2803
Langmuir 1991, 7, 2803-2807
Fluorescence Spectroscopy for a Cellulose Trioctadecanoate Monolayer at the Air/Water Interface Takahiro Itoh, Yoshinobu Tsujii,' Takeshi Fukuda, and Takeaki Miyamoto Institute for Chemical Research, Kyoto University, Uji, Kyoto 611, Japan
Shinzaburo Ito,' Tomoyuki Asada, and Masahide Yamamoto Department of Polymer Chemistry, Faculty of Engineering, Kyoto University, Sakyo-ku, Kyoto 606, Japan Received April 4, 1991. In Final Form: June J8, 1991 This paper describes an application of fluorescence spectroscopy for in situ studies of the structure of monolayer films spread at the airlwater interface. Cellulose trioctadecanoate, morphologically well characterized by electron microscopy, was employed as the sample polymer. The alkyl side chains were partially replaced with a fluorescentprobe, pyrenebutyryl group, and the emission properties of the pyrene units involving excimer formation were investigated as a function of surface pressure. The spectroscopic data were consistent with the film structures observed with an electron microscope. Decay curve measurements with a single photon counting technique revealed that the excited pyrenes in the monolayer are significantly quenched by oxygen in the air. This means that the molecules located just at the interface sensitivelyrespond to the change of the atmosphere. Thus, the fluorescence technique can be a unique and powerful method to study surface films in situ. Some interesting photophysical characteristicsof the surface films were disclosed.
Introduction The Langmuir-Blodgett (LB) method to fabricate ultrathin films has aroused wide interests from the scientific and technological standpoints.'I2 Recent studies have shown that a variety of polymers can be spread in a monolayer form at the air/water interface, and that the monolayers are transferable on solid substrates.3~~ Since such polymer films have many potential applicabilities, many attempts have been made to prepare well-ordered uniform films from polymeric materials.b13 (1) Kuhn, H.; MBbius, D.; Btlcher, H. In Physical Methods of Chemistry; Weissberger, A,, Rossiter, B. W., Eds.; Wiley: New York, 1972; Vol. 1, Part 3B, p 557. (2) Langmuir-Blodgett Films 4; Fukuda, K., Sugi, M., Eds.; Elsevier: London, 1989. (3) Breton, M. J. Macromol. Sci., Reo. Macromol. Chem. 1981, C21, 61.
(4) (a) Tredgold, R. H.; Winter, C. S. J. Phys. D Appl. Phys. 1982, 15, W5. (b) Tredaold, R. H.; Winter, C. S. Thin Solid Films 1983,99, 81. (c) Tredgold, R. H. Thin Solid Films 1987, 152, 223. (5) Takenaka, T.; Harada, K.; Mataumoto, M. J.Colloid Interface Sci. 1980, 73, 569. (6) (a) Kawaguchi, T.; Nakahara, H.; Fukuda, K. J. Colloid Interface Sci. 1986,104,290. (b) Kawaguchi, T.; Nakahara, H.; Fukuda, K. Thin Solid Films 1981, 133, 29. (7) (a) Watanabe, M.; Koeaka, Y.; Oguchi, K.; Sanui, K.; Ogata, N. Macromolecules 1988,21,2997. (b)Oguchi, K.; Yoden, T.; Kosaka, Y.; Watanabe, M.; Sanui, K.; Ogata, N. Thin Solid Films 1988, 161, 305. (8) Mumby, S. J.; Swalen, J. D.; Rabolt, J. F. Macromolecules 1986, 19, 1064. (9) (a) Lascheweky, A.; Ringedorf, H.; Schmidt, G.;Schneider, J. J. Am. Chem. SOC.1987,109,788. (b) Rinpdorf, H.; Schmidt, G.; Schneider, J. Thin Solid Films 1987, 152, 207. (10) (a) Schneider, J.; Ringsdorf, H.; Rabolt, J. F. Macromolecules I989,22,205. (b) Schneider, J.; Erdelen, C.; Rinpdorf, H.; Rabolt, J. F. Macromolecules 1989,22, 3475. (11)Naito, K. J. Colloid Interface Sci. 1989, 131, 218. (12) (a) Matsumoto, M.; Itoh, T.; Miyamoto, T.In Cellulosics Utilization;Inagaki, H.,Phillips, G. O., Eds.; Elsevier: London, 1989; p 151. (b) Itoh, T.; Suzuki, H.;Matsumoto, M.; Miyamoto, T. In Cellulose: Structural and Functional Aspects; Kennedy, J. F., Phillips, G. O., Williams, P. A,, Eds.; Ellis H o r w d : London, 1989; p 409. (13) (a) Ito, S.;Okubo, H.; Ohmori, S.; Yamamoto, M. Thin Solid Films 1989, 179, 445. (b) Ohmori, S.; Ito, S. Yamamoto, M.; Yonezawa, Y.; Hada, H. J.Chem. Soc., Chem. Commun. 1989,1293. (c) Ohmori, S.;Ito, S.; Yamamoto, M. Macromolecules 1990,23,4047.
