Conformational Dependence of Triton X-100 on Environment Studied

D2O, of 99.6% deuterated, is produced by Beijing Chemical Factory of China. 1H T1 ..... Crook, E. H.; Forduce, D. B.; Trebbi, G. F. J. Phys. Chem. 196...
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Langmuir 2000, 16, 3030-3035

Conformational Dependence of Triton X-100 on Environment Studied by 2D NOESY and 1H NMR Relaxation Han-Zhen Yuan,† Gong-Zhen Cheng,§ Sui Zhao,‡ Xi-Jia Miao,† Jia-Yong Yu,‡ Lian-Fang Shen,† and You-Ru Du*,† State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics, Wuhan Institute of Physics and Mathematics, The Chinese Academy of Sciences, Wuhan 430071, People’s Republic of China, Institute of Photochemistry, The Chinese Academy of Sciences, Beijing 100101, People’s Republic of China, and Department of Chemistry, Wuhan University, Wuhan 430072, People’s Republic of China Received May 11, 1999. In Final Form: November 8, 1999 Conformation of Triton X-100 (TX-100) in the bulk and in aqueous solutions at concentrations higher and lower than the critical micellar concentration (cmc) was studied by NMR relaxation, two-dimensional nuclear Overhauser enhancement spectroscopy (2D NOESY), and molecular simulation by HYPERCHEM. Results show that motion of TX-100 in dilute solution (0.5 cmc) is in the extreme narrowing condition. Molecules are in the single molecular state with an extended polyoxyethylene chain. In forming micelles, these hydrophilic polyoxyethylene chains, staying in the exterior of the micellar core, coil, bend, and align along the surface of the TX-100 micellar core, forming a layer thick in dimension and loose in structure around the micellar core with a certain number of water molecules included. This hydrophilic layer is in contact with the solvent, water, keeping the micellar solution stable. The micelle is spherical at 10 cmc. Motions of all of the protons of TX-100 in the bulk, even of those on the polyoxyethylene chain, are more restricted than motions of those in the micellar core. Exponential decay of the proton spin-spin relaxation of TX-100 in the bulk shows the absence of oriented self-aggregation. Very short internuclear distances of the protons between hydrophilic and hydrophobic parts of TX-100 bulk measured from the 2D NOESY spectrum show that the molecules are arranged randomly, closely, and uniformly, which is the origin for the high viscosity of TX-100 bulk. Results are discussed in terms of inter- and intramolecular interactions.

Introduction Triton X-100 (TX-100) is one of the nonionic surfactants widely used both in biochemical and chemical processes and has been commercially available for a long time. Its ability to undergo self-aggregation in aqueous solution has been intensively studied by many methods, including nuclear magnetic resonance (NMR).1-4 Although it is generally accepted that the nonpolar hydrocarbon chains are packed in the interior of the micellar core, whereas the hydrophilic polyoxyethylene chains stay outside the core and move relatively freely in the solvent, keeping the micelle stable in the solution,5 there are controversies on the penetration of water into the hydrophobic micellar core of the surfactants6-10 and the mobility of the hydrocarbon chain in the interior of the micellar core. The hydrocarbon part of the micelle was regarded as essentially the same as a liquid hydrocarbon;5 however, some authors confirm that the motion of these hydrocarbon * Corresponding author. Fax: 0086-27-87885291; e-mail: [email protected]. † Wuhan Institute of Physics and Mathematics. ‡ Institute of Photochemistry. § Wuhan University. (1) Kushner, L. M.; Hubbard, W. D. J. Phys. Chem. 1954, 58, 1163. (2) Crook, E. H.; Forduce, D. B.; Trebbi, G. F. J. Phys. Chem. 1963, 67, 1987. (3) Wright, A. K. J. Colloid Interface Sci. 1976, 55, 109. (4) Ribreiro, A. A.; Dennis, E. A. J. Phys. Chem. 1976, 80, 1746. (5) Stainby, G.; Alexander, A. E. Trans. Faraday Soc. 1950, 46, 587. (6) Podo, F.; Ray, A.; Nemethy, G. J. Am. Chem. Soc. 1973, 95, 6164. (7) Clemett, C. J. J. Chem. Soc. A 1970, 2251. (8) Corkill, C. M.; Goodman, J. E.; Wyer, J. Trans. Faraday Soc. 1969, 65, 9. (9) Gruen, D. W. R. J. Colloid Interface Sci. 1981, 84, 281. (10) Clifford, J. Trans. Faraday Soc. 1965, 61, 1276.

