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SINGLE MICROBUBBLE MEASUREMENTS OF LIPID MONOLAYER VISCOELASTIC PROPERTIES FOR SMALL AMPLITUDE OSCILLATIONS Jordan S Lum, Jacob D Dove, Todd W Murray, and Mark Andrew Borden Langmuir, Just Accepted Manuscript • DOI: 10.1021/acs.langmuir.6b01882 • Publication Date (Web): 23 Aug 2016 Downloaded from http://pubs.acs.org on August 24, 2016
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Langmuir
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SINGLE MICROBUBBLE MEASUREMENTS OF LIPID MONOLAYER
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VISCOELASTIC PROPERTIES FOR SMALL AMPLITUDE OSCILLATIONS
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Jordan S. Lum1, Jacob D. Dove1, Todd W. Murray1,2, Mark A. Borden*,1,2
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Department of Mechanical Engineering, University of Colorado, Boulder, CO 80309, USA
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Materials Science and Engineering Program, University of Colorado, Boulder, CO 80309, USA
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*Corresponding Author
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Mark A. Borden, PhD
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Department of Mechanical Engineering
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University of Colorado
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1111 Engineering Drive
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Boulder, CO 80309-0427
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Phone: 303.492.7750
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Fax: 303.492.3498
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Email:
[email protected] 17 18
KEYWORDS: surface elasticity, viscosity, resonance frequency, intermolecular forces
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ABSTRACT
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Lipid monolayer rheology plays an important role in a variety of interfacial phenomena,
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the physics of biological membranes, and the dynamic response of acoustic bubbles and drops.
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We show here measurements of lipid monolayer elasticity and viscosity for very small strains at
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megahertz frequency. Individual plasmonic microbubbles of 2-6 µm radius were photothermally
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activated with a short laser pulse, and the subsequent nanometer-scale radial oscillations during
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ring-down were monitored by optical scatter. This method provided average dynamic response
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measurements of single microbubbles. Each microbubble was modeled as an underdamped,
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linear oscillator to determine the damping ratio and eigenfrequency, and thus the lipid monolayer
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viscosity and elasticity. Our non-isothermal measurement technique revealed viscoelastic trends
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for different lipid shell compositions. We observed a significant increase in surface elasticity
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with lipid acyl chain length for 16 to 20 carbons, and this effect was explained by an
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intermolecular forces model that accounts for lipid composition, packing and hydration. The
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surface viscosity was found to be equivalent for these lipid shells.
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anomalous decrease in elasticity and increase in viscosity when increasing the acyl chain length
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from 20 to 22 carbons.
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technique to investigate lipid monolayer rheology in new regimes of frequency and strain,
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possibly elucidating phase behavior, as well as how the dynamic response of a microbubble can
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be tuned by the lipid intermolecular forces.
We also observed an
These results illustrate the use of a novel nondestructive optical
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Langmuir
INTRODUCTION
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Surfactant monolayer rheology plays an important role in a variety of interfacial phenomena,
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including surface-wave propagation, wetting, emulsification, emulsion stability, settling and
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multiphase flows.1 A particular class of surfactant monolayers – lipid monolayers – has been the
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focus of research on the biophysics of lung surfactant2 and cell membranes.3 In the ultrasonics
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community, there is interest in lipid-coated microbubbles and fluorocarbon nanodrops as imaging
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contrast agents, molecular probes, drug delivery vehicles and theranostics.4 The stability and
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dynamic behavior of such acoustic drops and bubbles depends on the lipid monolayer
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compression rheology.5,6 In these applications, and others employing microscale drops and
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bubbles, the relevant timescale for monolayer compression and expansion is given by the
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eigenfrequency for volumetric oscillations, which typically occurs in the megahertz range.5–7
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The most common method for measuring compression properties of lipid monolayers has been
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the Langmuir trough.8 Here, the compression elasticity (modulus) is calculated from the slope of
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the surface pressure-area isotherm, while the compression viscosity is obtained by the time
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variation of surface tension upon compression.1 Models of microbubble acoustic behavior have
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used surface elasticity values determined in this manner.9 The Langmuir trough is limited,
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however, to frequencies of a few hertz and below. The Langmuir trough is also inaccurate for
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measuring surface tension in condensed monolayers that may accumulate anisotropic stresses.10
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Sessile drop and captive bubble shape tensiometers, which rely on millimeter-scale particles to
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achieve a Bond number near unity, are likewise limited to frequencies below one hertz.11
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Capillary pressure tensiometer techniques can operate at frequencies up to 150 Hz,12 and light-
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scattering methods can access capillary waves at the liquid surface with frequencies up to 100
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kHz.1
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magnitude below the typical eigenfrequencies of microscale (femtoliter) drops and bubbles.
