2583
J. Phys. Chem. 1991, 95,2583-2587 phase. This possibility cannot be excluded for Winsor I1 microemulsions. Another conclusion from the ESEM analysis is that the “oil” region of the microemulsions seems to be well separated from the “water” region with little or no mixing between toluene and butanol. Furthermore, the invariance of the normalized modulation depths in the Winsor 111 microemulsions as a function of SM provides evidence at a molecular level that the interface of these microemulsions is little affected by SM variation.
Acknowledgment. Thanks are due to the Israel-Italy Scientific exchange program sponsored by the Italian CNR and the Israeli Ministry of Science and Development which made the above collaboration possible. This study was made possible in part by funds granted to D.G. through a fellowship program sponsored by the Charles H. Revson Foundation. The statements made and views expressed are solely the responsibility of the authors. Registry No. SDS, 151-21-3; 5-DSA, 29545-48-0; IO-DSA, 5061 398-4; I6-DSA, 53034-38-1; NaCI, 7647-14-5; toluene, 108-88-3; I-butanol, 71-36-3.
Bis( 1-pyrenyimethyi) Ether as an Excimer-Forming Probe of Hydrophobicaiiy Modified Poly( N-isopropyiacryiamides) in Water FranGoise M. Winnik,* Xerox Research Center of Canada, 2660 Speakman Drive, Mississauga, Ontario, Canada L5K 2Ll
Mitchell A. Winnik, Department of Chemistry and Erindale College, University of Toronto, Toronto, Ontario, Canada MJS 1A1
H. Ringsdorf, and J. Venzmer Institut fur Organische Chemie, Johannes Gutenberg- Universitat, Mainr, J.-J. Becherweg, 18-20 0-6500 Maim, FRG (Received: July 10, 1990)
The hydrophobic probe bis( 1-pyrenylmethyl) ether [dipyme] shows intramolecular excimer fluorescence (intensity I,) in competition with fluorescence from locally excited pyrene chromophore (“monomer” emission, intensity I M ) . The intensity of the ratio I E / I M is sensitive to solvent viscosity. In addition the fine structure of the emission from locally excited pyrene is sensitive to solvent polarity. Dipyme has been used to investigate the fluidity and micropolarity of the polymeric micelles that exist in aqueous solutionsof copolymers of N-isopropylacrylamide (NIPAM) and decyl-, tetradecyl-, or octadecyl-substituted acrylamides (NIPAM to N-alkylacrylamide molar ratios in the copolymers 1OO:l and 200:l). Solutions of the copolymers in water exhibit an LCST. Fluorescenceof dipyme dissolved in the polymeric micelles shows that macroscopic phase separation is accompanied by disruption of the hydrocarbon clusters. Potential artifacts associated with the use of dipyme in aqueous polymeric systems are discussed.
Introduction Water-soluble polymers containing a small number of hydrophobic alkyl or aryl substituents are often referred to as “hydrophobically modified” polymers. The tendency of these hydrocarbons to associate in water gives these polymers amphiphilic character. Amphiphilic polymers have received considerable attention recently not only because of their unusual rheological behavior’ but also because of their unique interactions with phospholipid vesicles and other biomembrane systems.* Relatively little is known about the nature of the hydrophobic aggregates formed from these and other hydrophobically modified watersoluble polymers. Questions of interest include the size, shape, and structure of the hydrophobic clusters formed in aqueous solutions of the polymers and the mobility of lipophilic species within the clusters. Fluorescence techniques are useful in obtaining this kind of information. Pyrene provides a very useful measure of local polar it^.^ However, because pyrene has a significant solubility in water, experiments at low cluster concentration are (1) For recent reviews, see: Polymers in Aqueous Media; Advances in Chemistry Series No. 223; Glass, J. E., Ed.,; American Chemical Society: Washington, DC, 1989. (2) For a review, see: Ringsdorf, H.; Schlarb, B.; Venzmer, J. Angew. Chem.. Inr. Ed. Engl. 1988. 27, 113 and references therein. (3) Kalyanasundaram, K.; Thomas, J. K. J . Am. Chem. Soc. 1977, 99, 2039. Nakajima, A. J . Lumin. 1977, IS,277. Dong, D. C.; Winnik, M.A. Can. J . Chem. 1985,62, 2560.
