Applications of Fluorescence Photobleaching Recovery

ACS Polymer GroupRaleigh, North CarolinaThursday, November 8

Gelation of “sticky” rods

Helical -polypeptide backbone,persistence length > 1000 Å

O

O

PBLG: poly(benzyl-L-glutamate)

PBLG is “sticky”but not very.

PSLG: a stickier wicket.

O

O

PSLG: poly(stearyl-L-glutamate)

Very rigid -helical polypeptide backbone Intramolecular

Hydrogen Bonds

Waxy Sidechains

C18 = Stearyl

•Self-solvating (well, sorta): Molten Rods—easier processing.•Oriented Films/waveguides/chiral separation•Polymer Physics of Rods in Solution, Gel & Melt•More rigid and more soluble than those high-$$$ Air Force &

NASA polymers•Hey…they don’t have to be rigid—helix-to-coil transition

Why everyone cares about these molecules again.

Idealized Rod Phase Diagram

ISO

a b

T LC

Very rigid rods! Mw Mw/Mn

by GPC/LS

x = L/d

a

(%)

b

(%)

avg

(%)

Onsager4/x

Flory

               

239,000 1.13 28 13.52

15.01.

5

14.31

14 27

208,000 1.08 24 15.21.

3

17.81.

3

16.52.

6

17 30

127,000 1.12 15 16.22.

1

25.31.

3

20.83.

4

27 46

Hypothetical Rod Phase Diagram With Gelation Transitions Overlaid

T

Iso LC

S1

P1

LL1

S2

LL2

P2

DSC can detect ISO-LC transition. Heat per molecule gives no trend.

0 10 20 30 40 5015

20

25

30

35

40

45

50

55

OrderedIsotropic

sarah file: s.conave

Schmidtke et al.

Figure 4a

Tm

Tc

Tem

pera

ture

(C

)

Weight % PSLG0 10 20 30 40 50

15

20

25

30

35

40

45

50

55

60

sarah file: s.conave

Schmidtke et al.Figure 4b

H (

mJ/

mg

PS

LG)

Weight % PSLG

Annealing perfects but does not alter gel structure. Suggests motion!

0 100 200 300 40030

35

40

45

50

55

Schmidtke et al.

Figure 5b

sarah file: s.annave

H (

mJ/

mg

PS

LG)

Time cured (minutes)0 100 200 300 400

25

30

35

40

45

50

Schmidtke et al.Figure 5a

sarah file: s.annave

Tem

pera

ture

(o C

)

Time cured (minutes)

Labeling Does Not Bug the Helix

Computer Simulation Intrinsic ViscosityStatic Light ScatteringDynamic Light

ScatteringPhase boundary

determinationsTemperature-

dependent studies

Modulation FPR Device

*

*

*

*

AOM

M

M

D

RR

DM

OBJ

S

PMT

PA OS

TA/PVD

L

Temperature-ramped modulation FPR

0 1000 2000 30000

1

2

3

4

Schmidtke et al.

Figure 6

scan1062

Melt at 30.9oC

2040 s

start ramp @ 0.3oC/min

Con

tras

t (A

C/D

C)

t (seconds)20 30 40 500

100

200

300

400

500

600

700

Schmidtke et al.Figure 7

combinehigh

DDSC

TRFPR: 30.7oC

DSC

DS

C (W

) or

DD

SC

+60

0 ( W

/ o C

)

T/ oC

20 30 40 50

1

2

AC

/DC

Everything can move, yet the structure remains

Sticky rods—remaining challenges

•Crosslink ‘em to lock in liquid crystal structure•Solution behavior above the gel transition•Rheological response•EM’s of these gels•Supercritical fluid drying of gels and LC’s

Problems related to entanglement

Bulk polymers: welding, processing, phase separation, failure

Solutions: rate of dissolving, intracellular transport, separations for genomics, joining PVC pipe!

Reptation—a theory to describe random snaking in polymer tangles

There once was a theorist from FranceWho wondered how molecules dance."They're like snakes," he observed,"as they follow a curve,the large ones can hardly advance."*

D ~ M-2deGennes

Need: better diffusion methodsProblem: lots of things appear to obey this scaling lawSuggests: look at the ignored systems, especially RODS

*With apologies to Walter Stockmayer

Entanglement in solution?Entanglement in solution?