Unfortunately, these techniques remain limited to frequencies at least an order of
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Our group recently described a novel optical method for measuring the photoacoustic response
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of plasmonic microbubbles,13–15 which is developed here to explore monolayer rheology at
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megahertz frequencies, albeit in the linear regime under non-isothermal conditions. The method
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employs a 0.5-ns pulsed laser to stimulate a microbubble loaded with 5-nm gold plasmonic
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nanoparticles. Heating of the gas core leads to rapid expansion and subsequent “ring down” of
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the microbubble, which is captured by light scatter from a second laser operating in continuous-
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wave mode.
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frequency and damping ratio, respectively. One advantage of this technique is that it measures
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the surfactant rheology at the characteristic response time of the particle to a thermal or
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mechanical impulse. This response time depends on the particle size, so it is important to
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measure over a range of megahertz frequencies.
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perturbations from the resting state, where the particle is in mechanical equilibrium and has a
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very low surface tension. Such perturbations are relevant not only for surfactant-stabilized
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bubbles and drops, but also natural lung surfactant2 and lipid rafts in the cell membrane.3
The lipid monolayer elasticity and viscosity are determined by the natural
Additionally, the technique investigates
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Another advantage of the optical technique is the high signal-to-noise ratio which manifests
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from averaging over thousands of nondestructive pulses per microbubble.14 This allows one to
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resolve sub-nanometer scale radial oscillations, or less than ~0.1% area strain, which is well
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within the linear regime.6 The intermolecular lipid displacements at such tiny strains are only
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~0.1 – 1.0 pm. Thus, the method can approximately measure the intermolecular forces between
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lipids. In the present study, the method is expanded to explore the intermolecular forces for a
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Langmuir
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homologous series of saturated diacyl phosphatidyl choline (PC) lipids commonly found in
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biological membranes and biomedical emulsions and foams.
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MATERIALS AND METHODS Fabrication of Plasmonic Lipid-Encapsulated Microbubbles
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Plasmonic microbubbles were fabricated using 1,2-dipalmitoyl-sn-glycero-3-phsophocholine
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(DPPC, 16 carbons per acyl chain), 1,2-distearoyl-sn-glycero-3-phsophocholine (DSPC, 18
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carbons per acyl chain), 1,2-diarachidoyl-sn-glycero-3-phsophocholine (DAPC, 20 carbons per
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acyl chain), or 1,2-dibehenoyl-sn-glycero-3-phsophocholine (DBPC, 22 carbons per acyl chain)
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purchased from NOF America (White Plains, NY, USA). The emulsifier, 1,2-distearoyl-sn-
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glycerol-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000] (DSPE-PEG2000, 18
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carbons
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phosphoethanolamine-N-[biotinyl(polyethylene
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purchased from Avanti Polar Lipids (Alabaster, AL, USA).
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perfluorobutane (PFB) at 99 wt% purity obtained from FluoroMed (Round Rock, TX, USA).
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The plasmonic nanoparticles, 5 nm avidin-tagged gold nanospheres, were purchased from
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Nanopartz (Loveland, CO, USA).
per
acyl
chain)
and
the
biotinylated
species,
glycol)-2000]
1,2-distearoyl-sn-glycerol-3(DSPE-PEG2000-B)
were
The gas core comprised
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The microbubble shells comprised either DPPC, DSPC, DAPC or DBPC combined with the
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emulsifier and biotinylated species at a molar ratio of 90:9:1, respectively. All microbubbles
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were fabricated using probe sonication and size isolated to 2-6 µm radius using differential
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centrifugation. The procedure for synthesizing plasmonic nanoparticle-coated microbubbles was
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previously described by Dove et al.13 Briefly, the lipids were suspended in 0.01 M NaCl
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phosphate-buffered saline (PBS) pH 7.4 (4 mg/mL total lipid) and heated 10% above the 5
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phosphatidylcholine main phase transition temperature to promote lipid mixing.
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suspension was then cooled and further refined using a Branson 450 Sonifier (Branson, Dandurt,
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CT) by completely submerging the probe sonicator tip and sonicating on a low setting. The
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headspace was then filled with PFB gas, and the sonicator tip was set to mechanically agitate the
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gas-liquid interface on the highest power setting to form microbubbles. Microbubbles were
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centrifuged at 300 RCF (relative centrifugal force) for 3 min (Eppendorf 5804, Hauppauge, NY,
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USA) and washed three times to remove free lipid. The resulting microbubbles were centrifuged
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at 100 RCF to allow removal of smaller bubbles (