CHART I TH3 HE-NH
I CHI
H3C -(CH21n
* n
I
n=9, m=100:
I
n=9, m=200:
PNIPAM-CqO/lOO PNIPAM-Cq0/200
n=13, m=100: n=13, m=200:
PNIPAM-Cjq/lW PNIPAM-Eqq/ZOO
n=17, m=100: n=17, m=200:
PNIPAM- Cqa/100 PNIPAM-Cq81200
-CO-CH
CHZ
- NH - CO -.ICH
CH2 L 1
rendered ambiguous because the spectra contain a superposition of signals from bound and unbound pyrene. Local mobility is often determined by measuring the fluorescence polarization anisotropy of probes such as perylene or 1,6-diphenyl-1,3,5-hexatriene (DPH).4 These measurements are convenient as long as the system is transparent. Both types of information are available also through the use of bis( 1-pyreny1methyl)ether(dipyme) as a fluorescent probe. Like the closely related 1,3-bis( 1-pyrenyl)propane (DPyP), dipyme forms an intramolecular excimer (Figure 1). The extent of excimer emission in both species depends upon the rate of conformational change. This motion is resisted by the local friction imposed by the environment. As a consequence, the excimer(4) Lakowicz, J. R. Principles of Fluorescence Spectroscopy; Plenum Press: New York, 1983; Chapter 5.
0022-3654/91/2095-2583$02.50/0 0 1991 American Chemical Society
2584 The Journal of Physical Chemistry, Vol. 95, No. 6, 1991 “’CHz
c-___,
I
+
/
Hz(
(Py Py)*
I
-hv
+
monomer emission
-hu’
excimer emission
Py-CHrO-CHrPy
(Dip!”)
Figure 1. Representation of intramolecular excimer formation in dipyme [bis( 1-pyrenylmethyl)ether].
to-monomer intensity ratio, IE/IM,provides a measure of the ‘microviscosity” of its environment and particularly powerful means of monitoring changes in microviscosity as the system is subjected to external stimuli. In addition the vibrational fine structure in the dipyme monomer emission is sensitive to the polarity of the probe microenvironment. This effect is much more prominent in pyrene itself, where molecular symmetry makes the (0,O)emission band (intensity 11)forbidden. Polar solvents provide anisotropic solvation of the pyrene, which relaxes the forbiddeness of the (0,O)transition relative to that of the vibrationally allowed band 13. Thus the intensity ratio 11/13reports on local polarity. This effect is absent in DPyP and most other 1-substituted pyrene derivatives, since the substituent in the 1-position effectively disrupts the electronic symmetry of the molecule. Curiously, however, this effect reappears in -OCH2-substituted pyrenes, where the &oxo substituent seems to have a symmetrizing effect on the electronic wave functions for the pyrene group. As a consequence, dipyme reports simultaneously on two features of its environment: the local friction (microviscosity) and the mic r ~ p o l a r i t y . ~ *This ~ paper describes the use of dipyme as a fluorescent probe to study the hydrophobic micellelike clusters formed with amphiphilic polymers in water. Qualitative changes in the microviscosity and in the polarity experienced by the probe are reported as a function of the structure of the copolymers and as a function of temperature for each copolymer. The polymers of interest here are derivatives of poly(N-isopropylacrylamide) (PNIPAM) containing a small percentage of pendant CnH2,,+lalkyl chains. They differ in the length of the alkyl chain (CIbC14, and C18)and in the ratio of n-alkylacrylamide to N-isopropylacrylamide (NIPAM) units (1:lOO and l:2OO).’ The structures of these polymers are shown in Chart I. Aqueous solutions of PNIPAM and its amphiphilic copolymers exhibit phase separation when heated above a critical temperature (lower critical solution temperature, LCST).*p9 In the case of PNIPAM itself, evidence from light-scattering experiments indicates that at the LCST individual chains collapse from an extended form into contracted coils.1o These in turn aggregate to create a macroscopically separated polymer-rich phase. The mechanism of phase separation in solutions of hydrophobically modified PNIPAMs is not known. Experiments using fluorescently labeled amphiphilic PNIPAMs have shown that a reorganization of the hydrophobic clusters occurs at the LCST.]] ( 5 ) Zachariasse. K. A.; Vaz, W. L.C.; Sotomayor, C.; Kllhnle, W. Biochim. Biophys. Acto 1982, 688. 323. (6) Georgescauld, D.;De”&J. P.;Lapouyade, R.; Babeau, A.; Richard, H.; Winnik, M.A. Photochem. Photobiol. 1980, 31, 539. (7) Ringsdorf. H.; Venner, J.; Winnik, F. M.Polym. Prepr., Am. Chem. Soc. Diu. Polym. Chem. 1990, 3/(1). 568. (8) Htskins, M.;Guillet, J. E. J . Mocromol. Sci. A2 1968, 1441. ( 9 ) Winnik, F. M.Mocromolecules 1990, 23, 233. (10) Fujishige, S.; Kubota, K.;Ando, I. J . Phys. Chem. 1989, 93, 331 1 . Yamamoto, I.; Iwazalri, K.; Hirotsu, S . J. Phys. Soc. Jpn. 1989, 58, 210.