chains is more restricted than in a liquid hydrocarbon of the same chain length.4,10 Conformation of the hydrophilic polyoxyethylene chains of the surfactant neither in micelles nor in the single molecular state in the aqueous solution is known.11 We have shown in our previous papers12,13 semiquantitatively that the first polyoxyethylene group and the phenoxy ring together with part of the p-tert-octyl chains participate in the formation of the micellar core, the surface of which is a rigid layer of compact texture with the oxyethylene groups toward the hydrophilic environment, water, and the alkyl chains toward the interior of the micellar core. Motion of the hydrocarbon chains included in the interior, although more free than that of the groups in the compact surface layer of the micellar core, is far from the extreme narrowing condition. It seems that the arrangement and relative mobility of different parts of the molecule in various media are still a considerable interest. Thanks to the development of the NMR techniques and computational chemistry, we are now able to study conformation of molecules in solution by the use of these methods. It is well-known that NMR method, especially two-dimensional nuclear Overhauser enhancement spectroscopy (2D NOESY), is an effective method to study the three-dimensional structure of large molecules such as proteins that have long motional correlation time of molecules.14,15 Cross-signals in a (11) Robson, R. J.; Dennis, E. A. J. Phys. Chem. 1977, 81, 1075. (12) Yuan, H. Zh.; Zhao, S.; Yu, J. Y.; Du, Y. R. Sci. China, Ser. A 1999, 42, 319. (13) Zhao, S.; Yuan, H. Zh.; Yu, J. Y.; Du, Y. R. Colloid Polym. Sci. 1998, 276, 1125. (14) Ernst, R. R.; Bodenhausen, G.; Wokaun, A. Principles of Nuclear Magnetic Resonance in One and Two Dimensions; Oxford University Press: New York, 1987.

10.1021/la990568p CCC: $19.00 © 2000 American Chemical Society Published on Web 02/17/2000

Conformation of Triton X-100

NOESY spectrum rely on the cross-relaxation of longitudinal magnetization during the mixing time. One can extract valuable information about internuclear distances from the intensity of the cross-peaks with one known distance in the molecule.16,17 Kolehmainen18 reported intra- and intermolecular interactions between different kinds of molecules by 2D NOESY in aromatic solubilizatecholate systems. Zhao and Fung19 observed a conformational change of the alkyl chain upon the formation of micelles in sodium cis-7-dodecane-1-yl sulfate system by 2D NOESY. Intermolecular information about direct and remote correlations within alkyl chains can also be resolved by heteronuclear Overhauser effect spectroscopy (HOESY).20 In this article, we present our recent results on the conformation of TX-100 in aqueous solution, in micelles, and in the bulk by NMR relaxation, 2D NOESY, and conformational calculation by using HYPERCHEM, a multifunctional molecular graphic package.21

Langmuir, Vol. 16, No. 7, 2000 3031 Table 1. 1H T1, T2 (ms), and TRa of Triton X-100 Bulk and at 10 and 0.5 cmc Concentrations Measured at Resonance Frequency of 500 MHz for 1H and 25 °C bulk

10 cmc

0.5 cmc

TX-100

T2

T1

TR

T2

T1

TR

T2

T1

TR

H1 H2 H3 H4 H5 H6 H7 H8

43 44 53 75 89 30 39 130

602 691 549 556 529 468 319 332

.07 .06 .10 .13 .17 .06 .12 .39

47 70 45 66 318 23 33 130

555 669 475 465 786 386 261 311

.08 .10 .10 .14 .40 .06 .13 .42

861 1073 508 524 606 383 306 601

766 1175 496 592 701 403 325 557

.91 1.12 1.02 .89 .86 .95 .94 1.08

a T ) T / T . Estimated errors of T and T values are smaller R 2 1 1 2 than 5 and 10%, respectively.

