J . Phys. Chem. 1989, 93, 6809-6813
or both of the other rotational moments. Consequently, the magnitude of the product of moments will be compensated and will not be changed by much. Furthermore, the rotational entropy is dependent on the natural logarithm of the product of rotational moments, and so even a large change in the product will have only a relatively minor effect on the rotational entropy. To determine the sensitivity of the calculations to variations in the M+-N internuclear distance, entropy values were calculated for plus and minus 0.2 A changes in the internuclear separation in the lead(+)-ammonia system. The rotational contribution changed by as much as 7% with resulting maximum changes in the vibrational contribution of only up to 0.7 cal/mol.K. The contribution of uncertainties in the internuclear separation distance to uncertainties in the calculated vibrational entropy changes are relatively small compared to the confidence level in the experimental determination of the overall entropy change. Using eq 3, we have calculated vibrational entropy changes of the first three stepwise additions of N H 3 onto alkali metal and Pb+ ions. The results are also shown in Tables 111-V. For any nonlinear molecule of N atoms, there will be 3 N - 6 vibrational degrees of freedom that contain the internal rotation. Addition of ammonia onto the bare atomic ion results in a net increase of
6809
three vibrational modes. Thereafter, further stepwise additions of ammonia each result in a net gain of six vibrational modes. Clearly, the vibrational entropy change must always be positive if the frequencies of the normal modes of vibration in the cluster ions and clustering ligand are not significantly changed upon ass~ciation.'~ Assuming the frequencies of the normal modes of vibration in the ammonia molecule are nearly the same both before and after binding to the cluster, then the magnitude of the vibrational entropy change for a given addition should be dependent only on the frequencies and number of the newly acquired vibrational modes. Therefore, the magnitude of the vibrational entropy change should be a small positive value. The positive values of the vibrational enthalpy changes calculated from eq 3 suggest that the total entropy changes determined in the experiments are quite reasonable. Acknowledgment. Financial support by the US.Department of Energy, Grants DE-AC02-82-ER60044 and DE-FGO288ER60668, is gratefully acknowledged. We thank B. J. Conklin for assistance in obtaining some of the data. Registry No. NH3, 7664-41-7; Pb+, 14701-27-0; Li+, 17341-24-1; Na', 17341-25-2; K+, 24203-36-9; Rb', 22537-38-8.
Formation of Clusters between Ionic Species and Sodium Dodecyl Sulfate below the Critical Micelle Concentration. Ethidium Ions and Divalent Metal Ions Stephen J. Atherton* and Christopher M. G. Dymond Center for Fast Kinetic Research, ENS 16N, University of Texas at Austin, Austin, Texas 78712 (Received: February 13. 1989; In Final Form: May 10, 1989)
Steady-state absorption and emission spectroscopies and time-resolved fluorescence spectroscopy have been used to study the formation of clusters between ionic species and sodium dodecyl sulfate (SDS) below the critical micelle concentration. Ethidium ions (E') are shown to form clusters with submicellar SDS, which at low SDS concentration may be multiply occupied by E+. This has allowed us to show that the ground state of E+ quenches the first excited singlet state of E+. The divalent metal ions Zn2+,CaZ+,and Mgz+ also form clusters with submicellar SDS, and it is shown that only one metal ion is required to induce the cluster. It is surmised that this is a general phenomenon for divalent metal ions.
Introduction Dissolution of many simple surfactants, e.g., sodium dodecyl sulfate (SDS), in water results in the formation of aggregates or micelles, if the surfactant concentration is above the critical micelle concentration (cmc). The structure of these micelles depends on factors such as the surfactant concentration, solution ionic strength, temperature, etc. For SDS in room-temperature water containing no added electrolyte, concentrations above the cmc (8.1 X M ) and below ca. 5 X M SDS result in the formation of roughly spherical micelles with an aggregation number of ca. 64.' At concentrations below the cmc, SDS is largely monomeric, and above the cmc, the concentration of monomers in equilibrium with micelles decreases as the SDS concentration increases.2 Ionic micelles have been used extensively as mediators of chemical reactions. In part, this is because their biphasic nature (hydrophobic/hydrophylic) and charged interface resembles biological membranes and macromolecules and also because of their ability to favorably mediate photoreactions, resulting in the storage of chemical energy. Studies concerning surfactant solutions below the cmc are scarce; however, there are a few reports of the ability of submicellar SDS to mediate reactions involving charged species.3 (1) Tanford, C. The Hydrophobic Effect; Wiley-Interscience: New York, 1980. (2) Hall, D. G.; Wyn-Jones, E. J. Mol. Liq. 1986, 32, 63.
