Surface Modifications and Applications of Organic & Inorganic Surfaces

Hyuk Yu(yu@chem.wisc.edu)

Department of Chemistry University of Wisconsin

Madison, WI 53706-1396

LSU Chemistry ColloquiumBaton Rouge, LA

March 5, 2004

Goals

•Outline what appears to be pivotal in the fundamental knowledge base and corresponding applications of surfaces.

•What we have learned.•What need to be learned.

Personal Perspective

•What the future holds, in terms of biomedicalapplications

Contributors & Collaborators Principal Contributors:

Dong X. Lin, Seagate, MinneapolisAbukar Wehelie, Intel, HoustonZhihao Yang, Eastman Kodak, Rochester, NYSangwook Park, LG Chem, KoreaJeffrey Galloway, Sandia National Labs.Dr. Keiji Tanaka, Kyushu Univ., JapanDr. Xiqun Jiang, Nanjing Univ., ChinaThorsteinn Adalsteinsson, MPI-Golum, GermanyDr. Junwei Li, Chemistry, Lehigh UniversityWeiguo Cheng, in residence, to leave for NALCO

Contributors & Collaborators

Principal Collaborators:Charles M. Strother, M.D., Radiology, U.W.-

Madison & Baylor College of Medicine, Houston

Richard Frayne, M.D., Ph.D., Radiology, U.W.-Madison & Univ. of Calgary

Orhan Unal, Medical Physics, U.W.-MadisonFrank Denes, C-PAM & Biological systems

Engineering, U.W.-Madison

Starting Point

•Liquid Surfaces: flat & smooth; facile dynamics to establish equi-chemical potential.•Amorphous Solid Surfaces: rough & irregular; dynamics-limited quasi-equilibrium; passivation required to make them smooth;

In case of amorphous polymer surfaces, the passivation route is effective through glass

transition

Water Surface: Air/Water Interface

Flat and smooth

“I plan to tell you of the behaviour of molecules and atoms that held at the surfaces of three-dimensional solids and liquids. . . . I will show you that we can have adsorbed films which really constitute two-dimensional gases, two-dimensional liquids and two-dimensional solids”

Irving Langmuir, Science 1936, 84, 379.

Air/Water Interface

• Pliny the elder (Gaius Plinius Secundus), AD 23-79 : “that all sea water is made smooth by oil, and so divers sprinkle oil on their face because it calms the rough element and carries light down with them”; . . . Historia Naturalis.

• Benjamin Franklin:. . .where the waves began to form, and there the oil, though not more than a teaspoonful, produced an instant calm, . . . perhaps half an acre as smooth as a looking glass”; Phil. Trans. Roy. Soc. (1774), 64, 445.

• Lord Rayleigh: “The earlier part of Miss Pockels’ letter covers nearly the same ground as some of my own recent work, . . . , raising many important questions.

I hope soon to find opportunity for repeating some of Miss Pockels’ experiment”

• Agnes Pockels: “MY LORD-Will you kindly excuse my venturing to trouble you with German letter on a scientific subject?. . .; Nature (1891) 43, 437.

• Irving Langmuir: “The constitution and fundamental properties of solids &

liquids. II Liquids”; J. Am. Chem. Soc. (1917) 39, 1848.

Historical Milestones

Early examples of chemistry on A/W monolayer

In low surface pressure, i.e., in low surface density, permanganate solution oxidizes oleic acid monolayer through its double bond at C 9 position.

At high surface pressure, oleic acid monolayer is no longer oxidized, for its chain conformation gives rise to a hydrocarbon insulation layer of 10Å thick.

This is a clear chemical evidence for flatness of Air/Water interface in macroscopic length scales.

Alexander & Rideal, Proc. R. Soc. London 1937, A163, 70.

Permanganate solution

Permanganate solution

oxidative cleavage of double bond on oleic acid

increase in surface pressure

no more cleavage!

10Å

Surface roughtness of A/W is 3.2Å by X-ray reflectometryBraslau et al., Phys. Rev. Lett. 1985, 54, 114.Braslau et al., Phys. Rev. A 1988, 38, 2457.

Changes on Polar Groups Covered Hydrocarbon Polymer (PE, PP PS) Surfaces

Time dependent aging of oxygen plasma treated the polymer surfaces

•Lower pressures, 100-250 mT:O2+ etching &

functionalization of electron deposited surfaces.•Higher pressures, 500-700 mT: oxygen atom etching

& functionalization of almost neutral surfaces

•Oxygen containing groups, e.g., carbonyl, carboxyl, alcohol, etc. remain on the surfaces.