Winnik et al.
Probe experiments reveal a number of interesting features about hydrophobic association in these systems. We have learned, for example, that the PNIPAM-Clo copolymers are unable to solubilize water-insoluble dyes, such as perylene, DPH, or dipyme. We found, as expected, that turbidity effects render polarization experiments with perylene or DPH useless above the LCST but that interesting information above the fate of the micellelike clusters in the CI4-and C18-substituted PNIPAM copolymers is available through examination of dipyme fluorescence. There is, unfortunately, a potential artifact associated with using either DPyP or dipyme to study hydrophobic clusters in water. This artifact derives from microcrystals of probe that are often formed during sample preparation. These microcrystals contribute a very strong excimer emission, and they resist separation by filtration or centrifugation. Since this problem appears in almost every application of these probes to aqueous solutions, part of this paper is devoted to the artifact. We provide appropriate spectra and describe the care necessary for ensuring that the probes are molecularly dispersed in the systems of interest. In a second part, this paper describes the use of dipyme to estimate the local viscasity and micropolarity in the amphiphilic NIPAM copolymer micelles below the LCST and in the polymer-rich phase that forms at the LCST. Experimental Section Materials. Spectral grade solvents were used for all spectroscopic measurements. Water was deionized with a Millipore Milli-Q water purification system. Dipyme was prepared and purified as described previously.6 n-Octy1/3-Dthioglucopyranoside (OTG) was purchased from Sigma Chemical Co. Sodium dodecyl sulfate (SDS, purum) was purchased from Fluka. Hexadecyltrimethylammonium chloride (HTAC) was obtained from Eastman Kodak Chemicals. The polymers samples were prepared by free radical polymerization in dioxane.]’ Their composition, determined by IH NMR” and expressed in N1PAM:n-alkyl chain molar units, was 240:l for PNIPAM-C,o/200, 114:l for PNIPAM-CIo/lOO, 220~1for PNIPAM-C,4/200, 108~1for PNIPAM-C14/100, 240:l for PNIPAM-Ct8/20O, and 126:l for PNIPAM-C18/ 100. The viscosity-averaged molecular weights of all the polymers were within the range 370000 f 20000. Therefore, assuming a degree of polymerization of about 3300, there are on average 33 and 16 hydrophobic groups per macromolecule in the copolymers with NIPAMmalkylacrylamide molar ratios of 1OO:l and 200:1, respectively. Instrumentation. UV spectra were measured with a Hewlett-Packard 8450A diode array spectrometer. Fluorescence spectra were recorded on a SPEX Fluorolog 212 spectrometer equipped with a DM3000F data system. The temperature of the water-jacketed cell holder was controlled with a Neslab circulating bath connected to a Neslab MTP-6 programmer. The temperature of the sample fluid was measured with a thermocouple immersed in the sample. Fluorescence Measurements. Excitation spectra were measured in the ratio mode. Emission spectra were not corrected. Slit widths were set at 3.6 (excitation) and 0.9 nm (emission). The excitation wavelength was 348 nm. The excimer to monomer ratios ( I E / I M ) were calculated as the ratio of the emission intensity a t 495 nm to that of the emission at 399 nm. The 11/13ratios were calculated as the ratio of the emission intensity at 378 nm to that of the emission a t 388 nm. Solutions were not degassed. For measurements at different temperatures samples were allowed to equilibrate for 10 min at a given temperature. The heating rate corresponded to approximately 0.2 OC min-I. Samples for Spectroscopic Analysis. Polymer stock solutions (5 g L-I) were prepared. They were kept at 5 OC to ensure complete dissolution of the polymers. Aliquots of the stock solutions were diluted to the desired concentration. Samples containing dipyme (2.6 X lo-’ mol L-]) were prepared by adding a concentrated solution of dipyme ( 5 wL, 1.3 X lo4 mol L-’in acetone) to aqueous solutions of the polymers (2.5 mL, 1.87 g L-I). ( 1 1) Ringsdorf, H.; Venzmer, J.; Winnik, F.M.Mocromolecules, in press.