Experimental Section TX-100 is the chemical pure reagent of Nailai Co. of Japan. D2O, of 99.6% deuterated, is produced by Beijing Chemical Factory of China. 1H T1 (spin-lattice) and T2 (spin-spin) NMR relaxation time measurements were performed on a Bruker ARX500 with a proton frequency of 500.13 MHz, using inversion recovery and CPMG pulse sequences, respectively, at 24 °C. Accumulations (16) were acquired for concentrated solution and the bulk. For dilute solution the number of accumulations was increased up to 80. D2O was used as solvent instead of water to weaken the water proton signal. Meanwhile, the presaturation method was used to further suppress the proton signal of the solvent. 2D NOESY experiments were performed with the standard three-pulse sequence.14 For 0.5 cmc solution, mixing time of 500 ms and 256 accumulations were used. Mixing times of 500, 200, 100, 50, 25, and 10 ms and 16 accumulations were used for TX-100 at a concentration of 10 cmc. Mixing times of 500 and 10 ms and 16 accumulations were used for TX-100 in the bulk. Spin-lattice and spin-spin relaxation times of TX100 bulk were also measured on a Varian Mercury 300 at 300.01 MHz for 1H at 25 and 50 °C.

Figure 1. Formula and proton numbering of Triton X-100 (TX-100) molecule.

Results 1

1. H Relaxation. Concentrations of TX-100 are expressed in terms of critical micellar concentration (cmc), 0.30mM.22 1H T1,T2, and the ratio TR (T2/T1) of all of the proton signals of TX-100 bulk and in 10 and 0.5 cmc solutions are listed in Table 1. H1 to H8 stand for the corresponding proton signals as indicated by the formula shown in Figure 1. It is obvious that TR of all of the proton signals of TX100 in the dilute solution (0.5 cmc) is about unit, but in the concentrated solution (10 cmc) they are about 0.1 with the exception of H5 and H8. It is surprising that TR of all of the protons of TX-100 in the bulk is about 0.1 with the exception of H8. This suggests that the motion of TX-100 in solutions of concentration lower than cmc is in the extreme narrowing condition, and that molecules are in the single molecular state. However, once the micelles are formed (concentration of 10 cmc), motions of most of (15) Macura, S.; Ernst, R. R. Mol. Phys. 1980, 41, 95. (16) Borgias, B. A.; Gochin, M.; Kerwood, D. J.; James, T. L. Prog. Nucl. Magn. Reson. Spectrosc. 1990, 22, 83. (17) Kessler, H.; Gehrke, M.; Griesinger, C. Angew. Chem., Int. Ed. Engl. 1988, 27, 490. (18) Kolehmainen, E. Magn. Reson. Chem. 1988, 26, 760. (19) Zhao, J.; Fung, B. M. J. Phys. Chem. 1993, 97, 5185. (20) Palmas, P.; Tekely, P.; Mutzenhardt, P.; Canet, D. J. Chem. Phys. 1993, 99, 4775. (21) Hypercube Inc. Hyperchem Release 4.5 for windows: Molecular Modelling System, 1995. (22) Fendler, J. H. Membrane Mimetic Chemistry; Johns Wiley and Sons: New York, 1982; p 9.

Figure 2. 2D NOESY spectrum of TX-100 in aqueous solution at a concentration of 0.5 cmc at mixing time of 500 ms.

the protons (hydrophobic and a part of the hydrophilic protons next to the phenoxy ring) are seriously restricted (TR ) 0.1). Meanwhile, motion of the polyoxyethylene protons located in the exterior of the micellar core is only relatively more restricted than that of TX-100 in dilute solution (0.5 cmc), but less restricted than those of the protons forming the micellar core. Motion of the end methyl groups is always relatively free because of their intrinsic internal rotation. Motions of all of the protons of TX-100 in the bulk are extremely restricted with no exception of the polyoxyethylene protons. 2. 2D NOESY Experiment. The contour plot of 2D NOESY spectrum for TX-100 dilute solution with mixing time of 500 ms is shown in Figure 2. Cross-peaks between protons on adjacent carbon atoms (H1-H2, H1-H7, H2H3, H3-H4, H6-H7, H6-H8, and H7-H8) are observed. Interaction of the hydrophilic polyoxyethylene chain with

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Yuan et al.