•Water contact angle comes almost to zero.•It changes significantly within a few days upon aging in

wet atmosphere, 18-95% relative humidity.

50

40

30

20

10

0

Contact Angle(

o )

160140120100806040200Aging time (day)

RH 18%, 100 mT y=42.5-28.9EXP(-x/12.4) RH 18%, 275 mT y=51.9-41.7EXP(-x/13.2)

Aging Time Dependence of Water Contact Angle

Oxygen plasma treated, aged in 18% RH

polymer interior

hydrophilic group Aging in contact with air

polymer interior

Functional group translocation hypothesis

Oxygen plasma generated hydrophilic surface functional groups, will likely migrate into the polymer interior driven by surface energy.

Surface topography of PS film prior to the plasma treatment

Surface topography of PS film, treated by O2 plasma at 100mT, 40W, 1min

Surface roughness of PS films

Film roughness in a spatial range of 2 m does not change with time, hence it is unlikely to be responsible for the contact angle changes.

Sample RMS roughness/nm

PS w/o plasma treatment 0.19

Right after plasma treatment, 100 mT oxygen1 day after the treatment, 100 mT oxygen2 months after the treament, 100 mT oxygen

0.220.210.27

Right after plasma treatment, 275 mT oxygen15 days after the treatment, 275 mT oxygen

0.770.74

The relative concentrations of different carbon binding states in O2 plasma treated PS surface vs. aging time

60

50

40

30

20

10

0

Percentage of different C1s binding state (%)

6420Aging time (day)

-CH2-

-O-C-

-C=O

-O-C=O

Plasma: 100 mT, 40W, 1min; aged in 18% RH; 45˚ takeoff angle

SFG vibrational spectra of PS surface with & without oxygen plasma treatments

40

30

20

10

0

SFG signal (a. u.)

330032003100300029002800wavenumber (cm-1)

PS surface without treatment PS Surface treated by oxygen Plasma

SFG vibrational spectra of the plasma treated PS surface at different aging times

14

12

10

8

6

4

2

0

SF intensity(a. u.)

330032003100300029002800Frequency(cm

-1)

1 day after plasma treatment 1.5 day after plasmam treatment 6 days after plasma treatment

C-H Symmetric Strench of CH3 and OCH2

Femi Resonance

Symmetric strench of CH2

Reorientation of Polar Functional Groups on PS Surface aged in air

Bare PS

Freshly treated PS

CH3 CH2OCH3C

O

Treated and aged PS in air

OCH3 C

OOCH3OH

CH3C

O

OCH2

CH3

CH2OCH3OCH2C

O

OH C

O

CH3

SummaryThe water contact angle of the plasma modified PS surface increases

markedly within two days of storage in air under 18-95% RH.

Surface roughness can be ruled out as the main cause for the change of water contact angle.

The depth profile of oxygen is essentially unaffected by aging as deduced from XPS measurements, indicating that reorganization of chain segments is confined in a layer thinner than the probing depth of XPS (3 nm).

Sum frequency generation (SFG) spectral changes with time provide direct support for the reactions in plasma treatment involving aromatic ring opening, followed by the formation of oxygen contained polar groups as side chains.

When the treated surfaces are aged in air at different RH, the intensity of CH2 vibration increases with time, while that of CH3 vibration decreases, indicating that the side chains with polar functional groups reorient toward the polymer interior, while the PS surface is covered mostly by polymer backbone. The final surfaces are not those of untreated PS.

Conversion of Inorganic Surfaces to Functional Biomembrane Mimetic Surfaces

Monolayers to Mimic Biomembrane Bilayers

AirAqueous Phase

Preserving Functionality of Membrane Proteins self-assembled on Phospholipid Monolayers?

• Advantages for using solid substrates:Stable and RobustVery large surface area (>300 cm2/g)

GLASS BEAD

Biomembrane Mimetic Surface Modification of Silica Substrates

Biomembrane Mimetic Surfaces

• Lipase on/in phospholipid self-assembled monolayers (SAMs)–Monolayer structure and dynamics–Diffusion of lipase in phospholipid SAMs–Lipase activity on/in phospholipid SAMs

(CH2)12 COOHHOOC

OHOHOHOHOHOH

OOOOOO

Si CH2CH2CH2NH2

Si CH2CH2CH2NH2

(CH3O)3SiCH2CH2CH2NH2

1,12-dodecanedicarboxylic acid

OO

C

OHPONO

O

OMonomyristoyl lysolecithin

(lyso-PC)

Lecithin-COOH (PC-COOH)

DCC/THF

DMAP/CHCl3

Carbonyldiimidazole1. 2.

