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Solvation Dynamics of DCM in a Polypeptide-Surfactant Aggregate: Gelatin-Sodium Dodecyl Sulfate Arnab Halder, Pratik Sen, Anupam Das Burman, and Kankan Bhattacharyya* Physical Chemistry Department, Indian Association for the Cultivation of Science, Jadavpur, Kolkata 700 032, India Received September 4, 2003. In Final Form: November 12, 2003 Solvation dynamics of 4-(dicyanomethylidene)-2-[p-(dimethylamino)styryl]-6-methyl-4H-pyran (DCM) is studied in a polypeptide-surfactant aggregate consisting of gelatin and sodium dodecyl sulfate (SDS) in potassium dihydrogen phosphate (KP) buffer. The average solvation time 〈τS〉 in gelatin-SDS aggregate at 45 °C is found to be 1780 ps, which is about 13 times slower than that in 15 mM SDS in KP buffer at the same temperature. The fluorescence anisotropy decay in gelatin-SDS aggregate is also different from that in SDS micelles in KP buffer. DCM displays negligible emission in the presence of gelatin in aqueous solution. Thus the solvation dynamics in the presence of gelatin and SDS is exclusively due to the probe (DCM) molecules at the gelatin-micelle interface. The slow solvation dynamics is ascribed to the restrictions imposed on the water molecules trapped between the polypeptide chain and micellar aggregates. The critical association concentration (cac) of SDS for gelatin is determined to be 0.5 ( 0.1 mM.
1. Introduction Dynamics of water molecules inside many biological and organized assemblies plays an important role in many natural processes.1 In bulk water, solvation dynamics of water occurs on a subpicosecond time scale.2,3 However, in the vicinity of many biological macromolecules, e.g., protein in native4 and denatured state,5 DNA,6 hydrophilic polymer,7 polymer-surfactant aggregate,8 reverse micelle,9-11 and micelles,12 the water molecules exhibit an ultraslow component of solvation dynamics on the 1001000 ps time scale. Such a slow component of solvation dynamics in organized assemblies was also detected in * Corresponding author: e-mail
[email protected]; fax (91)-33-2473-2805. (1) (a) Nandi, N.; Bhattacharyya, K.; Bagchi, B. Chem. Rev. 2000, 100, 2013. (b) Bhattacharyya, K. Acc. Chem. Res. 2003, 36, 95. (c) Pal, S. K.; Peon, J.; Bagchi, B.; Zewail, A. H. J. Phys. Chem. B 2002, 106, 12376. (d) Levinger, N. E. Curr. Opin. Colloid Interface Sci. 2000, 5, 118. (2) (a) Jarzeba, W.; Walker, G. C.; Johnson, A. E.; Kahlow, M. A.; Barbara, P. F. J. Phys. Chem. 1988, 92, 7039. (b) Jimenez, R.; Fleming, G. R.; Kumar, P. V.; Maroncelli, M. Nature 1994, 369, 471. (c) Nandi, N.; Roy, S.; Bagchi, B. J. Chem. Phys. 1995, 102, 1390. (3) (a) Horng, M.-L.; Gardecki, J. A.; Maroncelli, M. J. Phys. Chem. A 1997, 101, 1030. (b) Maroncelli, M.; Fleming, G. R. J. Chem. Phys. 1987, 86, 6221. (c) Maroncelli, M. J. Mol. Liq. 1993, 57, 1. (4) (a) Jordanides, X. J.; Lang, M. J.; Song, X.; Fleming, G. R. J. Phys. Chem. B 1999, 103, 7995. (b) Pal, S. K.; Peon, J.; Zewail, A. H. Proc. Natl. Acad. Sci. U.S.A. 2002, 99, 1763. (c) Mandal, D.; Sen, S.; Sukul, D.; Bhattacharyya, K.; Mandal, A. K.; Banerjee, R.; Roy, S. J. Phys. Chem. B 2002, 106, 10741. (d) Mukherjee, S.; Sen, P.; Halder, A.; Sen, S.; Dutta, P.; Bhattacharyya, K. Chem. Phys. Lett. 2003, 379, 