Hydrophobically Modified Poly(N4sopropylacrylamides)
The Journal of Physical Chemistry, Vol. 95, NO. 6, 1991 2585
a.
b.
b.
400
450
500
550
6 I
300
320
Figure 2. Fluorescence spectra of bis(1-pyrenylmethyl) ether in aqueous = solutions of PNIPAMC18/100(1.87 g L-l); temperature 20 O C ; bXc 348 nm: (a) 2 h after sample preparation; (b) 3 days after sample preparation.
The solutions were sonicated for 10 min in an ultrasonic bath (Cole-Parmer). They were kept at room temperature in the dark until equilibrated (2-4 days). Dilute solutions of the copolymers were prepared by adding to a given volume of water aliquots of the equilibrated samples. The solutions of dipyme in surfactants were prepared by adding a concentrated solution of dipyme ( 5 pL, 1.3 X 1O4 mol L-' in acetone) to aqueous solutions of the surfactants (2.5 mL, [SDS] 8.82 X mol L-I; [HTAC] 1.64 x lW3 mol LA*;[OTG] 1.67 X mol L-I). The solutions were sonicated for 10 min in an ultrasonic bath. They were kept at room temperature in the dark until equilibrated (2 days).
Results and Discussion Artgacts. Some of the very features that make dipyme and DPyP such attractive fluorescence probes make them a source of potential artifacts when applied to micellar systems in water. Because of their low water solubility these probes are normally introduced by injecting into the system a small amount of the probe dissolved in a water-miscible organic solvent such as ethanol, acetone, or tetrahydrofuran. Under these circumstances, microcrystals form, and these are the source of the problem, since they contribute a strong excimer emission. Although this problem was mentioned by Zachariasse and co-workers in their classic description of the behavior of these probes in biological memb r a n e ~and ~ by Kano and co-workers in their work on micelles,'* no spectra showing the artifacts were presented. One can find in the recent literature applications of these probes, where spectra are reported that resemble those in which microcrystals are present. I We encountered these artifacts when we began our experiments, and were able to overcome them. Examples are shown in Figures 2 and 3. In the lower part of Figure 2 we present a classic and artifact-free emission spectrum of dipyme dissolved in a hydrophobic aggregate, here formed by the C I Schains of PNIPAMCl8/10O. The spectrum has a broad excimer band with a max(12) Kano, K.; Ishibashi, T.; Ogawa, T. J. Phys. Chem. 1983, 87, 3010. Kano. K.; Ueno, Y.;Hashimoto, S. J . Phys. Chem. 1985, 89, 3161: (13) Parthasarathy, R.; Labes, M.M.Lrrngmuir 1990, 6, 542.
