Epilepsy Research (2013) 104, 35—39

journal homepage: www.elsevier.com/locate/epilepsyres

Deficiency of selenium and zinc as a causative factor

for idiopathic intractable epilepsy
Mehmet Seven a,∗, Sarenur Y. Basaran a, Mujgan Cengiz b, Seda Unal b,
Adnan Yuksel c

a
Istanbul University, Cerrahpaşa Medical Faculty, Department of Medical Genetics, Istanbul, Turkey
b
Istanbul University, Cerrahpaşa Medical Faculty, Department of Medical Biology, Istanbul, Turkey
c
Bezmialem University, Medical Faculty, Department of Pediatric Neurology, Istanbul, Turkey

Received 9 August 2011; received in revised form 24 September 2012; accepted 30 September 2012
Available online 25 October 2012

KEYWORDS Summary
Idiopathic epilepsy; Purpose: The accumulation of free radicals may lead to seizures and increase the risk of their
Intractability; recurrence. Glutathione peroxidase and superoxide dismutase are 2 major enzymes that are
Trace elements; involved in antioxidative defense mechanisms. Selenium (Se), zinc (Zn), and copper (Cu) are
Selenium; important trace elements that participate in the structure of these enzymes. The purpose of
Zinc; this study was to evaluate the possible associations between trace elements and idiopathic
Copper intractable epilepsy (IIE) by comparing the levels of Se, Zn, and Cu between patients with IIE
and healthy children.
Methods: Our study was designed as a case—control study with 70 IIE patients and 60 healthy
children who were matched for age, ethnicity, and socioeconomic status. The levels of serum
Se, Zn, and Cu were measured with an atomic absorption spectrophotometer. The results were
statistically analyzed with SPSS version 16.0.
Key findings: We found that the patients with IIE had significantly decreased levels of serum Se
and Zn compared to those of the control group (p < 0.05).
Significance: We believe that this study presents the first reports of decreased levels of Se and
Zn in patients with IIE. These results may provide new insights for delineating the etiological
basis of IIE and its potential therapeutic options.
© 2012 Published by Elsevier B.V.

Abbreviations: Ca, calcium; Cu, copper; GPx, glutathione per-
oxidase; IIE, idiopathic intractable epilepsy; Se, selenium; SOD,
superoxide dismutase; Zn, zinc.
∗ Corresponding author. Tel.: +90 212 414 30 00x22995;
fax: +90 212 414 31 84.
E-mail address: mimseven@istanbul.edu.tr (M. Seven).
Introduction
Epilepsy, which affects about 2—3% of the general popula-
tion and which has a prevalence of 0.8% in Turkish children
between the ages of 0—16, is a common chronic neurologi-
cal disorder with various etiological factors (Hauser et al.,
1996; Serdaroğlu et al., 2004). Epilepsy has been divided into

0920-1211/$ — see front matter © 2012 Published by Elsevier B.V.

http://dx.doi.org/10.1016/j.eplepsyres.2012.09.013
36 M. Seven et al.

