This presentation exemplifies the

latest Power Point dogma.

1
A full sentence at the top tells the main point.
Vertically aligned lists are avoided in favor of
diagonal arrangement to create visual tension.
A page number helps reader
ask questions.
Tasteful use of animation helps.
Occasionally, put in something strange or fun to keep
professors and other tired listeners awake.
dogma, n. A settled or established opinion, belief, or principle
You cannot measure absolute
molecular weights
Dow manager, 1983
Wrong!
Correct, even in 1983: you NEED NOT
always measure ABSOLUTE molecular
weightsbut you could have.
Correct in 2005: it is almost always
essential to measure absolute M s.

2
So, how would you analyze
polymers and nanoparticles for
polydispersity?
Here is the easiest equation in this talk.
SEC = GPC = GFC
Size Exclusion Chromatography
Gel Permeation Chromatography
Gel Filtration Chromatography
4
A riddle:
After a hurricane, many trees fall over and
bend into a river. Also, a cow and a dog
fall into that flooded river. Which one
reaches the ocean first, cow or dog?
Moo!
Woof!
5
In GPC, as when throwing a cat through
a tree, the big things come out first..
Solvent flow carries molecules from left to right; big ones come
out first while small ones get caught in the pores.
It is thought that particle volume controls the order of elution.
But what about shape?
6
Simple SEC is only simple when you
dont have to do it yourself.
degas
pump
injector
DRI
V
e
l
o
g
1
0
M

c

c

l
o
g
1
0
M

c

log
10
M

7
In simple GPC, you first make a
map (calibrate), then follow it.
V
e

log M
V
o

V
e

logM
A
DRI
V
e
A

DRI
A

M
A

8
Are you straight?
GPC Calibration
log(M
w
) = 0.0009V
e
3
- 0.0534V
e
2
+ 0.6303V
e
+ 6.8161
R
2
= 0.9996
0
20
40
60
80
100
120
14 16 18 20 22 24 26 28
V
e
[mL]
D
e
t
e
c
t
o
r

R
e
s
p
o
n
s
e
0
1
2
3
4
5
6
7
8
l
o
g
(
M
w
)
9
Use the right tool (column) for the job.
10
OK, but how did you get the M
values for the standards?
Osmometry
Scattering
MALDI
Analytical ultracentrifugation
End group analysis
Goal in this class: make it seem reasonable
that it IS possible to measure M values.
11
Osmometry is Real Science because it is
tied closely to thermodynamics.
Semipermeable membrane: stops polymers, passes solvent.
h
tV = n R T
n = g/M
c = g/V
t = c R T

...)
1
(
2
+ + c A
M
12
Light scattering operates on thermodynamics; think
of it as an osmometer without the membrane.
100,000
x
c
...) 2
1
(
2
,
1
+ + =
|
.
|

\
|
c
c

c A
M
cRT
c
I
p T
s
t
q
t 2
) 2 / sin(
4
u

o
n
q =
u
Teaching Light Scattering to Exemplify and Reinforce Basic Principles, D. S. Poche', P. S. Russo, B. Fong, E. Temyanko and H. Ricks, J. Chem. Ed., 1999, 76 (November), 1534-1538.
13
LS adds optical effects Size.
q = 0 in phase
I
s
maximum
q > 0 out of phase,
I
s
goes down 3
1
2 2
g
s
R q
I ~
14
Just add LS detectorand much
complexity.
degas
pump
injector
DRI
DRI
MALLS
15
This waterfall plot shows many slices of a chroma-
togram: 13 angles were recorded ~8 times per
second as the sample flowed by the MALS detector.

3D Plot - PBLG
4
5
6
7
8
9
10
11
12
13
14
15
16
Scattered intensity
16
The scattering envelope for a single
slice shows how I
s
decreases with angle.
0.0 0.2 0.4 0.6 0.8 1.0
0
20000
40000
60000
80000
100000
120000
140000
c = 0.044 mg/mL
M = 130000 g/mol
R
/
K
c
sin
2
(u /2)
InterceptMolecular weight
SlopeSize
17
We add a viscometer in some cases.
degas
pump
injector
DRI
LS90
o
AP q
viscometer
18
Intrinsic viscosity is a secondary molecular weight
method so good its almost like the real thing.
Mark-Houwink-Sakurada
relationship between the
intrinsic viscosity and the
molecular weight.


a
KM = ] [q
K and a are constants for a
given polymer, not strongly
dependent on solvent or
temperature, as long as were
talking about a good solvent.
These words have special
meaning in polymer science.
a
K M
/ 1
) / ] ([q =
19
Grubisic, Rempp & Benoit,
JPS Pt. B, 5, 753 (1967)
One of of the most important
papers in polymer science.
Imagine the work involved!
6 pages long w/ 2 figures.
Selected for JPS 50
th
Anniv. Issue.
Universal Calibration lets you get the molecular weight of
one kind of polymer knowing only the Mark-Houwink-
Sakurada values of a standard (look it up) and your
unknown (uh-oh).
20
Universal Calibration says that
whatever comes out at a particular
volume has the same product , [q]M.
[q] = KM
a