471. (5) (a) Pal, S. K.; Peon, J.; Zewail, A. H. Proc. Natl. Acad. Sci. U.S.A. 2002, 99, 15297. (b) Dutta, P.; Sen, P.; Halder, A.; Mukherjee, S.; Sen, S.; Bhattacharyya, K. Chem. Phys. Lett. 2003, 377, 229. (6) (a) Brauns, E. B.; Madaras, M. L.; Coleman, R. S.; Murphy, C. J.; Berg, M. A. Phys. Rev. Lett. 2002, 88, 158101-1. (b) Brauns, E. B.; Madaras, M. L.; Coleman, R. S.; Murphy, C. J.; Berg, M. A. J. Am. Chem. Soc. 1999, 121, 11644. (7) Frauchiger, L.; Shirota, H.; Uhrich, K. E.; Castner, E. W., Jr. J. Phys. Chem. B 2002, 106, 7463. (8) (a) Sen, S.; Sukul, D.; Dutta, P.; Bhattacharyya, K. J. Phys. Chem. B 2002, 106, 3763. (b) Dutta, P.; Sen, S.; Mukherjee, S.; Bhattacharyya, K. Chem. Phys. Lett. 2002, 359, 15. (c) Dutta, P.; Sukul, D.; Sen, S.; Bhattacharyya, K. Phys. Chem. Chem. Phys. 2003, 5, 4875. (9) (a) Lundgren, J. S.; Heitz, M. P.; Bright, F. V. Anal. Chem. 1995, 67, 3775. (b) Riter, R. E.; Willard, D. M.; Levinger; N. E. J. Phys. Chem. B 1998, 102, 2705. (c) Corbeil, E. M.; Levinger, N. E. Langmuir 2003, 19, 7264.
recent computer simulations13-15 and has been predicted by an analytical theoretical model.16 The protein-surfactant aggregates4d,5b,17 are the subject of many recent studies because of their biological implications, e.g., interaction of a protein with cell membrane surfactants. The interactions between the macromolecules (polymers or proteins) and the surfactants have been studied by various techniques such as fluorescence correlation spectroscopy,18,19 light scattering,22 small-angle neutron scattering,23 surface tension,24 NMR,25 fluorescence,8,17,21,26,27c and a theoretical model.20 It is interesting to study how (10) (a) Dutta, P.; Sen, P.; Mukherjee, S.; Halder, A.; Bhattacharyya, K. J. Phys. Chem. B 2003, 107, 10815. (b) Pal, S. K.; Mandal, D.; Sukul, D.; Bhattacharyya, K. Chem. Phys. Lett. 1999, 312, 178. (c) Sen, S.; Dutta, P.; Sukul, D.; Bhattacharyya, K. J. Phys. Chem. A 2002, 106, 6017. (d) Bhattacharyya, K.; Hara, K.; Kometani, N.; Yozu, Y.; Kajimoto, O. Chem. Phys. Lett. 2002, 361, 136. (11) (a) Riter, R. E.; Undiks, E. P.; Kimmel, J. R.; Pant, D. D.; Levinger, N. E. J. Phys. Chem. B 1998, 102, 7931. (b) Shirota, H.; Horie, K. J. Phys. Chem. B 1999, 103, 1437. (c) Hazra, P.; Chakrabarty, D.; Sarkar, N. Chem. Phys. Lett. 2003, 371, 553. (12) (a) Pal, S. K.; Sukul, D., Mandal, D.; Sen, S.; Bhattacharyya, K. Chem. Phys. Lett. 2000, 327, 91. (b) Hara, K.; Kuwabara, H.; Kajimoto, O. J. Phys. Chem. A 2001, 105, 7174. (c) Mandal, D.; Sen, S.; Bhattacharyya, K.; Tahara, T. Chem. Phys. Lett. 2002, 359, 77. (13) (a) Balasubramanian, S.; Pal, S.; Bagchi, B. Phys. Rev. Lett. 2002, 89, 115505-1. (b) Pal, S.; Balasubramanian, S.; Bagchi, B. J. Phys. Chem. B 2003, 107, 5194. (c) Bruce, C. D.; Senapati, S.; Berkowitz, M. L.; Perera, L.; Forbes, M. D. E. J. Phys. Chem. B 2002, 106, 10902. (14) (a) Faeder, J.; Albert, M. V.; Ladanyi, B. M. Langmuir 2003, 19, 2514. (b) Senapati, S.; Berkowitz, M. L. J. Chem. Phys. 2003, 118, 1937. (c) Senapati, S.; Chandra, A. J. Phys. Chem. A 2001, 105, 5106. (15) (a) Panatano, D. A.; Laria, D. J. Phys. Chem. B 2003, 107, 2971. (b) Thompson, W. H. J. Chem. Phys. 2002, 117, 6618. (c) Michael, D.; Benjamin, I. J. Chem. Phys. 2001, 114, 2817. (16) Nandi, N.; Bagchi, B. J. Phys. Chem. B 1997, 101, 