idiopathic, symptomatic, and cryptogenic forms. According indicated potential therapy options after the results
to the International League Against Epilepsy, the largest revealed a significant association between these trace ele-
group consists of patients with idiopathic epilepsy (Camfield ment levels and epilepsy.
and Camfield, 2006). Although many research studies have
been conducted on patients with epilepsy, a specific cause Methods
could only be identified in 30—35% of them (Holmes, 1996).
Genetic causes, including mutations in ion and non-ion
This study was composed of 70 (33 males and 37 females)
channel genes and mutations in genes involved in the antiox-
children with IIE (complex partial seizures, 27 patients;
idant system, have been suggested to be responsible for
myoclonic seizures, 13 patients; clonic seizures, 4 patients;
idiopathic epilepsy (Mizielinska, 2007; Lu and Wang, 2009;
tonic seizures, 7 patients; tonic—clonic seizures, 19
Naziroğlu, 2009). In addition, imbalances in trace elements
patients) who were aged between 12 months and 14
that are essential for the activation of enzymes in neuronal
years, and who had 1 or more seizures in the previ-
metabolism have been suggested to be another mechanism
ous 6 months despite being treated with at least 2
that may be involved (Hamed et al., 2004).
antiepileptic drugs, such as carbamazepine + valproate,
The equilibrium of trace elements is essential for a
carbamazepine + valproate + levetiracetam, or carba-
healthy nervous system because most of them contribute to
mazepine + valproate + diphenylhydantoin, according to
the activation of specific enzymes that play important roles
their epileptic seizure type. None of them had any identifi-
in many pathways of the central nervous system. Antiox-
able etiologies. The patients with idiopathic epilepsy who
idative defense mechanisms are an important pathway
responded to the drug therapy were excluded. The patients
involving trace elements. Glutathione peroxidase (GPx),
had no structural central nervous system abnormalities,
which is a selenium (Se)-dependent enzyme that is involved
infections, traumas, or hypoxic ischemic encephalopathy
in antioxidative defense mechanisms, controls the intracel-
histories. The results of the metabolic tests and physical
lular levels of hydrogen peroxide (Hamed et al., 2004). It
and neuroradiological examinations were all in the normal
is well known that the increased production of free radi-
range. The control group consisted of 60 healthy children
cals due to oxidative stress or the decreased functioning
(30 males and 30 females) who were aged between 12
of antioxidative defense mechanisms may lead to seizures
months and 14 years, and who had been admitted to our
or increase the risk of their recurrence (Mizielinska, 2007;
hospital for routine examinations. The groups in this study
Hamed et al., 2004; Willmore and Rubin, 1981; Savaskan
were matched for gender, age, and anthropometric mea-
et al., 2003; Ashrafi et al., 2007a) because oxidative stress
surements. Demographic features, such as socioeconomic
produces peroxidated membrane lipids and damages the
status and geographic area, were similar in the groups.
cells (Hayashi, 2009).
None of the patients were undergoing any dietary restric-
Copper (Cu) and zinc (Zn) are other trace elements that
tions for therapeutic reasons. Both the patient and control
affect the removal of free radicals through their involve-
groups were further divided into 2 subgroups according
ment in the structure of superoxide dismutase (SOD), which
to the following age distributions: 12—60 months and 61
is the most important antioxidant enzyme (Horning et al.,
months—14 years. Informed consents were obtained from
2000; Yüksel et al., 2000; da Cunha et al., 2003). Addition-
the parents of all of the subjects in the 2 groups. The study
ally, Cu induces neurotoxicity by impairing protein structure
was approved by the Ethical Committee of the Cerrahpasa
(Horning et al., 2000), and Zn regulates the central ner-
Medical School, Istanbul University, Turkey.
vous system through the modulation of glutamate and
Venous blood, which was collected in sterile silicone
␥-aminobutyric acid receptor activity (Peter et al., 1987).
tubes, was centrifuged at 2500 rpm for 20 min, and the
Excess intracellular Zn inhibits mitochondrial electron trans-
serum was collected in a sterile tube. The tubes were stored
port by dysregulating intracellular calcium (Ca) homeostasis
at −25 ◦ C until the levels of serum Se, Zn, and Cu were deter-
by impairing the membrane permeability of Ca. The net
mined. Before measurement, all of the tubes were allowed
result of this inhibition results in energy depletion and
to reach room temperature, and they were then treated with
reactive oxygen species accumulation, leading to neuronal
trichloroacetic acid (1:4). In order to induce precipitation,
damage (Dineley et al., 2005).
the serum was left at room temperature for 10 min.
Given the data that was obtained from previous stud-
The mixture was centrifuged at 3000 rpm for 5 min, and
ies on the relationship of Se, Cu, and Zn levels in inducing
the clear supernatant was analyzed with an atomic absorp-
seizures, we hypothesized that, in patients with idiopathic
tion spectrophotometer in order to determine the levels of
intractable epilepsy (IIE), the levels of these 3 trace ele-
Cu and Zn. The levels of Se were determined with a hydride
ments would be in the abnormal range. Studies of trace
generation atomic absorption spectrophotometer (Ericson
element associations in epilepsy usually involve patients
et al., 1986; Ashrafi et al., 2007b).
with unclassified epilepsy. We suggest that this represents
Statistical analyses were performed with SPSS version
a major confounding factor in these studies. Therefore,
16.0 (IBM Corporation, Armonk, NY, USA), and p values less
while designing this current study, we aimed to study Se,
than 0.05 on Student’s t-tests were accepted as statistically
Cu, and Zn levels in patients with IIE. Thus, this study
significant.
represents the first study that was conducted on a group
of patients with epilepsy who were classified according to
their type of epilepsy, and it aimed at assisting in the Results
delineation of the etiological background of this form of
epilepsy. Because the response of the patients to therapy The mean ages of the patients with IIE and the nor-
is also a major concern in this disease, the current study mal children were 85.7 ± 53.99 and 92.71 ± 51.75 months,
Idiopathic epilepsy and trace elements 37

Table 1 Age, gender, copper, zinc, and selenium levels of epileptic patients and control groups.