0022-3654/89/2093-6809$01.50/0 , I
,
Holzwarth et al.3c studied the rate of complex formation between the metal ions NiZ+and MnZ+and pyridine-2-azo-4,4-dimethylaniline (PADA) in aqueous SDS solution and found considerable enhancement of the reaction rate at SDS concentrations below the cmc. These enhancements were attributed to the formation of premicellar aggregates of SDS and PADA to which the metal ions were electrostatically attracted. Although more recent work (described below) would suggest that it is the metal ions that are inducing the submicellar aggregates, Bruhn and Holzwarth report that the neutral molecule Fe(phen)2(CN)2induces aggregates with submicellar SDS at SDS concentrations above half the c ~ c and , ~there ~ is a report of submicellar ag(3) (a) Cho, J.-R.; Morawetz, H. J . Am. Chem. SOC.1972, 94, 375. (b) Lachish, U.; Ottolenghi, M.; Rabani, J. J . Am. Chem. SOC.1977, 99, 8062. (c) Holzwarth, J.; Knoche, W.; Robinson, B. H. Ber. Bunsen-Ges.Phys. Chem. 1978,82, 1001. (d) Bruhn, H.; Holzwarth, J. Ber. Bunsen-Ges. Phys. Chem. 1978, 82, 1006. (e) Rodgers, M. A. J.; Foyt, D. C.; Zimek, 2.A. Radiat. Res. 1978,75,296. (f) Baxendale, J. H.; Rodgers, M. A. J. Chem. Phys. Lett. 1980, 72, 424. (9) Atherton, S. J.; Baxendale, J. H.; Hoey, B. M. J. Chem. Soc., Faraday Trans. 1 1982, 78, 2167. (h) Baxendale, J. H.; Rodgers, M. A. J. J. Phys. Chem. 1982, 86, 4906. (i) David, C.; Szalai, E.; BaeyensVolant, D. Ber. Bunsen-Ges. Phys. Chem. 1982,86, 710. 0') Kusumoto, Y.; Shizuka, M.; Satake, I. Chem. Phys. Lett. 1986, 125, 64. (k) Kusumoto, Y.; Shizuka, M.; Satake, I. Chem. Lett. 1986, 529. (I) Kusumoto, Y.; Watanabe, J.; Kurawaki, J.; Satake, I. Chem. Lett. 1987, 1417. (m) Miyoshi, N.; Hara, K.; Yokoyama, I.; Tomita, G.; Fukuda, M. Photochem. Photobiol. 1988,47, 685. Q - 1989 American Chemical Societv
6810
The Journal of Physical Chemistry, Vol. 93, No. 18, 1989
gregation between SDS and ~yclodextrin,~J a neutral hydrophobic molecule like PADA. Baxendale and Rodgers,' Following the work of Lachish et showed that Ru(bpy),,+ in submicellar SDS solutions induced the formation of aggregates of Ru(bpy),,+ and SDS, where the number of Ru(bpy),*+ per cluster decreased from ca. 8 to ca. 3 as the SDS concentration increased from 3 X lo-, to 8 X lo-, M. The time-resolved luminescence of Ru(bpy):+ in these clusters was shown to be biphasic with the faster component due to an annihilation reaction occurring when more than one Ru(bpy),,+ excited state occupies the same cluster. Later work by Atherton et aL38 showed that Cu2+ also caused the formation of clusters in submicellar SDS and that these clusters were capable of incorporating the uncharged ruthenium complex Ru(bpy),(CN),, resulting in enhanced quenching of Ru(bpy),(CN), luminescence. Ru(bpy),(CN), alone did not form aggregates with submicellar SDS; thus, it is the Cu2+ ions that are nucleating the clusters. Interestingly, no enhanced quenching of Ru(bpy)z(CN)2 luminescence by methylviologen (MV2+)was observed in submicellar SDS even though clustering of MV2+ with submicellar SDS is known to Unlike clusters of Cu2+ with SDS, MVzf clusters with SDS presumably are unable to incorporate the Ru(bpy),(CN), complex. The present report concerns the formation of aggregates between positively charged ethidium ions (E') and SDS below the cmc, as studied by steady-state absorption and emission spectroscopies and by time-resolved emission spectroscopy. In addition, it is shown that doubly charged metal ions are also able to form submicellar aggregates with SDS. We present evidence that only one metal ion is required to form the aggregate and that this appears to be independent of the nature of the metal ion. Experimental Section Ethidium bromide (EB) was from Sigma, SDS was BDH specially pure grade, other reagents were the best available grade, and all were used as received. Water was purified by a Millipore filtration system. Stock solutions were freshly prepared on the day of use. Steady-state absorption and emission spectra were measured on a Hewlett-Packard Model 8450A UV-vis spectrophotometer and a Perkin-Elmer Model LS5 fluorescence spectrophotometer, respectively. Time-resolved emission measurements were made with one of two instruments. In one, the second harmonic (532 nm, 30 ps) pulse from a Quantel Model YG402 Nd:YAG laser was the excitation source and the emission was measured at right angles to the excitation with a Hamamatsu streak camera (time resolution, ca. 10 ps). The second laser system comprised a Spectra Physics Series 3000 Nd:YAG laser mode-locked at 82 MHz. The second harmonic (532 nm) of this laser pumped a Spectra Physics Model 375B dye laser, containing pyridine dye. The 720-nm output from the dye was cavity dumped at 800 kHz and doubled to 360 nm to serve as the excitation source. The emission was monitored via single-photon counting as has been described p r e v i ~ u s l y . ~Appropriate cutoff filters were placed before the streak camera or photomultiplier tube in order to discriminate against scattered light. In both the single-photon counting and streak camera experiments, substitution of the sample by a scattering solution resulted in zero response at the sensitivities required for our measurements. All time-resolved data were passed to an on-line PDPl1/70 minicomputer for analysis. The computer system and analysis software have been described previ~usly.~ Results Steady-State Measurements. Figure 1 shows the visible ground-state absorption spectra of 3 X M E+ as a function of SDS concentration. The photophysics of E+ are known to be dependent on the basicity of the environment;6 thus, there is a (4) Atherton, S . J.; Beaumont, P. C. J . Phys. Chem. 1986, 90, 2252. ( 5 ) Foyt, D. C. Comput. Chem. 1981, 5,49. (6) Olmstead, J.; Kearns, D. R. Biochemistry 1977, 16, 3647.
Atherton and Dymond
:Z5C
.
XIIELEbGTH
Figure 1. Ground-state absorption spectra of 3 X M E+ as a function of SDS concentration: -, 0 SDS; 7.5 X lo4 M SDS; --,3 X lo-) M SDS;----,5 x 10-3 M SDS; 5 x 10-2 M SDS. -a-,
...,
2000
04 560
600
640
680
720
/
Wavelength/nm
M E+ as a Figure 2. Normalized fluorescence spectra of 3 X function of SDS concentration: -, 0 SDS; .-,3 X M SDS; --,lo-* M SDS.
