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Anal. Chem. 1989, 61, 2165-2188
Application of a Quartz-Crystal Microbalance for Detection of Phase Transitions in Liquid Crystals and Lipid Multibilayers Yoshio Okahata* and Hiroshi Ebato Department of Polymer Chemistry, Tokyo Institute of Technology, Ookayama, Meguro-ku, Tokyo 152, Japan
Resonance frequency Increased abruptly at the phase transltlon temperature ( T , ) from solld to llquid crystalllne state of llquld crystals (LCs) or llpld multlbllayer fllms cast on a quartz-crystal mlcrobalance ( O W ) electrode when the amblent temperature Increased gradually. The large frequency enhancement at the T , was observed In the case of smectlc LC-coated OCM compared with those of nematic and cholesterlc LC-coated QCM. Although the frequency enhancement at the T , of the LCtoated QCM was observed Independent of medla such as water and alr phases, the frequency of the llpld multlbllayertoated QCM Increased abrum at the T, only In a water phase, but not In an ak phase. Frequency enhancements at T, can be explained by sllpplng between layered structures of smectic LC and llpld multlbllayer In the fluid llquld crystalllne state above the T,. I n the case of llpld multlbllayers, the penetration of water Into lnterlayers (swelling) occurs at the T, and then the frequency Increase Is observed due to sllpplng between hydrated and fluld layers only In aqueous phases.
Piezoelectric quartz plates are known as a microbalance to detect the amount of substances deposited on the electrode of the quartz from the frequency decrease ( I ) . Natural protein-coated quartz-crystal microbalances (QCMs) have been studied as immunoassay procedures using antigen-antibody reactions (2-5). We have reported that the lipid multibilayer-coated QCM can detect selective adsorptions of various bitter or odor substances to lipid matrixes both in air and water phases (6-8). When the QCM was used as a substrate of Langmuir-Blodgett (LB) techniques in which lipid monolayers were transferred on the QCM plate from a water surface, it was found from the frequency change that LB films were transferred on the substrate with water which gradually evaporated through layers of the films when exposed to air (9). The fundamental principle in these studies involves frequency changes due to the surface mass increase or decrease on the QCM by deposition or release. In this paper, we would like to report that a new application of the QCM for detection of phase transitions from solid to liquid crystalline state of liquid crystals (LCs) and lipid multibilayer films cast on the electrode. Smectic (PCB), nematic (EBBA), and cholesteric (ChD) liquid crystal compounds, and 2C16PE phospholipid and polymeric 2ClsN+2C1/PSS- multibilayer films were cast on the electrode of the QCM (see Figure 1). The frequency of the QCM abruptly increased a t the phase transition temperature (TJ of the coating materials when the temperature was raised gradually in air or water phase. Crane and Fischer have pointed out that the resonance frequency of the shear node quartz crystal is influenced by the fluidity of the coating polymer in air (IO). Muramatsu and co-workers reported that the frequency shift was observed due to the change of a deficit resistance of a *To whom all correspondence should be addressed. 0003-2700/89/0361-2185$01.50/0
piezoelectric quartz when the quartz was in contact with high-viscosity liquids ( 1I).