Figure 4. Intensities of the cross-peaks in the 2D spectra of TX-100 micelles as a function of the mixing time: (A) H1 with the hydrophobic, (B) H1 with the hydrophilic protons.

Figure 3. 2D NOESY spectrum of TX-100 in aqueous solution at a concentration of 10 cmc at mixing times of (A) 500 ms, (B) 100 ms, (C) 50 ms, and (D) 10 ms.

water molecules is evidenced by the presence of crosspeaks between water protons and H3, H4, H5, and even the ortho-protons of the phenoxy ring, respectively. 2D NOESY spectra for TX-100 in aqueous solution at a concentration of 10 cmc were obtained with mixing times of 500, 200, 100, 50, 25, and 10 ms. Contour plots of spectra obtained with mixing times of 500, 100, 50, and 10 ms are shown in Figure 3. For mixing times of 500, 200, 100, 50, 25, and 10 ms, the ratios of the intensity of the diagonal peak of H1 to the intensity of the cross-peak between H1 and H2 are 1.6, 1.6, 3.1, 6.8, 9.8, and 33, respectively. The intensities of the cross-peaks (H1-H6, H1-H7, and H1H8) reach their maximum already at a mixing time of 100 ms. However, those of H1-H3 and H1-H4, especially H1-H5, still increase gradually, as shown by the increase of cross-peak intensity as a function of mixing time in Figure 4. This indicates that diffusion of magnetization along the hydrophobic chain is possible under this condition, although spin-diffusion does not completely couple all the nuclei in the micelle. It should be noted that a number of cross-peaks seen at early mixing times (shorter than 50 ms) are likely to be artifacts of the 2D experiment. These cross-peaks do not evolve as the magnetization of the other cross-peaks builds up, but simply decay with mixing time. In contrast to the case of dilute solution (0.5 cmc), significant interaction between H1 and all the other protons, especially the hydrophobic protons, occurs for TX-100 micelles. Cross-peaks of water neither with H4, H3 (the first oxyethylene protons), nor with the protons on the phenoxy ring and on the hydrophobic chain were found in spectra of TX-100 micelles in water solution (Figure 3). 2D NOESY spectra of TX-100 in the bulk at mixing times of 500 and 10 ms were obtained. Similar to the case of TX-100 in micelles, cross-peaks of all of the proton pairs were observed (Figure 5).

Intensities of the cross-peaks of the above three systems were measured. Distances between proton pairs in TX100 dilute solution, micelles, and in the bulk were calculated with the measured intensities of the cross-peaks at early mixing times to ensure that the distance specificity is not destroyed by spin diffusion.16 The following formula was used with the aid of the known distance H1-H2 in the molecule, 2.4 Å, and the corresponding cross-signal intensity

x

I12 rmn ) Imn r12

where, I12 and Imn are the intensities of the cross-peak between H1 and H2 and that between the two nuclei of interest, respectively. r12 and rmn are the corresponding internuclear distances. There are altogether eight resolvable resonance peaks in the TX-100 1H spectrum. They represent eight kinds of chemically equivalent groups of spins, namely, (CH3-)3- (H8), CH2- (H6), (CH3-)2- (H7), the ortho- (H2) and meta- (H1) protons on the phenoxy ring, OCH2- (H3), CH2O- (H4), and (CH2CH2O)8H- (H5). Chemical shifts of protons in (CH2CH2O)8 (labeled as a single group H5) are not equal, as indicated by the broad signals in the spectra, Figures 2 and 3. This results in the impossibility of determination of the interproton distances between each individual proton pair in the groups of chemically equivalent protons directly from the intensity of the cross-peak of the 2D NOESY spectrum. However, the average distance between groups of spins can be obtained from their cross-peak intensities.23,24 Therefore, the interproton distances calculated according to the above equation is actually the geometrically average distances between groups having chemically equivalent protons. Although these calculated internuclear distances are not precise, they can characterize the conformation of the system we are studying. It should be mentioned that the (23) Liu, H.; Thomas, P. D.; James, T. L. J. Magn. Reson. 1992, 98, 163. (24) Wuthrich, K.; Billeter, M.; Braun, W. J. Mol. Biol. 1983, 169, 949.