10954. (17) Deo, N.; Jockusch, S.; Turro, N. J.; Somasundaran, P. Langmuir 2003, 19, 5083. (18) Narenberg, R.; Kliger, J.; Horn, D. Angew. Chem., Int. Ed. 1999, 38 (8), 1626. (19) Qiu, Q.; Somasundaran, P.; Pethica, B. A. Langmuir 2002, 18, 3482. (20) Gilanyi, T. J. Phys. Chem. B 1999, 103, 2085. (21) (a) Fang, Y. J.; Winnik, F. M.; Clarke, R. J. Biophys. Chem. 2003, 104, 449. (b) Winnik, F. M.; Winnik, M. A.; Tazuke, S. J. Phys. Chem. 1987, 91, 594. (c) Anghel, D. F.; Toca-Herrera, J. L.; Winnik, F. M.; Rettig, W.; Klitzing, R. V. Langmuir 2002, 18, 5600. (d) Lissi E. A.; Abuin, E. J. Colloid Interface Sci. 1985, 105, 1. (22) (a) Herning, T.; Djabourov, M.; Leblond, J.; Takerkart, G. Polymer 1991, 32, 3211. (b) Xia, T.; Dubin, P. L.; Kim, Y. J. Phys. Chem. 1992, 96, 6805.
10.1021/la035647m CCC: $27.50 © 2004 American Chemical Society Published on Web 12/16/2003
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the water molecules are affected when they are trapped between a protein (or a polymer) chain and micelles (e.g., sodium dodecyl sulfate, SDS). In a protein (polymer)surfactant aggregate, the protein (polymer) chain wraps around the micelles and displaces some water molecules from the Stern layer of the micelle.8,17-21,25 This lowers the polarity of the Stern layer of the micelle. Very recently, it was shown that the solvation dynamics in a proteinsurfactant (lysozyme-SDS) aggregate5b and a polymersurfactant (PVP-SDS and PEG-SDS) aggregate8 is markedly slower than that in the corresponding protein or the polymer or in the micelle alone. In the present work, we have studied the solvation dynamics of a probe (DCM) in an aggregate consisting of a polypeptide (gelatin) and a surfactant (SDS). Gelatin, a polypeptide obtained from denatured collagen, is used extensively in food, pharmaceutical, and photographic industries.22-26 It has a nonuniform distribution of at least 18 amino acids, of which the principal constituents are glycine, proline, alanine, hydroxyproline, glutamic acid, aspartic acid, and arginine. Gelatin is a polypeptide that lacks secondary and tertiary structure above its gelation temperature (37 °C). Gelatin, having a range of both hydrophobic and hydrophilic side chains, is soluble in water around neutral pH. An aqueous solution of gelatin remains as a turbid gel below 37 °C. Above 37 °C, the gelatin solution becomes a clear liquid and the polymer assumes a random coil configuration. The anionic surfactant SDS binds to the positively charged sites of gelatin because of electrostatic attraction and to the hydrophobic parts of the chain by hydrophobic binding.22-26 In a gelatin-SDS aggregate, above a particular SDS concentration known as the critical association concentration (cac), the SDS molecules bind to the gelatin chain in the form of micelles. The cac is almost an order of magnitude lower than the critical micelle concentration (cmc) of SDS (8 mM in the absence of added salt). The cac is found to be independent of gelatin concentration.26 For the SDS-gelatin system, Griffiths et al.24a reported a cac of 0.4 mM, while Whitesides and Miller26 reported a cac of 0.9 mM with ANS as a fluorescent probe. According to a light scattering study,22a diffusion of a dilute solution of gelatin is bimodal with a fast and a slow component