The fine structure of a surface film is a critical factor determining the quality of the relevant LB films. Electron microscopy gives direct information on the fine structure.14 In a previous paper,12we applied the dark field imaging mode of electron microscopy to the surface film successfully transferred on a microscope grid. Those microscopic images offered various new insights into the characters of the surface film. The electron microscopic observation revealed that a cellulose ester derivative with relatively long alkyl side chains forms a homogeneous monolayer,12 while the monolayer of ita low molecular weight homologue, cellobiose ester derivative, is inhomogeneous with many holes.1s This indicates that the macromolecularity favors the formation of homogeneous monolayer films. However, an in situ observation of the film at the air/ water interface is impossible by electron microscopy. It is also difficult by electron microscopy to obtain molecularlevel information on a surface film, e.g., the conformation and dynamics of the polymer on the water surface. Recent instrumental developments have made in situ measurements of surface films possible, e.g., surface dynamic light scattering to study surface viscoelastic properties of polymer films,16 and high quality Raman spectroscopy to study molecular orientations in compressed monolayers.'* In this work, we have applied the fluorescence method to monolayers of cellulose derivatives.6J2 The extremely high sensitivity of light detection enables us to obtain sufficient signals, even when the specimen is an ultrathin film like a monolayer on the water surface. Using samples whose morphology has been wellcharacterized by electron microscopy, we have tried to obtain the fluorescence characteristics of the pyrenelabeled surface films. The pyrenyl chromophore is very useful as a fluorescent probe for studying the aggregation of polymers." ~~
~~
(14) Ries, H. E.; Walker, D. C. J. Colloid Sci. 1961, 16, 361. (15) Itoh, T.; Mataunoto, M.; Suzuki, H.; Miyamoto, T. Bull. Inet. Chem. Res., Kyoto Unio. 1990,68, 53. (16) Yoo, K.; Yu, H. Macromolecules 1989,22, 4019.
0 1991 American Chemical Society
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2804 Langmuir, Vol. 7, No. 11, 1991 Table I. Degree of Substitution (DS), DS of Pyrene Units and (DSpy),Molecular Weight per Glucose Unit (M,), Limiting Areas of Synthesized Polymers sample DSO DSeO M,, glucose unit limiting area, m2mgl 961.5 0.45 PO 3.0 0 961.7 0.44 P1 3.0 0.038 962.3 0.44 P2 3.0 0.21 P3 0
3.0
1.1
965.9
0.52
DS and DSp, are the number of total substituenta and pyrene-
butyryl groups per glucose unit, respectively.