ca. 35-nm red shift in the ground-state visible absorption band M SDS. As when E+ is associated with SDS micelles at 5 X the SDS concentration decreases below the cmc to 5 X lo-, M SDS, there is little change in the absorption spectrum; however, at 3 X lo-, M SDS, there is a further red shift in the absorption spectrum superimposed on a base line that rises toward shorter wavelengths. A plot of absorbance vs the reciprocal of the fourth power of the wavelength gives a straight line for wavelengths outside the absorption spectrum of E+ (650-800 nm), showing that this effect is caused by scattered light. At 7.5 X lo4 M SDS, the absorption appears as a mixture of E+ in pure water and the species responsible for the red-shifted band at 3 X M SDS and there is an increased contribution due to scattered light. These solutions are stable over a period of days, and no precipitation is observed. The fluorescence spectra of E+ as a function of SDS concentration are shown in Figure 2, where the gain conditions have been adjusted in order to bring all the spectra to roughly the same intensity. The effects of SDS concentration on the fluorescence of E+ are minor compared with the effects on the absorption spectra. In water, the fluorescence peaks at 617 nm and there is a slight red shift to 621 nm at M SDS. At 3 X lo-' M, the fluorescence is a little further red-shifted to 624 nm and the spectrum is slightly broader. The relative fluorescence intensity has been measured for E+ as a function of SDS concentration by exciting the samples at 490 nm and integrating (cutting and weighing) the area under the fluorescence curve from 560-760 nm. The excitation wavelength was chosen to be 490 nm since
The Journal of Physical Chemistry, Vol. 93, No. 18, 1989 6811
Cluster Formation between Ionic Species and SDS 1.5 ,
I
TABLE I: Decay Parameters" for the Fluorescence of 3 E+ as a Function of SDS Concentration ISDS1/10-' M 0 50 30 10 8 7 6
5
0.0
1 0
20
10
30
40
4 3 2 1 50
-3 M
[SDS]/lO
Figure 3. Integrated fluorescence intensity vs SDS concentration for 3 x 10-5 M E+.
0
2
4
6
8
10
Time/ns Figure 4. Fluorescence decay curves and weighted residuals for 3 X M E+ in water (top), 3 X lo-) M SDS (middle), and 5 X lo-* M SDS (bottom). Solid line through the data is the best fit to single-exponential (0 and 5 X M SDS) and double-exponential (3 X M SDS)
decay. it most nearly approximates an isosbestic point in the absorption spectra. These data are shown in Figure 3. As the SDS concentration increases, the fluorescence intensity decreases rapidly, M, and then rises to a reaching a minimum at ca. 2.5 X plateau value at roughly the cmc where the fluorescence intensity is a factor of ca. 2.5 larger than that in water. The increase in fluorescence intensity that occurs when E+ is associated with SDS micelles (above the cmc of SDS) is expected on the basis of reduced basicity of the environment, and as shown in text below, this is mirrored by an increase in the fluorescence lifetime. The decrease of fluorescence intensity at SDS concentrations below the cmc will be due in part to light scattering of the sample; however, we will show that the major reason for this decrease is clustering of E+ with SDS. Time-Resolved Experiments. Figure 4 shows the fluorescence time profiles for 3 X M E+ in water, 3 X M SDS, and 5 X 10-2 M SDS. The fluorescence follows single-exponentialdecay kinetics in both water and 5 X M $DS with lifetimes of 1.6 f 0.1 and 4.7 0.1 ns, respectively. At 3 X M SDS,
*
Fp
0.47 0.62 0.64 0.78 0.82 0.79 0.63
7Jns
0.19 0.1 1 0.21 0.19 0.09 0.08
0.05
F. 1 1 1 1 1 0.53 0.38 0.36 0.22 0.18 0.21 0.37
X
M
7Jns 1.6 4.7 4.7 4.7 4.6 4.2 2.2 2.0 1.2 1.6 1.6 1.4
"Ffand F, are the fractions of fluorescence decaying via the faster and slower processes, respectively, and 7f and 7, are the lifetimes of the faster and slower components, respectively.
