journal of

MOLECULAR

LIQUIDS ELSEVIER

Journal of Molecular Liquids 71 (1997) 49-59

EXCITED

STATE PROTON

~-CYCLODEXTRIN

TRANSFER

ENCAPSULATED

IN THE PRESENCE

OF

CARBAZOLE

OF UREA

Santi Kundu and Nitin Chattopadhyay § Department o f Chemistry, Jadavpur University, Calcutta - 700 032, India. Received 11 June 1996; accepted 7 November 1996 ABSTRACT

Excited state proton transfer (ESPT) reaction of carbazole (CAZL) in aqueous [3-cyclodextrin (13CD), in the presence of urea, has been studied by both steady state as well as time resolved methods. The unstructured anionic emission band from the excited fluorophore and the rate constants for the individual protropic processes reveal that urea expels the probe from the cyclodextrin microenvironment to the bulk aqueous phase. The same has also been established from the quenching study of the same fluorophore by iodide ion. © 1997 Elsevier Science B.V.

1.

INTRODUCTION

Urea causes denaturation of proteins and inhibits micellisation. This interesting observation provokes the photophysicists/photochemists to study the effect of urea on the photophysical and/or photochemical processes of different probes in different chosen environments [ 1-11 ]. Two propositions are put forward to explain the action of urea. The first one says that urea is a water structure breaker and according to the second one urea displaces some water molecules around a hydrophobic group and, thus, modifies the solvation of the latter. Surface

Present address : Laboratoryfor MolecularDynamicsand Spectroscopy,Departmentof Chemistry, Katholieke UniversiteitLeuven, 200F Celestijnenlaan,B-3001 Heverlee,Belgium. 0167-7322/9"//$17.00 © 1997 Elsevier Science B.V. All fights reserved Pll S0167-7322(96)00972-5

50 tension measurements [4] and computer simulations [12,13] go against the first mechanism revealing that there is hardly any difference between the water molecules in the vicinity of the urea molecules from those in the bulk. Rather, the latter studies indicate that urea displaces some water molecules from the neighbourhood of the hydrophobic group. Taking p-toluidinonaphthalenesulphonate (TNS) as probe in micellar systems, Bhattacharyya et al. [8,9] have shown that the second mechanism is more liable to operate. Following the excited state proton transfer (ESPT) of carbazole in micellar media, we are also in favour of the proposition that urea displaces water from the periphery of the hydrophobic zone [11]. Since the micelles are not very well defined and also not rigid in structure; cyclodextrins (CD), possessing well defined and stable structure, are now preferred systems to establish the mechanism of action of urea [9,14,15]. The kinetic aspects of the ESPT of a fluorescent probe, carbazole (CAZL), in aqueous as well as in aqueous cyclodextrin media have already been studied by us [16-20]. In the present communication we deal with this ESPT reaction in the presence of urea. Steady state and time resolved study strongly indicate that urea expels the fluorophore from the CD environment to the bulk aqueous phase. This has also been substantiated from the quenching study of the probe by the iodide ion.

2.

EXPERIMENTAL SECTION

2.1.

Materials

Carbazole (Aldrich) was purified by vacuum sublimation followed by recrystallisation from 90% ethanol. The purity of the compound was checked from the spectroscopic methods as well as TLC where only a single spot was noticed. 13-cyclodextrin (Aldrich) was recrystallised from water prior to experiment. Analytical grade NaOH and KI (both from BDH) were used as received. Triply distilled water was used for the experiments.

2.2.

Measurements

Shimadzu MPS 2000 absorption spectrophotometer and spex fluorolog spectrofluorimeter were used for the absorption and emission studies respectively. For the time resolved study, the time correlated single photon counting (TCSPC) technique was adopted [21]. The experimental setup is described elsewhere [22].

51 3.

RESULTS AND DISCUSSION

The variation of the steady state fluorescence spectra of a solution of carbazole (CAZL) in aqueous [3-CD and on subsequent addition of urea at a definite pH (=12) is shown in figure 1.

t, 3 2

t "r-

320

I 350 Wovelenq

I ~00 th (nm)

1 ~50

i 500

Fig. 1 : Steady state fluorescence spectra of carbazole in different aqueous environments at pH 12 : (1) in water, (2) in 5raM [3CD, (3) in 5raM [3CD plus 1M urea and (4) in 5raM [3CD plus 2M urea solution (ke, c,=,o, = 297 nm). CAZL concentration is same in all the cases. The figure dictates that although the alkali concentration is constant, the anion yield (emission maximum at = 415 nm) is enhanced at the cost of the neutral band (~ 360 nm). as CAZL is embedded within the CD cavity. The addition of urea to the resulting solution, however, reverses the intensities of the bands. The first part is just a reproduction of our earlier studies [18,19]. Upon complexation of CAZL with CDs ([3 and y), the deprotonation rate constant for the ESPT process is found to be enhanced by = 90% in the CD environments -- while the back protonation remains practically unaffected. The result was rationalised by the cooperative proton transfer mechanism according to which the imino proton of CAZL is dissociated by the alkali (OH') through the participation of the -OH groups attached at the periphery of the cyclodextnn moieties. From tig. 1 it is clear that wlalle ~L'D encapsulation favours the deprotonation process of CAZL, urea disfavours the same for the encapsulated system indicating that urea induces a remarkable change in the microenvironment around the probe.

