1042 Bull. Chem. Soc. Jpn. Vol. 83, No. 9 (2010)
Kinetics of Hydroxyl Radical Spin Adducts
3.3 © 107 M¹1 s¹1, which is approximately consistent with that
reported in a previous paper,27 was lower by two orders than
that of the reaction between DMPO and hydroxyl radical
(3.4 © 109 M¹1 s¹1). It is considered that H2O2 in the presence
of 300 mM of DMPO scavenged only a small amount of
hydroxyl radical even though H2O2 was generated by ultra-
sound irradiation of water. Furthermore, since the concentra-
tion of H2O2 used for photolysis (1 M) was only 3.4 times
higher than that of DMPO (300 mM), the generation of DMPO-
OH by photolysis of 1 M H2O2 was unlikely affected by the
presence of H2O2. SOD did not or very slightly affected the
decay of DMPO-OH, so that there are two possibilities
considered. One is that superoxide anion did not interact
with DMPO-OH, and the other is that very small amount of
superoxide anion was produced by ultrasound irradiation to
water. Since it was reported that superoxide anion produced
by xanthine-xanthine oxidase reaction induced the depletion
of DMPO-OH,28 it is likely that the decay of DMPO-OH
observed in this study was free from the effect of superoxide
anion. Indeed, in the present study, DMPO-OOH was not
detected as shown in Figure 1B while DMPO-OH was detected
in several dozen ¯M. Similarly, DMPO-H was not detected,
either. It was reported that hydrogen atom as well as hydroxyl
radical were generated by sonolysis of water when low-
frequency ultrasound (several dozen to hundred kHz) was
used.12,29,30 However, the 1650 kHz ultrasound used in this
study generated almost no hydrogen atom as reported in our
previous study.16
The half-life of DMPO-OH depended on its initial concen-
tration and varied from 16 to 42 min in this study. Although we
investigated potential factors affecting lifetime of DMPO-OH,
such as dissolved oxygen and H2O2, those factors did not seem
to influence the DMPO-OH decay curve. In addition, the decay
of DMPO-OH after the cessation of irradiation represents likely
spontaneous decay or mutual interaction with DMPO-OH
molecules themselves because the sample used in this study
contained only the ultrapure water, the remaining DMPO,
and DMPO-OH after ultrasound irradiation. Therefore, one
possibility is that the DMPO-OH decayed by second-order
reaction of two molecules of DMPO-OH so that the half-life
of DMPO-OH was in inverse proportion to the initial con-
centration of DMPO-OH. If the second-order reaction was
attributed to two molecules of DMPO-OH, the activation
energy of the reaction was low (26 kJ mol¹1) according to an
Arrhenius plot using the values in Table 2. Thus, we presumed
that the product from the second-order reaction might be a
dimer of DMPO-OH.
fore, it is suggested that the initial concentration of DMPO-OH
is an important factor affecting quantitative analysis of
hydroxyl radical.
As opposed to the potential factors discussed above
(dissolved oxygen and H2O2), the DMPO-OH decay curve
was affected by temperature. Since higher temperature causes
faster decay of DMPO-OH, it was confirmed that the second-
order reaction was accelerated by temperature. If the second-
order reaction for two molecules of DMPO-OH occurred, the
rate constants were quite low compared to those in the reactions
of hydroxyl radical and DMPO even though the temperature
was 75 °C, suggesting that the reaction was slow. On the other
hand, the influence of the temperature on the quantitative
analysis is not negligible as shown in Figure 4.
When the actually measured half-life of each concentration
of DMPO-OH and the calculated value using the equation
(y = 827.76/(x + 15.97)) of the decay curve of 50 ¯M DMPO-
OH were compared, there is little difference between the
two. This finding suggests that concentration of DMPO-OH
decreases according to the equation. For instance, the calcu-
lated half-life of each concentration of DMPO-OH is as
follows; 8.3 min for 100 ¯M DMPO, 4.1 min for 200 ¯M
DMPO-OH, and 2.1 min for 400 ¯M DMPO. The short
lifetime of several hundred ¯M DMPO-OH might be one of
the reasons for the saturation phenomenon of the actually
measured DMPO-OH. However, the spontaneous decay of
DMPO-OH is too slow to explain the whole reaction of
saturation of the actually measured DMPO-OH. Therefore,
hydroxyl radical generated constantly by ultrasound irradiation
might degrade DMPO-OH, which makes difficulty in measure-
ment of several hundred ¯M DMPO-OH correctly. Indeed,
hydroxyl radical is generated linearly with time (Figure 2)
but the actually measured concentration of DMPO-OH is
saturated at 80 ¯M. This finding is in accordance with previous
reports.12,16 Those studies investigated the relationship between
ultrasound irradiation time and concentration of DMPO-OH.
Iwasawa et al.16 reported that the concentration of DMPO-OH
was saturated at around 15 ¯M. The difference of the concen-
tration of saturation between the previous study and ours might
be due to the concentration of DMPO added to the reaction
system and the power of the ultrasound device. The factors
which cause the saturation of actually measured DMPO-OH
should be studied in the future for better understanding of a
generation system which could produce a large amount of
hydroxyl radical.
Optimal concentration of DMPO for photolysis of 1 M H2O2
was 300 mM under the conditions in which 50 ¯M DMPO-OH
was generated. Although the optimal conditions for photolysis
of H2O2 were coincident with those for sonolysis of water in
this study, optimal conditions should be evaluated for each
experiment because they are probably affected by the amount
and the generation rate of hydroxyl radical in each generation
system. As to decay of DMPO-OH generated by photolysis of
H2O2, a decay curve similar to that generated by sonolysis of
water was observed. Therefore, it is thought that the kinetics
related to the decay of DMPO-OH generated by sonolysis of
water and photolysis of H2O2 is almost the same. However,
kinetics of DMPO-OH may vary in other hydroxyl radical
generation systems, such as Fenton reaction or Haber-Weiss
Considering these findings, the difference of half-life
of DMPO-OH between the previous reports26,20,21 and the
present study was mainly due to the difference of the initial
concentration of DMPO-OH generated from different hydroxyl
radical generation systems. According to the equation
(y = 827.76/(x + 15.97)) used in this study, the 60 min half-
life means that the initial concentration of DMPO-OH was
about 14 ¯M. Similarly, as reported in a previous study on
radiolysis of water, DMPO-OH exhibited second-order radical
termination kinetics at initial concentrations of 13 ¯M or more,
followed by first-order termination kinetics at lower concen-
trations,31 which is in agreement with another study.32 There-