Tools for the Characterization of Biomass at the Nanometer Scale - The Nanoscience and Technology of Renewable Biomaterials - page 68

3.3.2.3 Raman and Infrared Spectroscopy

Raman and IR spectroscopy have both been suggested as ways to determine the ratio of
amorphous to crystalline cellulose I. In Raman, a linear correlation has been observed
between X-ray crystallinity and I 1481 /( I 1481 +

I 1462 ) , where the peaks at 1481 cm − 1 and
1462 cm − 1 are assigned to crystalline and amorphous cellulose, respectively (52). Using
IR, the height or area of the crystalline band at 1280 cm − 1 is compared with the relatively
constant band at 1200 cm − 1 (53). In both methods the intensity of the bands need to be
determined through mathematical deconvolution of the spectra because of overlap. The
correspondence of Raman-derived values and X-ray crystallinity values is better when
the Raman crystallinity is calibrated on a contiguous set of cellulose samples rather than
cellulose from widely varying sources. Also, there is some indication that the Raman
method is not sensitive above crystallinity index of 75. The FTIR method does not
account for any potential changes in the ratio of cellulose I α to I β but otherwise is a
reasonable method for routine characterization.

3.4 Microscopy and Spectroscopy

So far we have described a variety of techniques for measuring specific attributes of
biomass. The remainder of this chapter will give an overview of microscopic and spec-
troscopic techniques useful in generating images and chemical information, respectively,
from biomass samples.

3.4.1 Specimen Preparation
Biomass has several characteristics that can make it difficult to analyze. We have already
discussed water interactions and how the methods of freezing or removing water from
biomass can alter nanostructure. The modest pyrolysis temperature of 300 ◦ C (54) for
cellulose, as well as long experience with organic substrate damage by electrons and
x-rays shows that biomass constituents are susceptible to change during observation by
high energy probes. While the aromatic nature of lignin distinguishes it from carbohy-
drates, cellulose and hemicellulose are chemically very similar and so are difficult to
differentiate.
The obvious approach to analyzing biomass structure by selectively removing partic-
ular components has serious problems. Removing one polymer from a plant cell wall
almost always causes chemical and physical changes in the polymer removed, as well
as the residual material. For example, the relatively gentle acid chlorite delignification
procedure only removes about half of lignin before damage to hemicellulose becomes
apparent (55). Amorphous cellulose and hemicellulose are so chemically similar that
it is very difficult to dissolve one without disturbing the natural state of the other. In
addition, chemical crosslinks (lignin-carbohydrate complexes, or LCC) between lignin
and hemicellulose are well documented and hinder the removal of hemicellulose (56).
These chemical considerations, as well as the interpenetrating nature of the cellulose,
hemicellulose, and lignin polymer networks, makes it very difficult to analyze the native
structure of any one of the wood components by itself.