[q]
A
M
A
= [q]
S
M
S
= f (V
e
)

1 1 + +
=
S A
a
S S
a
A A
M K M K
Mark-Houwink Relation
Universal Calibration
A = analyte; S = standard
Combine to get these two
equations, useful only if
universal calibration works!
1
1
1
+
+
|
|
.
|

\
|
=
A
S
S
a
A
a
S
A
K
M K
M
21
They were young when GPC was.
They were young when GPC hit
middle age.
Here is a small subset of GPC spare parts.
To say nothing of unions, adapters, ferrules, tubing (low pressure and
high pressure), filters and their internal parts, frits, degassers, injector
spare parts, solvent inlet manifold parts, columns, pre-columns,
pressure transducers, sapphire plunger, and on it goes
24
Other SEC Deficiencies
0.05 M salt at 10 am, 0.1 M salt at 2 pm?
45
o
C at 8 am and 50
o
C at noon?
Non-size exclusion mechanisms: binding.
Big, bulky and slow (typically 30
minutes/sample).
Temperature/harsh solvents no fun.
You learn nothing by calibrating.
Columns are expensive, easily damaged.


25
Must we separate em to size em?
Your local constabulary probably
doesnt think so.

Atlanta, GA
I-85N at
Shallowford Rd.
Sat. 1/27/01 4 pm
26
Matrix Fluorescence Photobleaching Recovery* is an
LSU-invented method designed to compete with GPC
for certain systems (aqueous commodity polymers).
Indicates targeted M.
1000 10000 100000 1000000
0.0
0.2
0.4
0.6
0.8
1.0
Pullulan, 8% HPC Solution, M=12,200 and 48,000
CONTIN
Exponential
Exponential
F
A
r
b
i
t
r
a
r
y

U
n
i
t
s
M
*G. J. Doucet, D. Dorman, R. Cueto, D. Neau, P. S. Russo, D. DeKee, J. Pople Macromolecules 2006, 39(26), 9446 9455.
27
Ultimate Goal: A Black Box for MWD
Matrix FPR
Easily Maintained
Accurate
Precise
Simple Concept
Expedient
Easy System Switch
Basic Info Obtained
Miniaturizable
Detect Large Masses
Labeling Required
GPC
Accurate
Simple Concept
Miniaturizable
No Labeling Required
Broad Distributions
Pumps
Parts
DLS
Form Factor
Index Matching
Long Acquisition for
Multiangle Experiments
Precise
Accurate
DOSY
Easy System Switch
Precise
Accurate
Obtain Basic Info
Labeling Required
28
In other words, the search continues.
Two promising contenders are discussed next.
MALDI-TOF: most effective when the
molecules are small, biological and not
very polydisperse. Can be coupled to GPC!
FFF: like GPC only a flowing field replaces
the stationary phase, stuff comes out
backwards, and big stuff can be handled
as well as small.
29
MALDI-TOF stands for Matrix-Assisted Laser
Desorption Ionization Time-of-Flight Mass
Spectrometry.
http://www.astbury.leeds.ac.uk/Facil/MStut/mstutorial.htm
30
These data obtained at LSU: click
the figure to analyze these results.
0 2000 4000 6000 8000 10000 12000
0
100
200
300
400
File will download from the website
Compute Mn,Mw,Mz MANUALLY
Compute Mn,Mw,Mz in Excel or something else
You can subtract the mass of silver from each peak
We hope Dr. Tracy McCarley will give a tour of the MALDI
a.i.
C
o
u
n
t
s
Mass
Guess what M
w
/M
n
is.
31
Patience is a virtue.
You 4010 students will get to practice with a
MALDI dataset, but .

thats enough MALDI for now.
32
What about separating cows and
elephants? Either will float around the
trees. How do you separate them then?
Moo!
Eeee!
33
Field Flow Fractionation, thats how!
In FFF, large nanoparticles are made
to flow between plates; a field is
applied to separate them by size.
34
The most commonly used field is flow itself: one or
both plates are porous, and a cross-flow is arranged.
35
What happens because of the
cross-flow?
Little nanoparticles come out first!
At LSU, only one plate is porous.
1. Everyone calls it AF4 = Asymmetric Flow Field Flow Fractionation
2. How to you get a crossflow then?
36
Potential Advantages of FFF
Handles a wider range of particles.
May be easier for some aggressive solvents.

37
30 35 40
40
60
80
100
120
140
160
180
0.0
0.2
0.4
0.6
0.8
1.0
RMS radius vs Time
AF
4
separation of algoflon
tm
LS 90
o

r
m
s

r
a
d
i
u
s

(
n
m
)
time (min)

r
e
l
a
t
i
v
e

i
n
t
e
n
s
i
t
y
AF4 can even separate large PTFE
latex particles.
38
Conclusion
GPC is essential in any Nano Lab
GPC may eventually get replaced.
Matrix FPR
FFF
39