Age/gender/trace elements Epileptic patients Control groups p

N Mean/SD Range N Mean/SD Range

Age (month) 70 85.71 ± 53.99 12—168 60 92.71 ± 51.75 12—168 >0.05

Male (month) 33 92.03 ± 55.59 12—168 30 92.80 ± 56.17 12—168 >0.05
Female (month) 37 80.37 ± 52.67 12—168 30 92.63 ± 47.89 12—168 >0.05
Cu (mg/L) 0.96 ± 0.29 0.44—1.88 1.06 ± 0.55 0.28—1.96 >0.05
Zn (mg/L) 70 1.76 ± 0.46 0.80—2.80 60 2.03 ± 0.31 1.00—2.92* <0.05
Se (␮mol/L) 4.40 ± 1.75 1.04—9.36 5.77 ± 1.88 1.52—10.16* <0.05
* p < 0.05.

respectively. For the anthropometric measurements, the
average weights for the control and patient groups were
26.77 ± 13.23 kg and 22.80 ± 12.51 kg, respectively, and
the average heights for the control and patient groups
were 123.22 ± 26.16 cm and 114.57 ± 27.55 cm, respec-
tively. There were no significant differences in the
anthropometric measurements, the genders, and the ages
between the patient and control groups and between the
subgroups of these main groups (p > 0.05) (Table 1). The
physical examinations did not reveal any dietary restriction
signs in any of the patients. The electroencephalographic
findings varied according to the types and courses of
the seizures. Table 1 shows the Se, Cu, and Zn lev-
els and their ranges with respect to the genders of the
patient and control groups. When gender was not taken
into account in the patient group, the mean Se, Cu,
and Zn levels were 4.40 ± 1.75 ␮mol/L, 0.96 ± 0.29 mg/L,
and 1.76 ± 0.46 mg/L, respectively. The corresponding
values for the control group were 5.77 ± 1.88 ␮mol/L,
1.06 ± 0.55 mg/L, and 2.03 ± 0.31 mg/L, respectively. The
Se and Zn levels in the patients with epilepsy were
significantly lower than those of the control subjects
(p < 0.05). However, Cu levels showed similar distribu-
tions in both the patient and control groups (p > 0.05).
Significance did not change in terms of age and
gender.
The means and standard deviations of the Se, Cu, and
Zn levels according to the subgroups in the patient and con-
trol groups are shown in Table 2. There were no significant
differences in age and in the Cu levels according to age
distribution in both the patient and control groups (p > 0.05)
(Table 2).
The levels of Se and Zn were significantly lower in the
patient group compared to the control group. Additionally,
both of these trace elements were lower within the sub-
groups of the patient and control groups (p < 0.05).
Discussion
Epilepsy, which affects a significant ratio of the population
(∼2—3%), is a major health problem. The classification of
epilepsy according to a number of factors, such as etiologi-
cal background, affected region of the brain, seizure type,
and accompanying clinical features, has been attempted
by many researchers. Medical therapy with antiepileptic
drugs is effective in most patients, but 30% of the patients
do not benefit from drug therapy and therefore suffer
from intractable epilepsy (McEwan et al., 2004). Intractable
epilepsy is generally defined as epileptic attacks that are
unresponsive to medical treatment. However, there are no
consensus criteria regarding unresponsiveness (i.e., seizure-
free duration, number of administered antiepileptic drugs,
therapy duration, etc.). In a study that was conducted on
patients with intractable epilepsy, Ashrafi et al. (2007b) cat-
egorized the patients as intractable when they developed
at least 1 seizure in a 6-month period despite being treated
with at least 2 antiepileptic drugs.
There have been numerous reports on the association
of trace elements and epilepsy. The cascade of neurotoxic

Table 2 Ages and trace element concentrations in epileptic children and control subjects (ages: 12—60 months and 61
months—14 years).

Age/trace 12—60 months 61 months—14 years

elements
Epileptic patients Control groups Epileptic patients Control groups
(N = 29) (N = 23) (N = 41) (N = 37)
Mean/SD Range Mean/SD Range Mean/SD Range Mean/SD Range