52

The anionic emission band of CAZL, in the presence of I3CD, is found to be structured. Similar observation was also found in aqueous cetyltrimethylammoniumbromide (CTAB) and triton-X 100 (TX) micellar solutions [20]. This is, as suggested before, because of the restriction in molecular motion within the small space provided by the CD. It is interesting to note that the vibronic resolution is lost and the band resembles to that produced in pure aqueous medium as and when 2M urea is added to the system. This observation indicates that urea expels the fluorophore from the CD periphery to the bulk aqueous phase. The time resolved analyses, allows us to determine the individual rate constants for the excited state prototropic process in different environments. Figure 2 depicts the change in the fluorescence spectra of a I3CD embedded CAZL solution ([13CD]=5mM) in the presence of urea ([urea]=2M) with the variation of NaOH concentration. The excitation spectra were unchanged throughout, confirming that all the changes were due to the excited state process.

!

.2_

320

I 350

I L+O0 Wovetength

1 t~50 ( nm )

i 500

=

Fig.2 : Steady state fluorescence spectra of CAZL in the presence of5mM 13CD plus 2M urea as a function of NaOH concentration. NaOH concentrations in spectra 1 to 7 are 0.0, 0.005, 0.01, 0.015, 0.02, 0.03 and 0,04 M respectively.

A kinetic scheme for the excited state proton transfer (ESPT) reaction of CAZL, as described before [16], is as follows :

53 *

k

AH

ht~

+

OH-

11

AH

"

kt +kd

+

[OH

] =k'

i

OH-

,

'

A

_,

+

H20

1

kt +k d

k k

io

L

A

+

H20

20

where AH and A- represent CAZL and the corresponding anion, k~ and k_, are the rate constants for the forward and the backward reactions in the photoexcited state, k~0 and k20 are the corresponding rate constants in the ground state, kf and kf' are the rate constants for the fluorescence of CAZL and the anion respectively, k d and k d" are the respective non-radiative decay constants. The differential equations that describe the above kinetic scheme are : - d[AH°]/dt = (k~'+kf+kd) [AH'] - k2[A* ] .......................... (1) - d[A"]/dt = (k2+ke'+kd") [A"] - k,' [AH °] ....................... (2) where k I" = kt [OH]. Using the boundary conditions [AH'] = [AH']o and [A"] = 0 at t = 0 and taking X = kf+kd+k I ", Y = kf'+kd'+k 2 and Li, ~-2 = "~1"l, "t2"1 1

= ! {(X + Y) +_[(Y - X) 2 + 4k~k 2 ]2 } ..................... (3) 2 (where "t~ and z2 are the excited state lifetimes of the neutral and the anion of CAZL respectively) we arrive at the following equations : %t+%2 = k d + k f + k l [ O H ' ] + Y

.....................

%1%2= (kf + k~) Y + (kf" ÷ kd') kl [OH']

.............

(4) (5)

54

and ~., + ~-2 = (k,+~ + [ 1 - (~+kd)/(~.'+k~') ] Y + (k(+k~')" X,~

..... (6)

Now, one can estimate the individual rate constants from the lifetime values of the neutral and the anionic forms of CAZL at different concentrations of added alkali. The gradual addition of alkali led to the shortening of the neutral lifetime (at 360 nm) but the anion decay (at 415 nm) remained almost constant, as consistent with the earlier studies [16,17]. Figure 3

o

,

W

".o ~.o "~ *. "-~ ",.. ". °~°o

'

'..:, ".,'.....",,.. ".~.

"-°.

--..

7"-"

L

':

0

"[' •

U l~l

',,..,, ,i

,.

.°

.

• ,

~

i

.i

o*

•

oQ.

,o

•

,

o ,

°o

,

.g

.

.

.

.

0

.

o,

.

.

•

.

.

.60

.

.

.

.

.

.

.

.

i .,

t

,m

°

~ol

. . . .

.tSO ....

j.~

~.0

" ' ° . ~

-,..

,

:;

o°

o . °

o e .

. . . . . .

i*

, , , o . . . o ,

........................