Experimental Section Materials. Cellulose trioctadecanoates were prepared from regenerated cellulose according to the procedure of Malm et Details have been reported elsewhere.12 The cellulose esters partially substituted with octadecanoyl groups were labeled with pyrene chromophores by a further reaction with 1-pyrenebutyric acid (Aldrich) after Hayashi et al.20 An IR analysis showed that the total degree of substitution per glucose ring (DS)was 3.0 for all samples.21 That is, the three hydroxyl groups in a glucose ring are fully substituted by octadecanoyl and pyrenebutyryl groups. The DS value of pyrene unit per glucose ring, DSb, was determined by both UV absorption and NMR spectroscopies. The characteristics of the prepared polymersare listed in Table I. Preparation of Monolayer. A dilute solution of the sample polymer (0.01wt %) in benzene (Dojin, spectroscopic grade) was spread on the surface of pure water (293 K) in a Teflon-coated rectangular trough (15 cm x 60 cm, Kenkosha Model SI-2) equipped with a Wilhelmy-type film balance. The water used in the subphase was ion-exchanged,distilled, and passed through a water purification system (Barnstead Nanopure 11). The monolayer was compressed with a Teflon bar at a rate of 5-10 mm min-l. For the sake of comparison, built-up films were prepared which consisted of a bilayer of a sample polymer sandwiched between two protecting bilayers of polymer PO. These films were transferred on a quartz plate at a pressure of 10 mN m-l by the horizontal lifting method.n.23 Fluorescence Measurements. Two quartz optical fibers of 3 mm in diameter were set on the middle of the trough. One, the excitation light guide, was arranged at an angle of 45O to the surface plane. Excitation light from a Xenon flash lamp (Hamamatsu L2359 and C2190,5 w) was focused on the entrance end of the excitation fiber. The light of wavelengths between 250 and 340 nm was obtained by a suitable combination of optical filters. The other fiber, used for the observation of fluorescence from the surface film, was set normal to, and 5 mm above, the water surface. One end of the emission fiber was connected to the sample holder of a fluorescence spectrophotometbr (Hitachi Model 850). Fluorescencespectra were recorded through a cutoff filter of 350 nm to avoid the intense scattering of the excitation light. Measurement of fluorescence decay curves were performed by a single photon counting s y ~ t e m . ~ 'A, ~pulsed excitation light (315 nm) was obtained with a Spectra-Physics picosecond synchronously pumped, mode-locked, cavity-dumped dye laser, (17)(a)Winnik,F.M.; Winnik,M. A.;Tazuke,S. J.Phys. Chem. 1987, 91, 594. (b) Winnik, F. M.; Winnik, M. A.; Tazuke, S.; Ober, C . K. Macromolecules 1987,20,38. (c) Wilhelm, M.; Zhao, C.-L.; Wmg, Y.; Xu, R.; Winnik, M. A.; Mura, J.-L.; Riess, G.;Croucher, M. D. Macromolecules 1991,24,1033. (18)Kawai, T.; Umemura, J.; Takenaka, T. Chem. Phys. Lett. 1989, 162. ~- 243. ~
h9)Malm,C.J.; Mench, J. W.; Kendall, D. L.; Hiatt, G. D. Id. Eng. Chem. 1961,43,684. (20)Shimizu, Y.; Hayashi, J. Cellul. Chem. Technol. 1989,23,861. (21)Yamagishi, T.; Fukuda, T.; Miyamoto, T.; Takashina.. Y.:. Watanabe, J. Liq. Cryst., in press. (22) Kamata, T.; Umemura, J.;Takenaka, T. Chem. Lett. 1988,1231. (23)Itoh, T.; Tsujii, Y.; Fukuda, T.; Miyamoto, T.; Ito, S.; Yamamoto, M. Manuscript in preparation. (24) Yamazaki, I.; Tamai, N.; Kume, H.; Tsuchiya, H.; Oba, K. Reu. Sci. Instrum. 1986,56,1187. (25)(a) Ito, S.;Takami, K.; Yamamoto,M. Makromol. Chem.,Rapid Commun. 1989,10,79.(b) Ito,S.;Takami, K.;Tsujii, Y.; Yamamoto, M. Macromolecules 1990,23,2666.