To isolate spaghetti in "solution" with a fork is To isolate spaghetti in "solution" with a fork is difficult: hydrodynamic interactions interfere difficult: hydrodynamic interactions interfere with entanglement. After solvent is drained to with entanglement. After solvent is drained to obtain a "melt" the entire blob is easily handled. obtain a "melt" the entire blob is easily handled.

Collander

See, e.g., Lodge & Muthukumar, J. Phys. Chem. 1996, 100, 13275-13292

Our Hypothesis

Solutions containing rodlike diffusers may provide evidence for entanglement-like

phenomena in solution…or at least prove interesting, fun and challenging.

Evidence would be: sudden drop-offs in mobility with concentration, failure to

follow continuum mechanics.

Rods should make effective probes. But very challenging:

Early de Gennes paper on rod/coil diffusion: 19 citations

Same era de Gennes paper on coil/coil reptation: 479 citations

We may expect some problems!

Why it’s worth it: composite precursorfluids, dissolution rate, phase sep’n rate, relation to GPC, CGE of rods, intracellular transport.

Studying “entangled” rods by Optical Methods

Fluorescentlylabeled probe rod

Unlabeled rods

Solvent

Ld

1

1

3

3

3

L

L

L

1

1

2

2

2

dL

dL

dL

1

14

2

2

2

2

A

A

dLA

LC formation = 4/A2 5/dL2

Reduced # Density dL2/5

Doi-Edwards-Onsager Reference Volumes for Rods = number density = # of rods per unit volume

Desirable rod properties

Stiff—no bendingMonodisperse (all the same size)Non-aggregatingWater-solubleWe don’t have to make ‘em!

Let mother nature do the work: plants make viruses (unwillingly) with most of these qualities.

Seedlings

Sick Plants And close-up of mosaic pattern.

Doing our Part to Keep the “A” in LSU A&M

Two TMV’s in Transmission Electron Microscopy

An ear of corn has about as many kernels as TMVhas protein subunits (ca. 2130). The protein

subunits enfold a spiral-wound strand of RNA whichwill encode the next generation. TMV is more

extended than an ear of corn.

Better Views

http://www.uct.ac.za/depts/mmi/stannard/linda.html

TMV CharacterizationTMV Characterization

Sedimentation, Electron Microscopy & DLS Most TMV is intact. Some TMV is fragmented.

weaker, faster mode in CONTIN decay analysis

Intact TMV is easy to identify stronger, slower mode in CONTIN

It is easy to measure at low concentrations that avoid TMV-TMV interactions in DLS and we can use small amounts in FPR after dye attached.

Huh?

Tobacco from the Carolinas to Connecticut

If tobacco goes away…

“Traditionally in Kentucky great mounds of brush are piled and burned in February to prepare a bed for tobacco seedlings. I remember spending most of the day hauling and piling brush. My dad would start the fire in late afternoon and we would sit up most of a cold February or March night stoking the fire, watching the stars, and roasting hot dogs or marshmallows over the bonfire. Many times neighbors would stop by and sit with us for a spell around the fire, talking into the night.”

From: http://www.webcom.com/duane/farm2.html

Modulation FPR Device a’la Lanni & Ware

*

*

*

*

AOM

M

M

D

RR

DM

OBJ

S

PMT

PA OS

TA/PVD

L

SCOPE

Measuring Translational Diffusion

0 200 400 600 800 10000

1

2

3

4

DC

Sig

nal

Con

tras

t

t/s

0 2 4 6 8 10 12 140.000

0.005

0.010

0.015

0.020

0.025

0.030

0.035

0.040

/H

z

K2 / 105cm-2

FPR Data for TMV Solution: very low dye content.

0.1 1 10 1001

2

3

4

5

6

7

Source: Cush Monthly Report (5/1/01)

1/dL21/L3

Dse

lf /

10 -8

cm

2 s

-1

cTMV

/ mg mL-1

Dtracer self of TMV vs. cTMV

Rotation & Diffusion of TMV in Polymer Solutions

– – – Hard to see rotation in FPR; try scattering+ + + Fabulous new autocorrelators for scattering

10 decades of time in one measurement!– – – Contrast for scattering stinks: everything scatters, esp. in aqueous systems where refractive index matching cannot hide matrix.

Matrix Polymer

Solvent

TMV Probe