Complex time dependence of the EPR signal 793

Regulla D., Bartolotta A., Deffner U., Onori S., Pantaloni Singer L. S. (1961) Electron spin resonance of complexes of
M. and Wieser A. (1993) Calibration network based on aromatic hydrocarbons with iodine. J. Chem. Phys, 34,

alanine/ESR dosimetry. Appl. Radiat. Isot. 44, 23-31. 133-140.

Singer L. S. (1959) Synthetic ruby as a secondary standard Thompson D. S. and Waugh J. S. (1965) Adjustable ruby
for the measurement of intensities in electron para- intensity standard for ESR spectra. Rev. Sci. Instrum. 36,
magnetic resonance. J. Appl. Phys. 30, 1463-1464. 552.


192 Vitaly Yu. Nagy and Mare F, Desrosiers

Alanine
signal

50 100
Time after irradiation, h

150 = 200

24800 Ruby
signal
2 24700
& 24600
24500.
24400
50 100 «6150 = 200
Time after irradiation, h
Ratio of the
0.916 alanine signal to
0.915 the ruby signal
0.914
£0913
B gota

30 100

150
Time after irradiation, h

200

Fig. 2. Effect of the compensation of the EPR spectrometer
instability by means of an adjacent ruby reference sample
(data for the 100 kGy sample).

amplitudes with relative standard deviations as small
as 0.1% or even better for doses above 100 Gy.
Although the quoted value refers to immobile alanine
pellets, this approach should also improve precision
of dosimetry techniques using uniform exchangeable
pellets.

In principle, any material with stable paramagnetic
centers and a non-overlapping EPR signal can

be used as an adjacent reference sample. Ruby is
particularly convenient because, in addition to being
highly stable, its EPR signal is strongly anisotropic,
and one can, by means of rotating the crystal,
locate a narrow chromium(3+) line almost at
any place in the spectrum. For best results, the
microwave power and modulation amplitude used
must be within the strictly linear ranges of the
dependencies of the amplitudes of both the signals on
these parameters. Additionally, the spectrum sweep
time should be minimized such that the interval
between recording the two spectra is as short as.
possible.

An adjacent reference sample should be reliably
fixed in the cavity. There are several commercial
cavities equipped with a special insert for such a
sample in the front wall. Unfortunately, they do not
cover all of the frequently used microwave modes. A.
convenient assembly bearing a rotatable ruby crystal
for the Varian V4531 cavity has previously been
described (Thompson and Waugh, 1965). The
commercial removable ruby-bearing front wall for
the Varian E-231 cavity is also convenient. The use
of the second hole of a dual cavity to insert an
adjacent standard results in noticeably poorer
reproducibility. Most probably, this is because of the
switching of the current between the two pairs of
modulation coils, which causes permanent thermal
instability of the cavity.

Although, theoretically, the adjacent standard
technique may not provide complete correction of
errors under conditions of very large variations of
cavity Q-values (Nagy, 1994), it is well-suited for
EPR dosimetry, where both the shapes and the
dielectric characteristics of the samples under
comparison are exactly the same,

References

Anderson J. H. and Weil J. A. (1959) Paramagnetic
resonance of color centers in germanium-doped quartz.
J. Chem. Phys. 31, 427-434.

~ ASTM (1995) Annual Book of Standards. E 1607-94

Practice for Use of the Alanine-EPR Dosimetry System,
pp. 846-851, Vol. 12.02. ASTM, Philadelphia, Pa.

Callens F., Van Laere K., Mondelaers W., Matthys P. and
Boesman E. (1996) A study of the composite character of
the ESR spectrum of alanine. Appl. Radiat. Isot. 47,

000-000.

Desrosiers M. F., Burlinska G., Kuppusamy P., Zweier J.,
Yaczko D. M., Auteri F. P., McClelland M. R., Dick
C. E. and McLaughlin W. L, (1995) Research and
development activities in electron paramagnetic reson-
ance dosimetry. Radiat. Phys. Chem. 46, 1181-1184.