340
360
WAVELENGTH (nm)
WAVELENGTH (nm )
Figure 3. Excitation spectra of bis(1-pyrenylmethyl)ether in aqueous solutions of PNIPAM-Ci8/100 (1.87 g L-I) at 20 'C for the monomer
emission (-, monitored at 378 nm) and the excimer emission (---, monitored at 490 nm; monitored at 520 nm): (a) 2 h after sample preparation; (b) 3 days after sample preparation. -a,
imum at 490 nm. It has all the features of dipyme dissolved in a viscous organic solvent. When this sample was originally prepared, it gave the fluorescence spectrum shown in the top part of Figure 2. The shoulder to the red of the excimer maximum is the first indication of the presence of microcrystals. After the sample was allowed to stand at room temperature for 3 days, the dipyme emission evolved to that shown in the lower part of Figure 2, indicating that the dipyme had become molecularly dispersed. The presence of microcrystals is made even more evident in the corresponding excitation spectra. In Figure 3 (bottom) one sees that in a properly equilibrated sample the monomer and excimer emissions have nearly identical excitation spectra. This is an essential feature of any application where one wants to use the tE/tM ratio as a measure of the conformational mobility of the probe in the system. In Figure 3 (top) one sees that the monomer (380 nm), the 490-nm emission, and the 520-nm emission have very different excitation spectra. The noisy, poorly resolved excitation spectrum of the 520-nm emission is another indication that dipyme aggregates are present. It was not possible to obtain molecularly dispersed solutions of dipyme if we attempted to dissolve the dye into very dilute solutions of the CI4- or Cls-substituted PNIPAM copolymers. Such solutions could be prepared by mixing dipyme with more concentrated solutions (e.g., 2 g L-I) and then diluting them in water once the dipyme became molecularly dispersed. This phenomenon is symptomatic of a kinetic effect on the dissolution of dipyme. Dipyme in Aqueous Solutions of Amphiphilic NtPAM Copolymers. Aqueous solutions of amphiphilic NIPAM copolymers below the LCST: Polymer concentrations of 2 g L-'correspond to alkyl chain concentrations of ca. 1.6 X IO4 mol L-I for PNIPAM-C,/lOO and 8.2 X lW5 mol L-' for PNIPAM-Cn/2OO. In these samples the extent of dipyme solubilization was monitored by measuring both their emission and excitation spectra. Dissolution was judged to be complete when (1) the excimer to monomer intensities ratio (Ze/ZM) reached a constant value and (2) when identical excitation spectra were measured for the monomer and excimer emissions. This process took several days
Winnik et al.
2586 The Journal of Physical Chemistry, Vol. 95, No. 6, 1991 TABLE I: Spectroscopic Data for Dipyme in Aqueous Solutions of Amphipbilic PNIPAM Samples and in Surfactant Micelles
IE/IM sample copolymers PN I P A M C l a / 100 PN I PAM-C i8/200 PN 1PAM-C14/ 100 PNIPAM-C14/200 surfactants6
SDS HTAC OTG
LCST," OC 25.6 30.3 23.2 28.8
20
11/13
oc
35 "C
20 o c
35 OC
0.10 f 0.01 0.13 f 0.01 0.12 f 0.01 0.18 f 0.05
0.07 f 0.01 0.06 f 0.01 0.03 f 0.01 0.10 f 0.02
1.20 1.23 1.22 1.19
1.22 1.30 1.34 1.38
0.60 f 0.01 0.88 f 0.01 0.88 f 0.01
0.63 f 0.01 1.19 f 0.01 1.39 f 0.01
1.62 1.30 1.11
1.62 1.30 1.14
,
*From ref I 1, *SDS,sodium dodecyl sulfate; HTAC, hexadedyltrimethylammonium chloride; OTG, n-octyl j3-D-thioglucopyranoside; concentration > cmc.