EXPERIMENTAL SECTION Materials. Thermotropic liquid crystal compounds, p pentyloxy-p'-cyanobiphenyl (PCB, k s, 43 "C; s i, 68 "C), N-(p-ethoxybenzy1idene)-p'-butylaniline (EBBA, k n, 36 O C , n i: 80 "C), and cholesteryl decanoate (ChD, k ch, 82 "C; ch i, 91 "C), were purchased for analytical grade chemcials and used without further purification. Preparations of synthetic phospholipids, 1,3-dihexadecylglycerol-2-phosphoethanolamine(2Cl$'E), and polymeric bilayer-forming amphiphiles of dioctadecyldimethylammonium poly(styrenesulfonate) (2ClSN+2C1/PSS-)were reported elsewhere (6-8, 13). Chloroform solutions of liquid crystals (LCs) and bilayer-forming amphiphiles were cast on the electrode of both sides of the quartz, dried in air, and kept at 10 "C for a day. In the case of lipid multibilayers, the lipid coated QCM was aged in hot water at 60 "C for 30 min and then kept at room temperature for a day. PCB, EBBA, and ChD on the QCM were confirmed to show smectic, nematic, and cholesteric liquid crystalline states at the respective phase transition temperature (Tc). X-ray diffraction analyses showed that 2C16PE and 2C18Nf2Cl/PSS- amphiphiles form extended multilamellar structures of a lipid bilayer (3.3-3.9 nm spacing) parallel to the film plane (6-8,13). The multibilayer films on the QCM showed a sharp endothermic peak at 53 OC for 2ClePE and 42 "C for 2C18N+2C1/PSS-multibilayers from differential scanning calorimetry (DSC) in a water phase, which mean the phase transition from solid to liquid crystalline state of multibilayer films (6-8, 13). The polyion complex multibilayer film was used because it was physically stable and did not peel from the QCM plate even at harsh conditions in aqueous solutions. Apparatus. The experimentalapparatus comprised at 9-MHz, AT-cut quartz crystal plate (8 mm diameter) on both sides of which 20 mm2 of Ag electrodes had been deposited, and a homemade oscillator designed to drive the QCM at its resonance frequency (6-9). The QCM was driven at 5-V dc, and the frequency of the vibrating quartz was measured by an Iwatsu frequency counter (SC 7201 Model) attached to the microcomputer system (NEC,PC 8801 Model). The following equation has been obtained for the AT-cut shear mode QCM ( I ) :
--
-
--
where A F is the measured frequency shift (Hz), Fothe parent frequency of QCM (9 X lo6 Hz), Am the mass change (g), A the electrode area (0.20 cm2),pq the density of quartz (2.65 g c m 9 , and p the shear modulus (2.95 X 10" dyn cm-2). Calibration of the QbM used in our experimentsshowed that a frequencychange of 1Hz corresponded to a mass increase of 1.05 & 0.01 ng on the electrode of QCM (6-9) Am = -(1.05 & 0.01) X 10+ A F (2) LC compounds (smectic PCB, nematic EBBA, and cholesteric ChD) or lipid amphiphiles (2C16PEand 2ClsN+2C1/PSS-)were cast to be 15 f 2 pg (0.5 i 0.1 pm thick) on electrodes (20 X 2 mm2)of both sides of the QCM. The decrease of vibration frequency (14.3 f 0.1 kHz) was consistent with the mass deposited on the electrode in line with the calibration value of 1.05 f 0.02 ng/Hz of eq 2. The LCs- or lipid multibilayers-cast QCM was set in the sealed air vessel or immersed in a water cell and the ambient temperature was raised and lowered gradually in the 0 1989 American Chemical Society
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ANALYTICAL CHEMISTRY, VOL. 61, NO. 19, OCTOBER 1, 1989
Liquid crystal (LC) molecules
434:
PCB, k
364:
EBBA, k
CH3(CH2)&00
68%
i
4 s-
4
80°C
n-+
i
& 91 4:
82%
ChD, k
d ch+
i
Multibilager-forming amphiphiles
2Cl@E, Tc= 53 "C
"
2ClsN+2C1/PSS* , Tc= 42 OC
Flgure 1. Structures of liquid crystals (LC) and lipld multibllayers cast on the QCM plate.
E
Figure 3. Effect of orientation of smectic PCB liquid crystals on f r a quency changes at T , in air. PCB was cast on the surface of the Ag electrodes of the quartz plate which was treated with (a)octadecyltrichiorosllane (OTS) and (b) diphenyldichlorosilane (DPS).
