Spectrochimica Acta Part A 55 (1999) 883 – 894
Solid state 1H NMR studies of cell wall materials of
potatoes
Huiru Tang, Peter S. Belton *, Annie Ng, Keith W. Waldron, Peter Ryden
Institute of Food Research, Norwich Research Park, Colney Lane, Colney, Norwich NR4 7UA, UK
Received 23 April 1998; accepted 22 July 1998
Abstract
Cell wall materials from potatoes (Solanum tuberosum) prepared by two different methods have been studied using
NMR proton relaxation times. Spin lattice relaxation in both the rotating and laboratory frames as well as transverse
relaxation have been measured over a range of temperatures and hydration levels. It was observed that the sample
prepared using a DMSO extraction showed anomalous behaviour of spin lattice relaxation in the laboratory frame
probably due to residual solvent in the sample. Spin lattice relaxation in the laboratory frame is the result of
hydroxymethyl rotation and another unidentified high frequency motion. In the rotating frame relaxation is
adequately explained by hydroxymethyl rotation alone. In neither experiment is methyl group rotation observed,
calculation suggests that this is due to the low density of methyl groups in the sample. Non-freezing water in potato
cell walls, a-cellulose and pectin was found about 0.2, 0.04 and 0.18 g per gram dry matter, indicating preferable
hydration of pectin compared to cellulose. The effects of hydration are most noticeable in the measurements that
reflect low frequency motions, particularly transverse relaxation, where both second moments and the relative
intensity of signals arising from immobile material are reduced by hydration. © 1999 Elsevier Science B.V. All rights
reserved.
Keywords: Cell walls; Potato; Molecular dynamics; Hydration
1. Introduction
Cell walls not only play important roles in
various activities of plant cells, such as protection,
cell expansion and mass transportation [1], but
also act as one of the most important components
in many foods. Dietary fibres [2], which are be-
* Corresponding author. Fax: + 44-1603-458939; e-mail:
[email protected].
lieved to have a number of beneficial functions as
food, are primarily cell wall materials. Texture of
foods is probably also dependent on the state of
cell wall materials [3–5]. However, the complexity
of the assembly of cell walls [1,6,7] and the structure of each component have long been the major
difficulties in attempts to understand the behaviour of cell walls at the molecular level.
Cell walls chiefly consist of cellulose, hemicellulose, pectin, a small amount of proteins and
polyphenolic compounds [1,2,6,7]. Cellulose forms
1386-1425/99/$ - see front matter © 1999 Elsevier Science B.V. All rights reserved.
PII: S 1 3 8 6 - 1 4 2 5 ( 9 8 ) 0 0 2 3 5 - 2
884
H. Tang et al. / Spectrochimica Acta Part A 55 (1999) 883–894
the structural framework of cell walls whereas the
rest of the components interact with cellulose to
hold the whole structure together. However, these
models of cell walls, deduced largely from fractionation of plant cell wall materials and microscopic investigations, do not carry indications
about the molecular mobility of the cell wall
components, though the mechanical properties of
cell walls are likely to be dependent on these
properties.
NMR is a non-invasive and non-destructive
method of investigating molecular structure and
dynamics. It has been employed in obtaining information related to both structure and dynamics
of plant cell walls [8 – 17]. Relaxation time measurements with 1H NMR have been used to characterise the molecular mobility of cell wall
components [8 – 10] and the physical changes in
the cell walls of living cells. 13C solid state NMR
techniques have been employed to probe structural characteristics or different components of
cell walls [12 – 17]. The techniques used in studies
of synthetic polymers have proved to be useful in
cell wall research. An excellent example has been
given recently in mobility-resolved studies on hydrated cell walls of onion, tomato, pea stem and
tobacco leaves [17]. Examples of correlating proton relaxation times and structural features have
been given recently [14,15]. One problem with
most of the data published so far is that it has
been gathered on cell walls from different sources
and/or from different methods of preparation.
Measurements have also been made under different conditions of temperature and water content.
This makes it extremely difficult to compare the
results with each other.
In this paper we report on a systematic study of
the effects of hydration and temperature on the
proton relaxation behaviour of potato cell walls.
