Signaling preferences of substituted pyrrole coupled six-membered spirocyclic rhodamine probes towards Hg 2+ ion detection

Transcription

1 Electronic Supplementary Material (ESI) for rganic & Biomolecular Chemistry. This journal is The Royal Society of Chemistry 6 Signaling preferences of substituted pyrrole coupled six-membered spirocyclic rhodamine probes towards Hg + ion detection Biswonath Biswal, Debajani Mallick, Bamaprasad Bag* Colloids and Materials Chemistry Department, Academy of Scientific and Innovative Research, CSIR- Institute of Minerals and Materials Technology, P..: R.R.L., Bhubaneswar-75 3, disha, India. Fax: (+) ; Tel: (+ 9) , bpbag@immt.res.in Electronic Supplementary Information Experimental All the reagent grade chemicals were used without purification unless otherwise specified.,4-dimethyl pyrrole carboxaldehyde, rhodamine B base and the metal-perchlorate salts were obtained from Sigma Aldrich (India) and used as received. Anhydrous sodium sulphate, sodium borohydride, silica gel for column chromatography, acids and the solvents were received from Spectrochem Pvt Ltd (India). All the solvents were freshly distilled prior to use for absorption and fluorescence measurements. Fluorescence quantum yields were determined by comparing the corrected spectrum with that of rhodamine G (φ F =.95) in EtH by taking area under total emission using following eqn., eqn. S where φ S and φ R are radiative quantum yields, F S and F R are area under the fluorescence spectra, A S and A R are absorbances (at the excited wavelength) of respective samples and the reference respectively; η S and η R are the refractive indices of the solvent used for the sample and the reference. The quantum yield of Rhodamine G was measured using quinine sulfate in H S 4 as reference excited at (λ ex ) 35 nm. The standard quantum yield value thus obtained was used for the calculation of the quantum yield of the samples. The graphical representations of synthesized compounds are given in Scheme S for quick reference. H a Cl H H H a H H R Scheme S: Structure of the compounds R SP

2 Fig. S: H-MR spectrum of in CDCl 3. Fig. S: 3 C-MR spectrum of in CDCl 3. SP

3 Fig. S3: ESI-MS spectrum of Fig. S4: H-MR spectrum of a in CDCl 3. SP3

4 Fig. S5: 3 C-MR spectrum of a in CDCl 3. Fig. S6: ESI-MS spectrum of a SP4

5 Fig. S7: H-MR spectrum of in CD 3 C. Fig. S8: 3 C-MR spectrum of in CD 3 C. SP5

6 Fig. S9: ESI-MS spectrum of Abs Hg + (blank) + Mn + +Fe + +Co + +i + +Cu + +Zn + +Ag + +Cd + +Pb + +Hg + Abs Hg + (blank) +Mn + +Fe + +Co + +i + +Cu + +Zn + +Ag + +Pb + +Cd + +Hg Fig. S: Absorption spectra of and in presence of various metal ions. [ or ] = µm, MeC- H (: v/v, ph 7.). Fig. S: Change in colour in the solution of ( -5 M) in MeC-H (: v/v, ph 7.) upon addition of,,, 3 and 5 eq. of Hg + under normal light and upon irradiation at 45nm. SP6

7 Fig. S: Ground state HM and LUM of (left) and (right) as elucidated from DFT calculations. The energies of frontier orbitals and corresponding distribution of π-electrons shows that the electron densities located over xanthene ring in HM while those over spiro-ring in LUM. The HM-LUM energy gap shows that these probes do not absorb in the visible region, rather exhibits absorption transitions at 3 nm () and 33 nm() respectively, which is complemented well to the experimental observations. SP7

