PNNL-13501 - Pacific Northwest National Laboratory

In the current project, improved kinetic methods using
rapid-heating micro-cell reactors were introduced with the
goal of producing accurate kinetic measurement of
hydrolysis rates of sugar precursors. Experiments were
carried out in micro-reactors using quantitative 13 C
nuclear magnetic resonance (NMR). The purpose of
using NMR to monitor products was primarily to be able
to selectively detect monosaccharides in the presence of
aldehydic byproducts that may interfere with sugar
quantitation using conventional chromatography. Microcell
kinetic studies were carried out for a series of
oligomeric sugars using NMR and chromatographic
methods.
The key goal of fiber hydrolysis was to maximize the rate
of acid-catalyzed hydrolysis of starch, hemicellulose, and
cellulose with minimum or no ketone byproduct
formation. Byproduct ketones such as furfural and 2hydroxymethylfurfural
are produced by acid-catalyzed
dehydration of 5- and 6-carbon sugars, if excessive
conditions of acidity or temperature are employed. Thus,
a series of conditions of varying temperature and acidity
were examined using wet-milled corn fiber as well as a
series of disaccharide model structures. Near-optimal
hydrolysis conditions were applied to wet-milled corn
fiber and quantitative rates of sugar formation were
determined under conditions inducing no more than trace
formation of aldehydic dehydrations products. Analytical
procedures for carbohydrate products and for grain fiber
oils were refined.
Results and Accomplishments
Optimal conditions for hydrolysis of corn fiber were
identified and kinetics of formation of arabinose, xylose,
glucose, and galactose were determined. These
conditions allow precise sugar extraction to be designed.
Under optimal conditions (120°C, 0.2% v/v H2SO4),
arabinose is most rapidly formed, followed by xylose,
glucose, and galactose. Figure 1 shows about 25%
conversion of corn fiber by dry weight to
monosaccharides, with an additional 25% soluble
additional 25% soluble oligosaccharides produced (not
shown in Figure 1), corresponding primarily to
hemicellulose. Examination by NMR and
chromatography confirmed very low production of
aldehydic products under the identified conditions.
Kinetics of hydrolysis of disaccharide structures,
arabinobiose (Ab), xylobiose (Xb), cellobiose (Cb), and
lactose (L) were examined under optimal fiber conversion
conditions, as shown in Figure 2 for the hydrolysis of
xylobiose to xylose. Pseudo-first-order rate constants
422 FY 2000 Laboratory Directed Research and Development Annual Report
%Yield, g/g Fiber
Hydrolysis of Corn Fiber, 120 oC 0.2% Acid,
25mg Fiber/0.2mL Aqueous Phase
10
9
8
7
6
5
4
3
2
1
0
0 50 100 150
Time, min
Arabinose
Galactose
Xylose
Glucose
Figure 1. Hydrolysis of wet-milled corn fiber. Hydrolysis of
hemicellulose leads to early formation of arabinose.
0.01 6
0.01 4
0.01 2
0.01
0.008
0.006
0.004
0.002
0
Xylose
Xylobiose
0 10 20 30
Time, min
40 50 60
Figure 2. Kinetics of hydrolysis of xylobiose. At 120°C and
0.2%v/v H 2SO 4, xylobiose exhibits clean pseudo-first-order
disappearance with a rate constant 0.08 min -1 .
were found to be Ab < 0.1 min -1 , Xb 0.08 min -1 , Cb 0.01
min -1 , and L 0.03 min -1 . The kinetic methods used here
are now available for more exhaustive studies of the
response of fibers to selective hydrolysis conditions. A
typical 13 C NMR spectra of liquid corn fiber hydrolysate
is shown in Figure 3. Further work to determine the
kinetics of reaction of hemicellulose and xylan and
arabinan is in progress.
Finally, conditions for the analytical separation of
neutraceutical products (tocopherols, sitosterols) were
examined. Collection of 13 C NMR data for the subject
materials was begun, and chromatographic methods for
separation of the neutraceuticals were examined.
Procedures for isolation of neutraceuticals include
alkaline treatment of the wet-milled corn fiber to dissolve
hemicellulose and oils. Neutralization of this fraction
followed by ethanol extraction removes neutraceuticalcontaining
oils and yields isolated hemicellulose.
Characterization of parent corn fiber and hemicellulose by
solid-state 13 C NMR was carried out as a function of