corresponding to hydrodynamic radii of 210 and 750 Å, respectively. The latter is ascribed to formation of clusters of gelatin chains.22a When gelatin adsorbs SDS, the slow component disappears, presumably because of disruption of the clusters by electrostatic repulsion between the anionic SDS micelles.22a The hydrodynamic radius of gelatin-SDS aggregate is about 260 Å.22a According to a SANS study,23 the hydrodynamic radius of SDS micelles adsorbed to gelatin is about 24 Å, which is slightly larger than that of a free SDS micelle (20 Å). In a solution containing gelatin and SDS, when the polymer backbone is fully saturated and the concentration of free SDS exceeds the cmc value, SDS micelles are formed. The total (23) Cosgrove, T.; White, S. J.; Zarbakhsh, A.; Langmuir 1995, 11, 744. (24) (a) Griffiths, P. C.; Roe, J. A.; Jenkins, R. L.; Reeve, J.; Cheung, A. Y. F.; Hall, D. G.; Pitt, A. R.; Howe, A. M. Langmuir 2000, 16, 9983. (b) Arai, H.; Murata M.; Shinoda, K. J. Colloid Interface Sci. 1971, 37, 223. (25) Miller, D. D.; Lenhart, W.; Antalek, B. J.; Williams, A. J.; Hewitt, J. M. Langmuir 1994, 10, 68. (26) Whitesides, T. H.; Miller, D. D. Langmuir 1994, 10, 2899. (27) (a) Wittouck, N. W.; Negri, R. M.; De Schryver, F. C. J. Am. Chem. Soc. 1994, 116, 10601. (b) Quitevis, E. L.; Marcus, A. H.; Fayer, M. D. J. Phys. Chem. 1993, 97, 5762. (c) Sen, S.; Sukul, D.; Dutta, P.; Bhattacharyya, K. J. Phys. Chem. B 2001, 105, 7495. (d) Krishna, M. G. M.; Das, R.; Periasamy, N.; Nityananda, R. J. Chem. Phys. 2000, 112, 8502.
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concentration of SDS above which SDS micelles are formed obviously depends on gelatin concentration.26 In this work, we have compared solvation dynamics and fluorescence anisotropy decay in a gelatin-SDS aggregate with that in SDS micelles. 2. Experimental Section The dye DCM (laser grade, Exciton), SDS (Aldrich) and gelatin (Sigma, extracted from porcine skin, isoelectric point 4.9), were used as received. All measurements were done at 45 °C and at a pH ) 6.6 in 34 mM potassium dihydrogen phosphate (KP) buffer. The temperature (45 °C) is maintained via a circulator bath (Neslab, Endocal). The steady-state absorption and emission spectra were recorded on a Shimadzu UV-2401 spectrophotometer and a Perkin-Elmer 44B spectrofluorometer, respectively. For lifetime measurements, the samples were excited at 405 nm with a picosecond diode (IBH Nanoled-07). The emission was collected at a magic-angle polarization using a Hamamatsu MCP photomultiplier (2809U). The time-correlated single photon counting (TCSPC) setup consists of an Ortec 935 Quad CFD and a Tennelec TC 863 TAC. The data are collected with a PCA3 card (Oxford) as a multichannel analyzer. The typical full width at halfmaximum (fwhm) of the system response is about 80 ps. For anisotropy measurements, a polarizer was placed before the sample. The analyzer was rotated by 90° at regular intervals and the parallel (I|) and the perpendicular (I⊥) components of the fluorescence decay were collected for equal times, alternatively. Then, r(t) was calculated from
r(t) )
I|(t) - GI⊥ (t) I|(t) + 2GI⊥(t)
(1)
The G value of the setup was determined by use of a probe whose rotational relaxation time is very fast, e.g., Nile red in methanol.