Conformation of Triton X-100

Langmuir, Vol. 16, No. 7, 2000 3033 Table 2. Internuclear Distances, r (Å), between Protons of TX-100 in Solution at Concentrations of 0.5 and 10 cmc and the Bulk Measured from 2D NOESY Experiments with Various Mixing Times, and by HYPERCHEM Calculationa rcal H1-H2 2.4 H1-H3 5.3 H1-H4 6.9 H1-H5 19.0 H1-H6 3.9 H1-H7 3.8 H1-H8 4.9 H2-H3 3.4 H2-H4 4.7 H2-H5 16.8 H2-H6 5.9 H2-H7 5.7 H2-H8 6.3 H3-H4 2.8

rs500 rm50 rB10 2.4 3.2 3.2 2.8

2.4 4.5 5.2 7.4 2.9 3.2 4.8 3.0 4.2 6.4 4.1 4.3 5.1 2.9

2.4 4.9 3.2 3.3 4.8 3.1 4.8 4.7 5.2 2.8

H3-H5 H3-H6 H3-H7 H3-H8 H4-H5 H4-H6 H4-H7 H4-H8 H5-H6 H5-H7 H5-H8 H6-H7 H6-H8 H7-H8

rcal

rs500 rm50 rB10

14.8 7.3 7.7 7.6 13.7 9.4 9.3 9.3 20.9 22.2 21.1 3.3 3.4 5.0

3.2 3.2 4.1

4.8 6.2 5.5 6.4 5.3 5.9 5.5 7.8 6.8 7.2 3.2 3.7 4.3

3.8 4.2 5.1 4.1 6.0 5.6 5.2 5.1 4.9 3.1 4.4 4.2

a Internuclear distances are averaged values of different configurations due to the existence of flexible chains in the molecule. rcal, measured by HYPERCHEM; rs500, measured from 2D NOESY experiment with mixing time of 500 ms for 0.5 cmc solution; rm50, measured from 2D NOESY experiment with mixing time of 50 ms for solution of 10 cmc; rB10, measured from 2D NOESY experiment with mixing time of 10 ms for TX-100 bulk. Estimated error is 0.2 Å.

Figure 5. 2D NOESY spectrum of TX-100 bulk at mixing times of (A) 500 ms, (B) 10 ms.

proton signals of the polyoxyethylene chain, H5, overlap seriously. The intensities of cross-peaks of H5 with other protons are the contribution of the sum of the eight oxyethylene groups. The internuclear distances between H5 and other protons listed in Table 2 are the average values of the eight oxyethylene groups. Nine methyl protons of the end methyl groups (H8) and six methyl protons of the dimethyl groups next to the phenoxy ring (H7) are also taken into account in calculation of the internuclear distances from the intensities of the corresponding cross-peaks. Interproton distances calculated from the 2D NOESY spectra are listed in Table 2. To compare qualitatively the conformations of TX-100 in various environments, molecular structure of TX-100 in water solution was simulated by HYPERCHEM. Once the molecular model was built, it was then put in a periodic box filled with various amounts of water (200, 100, 50, 25, and 11 water molecules per TX-100 molecule), and the systems were then geometrically optimized using MM+ molecular mechanics program. All of the optimized structures of a single TX-100 molecule in different amounts of water show extended polyoxyethylene chains. The energies of the optimized structures are 38.27, 37.99, 43.06, 38.31, and 37.99 kcal/mol, respectively. Proton inter-