for solutions of PNIPAM-C14 and PNIPAM-C16. Suitable samples could not be prepared with PNIPAM or the PNIPAMClocopolymers. Dipyme microcrystals dominated the spectra even after the samples had been stored in the dark for weeks. This result demonstrates that neither PNIPAM nor its Cloderivatives are effective at solubilizing significant amounts of dipyme at room temperature. It leads us to suggest that neither polymer, below its LCST, has domains of significant hydrophobicity. Once the samples containing dipyme and the polymers carrying the C14 and pendant groups came to equilibrium, values were determined at 20 OC for the fluorescence intensity ratios 11/13and IE/IM. These data, collected in Table I, reveal the following trends: (1) The IE/IM values in the polymeric solutions are markedly lower than those measured in ionic or neutral surfactant micelles. They are of the same magnitude as those reported for dipyme in vesicles prepared from dioctadecyldimethylammonium bromide (IE/IM = 0.12,21 OC)I4 and dihexadecyl phosphate (IE/IM= 0.06, 21 OC)I4 and in phospholipid bilayer assemblies prepared from dipalmitoylphosphatidylcholine and egg phosphatidylcholine at 20 OCe6 (2) The 11/13and IE/IM ratios do not show a significant dependence either on the length of the alkyl group or on the level of alkyl group incorporation. The IE/IM results point to a rigid structure for the alkyl chain clusters formed in these solutions. It is well documentedS that bilayer vesicles possess a less fluid microenvironment in their hydrophobic interior than the core of a simple surfactant micelle. We had anticipated that the alkyl chain clusters formed within the polymer would be similar to surfactant micelles in terms of their structure and their mobility. While as yet little is known on the structure of these clusters, it is an interesting and significant observation that their microfluidity is more comparable to that of vesicles than micelles.'sJ6 A similar conclusion was drawn from fluorescence depolarization studies with DPH and perylene in these systems." The seoond set of observations, that the 11/13and IE/IM intensity ratios are essentially identical in both the C14 and the CI8polymers, indicates that the clusters formed are all very similar in their properties. The IE/IM values point to very similar microfluidity values, and the 11/13values indicate that in each of the types of clusters, the dipyme probe is located in a very similar and strongly hydrophobic environment. These do not change when the polymer concentration is changed. The polymer solutions containing di(14) Lukac, S. Photochem. Photobiol. 1982, 36, 13. Values of IE/lM depend upon how they are calculated (peak heights or peak areas) and upon instrumentation. These factors are taken into account when calculating and only OE/OMvalues can be compred from quantum yield ratios (OE/aM). one laboratory to another. Since IE/lMis proportional to a&, comparisons of different systems (e&. vesicles, micelles) are meaningful if the measurements are carried out in the same laboratory. (15) When one tries to infer absolute values of the microviscosities of surfactant micelles from examination of intramolecular excimer formation in probes of the form ArCH2XCH2Ar (X = CHI, 0,NCOCH,), the values one obtains depend upon where the probe is located in the micelle.'6 This issue would be important here if we were to try to infer the magnitude of the local viswsity from lE/IMvalues in the micellelike aggregate that exist in solutions of these polymers at temperatures below the LCST. (16) Goldenberg, M.;Emert, J.; Morawetz, H. J . Am. Chem. Soc. 1978, 100,7171. Emert, J.; Ekhrens, C.; Goldenberg, M.J . Am. Chem. Soc. 1979, 101, 771.
[DIPVME] (IO'mol C')
o,,*i
05
1p
2;o
I5
25
0.20
aos
0
0.5
A
PMIPAY-Cl~IZW PNIPAY-C~~/loQ
A
PNIPAY-C,~l+OO
1.5
1.0
2.0
POLYMER CONCENTRATION (gL-')
Figure 4. Plot of IE/lM for the emission of bis(l-pyrenylmethyl) ether in aqueous solutions of amphiphilic NIPAM copolymers as a function of polymer concentration; [n-alkyl chain]/[dipyme] = 615 (PNIPAMC,,/IOO) and = 315 (PNIPAM-C,,/ZOO) over the entire polymer concentration range. 0.15 1 PNIPAM -Cqe/200
0.05
01 20
25
30
35
TEMPERATURE ('C)
Figure 5. Plots of IE/lMfor the emission of bis(l-pyrenylmethyl) ether in aqueous solutions of amphiphilic NIPAM copolymers as a function of temperature (polymer concentration = 1.87 g L-l). For reasons of clarity, the curve corresponding to PNIPAM-C14/200 is not shown here.