3. Results 3.1. Steady-State Spectra. DCM is insoluble in water. Upon addition of gelatin, DCM becomes slightly soluble in water and the maximum concentration of DCM in 2 wt % gelatin in KP buffer is about 3 µM. In a 2 wt % gelatin solution in KP buffer at 45 °C, DCM exhibits an absorption maximum at 435 nm and displays negligible emission. It may be noted that the emission quantum yield of DCM in n-heptane is very small.10b Thus it seems that DCM resides in a hydrophobic and hydrocarbon environment inside gelatin. On addition of SDS to a 2 wt % gelatin in KP buffer at 45 °C, the absorbance, and hence concentration, of DCM remains constant up to 0.5 mM SDS. At an SDS concentration above ∼0.5 mM, the absorbance of DCM gradually increases. The increase in absorbance indicates solubilization of DCM molecules because of the formation of gelatin-SDS aggregate. Thus the cac of gelatin-SDS system is about 0.5 mM. The absorbance of DCM in a solution containing 2 wt % gelatin reaches its maximum value at a SDS concentration of 8 mM. At this concentration the absorption maximum of DCM is found to be at 480 nm, i.e., red-shifted by 45 nm from that in 2 wt % gelatin. This indicates that at this concentration the gelatin chain is saturated by the adsorption of the SDS micelles. In the presence of 8 mM SDS and 2 wt % gelatin in 34 mM KP buffer, the maximum concentration of DCM is 9 µM. On addition of SDS to a solution containing 2 wt % gelatin in KP buffer, emission intensity of DCM continues to be extremely weak up to about 0.5 mM SDS. At a SDS concentration above 0.5 mM, the emission intensity of DCM gradually increases and the emission maximum is
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Figure 2. Fluorescence decays of DCM at 45 °C in 34 mM KP buffer (pH 6.6) containing 2 wt % gelatin and 15 mM SDS at (i) 520 nm, (ii) 580 nm, and (iii) 700 nm. Figure 1. Variation of intensity of the emission spectra of DCM with increasing concentration SDS in 34 mM KP buffer (pH 6.6) at 45 °C in the presence (b) and absence (O) of 2 wt % gelatin. The initial part of the variation of emission intensity of DCM with SDS concentration in the presence of 2 wt % gelatin is shown in the inset.
found to be at 615 nm. The emission intensity of DCM in a gelatin solution reaches its maximum value at an SDS concentration of 10 mM and remains so up to 15 mM SDS. Further addition of SDS results in a decrease in emission intensity of DCM. This indicates disruption of the gelatinSDS aggregate and formation of nonadsorbed, free SDS micelles. The plot of emission intensity of DCM against SDS concentration in 2 wt % gelatin (Figure 1) shows a rapid increase above 0.5 ( 0.1 mM. The sharp increase in the emission intensity around 0.5 ( 0.1 mM SDS indicates formation of gelatin-SDS aggregates. From the steady-state emission measurement we conclude that the cac of SDS-gelatin system is 0.5 ( 0.1 mM. The cac obtained in this method is very close to the reported cac (0.4 mM) for the gelatin-SDS system.24a At room temperature, SDS is found to be sparingly soluble in an aqueous solution containing 34 mM KP buffer. However, SDS dissolves in aqueous KP buffer at 45 °C. From the sharp break in the emission intensity of DCM, the cmc of SDS in KP buffer at 45 °C is found to be ∼3 mM (Figure 1). This is lower than the cmc of SDS in the absence of any buffer at 20 °C (8 mM). 3.2. Time-Resolved Studies: Solvation Dynamics. In this section, we discuss our results on solvation dynamics of DCM in an aqueous solution containing 2 wt % gelatin and 15 mM SDS in 34 mM KP buffer (pH ) 6.6) at 45 °C. We will compare the results with the solvation dynamics of DCM in 15 mM SDS in a KP buffer at the same temperature. Since DCM bound to the gelatin exhibits negligible fluorescence, the solvation dynamics of DCM could not be studied in a gelatin environment. The emission decays of DCM in 2 wt % gelatin and 15 mM SDS in KP buffer at 45 °C exhibit a growth at the red end and a decay at the blue end. This is typical of solvation dynamics. In this case, at the blue end (520 nm) the decay is fitted to a biexponential with two components of 220 ps (85%) and 1500 ps (15%) (Figure 2). However, at the red end (e.g., at 700 nm), DCM exhibits a distinct rise of 130 ps followed by a decay of time constant 1900 ps (Figure 2). Following the procedure given by Maroncelli and Fleming,3b the time-resolved emission spectra (TRES) were constructed from the parameters of best fit to the fluorescence decays and the steady-state emission spectrum. The solvation dynamics is described by the decay
Figure 3. Time-resolved emission spectra of DCM at 45 °C in 34 mM KP buffer (pH 6.6) containing 2 wt % gelatin and 15 mM SDS at 10 ps (9), 250 ps (O), 1000 ps (2), and 10 000 ps (3).
