HgCl2NH