nuclear distances of these optimized structures were measured and are almost identical, one set of which is listed in Table 2 as rcalc. Here the internuclear distances between H5 and other protons are the average distances from protons on each oxyethylene group to the proton of interest (i.e., the distance between the center of gravity of the protons in the polyethylene chain and the protons of interest). It should be noted that these values being averaged cannot give the exact conformation of the molecule; however, they can give reasonable information about a change of conformation qualitatively. Internuclear distances of TX-100 in solution at a concentration of 0.5 cmc obtained from the 2D experiment agree quite well within experimental error with those calculated by HYPERCHEM. Cross-peaks of those internuclear distances greater than 5 Å were not observed in the 2D spectrum. This shows that TX-100 in dilute solution is in the single molecular state with extended hydrophilic chain. It further supports the observation of the relaxation experiment. For TX-100 micelles, internuclear distances were calculated from the 2D NOESY spectra with mixing time 50 ms to ensure that the distance specificity is restored to this slowly tumbling system. From Table 2 it is evident that, similar to the case of dilute solution, interproton distances of adjacent carbon atoms in micelles agree with those of the single molecule calculated by HYPERCHEM. However, cross-peaks, which were not observed in the case of dilute solution, appear in the 2D spectrum of TX100 micelles. Interproton distances of these proton pairs, especially those between hydrophilic protons (H3, H4, and H5) and protons on the hydrophobic chain (H6, H7, and H8), respectively, are significantly shorter than those of the single molecule calculated by HYPERCHEM where the hydrophilic polyoxyethylene chain is in the extended form. Relaxation and 2D NOESY experiments show that TX100 bulk is a slowly tumbling system. Therefore, interproton distances were calculated from 2D experiment with a mixing time of 10 ms. Similar results (Table 2) were obtained as for TX-100 micelles with the exception that

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the internuclear distances between the hydrophilic H5 and the hydrophobic protons, respectively, are even shorter. Although this approach does not give precise quantitative results, it does give a readily understood insight into the change of conformation of the surfactant molecule in this complex system qualitatively. Discussion 1. Structure of Micellar Core and Penetration of Water. Although controversies of different opinions on the penetration of water into the micellar core are existing, 2D NOESY spectrum of TX-100 in solution at a concentration of 10 cmc does not give evidence about interaction between a water proton and any protons of a TX-100 molecule other than H5. This gives direct evidence that penetration of water into the micellar core is excluded. No cross-peaks, even between the water proton and the protons of the first hydrophilic oxyethylene group (H4 and H3), were observed, although they are present in the 2D spectrum of dilute solution shown in Figure 2. This implies that H3 and H4 are involved in the surface layer of the micellar core, which is in agreement with our previous observation.13 Consequently, according to the prediction of Dennis et al.11 TX-100 micelles are spherical at a concentration of 10 cmc. 2. Conformation of Hydrophobic Part of TX-100 Micelles in Water Solution. Additional correlation peaks were observed when the concentration of TX-100 was increased from 0.5 to 10 cmc. This shows that the interaction between these proton pairs became strong enough to give rise to significant NOE between them. According to the NOE dependence on r-6, obviously, it originates from the intermolecular interaction in the micelles where the hydrophobic protons of different TX100 molecules are very closely packed. Internuclear distances of protons in the neighboring groups (H1-H7, H2-H3, H3-H4, H6-H7, and H6-H8) of TX-100 molecules in micelles obtained from the 2D experiment of 10 cmc solution are almost identical to those for 0.5 cmc solution and to those calculated by HYPERCHEM for a single molecule, respectively. However, those for protons in the next nearest neighboring groups (H1-H3, H1-H6, H2-H4, H2-H7, and H7-H8) are shorter by about 1 Å than those calculated by HYPERCHEM. The further the protons are apart from each other in the molecule, the more intense is the deviation of interproton distances measured by 2D experiment from the HYPERCHEM calculated values. It is well known that the NOESY (through space) effect can be either intra- or intermolecular in origin, and it is impossible to discriminate between them when one deals with interaction between molecules of the same kind. Therefore, we can only assume that the interproton distances obtained by the 2D NOESY experiment is the weighted average of these two interactions. Thus, the fact that interproton distances of neighboring groups obtained by 2D experiment are almost identical to those calculated by HYPERCHEM suggests that the average intermolecular distances between these protons are the same as those of the intramolecular ones. Because the NMR results reflect the average interproton distances due to the large number of conformations possible for the flexible aliphatic chain in this system, the significant shorter interproton distances beyond the neighboring groups measured by 2D experiment seem to originate from the following fact. The packing of the hydrophobic chains may not be strictly oriented. The end methyl group of one molecule may be packed near a phenoxy ring of another

