Macromolecules 1994,27, 77-86
77
Lateral Diffusion Study of Amphiphiles in Air-Water Monolayer Films of Polymerizable Surfactants F r a n k Caruso, F r a n z Grieser, a n d Peter J. Thistlethwaite' School of Chemistry, University of Melbourne, Parkville, Victoria 3052, Australia
D. Neil Furlong CSIRO, Division of Chemicals and Polymers, Private Bag 10, Clayton, Victoria 3168, Australia Received August 2, 1993; Revised Manuscript Received October 8, 1 9 9 9
ABSTRACT: The behavior of the chromophore N-[(l-pyrenylsulfonyl)dipalmitoyll-L-a-phosphatidylethanolamine (pyrene-DPPE) embedded in unpolymerized and polymerized air-water monolayers of 10,12nonacosadiynoic acid (diacetylene 16-8),octadecyl methacrylate (ODMA) and (octadecyldimethy1ammonio)ethyl methacrylate bromide (ODAEMA), has been investigated by using surface pressure-area isotherm measurements, and steady-state and time-resolved fluorescence spectroscopy. Steady-state fluorescence measurements revealed pyrene-DPPE to be homogeneously distributed in the diacetylene 16-8 monolayer only in the liquid-expanded (LE)phase on an alkaline subphase. The lateral diffusion coefficientsdetermined for the unpolymerized diacetylene 16-8 monolayer from steady-state measurements are in good agreement with literature values. Polymerization of the diacetylene 16-8 monolayer resulted in the formation of a liquid-condensed phase film and aggregation of pyrene-DPPE in the monolayer film, thus preventing a fluorescence quenching study. Pyrene-DPPE was found to be aggregated in both the monomer and polymer monolayersof ODMA. A derivative of ODMA incorporating a quaternary ammonium group (ODAEMA)was synthesized. ODAEMA showed LE isothermal behavior and gave a homogeneous distribution of pyreneDPPE in the monomer monolayer film. The lateral diffusion coefficients obtained in the unpolymerized ODAEMA monolayer film were found to decrease with increasing surface pressure, reflecting a decrease in monolayer fluidity with compression. In the polymerized film of ODAEMA, nonexponential fluorescence decay behavior of pyrene-DPPE was observed. The fluidity of the polymerized ODAEMA monolayer is estimated to be at least 1 order of magnitude less than that for the corresponding monomer monolayer.
1. Introduction In recent years there has been considerable interest in monolayers, bilayers, and Langmuir-Blodgett (LB) multilayers both as biomembrane models and because of their potential technological applications in the field of microelectronics.1-20 A concern common to both areas is that of film stability, and because of this, a great deal of attention has been focused on stabilizing organizedsystems of amphiphilic molecules via polymerization.11-20 Higher mechanical and/or better chemical and thermal stability can be achieved by polymerization of the reactive groups within the amphiphile. A wide variety of polymerizable amphiphiles have been synthesized and extensively investigated by different methods a t the air-water interface and in LB films.11*20 Polymerizable groups can be located in the head group and/or the alkyl chains of amphiphilic molecules, and include diene, diyne (diacetylene), acrylate, and methacrylate groups which can be polymerized upon exposure to UV light or by radical initiators. The position of the polymerizable group within the amphiphiles influences the phase transition and fluidity characteristics of the monomeric and polymerized ~ystems.~3J~-20 We have been particularly interested in the characterization of air-water monolayers by the measurement of the mutual lateral diffusion coefficient of suitable probequencher The fluorescencequenching technique employed is an alternative to the methods of fluorescence recovery after photobleaching (FRAPI2G23and pyrene excimer formation.24-26 The advantages of the fluorescence quenching approach have been described in recent papers.Sl0
* To whom correspondence should be addressed.
a Abstract
published in Aduance ACS Abstracts,
November 15,
1993.