In addition, the likely variability of proton relaxation behaviour with sample is assessed by comparison with a sample of potato cell walls
prepared by an alternative method to the main
sample. The results obtained have significance not
only for other proton relaxation studies of cell
walls but also for cross polarisation measurements
since the nature of the 13C cross polarisation
signal is dependent upon the proton relaxation
characteristics of the sample.
2. Experimental
2.1. Preparation of cell wall materials
Cell wall materials of potatoes (P) were obtained from two preparation methods described
previously [5,18] and referred to as PA [5] and PB
[18], respectively. PA was dried as described by
Newman et al. [19].
Cell walls were all dried in a vacuum oven over
P2O5, at 40°C, for at least 24 h. Samples were
used only after storage over P2O5 in a desiccator
under vacuum for at least 12 h. D2O exchanged
PB was prepared by submerging the corresponding materials in excess of D2O for at least 2 h
before lyophilisation. The process was repeated at
least three times. These samples were then dried
and stored as described earlier. Rehydration of
them with D2O were carried out in a sealed jar
with controlled relative humidity. Hydration levels, h, are defined as the number of grams of
water per gram of dry matter and determined
gravimetrically by the difference between the hydrated sample and the dry sample (h of the dry
cell walls is about 0.02 as measured with Karl–
Fisher titration method). The PB vacuum dried
from H2O is referred to as PBH whereas the D2O
exchanged PB containing D2O are referred to as
PB1 (h 0.02), PB2 (0.26), PB3 (0.4), PB4 (0.8)
and PB5 (4.1).
2.2. NMR measurements
All NMR measurements were carried out on a
Bruker MSL 100 spectrometer equipped with high
power probe heads, operating at 100.13 MHz for
1
H and 15.37 MHz for 2D. The 90° pulse length
was 1–1.5 ms for proton and 14 ms for deuterium
experiments. Temperatures were regulated using a
variable temperature unit, B-VT-1000. 1H relaxation time measurements were always started
from the lowest temperature in 10 K steps
whereas 2D NMR was conducted from 300 to 220
K in 3 K steps. Duration of 15 min was allowed
for temperature equilibration.
H. Tang et al. / Spectrochimica Acta Part A 55 (1999) 883–894
2.3. Non-freezing water
Proton spin-lattice relaxation time in the rotating frame, T1r, was measured using a standard
spin-lock sequence (90°-tspin-lock-aq). The 90° pulse
length was 4 ms for variable temperature and
hydration experiments whilst 2 and 6.25 ms were
also used for PB1. Single point sampling was
carried out on the top of each FID 5–10 ms after
the spin-lock pulse. A total of 64–128 data points
were collected with a t increment of 400–500 ms.
The spin-lock field used were equivalent to either
40 or 67 or 125 kHz. Proton residual second
moment, M2r, was measured from proton FID
using either one pulse or from a solid echo (90°0 t1-90°90-t2-aq), which can avoid signal loss during
the dead time. In the single pulse experiment,
dead time was about 4 ms and dwell time was
0.5–1 ms. In the solid echo experiments, t1 was
4–7 ms and t2 was 7–12 ms. The FIDs or the
decay part of echoes were fitted to a modified
Gaussian function [20]. The delay t2 in solid echo
experiments was chosen to be longer than probe
ring down time (3–4 ms). All relaxation time
values were obtained using a Levenburg-Marquardt non-linear curve-fitting routine installed in
TableCurve™ 2D (Jandel Scientific).
Non-freezing water was measured for a-cellulose (Sigma), pectin (Sigma from Citrus Fruit,
degree of esterification, 60%) and PB using 2D
NMR. On-resonance free induction decays (FIDs)
were acquired 50 ms after a single 90° pulse with a
recycle delay of 1 s. Signal intensities were corrected for temperature effect on magnetisation
using the factor "vo/2kT where " is Planck’s
constant, k the Boltzmann constant and vo the
Larmor frequency of 2D. Three PB samples were
measured with h (D2O) of 0.40, 0.81 and 4.10,
respectively; three cellulose samples (h 1.03, 1.76
and 3.62) and a pectin sample (h 1.18) were also
measured.