Yuan et al.

molecule. This intermolecular interaction may significantly contribute to the intensity of the cross-peak when the distance of the proton pairs are shorter than that of the corresponding proton pairs within the molecule. Second, the molecular mechanics computation for optimal conformation by HYPERCHEM is based on a single molecule put in various amounts of water (200, 100, 50, 25, and 11 water molecules per TX-100 molecule). In forming micelles, the hydrophobic protons are transferred to a hydrocarbon environment that is significantly less polar than that of water. The chains become more flexible and a part of them can be curved. Probably the optimal conformation of the hydrocarbon chains in the micellar core of hydrophobic environment is such that a part of the hydrocarbon chains are curved and bent back because of the lack of enough space in the center of the micellar core for the end methyl groups. This is an indication of the conformational change of the hydrophobic chain in micellization. This part of the shortening of interproton distances will also contribute to the overall weighted average value. The above discussion is in good agreement with our previous result. The inner hydrophobic protons farther from the dense phenoxy rings along the hydrophobic chain can also participate in the formation of the compact surface layer of the micellar core, although with less opportunity, for example, 35% for the end methyl protons.13 3. Conformation of Hydrophilic Polyoxyethylene Chain in Water Solution. We can deduce both from the proton spin-lattice and spin-spin relaxation times shown in Table 1 and the 2D NOESY spectrum (Figure 2) that TX-100 in dilute solution (0.5 cmc) is in the single molecular state with extended hydrophilic chain for the following reasons. That TR of all protons is about unit suggests free motion of small nonassociated molecules, the motion of which is in the extreme narrowing condition. Besides, the fact that internuclear distances between protons attached to adjacent carbon atoms obtained from the 2D NOESY spectrum agree with those calculated by HYPERCHEM, whereas no cross-peaks between protons situated 5 Å and farther apart could be detected, is in agreement with the simulated molecule with an extended hydrophilic chain. There is a decrease of the average internuclear distances between protons on the first segment of the polyoxyethylene chain (H3 and H4) and those on the hydrophobic part of the molecule (H1, H2, H6, H7, and H8), respectively, in micellization (Table 2). This implies that these hydrophilic protons have gotten closer to the hydrophobic part, which may lead to the participation of these protons in the formation of the surface layer of the micellar core, having been evidenced by the relaxation study mentioned above. The motion of protons of the hydrophilic polyoxyethylene chains (H5) of TX-100 micelles in aqueous solution at a concentration of 10 cmc is relatively less restricted (TR ) 0.4) than the motions of protons forming the micellar core (TR ) 0.1). However, it seems to be more restricted than the motion of the protons on the polyoxyethylene chains in dilute solution (TR ) 0.86). This implies that the polyoxyethylene chains are compact in certain degrees. The average internuclear distances between H5 and other protons (H1, H2, H3, H4, H6, H7, and H8) in micelles are very much shorter than those obtained for dilute solution where cross-peaks of these proton pairs could not be detected. They are also shorter than those calculated by HYPERCHEM. In both cases the polyoxyethylene chain of the single TX-100 molecule in water solution is in the extended form. These large differences suggest a confor-