of the solvent correlation function C(t), defined as
C(t) )
ν(t) - ν(∞) ν(0) - ν(∞)
(2)
where ν(0), ν(t), and ν(∞) are the peak frequencies at times 0, t, and ∞, respectively. TRES of DCM in 2 wt % gelatin and 15 mM SDS in KP buffer at 45 °C is shown in Figure 3. The decay of C(t) (Figure 4) is found to be biexponential with one component of 400 ps (75%) and another 5900 ps (25%), with an average solvation time 〈τs〉 ) 1780 ps (Table 1). The total Stokes shift is observed to be 500 ( 50 cm-1. The solvation dynamics of DCM in 15 mM SDS in 34 mM KP buffer is found to be very different from that in the presence of 2 wt % gelatin and 15 mM SDS. In a solution of 15 mM SDS in KP buffer at 45 °C, emission decays of DCM display marked wavelength dependence. For example, at 520 nm (the blue end) the fluorescence decay is biexponential with two decay components of 180 ps (94%) and 800 ps (6%), while at 700 nm (the red end) the decay of time constant 950 ps is preceded by a distinct rise with a time constant of 80 ps. The decay of C(t) for DCM, in KP buffered solution (pH ) 6.6) of 15 mM SDS, is found to be single-exponential with a time constant of 140 ps (Figure 4). This is much faster than the solvation dynamics in a gelatin-SDS aggregate. The total Stokes shift of DCM in SDS in KP buffer at 45 °C is found to be 300 ( 30 cm-1. 3.3. Fluorescence Anisotropy Decay. The fluorescence anisotropy decay of DCM in an aqueous solution of 2 wt % gelatin and 15 mM SDS in KP buffer exhibits a residual anisotropy of 0.1. The decay is fitted to a hindered
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Halder et al. Table 1. Decay Parameters of C(t) of DCMa
a
system
∆νb (cm-1)
a1
τ1b (ps)
a2
τ2b (ps)
〈τs〉b,c (ps)
2 wt % gelatin + 15 mM SDS 15 mM SDS
500 300
0.75 1.00
400 140
0.25
5900
1780 140
In 2 wt % gelatin solution with 15 mM SDS or in 15 mM SDS only in 34 mM KP buffer of pH 6.6 at 45 °C.
b
(10%. c 〈τs〉 ) a1τ1 + a2τ2.
Table 2. Parameters of Anisotropy Decay of DCMa system
r0
r
afastb
τfastb (ps)
aslowb
τslowb (ps)
τD (ps)
Dt × 1010 (m2 s-1)
2 wt % gelatin + 15 mM SDS 15 mM SDS
0.24 0.18
0.11 0.05
0.05 0.15
110 50
0.95 0.85
1270 630
1400 680
6.8 9.8
a
At 560 nm, in 2 wt % gelatin solution with 15 mM SDS or in 15 mM SDS only in 34 mM KP buffer of pH 6.6 at 45 °C. b (5%.
anisotropy (0.05). For SDS micelles in KP buffer the anisotropy decay of DCM is described by two components of 50 ps (15%) and 630 ps (85%) (Table 2). This is faster compared to the anisotropy decay in the gelatin-SDS aggregate. 4. Discussion
Figure 4. Decay of response function, C(t), of DCM at 45 °C in 34 mM KP buffer (pH 6.6) containing 2 wt % gelatin and 15 mM SDS (9) and in 34 mM KP buffer (pH 6.6) containing 15 mM SDS (O). The points denote the actual values of C(t) and the solid line denotes the best fit to a exponential decay. The initial parts of the decays of C(t) are shown in the inset.