Age (month) 33.41 ± 13.9 12—60 43.43 ± 30.6 12—60 122.9 ± 38. 60—168 123.3 ± 36.1 60—168
Cu (mg/L) 1.02 ± 0.34 0.52—1.88 1.15 ± 0.73 0.28—3.92 0.91 ± 0.24 0.48—1.52 1.00 ± 0.41 0.48—1.96
Zn (mg/L) 1.68 ± 0.48 0.88—2.72 1.97 ± 0.30 1.00—2.52* 1.82 ± 0.45 0.80—2.80 2.07 ± 0.31 1.08—2.92
Se (␮mol/L) 3.91 ± 1.17 1.72—6.24 5.85 ± 2.20 1.52—10.16* 4.75 ± 2.01 1.04—9.36 5.73 ± 1.69 1.96—9.94*
* p < 0.05.
38
events that lead to epileptic seizures is highly complex, but
the main event involves the accumulation of free oxygen
radicals (Hayashi, 2009). Adding more complexity to the
situation, oxygen radical formation has been found to be
both the cause and the result of epileptic seizures (Hamed
et al., 2004; Yüksel et al., 2000). Se, Zn, and Cu are 3
trace elements that are involved in the metabolism of oxy-
gen radicals. For example, Se is involved in the reduction of
peroxide by participating in the structure of GPx, which is
a very important antioxidant enzyme (Hamed et al., 2004).
Low levels of Se and GPx have been found in patients with
epilepsy (Ashrafi et al., 2007a; Yüksel et al., 2001). Se-
deficient rats have been found to be more susceptible to
excitotoxicity (Savaskan et al., 2003).
In another study that was conducted on 2 patients with
intractable epilepsy, the seizures ceased with Se supplemen-
tation in addition to antiepileptic therapy, which suggested
the benefit of this supplementation and which was confirmed
by the return of the seizures after cessation of the Se supple-
mentation (Ramaekers et al., 1994). In addition to these
observations, antiepileptic therapy was the main cause of
the decreased Se levels per se. Children with epilepsy have
also been found to suffer from relative malnutrition, which
adds to the severity of the intractability of their seizures
(Volpe et al., 2007).
SOD is another antioxidant enzyme that removes free
oxygen radicals from the intracellular environment. SOD
requires Cu and Zn for this action (Liochev and Fridovich,
2000). The intraventricular injection of Zn and Cu resulted
in severe seizures. The Zn:Cu ratio has been suggested to
be important in the pathogenesis of seizures. The neu-
roregulatory action of Zn has been suggested to be either
proconvulsive or anticonvulsive (Foresti et al., 2008; Elsas
et al., 2009). Increased levels of Cu have been observed in
some patients with epilepsy (Liu et al., 1998). However, the
results have been mostly conflicting and did not indicate
a clear association between Cu and Zn levels and epilepsy
(Verrotti et al., 2002).
Most of the epilepsy-related research that has been
done on trace elements has been conducted in patients
with epilepsy that was not classified. In the current study,
we included patients with idiopathic epilepsy who did not
respond to medical treatment that included at least 2
antiepileptic drugs and who developed at least 1 seizure in
the previous 6-month period. In order to clarify the effects
of nutrition, we subdivided the patients according to age
groups into those who were younger and older than 5 years
of age. This 5-year threshold was determined as this is the
peak age for the occurrence of most epileptic seizures.
With regard to diet, our patients did not exhibit any physi-
cal examination signs or anthropometric measurements that
revealed insufficient dietary intake or had they been on any
dietary restrictions for therapeutic reasons.
In our study, a significant decrease in Se and Zn levels was
found in patients with IIE, and there were no differences in
gender or age. The latter illustrated that nutritional status
did not make any differences in inducing seizures according
to Se and Zn levels, and, thus, our results did not support the
results of Volpe et al. (2007), which revealed an association
between malnutrition, seizures, and antiepileptic therapy.
The significantly decreased levels of Se in our study rein-
forced the results of Ashrafi et al. (2007a) who also found low
M. Seven et al.
levels of Se in patients with intractable epilepsy. Although
their data were from patients with general epilepsy, it was
important that a significant percentage of the patients in
that study (62.5%) had idiopathic epilepsy. Although our
study group was not identical to the study group of Ashrafi
et al. (2007a), our results seemed to positively correlate
with their results regarding low Se levels in patients with
mostly IIE.
The identification of the etiological background of IIE may
provide new insights and approaches for this disease. This is
the first study to evaluate trace element levels in patients
with IIE, and it showed that the serum Se and Zn levels in
patients with IIE were significantly lower than those of nor-
mal children of the same age. In addition, there were no
differences in the levels between patients who were younger
or older than 5 years of age. However, Cu levels were not
found to differ between the groups. These study results war-
rant future and larger studies of the associations between IIE
and trace element levels in order to delineate the etiological
basis of IIE and its possible therapeutic options. Additionally,
a future study that compares the pre- and post-treatment
levels of the trace elements in patients with IIE should
be performed given the relationships between antiepileptic
drugs and the levels of trace elements.
Conflict of interest
None of the authors has any conflict of interest to disclose.
We confirm that we have read the Journal’s position on
the issues involved in ethical publication and affirm that this
report is consistent with those guidelines.
Acknowledgement
We gratefully thank Prof. Dr. Ulvi AVCIATA (Dean of Yıldız
Technical University, Faculty of Arts and Sciences) for his
technical advice and for allowing the use of their equipment
for this study.
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