TERAHERTZ SPECTROSCOPY OF BIOMOLECULESIN WATER: L-PROLINE IN REVERSE MICELLES

NIST Colleagues: Craig Brown, Alan Migdall, Jerry Fraser, David Plusquellic

NIST Postdocs: Andrea Markelz, Matt Beard, Tim Korter, Okan Esenturk, Larry Iwaki, Karen Siegrist, Catherine Cooksey, Ahmasi Harris

Summer Students: Ari Evans (Cornell), Mary Kutteruf (UVa), Brendon Scheinman (Wash. U.), Ben Greer (Carnegie Melon)

Collaborators: Rad Balu and Susan Gregurick (UMBC), Joe Melinger (NRL)

Project Support and Funding

NIST Competence Program, Office of Law Enforcement and Safety,NIST STRS, DARPA, NAVY, DHS

• E. J. Heilweil and D. F. Plusquellic, “Terahertz Spectroscopy of Biomolecules,” book chapter in “Terahertz Spectroscopy: Principles and Applications,” Taylor and Francis, CRC Press, Susan Dexheimer, editor. Chapter 7, pages 269-298 (2008).

• David F. Plusquellic, Karen Siegrist, Edwin J. Heilweil, and Okan Esenturk, “Applications of Terahertz Spectroscopy in Biosystems,” review paper for Chemical Physics Physical Chemistry 8, 2412-2431 (2007).

Homeland Security

THz Metrology

BiomolecularPhysics

Pharmaceuticals

IMAGING

MATERIALS CHARACTERIZATION TIME-RESOLVED

SPECTROSCOPY

MODELING/THEORY

NIST Terahertz Project Objectives(1998- present)

• Investigate low frequency vibrational spectra of biomolecules, model biosystems and materials:

“THz spectroscopy probes structure, large-amplitude “torsional” modes and local environment, thus allowing the

conformational landscape to be directly mapped…”

• Examine hydrogen-bonding and bio-system dynamics • Bring together complimentary low-frequency spectroscopies

(e.g., Infrared, Raman, Inelastic Neutron Scattering)• Compare experiments to molecular modeling & theory• Advance THz imaging methods for bio-molecular and

materials applications (wafers, tissue, tablets, etc.)

Other Groups’ Biomolecular THz Work …

• P. Jepsen (Denmark) – Spectroscopy and modeling of peptides, sugars, biomolecules

• P. Bolivar (Germany) – DNA hybridization/chips• M. Havenith (Germany) – Spectroscopy of sugars in water• C. Schmuttenmaer (Yale) – Molecular liquids, biomolecules• M. Ito (Japan) – Methods development, biomaterials

spectroscopy• P. Taday (Teraview, UK) – Spectroscopy of

Pharmaceuticals, Rapid-scanning THz imager for tissue, tablets, materials characterization

• A. Markelz (SUNY Buffalo) – Spectroscopy of proteins, DNAs, etc.

• T. Korter (Syracuse) – Spectroscopy and theory for small biomolecules and explosives

Modified Fourier-Transform Infrared Spectrometer

MODIFICATIONS:• Silicon-coated mylar broadband beam-splitter• DTGS room temperature detector with HDPE window• Sensitivity from ~ 50 – 700 cm-1

Nicolet Magna 550 FTIR

Biomolecular THz Spectroscopy in AqueousReverse Micelles

Catherine Cooksey, NIST/NRC Postdoctoral Associate

AOTn-Heptane

Approach: • Encapsulate room temperature amino acids, proteins, DNAs in reverse water-alkane micellar structures to control water content and eliminate strong bulk THz water absorption • Also used in NMR and single molecule studies . . .

H2O and D2O in AOT and Brij-30 MicellesH2O / AOT

D2O / AOT

H2O / Brij-30

• Water in AOT anionic surfactant exhibits decreasing intensity and frequency shifts with higher w or micelle water loading

• Water encapsulated in the non-ionic surfactant Brij-30 shows “minimal” change in THz spectrum as the size is changed…

w=1,2,3,5,10,15,20

D2O / Brij-30

w=2: ~100 waters, d ~4 nm

Pathlength 4.2 mm

L-Proline-Water Inverse AOT Micelles[Pro]max ~ 11 Mol/liter

Solid in PE

L-Proline-Water Inverse AOT Micelles and Solid-State Spectrum

• L-Proline in AOT surfactant exhibits clear THz absorptions that correspond closely to those observed in the solid-state

• There appears to be significant red and blue-shifting of band frequencies arising from solvation interactions (e.g., hydrogen-bonding and solvent exclusion…

• Low frequency phonon bands of the solid become a broadened water hydrogen-bonding band

L-Proline-D2O in AOT Micelles[Pro]max ~ 11 Mol/liter

50 150 250 350 450 550 650

Ab

sorp

tio

n (

OD

)

0.000

0.050

0.100

0.150

0.200

Fructose in Water Inverse AOT MicellesCatherine Cooksey & Ben Greer (SURF student)

Wavenumber (cm-1)

50 150 250 350 450 550 650

Ab

sorp

tio

n (

OD

)

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

5% Solidin PE

??~5 mole/lw = 10

Phonons?