2.4. 1H relaxation time measurements
Proton T1’s were measured using an inversionrecovery (IR) pulse sequence (180°-t-90°-aq) or a
saturation recovery (SR) sequence, [(90°-t1-90°)n t-90°-aq] when T1 was longer than 2 s; single
point sampling was carried out on the top of the
FID 5 ms after the reading pulse. Delay t1 and t in
SR sequence were typically 25 – 30 ms and 0.2–1 s,
respectively with 64 – 128 data points acquired,
whereas in IR experiments the delay was calculated according to the following equation
3. Results and discussion
t =t0(10)(n − 1/12),
Two potato cell wall samples have been used in
this study. For presentation convenience they are
named PA and PB, respectively (see experimental
section for details) and their chemical composition is given in Table 1. Two thirds of potato cell
walls are pectic materials, of which one third is in
the form of uronic acids, about 60% of the uronic
acids are methylated as esters. About 30% of the
so that a sufficient spread of relaxation delays
enabled multiple-exponential decay to be detected. to was 1 – 3 ms and n is the number of
delays; typically 48 data points were acquired.
Recycle delay in SR experiments was 0.5–1 s
whilst it was chosen to be at least five times longer
than the longest component T1 in IR experiments.
Table 1
Carbohydrate composition of potato cell well materials
Samples
PAa
PB
a
885
Carbohydrate (mg/mg)
Degree of methylation (%)
Rha
Fuc
Ara
Xyl
Man
Gal
Glc
UA
14a
9.5
–
2.3
75
47.8
17
17.3
7
9.8
288
311.9
339
321.1
240
219.3
Deoxyhexose: mostly Rha, but contained some Fuc (see Ryden and Selvendran [18]).
63
60
886
H. Tang et al. / Spectrochimica Acta Part A 55 (1999) 883–894
Fig. 1. Spin-lattice relaxation rate of cell wall materials, R1L, as a function of temperature. The insert is an expansion of PB1 and
PBH data at 150–350 K. PA-potato cell walls prepared with method A, dried under vacuum; PBH-potato cell walls prepared with
method B, dried under vacuum; PB1-PB four-time freeze-dried from D2O followed with vacuum drying.
potato cell wall material is in the form of
cellulose.
The amount of non-freezing water measured
with 2H NMR at 260 K was found to be 0.16–
0.22 g per gram dry matter for PB. The amount of
non-freezing water for a-cellulose was 0.01–0.04
gram per gram of dry matter and for pectin,
having a similar degree of esterification to that in
PB, was 0.18 gram per gram. This indicates that
most of the non-freezing water in cell walls was
associated with the pectic materials whilst only
little was associated with cellulose. This is not
surprising since significant amounts of cellulose
are expected to be present in anhydrous crystalline form.
The spin-lattice relaxation in the laboratory
frame of vacuum dried potato cell walls, PA and
PBH (dried from H2O), showed bi-exponential
decays over the temperature range studied (100–
360 K). The long component is designated T1L
and short one T1S. The rates of these two relaxation components were called R1L and R1S, respectively. The relative fraction of T1L is referred
to as FT1L. For PB, neither D2O exchange nor
substantial D2O hydration (up to h 4.10)
changed the bi-exponential behaviour. If a common spin temperature is maintained throughout a
solid by rapid spin-diffusion, a single proton T1
would be observed. Bi-exponential relaxation behaviour of cell wall materials implies the existence
of at least two separate domains whose size is
such that spin diffusion exchange is slow on the
time scale of T1L.
Fig. 1 shows R1L values of PA and PB as a
function of temperature. It is apparent that the
relaxation behaviour of PA is dramatically different from either of the PB samples. There is some
convergence of behaviour at the high temperature
end of the graph but increasing divergence as the
temperature is lowered. Below 300 K the rate of
relaxation of PA increases. There appears to be a
levelling off around 150–200 K, followed by another very rapid increase. Low temperature maxima in spin lattice relaxation are often associated
with methyl group rotation and it is likely that the
low temperature peak is due to residual DMSO
used in the extraction process. It is also worthnoting that the CH3 groups in the form of esters
H. Tang et al. / Spectrochimica Acta Part A 55 (1999) 883–894
in pectin might be expected to show low temperature methyl maxima [21], however, these are not
observed for PBH and PB1 samples (we discuss
the reasons for this later). The possibility that
extraction with DMSO has in some way mobilised
pectin methoxy groups cannot be discounted but
evidence for residual DMSO has been obtained in
both proton and carbon magic angle spinning
spectra (P.S. Belton, R. Boetzel, A. Gil and P.