CO

CO O

O

O

O

OO

C

PON OC

COOH

O

NC

C

ONH2

OOOOOO

Si

Si

O

O

O

OO

C

P O NO

O

Preparation of Phospholipid SAMs on Silica

Self-Assembling of Phosphoipids on Native Oxide of Si Waferloading the immobilized lipid monolayer with free lipids(DLPC/NBD-PE)

Lateral Diffusion of a Probe (NBD-PE) in lipid SAMs

D=1.9±0.4x10-9 cm2/s (at 22°C)

References:•In transfered lipid bilayers, D=1~4x10-8 cm2/s at 30°C (M. Stelzle, R. Miechlich, E. Sackman, Biophys. J. 1992, 63, 1346.)•In DMPC liposomes, D=2x10-10 cm2/s at 22.5°C(A. B. Smith, H. M. McConnell, Proc. Natl. Acad. Sci. 1978, 75, 2759)

Fluorescent dye labeling of lipase

+ NH2

lipase w/ lysine residue

Fluorescein isothiocyanate isomer I (FITC)

mass ratio 1:10

24 hours @ 4°C, pH 10

1) Sephadex G-25 fine column

2) freeze dry

OO OH

N C S

COOH

NC

N FITC

S

H H

Lateral Diffusion of Lipase on Phospholipid SAMs

D=2.7±0.4x10-10 cm2/s

Hydrolysis Reaction of Umbelliferone Esters

OOOCORR: C16H33OOO+R-COOUmbelliferone stearate(non-fluorescent)

Umbelliferone(fluorescent)

Hydrolysis_

Hydrolysis of Umbelliferone Stearate by Lipase at Phospholipid SAM on Silica Gel

lipaseumbelliferone stearate

umbelliferonestearic acid

Experimental Test of Interfacial Activation of Lipase by the Lipid SAMs

UMB esters were loaded to the monolayer in CHCl3/MeOH

Hydrolysis condition: in 0.1M phosphate buffer, pH=7.0,

23°C1 mL of Pseudomonas sp. lipase

solution (ca.. 0.4 mg/mL) was added to hydrolyze

the UMB substrate.The fluorescence of elute was

monitored.

OOOumbelliferoneLipase

400

300

200

100

0

Fluorescence Intensity / a. u.

121086420

Flow Volume / ml

Control Exp.

Summary

A phospholipid monolayer on silica substrates (SiO2/Si) is constructed through a novel method, by a self-assembling process.

Such a monolayer is highly stable and structurally mimicks biomembranes.

The partially localized phospholipid SAMs show the dynamic properties (e.g. diffusion) close to those of biomembranes.

A biomembrane enzyme, lipase, localized to the monolayers are protected from denaturation.

Interfacial activation of lipase by the lipid SAMs is observed, indicating the viability of the monolayers as the biomembrane mimetic.

Endovascular biomedical applications

NH2

NH2NH2

+

OH

HO

N

NN O

OHO

O

HO

O

O

HO

1,1-'Carbonyldiimidazole Pyridine

OC

H

N

OHO

O

ONN

N

HO

OH

HOO

HO

H

GdO

O

N

NN

OO

O

OO

O

NH

CO GdCl3.6H2O

PE

PE PE

PE

NH2

NH2

Plasma

Hydrazine Plasma Functionalization of PE Surface

DTPA[Gd(III)] Complex & Water

Gd3+

NN

N

HO

HO

HO

HO

OHO

O

O

O

O

HO

H

H

O H

H

OH

H O

H

bulk water

inner-sphere

waterouter-sphere

water

Gd3+

O

O

O

O

O OH

HO

HO

HO

HO

N

N

N

H H

O

HO

H

H

OH

HO

H

H O

H

inner-sphere water

O

O

O

O

O OH

HO

HO

HO

O

N

NN

H H

O

HOH

H OH

Gd3+ bulk water

outer sphere water

NH2

NH2

NH2

NH

NH2

NH2

NH2

NH2

NH2

NH2

linker

HOH

H OH

Gd3+ has 7 unpaired 4f electrons, hence T1 of proton NMRof inner-sphere water is much shorter than bulk water.