30

40

50

Tiee/lO "9 sec

''f=

d :

i

Fig 3 : Fitted fluorescence decay curves of CAZL in presence of 5ram B e D plus 2M urea monitored at 360 rim. [OH] in 1 to 7 are 0.0, 0.005, 0.01, 0.015, 0.02, 0.03 and 0.04 M respectively. The corresponding T values obtained are 10.1, 8.25, 6.65, 5.20, 4.30, 3.31 and 2.76 nanoseconds respectively. For all the fits )~2 were within the range 1.0 to 1.2. The dotted profile corresponds to the lamp. The bottom part depicts the nature of the lit (standard deviation) for a sample decay (curve 4 of the upper picture).

55 shows the variation of neutral lifetime with the variation of alkali concentration. Following the kinetic scheme described above, we determined the rate constants for the excited state prototropic reaction. The values are reported in Table 1. Table 1

Rate constants for the ESPT reaction of carbazole.

Rate constants

Water

5mM [3CD

5mM I3CD + 2M urea

(ref.16)

(ref. 19)

(present work)

kl (MIs "l)

1.0 x 1010

1.9 (+ 0.1) x 10 l°

8.1 (± 0.2) x 109

k 2 (s "l)

8.5 x 106

8.0 (+ 0.2) x 106

8.2 (+ 0.2) x 106

The values reflect that while the forward rate constant for the ESPT process of CAZL is enhanced by 90% in the presence of 5mM 13CD; it falls within the range of that obtained in pure aqueous phase on addition of 2M urea to the solution. The rate constant for the backward process, however, remains practically unaffected in all the modifications of the environment. The rate analyses categorically reflects that in the presence of 2M urea, the ESPT reaction is taking place in an environment which is very similar to the bulk water. The slight lowering in the deprotonation rate constant is, presumably, because of the lowering in the bulk polarity due to the addition of reasonable amount of urea (2M). Thus, both the unresolved fluorescence band of the anion as well as the rate constant values establish that the carbazole molecule which, in association with the [3CD, was in a microenvironment different from water (likely, at the periphery of the CD [18,19]); in the presence of urea, goes into an environment which is nothing but the bulk aqueous phase modified by urea. That is, urea expels the probe molecule from the CD cavity to the bulk water. The expulsion of CAZL molecule from the CD environment has also been established by the fluorescence quenching study of the fluorophore by potassium iodide. The quenching rate constant was estimated following the steady state fluorescence quenching of the probe with added quencher and the lifetime of the former in the absence of the quencher. For the StemVolmer plots fluorescence ratios were calculated from the area under the fluorescence band after verifying that they gave identical values to the quantum yield ratios. The Stern-Volmer plot yielded straight lines for the three solutions, viz., in pure aqueous medium, in the

56

presence of 5mM 13CD and in the presence of 5mM 13CD plus 2M urea. (fig. 4). The values of the quenching rate constants (M"s "~)are given in Table 2.

,3.5 ,3.0 2.5 ~2.0 1.5 1.0 0.5 0.00

0.04 0.08 Conc. of K1 (M')

0.12

Fig 4 : Stem -Volmer plots of the fluorescence quenching of CAZL by KI in (a) water. (b) 5raM 13CD, (c) 5raM 13CDplus 2M urea and (d) 2M urea. Io and I denote the fluorescence yields of CAZL in the absence and presence of quencher (KI).

The value of the quenching rate constant in the presence of 13CD and urea together is much higher than that for the probe in 13CD solution only. This reflects that in the presence of added urea the fluorophore is no more within the CD cavity. The former rate constant value

57 Table 2

Rate constants for quenching of CAZL by KI.

Medium

Ksv

"to (ns)

Value of rate constant (M%-')

(a) Water

20.0

10.0

2.00 0:0.2) x 109

(b) 5mM ~CD

10.2

10.5

0.97 (i0.1) x 109

(c) 5raM IBCD + 2M urea

14.5

10.1

1.43 (+0.2) x 100

(d) 2M urea

13.6

9.9

1.37 (+0.2) x 100

is, however, well below the value obtained in pure aqueous environment. An independent quenching study of CAZL in aqueous urea solution ([urea]=2M) also yielded a linear SternVolmer plot (curve d of fig 4) giving a value of the rate constant as 1.37 x 109 Mqs -t. The agreement between this value and that obtained in 13CD-urea solution, unhesitatingly, establishes that the fluorophore molecule (CAZL), originally embedded within the 13CD cavity, is released to the bulk aqueous phase in the presence of urea.

4.

CONCLUSION

Both the excited state proton transfer and the fluorescence quenching studies establish that the fluorophore (CAZL) is expelled from the CD environment to the bulk aqueous phase.

5.

ACKNOWLEDGEMENTS

Thanks are due to Dr. S. C. Bera for his kind interest in the present work. Financial support from C.S.I.R., Govt. of India, is gratefully acknowledged.

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