Ryden, unpublished results). Because of the likely
contamination of the sample with DMSO no further work on type A preparations was pursued.
However the results do indicate that great care
must be taken in the choice of preparation
method and comparison between samples.
PB1 and PBH shows a relatively small change
over a wide range of temperature. Notable is the
absence of a low temperature maximum due to
methyl group rotation. The expected contribution
of methyl group rotation can be calculated according to Kubo and Tomita [22], for dipolar
relaxation characterised by an exponential correlation function.
R1 = C
n
tc
4t
+
1+ v 20t 2c 1 +4v 20t 2c
(1)
where tc is the rotational correlation time and can
be described by an Arhennius process thus
tc =t0 e(Ea/RT)
(2)
where R is the ideal gas constant and Ea is the
activation energy. Therefore a motion is characterised by a relaxation peak in R1 curve as a
function of temperature where v0tc is approximately 0.62 [20]. The expression as given assumes
only one correlation time and it would be expected that there would be distribution of correlation times in such a complex system as plant cell
walls. However, the single correlation time assumption allows an estimate of the maximum
contribution of methyl group rotation to relaxation. The relaxation constant C is associated
with the second moment reduction modulated by
motion and can be calculated [23,24] by:
C=
3 n m0
10 N 4p
2
6
g 4' 2r −
jk
(3)
887
where n is the number of protons involved in
motions, N the total proton number to be relaxed,
mo the permeability of a vacuum, g the magnetogyric ratio and rjk the interproton distance.
Chemical analysis (Table 1) of the potato cell
wall materials suggests that only 3% of the protons are present in methoxy groups. Rotational
motion of this type of methyl group often leads to
an R1 maximum at 80–100 K [21]. If interproton
distances (r) in methoxy groups of cell walls are
similar to that in methyl-a-D-galacturonic acid
methyl ester [25] which is 1.77 Å on average, then
the value of C can be estimated to be 1.67×108
s − 2. Assuming that no other groups contribute to
relaxation the predicted value of R1 at the maximum is about 0.4 s − 1. The motion of methyl
groups in rhamnose/fucose often results in a R1
maximum at higher temperature (120–200 K) because the protons there have immediate neighbours. However, the proportion of this type of
protons accounts for only less than 0.4% of all
protons. Given the same assumption about interproton distances and single correlation time, the
relaxation constant for them would be 3× 107 s − 2
and their predicted contribution to R1 is less than
0.1 s − 1. Moreover the value of 0.4 and 0.1 s − 1
are both over-estimates. Both types of methyl
groups probably will experience a variety of environments. The contribution to relaxation will thus
be smeared out over a considerable temperature
range. Given that the contribution to relaxation is
weak, it is not surprising that a maximum due to
methyl group rotation is not observed.
At high temperatures there is a clear tendency
towards an increase in relaxation rate in both PB1
and PBH. The behaviours of PB1 and PBH do
however differ slightly. At low temperatures PB1
is more slowly relaxing than PBH, but this is
reversed at high temperature. This can arise from
rapidly relaxing groups, containing non-exchangeable protons, acting as a relaxation sink for
groups containing exchangeable protons. The actual situation in cell walls is complex, but if only
two contributing groups are considered the behaviour may be illustrated by considering the
equation: R1L = PERE + PNRN, where the subscripts refer to exchangeable and non-exchangeable protons, P and R are the relative populations
888
H. Tang et al. / Spectrochimica Acta Part A 55 (1999) 883–894
and rates, respectively. The sum of PE and PN is
unity. If RN \RE and PN B1, then the effect of
term PNRN will be to make R1L BRN. When the
protons from the exchangeable protons are replaced by deuterium, PN =1 and R1L =RN, thus
the observed relaxation rate will increase. The
observation that the deuterium exchanged samples relaxes faster than the protonated sample is
thus explained if the relaxation of the exchangeable protons is slower than the non-exchangeable
protons. Similarly, at low temperatures RN B RE
and exchange decreases the relaxation rate.