Contrast Agent

DTPA

N

O

Gd3+

C

O/H2O

X-ray structure of Gd DTPA(H2O)2-, top view(hydrogen atoms are omitted)

R: rotational correlation time of the chelatekex: water/proton exchange rate

Imaging Mechanism•Interplay between the rotational mobility of ligand and the exchange rate of inner sphere water;

Caravan et al., Chem. Rev. 1999, 99, 2293-2352.

•Difference exists for the imageability between spatially confined DTPA[Gd(III)] within a thin slab from a surface and that spatially dispersed in bulk.

Scan Parameters

2D SPGR

TR = 18 ms, TE = 4.1 ms, Flip angle = 30º

Acquisition Matrix = 256 X 256 PE

Slice thickness = 2-3 mm

FOV = 16 cm X 16 cm and RBW = ± 32 kHz

2D SE

TR = 300 ms, TE = 9.0 ms, Flip angle = 30º

Acquisition Matrix = 256 X 256 PE

Slice thickness = 2-3 mm

FOV = 16 cm X 16 cm and RBW = ± 32 kHz

1.5 T GE CV Scanner (40mT/m, 150 mT/m/ms)

Agarose Gel Encapsulation of Functionalized PE Rods

Agarose gel

Gd3+

Gd3+

Gd3+

Gd3+

PE rod

1

2

3

1

2

3

1

2

3

Yogurt Saline( 0.9% NaCl, pH= 5.0)

Blood (human bloodbank)

MRI of Gel Encapsulated of PE Rods(soaked for 1 hour)

MRI of Gel Encapsulated of PE Rods(soaked for 10 hours)

Design III

III/5 after 30min6F Cath filled with 4% Gd

Catheter filled with Gd(III) solution

QuickTime™ and a decompressorare needed to see this picture.

Catheter coated with encapsulated DTPA-Gd3+ covalently linked

QuickTime™ and aCinepak decompressorare needed to see this picture.

SummaryWorking Hypothesis: For artificial surfaces to be biomembrane

mimetic, (structurally and functionally), they must manifest all the equilibrium and dynamics of at least monolayers of phospholipids.

MRI of endovascular medical devices, catheters & guide wires:

For diagnostic and interventional endovascular devices to be magnetically imageable, contrast agent functionalization on the surfaces with thin-layer hydrogel encapsulation suffices.

Conversion of Gold Surface to Viable Biomembrane Mimetic Surface

Lecithin-COOH (PC-COOH)

O

O

OO

OC

PON OC

COOH

O

H2NCH2CH2SH

EDC

HOBTCH2Cl2

O

O

OO

OC

PON OC

CONH

O

CH2CH2SH

Chemistry of SAMs on Gold Surface

O

OO

O

OC

PO

N

OC

CONH

O

CH2

CH2

SH

O

OO

O

OC

PO

N

OC

CONH

O

CH2

CH2

SH

O

OO

O

OC

PO

N

OC

CONH

O

CH2

CH2

SH

O

OO

O

OC

PO

N

OC

CONH

O

CH2

CH2

SHAu

Glass

Illustration of Phospholipid Monolayers through Thiol Linking on Gold

Surface Plasmon Resonance Reflectivity;Bare Gold & BSA

80Å thickness, native BSA: 140Åx40Å

Surface Plasmon Resonance Reflectivity;Bare Gold, PL Monolayer & BSA adsorbed

PL SAM thickness 20Å: tilt angle, 30˚BSA adsorbed thickness: 7Å, granular and heterogeneous.

Surface Plasmon Resonance Reflectivity;Bare Gold, PL Monolayer & Lipase adsorbed

PL SAM thickness: 20ÅLipase adsorbed layer: 25Å, native lipase 35Åx117Å

Summary

• Polycrystalline gold surface massively adsorbs a common serum protein, BSA.

• A self-assembled phospholipid monolayer on the polycrystalline gold substrate has a closed-packed and order structure as deduced by SPR, PM-FT-IRRAS and AFM measurements.

• The thiol-terminated phospholipid monolayer shows selective adsorption of a membrane protein, lipase, and resistant to BSA adsorption.

Personal Perspectives

Phospholipid self-assembled monolayers hold promise in modifying inorganic and metallic surfaces, but the nature of SAM seems empirical at best, hence some pivotal ingredients appear to be missing.

What have we learned? Chemistry of labeling & modification, together with characterization of the surfaces, are essential.

What must be learned? Dynamics & longevity.