however, the fluorescence decay is clearly more complex and can be fit with biexponential kinetics to give lifetimes of 0.18 0.1 and 1.8 0.1 ns, respectively. The smaller of these two lifetimes is considerably smaller than our instrument response function, and therefore, further fluorescence measurements in this region of SDS concentration were made with the streak camera. The M SDS residuals shown in Figure 4 (lower trace) for 5 X show some nonrandom effects at short times; however, we were unable to fit the data as a sum of exponentials and assume that laser instabilities are responsible for these effects. Our lifetime for E+ in water compares with 1.8 ns measured earlier.' The fluorescence decay of 3 X M E+ was measured as a function of SDS concentration, and the decay parameters are presented in Table I, where Ff and F, are the fractions of the fluorescence intensity that decay via the faster and slower decay channels, respectively, and T~ and T , are the lifetimes of the faster and slower components, respectively. All data at SDS concentrations below 8 X M were obtained with the streak camera with a time window of 1.2 ns. We would therefore expect a relatively large error in the values of the lifetime of the longer component in these cases. As the SDS concentration decreases below the cmc, the fluorescence decay becomes biphasic with the faster component having a lifetime considerably shorter than that of E+ in water and the slower component having a lifetime shorter than that of E+ in SDS above the cmc. With a further decrease in SDS concentration, the lifetimes of both fluorescence components decrease and the lifetime of the slower component becomes similar to that of E+ in pure water. In addition, the fraction of fluorescence intensity decaying via the faster process increases to a maximum of ca. 80% at an SDS concentration between 2 and 3 X M. As the SDS concentration decreases below 2 X loT3M, the fraction of fluorescence decaying via the faster channel now decreases, until at 0 SDS the fluorescence is well described by a single exponential. These results mirror the change in the steady-state fluorescence intensity with SDS concentration (Figure 3). We interpret these results in terms of the quenching of the fluorescence of excited E+ by ground-state E+ caused by clustering of E+ with monomer SDS below the cmc. These clusters of E+ and SDS are multiply occupied by E+, resulting in the close proximity of ground-state E+ to excited E+, which allows the quenching to proceed. Annihilation of E+ excited states under intense laser irradiation is ruled out since qualitatively the same effects are seen under the much less intense irradiation required for the steady-state and single-photon-counting measurements and also reduction of the laser power by 1 order of magnitude does not affect the results. Our interpretation of Figure 3 and Table I is as follows. E+ forms clusters with SDS below the cmc, similar to the situation shown previously for R~(bpy),*+.~'At concentrations of SDS
*
( 7 ) Burns, V. W. F. Arch. Biochem. Biophys. 1969, 183, 420.
6812 The Journal of Physical Chemistry, Vol. 93, No. 18, 1989 ? 5-
0.0
Atherton and Dymond 12
1 0
1
2
3
4
I
0.0 !
I
0
3
2
1
[z~~+I/Io-~M
4
[M 2f]/10-4M
Figure 5. Integrated fluorescence intensity vs ZnZt concentration for 3 X M Et in 0,2 X M; 0,4 X lo-' M; W, 5 X M; 0 , 3 X M; and A, M SDS.
Figure 6. Integrated fluorescence intensity vs metal ion concentration for 3 X M Et in 3 X lo-' M SDS. Key: 0, Mgzt; 0 , Zn2+.