The upward trend observed in both samples
might be attributed to trans-gauche rearrangements of hydroxymethylene groups, which can be
a source of spin-lattice relaxation [26 – 28]. These
typically exhibit rate maxima above room temperature. In practice, hydroxymethyl groups probably will experience a variety of environments, thus
the contribution to relaxation will appear over a
considerable temperature range this will result in
a smaller contribution to relaxation at the maximum. The behaviour of the short component, T1S,
follows general trends of the long relaxation time
process. Above 300 K the data is very scattered.
This is because the fraction of the short component is small and fitting error becomes large.
Nevertheless the overall trends are clear.
For PBH, FT1L accounted for 70 – 75% of the
population below 200 K and 85% above 300 K.
Since strong dipole – dipole coupling results in
faster spin diffusion, it would be expected to be
faster at lower temperature than at high temperature where occurrence of molecular motions often
weakens dipolar coupling. However, all three cell
wall materials showed more clearly defined bi-exponential relaxation behaviour for T1 at low temperature rather than at the high temperature
region. This implies that uniformity of T1 at
higher temperature arises from a convergence of
the relaxation times rather than from mixing by
spin diffusion.
Fig. 2a shows the hydration effects on, R1L, of
potato cell walls (PB) in the temperature region of
100–360 K. R1L increases as temperature increases, indicating a maximum at the high temperature although that is not reached at 360 K; there
is an indication of a flattening in the highest water
content sample. This motional process probably
corresponds to the reorientation of hydroxymethyl groups as discussed above. The general
effect of increasing hydration above 250 K is to
first increase then to decrease the relaxation rate
of the long component (R1L). Below 250 K the
values of R1L tend to converge. This is consistent
with the non-freezing water measurements which
imply that all the samples except PB1 have the
same liquid water content below the freezing transition. The results from PB5 are slightly higher,
this may be due to extensive ice crystal formation
causing damage to the cell walls. At low temperature the relative proportion of the long component is constant around 75% up to 200K, after
this it rises up to 90% (Fig. 2b). The scatter of the
data is considerable because of the problem of
fitting a curve where one component represents
most of the magnetisation. However it does appear that the least hydrated sample shows the
smallest change with temperature. As discussed
above the tendency to move towards single exponential relaxation at higher temperatures is inconsistent with the effects of dipolar spin diffusion. In
a hydrated system spin mixing may occur by
chemical exchange, but since all the exchangeable
protons have been replaced by deuterons in this
system this is not a viable mechanism.
In order to interpret the changes observed in
the relaxation curves at different hydration it is
necessary to consider the effects of hydration on
the polymers. In general hydration will result in
increased polymer mobility due to the plasticisation effects of water. Plasticisation will allow the
adjustment of polymer conformations, frozen in,
in the dehydrated state to minimum energy conformers. This will result in a decrease in the
distribution of conformers and thus to a decrease
in the distribution of correlation times. This results in a sharpening of the relaxation rate curves
and an increase in their intensity at the maxima.
At high hydration levels there will be increases in
polymer mobility such that the rate maxima may
be reduced to below 273 K. In this case there will
be a decrease in relaxation rate at the highest
temperatures for the most hydrated materials.
This is seen in samples PB4 and PB5. However,
below freezing temperature this effect is obscured
H. Tang et al. / Spectrochimica Acta Part A 55 (1999) 883–894
since freezing reduces the amount of liquid water
available to plasticise the polymers to the nonfreezing water content. At high temperatures and
high hydration levels, some of the polymers will
be dissolved, this will result in a considerable
increase in mobility.
