- Email: [email protected]
Dye aggregation in freezing aqueous solutions
Volume
119.
DYE
number 5
CHEMICAL
AGGREGATION
Rainer
IN FREEZING
PHYSICS LETTERS
AQUEOUS
13 September 1985
SOLUTIONS
SCHIRRA
Received 10 Apnl 1985; in final form 15 June 1985
The fluorescence wawr- The spectral usual cluster model hfeumes have been
and absorplion spectra of the dyesruff [hloflnwne T are strongly nffecwd by [he freaing of iIs solution m changes are though1 LObe caused by dlmerimuon of dyesluff ions uhich c;ln bc imerprcwd in ~crms of Ihc of wawr exerting hydrophobic interacllons The absorption and luminmcnce specua and thr fluoracrncc dekrminsd
1. Introduction The spectra of some aqueous dye solutions may be related to changes in the structure of water. The state of aggregation of ionic dyestuffs, in particular, ought to be sensitive to phase transitions in the solution if the van der Waals potentials exceed the Coulombrc repulsions. The abrupt change in the electronic spectra just as an aqueous solution of thioflavine T (3,6dimethyl2<4dimethylaminophenyl)-benzotbiazohum chloride, Cl. 49005) freezes, indicates the formation of dye aggregates synchronously coupled to a phase transition from lower to higher order_ Similarly ordered states of water molecules can also be sensed by thioflavine T in the interfacial water of clay minerals; thisphenomenon plays an essential role in the understanding of soil mechanics_ Perturbation of the water structure by the dye molecules IS thought to be negligible since the mean distance between these IS of the order of 100 run_
2. Apparatus The search for suitable dyestuffs was performed in an apparatus consisting of a N2-pumped dye laser, a commercial HejNz evaporating cryostat, a detecting monocluomator with photomultiplier for the obser-
vation of the fluorescence and a boxcar integrator for measuring fluorescence lifetimes. The solutions were placed in an open copper trough, insrde the cryostat_ Excitation and observation were performed at an angle of 45O with respect to the normal of the free surface. Absorption spectra were recorded with a UV/VIS spectrometer, using thin-walled plastic cells for frozen solutions.
3. Results Fig. 1 shows the fluorescence spectrum of 2.5 X lo-5 M aqueous thioflavine T. At room temperature (300 K), a single peak is observed without detectable shoulders. The fluorescence lifetime near the maximum of the transition is 1 .O f 0.2 ns This spectral distribution does not change on cooling untrl the freezing point is reached. The spectrum then changes over a range of 1 K during freezing to that labelled 273 K. (The range depends somewhat on the kinetics of freezing_) The spectra obtained on further cooling to 112 K and to 77 K are also illustrated. Like the fluorescence spectrum, the absorption spectrum (fig. 2) exhibits a slrnilar abrupt change on freezing; that is, on cooling the absorption only begms to change when the solution starts to freeze. The single peak at 412 run in the liquid phase is transformed on freezing to a maximum at 364 run, with severalshoulders at longer wavelengths. 463
Volume 119, number 5
CHEMICAL PHYSICS LETTERS
600
Wavelength
13 September
[nm]
Temperature
Fig L Tcmpenture dependence of the luminescence of thioffavine T in water; c = Z 5 X 10s M
spectrum
1985
I K]
Fig. 3. Temperature dependence of the lumine~nce in450.550 and 610 run; c = 2.5 x lo5 M. Note the abrupt appearanoeof the 550 nm emtiion at the freering point. tensity at
Fig 3 illustrates the temperature dependence of the three luminescence maxima of a 2.5 X 1O-5 M solution The 450 MI peak decays steeply on cooling from 300 K to the freezing point. On cooling below 273 K the 450 nm peak increases to a maximum at 160 K while the 550 MI peak reaches its maximum at 130 K. Expenments at differing concentrations revealed that the ratio of the 550 to the 450 nm fluorescence mtensity is greater the higher the concentration_ This is also true for the ratio of the intensity of the emission at 610 run to that at 450 run. The fluorescence lifetimes depend on temperature: at 460 run and 300 K: 1 .O f 0.2 ns, at 64 Kr 1 6 ns. The lifetime of the emission at 550 run rises to 5.0 ns at 70 K. The emission at 610 MI comes from a long-living state with a half-life of the order of some tenths of a second. a0
-xQ
-
Wavelength
Inml
Fig 2. Abrupt change of the absorptionof aqueous thiollavine T (2.5 X IO5 M) (1) 273 < T < 293 K; (2) 269 < T < 273 K. 464
4. Discussion