0024-929719412227-0077$04.50/0
The fluorescence quenching approach we have used is, however, dependent on there being a homogeneous distribution of probe and quencher within the matrix monolayer under investigation. Previous experience&'" has led us to formulate some qualitative empirical rules for predicting the miscibility of amphiphiles in monolayers and has suggested that complete miscibility of probe, quencher, and matrix is likely to be the exception rather than the rule. Accordingly, we have investigated the monolayer properties of three polymerizable amphiphiles in order to determine the mixing properties of the probequencher-matrix systems and to determine the extent to which the fluorescencequenching technique can be applied to yield reliable lateral diffusion coefficient data. Diyne and methacrylate amphiphiles, with polymerizable groups in the alkyl chain and the polar head group of the amphiphile, respectively, were employed as the monolayer matrices. Steady-state and time-resolved fluorescence spectroscopy were used to study the effect of polymerization on the homogeneity of distribution of the lipoidal probe pyrene-DPPE in the monolayer film. 2. Experimental Section 2.1. Materials. The chromophore N-[(l-pyrenylsulfony1)(pyrene-DPPE)and dipalmitoyl]-L-a-phosphatidylethanolamine the quencher 4-(N,N-dimethyl-N-hexadecylammonio)-2,2,6,6tetramethylpiperidinyl-1-oxy1iodide (CAT-16) were both purchased from Molecular Probes Inc. The alkylamine quencher, octadecyldimethylamine (ODDMA),was obtained from Pfaltz and Bauer Inc. All were used without further purification. The structures of the three different polymerizableamphiphilic compounds investigated are shown in Figure 1. lO,l2-Nonacosadiynoicacid (diacetylene 16-8) was obtained from Tokyo Kasei and was purified by recrystallization from diethyl ether (Ajax;AR grade) immediately before use to remove any polymer residue. 0 1994 American Chemical Society
Macromolecules. Vol. 27, No. I, 1994
78 Caruso et al.
layers were conducted on a 57.9 x 13.5 cm2 poly(tetrafluoroethylene) (PTFE) Langmuir trough, with a PTFE barrier, driven at a compressionrate of 0.03 nm2molecule-1 min-l. A 59.7 X 16.5 cm2 PTFE Langmuir trough with a compression rate of 0.05 nm2 molecule-’ min-l and a 47.0 X 15.0 cm2PTFE Langmuir trough (KSV-2200) (compression rate 0.05 nm2 molecule-’ min-‘) equipped with a quartz window in the bottom were used for steady-state monolayer fluorescence experiments. The KSVI 0 2200 trough was also used for time-resolved monolayer fluores0 I I cence experiments. All surface pressurearea measurements were made by the Wilhelmy hanging plate method.8 For PA measurements of the pure amphiphiles and steady-state monolayer fluorescence measurements, a 4.3-cm mica plate suspended from a Shinkoh 2-g capacity strain gauge was used. The apparent changes in 7 1 2 weight with monolayer compression were converted to voltages Figure 1. Structures of the three polymerizable amphiphilic by the strain gauge and recorded on an Apple Macintosh PC. A molecules. (1) 10,12-nonacosadiynoic acid (diacetylene 16-8); 3.0-cm roughened platinum plate suspended from a Cahn (2) Octadecyl methacrylate (ODMA);(3) (octadecyldimethylammicrobalance was used in the time-resolved monolayer fluoresmonio)ethyl methacrylate bromide (ODAEMA). cence measurements. The change in voltage from the microbalance was monitored by the KSV-2200 trough controller and Octadecyl methacrylate (ODMA)was purchased from Tokyo recorded on an IBM PC with software from KSV, Helsinki. Kasei and was used as received. Poly(octadecy1methacrylate) Experiments were initiated by filling the trough with the (PODMA) was obtained from Monomer-Polymer and Dajac appropriate subphase. Approximately10’’ molecules from 1mM Laboratories, Inc. The PODMA sample had a weight average chloroformsolutions mixed to the desired ratio were spread onto molecular weight of 175OOO and a number average molecular the subphase, using a 100-pL SGE syringe. The solvent was weight of 47 PODMA was purified by repeated precipithen allowed to evaporate for 10 min, after which the monolayer tation from the toluene solution by the addition of excess was compressed as desired. methanol. The precipitated polymer was freeze dried for 12 h, 2.3. Steady-State Monolayer Fluorescence Measurements. followed by vacuum dryingat room temperature for 72hto remove Steady-state fluorescencemeasurements were performed on two residual methanol. different experimental setups. The first setup, located at The The monomer (octadecy1dimethylammonio)ethyl methacryUniversity of Melbourne, employed a Perkin-Elmer LS-5 lumilate bromide (ODAEMA)was prepared by a modification of the nescence spectrophotometer, and details of the completesystem method of Nagai et a1.