Spin-lattice relaxation in the rotating frame
(T1r ) is sensitive to the motions comparable to the
889
frequency related to the spin-lock field [29] (104 –
105 Hz). Two T1r components can be detected for
all PB samples with the longer one designated
T1rL and short one T1rS. The corresponding rates
are R1rL and R1rS, respectively, and the fraction
of magnetisation relaxing with a time constant
T1rL is FT1rL. Fig. 3 shows R1rL of dry PB1 as a
function of temperature and spin-lock field. A
Fig. 2. (a) Spin-lattice relaxation rate of potato cell walls (PB), R1L, as a function of temperature and hydration levels. PB1-vacuum
dried PB following four-time freeze-drying from D2O; PB2-D2O hydrated PB1 with a hydration level h 0.26; PB3-D2O hydrated
PB1 with a hydration level h 0.40; PB4-D2O hydrated PB1 with a hydration level of 0.81; PB5-D2O hydrated PB1 with a hydration
level of 4.10. (b)Percentage proportion of the long component of spin-lattice relaxation time of potato cell walls (PB), FT1L, as a
function of temperature and hydration levels.
H. Tang et al. / Spectrochimica Acta Part A 55 (1999) 883–894
890
Fig. 3. Spin-lattice relaxation rate of vacuum dried potato cell walls, PB1, in the rotating frame, R1rL, as a function of temperature
and spin-lock field.
single broad relaxation peak is observable with all
three spin-lock frequencies. One of the possibilities is that the relaxation peak is related to the
motions of CH2OH/D groups [27,28] in cell walls.
R1r can be calculated according to Eq. (4),
tc
3
5
tc
R1r = C
+
2 2
2 1 +4v 1 t c 3 1 + v 20 t 2c
+
2
tc
3 1+ 4v 20 t 2c
n
(4)
where C and tc are defined similar to these in Eqs.
(1) and (2), but v1 is the effective field for spinlattice relaxation [29].
If hydroxymethylene groups are the only T1r
relaxation source and they have a single correlation time, R1r can be estimated using Eq. (4).
Typical values for hydroxymethylene groups in
monosaccharides have been found to be 34–40 kJ
mol − 1 for Ea and 1.3 – 2 ×10 − 12 s for t0 [26–28].
The maxima are estimated to be 400 – 650 s − 1 at
300–370 K, 300 – 450 s − 1 at 300 – 370 K and
150–300 s − 1 at 320 – 380 K when vSP are 40, 67
and 125 kHz, respectively. It is apparent that the
experimental rates are less than half that of the
estimated rates and the peaks appear at lower
temperatures than predicted. As the hydrox-
ymethyl groups are experiencing a range of environments they will have a distribution of
correlation times, the curves will therefore be
broader and with a less intense maximum than
predicted from the single correlation time model,
some support for the distribution of correlation
times can obtained from the frequency dependence of the relaxation rates, since there is a field
dependence on both sides of the relaxation maxima, which would only be expected if there were
more than one correlation time contributing to
motion. Given a distribution of correlation times,
the discrepancy between estimated maxima and
experimental ones is not a unreasonable outcome.
There may also be contributions from sugar ring
puckering. Solution state studies [30] indicate that
in solution rates are of the correct order of magnitude. But no direct evidence yet exists to indicate
that such motions are retained in the solid state.
Fig. 4a shows R1rL values of PB as a function
of temperature and hydration level. Below the
region 265–275 K there is a general convergence
of behaviour. This is due to the effects of freezing,
below the freezing point liquid water contents of
samples containing freezable water are all reduced
to the same level. The effects of freezing are
particularly notable in sample PB5 where a sharp
H. Tang et al. / Spectrochimica Acta Part A 55 (1999) 883–894
break is apparent around the freezing point. In
PB1 there is not a freezing effect and a maximum
is apparent at about 250 K. Above the freezing
point there is a marked effect of hydration, at 300
K there is a 5-fold difference in relaxation rate
between the wettest and driest samples. This is
much greater than the effects seen in the T1 data,
however, the general trends are the same: there is
891
an indication that the maxima are moving to a
lower temperature as hydration is increased but
they are not observed because of the freezing
effects.
The changes in the relative population of the
R1rL fraction are shown in Fig. 4b. Once again it
differs from the laboratory frame data, in the
laboratory frame the greatest differences in popu-
Fig. 4. (a)Spin-lattice relaxation rate of potato cell walls (PB) in the rotating frame, R1rL, as a function of temperature and
hydration levels (see Fig. 2 for keys). (b)Percentage proportion of the long component of spin-lattice relaxation time of potato cell
walls (PB) in the rotating frame, FT1rL, as a function of temperature and hydration levels (see Fig. 2 for keys).