The luminescence lifetimes demonstrate that the 450 and the 550 MI emission is fluorescence and
Volume
119, number 5
CHEMICAL
PHYSICS
the 610 nm emission is phosphorescence_ The peak emitted by dilute solutrons at 450 nm IS certainly monomer fluorescence; however, as is confirmed by the concentration dependence of the 550 nm/450 run intensity ratio, the 550 fluorescence is caused by aggregated dye molecules_ A small dimerization constant of about 3 X 1 O2 M-l can be determined from the slight concentration dependence ofthe extmction coefficient (c> 5 X 1 0m4 M) at room temperature. The formation of higher oligomers can be neglected because the association steps are thought to be successive for energetic reasons. Because of the abruptness of aggregation on freezing the formation of oligomers is forbidden for entropy consideratrons. Otherwise dye molecules in dilute solutions would have ta associate concertedly from a pre-oriented state which cannot be spectroscoprcally detected. In contrast to the exceedingly narrow bands of oligomeric and polymeric cyanines [ 1,2] the luminescence of thioflavme T consists of broad bands untypical of oligomerization and polymerization. Since the long-life transition (6i0 nm) is coupled with the existence of dimer fiuorcsccnce, this trrpler emission is likely to be the phosphorescence of a dlmerized species. The absorptron peak at 364 run seems to be produced by the dimerized dye molecules; the 394 run peak is evrdently the former 412 M-I maximum shifted by superposition (fig.2). The long-wavelength shoulder at 460 nm probably comes from a partially forbidden transition of dimerlzed dye molecules_ The increases to the maximum fluorescence intensity (fig. 3) on cooling from 273 to 160 K, and to 130 K, respectively, can be explained in terms of diminished thermal deactivation_ The unusual decrease which follows may originate from deactivation via the competing tnplet state. The dyestuff salt thioflavine T is, in contrast to symmetrical dyestuffs [3-6 J, a cationic molecule which exhibits such a slight tendency to associate that molecules can only aggregate during the freezing of an aqueous solution. In contrast to the well investigated cyanine dyestuffs (1.71, it can be assumed that thioflavine T forms sandwich dimers with an anti-conformation because of the increased positive charge density of the thiazolc ring. The attraction energy which has to exceed the
LETl-FXS
13 September
1985
Coulombic repulsion may be a n-rr interaction of the typical rm6 van der Waals type. This assumption is contirmed by the absence of acid H-atoms and the large displacement of spectra, indicating strong perturbation of the n-system. The hydrophobic groups at the periphery of the extended cations prevent adequate hydration. In contrast to their smaller anions, the dye cations are only partially or not integrated into the framework of the flickering clusters [S-10]. They preferentially occupy the spaces between the interfaces of the water clusters; there they create a high local concentration, which is one of the preconditions for aggregatron. Both the size of the clusters and their local concentration are enlarged with diminishing temperature until they reach their maxima at the freezing point. The association of the dye molecules is coupled to the cooperative phase transrtton [ 111 of the water clusters to the crystal lattice and is similarly abrupt in ice the interactions between the water molecules themselves are so strong that the already low degree of hydration of the dye molecules virtually disappears In consequence they attain a maximum degree of dimerization which does not depend upon the temperature T < 273 K but only upon the initial concentration. This opinion isconfirmed experimentally by another result: The fluorescence of frozen Nal-fluorescein solutions is almost completely quenched because of the formation of associates However, the addition of small amounts of methanol to the aqueous solution (molar ratro methanol I H20 = 1 : 100) efficiency prevents the quenching of rhe fluorescence. The alcohol molecules are thought to disturb the water clusters by participating in the H-bond system to the cost ofwater. The appearance of dimer fluorescence and dimer phosphorescence of thioflavine T is obviously an indicator of the phase transition of water, i.e. of a transitron from low to high order for water molecules, even when the pure water structure IS slightly disturbed.