8 Photon counts.x 4 (MeC) (MeC:H, : v/v) 8.x 3 6.x 3 4.x 3 (Ethanol) +Hg + (MeC) +Hg + (Ethanol) +Hg + (MeC:H, : v/v).x 3. Time (t) in ns Fig S3: Time-resolved fluorescence exponentially fitted decay profile of in absence and presence of Hg + ion in MeC, MeC-H (: v/v) and EtH medium. Table ST-: Fit-results to the exponential decay curve obtained with time-correlated single photon counting technique with single exponential fit equation A+B exp (-t/τ) Solvent MeC:H (: v/v) MeC EtH +Hg + +Hg + +Hg + τ (av., ns) A B σ (std. dev.) χ Abs..3. λ abs = 557 nm obs Abs λ abs = 557 nm obs [Hg(II)]/([Hg(II)] + []) Mole fraction [Pb + ]/ ([Pb + ]+[]) Fig. S4: Change in absorption as a function of mole fraction of added metal ion(job s plot) for determination of complexation stoichiometry, with Hg + and with Pb + ion respectively. SP8

9 5 Fluo. Int. (a. u., xe6) Eq. of Hg + added.6x -5.x -5 Eqn. Y = m*x+c Adj. R sq Value Std. err. /(I-I) Intercept -6.53E E-7 /(I-I) Slope E E- /(I-I ) 8.x -6 4.x /[Hg(II)].5 Fig. S5: Plot of change in fluorescence intensity of as a function of added [Hg(II)] ions in MeC- H (: v/v). Experimental conditions: [] = 5 7 M, λ ex = 5 nm, RT, ex. and em. b. p. = 5nm; Linear regression to the double reciprocal plot of fluorescence intensity change{/(i-i )} against added metal ion (/[Hg + ].5 ) for determination of complex stability constant for : (:Hg + ) complexation stoichiometry. SP9

10 .5 4 Abs Fluo. Int. (a. u., E5) Eq. of Hg + added Eq. of Hg + added Abs (A 557 ) Equation y = A + (A-A)/( + exp((x-x)/dx)) Adj. R-Square.9973 Value Standard Error A A x dx ln([hg + ]) (c).4 Fluo. Int. (arb. units, xe6) (d) Equation y = A + (A-A)/( + exp((x-x)/dx)).9975 Adj. R-Square Value Std. Err. E A E A.3758E E x E dx ln([hg + ]) Fig. S6: Plot of change in absorption and fluorescence intensity of as a function of added [Hg + ] ions in MeC-H (: v/v). Inset: change in colour of the solution of in presence of Hg + when irradiated with 45nm light. on-linear regression to the plot of (c) absorption and (d) fluorescence intensity against added metal ion (ln[hg + ]). Experimental conditions: emission [] = 5 7 M, λ ex = 5 nm, RT, ex. and em. b. p. = 5nm. SP

11 I 544 I 57 Fluo. Int. (arb units) 7 Eqv. Pb + Flou. Int. (arb.units, xe6) Flour. intensity a.u)..x -7 4.x -7 6.x -7 8.x x.x -6 3.x -6 4.x -6 5.x -6 6.x -6 7.x -6 8.x -6 [Pb + ] [Pb + ] I 544 I 57 Fluo. Int. (arb. units, xe6) Equation y = A + (A-A)/( + exp((x-x)/dx)) Adj. R-Square Value Standard Error C A C A 6.466E C x C dx ln([pb + ]) Fig. S7: Fluorescence spectra of as a function of equivalents of added Pb + ion and its corresponding spectral profile at 644nm and 67nm respectively. (Inset, a) Change in colour of the solution of in presence of Pb + when irradiated with 45 nm light. (c) on-linear regression to the plot of corresponding fluorescence intensity against added metal ion (ln[pb + ]). Experimental conditions: emission [] = 5 7 M, λ ex = 5 nm, RT, ex. and em. b. p. = 5nm. (c) SP