Figure 5. Fitted curve for fluorescence anisotropy decay of DCM at 45 °C in 34 mM KP buffer of pH 6.6 containing 2 wt % gelatin and 15 mM SDS (s) and in 15 mM SDS in 34 mM KP buffer of pH 6.6 (---).
rotor as
r(t) ) r(∞) + {r(0) - r(∞)}[afast exp(-t/τfast) + aslow exp(-t/τslow)] (3) The time constants for the decay of r(t) are found to be 110 ps (5%) and 1270 ps (95%) (Table 2). Figure 5 shows the fluorescence anisotropy decay of DCM at 45 °C in 34 mM KP buffer of pH 6.6 containing 15 mM SDS with 2 wt % gelatin and 15 mM SDS alone. The fluorescence anisotropy decay in 15 mM SDS in KP buffer is found to be much faster with a very small residual
This work shows that the solvation dynamics and fluorescence anisotropy decay of DCM in the presence of 2 wt % gelatin and 15 mM SDS in KP buffer are very different than those in 15 mM SDS micelles in KP buffer. DCM is almost nonfluorescent in the presence of gelatin. Thus the solvation dynamics and anisotropy decay in the presence of gelatin and SDS corresponds to the DCM molecules at the interface of the gelatin chains and SDS micelles in the gelatin-SDS aggregate. The most important finding of this study is the significant slowing down of the dynamics of the water molecules inside the gelatin-SDS aggregate compared to that in SDS micelles. In the gelatin-SDS aggregate, the solvation dynamics at 45 °C is found to be biexponential with a major component (75%) of 400 ps and a slower one of 5900 ps (25%). The solvation dynamics in SDS micelles at the same temperature and same buffer is found to be single-exponential with a component of 140 ps. The slow solvation inside the polypeptide (gelatin)-SDS aggregate is consistent with the previous reports on slow solvation in protein-SDS aggregate5b and in polymer-SDS aggregate.8 It seems that in the gelatin-SDS aggregate mobility of the water molecules trapped between the polypeptide chain and micellar aggregates is significantly retarded. It may be noted that the solvation dynamics of DCM in SDS micelles at a higher temperature (45 °C) and in the presence of KP buffer is faster than that carried out previously at 20 °C in the absence of KP buffer.12a We ascribe the faster dynamics mainly to the higher temperature. It may be recalled that the temperature dependence of solvation dynamics in micelles has been predicted by recent computer simulations on solvation dynamics in micelles.13b The anisotropy decay of DCM in gelatin-SDS aggregate is also found to be very different than in SDS micelles. The anisotropy decay may be analyzed by the “wobblingin-cone” model.27 According to this model, the decay part of the anisotropy given in eq 3 is a product of four independent motions: (i) wobbling motion, rw(t); (ii) translational motion, rt(t); (iii) overall rotation of the SDS micelles, rM(t); and (iv) overall rotation of the polypeptidesurfactant aggregate, rA(t):27c
{r(0) - r(∞)}[afast exp(-t/τfast) + aslow exp(-t/τslow)] ) rw(t)rt(t)rM(t)rA(t) (4) We have earlier discussed in detail analysis of anisotropy
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decay in a polymer-surfactant aggregate.27c Briefly, the slow component (τslow) of anisotropy decay is related to the time constant for translational diffusion (τD), overall rotation of the spherical micelle (τM), and overall rotation of the gelatin-SDS aggregate (τA) as27c
(τslow)-1 ) (τD)-1 + (τM)-1 + (τA)-1
(5)
The hydrodynamic radii (rh) of gelatin-SDS aggregate and SDS micelles in gelatin-SDS aggregate are about 26022a and 24 Å,23 respectively. τA may be calculated from27 3
τA )
4πηrh 3kT
τslow ≈ kbf-1
kbf )
(7)
For SDS micelles in gelatin-SDS aggregate, τM is calculated to be 14.3 ns. From eq 7, the value of τD is evaluated to be 1.4 ns. The translational diffusion coefficient (DT) is estimated from DT ) (rM2/6τD), where rM is the hydrodynamic radius of SDS micelles in gelatin-SDS aggregate. The value of the translational diffusion coefficient (DT) of DCM in gelatin-SDS aggregate is 6.8 × 10-10 m2 s-1 and that in SDS micelles is 9.8 × 10-10 m2 s-1. Thus translational diffusion of the probe along the surface of SDS micelles is slightly faster than that in gelatin-SDS aggregate. In summary, this analysis indicates that though the time constants of anisotropy decay are different