What the future holds? Making surfaces looking & acting like biomembranes may be the way for all future prosthetic implants.

I thank you for your patience.

Surface Modifications and Applications of Organic & Inorganic Surfaces

Addenda

LSU Chemistry ColloquiumBaton Rouge, LA

March 5, 2004

Addenda:not shown on 01/07/03Paper Trail-1

References: BSA on PEG Brushes & Mushrooms on Silica Surfaces

Zhihao Yang and Hyuk Yu, Preserving a Globular Protein Shape on Glass Slides: A Self-Assembled Monolayer Approach, Adv. Mater. 9, 426-429 (1997).

Mobility of Lipid on Phospholipid-Cholesterol Binary Monolayers at A/WKeiji Tanaka, Patricia A. Manning, Victor K. Lau and Hyuk Yu, Lipid Lateral Diffusion in Dilauroylphosphatidyl choline/Cholesterol Mixed Monolayers at the Air/Water Interface, Langmuir 15, 600-606 (1999).

Lipase Activity on Phospholipid Monolayers Assembled on Silica Surfaces.Zhihao Yang and Hyuk Yu, Biomembrane Mimetic Surfaces by Phospholipid Self-Assembled Monolayers on Silica Substrates, Langmuir 15, 1731-1737 (1999).

BSA Adsorption and Mobility on PEG Brushes & Mushrooms on Silica Surfaces.Zhihao Yang, Jeffrey A. Galloway and Hyuk Yu, Protein Interactions with Poly(ethylene glycol) Self-Assembled Monolayers on Glass Substrates: Diffusion and Adsorption, Langmuir 15, 8405-8411 (1999).

Protease Activity toward a Flexibly Attached Substrate on Silica Surfaces.Alan R. Esker, Philip F. Brode III, Donn N. Rubingh, Deborah S. Rauch, Hyuk Yu, Alice P. Gast, Channing R. Robertson, and Giuseppe Trigiante, Protease Activity on an Immobilized Substrate Modified by Polymers: Subtilisin BPN' Langmuir 16, 2198-2206 (2000).

Lipid Mobility on Multiply Stacked Bilayers vs. Monolayers at A/WThorsteinn Adalsteinsson and Hyuk Yu, Lipid Lateral Diffusion in Multi-Bilayers, and in Monolayers at the Air/Water and Heptane/Water Interfaces, Langmuir 16, 9410-9413 (2000).

Addenda: not shown on 01/07/03 Paper Trail-2

U.S.Patent on Magnetically Imageable Polymer Surface CoatingsRichard Frayne, Charles M. Strother, Orhan Uanl, Zhihao Yang, Abukar Wqehelie & Hyuk Yu, Magnetic Resonance Signal-Emitting Coatings, U.S. Patent 6,361,759: 03/26/2002

U.S.Patent on Biomembrane Mimetic SurfacesHyuk Yu, Charles M. Strother, Xiqun Jiang, Sangwook Park, Biomembrane Mimetic surface Coatings, Biomembrane Mimetic Surface Coatings, U.S. Patent 6,486,334: 11/26/02.

Polymer Surface RearrangementJunwei Li & Hyuk Yu, Surface Rearrangement of Oxygen Plasma-treated Polystyrene: Surface Dynamics and Humidity, to be submitted to Langmuir. Biomembrane Mimetic SurfaceXiqun Jiang, Sangwook Park, Zhihao Yang & Hyuk Yu, Self-assembled Monolayers of Phospholipids on Gold Surface: Toward Biomembrane Mimetic Surfaces, to be submitted to Langmuir.

AFM Image of Lipase Adsorbed on Lipid SAMs

6

4

2

015141312111098

nm

2D-Power Density Spectrum of the AFM Image

• A characteristic object sizing at 11 nm is indicated, which agrees with the size of native lipase molecules (~7 nm) with a 4 nm broadening from the AFM tip.

Bare Au + BSA

100 nm

(a)

(b)

200 nm

PC

(c)

200 nm

PC +BSA

100 nm

(d)

200 nm

PC + lipase

Reorientation Reversibility

100 mT oxygen1 min, 40W18% RH

80˚C water, 5 minDry in nitrogen18% RH

80˚C water, 5 minDry in nitrogen18% RH

Reorientation Reversibility

275 mT oxygen1 min, 40W51% RH

80˚C water, 5 minDry in nitrogen51% RH

80˚C water, 5 minDry in nitrogen51% RH