above ca. 7 X M, there is sufficient SDS to allow single occupancy of E+; however, as the SDS concentration is decreased, multiple occupancy of the clusters occurs and the fluorescence decay becomes complex due to fluorescence quenching of excited E+ by ground-state E+ occupying the same cluster. As the SDS concentration is decreased further, the average occupancy of the clusters rises, resulting in a decreased lifetime of the faster fluorescence decay component. Ultimately, there will be insufficient SDS to complex all E+, resulting in E+ free in solution and a component of fluorescence with the lifetime of E+ in water. As M, more and more the SDS concentration drops below 2 X E+ is uncomplexed with SDS and the portion of fluorescence decaying via quenching by ground-state E+ (faster channel) decreases. Addition of Divalent Metal Ions. It was shown previously that Cu2+ ions complex with submicellar SDS to form c l ~ s t e r s . ~ g However, no information was given concerning the nature of these clusters, for example, how many metal ions or how many SDS monomers were required to form a cluster. In an effort to gain more information concerning clusters of SDS with divalent metal ions, we have investigated the effects of divalent metal ions on the E+/SDS system. Zn2+ was chosen since it is close to Cu2+ in size but does not quench E+ fluorescence in water;* thus, only the effects of additional or competitive cluster formation should be observed. Figure 5 shows the effect on the fluorescence intensity of 3 X M E+ in solutions of (2-5) X lW3 M SDS caused by additions of Zn2+. In all cases, the addition of Zn2+ causes an increase in the fluorescence intensity of E+. The increase in fluorescence intensity for a given added Zn2+concentration is larger at higher SDS concentrations, and indeed, at 4 and 5 X 10" M SDS, the addition of as little as 4 X lo4 M Zn2+is sufficient to return the fluorescence intensity to the value observed in SDS solutions above the cmc. This effect of small additions of Zn2+ is also observed for the metal ions Mg2+ and Ca2+. Figure 6 shows the effect of additions of Zn2+ and Mg2+ to the fluorescence of 3 X M E+ in 3 X M SDS; within experimental error, the effect is the same. Essentially identical results were obtained with Ca2+ (data not shown). Although it is known that an increase in ionic strength of a surfactant solution has the effect of lowering the cmc, these small concentrations of metal ions are far below those required to appreciably alter the cmc of SDS. For example, 2 X M NaCl is required to lower the cmc of SDS to 4 X M.' Further, we have shown that lW3 M Na2S04has a negligible effect on the fluorescence intensity of 3 X M E+ in 3 X M SDS. Dodecyl sulfate solutions containing only divalent
TABLE II: Fraction of Et Fluorescence Decaying via the Slower M SDS,as Decay Channel (F,) for 3 and 6 X l e M Et in 3 X a Function of ZnZt Concentration
(8) Atherton, S. J.; Beaumont, P. C. Photobiochem. Photobiophys. 1984, 8, 103. (9) Williams, R. J.; Phillips, J. N.; Mysels, K. J. Trans. Faraday Soc. 1955, SI, 728.
0.17 0.20 0.30 0.40
0 2 2.5 3
4
0.41
4
5 6
0.52 0.58
5 6 7 7.5 8 9 10 12.5 15
0 1 2 3
0.09 0.14
0.13 0.15 0.22 0.30 0.31 0.39 0.41 0.41
0.38 0.49 0.53 0.59
counterions have cmc values of ca. M,'O and an electrostatic interaction is thought to result in these low values." In our experiments, dramatic effects are observed at concentrations of divalent ions well below this value; thus, we do not consider the formation of conventional micelles to be responsible. The fluorescence decay of E+ is similarly affected by addition of ZnZ+,in that the addition of Zn2+ appears to be equivalent to increasing the SDS concentration. With 3 X M SDS and in the absence of Zn2+, the fluorescence decay is biphasic with ca. 80% of the fluorescence decaying via the faster component. As the Zn2+concentration is raised, the fraction of fluorescence decaying via the faster channel decreases with a corresponding increase in that fraction decaying via the slower channel. The fractions of fluorescence decaying via the slower channel as a function of Zn2+ concentration for solutions of 3 and 6 X M E+ in 3 X M SDS are given in Table 11. For these measurements, both the faster and slower components of fluorescence were extrapolated to time zero, taken to be the middle of the exciting pulse. As the extent of the faster component decreases, the error on this extrapolation increases. For this reason, we confine our measurements to low ZnZ+concentrations.
Discussion It is clear from the above data that Et forms clusters with monomer SDS below the cmc. At low concentrations of SDS (less than ca. 6 X M), these clusters become multiply occupied by E+. It is this phenomenon that results in a decrease in the fluorescence intensity of E+, since an excited E+ in a multiply (10) Miyamoto, S. Bull. Chem. SOC.Jpn. 1960, 33, 375. (1 1) Gunnarsson, G.; Jonsson, B.; Wennerstrom, H . J . Phys. Chem. 1980, 84, 3144.
The Journal of Physical Chemistry, Vol. 93, No. 18, 1989 6813
Cluster Formation between Ionic Species and SDS occupied cluster will be close to a ground-state E+ that quenches the fluorescence. We are able to observe this process via the streak camera as a very fast fluorescence decay with a lifetime