- -
-
crystal to isotropic state (s i, n i, and ch i). The temperature where the frequency changed was independent of the raising rate of temperatures in the range 0.1-3 OC min-I. After temperatures were raised and then lowered in air and water phases, the frequency reverted to the original value, which means the coating LC materials did not peel from the substrate during experiments. The magnitude of the frequency enhancement of T, of the smectic PBC-coated quartz was 2-5 times larger than those of the nematic EBBA- and cholesteric ChD-coated QCMs. This means that the multilayered structure of smectic LCs parallel to the QCM plate plays an important role for the frequency change at T,in addition to the fluidity or elasticity change from solid to nematic or cholesteric state of LC coatings. The orientation of smectic LCs is known to be affected with the surface treatment of the substrate (14,15). Figure 3 shows effects of the surface treatment of the electrode of the QCM on the frequency change at Tcof smectic PCB LCs. When the Ag electrode surface was modified with long alkyl chain monolayers of octadecyltrichlorosilane (OTS), the large frequency enhancement was observed a t T,(k s) = 42 OC, in which LC molecules are thought to orient parallel to the long axis of alkyl chains of OTS as shown schematically in Figure 3a. On the contrary, the magnitude of the frequency enhancement reduced about a half when the surface was treated with diphenyldichlorosilane (DPS), in which LC molecules are supposed to orient parallel to the film plane due to the bulky phenyl groups of DPS surfaces (see Figure 3b) (15). These results imply that well-oriented layered structures parallel to the QCM plate are important to cause the large frequency change a t T,. Figure 4 shows the frequency decrease against the cast amount of smectic PCB on the QCM at temperatures below and above T, (20 and 50 "C, respectively). In the solid state below T, (at 20 "C), the frequency decreases linearly with increasing the mass of PCB deposited on the electrode in both cases of the OTS-and DPS-treated surface. The slope was calculated to be 1.1f 0.2 ng/Hz which is consistent with eq 2. However, in the fluid smectic liquid crystalline state of PCB deposited on the OTS-treated electrode at 50 "C, the frequency hardly decreased proportionally to the cast amount above 5 pg, which means that the QCM scarcely detects the mass above 5 pg of the fluid smectic layered structure on the electrode. In other words, unlike the solid LCs, the fluid or elastic layered structure is thought not to vibrate with the QCM plate. On the contrary, the frequency decreased almost
-
Temp / 'C Figure 2. Frequency increases of LC (15 pgbcoated QCMs at the phase transition from solid to liquid crystalline state in air: (a) smectic PBccOated, (b) nematic EBBAcoated, and (c)chdesteric ChDCoated QCMs. Closed circles show the frequency changes of the uncoated QCM in air. The similar frequency changes at T , of LC-coated and
uncoated QCMs were observed In a water phase.
range of 0.1-30 "C min-l. The resonance frequency at each temperature was measured.
RESULTS AND DISCUSSION Phase Transition of Liquid Crystals (LCs). Figure 2 shows frequency changes of the LC-coated QCM when ambient temperatures are raised gradually at a rate of 1 "C min-' in air phase. The frequency abruptly increased near the respective phase transition temperature from solid to liquid crystalline state (T,of k s, k n, and k ch) independent of media such as air phase and aqueous solution. The frequency hardly changed at the phase transition from liquid
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ANALYTICAL CHEMISTRY, VOL. 81, NO. 19, OCTOBER 1, 1989
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i'"Bpli'"'A
20
I
0 Cast A m w t / P g
Figure 4. Frequency decreases dependent on the cast amount of PCB liquid crystals on the electrode of the QCM at 20 and 50 OC in air. The electrode surface was treated wlth (a) octadecyltrichlorosilane (OTS) or (b) diphenyldichlorosilene (DPS).
8
I
I
I
2
3 4 T i m / min
5
6
Flgure 8. Repeated frequency changes of the 2,ePE ( T , = 53 OC)coated QCM when it was immersed into the 50 and 70 OC water baths alternately at the arrow.
fl
70
40
60
80
Temp./ "C Flgure 5. Frequency increases at T , of (a) the P C # E and (b) the 2C18N+2C1/PSS-multibilayer (15 pg)-coated QCMs in a water phase. The frequency enhancement and decrease at T , were observed reversibly when temperatures were raised and lowered in water.