Wavenumber (cm-1)

•P3HT = Poly(3-hexylthiophene-2,5-diyl)

Regio-regular structure, average Mw ~87,000 (Sigma-Aldrich)

< 200 nm thick CVD films

•Conductive polymer organic semi-conductor

•Scientific and industrial interest ..

High efficiency solar cells,

flexible electronics , displays

•Non-contact, All optical measurement

•THz carrier concentration and mobility

•Frequency-dependent mobility

THz Measurement of Carrier Mobility in Semiconductor Polymer Films

With Okan Esenturk and Joe Melinger (NRL)

erahertz Signals Carrier Mobility

= mobility, = photogeneration efficiency

T /To = differential transmission,

h = Plank constant, light frequency,

N = refractive index of substrate,

e = electric charge, F = Fluence,

Z0 = free space impedance

eF(1-e-d) Z0

|T/To| h (1+N) =

(Hegmann et al. J. Appl. Phys. 98, 033701, 2005)

-6 -4 -2 0 2 4 6

-10

-5

0

5

10

Sig

nal

(a.

u)

Time (ps)

Ref THz-TDS

5

-5 0 5 10 15-0.6

-0.5

-0.4

-0.3

-0.2

-0.1

0.0

T/T

o (

%)

Relative Probe Time (ps)

Signal Comparison for P3HT versus PBTTT:PBTTT exhibits higher conductivity…

Esenturk, et. al. J. Phys. Chem. C (Letters), in press

-5 0 5 10 15 20 25 30 35 40 45

-60

-50

-40

-30

-20

-10

0

-2 0 2 4

Blend Ratios 75:25 50:50 45:55 20:80 0:100

T (

a.u

.)

Time Delay (ps)

10 15 20 25 30 35 40

40

50

60

70

80

Power dependence neat C60 film

Pea

k S

igna

l

Power (mW)

Blended Zn-Pthalocyanene/C60 Films

• ~ 1:1 blend has highest mobility

• Tightly bound excitons

• Peak ~t=0 is from C60

• Is dissociation intermolecular?

h

e-

Nano-layered ZnPthalocyanine/C60 Films

0 10 20 30 40

-70

-60

-50

-40

-30

-20

-10

0

T (

a.u

.)

Time Delay (ps)

40 nm 50:50 Blend 20 nm 10 nm 5 nm

Ratio exp calc -------- ----- ------5/10 2.0 2.05/20 4.1 4.15/40 8.8 8.7

10/20 2.0 2.010/40 4.4 4.3

20/40 2.2 2.1

Exp ratio = I1 / I2 at 30 psCalc ratio = nint1 / nint2 for tfilm = 440 nm• Tightly bound excitons -> free carriers

• Exciton diffusion length is ~nm in few ps • Thinner alternating layer structure -> higher mobility

Carrier Population Persists Beyond 0.5 ns

0 100 200 300 400 500

-60

-50

-40

-30

-20

-10

0

-5 0 5 10 15 20-1.0

-0.8

-0.6

-0.4

-0.2

0.0

10 nm

5 nm

Diffe

ren

tia

l T

ran

sm

issio

n (T/T

o,

a.u

.)

Time Delay (ps)

T/T

o (N

orm

aliz

ed)

• Amplitude ratio extends beyond 0.5 ns

• Similar carrier diffusion and recombination processes

0 10 20 30 40

0

5

10

15

2020 nm

295 K 78 K

T/T

o

Time Delay (ps)

0 10 20 30 40

0

5

10

15

20 C60

295 78

T/T

o

Time Delay (ps)

0 10 20 30 40

0

5

10

1540 nm

295 K 78 K

Lock

-in S

igna

l (V

)

Time Delay (ps)

0 10 20 30 40

0

5

10

15

20

255 nm

350 K 295 K 78 K

T/T

o (a

.u.)

Time Delay (ps)

Photoconductivity versus Temperature

Summary THz FTIR and TDS spectrometers can collect low-frequency vibrational spectra of biomolecular solids and as solutes in dispersed aqueous micelle samples

Low frequency THz aqueous spectra obtained for L-Proline and Fructose model species Novel Time-Resolved THz measurements of conducting nanometer

organic thin-films reveals carrier mobility and efficiencies forscreening materials for semiconductor device applications

Future Prospects:

THz methods will be useful for determining water-phase biomolecular structure and solvent interactions

Modeling and theoretical advances (e.g., modified potential functions; add anharmonicity?) are needed to identify spectral features of hydrogen-bonded systems

Try similar approach on peptides and small proteins…

THz Spectra of Water in AOT Micelles

Optical pathlength = 4.2 mm

(~100 waters; d~4 nm)