Nagy V. (1994) Quantitative EPR: some of the most difficult
problems. App!. Magn. Reson. 6, 259-285,

Nette H. P., Onori S., Fattibene P., Reguila D. and Wieser
A. (1993) Coordinated research efforts for establishing an
international radiotherapy dose intercomparison service
based on the alanine/ESR system. Appl. Radiat. Isot. 44,
7.

Pilbrow J. R., Hutton D. R., Zhong Y. C., Nobie C. J. and
Song R. (1996) Pulsed ESR investigation of hyperfine
structure in y-irradiated alanine. Appl. Radiat. Isot. 47,
000-000.



Appl. Radiat. Isot. Vol. 47, No. 8, pp. 789-793, 1996
Published by Elsevier Science Ltd

Printed in Great Britain. All rights reserved
0969-8043/96 $15.00 + 0.00

© vv

‘S0969-8043(96)00053-X.

Complex Time Dependence of the EPR Signal
of Irradiated L-«-alanine

VITALY YU. NAGY and MARC F, DESROSIERS

Ionizing Radiation Division, Physics Laboratory, National Institute of Standards and Technology,
Gaithersburg, MD 20899-0001, U.S.A.

(Received 9 February 1996; in revised form 26 March 1996)

Measurements of the EPR signal amplitude of y-irradiated L-a-alanine with use of an adjacent reference
sample have revealed variations in the signal intensity within hours and days after irradiation. The
character of the time dependence of the amplitude varies. with dose and the amplitude changes reach
1-1.5%. This observation favors the hypothesis that irradiated alanine contains several paramagnetic
centers. Usefulness of adjacent reference samples in alanine dosimetry is also demonstrated. Published
by Elsevier Science Ltd

Introduction

The alanine system is now used for a transfer
dosimetry in’ primary and secondary standards
laboratories and as a routine tool in some industrial

each time immediately after the signal of a test
sample, the ratio of the signals in question will
provide a characteristic of the test sample intensity
that is free (or nearly free) of errors due to long-term
drift and other uncontrollable changes in the system

applications (Regulla er al., 1993; Nette et al., 1993;
ASTM, 1995). Improvements of its accuracy and
precision were so successful that, at present, even
errors below 1% are of importance. Under these
conditions, uncontrollable long-term drifting of
electron paramagnetic response (EPR) spectrometers,
which can be neglected in most other EPR
applications, becomes an important component of
inaccuracies. In fact, even sophisticated modern EPR
spectrometers typically do not provide a long-term
stability of a signal with variations in intensity less
than 0.5%, and signal variations as large as several
percent occasionally occur.

In order to further improve alanine dosimetry, it is
necessary to take these instabilities into account. An
effective method to do this has been proposed long
ago, when instabilities of EPR spectrometers were
much greater. It consists of placing a second,
immobile, paramagnetic sample with a different EPR
signal into the microwave cavity to serve as a

sensitivity.

Upon applying this technique to alanine dosimetry,
we uncovered a time dependence of the signal of
irradiated alanine during, hours and for several days
after irradiation, which is'gignificant for precise dose
measurements, These effects are especially important
in transfer dosimetry where several days can elapse
between exposure and instrumental evaluations.

Experimental

Algnine pellets* (4.9 mm diameter, 2.7 mm height)
containing 90% {by mass) L-a-alanine (Aldrich,
99+%) and 10% polyethylene (Polysciences,
MW = 700, 60 ym) as a binder, were irradiated with
“Co y-rays to absorbed doses from 0.01 to 100 kGy
under electron equilibrium conditions and controlled
temperature,