%(N~-Dimethy~o)ethylmethacrylate have been given previously.”’O Briefly, two silica fiber optic (Aldrich Chemical Co.) (2.9 g, 0.010 mol) was quaternized by bundles were used to transfer the exciting light and the reacting with freshly distilled l-brommtadecane (Fluka) (7.3g, fluorescenceto and from the monolayer. Since the fluorescence 0.011 mol), in acetone (20 mL), at room temperature for 7 days signal from the monolayer was small, it was necessary to subtract in the presence of 2000ppm p-methoxyphenol (inhibitor). After the background signal due to scatter from the subphase of the removal of acetone, anhydrous ether was added, and the exciting light and/or fluorescence from the PTFE. The backprecipitated solid was filtered and then repeatedly recrystallized ground signal, monitored at the same emission wavelength as from ethyl acetate. The purified product was obtained am a white that of pyrene-DPPE, was recorded for 10 min, averaged, and powder, and ita structurewas established by ‘H NMR. lH NMR then subtracted from the fluorescencesignalwhen the monolayer spectra were recorded as solutions in CDCls on a Bruker AC 200 was present. Fluorescence intensity curves as a function of spectrometer operating at 200 MHz with use of the solvent proton monolayer compression were obtained using this experimental signal as reference. The resonances were assigned as follows: 6 setup. Thus the fluorescence intensities at various surface 6.08 (m, lH, proton tram to a methyl group), 5.61 (m, lH, proton pressures (I values) were obtained for each of the monolayersof cis to methyl group), 4.61 (m, 2H, N-CHz-CHd), 4.09 (m, 2H, different quencher concentration. The l o value at a givensurface N-CH*-CH2-0), 3.55 (m, 2H, CH~(CH~)&H&HZ-N), 3.46 (m, pressure was obtained from a monolayer containing only pyrene6H, N-CHs), 1.91 (bs, 3H, OC(O)C(CHdCH3), 1.65 (m, 2H, DPPE in the matrix monolayers. Addition of the quenchers CHs(CH2)l&HZCHrN), 1.10-1.30 (m,30H,CH~(CH~)I~CH~CHZ-CAT-16 and ODDMA to the monolayer films of the matrices did N), 0.93 (bt, 3H, C H ~ ( C H Z ) ~ ~ C H ~ C H ~ N ) . not significantly affect the packing of the molecules, and in all Polymerization of the monomer ODAEMA was accomplished cases, with the exception of the ODAEMA matrix, the *-A by catalytic polymerization induced by 2,2’-azobis[isobutyroniisotherms were within f0.02 nm2 molecule-’ of each other in trile] (AIBN). ODAEMA (1.2 g, 2.5 mmol) and AIBN (8.0 mg, both the absence and presence of quencher. Addition of CAT0.05 mmol, 2 mol %) were placed in a polymerization flask to 16 quencher to the monolayer film of ODAEMA caused an which HzO (90mL) was added. The mixture was agitated to increase in area, at a particular surface pressure, with the F A obtain a homogeneous solution. The flask was degassed by the isotherms differing in average area per molecule by ca. 0.07 nm2 freeze-thaw technique, sealed in a nitrogen atmosphere, and at the high end of the concentration range of CAT-16(25mol 5% ). placed in a constant temperature bath at 70 “C for 17 h. The This resulted in a different surface concentration of pyrenepolymer was freeze dried for 24 h, followed by vacuum drying for DPPE moleculesfor the reference monolayer and for monolayers 3 h to give a white powder (1.0 9). Polymerization of ODAEMA containing quencher. To account for this variation in average was determined by 1H NMR spectra by following the decrease area per molecule, and for the dependence of the fluorescence in intensity of the vinyl protons (6 6.08 and 5.61 ppm), and the lifetime of pyrene-DPPE on the average area per molecule, it completionof polymerizationwas indicated by the disappearance was necessary to apply a correction for the pyrene-DPPE/CAT16/ODAEMAsystem in the followingmanner. All fluorescence of the vinyl proton peaks. Further evidence for polymerization was the line broadening of peaks observed in the ‘H NMR intensities were scaled to constant surface concentration of pyrene-DPPE and then further corrected for the variation of spectrum. fluorescencelifetime of pyrene-DPPE with the average area per Sodium perchlorate (AR grade) was purchased from Merck molecule (see later). and sodium hydroxide (AR grade) from Ajax Chemicals. All nonaqueous solvents were spectroscopicgrade and were obtained In the second setup (University of Uppsala), a silica lens and from Ajax Chemicals or Merck. “Milli-Q” water was used to mirrors were used to focus the excitation light from a pulsed prepare the subphase (conductivity