Conformation of Triton X-100

mational change of the polyoxyethylene chain in the formation of micelles. The polyoxyethylene chains might become coiled or bent and aligned along the surface of the TX-100 micellar core in micellization. A layer thick in dimension and loose in structure surrounding the micellar core is formed. The loose layer allows a certain amount of water molecules to be included, forming a thick hydrophilic layer that keeps in contact with the solvent water, rendering the micellar solution stable. This conformation provides the possibility of strong intermolecular interaction between the hydrophobic protons in the micellar core and those of the polyoxyethylene chains. It supports the somewhat restricted motion of the polyoxyethylene chain of TX-100 in micellar form mentioned above. Interproton distance between HW (proton of water) and H5 of TX-100 in aqueous solution at a concentration of 10 cmc (4.9 Å) is larger than that of TX-100 in the dilute solution of 0.5 cmc (3.6 Å). This shows that the hydrophilic polyoxyethylene chain is surrounded by fewer water molecules in the exterior part of the micelle than that in the single molecular state. 4. Conformation of TX-100 Molecule in the Bulk. Proton spin-lattice and spin-spin relaxation measurements of TX-100 in the bulk show that TR of protons of the hydrophobic part, H3 and H4, are almost identical with the corresponding values of TX-100 in the micellar state. This suggests that motions of these protons in TX100 are also restricted. However, one can find that their spin-spin relaxation processes behave significantly differently if simulation of the decay curves were traced. Protons forming the surface layer of the micellar core (H1, H2, and H3) of TX-100 decay biexponentially, whereas those of TX-100 bulk decay exponentially. This is obviously shown in Figure 6. The lack of biexponential behavior in the spin-spin relaxation of TX-100 bulk shows that oriented self-aggregation does not occur. Besides, it is worth noticing that the TR value of H5 in TX-100 bulk is considerably smaller than that in TX-100 micelle. Motion of the polyoxyethylene protons of TX-100 in the bulk is even more restricted than that in the micelle. The restricted motions of TX-100 molecules in the bulk might arise from the strong interaction among molecules, resulting in the formation of a very dense state. To verify this fact, spin-lattice and spin-spin relaxation times of TX-100 bulk were also measured at 300 MHz. T1 and T2 values measured at 25 and 50 °C are listed in Table 3. T1 values measured at 25 °C at 300 MHz (Table 3) are shorter than those measured at 500 MHz (Table 1), whereas T2 values are of the same order, which is characteristic for relaxation of liquids measured at different resonance frequencies. T1 and especially T2 values increased as the temperature was increased to 50 °C, resulting in the significant increase in TR values for H5-H8 (0.94-1.03 respectively in Table 3). This is typical for molecular motion in the extreme narrowing condition. That is to say, motions of protons farther from the phenoxy ring are almost in the extreme narrowing condition at higher temperatures. The strong interactions between molecules are weakened because of thermal motion. Molecules tumble faster and τc becomes shorter, so the viscous TX100 becomes thinner. The increase of TR values for the phenoxy protons and for the protons of the first oxyethylene group next to the phenoxy ring is not so obvious because of the strong interaction among the phenoxy rings themselves. The appearance of strong cross-peaks between pairs of all the protons observed in the 2D NOESY spectrum of TX-100 bulk (Figure 5) supports that TX-100 bulk is dense at room temperature. Besides, the internuclear distances

Langmuir, Vol. 16, No. 7, 2000 3035

Figure 6. Simulation of spin-spin relaxation data at resonance frequency of 500 MHz for H1, H2, and H3 of TX-100 micelle and of TX-100 bulk, respectively. Dotted curves represent exponential simulation and solid curves represent biexponential simulation. Table 3. 1H T1, T2 (ms), and TRa of TX-100 Bulk at 25 and 50 °C with Resonance Frequency of 300 MHz for 1H T1 H1 H2 H3 H4 H5 H6 H7 H8

T2

TR

25 °C

50 °C

25 °C

50 °C

25 °C

50 °C

369 412 297 269 327 262 185 235

458 556 309 332 475 195 182 360

39 35 34 37 69 67 38 42

88 64 86 135 445 123 186 370

0.11 0.08 0.11 0.14 0.21 0.26 0.21 0.26

0.19 0.12 0.28 0.41 0.94 0.63 1.02 1.03

a T ) T /T . Estimated errors of T and T values are smaller R 2 1 1 2 than 5 and 10%, respectively.

of all of the proton pairs calculated from the intensities of the corresponding cross-peaks are very close, ranging from 2.8 to 3.6 Å. Such short distances between protons on different parts of a long molecular chain are impossible, even when the chain is coiled. Therefore a large part of the cross-peaks should arise from the intermolecular interaction of randomly oriented molecules. This shows that the molecules in the bulk are randomly, uniformly, and closely packed, forming a liquid of high viscosity. That is really how pure TX-100 looks. Acknowledgment. This project was supported by the National Climbing Project (B) “The Fundamental Research of Alkali-Surfactant-Polymer Flooding for Enhanced Oil Recovery”. LA990568P