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Comparative Analysis of Cellulose Preparation Techniques for Use with 13C, 14C, and 18O Isotopic Measurements
journal contribution
posted on 2005-11-15, 00:00 authored by Julia B. Gaudinski, Todd E. Dawson, Sylvie Quideau, Edward A. G. Schuur, John S. Roden, Susan E. Trumbore, Darren R. Sandquist, Se-Woung Oh, Roderick E. WasylishenA number of operationally defined methods exist for
pretreating plant tissues in order to measure C, N, and O
isotopes. Because these isotope measurements are used
to infer information about environmental conditions that
existed at the time of tissue growth, it is important that
these pretreatments remove compounds that may have
exchanged isotopes or have been synthesized after the
original formation of these tissues. In stable isotope
studies, many pretreatment methods focus on isolating
“cellulose” from the bulk tissue sample because cellulose
does not exchange C and O isotopes after original synthesis. We investigated the efficacy of three commonly
applied pretreatment methods, the Brendel method and
two variants of the Brendel method, the Jayme−Wise
method and successive acid/base/acid washes, for use
on three tissue types (wood, leaves, roots). We then
compared the effect of each method on C and O isotope
composition (13C, 14C, 18O), C and N content, and chemical composition of the residue produced (using 13C
nuclear magnetic resonance (NMR)). Our results raised
concerns over use of the Brendel method as published,
as it both added C and N to the sample and left a residue
that contains remnant lipids and waxes. Furthermore, this
method resulted in 18O values that are enriched relative
to the other methods. Modifying the Brendel method by
adding a NaOH step (wash) solved many of these problems. We also found that processed residues vary by
tissue type. For wood and root tissues, the 13C NMR
spectra and the 18O and 13C data showed only small
differences between residues for the Jayme−Wise and
modified Brendel methods. However, for leaf tissue, 13C
NMR data showed that Jayme−Wise pretreatments produced residues that are more chemically similar to
cellulose than the other methods. The acid/base/acid
washing method generated 13C NMR spectra with incomplete removal of lignin for all tissues tested and both
isotopic, and 13C NMR results confirmed that this method
should not be used if purified cellulose is desired.