linearly with increasing the cast amount even in the fluid liquid crystalline state at 50 OC when the surface was treated with the bulky DPS (see illustrations in Figure 3). From these findings, the large, abrupt frequency change at the T,of smectic PCB on the QCM is explained as follows. In the solid state of LC molecules below T,, the solid coating vibrates with the QCM plate and the frequency decreases proportional to the surface mass deposited. When LC coatings become fluid above T,,the multilayered smectic phases may slip between layers and the upper part of LC coatings cannot vibrate with the QCM plate. As a result, the frequency drastically increases at the phase transition temperature (k s) of the coating LCs. When smectic PCB molecules do not orient perpendicularto the QCM plate on the DPS-treated surface (see Figure 3b) or when the nematic EBBA and cholesteric ChD are deposited on the QCM (see parts b and c of Figure 2, respectively), the frequency changes at T,are very small because LCs do not form the layered structure parallel to the QCM plate and the slipping is difficult to occur even in the fluid state. The frequency is also affected by the viscoelastisity change of coatings on the QCM; however, the magnitude seems to be small compared with that due to the slipping of layered structures. Phase Transition of Lipid Bilayers in Water. Figure 5 shows the frequency change of 2C1$E and 2Cla+2C1/PSSbilayer-coated QCM when the temperature was increased gradually at a rate of 5 "C mi& in both water and air phases. The frequency increased immediately at the respective T,from solid to liquid crystalline state of coating multibilayers only in water, but not in air. The magnitude of the frequency enhancement at T,of the 2C16Pe-coatedQCM was 2 times
-
1
10 20 3( Cast Amount / w Flgure 7. Frequency decreases dependent on the cast amount of 2C16F€multlMlayers (T, = 53 "C) on the quae: (a) at 25 OC in water, (b) at 70 OC in water, and (c) In air hxlependent of temperatires (20-70 "C).
larger than that of the polymeric 2C18N+2C1/PSS--coated QCM. In the case of the LC-coated quartz, the frequency change was observed independent of media. Figure 6 shows the reversible frequency changes of 2C1$E multibilayer (T, = 53 OC)-coated piezoelectric crystal when the QCM was immersed in water phase at 50 and 70 OC repeatedly. The frequency changed reversibly in the range of 20 OC below and above the T,at least more than 10 times. The similar reversible frequency changes below and above the T,were also observed in the polymeric 2ClaN+2C1/PSS-coated QCM although the magnitude was relatively small. When the 2C16PEmultibilayer-coated QCM was covered with hydrophobic polymers such as polystyrene, the frequency change at T,in a water phase was hardly observed. This clearly indicates that the frequency change at T, of lipid multibilayers occurred only when the multibilayers contact with aqueous media. Figure 7 shows the relationship between the frequency decreased and the casting amount of 2C16PE (T,= 53 OC) multibilayers on the QCM. The frequency decreased linearly with increasing the casting amount of the solid bilayers below T,at 25 OC and the slope (1.0 ng/Hz) was consistent with eq 2, which indicates that all the mass of the solid multibilayers on the electrode vibrate with the QCM. On the contrary, in the case of the fluid liquid crystalline multibilayers at 70 "C, the frequency hardly increased with increasing the mass of multibilayers above 10 pg. This means that the upper layers above 10 pg of the fluid 2C16PE multibilayers are slipping between layers in the liquid crystalline state and the weight above 10 pg is not monitored. The s i m i i frequency behaviors were observed for the polymeric 2Cla+2C1/PSS- multibilayer film. In an air phase, the frequency decreased linearly with increasing the mass on the QCM and the frequency decrease due to the slipping was not observed. Dynamic mechanical measurements of 2ClaN+2C1/PSSfilms were made with a nonresonance, forced vibration in-