Acknowledgement The author wishes to thank Professor H. Moesta for his helpful discussions and the Fonds der Chemischen Industrie for tinancral support.
465
Volume
119. number 5
CHEMICAL
PHYSICS
References [ 1] G. Scheibe, Angew. Chem- 42 (1939) 631. [Z] Th. Fdrster, Fluoreszenz organivher Verbidungen (Vnndcnhoeck & Ruprecht, mttingen. 1980). [3] AX. Ghosh and P. Mukeiee. J. Am. Chem_ Sot. 92 (1970) 6403. (41 W. Spencer and J.R. Sutter. J. Phys. Chem. 83 (1979) 1573. [S] S L_ Fomili. G- Sgroi and V. Izzo, J. Chem. Sot_ Faraday Trans. I79 (1983) 1085
LETTERS
[6] T G. Dewey. P.S. Wikon and D.H. Turner, J. Am. Chem. sot. 100 (1978) 4550. 171 G. Scheibe. 2. Elektiochem. 52 (1948) 28X [S] H.S. Frank and W-Y. Wen. Disc. Faraday SOC_ 24 (1957) 133. 19) G. N6methy and HA. Schemga,I. Chem. Phyn 36 (1962) 3382. !?O] W. Luck, Z. Elektrochem. 66 (1962) 766. [ll] J_i. Frenkel, Kinetwhe Theorie der FlUssigkeiten (Deutscher Verlag der Wissenrhaften, Berlm. 1957).
D.B. Chesnut and C.K_ Foley, Some simple basis sets for accurate 13C chemical shift calculations, Chem. Phys. Letters 118 (1985) 316. In the caption of table 4, the expression for the shift anisotropy should read: PO = u,, - IT,_,or Au = 033 - ;(a22 + all).
466
13 September 1985
119.
DYE
number 5
CHEMICAL
AGGREGATION
Rainer
IN FREEZING
PHYSICS LETTERS
AQUEOUS
13 September 1985
SOLUTIONS
SCHIRRA
Received 10 Apnl 1985; in final form 15 June 1985
The fluorescence wawr- The spectral usual cluster model hfeumes have been
and absorplion spectra of the dyesruff [hloflnwne T are strongly nffecwd by [he freaing of iIs solution m changes are though1 LObe caused by dlmerimuon of dyesluff ions uhich c;ln bc imerprcwd in ~crms of Ihc of wawr exerting hydrophobic interacllons The absorption and luminmcnce specua and thr fluoracrncc dekrminsd
1. Introduction The spectra of some aqueous dye solutions may be related to changes in the structure of water. The state of aggregation of ionic dyestuffs, in particular, ought to be sensitive to phase transitions in the solution if the van der Waals potentials exceed the Coulombrc repulsions. The abrupt change in the electronic spectra just as an aqueous solution of thioflavine T (3,6dimethyl2<4dimethylaminophenyl)-benzotbiazohum chloride, Cl. 49005) freezes, indicates the formation of dye aggregates synchronously coupled to a phase transition from lower to higher order_ Similarly ordered states of water molecules can also be sensed by thioflavine T in the interfacial water of clay minerals; thisphenomenon plays an essential role in the understanding of soil mechanics_ Perturbation of the water structure by the dye molecules IS thought to be negligible since the mean distance between these IS of the order of 100 run_
2. Apparatus The search for suitable dyestuffs was performed in an apparatus consisting of a N2-pumped dye laser, a commercial HejNz evaporating cryostat, a detecting monocluomator with photomultiplier for the obser-
vation of the fluorescence and a boxcar integrator for measuring fluorescence lifetimes. The solutions were placed in an open copper trough, insrde the cryostat_ Excitation and observation were performed at an angle of 45O with respect to the normal of the free surface. Absorption spectra were recorded with a UV/VIS spectrometer, using thin-walled plastic cells for frozen solutions.
