Tools for the Characterization of Biomass at the Nanometer Scale - The Nanoscience and Technology of Renewable Biomaterials - page 64

Accurate concentration measurement is critical when using the solute exclusion tech-
nique. Probe molecule concentrations have been measured by interference refractometer
(8), spectropolarimeter (12, 13), and HPLC using a refractive index detector (14).
The validity and accuracy of the solute exclusion technique for pore size and volume
distribution measurements are based on two assumptions: (1) the concentration of a
probe molecule in the accessible pores is the same as in bulk solution surrounding the
pulp specimen, and (2) probe molecules can fully penetrate the pore to get full access
to the pore water. To meet these two assumptions, the probe molecules should not
adsorb on nor chemically react with the substrate. They should be spherical and must
be available in a large variety of highly monodisperse molecular sizes. Cross-linked
dextrans (10) and poly(ethyleneglycol)s (11) were originally proposed. Both of these
probe types have seen continued use (12-15). Gel permeation chromatography indi-
cated that complete penetration of a pore is not possible (16). The concentration of
the probe molecule in pores is shown both theoretically (17, 18) and experimentally
(19-21) to vary with pore shape and relative size of the pore and probe molecule.
Probe molecules with low hydrogen bonding capability have limited potential to access
water in pores, and electrostatic charge on the probe can effect the ability to penetrate
pores (22).
Other problems with determining pore size and volume distribution with the solute
exclusion technique are the 'ink-bottle' effect and osmotic pressure (23) (Figure 3.1).
Probe molecules can be excluded from the water in pores with a narrow opening but
wider space inside the pore. If the pore substrate contains ionized groups, even nonionic
solutes can be excluded from pores by osmotic pressure. In this case, using molecules
that interact and adsorb on fibers has been suggested as they can be forced to enter pores
(24). Cell walls appear to have a lamellar structure (25), and the simple slit model for
cell wall pores is a reasonable assumption (8, 10).
In view of the discussion above, we should emphasize the terms 'effective pore size'
and 'accessible water' when measured by the solute exclusion technique. The solute
exclusion technique is a valuable tool for determining the total pore volume accessible
to a probe molecule of given size. It can also quantify relative changes in the porous
structure from various treatments and between different substrates. Solute exclusion is
not an acceptable tool for the determination of absolute pore size and volume distribution.
In this sense this technique fits the needs for practical applications in bionanosensing,
such as determining enzyme accessibility to substrates and comparing the effectiveness of
various mechanical and chemical pretreatment processes and substrates for lignocellulose
bioconversion.
Despite the shortcoming of the solute exclusion technique, it is very useful for under-
standing the effects of molecular size on accessibility (26, 27). In addition, a very useful
pore structure model is based on the results of the method (10).

3.3.1.2 Porosimetry by Differential Scanning Calorimetry

The differential scanning calorimetry (DSC) technique for pore size distribution mea-
surements is based on the principle that water contained inside pores has a lower freezing
point than that of bulk water. This technique, called thermoporosimetry, was initially
developed for measuring pore size distribution in other materials (28-30) but has been