Area
( mzmg-')
Figure 1. Surface pressure-area isotherms of pyrene-labeled cellulose derivatives at 293 K. T h e letters indicate the points observed by the electron microscope. and was introduced to the entrance terminal of the excitation light guide. The emission through the light guide was detected with a photomultiplier (Hamamatsu R3234). Time-correlated pulses were obtained by conventional Ortec photon counting electronics and accumulated on a multichannel analyzer (Norland IT-5300). The full width at half-maximum of the overall excitation pulse was 500 ps. Fluorescent molecules produced in a monolayer will be liable to be quenched by oxygen. When necessary, nitrogen gas was gently flowed over the water surface to prevent the quenching. Electron Microscopy. Specimens for electron microscopy were prepared by lifting the surface film horizontally on a microscope grid covered with a carbon supporting film. The surface film thus transferred on the carbon film was observed by applying the dark-field imaging mode of a J E M 200-CX electron microscope (JEOL)with a direct magnification of 2800 times. In the dark-field imaging mode, the main electron beam is blocked by an objective aperture, and only the electrons scattered from the monolayer are focused on a screen. This operation enables ustoobserve themonolayerwithasufficientcontrast. Thedetails were described previously.12+2s
Results and Discussion Figure 1shows the surface pressure-area isotherms of samples PO,P1,P2,and P3. The isotherms begin to rise at around 0.6 m2 m g l and then increase steeply with further compression. The cohesive force between the long alkyl chains gives rise to a "condensed-type" monolayer. While the monolayer of P3 is collapsed at 20 mN m-l and ita limiting area slightly increases (from 0.45 to 0.52 m2 mg-l), P1 and P2 show a similar isotherm to that of PO. This indicates that the introduction of pyrene units as much as 7 94 (by side-chain mole base) does not essentially alter the monolayer characteristics. Electron microscopy provides direct information on the morphology of surface films. Figure 2 shows dark-field images of the PO monolayer transferred at 0, 10, and 30 mN m-l, respectively: The observed points are indicated by a, b, and c in Figure 1. At point a, the surface film takes the so-called "island structure"; a heterogeneous film consisting of aggregates of nonuniform size can be seen. With increasing compression, the film structure becomes more and more uniform and a perfectly homogeneous monolayer is obtained at 10 mN m-l (point b). Even (26)Uyeda, N.;Takenaka, T.; Aoyama, K.; Mataumoto,M.; Fujiyoshi, Y.Nature 1987,327,319.
Cellulose Trioctadecanoate Monolayer
Langmuir, Vol. 7, No. 11,1991 2805
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UK)
Wavelength (nm)
Figure 3. Fluorescence spectra of surface films prepared from (A) P1,(B) P2,and (C) P3. The spectra were recorded with a bandwidth of 5 nm. The broken lines are the spectra observed for the built-up films on quartz plates.
C
Figure 2. Dark-field images of the cellulose trioctadecanoate monolayertransferred at the surface pressures of (a) 0 mN m-l, (b) 10 mN m-l, and (c) 30 mN m-l.
