Electronic Supplementary Information

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1 Electronic Supplementary Material (ESI) for Organic & Biomolecular Chemistry. This journal is The Royal Society of Chemistry 7 Electronic Supplementary Information Modulation in the Acidity Constant of Acridine Dye with Cucurbiturils: Stimuli- Responsive pk a Tuning and Dye Relocation into Live Cells Raman Khurana, a,b Nilotpal Barooah, a Achikanath C. Bhasikuttan, a,b Jyotirmayee Mohanty a,b * a Radiation & Photochemistry Division, Bhabha Atomic Research Centre, Mumbai 4 85, India; b Homi Bhabha National Institute, Anushaktinagar, Mumbai, 4 94, India nm.6..8 pk a = ph /nm Figure S. Absorption spectra of acridine (.45-6 M) in water at different phs. ().6, () 4., (3) 4.5, (4) 5., (5) 5.6, (6) 6., (7) 7., (8) 8. and (9) 9.. Inset: variation in absorbance with ph at 354 nm.

2 .4 (A) 7 6 (B) I fl I fl [CB7] / M / nm / nm Figure S. (A) Absorption spectra of Ac (4.5-6 M) in aqueous solution at different CB7 concentrations at ph. CB7/M: (), () 5, (3), (4) 77, (5) 3, (6) 57 and (7) 6. (B) Fluorescence spectra of Ac (4.5-6 M) in aqueous solution at different CB7 concentrations at ph. CB7/mM: ()., ()., (3).4, (4)., (5).6, (6)., (7).46, (8).9 and (9) mM mm M / nm / nm Figure S3. CB7 Concentration dependent normalized absorption spectra of acridine dye at ph ~3.5. Method M: : host: guest binding model In the present systems, the binding constants (K) for the different forms of the dye with the CB7 host were estimated at suitable ph conditions by the fluorescence titration method assuming : complexation stoichiometry according to. S, which afforded satisfactory fitting results. -3 I f [ Dye] [ 7 : ] CB Dye I Dye ICB7: Dye (S) [ Dye] [ Dye] Where, I Dye and I CB : Dye 7 are the extrapolated fluorescence intensities of the uncomplexed and complexed form of the dye, respectively, Dye and CB7 are the respective total concentrations of dye (AcH +, Ac forms, as applicable) and CB7

3 host, and [Dye] is the concentration of uncomplexed dye in the solution. Exchange of the dye during its excited-state lifetime (<3 ns), i.e., the conversion of the uncomplexed dye to the complexed one or vice versa, can be excluded since the corresponding rate constants are very small for macrocyclic host molecules.,3 For fitting, the change in the fluorescence intensity ( I f ) was plotted against the total host concentration and the obtained titration curve was fitted according to the rearranged. S:, 3 [ Dye] I I I where, [Dye] is expressed as f CB7: Dye Dye [ Dye] (S) [ ] { K[ Dye] K[ SCX 6] ( K[ Dye] K [ CB7] ) 4K[ Dye] [ CB7] Dye } / K (S3) Method M: : host: guest binding model 4 CB + Dye K CB : Dye K CB : Dye + Dye CB : (Dye) (S4a) (S4b) K (ternary complex) = K K The fluorescence intensity of the system, I, is a function of the intensities of the free guest (I g ) and the : and : host-guest complexes (I and I g, respectively): I x I x I x I (S5) g g g g x g, x and x g are the mole fractions of the free guest and the : and : host-guest complexes. The mole fractions are further defined as follows: G G x g (S6) x HG G K H G K G G x (S7) K K G

4 x HG G H G G K K x g (S8) K G K K G Thus, the fluorescence intensity of the system is obtained throu substitution of. (S6-S8) into. (S5). G G K G K K G Ig K H I K K H G K G K K G Ig I (S9) The K value of CB8:(AcH + ) has been estimated from the binding isotherm (inset of Fig. 3B) by using. S9. Time (min) Time (min) (A). -. (B) µcal/sec µcal/sec kcal mol - of injectant -. kcal mol - of injectant Molar Ratio Molar Ratio Figure S4. (A) ITC isotherm for titration of AcH + with CB7 in water at 5 C. (B) ITC isotherm for titration of CB8 with AcH + in water at 5 C.

5 (A) (B) Counts Counts L L 5 5 Time / ns 5 5 Time / ns Figure S5. Decay traces of AcH + in solutions at ph ~3.5 in the absence () and presence () of 5 M CB7 (A) and 7 M CB8 (B). L represents excitation lamp profile. ex = 374 nm. CB8-CH CB8-CH CB8-CH H a (c) CB7-CH H c,d CB7-CH CB7-CH (b) Ha H e H a Hc H a H c H c,d H d N H d (a) H e He H H e ppm Figure S6. H NMR spectra (5 MHz) of M acridine dye in the absence (a) and in the presence (b) of mm CB7 and (c) 8 M CB8 in D O at pd 4.5. Inset: Pictorial representation of ACH +. Method M3: The ph-dependent absorbance changes at 355 nm (inset of Fig. S) were fitted according to the following relation (. S), 3 A AcH AAc A obs (S) phpka pka ph { } { } where A obs is the observed absorbance at any ph, and extrapolated absorbances of the AcH + A and A are the AcH and Ac forms, respectively. From this analysis, the pk a value of the dye in its ground state was found to be 5.4 ±., which matches well with the reported value. 3, 5 Ac

6 6 (A) 8 (B) I fl I fl / nm / nm Figure S7. Fluorescence spectra of CB7:AcH + complex (A) and CB8:AcH + complex (B) with increasing temperature. T/ C: (), () 3, (3) 4, (4) 5, (5) 6, (6) 7 and (8) 8. (a) (b) (c) Ac (Ac form) Ac-CB7 (AcH + form) Ac-CB7-AD (Ac form) Figure S8. Fluorescence microscopic images recorded from CHO cell lines at ph 7.4 after treating them with uncomplexed acridine dye ( M) (a); acridine dye (M)-CB7 (mm) (b); and acridine dye-cb7 (mm)-ad (M) (c). References. J. Mohanty, A. C. Bhasikuttan, W. M. Nau and H. Pal, J. Phys. Chem. B, 6,, M. Shaikh, J. Mohanty, P. K. Sin, W. M. Nau and H. Pal, Photochem. Photobiol. Sci., 8, 7, A. Jadhav, V. S. Kalyani, N. Barooah, D. D. Malkhede and J. Mohanty, ChemPhysChem, 5, 6, M. Sayed, F. Biedermann, V. D. Uzunova, K. I. Assaf, A. C. Bhasikuttan, H. Pal, W. M. Nau and J. Mohanty, Chem. Eur. J., 5,, M.-J. Ji, J.-G. Kim and U. S. Shin, Bull. Korean Chem. Soc.,, 33, 489.