The pellets were prepared by the following

procedure. The bulk alanine crystals were ground
with a centrifugal mill (Brinkmann) fixed with a
0.5 mm ring sieve. Alanine crystals in the 53-125 ym
range were selected with a vibrating sieve
(Brinkmann). Appropriate weights of sieved alanine
and polyethylene were blended in a powder blender
(Paterson-Kelly}. The mixture was pressed into
pellets with a Manesty hand-tabletting press to
produce alanine dosimeters 4.9 mm in diameter with
an average height of 2.7 mm. The dosimeters were
placed in a 130°C oven for 30 min, foliowed by 5 min

reference during the measurements (Anderson et al.,
1959; Singer, 1959, 1961). Variations in the intensity
of the signal of the reference sample reflect
instrument instabilities, and, if this signal is recorded

*The mention of commercial products throughout this
paper does not imply recommendation or endorsement
by the National Institute of Standards and Technology,
nor does it imply that products identified are necessarily
‘the best available for this purpose.

789



790

in an 85°C oven. The dosimeters were stored in the
dark space at ambient temperature under controlled
humidity conditions (45 + 5)% r.h. For replicate
checking, irradiation of samples of the pure alanine
powder used in preparing the pellets has been
performed under the same conditions.

EPR measurements were taken at room tempera-
ture and relative humidity 55-60% with a Bruker
ECS106 spectrometer equipped with a 4108TMH/
8909 cavity. Because this cavity has no additional
holes in the side walls, the exposure of alanine
samples to light during measurements was negligible.
In the course of measurements, an alanine sample
was kept intact in the cavity, which also contained an
immobile smail single crystal of synthetic ruby (Al,0,
containing about 0.05% Cr*). The spectra of both
samples were recorded at time intervals ranging from
7 min to 1.5 h depending on the rate of change in the
alanine signal intensity.

The ruby signal was recorded each time immedi-
ately after the alanine measurement at exactly the
same settings of instrumental variables, except the
Zeeman field range and, in most cases, the receiver
gain. In each experiment, receiver gains for alanine
and ruby remained unchanged during the whole
measurement session. The microwave ‘power was at
the level of 0.25 mW, which does not cause saturation
of either of the two signals. Modulation amplitude
of 1.4mT was used in all of the measurements
to provide a good signal-to-noise ratio for the
lower-dose samples and comparability of results for
different doses. Prior to the measurement period,
the spectrometer was kept operating at the same
parameters (including the modulation amplitude) as
used in the experiment, for at least several hours.
First-derivative amplitudes were used as character-
istics of intensity for both signals. For alanine, the
central line in the powder spectrum was monitored,
whereas, for ruby, the line with the greatest intensity
at a selected crystal orientation was used.

Field sweep rate and time constant were selected to,

meet the manufacturer's guideline (time constant ten
times less than the peak-to-peak sweep time).

Results and Discussion

Time dependencies of the amplitude of the central
line in the irradiated alanine spectrum normalized by
the intensity of the ruby signal are presented in Fig. 1.
The data show that the intensity of the signal varies
with time and absorbed dose. At any dose, we
observed a sharp decrease in the signal intensity for
the first 1.5-2.5 h after irradiation: Because it took
several minutes to deliver a sample from the radiation
source to the EPR spectrometer and make the
necessary adjustments of the spectrometer, signal
evolution could not be observed from the very
beginning. Furthermore, because we tried to start
measurements as soon as possible in each case, the
initial moment of the measurement start was not

Vitaly Yu. Nagy and Marc F, Desrosiers

exactly the same in all the experiments, and one
should not directly compare the magnitudes of the
initial drops on the plots.

In the case of the 10 Gy sample, the overall drop
observed following the 13th minute after irradiation
is about 1.2%. Reasoning from the general principles
of chemical kinetics, one should assume an even
steeper signal decrease at the very initial period after
irradiation. The total percentage magnitude of the
dip seems to decrease with increasing dose, although
it is difficult to draw a firm conclusion on this point
without values of the signal just at the end of
irradiation.

In the dose range 10-1000 Gy, the signal remains
virtually stable after this initial drop, at least for 1-2
days. However, at the. doses above 1 kGy, a
subsequent increase in the intensity occurs. At 2 kGy,
the signal seems to increase steadily, at least for the
first 4 days. In the range 5-20 kGy, the signal
dependence on time exhibits a plateau beginning at
about the 70th hour (3 days after irradiation). The
signal remains constant for at least 3 more days. The
greatest observed. magnitude of the increase from the
dip to the saturation level .is about 1.5%.