12 .3.5. Abs eq..5 [Cu + ] Wavelength (nm) Fig. S8: Absorption spectra of ( -4 M) in MeC-H (: v/v, PBS, ph 7.) as a function of equivalents of added Cu + ion Flou. Int. (arb. units, xe4) Equation y = a + b*x Adj. R-Square Value Standard Error a Intercept b Slope.7744E.84963E Fluo. Int. (arb. units, E5) Equation y = a + b*x Adj. R-Sq.9976 Value Standard Error I Intercept I Slope.787E 3.375E9.E+ 5.E-9.E-8.5E-8.E-8..x -8 4.x -8 6.x -8 8.x -8.x -7 [Hg + ] [Hg + ] Fig. S9: Linear regression of fluorescence spectral intensities of and with added Hg + ion for determination of sensitivity of Hg + detection. Fluo. Int. (arb. units) 3.x 6.5x 6.x 6.5x 6.x 6 5.x 5 MeC:H (9: v/v) MeC:H (8: v/v) MeC:H (7:3 v/v) MeC:H (6:4 v/v) MeC:H (5:5 v/v) MeC:H (4:6 v/v) MeC:H (3:7 v/v) MeC:H (:8 v/v) MeC:H (:9 v/v) Wavelength (nm) Fig. S: Fluorescence spectra of in presence of Hg(Cl 4 ) (5 eq.) in varied proportion of binary solvent composition (MeC:H, v/v, PBS). [] = M, λ ex = 5nm, RT, ex. and em. b p = 5nm. SP

13 85 85 T(%) T(%) Hg(II) [.5 eq.] + Hg(II) [. eq.] + Hg(II) [. eq.] + Hg(II) [complex] Hg(II) [.5 eq.] + Hg(II) [. eq.] + Hg(II) [. eq.] + Hg(II) [complex] Wavenumber (cm ) Wavenumber (cm ) (c) (d) T (%) T (%) 4 T (%) Hg(II) (.5 eq.) + Hg(II) ( eq.) + Hg(II) (complex) Wavenumber (cm - ) Wavenumber (cm - ) 4 + Hg(II) (.5 eq.) + Hg(II) ( eq.) + Hg(II) (complex) Wavenumber (cm - ) Fig. S: FT-IR spectra of (a and b) and (c and d) alone and in presence of Hg + ion in solution. SP3

14 k k (d) -.6 (Abs) k (c) ln (A 557 ) Time(sec.) Equation y = a + b*x Adj. R-Square.985 Value Standard Error Intercept E-4 Slope.6687E E (c) (d) ln (A 557 ) Equation y = a + b*x Adj. R-Square Value Standard Error Intercept E-5 Slope.3668E E-8 ln (A 557 ) Equation y = a + b*x Adj. R-Square Value Standard Error Intercept E-4 Slope E-5.463E-7 Fig. S: Change in absorption (A 557 ) in ( M) when added Hg + ion as a function of time (s) in MeC-H (: v/v, PBS, ph 7.). Figures, (c) and (d) represents linear regression plots of ln(a 557 ) against time for determination of rate constant (k) of first-order kinetics at the desired time interval. (Abs.) R Time(s) H ln (A) Equation y = a + b*x Adj. R-Square.946 Value Standard Error Intercept Slope E-4 Fig. S3: Change in absorption (A 557 ) in a five membered spiro-ring based probe R (4 M) when Hg + ion added (: stoichiometry) as a function of time (s) in MeC-H (: v/v, PBS, ph 7.). SP4

15 Abs.4.4 (R) H H ln (A) k Equation y = a + b*x Adj. R-Square.966 Value Standard Error Intercept Slope E (c) (d) ln (A) Equation y = a + b*x Adj. R-Square.7 Value Standard Error Intercept E-4 Slope E E k ln (A) Equation k 3 y = a + b*x Adj. R-sq Value Standard Error Intercept Slope E Fig. S4: Change in absorption (A 557 ) in another five membered spiro-ring based probe R ( M) when Hg + ion added in (: ligand-metal stoichiometry) as a function of time (sec) in MeC-H (: v/v, PBS, ph 7.). Figures, (c) and (d) represents corresponding linear regression plots of ln(a 557 ) against time(s) for determination of fractional rate constants (k) of first-order kinetics at the desired time interval ph 4. ln (A 557 ) Equation y = a + b*x Adj. R-Square Fig. S5: Change in absorption (A 557 ) in ( M) when Hg + ion (: stoichiometry) added as a function of time (s) in MeC-H (: v/v, PBS) at ph = 4.. SP5 Value Standard Error Intercept Slope 3.483E E-5