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ANALYTICAL CHEMISTRY, VOL. 61, NO. 19, OCTOBER 1, 1989
strument, Rheovibron viscoelastmeter, at 110 Hz in both water and air phases (16). In a water phase, the dynamic storage modulus (E? decreased abruptly by about 2 orders of magnitude near T, = 45 OC. In the tan b curve narrow peak was found at T,. The characteristic viscoelastic behavior at T , of the self-standing multibilayer-immobilized film was not found in air phase. When the 2C16PEmultibilayer-coatedQCM was immersed in water at temperatures above T,, the frequency gradually decreased with time [AF = -(60 f 10) Hz] which indicates the swelling of hydrophilic interlayers of the fluid lipid bilayers with water above T,. In the solid state of multibilayers, the frequency change due to the swelling was hardly observed (less than -5 Hz). When the polymeric 2C18N+2C1/PSS--coated QCM was immersed in an aqueous phase, the similar swelling behavior was observed above the T,, though the magnitude of the swelling was relatively small [AF= 4 2 0 f 10) Hz]. The frequency decrease above the T , due to the swelling was not clearly observed when the temperature was raised slowly in Figure 5, because the swelling amount (-60 Hz) was very small compared with the frequency enhancement at the T, (+300O-5000 Hz). From these findings, the frequency change of lipid multibilayer-coated QCM at the T, can be explained as follows. When lyotropic lipid multibilayers become fluid above the T, in aqueous phases, the fluid multibilayers seem to swell slightly with water penetrated into hydrophilic interlayers and slip between fluid and swelled layers, because the frequency change due to T, was not observed in the absence of aqueous media. When the polymeric 2C18N+2C1/PSS--coatedQCM was employed in water, the frequency enhancement at T, and the swelling amount in the fluid state were smaller than those of the 2C16PE-coatedQCM (see Figure 5). In the polyion complex type multibilayers, cationic 2ClsN+2C1amphiphiles form multibilayer structures complexing with PSS- polyanions at
hydrophilic head groups (6-8, 13), and the slipping and swelling between multibilayers seem to be difficult relative to the monomeric 2CI6PE multibilayers.
SUMMARY It was first found that piezoelectric quartz microbalances can detect the phase transition from solid to liquid crystalline state of the coating layered materials on the electrode by observing frequency changes. The general fluidity or viscoelastic change of coating materials can ah0 be detected from the frequency change, although the magnitude of that is relatively small compared with that due to the phase transition (slipping between layered structures). The obtained results are useful as a new application of piezoelectric crystals as well as a microbalance sensitive to the surface mass deposition. LITERATURE CITED (1) Sauerbrey, G. Z.Phys. 1959, 155, 206. (2) Ngeh-Ngwalnbi, J.: Foley, P. H.; Kuan, S. S.: Guilbault, G. G. J. Am. Chem. SOC. 1988, 108, 5444. (3) Muramatsu, H.; Dicks, J. M.; Tamiya, E.; Karube, 1. Anal. Chem. 1987, 59, 2760. (4) Roederer. J. E.; Bastiaans, G. J. Anal. Chem. 1983, 55, 2333. (5) Ebersole, R . C.; Ward, M. D.J. Am. Chem. Soc. 1988. 110, 6623. (6) Okahata, Y.; Ebato, H.; Taguchi, K. J. Chem. SOC.,Chem. Commun. 1987, 1363. (7) Okahata, Y.; Shimlzu, 0. Langmuir 1987, 3 , 1171. ( 8 ) Okahata, Y.: Ebato. H.; Ye, X. J . Chem. SOC., Chem. Commun. lB88- 1037. (9) Okahata, Y.; Arlga, K. J . Chem. Soc., Ct".&mmun. 1987, 1535. (IO) Crane, R. A.; Fischer, G. J. f h y s . D : Appl. W y s . 1979, 12, 2019. (11) Muramatsu, H.; Tamlya, E.: Karube, I. Anal. Chem. 1988. 60, 2142. (12) Skei, T.; Okahata, Y. J . Microencapsulation 1985, 2 , 13. (13) Okahata, Y.; Taguchi, K.: Sekl, T. J . Chem. Soc., Chem. Commun. 1985, 1122. (14) Kahn, F. J.; Tayler, G. N.; Schonhorn, H. R o c . Iff€ 1973, 61, 823. (15) Kahn, F. J. Appl. Phys. Left. 1973. 22, 366. (16) Taguchl, K.; Yano, S.; Hiratani, K.: Minoura, N.; Okahata, Y. Macromolecules 1988, 21, 3336.
RECE~VED for review March 17,1989. Accepted June 28,1989.