3. Results Fig. 1 shows the fluorescence spectrum of 2.5 X lo-5 M aqueous thioflavine T. At room temperature (300 K), a single peak is observed without detectable shoulders. The fluorescence lifetime near the maximum of the transition is 1 .O f 0.2 ns This spectral distribution does not change on cooling untrl the freezing point is reached. The spectrum then changes over a range of 1 K during freezing to that labelled 273 K. (The range depends somewhat on the kinetics of freezing_) The spectra obtained on further cooling to 112 K and to 77 K are also illustrated. Like the fluorescence spectrum, the absorption spectrum (fig. 2) exhibits a slrnilar abrupt change on freezing; that is, on cooling the absorption only begms to change when the solution starts to freeze. The single peak at 412 run in the liquid phase is transformed on freezing to a maximum at 364 run, with severalshoulders at longer wavelengths. 463
Volume 119, number 5
CHEMICAL PHYSICS LETTERS
600
Wavelength
13 September
[nm]
Temperature
Fig L Tcmpenture dependence of the luminescence of thioffavine T in water; c = Z 5 X 10s M
spectrum
1985
I K]
Fig. 3. Temperature dependence of the lumine~nce in450.550 and 610 run; c = 2.5 x lo5 M. Note the abrupt appearanoeof the 550 nm emtiion at the freering point. tensity at
Fig 3 illustrates the temperature dependence of the three luminescence maxima of a 2.5 X 1O-5 M solution The 450 MI peak decays steeply on cooling from 300 K to the freezing point. On cooling below 273 K the 450 nm peak increases to a maximum at 160 K while the 550 MI peak reaches its maximum at 130 K. Expenments at differing concentrations revealed that the ratio of the 550 to the 450 nm fluorescence mtensity is greater the higher the concentration_ This is also true for the ratio of the intensity of the emission at 610 run to that at 450 run. The fluorescence lifetimes depend on temperature: at 460 run and 300 K: 1 .O f 0.2 ns, at 64 Kr 1 6 ns. The lifetime of the emission at 550 run rises to 5.0 ns at 70 K. The emission at 610 MI comes from a long-living state with a half-life of the order of some tenths of a second. a0
-xQ
-
Wavelength
Inml
Fig 2. Abrupt change of the absorptionof aqueous thiollavine T (2.5 X IO5 M) (1) 273 < T < 293 K; (2) 269 < T < 273 K. 464
4. Discussion
The luminescence lifetimes demonstrate that the 450 and the 550 MI emission is fluorescence and
Volume
119, number 5
CHEMICAL
PHYSICS
the 610 nm emission is phosphorescence_ The peak emitted by dilute solutrons at 450 nm IS certainly monomer fluorescence; however, as is confirmed by the concentration dependence of the 550 nm/450 run intensity ratio, the 550 fluorescence is caused by aggregated dye molecules_ A small dimerization constant of about 3 X 1 O2 M-l can be determined from the slight concentration dependence ofthe extmction coefficient (c> 5 X 1 0m4 M) at room temperature. The formation of higher oligomers can be neglected because the association steps are thought to be successive for energetic reasons. Because of the abruptness of aggregation on freezing the formation of oligomers is forbidden for entropy consideratrons. Otherwise dye molecules in dilute solutions would have ta associate concertedly from a pre-oriented state which cannot be spectroscoprcally detected. In contrast to the exceedingly narrow bands of oligomeric and polymeric cyanines [ 1,2] the luminescence of thioflavme T consists of broad bands untypical of oligomerization and polymerization. Since the long-life transition (6i0 nm) is coupled with the existence of dimer fiuorcsccnce, this trrpler emission is likely to be the phosphorescence of a dlmerized species. The absorptron peak at 364 run seems to be produced by the dimerized dye molecules; the 394 run peak is evrdently the former 412 M-I maximum shifted by superposition (fig.2). The long-wavelength shoulder at 460 nm probably comes from a partially forbidden transition of dimerlzed dye molecules_ The increases to the maximum fluorescence intensity (fig. 