though the isotherm goes on increasing with further compression, the monolayer at point c has some stripes perhaps due to a folding of the monolayer. Thus, the structure has already changed to a partially collapsed form at 30 mN m-l. On the basis of the mentioned morphological knowledge, we investigated the fluorescence characteristics of the monolayer in situ at the air/water interface. Figure 3 shows the fluorescence spectra of the monolayers prepared with samples P1,P2,and P3 at a surface pressure of 10 mN m-1 (point b in Figure 1). Sample P1, which has the lowest pyrene content, shows only monomer emission at 377 and 397 nm (Figure 3A). With an increase in pyrene content, excimer emission begins to appear at
around 480 nm. The excimer is formed between a pair of pyrene chromophores which happen to take a parallel arrangement of aromatic rings. The chance of such a geometrical arrangement increases with pyrene concentration. In a monolayer of a polymeric material, a statistically random distribution of chromophoric units along the chain results in a relatively small number of excimer sites compared with that in the LB films of longchain fatty acids, in which pyrene units often take an aggregated s t r u c t ~ r e . ~In~ this , ~ ~system, the excitation spectra of the films built up on a quartz plate indicate no aggregation of pyrenyl groups in the ground state.23 The DSp, value of P3 is large enough that most of the pyrene units have a pyrene moiety at the adjacent position, in any case, and the excimer emission predominates (Figure 3C). Notably, these fluorescence spectra on the water surface are similar to those of the built-up films, as shown by the broken lines in Figure 3. The deposition was carried out at the same surface pressure of 10 mN m-l. This indicates that the molecular arrangement formed at the air/water surface is maintained in the transferred films. Figure 4 shows the fluorescence spectra of the P2 film as a function of the surface pressures, in which the intensity is normalized at the maximum peak height. The spectra are almost independent of surface pressures ranging from 0 to 10 mN m-l. This result can be compared to the microscope images, showing that the surface film takes an island structure at 0 mN m-l, and then forms a homogeneous monolayer at 10 mN m-l. The electron microscopy also demonstrated that the homogeneous monolayer has a thickness of 2.4 nm,% consistently with the molecular model in which the alkyl chains stand normal to the water surface in a closely packed structure. Surprisingly, the film thickness at 0 mN m-l was measured to be 1.7 nm, not so much different from that of the homogeneousmonolayer. This shows that, at 0 mN m-l, the alkyl chains in the "island" already take a similar conformation to that (27) Yamazaki, I.; Tamai, N.;Yamazaki, T. J. Phys. Chem. 1987,92, 3572. (28) Miyamoto, T.; Itoh, T. Koubunshi Kakou 1989,38,225.
Ztoh et al.
2806 Langmuir, Vol. 7, No.11, 1991
5W
100
Wavelength (nm)
I
Figure 4. Spectral changes with the compreesion of the P2 monolayer: (a) 0 mN m-l; (b) 10 mN m-l; (c) collapsed film.
0
A
I
I
1 2 3 Surface Concentration (mg m-21
4
0
Figure 6. Fluorescence intensity at 377 nm and surface pressure 88 a function of surface concentration of the P1 polymer: ( 0 )in nitrogen atmosphere; (0)in the air.
"I 0
100
200
Channel Number ( 2.6 nwch 1
Figure 5. Fluorescence decay curves of the P1 monolayer: (a) 1mN m-1; (b) collapsed film; (a') the same as curve a, but under a nitrogen atmosphere.
in the monolayer. The fluorescence measurement also indicates that the microscopic environment around the pyrene probe remains unchanged throughout the compression process between 0 and 10 mN m-l. The excimer emission, however, becomes stronger at elevated pressures. In particular, the intensity from the collapsed film is much higher than that from the uncollapsed one. An increase in excimer emission intensity would mean an increase in excimer site density. In a regular multilayer film, no excimer is expected to be formed between adjacent layers, because an octadecanoyl group is much longer than a pyrenebutyryl Therefore, the monolayer presumably collapses into a rather irregular structure, not into a regular multilayer structure. This observation suggests that the excimer emission can be a useful measure of monolayer structural orders. Decay curve measurements gave us more direct information on the dissipation processes of the excited pyrenes. Despite the weak intensity from a surface film, a sufficient number of photons could be detected with a picosecond laser system. Figure 5shows the decay curves of P1. Since this sample formed no excimer, we had expected it possible to estimate the intrinsic lifetime of the pyrene chromophore from the slope of a single exponentialdecay curve. However, the decay curve observed at 1 mN rapidly fell at an early stage after excitation (the decay a in Figure 5). This indicates that there is a certain quenching process for the excited pyrenes in the monolayer system. Up to collapse, the decay curve showed little dependence on the surface pressure. Furthermore, with a further compression extending to the collapsing region, the decay time tended to become longer than that of the monolayer. Measurements under nitrogen atmosphere have disclosed the origin of the quenching process: When agentle stream of nitrogen gas was introduced on the surface film, the decay time