16 ln (A 557 ) ph4. ln (A 557 ) ph Equation y = a + b*x Adj. R-Square.985 Value Standard Error Intercept Slope E E Equation y = a + b*x Adj. R-Square Value Standard Error Intercept Slope 8.463E E (c).5 (d) ln (A 557 ) ph. A ph 4. ph 7. ph. -4. Equation y = a + b*x Adj. R-Square Value Standard Error Intercept Slope 5.937E E Fig. S6: Determination of rate of first order kinetics between ( µm) and Hg + ( eq.) in at 4., 7. and (c). ph from linear regression of the plot of log(a-a ) as a function of time (s). λ obs (abs) = 557nm. (d) Spectral profile of absorbance versus time (as given in Fig. 5b) for immediate reference. 7 Fluo. Int.(arb. units, xe6) Hg + +Ac - +Hg + +Hg + +Hg + +I - +Hg + +SC - +Hg + +Ac - +Hg + +En +Hg + +EDTA Fig. S7: Fluorescence spectra of saturated solution containing ( eq.) and Hg + (5 eq.) upon addition of anions and complexing reagents such as, SC -, EDTA, En, I -, acetate ion (eq.) in MeC-H (:v/v); [] = M, λ ex = 5nm, RT, ex. and em. b p = 5nm. SP6

17 Fluo. Int. (arb. units) + X +Hg + + X +Hg Fig. S8: Symbolic fluorescence spectra of (M) with mixed metal ions (X = Pb +, Zn +, Cu +, i +, Fe +, Cd +, Co +, Mn +, and Ag +, M, respectively) and upon addition of eq. of Hg + ion, λ ex = 5 nm..5. Abs..5 ph- ph- ph-3 ph-4 ph-5 ph-6 ph-7 Fluo. Int.(x E5, arb.units) ph ph ph 3 ph 4 ph 5 ph 6 ph Fig. S9: Variation in absorption fluorescence intensity of ( M) in MeC-H (:9 v/v, PBS buffer) under different ph conditions. Abs Hg(II) [A + Hg(II) /A ] / Fluo. Int.(x E6, arb. units) ph-6 ph-7 ph-8 ph-9 ph ph Fig. S3: Change in absorption(a 557 ) of ( M) in the presence of Hg + ion( eq.) in MeC-H (:9 v/v, PBS buffer) under different ph conditions. Variation of fluorescence intensity (I 58 ) of ( M) in presence of Hg + ion (5 eq.) in MeC-H (: v/v, PBS buffer) at different ph, λ ex = 5nm. SP7

18 Abs..3.. ph ph ph 3 ph 4 ph 5 ph 6 ph 7 Fluo. Int.(x 6, arb. units) ph ph ph 3 ph 4 ph 5 ph 6 ph Fig. S3: Variation in absorption fluorescence intensity of ( M) in MeC-H (:9 v/v, PBS buffer) under different ph conditions. Fig. S3: Colorimetric mapping of alone, +Hg + and (c) +Hg + in MeC-H (:9 v/v, PBS buffer) under different ph (ph = -8) conditions. [ or ] = M. SP8

19 Hg + H Hg + H Fig. S33: Proposed mechanism of Hg + coordination to and. Bio-imaging with for Hg + detection in E. Coli The biomass not treated with Hg + shows no fluorescence whereas treated biomass produced high fluorescence of rhodamine implying the significance of quantification of Hg + in biological system. The figure depicted below shows that Hg + could be possibly detected with even a low resolution fluorescence microscope. (c) (d) Fig. S34: Bright-field images of E. Coli alone and after incubation with. Their fluorescence image are on incubation with before (c) and after (d) addition of Hg + ion, λ ex 5nm. SP9