in gelatinSDS aggregate and SDS micelles, the translational diffusion coefficients are quite close. While solvation dynamics of water molecules around DCM in a gelatinSDS aggregate is significantly slower (∼13 times) than that in SDS micelles, the translational diffusion coefficients of the fluorescence probe differ only by 1.3 times. If the solvation dynamics in the gelatin-SDS aggregate were due exclusively to the polar residues and headgroups of gelatin and SDS, they should have been on the order of chain dynamics of the surfactants. The chain dynamics occur on a 100 ns time scale.28 The observed average solvation time in this work (1.78 ns) is very much smaller than this. This suggests that the solvation dynamics is not due to polymer chain dynamics and is mainly due to trapped water molecules. Of course, one cannot rule out dynamics of local segments of the polypeptide/surfactant. According to a theoretical model proposed by Nandi and Bagchi,16 the slow solvation in the vicinity of a biological macromolecule arises as a result of a dynamic exchange between bound and free water molecules. The “bound” water molecules refer to those that are hydrogen-bonded to the macromolecule. The magnitude of the slow component of solvent relaxation depends on the difference (28) (a) Cassol, R.; Ge, M.-T.; Ferrarini, A.; Freed, J. H. J. Phys. Chem. B 1997, 101, 8782. (b) Sung-Suh, M. M.; Kevan, L. J. Phys. Chem. A 1997, 101, 1414.
(8)
where kbf is the rate constant for bound-to-free interconversion:
(6)
For gelatin-SDS aggregate τA is calculated to be more than 18 000 ns, i.e., exceedingly slow, and hence may be neglected. Thus one may rewrite eq 5 as
(τslow)-1 ≈ (τD)-1 + (τM)-1
between the free energy of the bound (G0b) and the free water molecules (G0f ), with G0b < G0f . The marked slowing down of the solvation dynamics in the gelatin-SDS aggregate in comparison with the SDS micelles may be attributed to an increase in the binding energy of the “bound” water molecules. In the limit of very high binding energy (i.e., |∆G0bf|), the slow component of solvation (τslow) is given by1c,16
( ) (
)
kBT -(∆G0bf + ∆G*) exp h RT
(9)
where ∆G* is the activation energy for the conversion of free-to-bound water molecules. Using eqs 8 and 9 and the average solvation times (〈τs〉, Table 1), one may calculate the binding energy |∆G0bf| in different systems. We have used ∆G* ≈ 900 cal mol-1. For SDS micelles, |∆G0bf| is found to be 3.4 kcal mol-1, whereas for the gelatin-SDS aggregate it is 5.0 kcal mol-1. It should be noted that dynamic exchange between water molecules trapped between SDS and gelatin chains may involve local motion of the macromolecular chains (SDS/ gelatin). Such a rearrangement may occur on a slow time scale and this may also be responsible for the slower solvation dynamics in the gelatin-SDS aggregate compared to SDS micelles at the same temperature. 5. Conclusion This work demonstrates that the solvation dynamics of DCM in a polypeptide-surfactant (gelatin-SDS) aggregate is substantially slower than that in SDS micelles. At 45 °C the cac of the SDS-gelatin system is found to be about 0.5 mM. This is smaller than the cmc of SDS (3 mM) in the presence of KP buffer. The slow dynamics of water molecules in gelatin-SDS aggregate is attributed to the strong binding of water molecules with the macromolecular assemblies and hindered dynamic exchange. The anisotropy decay of DCM in gelatin-SDS aggregate exhibits large residual anisotropy with a longer relaxation time compared to SDS micelles. The translational diffusion coefficient of the probe in gelatin-SDS aggregate is only about 1.3 times smaller than that in SDS micelles. It would be interesting to study the dynamics of the water molecules trapped between polypeptide and surfactant by other techniques, e.g., NMR and NOE. Acknowledgment. Thanks are due to the Department of Science and Technology (DST), Government of India, the “Femtosecond Laser Facility” and to the Council of Scientific and Industrial Research (CSIR) and the Department of Science and Technology (DST), Government of India, for generous research grants. A.H. thanks CSIR for a fellowship, and P.S. thanks DST for a fellowship. A.D.B. thanks the Government of India for a postdoctoral fellowship. LA035647M