drastically increased (the decay a' in Figure 5); curves a and a' are for the same monolayer from P1 at 1mN m-l. Evidently, oxygen in the air quenches the excited state of pyrene chromophores. In fluid solution, the fluorescence quenching of aromatic compounds has been well k n o w n . ~ ~ Despite the long alkyl chains surrounding the pyrene units, their shieldingeffect against oxygen is insufficient. When the monolayer is collapsed by compression, the pyrene units are effectivelyscreened from oxygen molecules owing to the folded film structure, thus decreasing the relative concentrationof pyrene unitson the film surface. A simii behavior of the decay curves was observed for the P2 film. Thus, excited molecules in a monolayer sensitivelyrespond to changes of surroundings. Such dynamic quenching is a minor process for a thicker film in which the surface area is relatively small for the volume. This is one characteristicof photophysicalprocesses in ultrathin films. To make quantitative discussion on the fluorescence intensity, measurements should be made under a nitrogen atmosphere. Figure 6 shows the fluorescence intensity at 377 nm (monomer emission) versus surface concentration of sample P1. Below 2.0 mg m-2, the data were seriously scattered, depending on the position of observation. Reliable data could be obtained only at concentrations higher than 2.0 mg m-2. This value corresponds to the critical concentration at which the surface pressure begins to show a steep rise. In the range of 2.0-2.7 mg m-2, the intensity is proportional to the surface concentration, as shown by the solid line. This means that under a nitrogen atmosphere there is no extrinsic dissipation process for the excited pyrene moleculesand the fluorescenceintensity is given simplyby the totalnumber of fluorescent molecules present (within the scope of the optical fiber). With a further compression beyond 2.7 mg m-2, the plots deviate from the solid line. Referring to the surface pressure, we may ascribe this deviation to collapsing of the monolayer. The collapse probably proceeds from weak positions of the film. Additional measurement in fact showed that the intensity is higher around the moving bar than near the center of the trough, indicating that collapsing of the film proceeds predominantly around the bar. Therefore, in the collapsing region, the average concentration indicated by the abscissa scale in the figure becomes higher than the effective concentration at the center of trough. (29)Birke, J. B. Photophysics of Aromatic Molecules; Wdey-Interscience: London, 1970; Chapter 10. (30) (a) Bowen, E. J. Trans. Faraday SOC.1954,50,97. (b) Ware, W. R. J. Phys. Chem. 1962,66, 455. (c) Stevens, B.; Dubois, J. T.Trans. FaradaySoc. 1963,59,2813.
Cellulose Trioctadecanoate Monolayer
A similar measurement carried out in the air did not show such a linear relationship between intensity and surface concentration. Quenching of the excited pyrenes by oxygen drastically weakens the fluorescence intensity. In the monolayer region the intensity is decreased by a factor of 1 1 3 to I / 4 , while in the collapsed region, by a factor of about l / 2 . The relatively larger intensity in the collapsed region indicates a significant shielding effect due to a folded structure of the film. This is consistent with the relatively long lifetimes observed for the collapsed films (see Figure 5). Concluding Remarks The structure of cellulose trioctadecanoate monolayers formed at the air/water interface was studied in situ by fluorescence spectroscopy. The results were consistent with the film morphology observed by electron microscopy. This work also has provided new pieces of information on
Langmuir, Vol. 7, No. 11, 1991 2807 the fluorescence characteristics of the monolayer. Above all, it has revealed that the fluorescenceresponse to oxygen is particularly sensitive, which affords new means to study the packing state around the probe molecules. Thus, fluorescence spectroscopy provides a unique and powerful method for studying monolayers in situ.
Acknowledgment. We thank Dr. M. Matsumoto, Institute for Chemical Research, Kyoto University, for the electron microscopic observation. This work was partially supported by a Grant-in-Aid for Encouragement of Young Scientists (No. 02750648), a Grant-in-Aid for Scientific Research (No. 01550692) and a Grant-in-Aid for Scientific Research on Priority Areas, New Functionality Materials-Design, Preparation and Control (No. 02205071), from the Ministry of Education, Science and Culture of Japan. Registry No. Cellulose trioctadecanoate, 39320-20-2.