pyme were diluted with water to concentrations as low as 0.01 g L-' ([dipyme] ca. 1.4 X mol L-I). The lower limit was imposed by the ability to measure reliably the emission of such low concentrations of dipyme. The only change in IE/IM detected as a function of polymer concentration (Figure 4) occurred at the lowest concentrations of probe and polymer, where the spectra are quite noisy. The fact that IE/IM remains constant over so wide a range of concentrations is consistent with the idea that cluster formation is exclusively an intramolecular, single-polymer phen0menon.l' We cannot, however, rigorously rule out contributions of interpolymer association. (17) Anionic copolymers containing long alkyl chains have been shown to form single polymer micelles with rather rigid hydrophobic clusters. See for example: Chu, D.-Y.; Thomas, J. K. Macromolecules 1987, 20, 2133. Binan-Limbel€, W.; a n a , R. Mucromolecules 1990,23, 2731. McGlade, M. J.; Randall, F. J.; Tchevrekdjian, N. Macromolecules 1987, 20, 1782.
Hydrophobically Modified Poly(N4sopropylacrylamides) 1.40 PNIPAY -Cj,/lOO
7
PNIPAY-CjaIlOO
The Journal of Physical Chemistry, Vol. 95, NO. 6, 1991 2sS7 Above the LCST, the various solutions undergo phase separation to form a polymer-rich phase and an aqueous phase containing very little polymer. Previous experiments with amphiphilic NIPAM copolymers containing both CISand pyrene substituents led to the conclusion that phase separation above the LCST triggers the disruption of the hydrophobic clusters and a more isotropic distribution of the alkyl and aryl pendant groups within the polymer-rich phase. Another pertinent observation is that above the LCST, PNIPAM is able to solubilize dyes such as perylene and dipyme, which recrystallize upon cooling the solutions below the LCST. These results all point to PNIPAM forming a polymer-rich phase above the LCST that is much less polar than the aqueous PNIPAM solution below the LCST but that is still substantially more polar than the interior of the hydrocarbon clusters formed through the association of pendant C14 or C18 substituents. The results presented in Table I are consistent with disruption of the hydrocarbon clusters at temperatures above the LCST. The differences in 11/13values observed at these temperatures then reflect differences in the mean polarity of the polymer-rich phase as it is affected by the presence of the C14 or CISchains. One feature of the data shown in Figures 5 and 6 is particularly significant. While the changes in IE/IM and 11/13for dipyme occur in the temperature region of the LCST, these changes do not correspond exactly to the changes in turbidity observed. For example, in PNIPAM-C18/200, the onset of the change in I E / I M m u r s at 28 "C, some 2 OC below the LCST (30.3 "C). On the other hand, for PNIPAM-C14/100, the I E / I M change commences at 25 "C, above the LCST (23.2 "C) for this polymer. These results emphasize that the probe is sensitive to very local phenomena and reports on changes that occur at the level of the hydrocarbon clusters themselves. Turbidity is a more global phenomenon. It describes the onset of concentration fluctuations over a spacial scale of 1000 A or larger.
Summary Dipyme is a useful fluorescent probe of hydrocarbon chain association in copolymers of N-isopropylacrylamide containing 0.5-1 mol % pendant CI4HBor ClsH3, substituents. In aqueous solution these pendant groups associate to form micellelike clusters at room temperature. Unlike common surfactant micelles, these clusters are more rigid structures that permit much less probe mobility. When heated to 35 "C (above the LCST), these polymers undergo phase separation, accompanied by disruption of the hydrocarbon clusters. A h v e the LCST, the dipyme fluorescence indicates the polymer-rich phase is a more polar and less mobile environment than that of the hydrocarbon clusters below the LCST. Acknowledgment. Financial support for this work was provided in part by the Bundesminister fur Forschung und Technologie (H.R. and J.V.), by NSERC Canada (M.A.W. and J.V.), and by the Ontario Centre for Materials Research (M.A.W.). (18) Turro, N. J.; Aikawa, M.; Yekta, A. J . Am. Chem. Soc. 1970, 101, 112.
Registry No. OTG, 85618-21-9; SDS, 151-21-3; HTAC, 112-02-7; PNIPAM-Cl,/IOO, 129674-12-0; PNIPAM-C14/100, 129674-13-1; PNIPAM-C18,129674-14-2; dipyme, 74833-81-1.