3) on cooling from 273 to 160 K, and to 130 K, respectively, can be explained in terms of diminished thermal deactivation_ The unusual decrease which follows may originate from deactivation via the competing tnplet state. The dyestuff salt thioflavine T is, in contrast to symmetrical dyestuffs [3-6 J, a cationic molecule which exhibits such a slight tendency to associate that molecules can only aggregate during the freezing of an aqueous solution. In contrast to the well investigated cyanine dyestuffs (1.71, it can be assumed that thioflavine T forms sandwich dimers with an anti-conformation because of the increased positive charge density of the thiazolc ring. The attraction energy which has to exceed the
LETl-FXS
13 September
1985
Coulombic repulsion may be a n-rr interaction of the typical rm6 van der Waals type. This assumption is contirmed by the absence of acid H-atoms and the large displacement of spectra, indicating strong perturbation of the n-system. The hydrophobic groups at the periphery of the extended cations prevent adequate hydration. In contrast to their smaller anions, the dye cations are only partially or not integrated into the framework of the flickering clusters [S-10]. They preferentially occupy the spaces between the interfaces of the water clusters; there they create a high local concentration, which is one of the preconditions for aggregatron. Both the size of the clusters and their local concentration are enlarged with diminishing temperature until they reach their maxima at the freezing point. The association of the dye molecules is coupled to the cooperative phase transrtton [ 111 of the water clusters to the crystal lattice and is similarly abrupt in ice the interactions between the water molecules themselves are so strong that the already low degree of hydration of the dye molecules virtually disappears In consequence they attain a maximum degree of dimerization which does not depend upon the temperature T < 273 K but only upon the initial concentration. This opinion isconfirmed experimentally by another result: The fluorescence of frozen Nal-fluorescein solutions is almost completely quenched because of the formation of associates However, the addition of small amounts of methanol to the aqueous solution (molar ratro methanol I H20 = 1 : 100) efficiency prevents the quenching of rhe fluorescence. The alcohol molecules are thought to disturb the water clusters by participating in the H-bond system to the cost ofwater. The appearance of dimer fluorescence and dimer phosphorescence of thioflavine T is obviously an indicator of the phase transition of water, i.e. of a transitron from low to high order for water molecules, even when the pure water structure IS slightly disturbed.
Acknowledgement The author wishes to thank Professor H. Moesta for his helpful discussions and the Fonds der Chemischen Industrie for tinancral support.
465
Volume
119. number 5
CHEMICAL
PHYSICS
References [ 1] G. Scheibe, Angew. Chem- 42 (1939) 631. [Z] Th. Fdrster, Fluoreszenz organivher Verbidungen (Vnndcnhoeck & Ruprecht, mttingen. 1980). [3] AX. Ghosh and P. Mukeiee. J. Am. Chem_ Sot. 92 (1970) 6403. (41 W. Spencer and J.R. Sutter. J. Phys. Chem. 83 (1979) 1573. [S] S L_ Fomili. G- Sgroi and V. Izzo, J. Chem. Sot_ Faraday Trans. I79 (1983) 1085
LETTERS
[6] T G. Dewey. P.S. Wikon and D.H. Turner, J. Am. Chem. sot. 100 (1978) 4550. 171 G. Scheibe. 2. Elektiochem. 52 (1948) 28X [S] H.S. Frank and W-Y. Wen. Disc. Faraday SOC_ 24 (1957) 133. 19) G. N6methy and HA. Schemga,I. Chem. Phyn 36 (1962) 3382. !?O] W. Luck, Z. Elektrochem. 66 (1962) 766. [ll] J_i. Frenkel, Kinetwhe Theorie der FlUssigkeiten (Deutscher Verlag der Wissenrhaften, Berlm. 1957).
D.B. Chesnut and C.K_ Foley, Some simple basis sets for accurate 13C chemical shift calculations, Chem. Phys. Letters 118 (1985) 316. In the caption of table 4, the expression for the shift anisotropy should read: PO = u,, - IT,_,or Au = 033 - ;(a22 + all).
466
13 September 1985