H. Tang et al. / Spectrochimica Acta Part A 55 (1999) 883–894
892
Fig. 5. Proton FIDs of potato cell walls (PB) at 300 K at different hydration levels (see Fig. 2 for keys), solid lines are fitted values.
lation are seen in the low temperature region
whereas in the rotating frame data the reverse is
true. The trend is quite weak for samples PB1 and
PB2, but for PB3 and PB4 there is a strong shift
in population above 250 K. Since the laboratory
frame relaxation rates are slower than the rotating
frame rates it would be expected that spin mixing
by spin diffusion would be more efficient in the
former. The almost single exponential relaxation
shown at low temperatures for the rotating frame
results suggests therefore that the differences seen
in T1 are not seen in T1r and that at low temperatures all sites in the sample have similar relaxation rates. PB5 shows distinctive behaviour. This
may be the result of the effects of large scale ice
crystal formation or could possibly be the effects
of chemical degradation or polymer solublisation
at high temperatures and water contents.
The baseline corrected FID can be fitted with
the function [20] in Eq. (5) as shown in Fig. 5:
I(t)=I1 exp
−a 2t 2
sin bt
t
+ I2 exp −
2
bt
T2e
(5)
Two components are assumed with I1 and I2
representing their signal intensities. The first term
consists of a Gaussian relaxation term modulated
by a sinusoidal term with a and b representing
relaxation parameters, this is typical of the transverse relaxation of protons in a rigid solid. The
second component is an exponential term and is
typical of protons in the motionally narrowed
regime of motion, it is characterised by a time
constant T2e. A useful way of characterising the
Gaussian term is by the second moment M2r
which can be calculated from the value of a and b
according to [20]: M2r = a 2 + b 2/3. The relative
proportion of the first component, FG, can be
calculated as FG = I1/(I1 + I2).
At a constant proton density, the value of M2r
depends on the degree to which motion in the
system results in reducing static dipolar interactions. Typically the value of M2r reduces as temperature increases. PB dried from D2O and H2O
(data not shown) showed slightly different M2r
reductions, of about 4 G2 (from 14 to 10) and 3
G2 (from 14 to 11), respectively, over the temperature range of 100–360 K. As correlation times
for motion move from the rigid lattice regime into
the motionally narrowed regime they begin to
contribute to spin lattice relaxation processes.
Fig. 6a shows the temperature dependence of
M2r of PB as a function of temperature and
hydration levels. There is a 3 G2 difference be-
H. Tang et al. / Spectrochimica Acta Part A 55 (1999) 883–894
tween PB1 and wet PB samples at very low temperature (100 – 200 K). For the most hydrated
sample there is a very strong reduction in the
second moment. This may well be associated with
the partial dissolution and or chemical degradation of the cell wall components disrupting the
structure of the wall.
The proportion of the Gaussian component is
shown in Fig. 6b. Whilst there is a modest effect
893
for dry PB at about 250–280 K, increasing hydration led to much greater effects. For the more
hydrated materials there is a sharp effect at the
freezing point. Below this temperature, as expected, all the samples behave in roughly the same
way due to their similar unfrozen water contents.
Above the freezing point increasing hydration led
to progressively smaller proportions of the Gaussian component. When h was about 4.10, FG
Fig. 6. (a)Proton second moments of PB as a function of temperature and hydration levels (see Fig. 2 for keys). (b)Percentage
proportion of the rigid component in the FID of PB as a function of temperature and hydration levels (see Fig. 2 for keys).
894
H. Tang et al. / Spectrochimica Acta Part A 55 (1999) 883–894
dropped to about 30% of the magnetisation, this
is equivalent to the amount of cellulose protons in
the system and may indicate that partial dissolution and disruption of the cell walls leaves only
the cellulose component unaffected. The relaxation constant, T2e, of the exponential component
showed a consistent increase, with temperature
and hydration above 270 K, suggesting the motion of the polymers is enhanced when hydration
level or temperature increased.
Acknowledgements
This work was supported by a Competitive
Strategic Grant of the BBSRC. We also thank an
anonymous referee for a number of constructive
comments.
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