The situation becomes even more complicated at
higher doses: after the drop and the increase, the
signal begins to fade again. The time for the
maximum seems to depend on the dose. At 25 kGy,
it is observed at about the 80th hour and at 100 kGy
at about 40 h. The decrease in 250 h is about 0.55%.

The described dependencies of the signal were.
observed both for pellets containing polyethylene as
a binder and for. pure alanine powders without
additives. t

The results clearly‘indicate that the widespread
concept of the irradiation product of alanine as a
unique and stable radical is overly simplified.
Obviously, there are chemical processes occurring in
alanine after irradiation that differ with absorbed
dose. This is consistent with the results of the other
studies that suggest that several paramagnetic centers
are present in the irradiated alanine (Callens er al.,
1996; Pilbrow et al., 1996; Desrosiers et'al., 1995).
Therefore, we conclude that the radicals present in
freshly irradiated alanine samples are not a unique
chemical species and that the system is*much more
dynamic than was previously assumed.

Practical conclusions from these observations are
straightforward. When precise dose determinations
are needed, one should not conduct any measure-
ments during the first 2-3 h after irradiation; this
is true for any dose. For measuring doses below
1-2kGy, the period 324h after the end of
irradiation is favorable. For doses in the range 5 kGy
to 20 kGy, it is better to perform measurements 3
days after irradiation, that is, in the plateau region.
Finally, to determine precisely doses above 20-
25 kGy, it is recommended that measurements be
performed in the region of the maximum; it occurs at
60-120 h for the dose 25 kGy and at 30-70h for



‘Complex time dependence of the EPR signal 791

the dose 100 kGy. These peculiarities should be
taken into consideration in designing accurate dose
measurement and calibration procedures. ‘

In conclusion, we consider use of an adjacent
reference sample in general. This old technique
has proved to be excellent in its applicability and
usefulness to us in alanine dosimetry. In fact,
although the EPR spectrometer we used in this study
was relatively stable (variations in the ruby signal

10 Gy
1.008

28

1.002

;

0.998

Rel. amplitude, arb. units

5 10 15 20 25 30 35 40
Time after irradiation, h

100 Gy
1.010

1.008
1,006
1.004

1.002

Rel. amplitude, arb. units

1.000

5 10 15 20 25
Time after irradiation, h

1.010 2kGy
1.008,
1.006.
1.004

1.002

Rel. amplitude, arb. units

1.000

20-40" 60; 60-100
Time after irradiation, h

intensity were typically below 1% for periods of
several days), we would not have been able to
measure the time dependencies in a statistically
significant form without simultaneously monitoring
the ruby signal. As an illustration, we present the
plots of the intensities of the alanine and ruby signals
along with their ratios in Fig. 2. It can be also
seen from Fig. 1 that, using this technique, one can
achieve reproducibility in measuring alanine signal

5kG
1.005 y

units

1.004

arb.

3 1.003
1.002
1.001

1.000

Rel. amplitude,

20 40 60 80
Time after irradiation, h

1.007 25 kGy

1.006
1.005
1.004
1.003
1.002
1.001 x
1.000

implitude, arb. units

Rel. ai

20 40 60 80 100 120 140
Time after irradiation, h

100 kGy
1.002

1.001
1.000
0.999

0.998

Rel. amplitude, arb. units

50 100 150 200
Time after irradiation, h

Fig. 1. Time dependence of the amplitude of the central line in the EPR spectra of irradiated
polycrystalline L-2-alanine. The plotted relative amplitude is the result of the division of the alanine signal
peak-to-peak amplitude by the peak-to-peak amplitude of the Cr(3 + ) signal in the adjacent reference
sample. In each experiment, the averaged relative amplitude at the first minimum was taken to be 1.000.